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BCL2 Family of Apoptosis-Related Genes: Functions and Clinical Implications in Cancer
Abstract
One of the most effective ways to combat different types of cancer is through early diagnosis and administration of effective treatment, followed by efficient monitoring that will allow physicians to detect relapsing disease and treat it at the earliest possible time. Apoptosis, a normal physiological form of cell death, is critically involved in the regulation of cellular homeostasis. Dysregulation of programmed cell death mechanisms plays an important role in the pathogenesis and progression of cancer as well as in the responses of tumours to therapeutic interventions. Many members of the BCL2 (B-cell CLL/lymphoma 2; Bcl-2) family of apoptosis-related genes have been found to be differentially expressed in various malignancies, and some are useful prognostic cancer biomarkers. We have recently cloned a new member of this family, BCL2L12, which was found to be differentially expressed in many tumours. Most of the BCL2 family genes have been found to play a central regulatory role in apoptosis induction. Results have made it clear that a number of coordinating alterations in the BCL2 family of genes must occur to inhibit apoptosis and provoke carcinogenesis in a wide variety of cancers. However, more research is required to increase our understanding of the extent to which and the mechanisms by which they are involved in cancer development, providing the basis for earlier and more accurate cancer diagnosis, prognosis and therapeutic intervention that targets the apoptosis pathways. In the present review, we describe current knowledge of the function and molecular characteristics of a series of classic but also newly discovered genes of the BCL2 family as well as their implications in cancer development, prognosis and treatment.
... In this study, we show for the first time that BCL2 and NOTCH1 are direct MIR449A targets. BCL2 is an antiapoptotic gene capable of antagonizing the p53 pathway, and has been described as over expressed in different types of lymphoma and AML (Nagy et al, 2003;Thomadaki & Scorilas, 2006). The NOTCH1 gene, which is part of a type 1 transmembrane protein family, is involved in regulation of self-renewal, apoptosis and differentiation. ...
... An important failsafe program against tumourigenesis is provided by the gatekeeper protein p53, and it is generally accepted that this barrier must be overcome during the ontogenesis of a tumour. Interestingly, both NOTCH1 (Beverly et al, 2005;Rosati et al, 2009;Secchiero et al, 2009) and BCL2 (Thomadaki & Scorilas, 2006) have been implicated in suppression of p53 activity in leukaemic cells. Therefore, our findings position repression of MIR449A as a means by which EVI1-positive leukaemic cells circumvent the p53 anti-tumour barrier and it is tempting to speculate that this suppression of the p53 pathway relies on up regulation of both BCL2 and NOTCH1 expression. ...
361
Chromosomal rearrangements involving the EVI1 gene are a recurrent finding in malignant myeloid disorders. These translocations or inversions contribute to ectopic expression or to the formation of fusion genes involving the EVI1 gene. EVI1 transcriptional activation has been reported in up to 10% of acute myeloid leukemia (AML) and is a prognostic marker of poor outcome. MicroRNA (miRNA) deregulation was recently identified as a major contributor to cancer initiation and progression. As miRNA genes were shown to be directly regulated by activated proto-oncogenes, we aimed to identify miRNAs under direct or indirect control of EVI1. To this purpose, we analyzed the expression of 366 miRNAs in 38 EVI1 rearranged/overexpressing patient samples, 6 normal bone marrow controls and 2 EVI1 knockdown model systems (siRNA mediated EVI1 knockdown in the EVI1 rearranged/overexpressing cell lines Kasumi-3 and UCSD-AML1). In total, 24 upregulated and 25 downregulated miRNAs (p<0.05) were shown to be related to the EVI1 expression status. Amongst these, miR-449a was selected for further study based on its homology to the known cancer associated miRNA miR-34a. Downregulation of miR-449a by EVI1 was further confirmed in the leukemic cell line U937 with tetracycline controllabel (tet-off) EVI1 overexpression. Next, direct transcriptional regulation of miR-449a expression by EVI1 was demonstrated by chromatin immunoprecipitation (ChIP). To test the functional consequences of downregulation of miR-449a in AML cells, reconstitution of the expression of miR-449a in the Kasumi-3 and UCSD-AML1 cell lines was performed, which resulted in significantly decreased cell viability, increased apoptosis and differentiation towards the megakaryocytic and monocytic lineages. Interestingly, siRNA mediated knockdown of EVI1 expression in Kasumi-3 or UCSD-AML1 almost completely abrogated the miR-449a induced reduction in cell viability, while electroporation of both cell lines with EVI1 siRNAs alone had essentially no effect on cell viability. These data strongly suggest that repression of miR-449a expression is essential for the survival and growth of EVI1 overexpressing cells and that this requirement is specifically imposed by EVI1 itself. We next demonstrated that the predicted miR-449a targets NOTCH1 and BCL2 were bona fide miR-449a targets using promoter reporter assays. To asses the contribution of these target genes to the observed phenotype upon miR-449a upregulation, knockdown of NOTCH1 and BCL2 was performed, revealing similar effects on cell viability and apoptosis. These results indicated that the effects seen upon treatment of cells with a precursor miR-449a are at least partly mediated through NOTCH1 and BCL2. In conclusion, we provided for the first time evidence that EVI1 mediated downregulation of miR-449a leads to NOTCH1 and BCL2 upregulation and is required for sustained proliferation and survival of EVI1 overexpressing cells. These data also open new perspectives for therapeutic intervention through modulation of miR-449a and/or its target genes.
Disclosures
No relevant conflicts of interest to declare.
... The protein contains each of the four BCL2 homology (BH) domains, BH1 through BH4. The protein can interact and form homo-and heterodimers with pro-apoptotic BCL2 protein family members through the hydrophobic gap that BH1, BH2, and BH3 form [51,52]. Numerous cell types have been shown to undergo apoptosis when exposed to DLM. After three hours of DLM administration, BCL2 expression is considerably decreased, whereas p38 MAP kinase and Bax expression are enhanced in a concentration-dependent manner, according to Western blot analysis [53]. ...
Background
Deltamethrin (DLM) represents one of the most commonly used pesticides. DLM passes through milk, vegetables, and fruits to humans or through animals (veterinary drugs and feeding on contaminated forage) to milk; it can escape from skin to blood and be secreted in breast milk in lactating women. It was believed to have neurotoxic, nephrotoxic, and hepatotoxic properties.
Methods
In order to investigate deltamethrin-induced hepatotoxicity, 64 rats were divided into 8 groups. The control group did not receive any treatment. D 30 mg/kg DLM (body weight) dissolved in corn oil, B 1 mL whey (10¹⁰ cfu/ml of Bifidobacterium logum ATCC 15707), S 1 mL whey (0.5 ppm selenium), BS 1 mL whey (10¹⁰ cfu/mL of Bifidobacterium logum ATCC 15707 + 0.5 ppm selenium), BD 1 mL whey (10¹⁰ cfu/mL of Bifidobacterium logum ATCC 15707 + DLM), SD 1 mL whey (0.5 ppm selenium) + DLM, and BSD 1 mL whey (10¹⁰ cfu/mL of Bifidobacterium logum ATCC 15707) + 0.5 ppm selenium + DLM.
Results
Results marked that manipulation of bifidobacteria, or selenium triggered significant improvement in AST, ALT, GSH, TNF-α, NF-KB and BCL2 as well as reduction in histopathological necrosis, congestion, and degradation.
Conclusion
Whey beverage fortified with Bifidobacterium longum and selenium implicated reduction in oxidative stress, histopathological degradation that accomplished DLM toxicity. Utilization of whey (a byproduct from cheese making) is considered a recycling process which supports ecofriendly practices and sustainability.
... In addition, histological studies on the kidney of CCL 4 -treated mice were similar with other studies. [50][51][52][53] ...
Background
Hepatotoxicity caused by CCL 4 is well known. Geraniol (GNL) has high antioxidant effect that can induces liver regeneration. However, the protective effect of GNL effect on CCL 4 -induced hepatorenal toxicity in pregnant mice has not yet been studied.
Objective
To investigate whether GNL could protect against oxidative stress induced by CCL 4 via the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway, which is regulated by phosphatidylinositol 3 kinase/protein kinase B (PI3K/AKT), and has been found to have protective effects on renal and hepatic tissues.
Materials and Methods
Forty-eight female albino mice weighing 25–30 g were randomly allocated to 4 groups: Group I served as a control; Group II received a toxicity-inducing single dose of 15 μL of CCL 4 on the 4 th day after mating; Group III received 40 mg/kg GNL + CCL 4 (with GNL from the 1 st day of assimilation to delivery); and Group IV received GNL alone from the 1 st day of assimilation to the end of the delivery period. GNL was evaluated for its protective effects on hepatotoxicity in CCL 4 -treated pregnant mice. Litter size, weight, survival rate, and resorption were recorded. In addition, H & E staining was done for liver and kidney pathology as well as biochemical markers and oxidative markers malondialdehyde, superoxide dismutase, and catalase were analyzed.
Results
CCL 4 significantly reduced survival rate and increased resorption after exposure. Alanine transaminase and aspartate aminotransferase concentrations in the serum, tissue MDA, blood urea nitrogen, and creatinine were increased after CCL 4 exposure. GNL improved enzyme and antioxidant levels and prevented CCL 4 -induced hepatic injury in mice. Caspase-3 cleavage was decreased by GNL, which increased PI3K, phosphorylated AKT, Nrf2, and B-cell lymphoma 2.
Conclusion
GNL demonstrates a protective effect against CCl4-induced hepatorenal toxicity, mediated through the activation of the PI3K/AKT signaling pathway and the upregulation of Nrf2. These findings highlight the potential therapeutic implications of GNL in mitigating oxidative stress and inflammation in liver and kidney tissues.
... Bax and Bcl-2 proteins are the main regulatory factors in mitochondrionmediated apoptosis [38] . The Caspase-3 pathway is an important link to apoptosis [39] , and the Caspase family is also an important regulator in the Bcl-2/Bax signaling pathway [40] . Tanshinone I was reported to induce apoptosis of EC cells by inhibiting JAK/STAT signaling pathway and increasing the expression of Caspase-3 [41] . ...
The pathogenesis of endometrial cancer (EC) remains unclear. Currently, there is a lack of available drugs that specifically target EC. Here, we demonstrated that both DBC1 and SIRT1 as oncogenic factors were involved in the development and progression of EC and associated with histological grade, lymph node metastasis, depth of myometrial invasion and FIGO stage, the protein interaction ability of DBC1 and SIRT1 in cancer group decreased compared with the control group, and tanshinone IIA promoted apoptosis and inhibited proliferation of Ishikawa cells through P53-Cleaved caspase-3 pathway and Bcl-2 pathway, a process possibly regulated by CK2/DBC1/SIRT1/P53 signaling pathway. In the xenograft model of Ishikawa origin, high-concentration tanshinone IIA also down-regulated the expression of DBC1 and SIRT1, and inhibited the growth of tumors. Our results indicate that DBC1 and SIRT1 may serve as novel molecular targets and tanshinone IIA may play a role in the treatment of EC.
... The dysregulation of Bcl-2 family proteins, including Bcl-2 as an anti-apoptotic regulator protein and Bax as a pro-apoptotic regulator, is involved in cancer cell apoptosis and modifying programmed cell death pathways by numerous mechanisms, including mitochondrial pathway. 35,36 miR-21 was found to function as a potent oncomiR that regulates cell growth and inhibits apoptosis through regulating the expression of multiple target genes, including PTEN, RECK, and Bcl-2 in NSCLC. 37 Furthermore, it was shown that miR-145 overexpression reduced the expression of Bcl-2 in glioma cancer and induced cell apoptosis. ...
Introduction: Around the world, gastric cancer (GC) is the third leading cause of cancer-related deaths. microRNAs are a group of regulatory non-coding RNAs that are involved in GC progression. miR-145 as a tumor suppressor and miR-21 as an oncomiR were shown to be dysregulated in many cancers, including GC. This research aimed to enhance the expression of miR-145 while reducing the expression of miR-21 and examine their impact on the proliferation, apoptosis, and migration of GC cells. Methods: KATO III cells with high expression levels of miR-21-5p and low expression of miR-145-5p were selected. These cells were then transfected with either miR-145-5p mimics or anti-miR-21-5p, alone or in combination. Afterward, the cell survival rate was determined using the MTT assay, while apoptosis induction was investigated through V-FITC/PI and DAPI staining. Additionally, cell migration was examined using the wound healing assay, and cell cycle progression was analyzed through flow cytometry. Furthermore, gene expression levels were quantified utilizing the qRT-PCR technique. Results: The study's findings indicated that the co-replacement of miR-145-5p and anti-miR-21-5p led to a decrease in cell viability and the induction of apoptosis in GC cells. This was achieved via modulating the expression of Bax and Bcl-2, major cell survival regulators. Additionally, the combination therapy significantly increased sub-G1 cell cycle arrest and reduced cell migration by downregulating MMP-9 expression as an epithelial-mesenchymal transition marker. This study provides evidence for the therapeutic possibility of the combination of miR-145-5p and anti-miR-21-5p and also suggests that they could inhibit cell proliferation by modulating the PTEN/AKT1 signaling pathway. Conclusions: Our research revealed that utilizing miR-145-5p and anti-miR-21-5p together could be a promising therapeutic approach for treating GC.
... BNIP2 belongs to the BCL2 family and plays a role in the mitochondrial apoptotic pathway that underlies chemotherapeutic-induced apoptosis 63 . Moreover, caspases-mediated cleavage of BNIP2 appears to be crucial for the pro-apoptotic function of this protein 64 . ...
We have previously shown that high expression of prolactin-induced protein (PIP) correlates with the response of breast cancer (BC) patients to standard adjuvant chemotherapy (doxorubicin and cyclophosphamide), which suggests that the absence of this glycoprotein is associated with resistance of tumor cells to chemotherapy. Therefore, in the present study, we analyzed the impact of PIP expression on resistance of BC cells to anti-cancer drugs and its biological role in BC progression. Expression of PIP and apoptotic genes in BC cell lines was analyzed using real-time PCR and Western blotting. PIP was detected in BC tissue specimens using immunohistochemistry. The tumorigenicity of cancer cells was analyzed by the in vivo tumor growth assay. Apoptotic cells were detected based on caspase-3 activation, Annexin V binding and TUNEL assay. The interaction of PIP with BC cells was analyzed using flow cytometry. Using two cellular models of BC (i.e. T47D cells with the knockdown of the PIP gene and MDA-MB-231 cells overexpressing PIP), we found that high expression of PIP resulted in (1) increased sensitivity of BC cells to apoptosis induced by doxorubicin (DOX), 4-hydroperoxycyclophosphamide (4-HC), and paclitaxel (PAX), and (2) improved efficacy of anti-cancer therapy with DOX in the xenograft mice model. Accordingly, a clinical study revealed that BC patients with higher PIP expression were characterized by longer 5-year overall survival and disease-free survival. Subsequent studies showed that PIP up-regulated the expression of the following pro-apoptotic genes: CRADD, DAPK1, FASLG, CD40 and BNIP2. This pro-apoptotic activity is mediated by secreted PIP and most probably involves the specific surface receptor. This study demonstrates that a high expression level of PIP sensitizes BC cells to anti-cancer drugs. Increased sensitivity to chemotherapy is the result of pro-apoptotic activity of PIP, which is evidenced by up-regulation of specific pro-apoptotic genes. As high expression of PIP significantly correlated with a better response of patients to anti-cancer drugs, this glycoprotein can be a marker for the prognostic evaluation of adjuvant chemotherapy.
... Bcl-2 is an anti-apoptotic protein that suppresses apoptosis by inhibiting the action of Bcl-2 associated X protein (BAX), a member of the Bcl-2 family that promotes apoptosis via the mitochondrial-mediated pathway. The ratio of Bcl-2 to BAX expression indicates cellular susceptibility to apoptosis [17][18][19]. In a report studying the effects of an autophagy inhibitor in the induced-OA animal models, increased autophagy in cartilages of knee joints was associated with inhibition of the Akt/ mTOR signaling pathway and increased Bcl-2/Bax ratio [20]. ...
Objective:
This study aimed to evaluate the effectiveness of metformin versus placebo in overweight patients with knee osteoarthritis (OA). In addition, to assess the effects of inflammatory mediators and apoptotic proteins in the pathogenesis of OA, the genetic polymorphisms of two genes, one related to apoptosis (rs2279115 of Bcl-2) and the other related to inflammation (rs2277680 of CXCL-16), were investigated.
Methods:
In this double-blind placebo-controlled clinical trial, patients were randomly divided to two groups, one group receiving metformin (n = 44) and the other one receiving an identical inert placebo (n = 44) for 4 consecutive months (starting dose 0.5 g/day for the first week, increase to 1 g/day for the second week, and further increase to 1.5 g/day for the remaining period). Another group of healthy individuals (n = 92) with no history and diagnosis of OA were included in this study in order to evaluate the role of genetics in OA. The outcome of treatment regimen was evaluated using the Knee Injury and Osteoarthritis Outcome Score (KOOS) questionnaire. The frequency of variants of rs2277680 (A181V) and rs2279115 (938C>A) were determined in extracted DNAs using PCR-RFLP method.
Results:
Our results indicated an increase in scores of pain (P ≤ 0.0001), activity of daily living (ADL) (P ≤ 0.0001), sport and recreation (Sport/Rec) (P ≤ 0.0001), and quality of life (QOL) (P = 0.003) and total scores of the KOOS questionnaire in the metformin group compared to the placebo group. Susceptibility to OA was associated with age, gender, family history, CC genotype of 938C>A (Pa = 0.001; OR = 5.2; 95% CI = 2.0-13.7), and GG+GA genotypes of A181V (Pa = 0.04; OR = 2.1; 95% CI = 1.1-10.5). The C allele of 938C>A (Pa = 0.04; OR = 2.2; 95% CI = 1.1-9.8) and G allele of A181V (Pa = 0.02; OR = 2.2; 95% CI = 1.1-4.8) were also associated with OA.
Conclusion:
Our findings support the possible beneficial effects of metformin on improving pain, ADL, Sport/Rec, and QOL in OA patients. Our findings support the association between the CC genotype of Bcl-2 and GG+GA genotypes of CXCL-16 and OA.
... The term "structural homology" across family members refers to one to four areas that are highly homologous to BCL-2, known as the BH (BCL-2-homology region) domains (BH1, BH2, BH3, and BH4). The importance of BH domains for function and heterodimerization amongst members of the same family has been demonstrated via investigations on mutation and deletion [39]. The BCL-X gene, also known as BCL-2L1 (BCL-2-like 1), has a 44% homology to BCL-2. ...
Apoptosis, also known as the programmed death of cells, is responsible for maintaining the homeostasis of tissues and this function is carried out by caspases. The process of apoptosis is carried out via two distinct pathways: the extrinsic pathway, which is governed by death receptors, and the intrinsic pathway, also known as the mitochondrial pathway. The BCL-2 protein family encoded by the BCL-2 gene, located at the 18q21.33 chromosomal location, is in charge of regulating the intrinsic pathway, which is responsible for inducing cell death via the permeabilization of the mitochondrial membrane and the release of apoptosis - inducing components. The BCL-2 homology (BH1, BH2, BH3, BH4) domains of this family proteins are crucial for their functioning and their common BH domains allow interactions between members of the same family and can also serve as indications of pro- or anti-apoptotic activity. A direct correlation may be shown between the overexpression of BCL-2 and the postponement of cell death. It has been determined that a change in the expression of BCL-2 is the root cause of a variety of malignancies, including lung, breast, melanoma, and chronic lymphocytic leukemia, Multiple Sclerosis, Diabetes. In this review, we discuss the therapeutic potential of regulating BCL-2 family connections and their relevance to health and disease.
... Under the stimulation of oxidative stress caused by Cd, the hepatic Bax increased, then the downstream Caspase-3 was up-regulated, and eventually, apoptosis occurred [32]. Anti-apoptotic Bcl-2 plays a central regulatory role in apoptosis [33]. Accordingly, we examined the effect of OPs on Cd-induced hepatic apoptosis by measuring the levels of pro-apoptotic factors (Bax and caspase-3) and anti-apoptotic factor Bcl-2 in Cd-exposed mice. ...
Cadmium (Cd) is a widespread environmental toxicant that can cause severe hepatic injury. Oyster protein hydrolysates (OPs) have potential effects on preventing liver disease. In this study, thirty mice were randomly divided into five groups: the control, Cd, Cd + ethylenediaminetetraacetic acid (EDTA, 100 mg/kg), and low/high dose of OPs-treatment groups (100 mg/kg or 300 mg/kg). After continuous administration for 7 days, the ameliorative effect of OPs on Cd-induced acute hepatic injury in Cd-exposed mice was assessed. The results showed that OPs significantly improved the liver function profiles (serum ALT, AST, LDH, and ALP) in Cd-exposed mice. Histopathological analysis showed that OPs decreased apoptotic bodies, hemorrhage, lymphocyte accumulation, and inflammatory cell infiltration around central veins. OPs significantly retained the activities of SOD, CAT, and GSH-Px, and decreased the elevated hepatic MDA content in Cd-exposed mice. In addition, OPs exhibited a reductive effect on the inflammatory responses (IL-1β, IL-6, and TNF-α) and inhibitory effects on the expression of inflammation-related proteins (MIP-2 and COX-2) and the ERK/NF-κB signaling pathway. OPs suppressed the development of hepatocyte apoptosis (Bax, caspase-3, and Blc-2) and the activation of the PI3K/AKT signaling pathway in Cd-exposed mice. In conclusion, OPs ameliorated the Cd-induced hepatic injury by inhibiting oxidative damage and inflammatory responses, as well as the development of hepatocyte apoptosis via regulating the ERK/NF-κB and PI3K/AKT-related signaling pathways.
... However, it is not clear how ELF-EMF affects cell response. Apoptosis is essential to maintain the homeostatic balance, and any failure in this process might increase the risk of tumor development 46 . Excessive cell growth and abrogated function of cell cycle factors are the main characteristics of cancer cells. ...
Extremely low-frequency electromagnetic field (ELF-EMF) induces biological effects on different cells through various signaling pathways. To study the impact of the ELF-EMF on living cells under an optimal physiological condition, we have designed and constructed a novel system that eliminates several limitations of other ELF-EMF systems. Apoptosis and cell number were assessed by flow cytometry and the Trypan Blue dye exclusion method, respectively. In vitro cell survival was evaluated by colony formation assay. The distribution of cells in the cell cycle, intracellular ROS level, and autophagy were analyzed by flow cytometer. Suspended cells differentiation was assessed by phagocytosis of latex particles and NBT reduction assay. Our results showed that response to the exposure to ELF-EMF is specific and depends on the biological state of the cell. For DU145, HUVEC, and K562 cell lines the optimum results were obtained at the frequency of 0.01 Hz, while for MDA-MB-231, the optimum response was obtained at 1 Hz. Long-term exposure to ELF-EMF in adherent cells effectively inhibited proliferation by arresting the cell population at the cell cycle G2/M phase and increased intracellular ROS level, leading to morphological changes and cell death. The K562 cells exposed to the ELF-EMF differentiate via induction of autophagy and decreasing the cell number. Our novel ELF-EMF instrument could change morphological and cell behaviors, including proliferation, differentiation, and cell death.
... Bcl-2 protein is encoded by BCL2 gene, a proto-oncogene that promotes tumorigenesis by inhibiting cell death; the antiapoptotic action of Bcl-2 protects cells against the effect of both endogenous and exogenous factors, including chemotherapeutic drugs and glucocorticoids [27]. On the other hand, the loss of Bcl-2 expression might indicate that the tumor shifted toward different pro-survival pathways associated with more aggressive behavior; on this account, Bcl-2 loss has been proposed as a possible marker of tumor aggressiveness [28]. ...
Background
Uterine leiomyosarcoma (uLMS) may show loss of expression of B-cell lymphoma-2 (Bcl-2) protein. It has been suggested that Bcl-2 loss may both be a diagnostic marker and an unfavorable prognostic marker in uLMS.
Objective
To define the diagnostic and prognostic value of Bcl-2 loss in uLMS through a systematic review and meta-analysis.
Methods
Electronic databases were searched from their inception to May 2020 for all studies assessing the diagnostic and prognostic value of Bcl-2 loss of immunohistochemical expression in uLMS. Data were extracted to calculate odds ratio (OR) for the association of Bcl-2 with uLMS vs leiomyoma variants and smooth-muscle tumors of uncertain malignant potential (STUMP), and hazard ratio (HR) for overall survival; a p value < 0.05 was considered significant.
Results
Eight studies with 388 patients were included. Loss of Bcl-2 expression in uLMS was not significantly associated with a diagnosis of uLMS vs leiomyoma variants and STUMP (OR = 2.981; p = 0.48). Bcl-2 loss was significantly associated with shorter overall survival in uLMS (HR = 3.722; p = 0.006). High statistical heterogeneity was observed in both analyses.
Conclusion
Loss of Bcl-2 expression appears as a significant prognostic but not diagnostic marker in uLMS. The high heterogeneity observed highlights the need for further research and larger studies.
... They are divided into two groups, one containing anti-apoptotic proteins (Bcl-2) and the other containing pro-apoptotic proteins (Bax). Functions of these proteins, for example, controlling permeability of mitochondrial membrane, releasing mitochondrial apoptogenic factors into the cytoplasm, and regulating apoptosis, are dependent on I their ability to homo-or heterodimerize with BH3 domains, and (ii) the integration of the carboxy-terminal hydrophobic domain, or TM (transmembrane) domain, into specific cytoplasmic membranes [5,6]. ...
In today's medical environment, natural products have made a substantial contribution to the therapeutic approach in the treatment of diseases ranging from the simple to the complex. The old or traditional approach of standardization in medicinal plant research is a time-consuming, costly, and to some extent antiquated process. As a result, a computational technique that includes an in silico molecular docking simulation study has become an important tool for drug development, standardisation, and screening of phytochemicals. To investigate the cardioprotective research and the interaction of the strong chemical against Bax and Bcl-2 cardiomyocyte gene, docking was conducted using multiple Protein Data Bank files (3EOO, 3D2U, 2I42, and 3D2Y). The Anthraquinone has shown more potent interaction with apoptotic regulators Bcl-2 and Bax genes by showing good binding energy. The study also evident that Anthraquinone (UBA) was an ideal drug agent with better drug likeliness. Further, the compound can be used as therapeutic molecule for myocardial infarction. However, the results are preliminary and experimental evaluation will be carried out in near future.
... BCL-2 prevents MOMP by sequestering the pro-apoptotic proteins [102]. This action of BCL-2 and other anti-apoptotic members of this family blocks the release of cytochrome c, a hallmark of mitochondrial apoptosis [103], which consequently prevents apoptosis activation and increases cell survival [104,105]. This anti-apoptotic activity contributes to cancer development and also resistance to chemotherapy, as cancer cells often control the death mechanisms by elevating the expression of BCL-2, BCL-XL, and MCL-1, which neutralises the apoptotic action of the BH3-only proteins of the BCL-2 family [106]. ...
Acute myeloid leukemia (AML) is a highly heterogeneous malignancy characterized by the clonal expansion of myeloid stem and progenitor cells in the bone marrow, peripheral blood, and other tissues. AML results from the acquisition of gene mutations or chromosomal abnormalities that induce proliferation or block differentiation of hematopoietic progenitors. A combination of cytogenetic profiling and gene mutation analyses are essential for the proper diagnosis, classification, prognosis, and treatment of AML. In the present review, we provide a summary of genomic abnormalities in AML that have emerged as both markers of disease and therapeutic targets. We discuss the abnormalities of RARA, FLT3, BCL2, IDH1, and IDH2, their significance as therapeutic targets in AML, and how various mechanisms cause resistance to the currently FDA-approved inhibitors. We also discuss the limitations of current genomic approaches for producing a comprehensive picture of the activated signaling pathways at diagnosis or at relapse in AML patients, and how innovative technologies combining genomic and functional methods will improve the discovery of novel therapeutic targets in AML. The ultimate goal is to optimize a personalized medicine approach for AML patients and possibly those with other types of cancers.
... Caspases regulate the intrinsic and extrinsic apoptosis pathways, resulting in DNA fragmentation, cytoskeletal breakdown and protein degradation (73,74). This current study showed that the expression of Caspase 3 was decreased after knockdown of BCLAF1 expression. ...
Cancer arises from a multi‑step cellular transformation process where some mutations may be inherited, while others are acquired during the process of malignant transformation. Aberrations in the BCL2 associated transcription factor 1 (BCLAF1) gene have previously been identified in patients with cancer and the aim of the present study was to identify structural variants (SVs) and the effects of BCLAF1 gene silencing on cell transformation. Whole‑genome sequencing was performed on DNA isolated from tumour biopsies with a histologically confirmed diagnosis of oesophageal squamous cell carcinoma (OSCC). Paired‑end sequencing was performed on the Illumina HiSeq2000, with 300 bp reads. Reads were aligned to the Homo sapiens reference genome (NCBI37) using ELAND and CASAVA software. SVs reported from the alignment were collated with gene loci, using the variant effect predictor of Ensembl. The affected genes were subsequently cross‑checked against the Genetic Association Database for disease and cancer associations. BCLAF1 deletion was identified as a noteworthy SV that could be associated with OSCC. Transient small interfering RNA‑mediated knockdown of BCLAF1 resulted in the altered expression of several downstream genes, including downregulation of the proapoptotic genes Caspase‑3 and BAX and the DNA damage repair genes exonuclease 1, ATR‑interacting protein and transcription regulator protein BACH1. BCLAF1 deficiency also attenuated P53 gene expression. Inhibition of BCLAF1 expression also resulted in increased colony formation. These results provide evidence that the abrogation of BCLAF1 expression results in the dysregulation of several cancer signalling pathways and abnormal cell proliferation.
... We analyzed the protective effect of rhFGF12 by detecting apoptosis-related indicators. Bcl-2/Bax is a powerful indicator of cell anti-apoptosis (Thomadaki and Scorilas 2006). The results showed that the rhFGF12 played a protective role by inhibiting apoptosis (Fig. 4c). ...
In recent years, an increasing number of studies have shown that fibroblast growth factor 12 (FGF12) plays important roles in regulating neural development and function. Importantly, changes of FGF12 expression are thought to be related to the pathophysiology of many neurological diseases. However, little research has been performed to explore the protective effect of FGF12 on nerve damage. This study aims to explore its neuroprotective effects using our recombinant humanized FGF12 (rhFGF12). The hFGF12 gene was cloned and ligated into an expression vector to construct a recombinant plasmid pET-3a-hFGF12. Single colonies were screened to obtain high expression engineering strains, and fermentation and purification protocols for rhFGF12 were designed and optimized. The biological activities and related mechanisms of rhFGF12 were investigated by MTT assay using NIH3T3 and PC12 cell lines. The in vitro neurotoxicity model of H2O2-induced oxidative injury in PC12 cells was established to explore the protective effects of rhFGF12. The results indicate that the beneficial effects of rhFGF12 were most likely achieved by promoting cell proliferation and reducing apoptosis. Moreover, a transgenic zebrafish (islet) with strong GFP fluorescence in the motor neurons of the hindbrain was used to establish a central injury model caused by mycophenolate mofetil (MMF). The results suggested that rhFGF12 could ameliorate central injury induced by MMF in zebrafish. In conclusion, we have established an efficient method to express and purify active rhFGF12 using an Escherichia coli expression system. Besides, rhFGF12 plays a protective effect of on nerve damage, and it provides a promising therapeutic approach for nerve injury.
Key points
• Effective expression and purification of bioactive rhFGF12 protein in E. coli.
• ERK/MAPK pathway is involved in rhFGF12-stimulated proliferation on PC12 cells.
• The rhFGF12 has the neuroprotective effects by inhibiting apoptosis.
... 33 Inactivation of the apoptotic pathways and overexpression of anti-apoptotic proteins lead to apoptosis imbalance and survival of HCC cells. 34 Our results showed that the expressions of the pro-apoptotic proteins Bax and caspase-3 were upregulated, and the expression of the anti-apoptotic protein Bcl-2 was decreased in H22 xenograft tumors after TB-TF administration. TB-TF significantly promoted the apoptosis of H22 cells, which was also confirmed by flow cytometry analysis. ...
he fruits of Terminalia bellirica (Gaertn.) Roxb. (TB) are used as a multi-use therapeutic herbal product in
the Tibetan medicinal system and are prescribed as a general health tonic in the traditional Ayurvedic med�icinal system. It has been demonstrated that these fruits have a variety of pharmacological activities, includ�ing anti-tumor, anti-oxidative, anti-inflammatory, hepatoprotective and immunoregulatory effects, etc.
However, the therapeutic effects of tannins in TB on HCC and the underlying mechanisms remain unchar�acterized. In the current study, we aimed to identify the anti-tumor effect of tannins in TB by employing a
H22 xenograft mouse model and by performing cell-based in vitro studies with the assistance of the
network pharmacology analysis. The crude extract of TB was purified to yield total tannin fraction (TB-TF),
and our results found that TB-TF significantly inhibited the tumor growth of H22 xenografts in mice by
inducing apoptosis and reducing angiogenesis. A total of 90 compounds were then identified in TB-TF by
UPLC-MS/MS, and 27 were found in serum after oral administration of TB-TF in mice. The network
pharmacology analysis based on these absorbed components was performed and, along with experimental
evidence, it revealed that the ERBB, PI3K-Akt, and MAPK signaling pathways may be involved in the anti�tumor effect of TB-TF on HCC. Furthermore, we suggested that TB-TF effectively modulated the immuno�suppressive tumor microenvironment in H22 xenograft mice. In summary, our study demonstrated that
TB-TF could be developed as a functional food, which is not only a promising anti-cancer reagent but also
a potential candidate with bright prospects for the emerging trends of immunotherapy for HCC.
... Cyclin D1, one of cell cycle regulatory proteins, plays a key role in determining cell cycle transition from G1 to S phase. Whereas, caspase exists in the form of zymogens in healthy cells, which can be activated by interacting with specific adaptor proteins to promote conformational changes and autocatalytic processes, or proteolysis by already active caspase [43][44][45][46][47]. In our present study, we observed that the expression of Bcl-2 and Cyclin D1 were reduced by RNAi targeting against Aurora A in the U251 cells, on the contrary, the expression level of caspase-3 was upregulated obviously compared with the control groups (Fig. 5). ...
... 33 Inactivation of the apoptotic pathways and overexpression of anti-apoptotic proteins lead to apoptosis imbalance and survival of HCC cells. 34 Our results showed that the expressions of the pro-apoptotic proteins Bax and caspase-3 were upregulated, and the expression of the anti-apoptotic protein Bcl-2 was decreased in H22 xenograft tumors after TB-TF administration. TB-TF significantly promoted the apoptosis of H22 cells, which was also confirmed by flow cytometry analysis. ...
The fruits of Terminalia bellirica (Gaertn.) Roxb. (TB) are used as a multi-use therapeutic herbal product in the Tibetan medicinal system and are prescribed as a general health tonic in the traditional Ayurvedic medicinal system. It has been demonstrated that these fruits have a variety of pharmacological activities, including anti-tumor, anti-oxidative, anti-inflammatory, hepatoprotective and immunoregulatory effects, etc. However, the therapeutic effects of tannins in TB on HCC and the underlying mechanisms remain uncharacterized. In the current study, we aimed to identify the anti-tumor effect of tannins in TB by employing a H22 xenograft mouse model and by performing cell-based in vitro studies with the assistance of the network pharmacology analysis. The crude extract of TB was purified to yield total tannin fraction (TB-TF), and our results found that TB-TF significantly inhibited the tumor growth of H22 xenografts in mice by inducing apoptosis and reducing angiogenesis. A total of 90 compounds were then identified in TB-TF by UPLC-MS/MS, and 27 were found in serum after oral administration of TB-TF in mice. The network pharmacology analysis based on these absorbed components was performed and, along with experimental evidence, it revealed that the ERBB, PI3K-Akt, and MAPK signaling pathways may be involved in the anti-tumor effect of TB-TF on HCC. Furthermore, we suggested that TB-TF effectively modulated the immunosuppressive tumor microenvironment in H22 xenograft mice. In summary, our study demonstrated that TB-TF could be developed as a functional food, which is not only a promising anti-cancer reagent but also a potential candidate with bright prospects for the emerging trends of immunotherapy for HCC.
... In the present study, miR-23a-3p was predicted and verified to target BCL2, an anti-apoptotic gene family member, in HLE-B3 cells, which may improve the current understanding of the role of miR-23a-3p in numerous types of human disease (24,25). In a previous study, BCL2 reduced cell apoptosis by acting via cellular signal transduction pathways or inhibiting lipid oxidation via inhibition of oxygen free radicals (26). ...
Cataracts account for ~50% of the cases of blindness in individuals worldwide. The apoptosis of lens epithelial cells (LECs) occurs during the formation of cataracts, which is a non-congenital condition. Numerous microRNAs (miRs) have been reported to regulate apoptosis in LECs. For instance, miR-23a expression levels were shown to be upregulated in cataractous lenses; however, the function of miR-23a in cataracts remains undetermined. To establish an in vitro model of cataracts, human LECs, HLE-B3 cells, were induced with 200 µmol/l H2O2 for 24 h. HLE-B3 cells were transfected with the miR-negative control (NC) mimic, miR-23a-3p mimic, miR-NC inhibitor, miR-23a-3p inhibitor, small interfering RNA (siRNA) targeting BCL2 (siRNA-BCL2) and siRNA-NC. The expression levels of miR-23a-3p were detected using reverse transcription-quantitative PCR. The interaction between miR-23a-3p and the 3'-untranslated region (UTR) of the target mRNA BCL2 was predicted by TargetScan 7.1, and further validated using a dual luciferase reporter assay. The BCL2 protein expression levels were analyzed using western blotting, cell proliferation was determined using a CCK-8 assay and the levels of cell apoptosis were analyzed using flow cytometric analysis. The results of the present study revealed that the expression levels of miR-23a-3p were significantly upregulated, while the expression levels of BCL2 were significantly downregulated in H2O2-induced HLE-B3 cells compared to untreated control cells. BCL2 was shown to be a target of miR-23a-3p. The miR-23a-3p inhibitor subsequently attenuated H2O2-induced apoptosis and increased the proliferation of HLE-B3 cells, which was partially reversed by siRNA-BCL2. In conclusion, the findings of the current study suggested that the inhibition of miR-23a-3p may attenuate H2O2-induced cataract formation by targeting BCL2, thus providing a novel therapeutic target for the treatment of patients with cataracts in the clinic.
... a variety of apoptotic stimuli, including those associated with cancer chemotherapeutic agents, and confers on cancer cells resistance to current therapeutic agents. That is why, Bcl-2 proteins are a promising molecular target for the several pharmaceutical companies and academia to develop promising anti-cancer agents(Thomadaki & Scorilas, 2006). Within the following context, a descriptive review of the small molecules within clinical development targeting Bcl-2 family proteins(Figure 3). ...
... Apoptosis is mediated via two distinct pathways: the intrinsic or mitochondrial pathway, triggered by intracellular signals, and the extrinsic or death receptor pathway, triggered by extracellular signals [7]. The strict regulation of the intrinsic apoptotic pathway is partly achieved through the action of the BCL2 family proteins [8]. BCL2 family members can be characterized as pro-or anti-apoptotic, depending on their effect on the pathway. ...
The utility of circular RNAs (circRNAs) as molecular biomarkers has recently emerged. However, only a handful of them have already been studied in colorectal cancer (CRC). The purpose of this study was to identify new circRNAs deriving from BCL2L12, a member of the BCL2 apoptosis-related family, and investigate their potential as biomarkers in CRC. Total RNA extracts from CRC cell lines and tissue samples were reversely transcribed. By combining PCR with divergent primers and nested PCR followed by Sanger sequencing, we were able to discover two BCL2L12 circRNAs. Subsequently, bioinformatical tools were used to predict the interactions of these circRNAs with microRNAs (miRNAs) and RNA-binding proteins (RBPs). Following a PCR-based pre-amplification, real-time qPCR was carried out for the quantification of each circRNA in CRC samples and cell lines. Biostatistical analysis was used to assess their potential prognostic value in CRC. Both novel BCL2L12 circRNAs likely interact with particular miRNAs and RBPs. Interestingly, circ-BCL2L12-2 expression is inversely associated with TNM stage, while circ-BCL2L12-1 overexpression is associated with shorter overall survival in CRC, particularly among TNM stage II patients. Overall, we identified two novel BCL2L12 circRNAs, one of which can further stratify TNM stage II patients into two subgroups with substantially distinct prognosis.
... An explanation interpretation of the expression profiling results is the role of MEG3 in coordinating vascular remodeling. MEG3 depletion caused the upregulation of two important antiapoptotic proteins, bcl-2-related protein A1 (BCL2A1) and baculoviral IAP repeat containing 3 (BIRC3) [ 12]. VSMC apoptosis can perform major changes in arterial architecture, especially when coordinated with matrix turnover and VSMC proliferation. ...
Background: Ischemic stroke (IS) is a highly heterogeneous disease with variable pathogenesis. Due to the lack of early predictive markers, the mortality rate of IS remains high worldwide. This study focused on the expression pattern of long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) and its prognostic value in IS. Methods: Twenty-four ICR mice (30-35g, 12 males and 12 females) were involved in this study. Middle cerebral artery occlusion (MCAO) was established to simulate an IS environment. Oxygen-glucose deprivation/reoxygenation (OGD/R) in cells were also performed. A total of 215 IS patients and 153 age and gender-matched controls were included. Quantitative Real-time PCR (qRT-PCR) was performed to detect the expression of MEG3 in peripheral blood leukocytes. Results: The MEG3 expression in MCAO group was significantly higher than control group (P=0.004), and the survival time of high MEG3 group was significantly lower than that of the low MEG3 group (P=0.042). OGD/R promotes cell apoptosis, while si-MEG3 inhibits apoptosis. Results of WB showed that, expression level of Bax was down regulated by si-MEG3, while OGD/R increased its expression level. MEG3 was significantly up-regulated in IS patients and associated with the National Institutes of Health Stroke Scale (NIHSS) (r=0.347, P<0.001), modified Rankin Scale (mRS) (r=0.385, P<0.001), high sensitivity C-reactive protein (Hs-CRP) (r=0.221, P=0.002) level and infarct volume (r=0.201, P=0.006). Overall survival analysis showed that patients with higher MEG3 expression had a relatively poor prognosis (P<0.001). Meanwhile, multivariate analysis revealed that MEG3 was an independent prognostic marker of functional outcome and death in patients with IS. Conclusions: Current study suggested that MEG3 might be considered as a potential prognostic indicator in IS.
... Inhibition of Ptgs2 effectively increased the sensitivity of tumors to drugs [140]. Similarly, Bcl-2, Kit, K-Ras, and Fos genes have been found to be associated with cancer [141][142][143][144]. These genes play an important role in the sleep-wake cycle regulation and are shown to be correlated with sleep [43,44,[145][146][147]. ...
Sleep is essential for the survival of most living beings. Numerous researchers have identified a series of genes that are thought to regulate "sleep-state" or the "deprived state". As sleep has a significant effect on physiology, we believe that lack of total sleep, or particularly rapid eye movement (REM) sleep, for a prolonged period would have a profound impact on various body tissues. Therefore, using the microarray method, we sought to determine which genes and processes are affected in the brain and liver of rats following nine days of REM sleep deprivation. Our findings showed that REM sleep deprivation affected a total of 652 genes in the brain and 426 genes in the liver. Only 23 genes were affected commonly, 10 oppositely, and 13 similarly across brain and liver tissue. Our results suggest that nine-day REM sleep deprivation differentially affects genes and processes in the brain and liver of rats.
... The BCL2L15 protein is localized to the cytoplasm of intestinal epithelial cells (Ozoren et al., 2009). It does not possess any signal peptide or C-terminal membrane anchor and, consequently, it is not associated with any cellular organelles (Coultas et al., 2003;Ozoren et al., 2009), unlike other members of the BCL2 family (Thomadaki and Scorilas, 2006). However, according to Gene Ontology analysis, it may also be localized in the nucleus, while there are also indications regarding its localization to other cellular organelles and to the cytoskeleton (Gaudet et al., 2011). ...
... Our study on DMAMCL in OS cells shows a similar cell death mechanism. BCL-2 family proteins play a pivotal role in mediating the apoptosis process [25,35,36]. Previous studies discovered that DMAMCL and its analogues or prodrugs induced apoptosis in glioma cells, leukemia cells, and leukemia stem cells by regulating the expression of BCL-2 family members [16,29,37]. ...
Osteosarcoma (OS) is the most common primary malignancy of bone that mostly affects children, adolescents, and young people. Despite advances have been made in multimodal therapy of OS, the long-term survival rate has reached a plateau, and the main obstacles are bad response to chemotherapy and gained chemoresistance. In this study, we tested the therapeutic effect of a newly reported drug, DMAMCL, on OS. Five human OS cell lines (143B, MNNG, MG63, Saos-2, U-2OS), and the mouse fibroblast cell line (NIH3T3) and human retinal epithelial cell (ARPE19) were used. The anti-tumor effect of DMAMCL was studied by MTS assay or IncuCyte-Zoom (in vitro), and Xenograft-mice-model (in vivo). Changes of cell cycle, apoptotic cells, caspase3/7 activities, and stemness after DMAMCL treatment were investigated. BAX siRNAs were used to knockdown the expression of BAX. Expressions of CyclinB1, CDC2, BCL-2 family, PARP, CD133, and Nanog were measured by Western Blotting. DMAMCL-induced dose-dependent OS cell death in vitro, and suppressed tumor growth and extended the survival of xenograft-bearing mice. DMAMCL-induced G2/M phase arrest in vitro, and apoptosis both in vitro and in vivo. Down-regulation of BAX expression attenuated the DMAMCL-induced OS cell death in vitro. We also found that DMAMCL inhibited the stemness in OS cells. These results indicated that DMAMCL possess therapeutic value in OS and may be a promising candidate for the new drug discovery for OS therapy.
... However, transcription of the human BCL2L12 gene and maturation of its primary transcripts generates another sixty-two BCL2L12 transcripts, nineteen of which are predicted to encode distinct protein isoforms, whereas the rest of them are candidates for nonsense-mediated mRNA decay (Adamopoulos et al., 2016;Kontos and Scorilas, 2012). In fact, alternative splicing is characteristic for the most members of this family (Thomadaki and Scorilas, 2006;Artemaki et al., 2019). With regard to the BCL2L12 splice variants, these result from retention of intron 1, intron 2, or both of them; the use of two distinct 5′ alternative splice sites in exon 3, two distinct 5′ alternative splice sites in exon 3, and/or two distinct 3′ alternative splice sites in exon 5; and/or exon skipping of one or multiple exons (Adamopoulos et al., 2016;Kontos and Scorilas, 2012). ...
The BCL2L12, one of the latest discovered members of the BCL2 family, has both pro- and anti-apoptotic roles that are cell-type-dependent. Its role in tumorigenesis is highly implicated. Sixty-three splice variants of this gene have been identified so far with significant differences in expression patterns between various cancer cell lines. Presently, little is known regarding the regulation of expression of the BCL2L12 gene. For the vast majority of BCL2L12 gene splice variants, the 5'- and 3'-untranslated regions as well as their transcriptional regulation have not been determined yet. The aim of this study was to get insight into the regulation of the BCL2L12 gene transcription in human chronic myelogenous leukemia (K562) cell line. Our results point to the activity of novel transcription start site of the BCL2L12 gene and indicate that Sp1 and GATA-1 transcription factors could be involved in the regulation of BCL2L12 gene expression in K562 cells. Previously reported active promoter of BCL2L12 gene differs from the one we described in our study. If this novel BCL2L12 promoter is confirmed to be active in other malignancies, transcripts generated from this region could be considered as new cancer-specific biomarkers. The results of our study contribute to the better understanding of the transcriptional regulation of the BCL2L12 gene.
... Apoptosis is essential to maintain the homeostatic balance and any failure in this process might increase the risk of tumor development. Activation of apoptosis signaling pathways is one of the most effective therapeutic strategies in cancer treatment (44). Reported evidence correlating the Figure 5: Induction of apoptosis in MC4-L2 cells exposed to 1 Hz and 100 mT ELF-MF at varying times. ...
Background
Extremely low-frequency magnetic field (ELF-MF) significantly induces apoptosis in cancer cells. To study the biological effects of the ELF-MF on tumor cells, we have designed and constructed a new exposure system that provides a uniform magnetic field with negligible temperature fluctuations during the exposure. Additionally, it provides ideal physiological conditions for live cells inside the incubator. This ELF-MF exposure system eliminates several limitations and disadvantages of other low frequency magnetic field systems; it generates a magnetic field with a frequency of 0 to 70 Hz with a maximum magnetic flux density of 150 mT.
Methods
The capabilities of the setup were examined using a 1 Hz, 100 mT magnetic field, at various exposure times, to induce apoptosis-mediated cell death in the MC4-L2 cell line. After exposure, apoptosis was assessed by flow cytometry.
Results
A biphasic response was observed in cells exposed to ELF-MF: at first a decreasing apoptotic rate during 2-12 hours exposure time was detected, after which apoptosis gradually increased during 24-120 hours of exposure.
Conclusions
We show that ELF-MF exposure with a frequency of 1 Hz and intensity of 100 mT induces apoptosis in MC4-L2 cancer cells in a time-dependent manner. These results show the significance of the long term studies of the ELF exposure effects.
... In this study, the OR and P values of the patients genotyped as GT and TT in their rs1894720 SNP of MIAT indicated that they were associated Bcl2l2 is a member of the Bcl-2 protein family. As an anti-apoptotic regulator, Bcl2l2 can promote cell survival and decrease cell apoptosis under cytotoxic conditions [37,38]. Bcl2l2 is also aberrantly expressed in a wide range of cancer cells and has been implicated in carcinogenesis [39]. ...
Introduction
Cataracts caused by old age are one of the most frequent causes for blindness and poor vision worldwide. In this study, we aimed to clarify the possible role of rs1894720 polymorphism in the pathogenesis of age-related cataract.
Material and methods
Rs1894720 polymorphism genotype was detected by TaqMan. Bioinformatics analysis, luciferase assay, real-time PCR, western blot, and protein density analysis were conducted to establish the correlations between MIAT and miR-26b as well as between BCL2L2 and miR-26b. Flow cytometry and MTT assay were also performed to observe the effect of MIAT/miR-26b/BCL2L2 signalling pathway on the status of cell apoptosis and viability.
Results
MIAT functioned as an endogenous competing RNA to sponge miR-26b. In addition, BCL2L2 was identified as a target of miR-26b. Therefore, the expression of miR-26b was obviously suppressed by MIAT or anti-miR-26b, while the mRNA and protein expression of BCL2L2 was up-regulated in the presence of MIAT or anti-miR-26b. Moreover, the positive effect of MIAT on BCL2L2 expression was exerted via inhibition of the expression of miR-26b. In addition, the cells transfected with MIAT or anti-miR-26b showed suppressed expression of caspase-3 and reduced apoptosis index but higher cell viability, indicating that MIAT could suppress cell apoptosis via inhibition of miR-26b expression. Furthermore, the subjects carrying the GT and TT genotypes of single-nucleotide polymorphism (SNP) rs1894720 were associated with a higher risk of age-related cataracts, as indicated by their odds ratio (OR) and p values.
Conclusions
Rs1894720 SNP could down-regulate the expression of MIAT, thus leading to reduced BCL2L2 expression and enhanced epithelial cell apoptosis in the lens, eventually increasing the incidence of age-related cataract.
... Besides, our study indicated that the local environment chemicals, especially sugar content and oxygen content, regulated the expression of MEG3. Highly expressed MEG3 inhibits two important anti-apoptotic proteins, BCL-2-related protein A1 and baculoviral IAP repeat-containing 3 12) on the one hand, and the other hand, as our results showed, MEG3 can also stimulate expressions of apoptosis-related proteins, such as Bax Advance Publication Journal of Atherosclerosis and Thrombosis Accepted for publication: September 24, 2019 Published online: October 25, 2019 ...
Aim: This study focused on the expression pattern of long non-coding RNA maternally expressed gene 3 (MEG3) and its value in ischemic stroke (IS).
Methods: The expression pattern and the roles of MEG3 in the development of IS were explored in mice IS model and human brain microvascular endothelial cells (hBMECs). A case-control study, including 215 IS patients and 153 controls, was also conducted to investigate its prognostic value.
Results: In vivo study showed that MEG3 increased significantly in the IS group (P=0.004), and its level remained stable within 3 to 48h after the onset of IS. Besides, the survival time of the mouse in the high MEG3 group was significantly lower than that in the low MEG3 group (P=0.042). In vitro study showed that oxygen-glucose deprivation (OGD) treatment significantly up-regulated expressions of MEG3, Bax, and cleaved caspase-3, and further promoted apoptosis of hBMECs, while si-MEG3 blocked these effects. A human study showed that MEG3 increased markedly within 48h of IS onset and was positively associated with the National Institutes of Health Stroke Scale (r=0.347, P＜0.001), modified Rankin Scale (r=0.385, P＜0.001), high-sensitivity C-reactive protein (r=0.221, P=0.002) level, and infarct volume (r=0.201, P=0.006). Overall survival analysis showed that patients with higher MEG3 expression within 48h had a relatively poor prognosis (P＜0.001). Meanwhile, multivariate analysis revealed that MEG3 was an independent prognostic marker for unfavorable functional outcome and death in IS patients.
Conclusions: This study suggested that MEG3 might be considered as an intervention point and potential prognostic indicator for IS.
... The additional exploration revealed an effect on anti-apoptotic gene, Bcl-2, upregulation. Bcl-2 family is apoptotic related genes controlling program cell death (Czabotar et al., 2014;Thomadaki and Scorilas, 2006) which consists of both anti-apoptotic and pro-apoptotic genes (Danial, 2007). The expressions of Bcl-2 family such as B cell lymphoma 2 (Bcl-2) which is anti-apoptotic gene marker was found to block the program rather than promote cell proliferation rate (Hardwick and Soane, 2013;Korsmeyer, 1999). ...
Establishing the intervention to enhance proliferation and differentiation potential is crucial for the clinical translation of stem cell-based therapy. In this study, the effects of simvastatin on these regards were explored. Canine bone marrow-derived mesenchymal stem cells (cBM-MSCs) were treated with 4 doses of simvastatin, 0.1, 1, 10, and 100 nM. Simvastatin in low-dose range, 0.1 and 1 nM, enhanced dose-dependent cell proliferation at day 5 and 7. Exploration of the mechanisms revealed that simvastatin in low-dose range dose-dependently upregulated sets of cell cycle regulators, Cyclin D1 and Cyclin D2; proliferation marker, Ki-67; and anti-apoptotic gene; Bcl-2. Interestingly, pluripotent markers, Rex1 and Oct4, were dramatically increased upon the low-dose treatment. Contrastingly, treatment with high-dose simvastatin suppressed the expression of those genes. Thus, the results suggested beneficial effects of simvastatin on cBM-MSCs proliferation and expansion. Further study regarding differentiation potential and underlying mechanisms will accelerate the clinical application of the molecule on veterinary stem cell-based therapy. : Cell biology; Cell culture; Cell death; Cytotoxicity; Stem cell research; Regenerative medicine, Canine bone marrow-derived mesenchymal stem cells (cBM-MSCs); Simvastatin; Proliferation; Stemness genes Keywords: Cell biology, Cell culture, Cell death, Cytotoxicity, Stem cell research, Regenerative medicine, Canine bone marrow-derived mesenchymal stem cells (cBM-MSCs), Simvastatin, Proliferation, Stemness genes
... Although autophagy is recognized as a cell survival process that promotes tumor development, it is also utilized as a caspase-independent form of programmed cell death (Lin end Baehrecke, 2015). Bcl-2 has been recognized as a negative regulator of both apoptosis (Thomadaki and Scorilas, 2006) and autophagy (Pattingre et al., 2005). When apoptosis is blocked, various apoptotic stimuli activate autophagy, resulting in the induction of autophagic cell death (Shimizu, 2015). ...
Oleanolic acid (OA) is a natural triterpenoid that possesses numerous beneficial health effects such as antioxidant, anti-inflammatory and anti-apoptotic activities. In this study, we investigated the therapeutic effect of OA (10 and 40 mg/kg) on cisplatin (CP)-induced (13 mg/kg) nephrotoxicity. Treatment with OA 40 mg/kg once daily for 2 days, 48 h after CP-intoxication, ameliorated the increased serum markers and histological features of kidney injury. CP administration increased renal expression of antioxidant and anti-inflammatory markers, which was reduced by OA. The increase in proapoptotic caspase-3 and -9 activations, with concomitant increase in poly (ADP-ribose) polymerase (PARP) cleavage, were dose-dependently inhibited by OA. Treatment with OA also ameliorated microtubule-associated protein 1A/1B-light chain 3B (LC3B)-II and autophagy-related protein (Atg) 5 expression induced by CP. The suppression of CP-induced oxidative stress, apoptosis, autophagy and inflammatory response by OA coincided with the inhibition of extracellular-regulated kinase (ERK) 1/2, signal transducer and activator of transcription (STAT) 3 and nuclear factor-kappa B (NF-κB). Interestingly, OA increased CP cytotoxicity in HeLa cervical cancer cells by inducing cytotoxic autophagy. The chemosensitization of HeLa cells to CP suggests a potential beneficial effect of OA in cervical cancer patients due to reduced CP dosage requirements, which requires further investigation.
... Disorders of apoptosis are related to malignancy. Various molecules, such as most anti-cancer therapeutic agents and chitosan nanoparticles can be used to induce apoptosis in cancer cells [26,27]. In this study, flow cytometry analysis was conducted using Annexin V-FITC/PI staining to investigate the cell apoptosis. ...
Cisplatin is widely used as an anticancer drug in chemotherapy of human cancers. In the field of cancer therapy, nanoparticles modified with biocompatible copolymers are suitable vehicles to effectively deliver smaller doses of hydrophobic drugs such as cisplatin in the body. In this study, we investigated whether cisplatin-loaded iron oxide nanoparticles (IONPs) modified with chitosan can exert cytotoxic effects in the human breast cancer cell line MDA-MB-231. IONPs was synthesized using eucalyptus leaf extract as a reducing and stabilizing agent. MDA-MB-231 cells were treated with different concentrations of cisplatin, cisplatin-IONPs and cisplatin-IONPs-chitosan for 24 h. Apoptosis was confirmed by flow cytometry, whereas The mRNA and protein expression of pro- and anti-apoptotic molecules were measured using Real time RT-PCR and western blotting. Treatment with both cisplatin-IONPs and cisplatin-IONPs-chitosan showed a significantly higher cytotoxic effect in comparison to the free drug alone in MDA-MB-231 cells. The levels of apoptosis in cells treated with a combination of cisplatin-IONPs-chitosan were significantly higher compared with cisplatin-IONPs and cisplatin alone. The results of this study showed that the interaction between cisplatin and iron oxide nanoparticles modified with chitosan could enhance responsiveness to cisplatin in breast cancer cells.
... Which includes a series of biochemical changes such as disruption of the mitochondrial membrane and release of apoptogenic molecules to cytosol, activation of caspase cascade, induction of DNA fragmentation. The members of the Bcl-2 family mainly regulate the intrinsic apoptotic signaling pathway, which comprises of both pro-apoptotic (Bax and Bad) and anti-apoptotic (Bcl-2) proteins (Thomadaki and Scorilas 2006). Balance between these two groups decides the fate of the cells. ...
The novel series of piperidine conjugated benzophenone analogs with amide link 11a–l were synthesized in a multistep process. The structures of these compounds were confirmed by IR, ¹H, ¹³C, NMR, and mass spectra and also by elemental analyses. The newly synthesized molecules were screened for selectivity against cancers of different origin through cell based assay system using B16F10, A375, A549, HepG2, ACHN, and MCF7 cells. The results postulated that compound 11f with two bromo groups at the para position in rings A and E and two methyl groups at ortho position in rings B and D evokes target specific action against melanoma highlighting the importance of substituted groups. Down the line studies further inferred compound 11f evokes the apoptotic cellular event leading to cell death. Investigation of eventual mechanism revealed that compound 11f turned out to be a dual inhibitor of B-cell lymphoma-2 and X-linked inhibitors of apoptosis causing the up regulation of Bax and Bad. Further, the antiproliferative effects were mimicked in murine melanoma with similar mechanisms. Molecular docking experiments further confirmed that compound 11f possessed a superior affinity for of B-cell lymphoma-2 and X-linked inhibitors of apoptosis through strong hydrogen bonds. The study implies the identification of compound 11f with selective target against melanoma by inducing apoptogenic effect, which could be the future hope for the treatment of skin cancer.
... Bcl-2 is an anti-apoptosis regulator to keep apoptogenic factors sequestered in the mitochondria. In contrast, Bax can induce the release of apoptogenic factors, and thus can be pro-apoptotic [49]. Alcohol exposure increased Bax expression but inhibited the expression of Bcl-2. ...
Natural polysaccharides, particularly galactomannans, are potential candidates for treatment of alcoholic liver diseases (ALD). However, applications are restricted due to the physicochemical properties associated with the high molecular weight. In this work, guar gum galactomannans were partially hydrolyzed by β-mannanase, and the molecular mechanisms of hepatoprotective effects were elucidated both in vitro and in vivo. Release of lactate dehydrogenase and cytochrome C were attenuated by partially hydrolyzed guar gum (PHGG) in HepG2 cells, due to protected cell and mitochondrial membrane integrity. PHGG co-administration decreased serum amino transaminases and cholinesterase levels of acute alcohol intoxicated mice, while hepatic pathologic morphology was depleted. Activity of superoxide dismutase, catalase, and glutathione peroxidase was recovered to 198.2, 34.5, 236.0 U/mg protein, respectively, while malondialdehyde level was decreased by 76.3% (PHGG, 1000 mg/kg∙day). Co-administration of PHGG induced a 4.4-fold increment of p-AMPK expression, and lipid metabolism was mediated. PHGG alleviated toll-like-receptor-4-mediated inflammation via the signaling cascade of MyD88 and IκBα, decreasing cytokine production. Moreover, mediated expression of Bcl-2 and Bax was responsible for inhibited acute alcohol-induced apoptosis with suppressed cleavage of caspase 3 and PARP. Findings gained suggest that PHGG can be used as functional food supplement for the treatment of acute alcohol-induced liver injury.
... Various factors such as Bcl-2 family members play a major role in the intrinsic mitochondrial apoptotic cascade [23]. The members of Bcl-2 family are divided in two main groups, proapoptotic (Bax, Bad, Bak, Bim, Bid, Bik, Noxa, Puma) and antiapoptotic (Bcl-2, Bcl-x L , Bcl-w, Mcl-1), which are characterized by sharing at least one region of Bcl-2 homology sequence termed BH1-BH4 [24,25]. The over expression of survival proteins not only contributes to the progression of cancer, but also confers resistance to the therapeutic treatments [25]. ...
... Similar to p53, Bcl-2, it reg-ulates some important cell cycle events such as different types of apoptosis and necrosis. The mutated Bcl-2 is seen in many types of cancers, such as colorectal cancer (5,6). ...
Background: Colorectal cancer is one of the most common causes of mortality in the world.Objectives: The aim of this study was to investigate the histopathologic changes including hyperchromatism, tissue lymphocyteinfiltrations (TILs), aberrant crypt foci (ACF), microvessel density (MVD), p53, Bcl-2 and CD31 changes during colorectal cancer development.Methods: Subcutaneous injections of dimethyl hydrazine DMH were administered to rats (40 mg/kg body weight) for 10 weeks.Rats were fed by food and water until 40th week and sacrificed two by two within 10, 15, 20, 25, 30 and 40 weeks after the start oftreatment. Thin paraffinized sections were applied to anti-CD31, anti-Bcl-2 and anti-p53 staining procedures. MVD and ACF werereported as mean value of three HPFs.Results: Hyperchromatism, TILs and angiogenesis were the most common initial histologic changes which started at 10th week ofDMH treatment. Hyperchromatism’s severity increased earlier than other changes and reached the highest value at the 25th week.The highest value of all variants occurred in the 40th week except the TILs which started to achieve the highest value in week 30 andincreased until 40th week. A diminished amount of p53 was observed at week 40, however, increased intensity of CD31 and Bcl-2were seen between 30th and 40th week.Conclusions: In conclusion, TILs and angiogenesis might be the important earliest factors contributing to colorectal cancer progression.
... Various factors such as Bcl-2 family members play a major role in the intrinsic mitochondrial apoptotic cascade [23]. The members of Bcl-2 family are divided in two main groups, proapoptotic (Bax, Bad, Bak, Bim, Bid, Bik, Noxa, Puma) and antiapoptotic (Bcl-2, Bcl-x L , Bcl-w, Mcl-1), which are characterized by sharing at least one region of Bcl-2 homology sequence termed BH1-BH4 [24,25]. The over expression of survival proteins not only contributes to the progression of cancer, but also confers resistance to the therapeutic treatments [25]. ...
Alpha-lipoic acid (ALA), a naturally-occurring antioxidant, inhibits proliferation and induces apoptosis in various cancer cell lines without effects on normal non-transformed cells. The aim of this study was to examine the effects of alpha-lipoic acid (ALA), alone and combined with 5-fluorouracil (5-FU), on Bcl-2/Bax expression in human colon cancer Caco-2 cell line as well as to investigate possible molecular mechanisms and pathways involved in ALA-mediated effects. In the present study ALA and 5-FU showed a tendency to decrease Bcl-2 and increase Bax expression. ALA exerted higher inhibitory effects on Bcl-2 expression, while the significant increase of Bax expression was shown after the treatment with the combination of ALA and 5-FU. The binding modes of ALA and 5-FU with both targets were shown to be closely similar, and some interactions the same like those of known BH3 mimetics. Thus, ALA may be considered as potential BH3 mimetic. Additionally, with in silico calculated physico-chemical properties taken into account, it was confirmed that ALA may easily be delivered to its intracellular and membrane targets.
The use of anti-oxidants could thus prove an effective medication to prevent or facilitate recovery from oxidative stress-induced sensorineural hearing loss (SNHL). One promising strategy to prevent SNHL is developing probucol (PB)-based nanoparticles using encapsulation technology and administering them to the inner ear via the established intratympanic route. The preclinical, clinical and epidemiological studies support that probucol is a proven anti-oxidant that could effectively prevent oxidative stress in different study models. Such findings suggest its applicability in preventing oxidative stress within the inner ear and its associated neural cells. However, several hurdles, such as overcoming the blood-labyrinth barrier, ensuring sustained release, minimizing systemic side effects, and optimizing targeted delivery in the intricate inner ear structures, must be overcome to efficiently deliver PB to the inner ear. This review explores the background and pathogenesis of hearing loss, the potential of PB in treating oxidative stress and its cellular mechanisms, and the obstacles linked to inner ear drug delivery for effectively introducing PB to the inner ear.
Apoptosis, also known as the programmed death of cells, is responsible for maintaining the homeostasis of tissues, and this function is carried out by caspases. The process of apoptosis is carried out via two distinct pathways: the extrinsic pathway, which is governed by death receptors, and the intrinsic pathway, also known as the mitochondrial pathway. The BCL-2 protein family encoded by the BCL-2 gene, located at the 18q21.33 chromosomal location, is in charge of regulating the intrinsic pathway, which is responsible for inducing cell death via the permeabilization of the mitochondrial membrane and the release of apoptosis-inducing components. The BCL-2 homology (BH1, BH2, BH3, BH4) domains of this family proteins are crucial for their functioning, and their common BH domains allow interactions between members of the same family and can also serve as indications of pro- or anti-apoptotic activity. A direct correlation may be shown between the overexpression of BCL-2 and the postponement of cell death. It has been determined that a change in the expression of BCL-2 is the root cause of a variety of malignancies, including lung, breast, melanoma, and chronic lymphocytic leukemia, multiple sclerosis, diabetes. In this review, we addressed the genetic information and structural homology of BCL-2 family members. Further, we elucidate the pro-apoptotic and anti-apoptotic roles of the family members. This review highlights the most recent developments in the BCL-2 protein family and presents evidence that targeting this family proteins may have a positive impact on the treatment of medical problems that are still underserved.
Background:
One of the most important research activities around the world is the screening of various plant components for novel anticancer medicines. The anticancer activities of Aconitum heterophyllum were studied in human breast cancer MDA-MB-231 cells in this study. Since tumorigenesis is thought to be the result of a series of progressive gene alterations, including oncogene activation and tumour suppressor gene inactivation, the expression of genes like p53, p21, STAT, and Bcl-2, which are thought to be important in tumorigenesis and cell death, was determined. In the present study there was an upregulation in the level expression of p53and p21 and down regulation in the expression of BCL2 and STAT. However, there is increase and decrease level of gene expression in Aconitum heterophyllum roots loaded Phyto-Niosomes (nEEAH), when compared to ethanolic root extract of Aconitum heterophyllum EEAH extract treated MDA-MB-231 cell lines.
Methods:
The enzymatic antioxidants such as CAT, SOD, GR, GST, and GPX as well as non-enzymatic antioxidants such as glutathione, Vitamin E and Vitamin C were estimated in the treated MDA-MB-231 cells at the end of incubation. The RT-PCR technique was performed to study the expression patterns of apoptotic genes such as p53 and p21 and anti-apoptotic genes BCL2 and STAT in the drug treated MDA-MB-231 cells.
Results:
In the present study there was a significant (p<0.05) increase in CAT and glutathione levels and a decrease in Vit C, Vit E and SOD, GR, GST, GPX levels in the untreated MDA-MB-231 cells. Increased apoptotic gene expression and decreased anti-apoptotic gene expression suggest the anti-proliferative nature of the drug extract was comparable to the doxorubicin the positive drug used in the present study.
Conclusion:
It can be concluded that the ethanolic extract of Aconitum heterophyllum roots loaded Phyto-Niosomes (nEEAH), when compared to ethanolic root extract of Aconitum heterophyllum EEAH extract treated MDA-MB-231 cell lines exert its anti-cancer activity by activating the apoptotic genes, suppressing anti-apoptotic genes as well as modulating the antioxidant enzymes.
Cancer is the name given to all malignant tumors, the main reason for which is uncontrolled growth, and the tumor, which has become a mass as a result of uncontrolled cell proliferation, also attacks the surrounding cells and envelops the whole body (metastasis) in the later stages of the disease. Although cancer is an important health problem, it is not a common disease in childhood. On the other hand, statistics show that cancer affects one in three adults, causes up to 20% of all deaths, and covers about 10% of treatment costs in developed countries. Although it is known that cancer develops under the influence of genetic and environmental factors, environmental factors are more prominent in the formation of some types of cancer. Breast cancer is one of the cancer types known to have tumor suppressor genes in its etiology. These tumor suppressor genes are BRCA1 and BRCA2 genes. Studies have shown that these two genes are particularly effective in the development of familial breast cancers. These types of cancers occur much earlier than non-familial cancers. The research, two genes; It has shown that it is especially effective in the development of familial breast cancers.
Bladder cancer (BC) is a primary source of malignancy-associated death, and the mortality rate is high due to its prevalence of metastasis. Corilagin (CLG), a bioactive constituent of numerous medicinal plants, exerts assorted pharmacological actions comprising anti-cancer, apoptotic, anti-inflammatory, and hepatoprotective. CLG possesses a substantial anti-tumor prospective and less noxiousness in normal cells in vitro. However, the molecular mechanisms of CLG on BC cells are not studied well. The current research explored the molecular process intricate in the anticancer and anti-proliferative actions of CLG on the relocation of BC cells T24 and TSGH 8301. The cytotoxicity, apoptosis, adhesion, and migration of CLG on BC cells T24 and TSGH 8301 were evaluated by MTT assay, DAPI, Rh-123, cell adhesion, and cell migration assay. The results point out that CLG inhibits the viability, adhesion, movement, incursion, and inflammation, whereas persuades BC cells apoptosis in a concentration-dependent mode. Besides, CLG treated with T24 and TSGH-8301 cells subdue inflammatory and PI3K/Akt signaling pathways. CLG is accomplished of impeding BC cell migration, invasion, and metastasis through the repression of the NF-κB mediated P13K/Akt signaling. Our findings offer a unique vision into the demonstration of the anti-cancer potential of CLG on BC cells.
Clostridium perfringens (C. perfringens) type C (CPC) is one of the chief pathogens that causes diarrhea in piglets, and C. perfringens beta2 (CPB2) toxin is the main virulence factor of CPC. Our previous research demonstrated that ssc-microR-132 was differentially expressed in ileal tissues of CPC-mediated diarrheic piglets and healthy piglets, which implied a potential role of ssc-microR-132 in this process. Here, we found that ssc-microR-132 was notably down-regulated in CPB2-exposed intestinal porcine epithelial cells (IPEC-J2), which was consistent with the ileal tissue expression. Moreover, ssc-microR-132 upregulation alleviated CPB2-induced inflammatory damage and apoptosis in IPEC-J2, whereas ssc-microR-132 knockdown presented the opposite effects. Furthermore, the dual-luciferase reporter assay indicated that ssc-microR-132 directly targeted Dachshund homolog 1 (DACH1). Moreover, DACH1 overexpression intensified CPB2-induced inflammatory injury and apoptosis in IPEC-J2. Remarkably, the introduction of DACH1 weakened the anti-inflammatory and anti-apoptotic effects of ssc-microR-132 in CPB2-exposed IPEC-J2. Overall, the results reveal that ssc-microR-132 targeted DACH1 to alleviate CPB2-mediated inflammation and apoptosis in IPEC-J2.
Background:
Periodontitis is an inflammatory disease initiated by dental deposits. Microorganisms in the dental biofilm induce cell death in epithelial cells, contributing to the breakdown of epithelial barrier function. Recently, dental calculus has also been implicated in pyroptotic cell death in oral epithelium. We analyzed the cytotoxic effects of dental calculus and freeze-dried periodontopathic bacteria on oral epithelial cells and macrophages.
Methods:
HSC-2 human oral squamous carcinoma cells and PMA-differentiated THP-1 macrophages were exposed to dental calculus or one of two species of freeze-dried bacterium, Aggregatibacter actinomycetemcomitans and Fusobacterium nucleatum. Following incubation for 24 h, we measured cytotoxicity via lactate dehydrogenase release. Cells were then incubated with glyburide, an NLRP3 inflammasome inhibitor, to assess the potential role of pyroptosis. We also conducted a permeability assay to analyze the effects on epithelial barrier function.
Results:
Dental calculus induced dose-dependent cell death in HSC-2 cells, whereas cell death induced by freeze-dried bacteria was insignificant. Conversely, freeze-dried bacteria induced more cell death than dental calculus in THP-1 macrophages. Cell death induced by dental calculus but not by freeze-dried bacteria was inhibited by glyburide, indicating that these are different types of cell death. In the permeability assays, dental calculus but not freeze-dried bacteria attenuated the barrier function of HSC-2 cell monolayers.
Conclusion:
Due to the low sensitivity of HSC-2 cells to microbial cytotoxicity, dental calculus had stronger cytotoxic effects on HSC-2 cell monolayers than freeze-dried A. actinomycetemcomitans and F. nucleatum, suggesting that it plays a critical role in the breakdown of crevicular/pocket epithelium integrity. This article is protected by copyright. All rights reserved.
Ochratoxin A (OTA) is a fungal toxin that causes serious threat to human health. OTA could lead to the injury of various tissues, especially kidney injury. However, the toxic effects of OTA on human kidney tubular epithelial cell (HK-2) and the possible mechanism remains poorly understood. This study was to investigate the toxic effects of OTA on HK-2 and elucidate the molecular mechanism. HK-2 cells were treated OTA to evaluate the effect of OTA on cell viability and apoptosis. OTA inhibited the growth of HK-2 in a concentration-dependent manner. With the concentration increased, OTA significantly lead to the apoptosis of HK-2. OTA could increase the levels of reactive oxygen species (ROS) and Malondialdehyde (MDA). Superoxide dismutase (SOD) and glutathione (GSH) activities were decreased by OTA. Furthermore, OTA increased Caspase-3 and Bax expression and decreased BCL2 expression. Compared to the control group, the expression of PTEN was increased and the expression of PI3K and AKT were decreased in OTA treated groups. In addition, we found OTA could disrupt the formation of lipid raft by attenuating sphingomyelin and cholesterol levels. In conclusion, our results indicated that OTA induces apoptosis in HK-2 through regulating PTEN/AKT signaling pathway via disrupting lipid raft formation.
Aims
Several studies suggested that high expression of Bcl2 and/or p53 in Hodgkin/Reed-Sternberg cells is an unfavorable prognostic factor in Hodgkin lymphoma (HL). However, results in this field appear contrasting.
We aimed to assess the prognostic value of p53 and Bcl2 in HL through a systematic review and meta-analysis.
Methods
Electronic databases were searched from January 2000 to December 2020 for all studies assessing the prognostic value of p53 and Bcl2 in HL. The association of high p53 or Bcl2 expression with overall survival (OS), progression-free survival (PFS) and response to treatment was assessed by using hazard ratio (HR) and odds ratio (OR).
Results
Eighteen studies were included. Bcl2 overexpression was significantly associated with decreased PFS (HR = 2.202; p < 0.0001), while the associations with decreased OS (HR = 1.565; p = 0.257) and refractoriness to treatment (OR = 0.482; p = 0.068) were non-significant.
p53 overexpression was not significantly associated with refractoriness to treatment (OR = 0.904; p = 0.155); the analysis of OS and PFS was not feasible, but published data suggested the absence of a significant association.
Conclusions
In HL, Bcl2 overexpression is associated with decreased PFS, while a significant prognostic value could not be demonstrated for p53. Defining optimal criteria for interpreting Bcl2 and p53 immunostaining is necessary to draw definitive conclusions.
Nanoparticle‐based nucleic acid conjugates (NP‐NACs) hold great promise for theragnostic applications. However, several limitations have hindered the realization of their full potential in the clinical treatment of cancer and other diseases. In diagnoses, NP‐NACs suffer from low signal‐to‐noise ratios, while the efficiency of NP‐NACs‐mediated cancer therapies has been limited by the adaptation of alternative prosurvival pathways in cancer cells. The recent emergence of personalized and precision medicine has outlined the importance of having both accurate diagnosis and efficient therapeutics in a single platform. As such, the controlled assembly of hybrid graphene oxide/gold nanoparticle (Au@GO NP)‐based cancer‐specific NACs (Au@GO NP‐NACs) for multimodal imaging and combined therapeutics is reported. The developed Au@GO NP‐NACs show excellent surface‐enhanced Raman scattering (SERS)‐mediated live‐cell cancer detection and multimodal synergistic cancer therapy through the use of photothermal, genetic, and chemotherapeutic strategies. Synergistic and selective killing of cancer cells are then demonstrated using in vitro microfluidic models. Moreover, with the distinctive advantages of the Au@GO NP‐NACs for cancer theragnostics, precision cancer treatment through the detection of cancer cells in vivo using SERS followed by efficient ablation of tumors is shown. Therefore, the Au@GO NP‐NACs can pave a new road for advanced disease theragnostics.
Age-related cataract remains a serious problem in the aged all over the world. MicroRNAs are abnormally expressed in various diseases including age-related cataract. MicroRNA-15a (MicroRNA-15a) has been involved in various diseases and plays crucial roles in many cellular processes. However, the mechanism of MicroRNA-15a in the genesis of cataract remains barely known. We therefore aimed to investigate the role of MicroRNA-15a in the cataract. Herein, human lens epithelial cells, HLE-B3 cells were treated with 200μmol/L H2O2 for 24h. H2O2 was utilized in our study to induce HLE-B3 cells injury. We observed that cell apoptosis was induced by the treatment of H2O2 and meanwhile, cell proliferation was repressed by 200μmol/L H2O2. Then, it was found that microRNA-15a was significantly increased with the H2O2 exposure in vitro. Importantly, B-cell lymphoma-2 (BCL2) and E2F transcription factor 3 (E2F3) exert crucial roles in cell apoptosis and cell proliferation. We found that BCL2 and E2F3 were greatly reduced by 200μmol/L H2O2 inhuman lens epithelial cells. In addition, microRNA-15a overexpression induced cell apoptosis and repressed cell proliferation through suppressing BCL2 and E2F3. Subsequently, BCL2 and E2F3 were predicted as a direct target of MicroRNA-15a. The direct correlation between microRNA-15a and BCL2/E2F3 was confirmed by dual luciferase reporter assay. In conclusion, we demonstrated that microRNA-15a triggered apoptosis and repressed the proliferation of HLE-B3 cells by modulating BCL2 and E2F3.
Genetic analysis of programmed cell death inDrosophila reveals many similarities with mammals. Heretofore, a missing link in the fly has been the absence of any Bcl-2/Bax family members, proteins that function in mammals as regulators of mitochondrial cytochrome c release. ADrosophila homologue of the human killer protein Bok (DBok) was identified. The predicted structure of DBok is similar to pore-forming Bcl-2/Bax family members. DBok induces apoptosis in insect and human cells, which is suppressible by anti-apoptotic human Bcl-2 family proteins. A caspase inhibitor suppressed DBok-induced apoptosis but did not prevent DBok-induced cell death. Moreover, DBok targets mitochondria and triggers cytochrome c release through a caspase-independent mechanism. These characteristics of DBok reveal evolutionary conservation of cell death mechanisms in flies and humans.
We identified and cloned a novel murine member of the pro-apoptotic Bcl-2 family. This protein, designated Blk, is structurally and functionally related to human Bik and localized to the mitochondrial membrane. Blk contains a conserved BH3 domain and can interact with the anti-apoptotic proteins Bcl-2 and Bcl-xL. Ectopic expression of Blk in mammalian cells induces apoptosis, which can be inhibited by mutations in the BH3 domain and by overexpression of Bcl-2 or Bcl-xL but not by CrmA. The apoptotic activity of Blk is also inhibited by a dominant negative caspase-9, suggesting that Blk induces apoptosis through activation of the cytochromec-Apaf-1-caspase-9 pathway.
Bcl-2-related anti- and proapoptotic proteins are important in the decision step of the intracellular death program upstream from the caspase proteases. Targeted overexpression of Bcl-2 in ovarian so- matic cells of transgenic mice leads to decreased apoptosis of gran- ulosa cells and is associated with higher ovulation rate, increased litter size, and ovarian teratoma formation. The ability of exogenous Bcl-2 proteins to promote follicle cell survival suggests that the trans- gene can bind to endogenous ovarian Bcl-2 family members and mod- ulate the intracellular apoptosis process in favor of cell survival. We used the yeast two-hybrid system to search for ovarian Bcl-2 inter- acting proteins. The screening of an ovarian fusion complementary DNA library yielded several clones encoding for the death agonist Bcl-XL/Bcl-2-associated death promoter (BAD). Dimerization of Bcl- 2-related proteins mediated by the consensus Bcl-2 homology (BH) domains is essential for their apoptosis-regulating function. Consis- tent with these observations, yeast two-hybrid assays indicated that the interaction of Bcl-2 with BAD is dependent on both BH4 and BH2 domains of Bcl-2. Northern blot analysis showed a wide distribution of BAD messenger RNA (mRNA) in diverse tissues with highest levels in the lung, ovary, uterus, and brain. In situ hybridization analysis indicated BAD mRNA expression in granulosa cells of different sizes of follicles and also in the theca and interstitial cells. BAD mRNA was expressed in the ovaries between postnatal 15-27 days and did not alter during the developmentally occurring apoptosis found about postnatal day 18 when the first group of early antral follicles were formed. Similarly, BAD mRNA levels did not change during follicle atresia induced by estrogen withdrawal in immature rats. To study the role of BAD in the ovary, BAD complementary DNA was trans- fected into primary cultures of granulosa cells and in a gonadal tumor cell line. Overexpression of BAD induced apoptosis in both cell types, and the effect of BAD was reversed by a membrane-permeable caspase inhibitor, indicating that BAD induces apoptosis via the ac- tivation of caspase cysteine proteases. In summary, the death agonist BAD was identified as a Bcl-2-interacting protein in the ovary, and BAD mRNA is constitutively expressed in granulosa cells, suggesting that BAD is an essential part of the ovarian cell death process. Be- cause BAD overexpression in granulosa cells leads to apoptosis, fu- ture studies on ovarian BAD binding proteins and hormonal regula- tion of the interactions among different Bcl-2 family members could provide a better understanding of the cellular mechanism of ovarian follicle atresia. (Endocrinology 138: 5497-5504, 1997)
The BCL-2 family of proteins is composed of both pro- and antiapoptotic regulators, although its most critical biochemical
functions remain uncertain. The structural similarity between the BCL-XL monomer and several ion-pore-forming bacterial toxins has prompted electrophysiologic studies. Both BAX and BCL-2 insert
into KCl-loaded vesicles in a pH-dependent fashion and demonstrate macroscopic ion efflux. Release is maximum at ≈pH 4.0 for
both proteins; however, BAX demonstrates a broader pH range of activity. Both purified proteins also insert into planar lipid
bilayers at pH 4.0. Single-channel recordings revealed a minimal channel conductance for BAX of 22 pS that evolved to channel
currents with at least three subconductance levels. The final, apparently stable BAX channel had a conductance of 0.731 nS
at pH 4.0 that changed to 0.329 nS when shifted to pH 7.0 but remained mildly Cl− selective and predominantly open. When BAX-incorporated lipid vesicles were fused to planar lipid bilayers at pH 7.0, a Cl−-selective (PK/PCl = 0.3) 1.5-nS channel displaying mild inward rectification was noted. In contrast, BCL-2 formed mildly K+-selective (PK/PCl = 3.9) channels with a most prominent initial conductance of 80 pS that increased to 1.90 nS. Fusion of BCL-2-incorporated
lipid vesicles into planar bilayers at pH 7.0 also revealed mild K+ selectivity (PK/PCl = 2.4) with a maximum conductance of 1.08 nS. BAX and BCL-2 each form channels in artificial membranes that have distinct
characteristics including ion selectivity, conductance, voltage dependence, and rectification. Thus, one role of these molecules
may include pore activity at selected membrane sites.
A critical function of tumor suppressor p53 is the induction of apoptosis in cells exposed to noxious stresses. We report
a previously unidentified pro-apoptotic gene, Noxa. Expression of Noxa induction in primary mouse cells exposed to x-ray irradiation was dependent on p53. Noxa encodes a Bcl-2 homology 3 (BH3)–only member of the Bcl-2 family of proteins; this member contains the BH3 region but not
other BH domains. When ectopically expressed, Noxa underwent BH3 motif–dependent localization to mitochondria and interacted
with anti-apoptotic Bcl-2 family members, resulting in the activation of caspase-9. We also demonstrate that blocking the
endogenous Noxa induction results in the suppression of apoptosis. Noxa may thus represent a mediator of p53-dependent apoptosis.
The tumour-suppressor gene p53 is frequently mutated in human cancers and is important in the cellular response to DNA damage. Although the p53 family members p63 and p73 are structurally related to p53, they have not been directly linked to tumour suppression, although they have been implicated in apoptosis. Given the similarity between this family of genes and the ability of p63 and p73 to transactivate p53 target genes, we explore here their role in DNA damage-induced apoptosis. Mouse embryo fibroblasts deficient for one or a combination of p53 family members were sensitized to undergo apoptosis through the expression of the adenovirus E1A oncogene. While using the E1A system facilitated our ability to perform biochemical analyses, we also examined the functions of p63 and p73 using an in vivo system in which apoptosis has been shown to be dependent on p53. Using both systems, we show here that the combined loss of p63 and p73 results in the failure of cells containing functional p53 to undergo apoptosis in response to DNA damage.
Cubital tunnel surgery should be considered a failure if patients have no relief of their symptoms or if the symptoms recur shortly after the surgery. Choice of treatment should be based on careful examination and evaluation of patient expectations, general medical condition, level of activity, and duration and severity of symptoms. Failure of the initial procedure may be the result of inadequate release, instability, subluxation, inadvertent creation of a new site of compression, and intraoperative nerve injury.
Certain clinical manifestations specify the cause for failure. A positive Tinel’s sign, for example, may indicate the exact location of persistent nerve compression. A palpable mobile mass on the medial aspect of the elbow is consistent with recurrent subluxation. Localized point tenderness along the course of the incision may indicate a neuroma secondary to injury to the medial antebrachial cutaneous nerve. In all cases, it is imperative that the surgeon be familiar with all the possible anatomic sources of compression as well as the variations in the ulnar nerve and the medial antebrachial cutaneous nerve. Once operative failure has been determined, efforts should be directed at completely releasing the nerve through external neurolysis, eliminating any mechanical stretch, and releasing any sites of compression or kinking. Some improvement should be expected if the surgeon thoroughly understands the anatomy, chooses the appropriate revision technique based on patient history, and adheres to the technical details of the chosen revision technique.
Cervical carcinomas develop as a result of multiple genetic alterations. As the genetic alterations are the cause of malignant transformation, it is likely that specific genetic alterations lead to specific clinical behaviour. The aim of this study was (i) to localise chromosome arms that harbour likely tumour‐suppressor genes, by analysing loss of heterozygosity (LOH) and (ii) to study the association of LOH with clinicopathological parameters. To define the regions of interest, we studied the presence of loss of heterozygosity at all chromosomes in 67 cervical carcinomas (stages IB and IIA) with 81 polymorphic markers. In addition, all frequent allelic imbalances were correlated with HPV status and clinicopathologic parameters including survival, FIGO‐stage, lymph‐node metastasis, tumour size, number of mitoses, vaso‐invasion and histologic type. LOH at a frequency over 25% was observed at sites on 9 chromosome arms: 3p21, 4p16.1‐15, 6p, 6q22.3‐23.1, 11q22‐24, 15q11‐21.1, 17p13.3, 18q22‐qter and Xq. LOH of chromosome 6q14‐16.2, 6p22 and 17p13 correlated marginally with HPV‐16 positivity. LOH on chromosome 3p21 was weakly correlated with high mitotic activity, while LOH on chromosomes 11q23.3, 15q21.1 and 17p13 correlated with low mitotic activity. LOH at chromosome 17p13 associated marginally with FIGO stage I, while LOH at chromosome 15q associated weakly with FIGO stage II. When chromosome 18q showed LOH in the tumour, the patients had decreased survival (p = 0.024). We conclude that, in carcinoma of the uterine cervix, a novel tumour‐suppressor gene may be present on chromosome 15q21 and that patients with LOH on chromosome 18q have relatively poor survival (p = 0.025). Int. J. Cancer (Pred. Oncol.) 79:411–417, 1998. © 1998 Wiley‐Liss, Inc.
Background
Tumor cell heterogeneity and resistance to chemotherapy‐mediated cell death are major obstacles in cancer therapy. It has been reported that expression of the pro‐apoptotic molecule Bax can induce cell death or sensitize tumor cells to chemotherapy in stable cell clones derived from tumor cells. However, these studies are limited in that they cannot represent the heterogeneity of cancer cells observed in vivo. In this study, we have further explored the therapeutic potential of Bax.
Methods
Using an inducible recombinant Bax adenovirus, we screened a panel of ovarian cancer cell lines and primary patient‐derived ovarian tumor cells for their sensitivity to Bax‐mediated cytotoxicity. Apoptotic cell death was evaluated qualitatively with Hoechst staining and quantitatively with MTS and Annexin V‐based assays. Endogenous levels of both Bcl‐2 and Bax protein and p53 status were evaluated. The potential of bax to sensitize ovarian cancer lines to chemotherapy was also tested. Dose–response curves were generated to evaluate cell death.
Results
Overexpression of Bax directly induced apoptosis in both ovarian cancer cell lines and the patient‐derived primary cancer cells. However, the sensitivity of these cells to Bax varied and appeared to be independent of both the status of p53 and the endogenous levels of bcl‐2 or Bax, critical molecules in the apoptotic pathway. Importantly, overexpression of Bax significantly enhanced chemotherapy‐induced cytotoxicity in both established cell lines and primary ovarian carcinoma cells.
Conclusions
These studies suggest that overexpression of Bax alone or in combination with chemotherapy may provide a means to overcome the problems imposed by the heterogeneous nature of tumors, ultimately augmenting the efficacy of chemotherapy in patients suffering from ovarian cancer. Copyright © 2000 John Wiley & Sons, Ltd.
We have previously demonstrated that CD4 cross-linking (CD4XL) results in apoptosis of CD4+ T cells and augmentation of Fas antigen (CD95, APO-1) expression in CD4+ and CD8+ T cells. Here we demonstrate that CD4XL mediated by both, anti-CD4 monoclonal antibody (MoAb) or human immunodeficiency virus (HIV) envelope protein gp120 reduces the expression of the proto-oncogene Bcl-2 in CD4+ T cells, but not in CD8+ T cells, concurrently with the induction of CD4+ T cell-apoptosis. Additionally, the Bcl-2dim population expressed high levels of Fas antigen. Bax, an antagonist of Bcl-2, was brightly expressed even in the Bcl-2dim population. Addition of interleukin (IL)-2 rescued CD4+ T cells from CD4XL-induced Bcl-2 downmodulation and apoptosis induction. These results support the hypothesis that CD4 ligation by HIV-1 envelope protein in vivo in HIV-infected patients selectively reduces Bcl-2 protein in CD4+ T lymphocytes, thereby facilitating Fas/Fas-ligand triggered apoptosis; furthermore the findings reported expand the rationale for use of IL-2 in HIV disease.
Human T lymphotropic virus type I (HTLV-I) is the etiological agent of adult T-cell lymphocytic leukemia (ATLL), whereas HTLV-II has not been associated with hematopoietic malignancies. The control of apoptotic pathways has emerged as a critical step in the development of many cancer types. As a result, the underlying mechanism of long-term survival of HTLV-I and HTLV-II was studied in infected T cells in vitro and in ex vivo ATLL samples. Results indicate that HTLV-I– and HTLV-II–infected T cells in vitro express high levels of the antiapoptotic protein Bcl compared with other human leukemic T cell lines or uninfected peripheral blood mononuclear cells. The levels of proapoptotic proteins Bax, BAD, and Bak were not significantly altered. HTLV-I and HTLV-II viral transactivators, Tax1 and Tax2, are known to increase expression of cellular genes. These proteins were tested for increased transcription from the human Bcl2 and Bcl-XL promoters. Whereas no effect was observed on the Bcl2 promoter, both Tax1 and Tax2 increased transcription of the Bcl-XL promoter in T cells, although Tax1 appeared to be more efficient than Tax2. The biological significance of these observations was validated by the finding of an increased expression of Bcl-XL in ex vivo ATLL cells, especially from patients unresponsive to various chemotherapy regimens. Altogether, these data suggest that overexpression of Bcl-XL in vivomay be in part responsible for the resistance of ATLL cells to chemotherapy. In addition, inefficient activation of the Bcl-XL promoter by Tax2 may result in a shorter survival time of HTLV-II–infected cells in vivo and a diminished risk of leukemia development.
Mcl-1 is a member of the Bcl-2 family that is expressed in early monocyte differentiation and that can promote viability on transfection into immature myeloid cells. However, the effects of Mcl-1 are generally short lived compared with those of Bcl-2 and are not obvious in some transfectants. To further explore the effects of this gene, mice were produced that expressed Mcl-1 as a transgene in hematolymphoid tissues. The Mcl-1 transgene was found to cause moderate viability enhancement in a wide range of hematopoietic cell types, including lymphoid (B and T) as well as myeloid cells at both immature and mature stages of differentiation. However, enhanced hematopoietic capacity in transgenic bone marrow and spleen was not reflected in any change in pool sizes in the peripheral blood. In addition, among transgenic cells, mature T cells remained long lived compared with B cells and macrophages could live longer than either of these. Interestingly, when hematopoietic cells were maintained in tissue culture in the presence of interleukin-3, Mcl-1 enhanced the probability of outgrowth of continuously proliferating myeloid cell lines. Thus, Mcl-1 transgenic cells remained subject to normal in vivo homeostatic mechanisms controlling viable cell number, but these constraints could be overridden under specific conditions in vitro. Within the organism, Bcl-2 family members may act at “viability gates” along the differentiation continuum, functioning as part of a system for controlled hematopoietic cell amplification. Enforced expression of even a moderate viability-promoting member of this family such as Mcl-1, within a conducive intra- and extracellular environment in isolation from normal homeostatic constraints, can substantially increase the probability of cell immortalization.
© 1998 by The American Society of Hematology.
Human neutrophils possess a very short half-life because they constitutively undergo apoptosis. Cytokines, such as granulocyte-macrophage colony-stimulating factor (GM-CSF), and other agents can rescue neutrophils from apoptosis but the molecular mechanisms involved in this rescue are undefined. Here, we show by Western blotting that human neutrophils do not express Bcl-2 or Bcl-X but constitutively express Bax. However, cellular levels of these proteins are unaffected by agents which either accelerate or delay neutrophil apoptosis. In contrast, neutrophils express the antiapoptotic protein Mcl-1 and levels of this protein correlate with neutrophil survival. Thus, cellular levels of Mcl-1 decline as neutrophils undergo apoptosis and are enhanced by agents (eg, GM-CSF, interleukin-1β, sodium butyrate, and lipopolysaccharide) that promote neutrophil survival. Neutrophils only possess few, small mitochondria, and much of the Mcl-1 protein seems to be located in nuclear fractions. These observations provide the first evidence implicating a Bcl-2 family member in the regulation of neutrophil survival. Moreover, this work also provides a potential mechanism whereby cytokine-regulated gene expression regulates the functional lifespan of neutrophils and hence their ability to function for extended time periods during acute inflammation.
The BCL-2 proto-oncogene encodes a mitochondrial protein that blocks programmed cell death. High amounts of bcl-2 protein are found not only in lymphoid malignancies, but also in normal tissues characterized by apoptotic cell death, including bone marrow. Using a monoclonal antibody to bcl-2 protein, we analyzed 82 samples of newly diagnosed acute myeloid leukemia. The number of bcl-2+ cells in each sample was heterogeneous (range, 0% to 95%), with a mean of 23%. The percentage of bcl-2+ cells was higher in M4 and M5 types, according to French- American-British classification, and in cases with high white blood cell counts. bcl-2 expression was also correlated with that of the stem cell marker CD34. In vitro survival of leukemic cells maintained in liquid culture in the absence of growth factors was significantly longer in cases with a high percentage of bcl-2+ cells. High expression of bcl-2 was associated with a low complete remission rate after intensive chemotherapy (29% in cases with 20% or more positive cells v 85% in cases with less than 20% positive cells, P < 10(-5)) and with a significantly shorter survival. In multivariate analysis, the percentage of bcl-2+ cells (or the blast survival in culture), age, and the percentage of CD34+ cells were independently associated with poor survival.
Hrk is a newly described proapoptotic member of the Bcl-2 family that is mainly expressed in hematopoietic tissues and cultured neurons. In this study we have examined the expression and activity of Hrk in hematopoietic progenitors. To address these issues, we used 3 growth factor-dependent murine hematopoietic cell lines, HCD-57, FDCP-Mix, and FL5.12. The expression of Hrk was undetectable in cells cultured with growth factors, but it was rapidly up-regulated on growth factor withdrawal. In contrast, the expression of Bcl-xL decreased and that of proapoptotic Bax, Bad, and Bak was unchanged or down-regulated after removal of growth factors. This pattern of expression correlated with the induction of apoptosis. Hrk was also up-regulated in human cell lines and in bone marrow-derived CD34+ cells cultured in the absence of growth factors. In addition, the levels of Hrk were up-regulated after treatment with the chemotherapeutic drug etoposide. Expression of prosurvival Bcl-xL or Bcl-2 proteins blocked the induction of Hrk. Hrk was induced in FDCP-Mix cells treated with ionomicin in the presence of IL-3, suggesting that cytosolic calcium may regulate the expression of this proapoptotic protein. Furthermore, ectopic expression of Hrk induced cell death of hematopoietic progenitors in the presence of IL-3. Thus, Hrk is specifically and rapidly induced in hematopoietic progenitors after growth factor deprivation or treatment with chemotherapeutic drugs, and this may be sufficient to induce apoptosis in these cells.
The value of various prognostic factors in breast cancer patients has been determined in a number of studies, One hundred thirty-eight Greek women were followed up over a 5-year period after surgery for breast cancer, Amplification and overexpression of c-erbB-2 was found in 22.4% and 29.7% of the respective cases, and the concentration of total cytosolic Cathepsin-D (CD) in 46.4% of them was high (greater than or equal to 60 pmol/mg protein). The examined biological variables were compared with standard clinicopathological prognostic factors for the disease and related to early relapse (ER; before 3 years), relapse-free survival (RFS; median, 5 years), and overall survival (OS; median, 5 years), It was found that high CD levels significantly shorten ER of both node-negative and node-positive patients (P < 0.0001 and P = 0.002, respectively) and have prognostic value for RFS and OS of node-negative patients (P = 0.0012 and P = 0.0288, respectively), but lose their value as relapse predictors for nodepositive patients for periods longer than 3 years, Overexpression of c-erbB-2 was found to be predictive for OS of node-positive and -negative patients (P = 0.0048 and P = 0.0285, respectively), but its predictive power was weak for ER (P = 0.0456) and RFS (P = 0.0455) of node-negative patients and disappeared for node-positive patients. c-erbB-2 amplification offers minimal assistance to the prediction. In conclusion, high CD concentration is indicative of ER of patients, and c-erbB-2 overexpression correlates with OS of patients.
Most members of the Bcl-2 protein family of apoptosis regulating proteins contain two evolutionarily conserved domains, termed BH1 and BH2. Both BH1 and BH2 in the Bcl-2 protein are required for its function as an inhibitor of cell death and for heterodimerization with the proapoptotic protein Bax. In this report, we mapped the region in Bax required for heterodimerization with Bcl-2 and homodimerization with Bax, using yeast two-hybrid and in vitro protein-protein interaction assays. Neither the BH1 nor the BH2 domain of Bax was required for binding to the wild-type Bcl-2 and Bax proteins. Moreover, Bax (Delta BH1) and Bax (Delta BH2) mutant proteins bound efficiently to themselves and each other, further confirming the lack of requirement for BH1 and BH2 for Bax/Bax homodimerization. Bax/Bax homodimerization was not dependent on the inclusion of the NH2-terminal 58 amino acids of the Bax protein in each dimerization partner, unlike Bcl-2/Bcl-2 homodimers which involve head-to-tail interactions between the region of Bcl-2 where BH1 and BH2 resides, and an NH2-terminal domain in Bcl-2 that contains another domain BH4 which is conserved among antiapoptotic members of the Bcl-2 family. Similarly, heterodimerization with Bcl-2 occurred without the NH2-terminal domain of either Bax or Bcl-2, suggesting a tail-to-tail interaction. The essential region in Bax required for both homodimerization with Bax and heterodimerization with Bcl-2 was mapped to residues 59-101. This region in Bax contains a stretch of 15 amino acids that is highly homologous in several members of the Bcl-2 protein family, suggesting the existence of a novel functional domain which we have termed BH3. Deletion of this 15-amino acid region abolished the ability of Bax to dimerize with itself and to heterodimerize with Bcl-2. The findings suggest that the structural features of Bax and Bcl-2 that allow them to participate in homo- and heterodimerization phenomena are markedly different, despite their amino-acid sequence similarity.
Bim is a proapoptotic member of the Bcl-2 family that shares only the BH3 domain with this family. Three Bim proteins Bim-EL, Bim-L and Bim-S are synthesized from the same transcript. We report here that Bim-EL when phosphorylated by Erk1/2 is rapidly degraded via the proteasome pathway. Using different cellular models we evidence that serine 69 is both necessary and sufficient for Erk1/2-mediated phosphorylation and degradation of Bim-EL. In K562 cells, Phorbol 12-myristate 13-acetate activates Erk1/2 and consequently increases Bim-EL phosphorylation and degradation by the proteasome, resulting in cell survival, while the Bcr-Abl inhibitor imatinib abrogates Bim-EL phosphorylation and degradation and induces caspase activation and apoptosis. We also show that Bim-EL(S69G) promotes apoptosis more efficiently than Bim-EL-WT in K562 cells. Altogether, our findings demonstrate that phosphorylation of Bim-EL by Erk1/2 on serine 69 selectively leads to its proteasomal degradation and therefore represents a new and important mechanism of Bim regulation.
From an acute B-cell leukemia cell line, a DNA probe was obtained that was specific for chromosome 18 and flanked the heavy chain joining region of the immunoglobulin heavy chain locus on chromosome 14. This probe detected rearrangement of the homologous DNA segment in the leukemic cells and in follicular lymphoma cells with the t(14:18) chromosome translocation but not in other neoplastic or normal B or T cells. The probe appears to identify bcl-2, a gene locus on chromosome 18 (band q21) that is unrelated to known oncogenes and may be important in the pathogenesis of B-cell neoplasms with this translocation.
Extracellular survival factors alter a cell's susceptibility to apoptosis, often through posttranslational mechanisms. However, no consistent relationship has been established between such survival signals and the BCL-2 family, where the balance of death agonists versus antagonists determines susceptibility. One distant member, BAD, heterodimerizes with BCL-X(L) or BCL-2, neutralizing their protective effect and promoting cell death. In the presence of survival factor IL-3, cells phosphorylated BAD on two serine residues embedded in 14-3-3 consensus binding sites. Only the nonphosphorylated BAD heterodimerized with BCL-X(L) at membrane sites to promote cell death. Phosphorylated BAD was sequestered in the cytosol bound to 14-3-3. Substitution of serine phosphorylation sites further enhanced BAD's death-promoting activity. The rapid phosphorylation of BAD following IL-3 connects a proximal survival signal with the BCL-2 family, modulating this checkpoint for apoptosis.
In order to resolve the mechanisms and reasons of cellular fragmentation it is crucial to understand what genes may be responsible for regulation of this process. We report herein that human oocytes and preimplantation embryos possess abundant levels of transcripts encoding cell death suppressors, Mcl‐1, Bcl‐x and Bag‐1, and the cell death inducer genes, Bax and Caspase‐2. Lower but detectable levels of mRNA expression for the Bfl‐1/a1, Bcl‐w, Harakiri (Hrk) and Caspase‐3 genes were also detected during all developmental stages. We also performed analysis of gene expression in single human embryos exhibiting various degrees of fragmentation at the 2‐, 4‐ and 8‐cell stages. At the 4‐cell stage, embryos displaying 30–50% fragmentation showed a significant increase in Hrk mRNA levels (P = 0.016). Immunostaining with anti‐Hrk antibody confirmed increased staining in some, but not all, fragmented embryos. While Caspase‐3 transcripts were elevated in both 4‐ and 8‐cell embryos exhibiting a severe degree of fragmentation, this difference did not reach statistical significance. However, accumulation of Caspase‐3 mRNA in fragmented embryos was paralleled by an induction of Caspase‐3‐like activity. These findings suggest that cellular fragmentation in a subset of human preimplantation embryos could be regulated by certain components of a genetic programme of cell death.
Most of human follicular lymphomas possess the t(14;18) chromosome translocation that juxtaposes the IgH gene to the 3' region of bcl-2 in a head-to-tail configuration. Here we show that the rearrangement of the bcl-2 gene occurs in a significant fraction (approximately of 10%) of B cell CLL. In all cases analyzed, breakpoints on chromosome 18 clustered at the 5' flanking region of the bcl-2 gene, and no rearrangements were found at the major or minor breakpoint clustering region (3' region of bcl-2 gene) typical of the t(14;18) chromosome translocation. All of the rearranged bcl-2 genes were juxtaposed with the Ig lambda or K genes in a head-to-head configuration. These results imply that the bcl-2 gene is preferentially linked to the IgL genes in CLL and could function in leukemogenesis.
A family of genes related to the bcl-2 protooncogene has recently emerged. One member of this family, mcl-1, was cloned from a human myeloblastic leukemia cell line (ML-1) undergoing differentiation. The intracellular localization of mcl-1, as well as the kinetics of its expression during differentiation, have now been studied. These studies show that the intracellular distribution of mcl-1 overlaps with, but is not identical to, that of bcl-2: mcl-1 is similar to bcl-2 in that the mcl-1 protein has a prominent mitochondrial localization, and in that it associates with membranes through its carboxyl hydrophobic tail. mcl-1 differs from bcl-2, however, in its relative distribution among other (nonmitochondrial/heavy membrane) compartments, mcl-1 also being abundant in the light membrane fraction of immature ML-1 cells while bcl-2 is abundant in the nuclear fraction. Similarly, in differentiating ML-1 cells, the timing of expression of mcl-1 overlaps with, but is not identical to, that of bcl-2: the mcl-1 protein increases rapidly as cells initiate differentiation, and mcl-1 is a labile protein. In contrast, bcl-2 decreases gradually as cells complete differentiation. Overall, the mcl-1 and bcl-2 proteins have some properties in common and others tht are distinct. A burst of expression of mcl-1, prominently associated with mitochondria, complements the continued expression of bcl-2 in ML-1 cells differentiating along the monocyte/macrophage pathway.
Ultraviolet B (UVB) damage is recognized as the most important etiological factor in the development of skin cancer. Human papillomaviruses (HPV) have also been implicated in the disease, although the mechanism of action of these viruses remains unknown. We present evidence here that Bak protein is involved in signaling apoptosis in the skin in response to UVB damage, and that cutaneous HPV E6 proteins target and abrogate Bak function by promoting its proteolytic degradation both in vitro and in regenerated epithelium. Additionally, HPV positive skin cancers had undetectable levels of Bak in contrast to HPV negative cancers, which expressed Bak. This study supports a link between the virus and UVB in the induction of HPV-associated skin cancer and reveals a survival mechanism of virally infected cells.
Keywords
• HPV
• skin cancer
• apoptosis
• UV
• proteolysis
Here we report the genomic cloning and characterization of the murine A1 genes, which belong to the bcl-2 gene family. Southern analysis indicated the existence of at least four A1 genes in the murine genome and four different A1 genes, designated A1-a, -b, -c and -d, were cloned from the murine genomic library. The A1-a, -b and -d genes consisted of two exons, whereas the A1-c gene contained 1 bp insertion in the coding region which may result in an aberrant and truncated protein by frame-shift. With the exception of A1-c, the coding regions among A1 genes are highly conserved at >97% at the nucleotide level and at .96% at the amino acid level. A1-a, -b and -d genes appeared to be expressed specifically in organs containing many neutrophils. In neutrophils, A1-a, -b and -d transcripts were detected at a comparable level. Our data suggest that the multiple A1 genes in mice were generated by gene duplication and each of them may function as antiapoptotic molecules in neutrophils.
The Bcl-2 family consists of about 20 homologues of important pro- and anti-apoptotic regulators of programmed cell death. The established mode of function of the individual members is to either preserve or disturb mitochondrial integrity, thereby inducing or preventing release of apoptogenic factors like Cytochrome c (Cyt c) from mitochondria. Recent findings also indicate further Bcl-2-controlled mitochondria-independent apoptosis pathways. Bcl-2 represents the founding member of the new and growing class of cell death inhibiting oncoproteins. In this review, we try to briefly summarize current models of Bcl-2 family function and to outline the work demonstrating the influence of deregulated Bcl-2 family member expression on tumorigenesis and cancer therapy. Since several Bcl-2 homologues, in addition to influencing apoptotic behaviour, also impinge on cell cycle progression, we discuss possible implications of this additional role for the expression of Bcl-2 family members in tumor cells.
In a cell-free apoptosis system, mitochondria spontaneously released cytochrome c, which activated DEVD-specific caspases, leading to fodrin cleavage and apoptotic nuclear morphology. Bcl-2 acted in situ on mitochondria to prevent the release of cytochrome c and thus caspase activation. During apoptosis in intact cells, cytochrome c translocation was similarly blocked by Bcl-2 but not by a caspase inhibitor, zVAD-fmk. In vitro, exogenous cytochrome c bypassed the inhibitory effect of Bcl-2. Cytochrome c release was unaccompanied by changes in mitochondrial membrane potential. Thus, Bcl-2 acts to inhibit cytochrome c translocation, thereby blocking caspase activation and the apoptotic process.
Members of the BCL2 family of proteins are key regulators of programmed cell death, acting either as apoptotic agonists or antagonists. Here we describe the solution structure of BID, presenting the structure of a proapoptotic BCL2 family member. An analysis of sequence/structure of BCL2 family members allows us to define a structural superfamily, which has implications for general mechanisms for regulating proapoptotic activity. It appears two criteria must be met for proapoptotic function within the BCL2 family: targeting of molecules to intracellular membranes, and exposure of the BH3 death domain. BID’s activity is regulated by a Caspase 8–mediated cleavage event, exposing the BH3 domain and significantly changing the surface charge and hydrophobicity, resulting in a change of cellular localization.
There are at present disparate published results with regard to the relevance of the Bcl-2 gene family, levels of apoptosis, and cell proliferation in the development and progression of renal cell carcinoma (RCC). The present study v analyses the interrelationship between the expression of representatives of the anti-apoptotic (Bcl-2, Bcl-X-L) or pro-apoptotic (Bax) Bcl-2 proteins, incidence of apoptosis, and mitosis in a selected small group of 22 graded RCCs that had paired normal renal tissue, or non-neoplastic tissue in the renal biopsy specimen. The cases were chosen to determine the feasibility of measuring these parameters as potential surrogate markers of progression or treatment failure of the cancers. The results showed that in approximately 50% of the RCCs, where Bcl-2 and/or Bcl-X-L expression was high, apoptosis it-as not detected, and when expression of these proteins was low or not found, increased levels of apoptosis were seen. In most of the remaining 50% of samples, high levels of Bcl-X-L but not Bcl-2 were negatively correlated with low levels of apoptosis (Bcl-X-L: r = -0.437, P = 0.07 and Bcl-2: r = + 0.560, P = 0.02). For the same group of samples, high Bax expression was found in association with apoptosis (r = + 0.578, P = 0,02). A novel finding was an association between low expression of Bcl-2 an/or Bcl-X-L in normal tissue and the level of expression of these proteins in the RCCs, an intrinsic variation that may be an individual patient factor. The results indicate that, in RCCs with increased expression of Bcl-2 and/or Bcl-X-L, levels of apoptosis are minimal and these combined factors may assist in progression of the cancers and resistance to treatments.
The two anti-apoptotic proteins Bcl-2 and Bcl-xL, and the two pro-apoptotic proteins Bax and Bcl-xS were measured by Western blotting in 51 neoplastic and 8 normal endometrial samples. The corresponding mRNA levels were analyzed by semiquantitative reverse transcriptase-polymerase chain reaction in a subgroup of 19 endometrial carcinomas. Neoplastic tissues had higher amounts of Bcl-2 protein than normal tissues (p < 0.051). Bcl-xL followed the same trend since its levels were higher in tumor than in normal samples (p < 0.048). Interestingly, Bcl-2 and Bcl-xL protein content showed a trend towards an inverse correlation (r = 0.27, p < 0.052). mRNA and protein levels directly correlated only with Bcl-2 (r = 0.63, p < 0.0032). Despite the fact that the amounts of Bcl-2, Bax and Bcl-xL proteins in the neoplastic population were not significantly differently distributed according to the clinicopathological features of the patients, the differences observed between normal and neoplastic samples suggest that these proteins may play a role in endometrial carcinoma: long-term follow-up studies will be required to confirm this hypothesis.
The proapoptotic Bax protein induces cell death by acting on mitochondria. Bax binds to the permeability transition pore complex
(PTPC), a composite proteaceous channel that is involved in the regulation of mitochondrial membrane permeability. Immunodepletion
of Bax from PTPC or purification of PTPC from Bax-deficient mice yielded a PTPC that could not permeabilize membranes in response
to atractyloside, a proapoptotic ligand of the adenine nucleotide translocator (ANT). Bax and ANT coimmunoprecipitated and
interacted in the yeast two-hybrid system. Ectopic expression of Bax induced cell death in wild-type but not in ANT-deficient
yeast. Recombinant Bax and purified ANT, but neither of them alone, efficiently formed atractyloside-responsive channels in
artificial membranes. Hence, the proapoptotic molecule Bax and the constitutive mitochondrial protein ANT cooperate within
the PTPC to increase mitochondrial membrane permeability and to trigger cell death.
The transcription factor c-Jun mediates several cellular processes, including proliferation and survival, and is upregulated in many carcinomas. Liver-specific inactivation of c-Jun at different stages of tumor development was used to study its role in chemically induced hepatocellular carcinomas (HCCs) in mice. The requirement for c-jun was restricted to early stages of tumor development, and the number and size of hepatic tumors was dramatically reduced when c-jun was inactivated after the tumor had initiated. The impaired tumor development correlated with increased levels of p53 and its target gene noxa, resulting in the induction of apoptosis without affecting cell proliferation. Primary hepatocytes lacking c-Jun showed increased sensitivity to TNF-α-induced apoptosis, which was abrogated in the absence of p53. These data indicate that c-Jun prevents apoptosis by antagonizing p53 activity, illustrating a mechanism that might contribute to the early stages of human HCC development.
We have isolated a novel cDNA that encodes a product showing significant sequence homology (56% identity) to human NIP3, a protein thought to interact with adenovirus E1B19kD and human BCL2 proteins. This cDNA contains an open reading frame of 657 nucleotides encoding a 219 amino acid polypeptide. The gene, designated BNIP3L, was expressed in all 16 normal human tissues examined; we mapped it to chromosome band 8p21 by fluorescence in situ hybridization. Introduction of the BNIP3L gene into six different cancer-cell lines caused significant growth suppression in each of them, while no such effect occurred when the antisense cDNA or the vector DNA was transfected, indicating that BNIP3L may function as a tumor suppressor. Genes Chromosomes Cancer 21:230–235, 1998. © 1998 Wiley-Liss, Inc.
Raf-1 kinase was shown to bind via its catalytic domain (Cat) to Bcl-2 in a BH4 domain-dependent manner. Using a green fluorescent protein (GFP)-Raf-1 (Cat) fusion protein, Bcl-2 but not Bcl-2(Δ BH4) was found to target Raf-1 to mitochondria in cells. Targeting Raf-1 (Cat) to mitochondrial membranes by fusing with the transmembrane domain of an outer mitochondrial membrane protein protected cells from apoptosis and resulted in phosphorylation of BAD protein, whereas plasma-membrane targeted Raf-1 failed to phosphorylate BAD and did not protect against cell death. Moreover, a Bcl-2 binding protein, BAG-1, was shown to not only bind Raf-1 but also increase the activity of this kinase through a protein—protein interaction. The findings suggest that Bcl-2 targets Raf-1 to mitochondria, allowing this kinase to contribute to cellular survival by phosphorylating BAD or possibly other protein substrates in the vicinity of Bcl-2.
The anti-apoptotic molecule BCL2 delays cell-cycle entry from quiescence. We have recently cloned a new member of the BCL2 family of apoptosis-related genes, BCL2L12. In the present study, the expression of BCL2 and BCL2L12 genes during cisplatin-induced apoptosis in HL-60 leukemic cells was investigated. The kinetics of apoptosis induction and cell toxicity were evaluated by DNA laddering and the MTT method, respectively. BCL2 and BCL2L12 expression was analyzed by RT-PCR using gene-specific primers. The ratio of apoptotic cells increased with increasing concentrations of cisplatin and exposure time of cell culture to the drug. Gradual, time-dependent downregulation of BCL2 gene was observed during cisplatin treatment. Up-regulation of BCL2L12 was observed 3 h (no DNA fragmentation) and 6 h (initiation of the DNA fragmentation) after treatment with cisplatin, followed by a decrease of its expression after 12-h continuous treatment with the drug. It is known that the main anti-carcinogenic effect of cisplatin is due to the induction of cell apoptosis. Present results indicate that downregulation of the BCL2 and upregulation of the BCL2L12 gene may be the underlying mechanisms.
We have previously shown that the pro-apoptotic BAX protein is differentially expressed in breast cancer and in other epithelial tumors. In this line, a reduced BAX protein expression is a negative prognostic factor in various carcinomas including breast cancer. For p53, a trancriptional activator of BAX in apoptosis, mutations in the coding sequence were shown to modulate BAX protein expression in cell line models on the transcriptional level. We therefore investigated the BAX gene in 68 breast cancer specimens for the presence of mutations in the coding sequence by single-strand conformation polymorphism (SSCP)-PCR and direct sequencing. The expression of BAX protein was assessed by immunohistochemistry. In addition, we screened for mutations in the exons 5–8 of the p53 gene by SSCP-PCR to assess whether mutations in the DNA-binding domain of this upstream regulator of BAX gene transcription are responsible for differences in BAX protein expression. As previously observed, BAX was differentially expressed in the breast cancer samples, but no mutations in the coding sequence of the BAX gene were found besides a polymorphism in exon 6 at the position 552 (G->A) and additional intronic polymorphisms. In contrast, we identified 16 of 68 (23.5%) tumors to bear mutations in the p53 gene. In the subset of BAX-expressing tumors, the mutational inactivation of p53 did result in a reduced BAX protein expression (Fisher exact test, p = 0.047). Nevertheless, we identified a subset of BAX-negative tumors lacking BAX or p53 mutations. Thus, additional, not yet identified regulators, apart from p53, appear to be involved in the regulation of BAX protein expression. Int. J. Cancer 87:517–521, 2000. © 2000 Wiley-Liss, Inc.
Cervical carcinomas develop as a result of multiple genetic alterations. As the genetic alterations are the cause of malignant transformation, it is likely that specific genetic alterations lead to specific clinical behaviour. The aim of this study was (i) to localise chromosome arms that harbour likely tumour-suppressor genes, by analysing loss of heterozygosity (LOH) and (ii) to study the association of LOH with clinicopathological parameters. To define the regions of interest, we studied the presence of loss of heterozygosity at all chromosomes in 67 cervical carcinomas (stages IB and IIA) with 81 polymorphic markers. In addition, all frequent allelic imbalances were correlated with HPV status and clinicopathologic parameters including survival, FIGO-stage, lymph-node metastasis, tumour size, number of mitoses, vaso-invasion and histologic type. LOH at a frequency over 25% was observed at sites on 9 chromosome arms: 3p21, 4p16.1-15, 6p, 6q22.3-23.1, 11q22-24, 15q11-21.1, 17p13.3, 18q22-qter and Xq. LOH of chromosome 6q14-16.2, 6p22 and 17p13 correlated marginally with HPV-16 positivity. LOH on chromosome 3p21 was weakly correlated with high mitotic activity, while LOH on chromosomes 11q23.3, 15q21.1 and 17p13 correlated with low mitotic activity. LOH at chromosome 17p13 associated marginally with FIGO stage I, while LOH at chromosome 15q associated weakly with FIGO stage II. When chromosome 18q showed LOH in the tumour, the patients had decreased survival (p = 0.024). We conclude that, in carcinoma of the uterine cervix, a novel tumour-suppressor gene may be present on chromosome 15q21 and that patients with LOH on chromosome 18q have relatively poor survival (p = 0.025). Int. J. Cancer (Pred. Oncol.) 79:411–417, 1998. © 1998 Wiley-Liss, Inc.
Apoptosis is an evolutionarily conserved process for killing unwanted cells. Genetic and biochemical experiments have indicated that three groups of proteins are necessary for activation of the cell-death effector machinery: cysteine proteases, their adaptors, and proapoptotic Bcl-2 family members. Antiapoptotic Bcl-2 family members are needed for cell survival. We have cloned Bim, a proapoptotic Bcl-2 family member that shares with the family only a 9-16 aa region of homology [Bcl-3 homology region (BH3)], but is otherwise unique. Bim requires its BH3 region for binding to Bcl-2 and activation of apoptosis. Analysis of Bim-deficient mice has shown that Bim is essential for the execution of some but not all apoptotic stimuli that can be antagonized by Bcl-2. Bim-deficient mice have increased numbers of lymphocytes, plasma cells, and myeloid cells, and most develop fatal autoimmune glomerulonephritis. In healthy cells, Bim is bound to the microtubule-associated dynein motor complex, and is thereby sequestered from Bcl-2. Certain apoptotic signals unleash Bim and allow it to translocate to intracellular membranes, where it interacts with Bcl-2 or its homologues. These results indicate that BH3-only proteins are essential inducers of apoptosis that can be unleashed by certain death signals. Unleashed BH3-only proteins neutralize the prosurvival function of Bcl-2-like molecules, and this is thought to liberate Apaf-l-like adapters to activate caspase zymogens, which then initiate cell degradation.
mcl-1 was identified as an “early-induction” gene that increases in expression during the differentiation of ML-1 human myeloblastic leukemia cells. The mcl-1 gene product proved to be a member of the bcl-2 gene family and, like bcl-2, to have the capacity to promote cell viability. The pattern of expression of mcl-1 has now been characterized, the aim being to determine whether increased expression is consistently associated with differentiation-induction and whether expression is also associated with other changes in proliferative state or cell viability. Expression of the mcl-1 mRNA was found to increase rapidly in ML-1 cells exposed to inducers of monocyte/macrophage differentiation (phorbol esters or lymphocyte conditioned medium), but not cells exposed to an inducer of granulocyte differentiation (retinoic acid). Expression also increased rapidly in response to certain cytotoxic agents (colchicine and vinblastine), but did not increase during serum stimulation or growth-arrest in reduced serum. Increased expression of mcl-1 occurred during the initiation of cell differentiation or death and was not inhibited by cycloheximide, in agreement with the designation of mcl-1 as an early-induction gene. Increased transcription contributed to the increase in expression, and turnover of the mcl-1 mRNA was rapid. These findings suggest that mcl-1 may serve as a modulator of cell viability that can undergo rapid upregulation as well as downregulation, with upregulation harbingering the initiation of cell differentiation or death. © 1996 Wiley-Liss, Inc.
We have recently reported that Mcl-1, an anti-apoptotic member of the Bcl-2 family, is upregulated by interleukin (IL)-6 in human myeloma cells through the janus kinase/signal transducers and activators of transduction (JAK/STAT) pathway. In the current study, we have explored the effects of interferon (IFN)-α, a cytokine which has been shown to increase myeloma cell survival. Our results demonstrate that IFN-α potently upregulates Mcl-1 on both myeloma cell lines and purified native myeloma cells. Of note, this upregulation is not due to an induction of an IL-6 autocrine loop. Furthermore, we showed that IL-6 and IFN-α had no additive effect on Mcl-1 upregulation, suggesting that both cytokines act through a common mechanism. Finally, the analysis of signalling transduction pathways strongly suggests that Mcl-1 upregulation induced by IFN-α depends on STAT3 activation. Altogether, our data show that IFN-α has an IL-6-like effect on human myeloma cells and suggest that it could be deleterious in some patients.
mcl-1 is an immediate-early gene activated by the granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 3 (IL-3)
signaling pathways and plays an important role in the viability response of these cytokines. In this study, we demonstrated
that cytokine stimulation of mcl-1 mRNA and protein expression were attenuated by pretreatment of cells with phosphatidylinositol 3-kinase (PI3-K) inhibitors.
Reporter gene assays further showed that the PI3-K/Akt signaling pathway was involved in IL-3 activation of mcl-1 gene transcription. Analysis of the mcl-1 promoter revealed that both promoter elements, SIE at position −87 and CRE-2 at −70, contribute to IL-3 stimulation of mcl-1 gene expression. Although either the SIE site or the CRE-2 site alone was sufficient to confer IL-3 inducibility on a heterologous
promoter, only IL-3 activation of the CRE-2 reporter was mediated via the PI3-K/Akt pathway. The SIE binding activity was
constitutively high in cells deprived of or stimulated by IL-3. In contrast, the CRE-2 binding activity was low in cytokine-starved
cells and was strongly induced within 1 h following cytokine treatment of cells. In addition, cytokine induction of the CRE-2
but not of the SIE binding activity was dependent on activation of the PI3-K/Akt signaling pathway. Lastly, we showed that
CREB was one component of the CRE-2 binding complex and played a role in IL-3 activation of themcl-1 reporter gene. Taken together, our results suggest that both PI3-K/Akt-dependent and -independent pathways contribute to
the IL-3 activation of mcl-1 gene expression. Activation ofmcl-1 by the PI3-K/Akt-dependent pathway is through a transcription factor complex containing CREB.