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Immunohistochemistry in human liver cirrhosis. Immunohistochemistry on serial liver sections obtained from a cirrhotic patient with ductular proliferation. A) Representative images show negative control (C-) and immunostaining for cytokeratin 7 (CK-7), cytokeratin 19 (CK-19) and for SerpinB3. B) Serial liver sections immunostained for SerpinB3, EpCAM, CD-90, CD-34 and CD-117. Original magnification as indicated.
Source publication
In the setting of liver injury hepatic progenitor cells are activated, counterbalancing the inhibited regenerative capacity of mature hepatocytes. Chronic activation of this compartment may give rise to a subset of liver tumours with poor prognosis. SerpinB3, a serpin over-expressed in injured liver and in primary liver cancer, has been shown to in...
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Human epithelial cell adhesion molecule hEpCAM is a tumour-associated antigen frequently expressed in carcinomas, which promotes proliferation after regulated intramembrane proteolysis. Here, we describe extracellular shedding of hEpCAM at two α-sites through ADAM proteases and at one β-site through BACE1. Transmembrane cleavage by γ-secretase occu...
Citations
... As aforementioned, the inhibition of lysosomal proteases was proposed as an additional carcinogenic mechanism, through the induction of a constitutive and chronic activation of the endoplasmic reticulum stress-related unfolded protein response [82]. Regarding its role in primary liver cancers, SerpinB3 was found to be expressed in aggressive forms of all these tumors, since its expression is observed in the hepatic stem cell compartment of both fetal and adult cirrhotic livers [86]. ...
SerpinB3, a member of the serine-protease inhibitor family, has emerged as a crucial player in various physiological and pathological processes. Initially identified as an oncogenic factor in squamous cell carcinomas, SerpinB3's intricate involvement extends from fibrosis progression and cancer to cell protection in acute oxidative stress conditions. This review explores the multi-faceted roles of SerpinB3, focusing on its implications in fibrosis, metabolic syndrome, carcino-genesis, and immune system impairment. Furthermore, its involvement in tissue protection from oxidative stress and wound healing underscores its potential as diagnostic and therapeutic tool. Recent studies have described the therapeutic potential of targeting SerpinB3 through its upstream regulators, offering novel strategies for cancer treatment development. Overall, this review underscores the importance of further research to fully elucidate the mechanisms of ac-tion of SerpinB3 and to exploit its therapeutic potential across various medical conditions.
... Strategies by which SERPINB3 leads to the development of primary liver cancers are multiple and are supported by the previously described findings, including apoptosis resistance, increased proliferation and EMT induction, impaired immune response, and induction of stemness features. While these properties confer cell protection in acute damage conditions [126][127][128], they favor malignant transformation in case of chronic injuries. It is worth noting that SERPINB3 has been detected in the more aggressive forms of all the types of primary liver cancers, including HCC, cholangiocarcinoma, and hepatoblastoma and this is not surprising, considering that this SERPIN is expressed in the hepatic stem cells compartment, consisting in the epithelial cell adhesion molecule (EpCAM) positive fraction, sorted from both foetal and adult cirrhotic livers [129]. ...
Chronic liver diseases (CLDs), which are typically characterized by fibrogenic progression towards liver cirrhosis and related complications eventually leading to organ failure and can also lead to the development of primary liver cancers, represent a major burden for human health on a worldwide basis. Although the present knowledge on the pathogenesis of CLDs progression and primary liver cancers development has remarkably increased in the last decades, critical molecular mediators remain incompletely understood, and approved antifibrotic therapies to efficiently counteract CLDs progression and liver cancer are lacking. In the present review, this study will specifically analyse the putative contribution of SERPINB3, a member of the superfamily of serine-protease inhibitors (SERPINs), which has been shown to exert significant pro-inflammatory and pro-fibrogenic roles in progressive CLDs as well as to be involved in the development of primary liver cancers, including hepatocellular carcinoma (HCC), cholangiocarcinoma, and hepatoblastoma.
... Squamous cell carcinoma antigen (SCCA1), also known as Ser-pinB3, is a serine protease inhibitor, described in the liver stemprogenitor cell compartment. This protein, not detectable in normal liver, has been indeed identified in EpCam positive cell sorted from foetal and cirrhotic human livers [12] . SerpinB3 has also been detected in preneoplastic liver lesions [13] and in HCC, especially in patients with poor prognosis [14][15][16] . ...
... In addition, this molecule activates the Wnt/ β−catenin signaling, leading to more aggressive forms not only in tumors of the liver [14 , 15] , but also in colorectal and oesophageal cancers [28 , 16] . Furthermore, given the expression of SCCA in the liver progenitor cell compartment [12] , this serine protease inhibitor could be useful to identify a subset of e-CCA endowed with a more invasive phenotype originating from a stem cell niche located in the peribiliary glands. ...
Background
Cholangiocarcinoma (CCA) is a rare biliary tract tumor with poor prognosis that often is challenging to diagnose and the majority of patients present with advanced stage. Squamous cell carcinoma antigen 1 (SCCA1) overexpression has been found in different tumors associated with poor prognosis and chemoresistance.
Aims
To assess the presence and possible prognostic role of SCCA1/2 isoforms in bile and serum of patients with CCA.
Methods
Forty seven surgical patients (36 with CCA and 11 with benign diseases) were prospectively included in the study. Serum and bile specimens were collected at the time of surgery and free and IgM-complexed SCCA was quantified by ELISA (Xeptagen, srl).
Results
Free or IgM linked SCCA was rarely found in serum, while SCCA was detectable in bile samples of patients with CCA, especially in those with extrahepatic form (43% vs 17%, p = 0.008), but not in controls. Despite similar tumor stage, these positive patients presented a trend toward a higher percentage of portal invasion (27% vs 15%) and of tumor recurrence than negative cases (62% vs 40%), although the difference was not statistically significant.
Conclusion
These preliminary results indicate that bile testing for SCCA is a specific marker of extrahepatic CCA, with potential prognostic value.
... Based on the decrease in cell viability we observed after SerpinB3 knockdown and the well-established role for SerpinB3 in inhibiting cell death, we asked whether this correlates with an increase in cell death (Villano et al., 2014). Moreover, as our NanoString analysis of cancer pathways also revealed an increase in apoptosis in cells depleted of SerpinB3 ( Figure 2E), we validated this increase in apoptosis after SerpinB3 knockdown as read out by annexin V/propidium iodide (PI) double-positive cells in multiple CSC models ( Figures 3A and S3A). ...
... A recent study in cholangiocarcinoma found that SerpinB3 was expressed in a stem-like subset of cells and that knockdown of SerpinB3 resulted in decreased invasion and proliferation (Correnti et al., 2021). In addition to its roles in cancer, SerpinB3 is Cell Reports 40, 111348, September 13, 2022 7 Article ll expressed in hepatic stem cells, where SerpinB3 expression correlates with decreased activated caspase 3 (Villano et al., 2014). SerpinB3 has been linked to the inhibition of apoptosis in the settings of endoplasmic reticulum stress (Verfaillie et al., 2013), tumor necrosis factor-a (TNF-a) release , radiation , and ultraviolet radiation (Katagiri et al., 2006), but the exact mechanism is unknown. ...
Despite therapeutic interventions for glioblastoma (GBM), cancer stem cells (CSCs) drive recurrence. The precise mechanisms underlying CSC resistance, namely inhibition of cell death, are unclear. We built on previous observations that the high cell surface expression of junctional adhesion molecule-A drives CSC maintenance and identified downstream signaling networks, including the cysteine protease inhibitor SerpinB3. Using genetic depletion approaches, we found that SerpinB3 is necessary for CSC maintenance, survival, and tumor growth, as well as CSC pathway activation. Knockdown of SerpinB3 also increased apoptosis and susceptibility to radiation therapy. SerpinB3 was essential to buffer cathepsin L-mediated cell death, which was enhanced with radiation. Finally, we found that SerpinB3 knockdown increased the efficacy of radiation in pre-clinical models. Taken together, our findings identify a GBM CSC-specific survival mechanism involving a cysteine protease inhibitor, SerpinB3, and provide a potential target to improve the efficacy of GBM therapies against therapeutically resistant CSCs.
... They can also act independently of their protease inhibitory functions, e.g., as chaperones or hormonal transporters [4]. In particular, SerpinB3 (SB3) can be physiologically detected in the superficial and intermediate layers of the normal squamous epithelium but it is also highly expressed in the hepatic stem / progenitor cell compartment that is composed of quiescent cells that proliferate under conditions of oxidative stress [5]. This molecule has shown a protective anti-apoptotic function, unrelated to its proteinase inhibition activity, in a variety of stressful conditions [6]. ...
Extracellular vesicles (EVs) are increasingly studied as vectors for drug delivery because they can transfer a variety of molecules across biological barriers. SerpinB3 is a serine protease inhibitor that has shown a protective anti-apoptotic function in a variety of stressful conditions. The aim of this study was to evaluate protection from oxidative stress-induced damage, using extracellular vesicles that overexpress SerpinB3 (EVs-SB3) in order to enhance the effect of extracellular vesicles on cellular homeostasis. EVs-SB3s were obtained from HepG2 cells engineered to overexpress SerpinB3 and they revealed significant proteomic changes, mostly characterized by a reduced expression of other proteins compared with EVs from non-engineered cells. These EV preparations showed a significantly higher protection from H2O2 induced oxidative stress in both the hepatoma cell line and in primary cardiomyocytes, compared to cells treated with naïve EVs or SerpinB3 alone, used at the same concentration. In conclusion, the induction of SerpinB3 transgene expression results in the secretion of EVs enriched with the protein product that exhibits enhanced cytoprotective activity, compared with naïve EVs or the nude SerpinB3 protein.
... In line with previous findings, here we confirm the relationship between miR-122/SerpinB3 axis and stemness markers in HCC specimens, as well as the opposite modulation of stem cell features exerted by miR-122 and SerpinB3 in HCC cell lines, suggesting the targeting of SerpinB3 as a possible mechanism contributing to miR-122 active role in reducing HCC stemness. In this context, we previously reported that SerpinB3 is highly expressed in the hepatic stem/progenitor cell compartment of both fetal and adult livers [35]. In addition, an involvement of miR-122 in stem-associated phenotypes was previously reported in HCCs, where it inhibited cancer stem cell (CSC) glycolysis through the direct targeting of PDK4 [36][37][38], suggesting that miR-122 is fundamental for metabolic reprogramming of HCC cells. ...
The only first-line treatment approved for advanced hepatocellular carcinoma (HCC) is sorafenib. Since many patients experience drug resistance, the discovery of more effective therapeutic strategies represents an unmet clinical need. MicroRNA (MiR)-122 is downregulated in most HCCs, while oncogenic SerpinB3 is upregulated. Here, we assessed the relationship between miR-122 and SerpinB3 and their influence on cell phenotype and sorafenib resistance in HCC. A bioinformatics analysis identified SerpinB3 among hypothetical miR-122 targets. In SerpinB3-overexpressing HepG2 cells, miR-122 transfection decreased SerpinB3 mRNA and protein levels, whereas miR-122 inhibition increased SerpinB3 expression. Luciferase assay demonstrated the interaction between miR-122 and SerpinB3 mRNA. In an HCC rat model, high miR-122 levels were associated with negative SerpinB3 expression, while low miR-122 levels correlated with SerpinB3 positivity. A negative correlation between miR-122 and SerpinB3 or stem cell markers was found in HCC patients. Anti-miR-122 transfection increased cell viability in sorafenib-treated Huh-7 cells, while miR-122 overexpression increased sorafenib sensitivity in treated cells, but not in those overexpressing SerpinB3. In conclusion, we demonstrated that miR-122 targets SerpinB3, and its low levels are associated with SerpinB3 positivity and a stem-like phenotype in HCC. MiR-122 replacement therapy in combination with sorafenib deserves attention as a possible therapeutic strategy in SerpinB3-negative HCCs.
... The presence of SerpinB3 has been further described in liver stem/progenitor cells positive for the hepatic epithelial cell adhesion molecule (EpCAM), both in human fetal livers and in adult livers with cirrhosis, and these findings were corroborated by the induction of this serpin in a mouse model of liver stem/progenitor cell activation [39] . Liver tumors with stemness signature are highly aggressive, and along this line the highest levels of SerpinB3 have been found overexpressed, together with TGF-β1, in the subset of aggressive forms of hepatocellular carcinoma, characterized by early tumor recurrence after surgical resection [8] . ...
Hepatocellular carcinoma (HCC) is one of the most relevant sanitary problems for its prevalence and poor prognosis. This tumor is characterized by highly heterogeneous features, both at clinical and molecular level. SerpinB3 (squamous cell carcinoma antigen-1 or SCCA1) is a serine-protease inhibitor that protects cells from oxidative stress conditions, but in chronic liver damage it may lead to HCC through different strategies, including inhibition of apoptosis, induction of epithelial to mesenchymal transition, cell proliferation and invasiveness. Mechanisms of tumor growth promotion induced by SerpinB3 encompass the inhibition of intratumor infiltration of natural killer cells and the up-regulation of Myc oncogene. Recently this serpin has also been identified as a Ras-responsive factor and modulator of metabolic pathways. In the liver SerpinB3 is undetectable in normal hepatocytes, but its expression progressively increases in chronic liver diseases, dysplastic nodules and hepatocellular carcinoma, especially in those with poor prognosis, in which it could also exert immunomodulatory effects. In serum SerpinB3/4 isoforms (or SCCA) circulate bound to IgMs (SCCA-IgM) in patients with HCC, and in patients with cirrhosis their levels have been found correlated to the risk of HCC development. Preliminary findings in patients with HCC revealed that SCCA-IgM levels are predictive of HCC prognosis.
... In the liver, SerpinB3, a serine protease inhibitor overexpressed in several tumors of epithelial origin [9], was not detected in normal hepatocytes, while its expression was found progressively increased in chronic liver diseases [10], dysplastic nodules [11] and HCC [12,13]. SerpinB3 overexpression was also recently described in the liver progenitor cell compartment [14], where DPPIV/CD26 was identified as a liver progenitor cell marker [15]. Beyond its antiprotease activity, SerpinB3 makes cells more resistant to apoptosis [16], induces epithelial-mesenchymal transition (EMT), cell invasiveness and proliferation [17]. ...
... The results of our study not only confirm that this molecule is expressed in HCCs [26], in parallel with SerpinB3 [27], but also that the expression of both molecules was correlated with the grade of tumor differentiation, with the highest values detected in GI tumors. It is worth to note that both molecules were also expressed in the small bile ducts that likely reflect the activation of the liver stem cell compartment of peritumoral liver tissue [14,15]. In line with these clinical observations, our in vitro experiments showed a significant up-regulation of DPPIV/CD26 in cells overexpressing SerpinB3. ...
Aims:
In hepatocellular carcinoma (HCC), the regulatory protease Dipeptidyl-peptidase IV (DPPIV/CD26), that possesses pro-apoptotic properties, has been found abnormally regulated. The protease inhibitor SerpinB3, exerting anti-apoptotic activity, has also been described to be upregulated, especially in HCCs with poor prognosis. The aim of this study was to investigate the possible relationship between these two molecules in HCC patients and in experimental models.
Materials and methods:
DPPIV/CD26 and SerpinB3 expression was measured in liver specimens of 67 patients with HCC. HepG2 and Huh7 cells, stably transfected to overexpress SerpinB3, and respective control cells were used to assess biological and metabolic modifications of DPPIV/CD26 activity induced by this serpin.
Key findings:
DPPIV/CD26 and SerpinB3 were localized in the same tumoral areas and both molecules was directly correlated with the grade of tumor differentiation, with the highest values detected in GI tumors. Cell lines over-expressing SerpinB3 displayed upregulation of DPPIV/CD26, likely as a feedback mechanism, due to the DPPIV/CD26 protease activity inhibition by SerpinB3, as confirmed by the similar behavior induced by the inhibitor Sitagliptin. Moreover, they exhibited lower glycogen storage and higher lipid accumulation, typical effects of DPPIV/CD26.
Significance:
A close connection between SerpinB3 and DPPPIV has been identified, but further studies are required to better understand the mechanism by which these proteins communicate and exert metabolic effects in HCC.
... 18 To characterize the mouse-homologous serpin isoforms, the following set of conserved primers was used: sense: 5′-TTT TACA CAAG TCCT TTGTGGAGG-3′, antisense: 5′-CTG GACA CATG GAAG AGACACCAC-3′, which allows the amplification of the mouse Serpinb3a, -b3b and -b3c, homologous to human SerpinB3 and SerpinB4. 19 To characterize the expression of human SerpinB3 in human cell lines the following set of specific primer was used: sense: 5'-aactcctgggtggaaagtcaa-3'; antisense: 5'-accaatgtggtattgctgccaa-3'. β-Actin (for mouse models) and GAPDH (for human cell lines) were used as internal reference and coamplified with target samples using identical qRT-PCR conditions. ...
Iron overload results in cellular toxicity, tissue injury, organ fibrosis and increased risk of neoplastic transformation. SerpinB3 is a serine protease inhibitor overexpressed in the liver in oxidative stress conditions, able to induce fibrosis and increased risk of malignant transformation. Aim of the present study was to assess the effect of iron overload on SerpinB3 expression in the liver using in vivo and in vitro models.
The expression of Serpinb3 was assessed in the liver of hemojuvelin knockout mice (Hjv−/−), an established model of hereditary hemochromatosis, and of wild type control mice, following dietary or pharmacological iron manipulation. To assess the direct effect of iron in vitro, cell lines were treated with different concentration of hemin or with an iron chelator.
Hepatic Serpinb3 mRNA and protein were highly expressed in Hjv−/− mice, but not in wild type controls fed with a standard diet. Serpinb3 became detectable in wild type mice fed with a high iron diet or injected with iron dextran; these treatments further induced Serpinb3 expression in Hjv−/− mice. Livers expressing Serpinb3 showed a positive staining also for HIF-2α in the same areas. Hemin promoted induction of SerpinB3 mRNA in HeLa and HA22T/VGH cells, but a mild stimulation of SerpinB3 promoter activity in HeLa and Huh7 cells. In conclusion, Serpinb3 is strongly induced by iron in the mouse liver. The molecular link between iron, ROS and SerpinB3 seems to be HIF-2α, which is induced by iron overload and was previously found capable of up-regulating SerpinB3 at the transcriptional level.
... This may denote failure of hepatocyte regeneration as it is associated with the occasional detection of hepatic progenitor cells. It is reported that impairment of hepatocytes regeneration may trigger the expansion of stem/progenitor cells counterbalancing the inhibited regenerative ability of mature hepatocytes [58,59]. This hepatocyte fragmentation or splitting can be considered clonal fragmentation or kind of regeneration. ...
Hepatitis C virus represents one of the rising causes of hepatocellular carcinoma (HCC). Although the early diagnosis of HCC is vital for successful curative treatment, the majority of lesions are diagnosed in an irredeemable phase. This work deals with a comparative ultrastructural study of experimentally gradually induced HCC, surgically resected HCC, and potential premalignant lesions from HCV-infected patients, with the prospect to detect cellular criteria denoting premalignant transformation. Among the main detected pathological changes which are postulated to precede frank HCC: failure of normal hepatocyte regeneration with star shape clonal fragmentation, frequent elucidation of hepatic progenitor cells and Hering canals, hepatocytes of different electron density loaded with small sized rounded monotonous mitochondria, increase junctional complexes bordering bile canaliculi and in between hepatocyte membranes, abundant cellular proteinaceous material with hypertrophied or vesiculated rough endoplasmic reticulum (RER), sequestrated nucleus with proteinaceous granular material or hypertrophied RER, formation of lipolysosomes, large autophagosomes, and micro-vesicular fat deposition.
In conclusion, the present work has visualized new hepatocytic division or regenerative process that mimic splitting or clonal fragmentation that occurs in primitive creature. Also, new observations that may be of value or assist in predicting HCC and identifying the appropriate patient for surveillance have been reported. Moreover, it has pointed to the possible malignant potentiality of liver stem/progenitor cells.
For reliability, the results can be subjected to cohort longitudinal study.
