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Wound healing assay in MCF-7 cells. (a) MCF-7 breast cancer cell were transfected with pFlag-Twist, as well as its negative control pFlag-CMV. Twist greatly increases the motility of MCF-7 cells. (b, c) Quantitative analyses of relative migration distance at 24 and 48 h, respectively. Data shown are means AE SD of at least three independent experiments. ** P < .01, (Student's t-test) as compared to control cells
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Breast cancer, the most common malignancy among women worldwide, is a heterogeneous disease, and it therefore has remarkably different biological characteristics and clinical behavior. Breast cancer has been divided into several different molecular subtypes based on the status of estrogen receptor (ER), progesterone receptor (PR), human epidermal g...
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Context 1
... were obtained using the Student's t-test analysis from time 0 (step 4) to the last time point, Obtain the distance of each scratch closure on the basis of the distances that are measured by software. Measure at least five readings of distance for each sample and repeat each experiment at least three times. A representative result is shown in Fig. ...
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Background
The therapeutic use of Helleborus niger L. is manifold due to its specific phytochemical composition. Two compound groups, the ranunculin derivates including protoanemonin and the steroidal saponins, are also associated with toxicity (genotoxicity, disintegration of membrane structures). Therefore, in vitro investigations were performed...
Citations
... Wound healing assay, cell migration, and invasion assays Wound healing assays, cell migration, and invasion assays were performed according to both the manufacturer's instructions and previous work. 53 ...
Estrogen receptor α (ERα) serves as an essential therapeutic predictor for breast cancer (BC) patients and is regulated by epigenetic modification. Abnormal methylation of cytosine phosphoric acid guanine islands in the estrogen receptor 1 (ESR1) gene promoter could silence or decrease ERα expression. In ERα‐negative BC, we previously found snail family transcriptional repressor 2 (SNAI2), a zinc‐finger transcriptional factor, recruited lysine‐specific demethylase 1 to the promoter to transcriptionally suppress ERα expression by demethylating histone H3 lysine 4 dimethylation (H3K4me2). However, the role of SNAI2 in ERα‐positive BC remains elusive. In this study, we observed a positive correlation between SNAI2 and ESR1 methylation, and SNAI2 promoted ESR1 methylation by recruiting DNA methyltransferase 3 beta (DNMT3B) rather than DNA methyltransferase 1 (DNMT1) in ERα‐positive BC cells. Subsequent enrichment analysis illustrated that ESR1 methylation is strongly correlated with cell adhesion and junction. Knocking down DNMT3B could partially reverse SNAI2 overexpression‐induced cell proliferation, migration, and invasion. Moreover, high DNMT3B expression predicted poor relapse‐free survival and overall survival in ERα‐positive BC patients. In conclusion, this study demonstrated the novel mechanisms of the ESR1 methylation mediated with the SNAI2/DNMT3B complex and enhanced awareness of ESR1 methylation's role in promoting epithelial–mesenchymal transition in BC.
... Changes in the expression of EMT-related markers are regulated by transcription factors such as Twist, Snail, and Zeb, which are activated in the early stage of EMT to coordinate the suppression of epithelial genes and the activation of the mesenchymal gene (Bai et al., 2017;Przygodzka et al., 2019). Therefore, blocking the occurrence of EMT is an effective treatment to limit the spread of tumor cells. ...
Modified Sijunzi Decoction (MSJZD) is an empirical prescription of Traditional Chinese Medicine (TCM) and has been corroborated to be effective in multiple human diseases, but its role in non-small cell lung cancer (NSCLC) is enigmatic. Here we mainly analyze the function and mechanism of MSJZD in NSCLC. In this study, we used a method that coupled ultra-performance liquid chromatography to quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) to investigate the major constituents in MSJZD with positive and negative ion modes. Additionally, in in vitro experiments, the effects of serum-containing MSJZD on the biological behavior of NSCLC cells induced by TGF-β1 were assessed by cell function experiments. Then, the influences of serum-containing MSJZD on epithelial-mesenchymal transition (EMT)-related markers were examined by immunofluorescence and western blot assays. Also, the AKT/GSK3β pathway and apoptosis-related markers were estimated by western blotting. Tumor xenografts were generated by subcutaneously injecting A549 cells into BALB/c nude mice to determine the effects of MSJZD in vivo. We first analyzed the composition of MSJZD. In positive ion mode, 47 kinds of components were identified. In negative ion mode, 45 kinds of components were identified. We also found that TGF-β1 contributed to inducing cell morphological changes and EMT progression. In vitro, surprisingly, cell proliferation, migration as well as invasion in NSCLC cells induced by TGF-β1, could be weakened by serum-containing MSJZD, and apoptosis was intensified. Moreover, serum-containing MSJZD weakened EMT passage and AKT/GSK3β pathway activation and induced apoptosis-related markers in NSCLC cells triggered by TGF-β1. In vivo, we discovered that MSJZD attenuated the tumor growth, promoted histopathological damage, and induced apoptosis in A549 tumor-bearing mice. Importantly, MSJZD has also restrained the development of EMT, AKT/GSK3β pathway, and TGF-β1 expression levels in nude mice. These findings demonstrated that MSJZD significantly weakened NSCLC progression by modulating EMT and AKT/GSK3β pathway.
... 26 Generally, overexpression of Ncadherin and low-expression of E-cadherin are the hallmarks of EMT and metastasis, and Twist plays a key role behind this. 27,28 Twist plays a key function in Ecadherin adjustment. Twist, as a basic helix-loop-helix transcription factor, can recognize and combine with the E-box sequence on the promoter of the target gene, consequently affecting the E-box promoter expression at the transcriptional level, inhibiting the expression of Ecadherin, and inducing the formation of EMT. ...
Aim:
To investigate the roles of cell migration and invasion mediated by Twist in endometriosis.
Methods:
The protein levels and locations of Twist, N-cadherin and E-cadherin were measured by Western blot and immunohistochemistry in ectopic endometrium and eutopic endometrium of ovarian endometriosis as well as normal endometrium of nonendometriosis patients. The messenger RNA (mRNA) expressions of Twist, N-cadherin and E-cadherin in these tissues were measured by quantitative reverse transcription polymerase chain reaction. Stable overexpression of Twist in eutopic endometrial stromal cells was transfected with a plasmid-mediated delivery system. The protein and mRNA expressions of N-cadherin and E-cadherin were detected by western blot and reverse transcription polymerase chain reaction. The changes of migration and invasion of endometrial stromal cells were explored by transwell.
Results:
Levels of protein and mRNA of Twist and N-cadherin showed the highest expression in ectopic endometrium of ovarian endometriosis, while lowest in normal endometrium of nonendometriosis patients. On the contrary, the expression of E-cadherin showed highest in normal endometrium of nonendometriosis patients. The overexpression of Twist after transfection significantly upregulated the protein and mRNA expression of N-cadherin, while downregulated the protein and mRNA expression of E-cadherin. There is significant difference between groups. For transwell, the overexpression of Twist in eutopic endometrial stromal cell significantly promoted cell migration and invasion.
Conclusion:
Twist might be related with the increase of migration and invasion in endometrial stromal cells, mediated by epithelial-to-mesenchymal transition.
... At the indicated time points, cells were fixed with methanol and stained using crystal violet. Images were obtained using a camera, with colonies counted using Image-Pro Plus 6.0 analysis software [19]. ...
Exosomes have recently become the subject of increasing research interest. Interactions between tumor and host cells via exosomes play crucial roles in the initiation, progression and invasiveness of breast cancer. In our study, we used exosomes isolated from a co-culture model of THP-1-derived macrophages exposed to apoptotic MCF-7 or MDA-MB-231 breast cancer cell line cells to investigate their effects on naïve MCF-7 or MDA-MB-231 cells in vitro and in vivo. This post-chemotherapy tumor microenvironment model allowed us to explore possible mechanisms that explain increased proliferation and metastasis of breast cancer seen in some patients. Our results suggest that while exosomes derived from macrophages normally inhibit proliferation and metastasis of MCF-7 or MDA-MB-231 cells, exposure of macrophages to breast cancer cells that have experienced chemotherapy are modified them to promote these processes. Exosomes from macrophages exposed to apoptotic cancer cells have increased amounts of IL-6 that increases the phosphorylation of STAT3, which likely explains the increased transcription of STAT3 target genes such as CyclinD1, MMP2 and MMP9. These observations suggest that the inhibition of exosome secretion and STAT3 signaling pathway activation might suppress the growth and metastasis of malignant tumors, and provide new targets for therapeutic treatment of malignant tumors after chemotherapy.
Within the developing head, tissues undergo cell-fate transitions to shape the forming structures. This starts with the neural crest, which undergoes epithelial-to-mesenchymal transition (EMT) to form, amongst other tissues, many of the skeletal tissues of the head. In the eye and ear, these neural crest cells then transform back into an epithelium, via mesenchymal-to-epithelial transition (MET), highlighting the flexibility of this population. Elsewhere in the head, the epithelium loses its integrity and transforms into mesenchyme. Here, we review these craniofacial transitions, looking at why they happen, the factors that trigger them, and the cell and molecular changes they involve. We also discuss the consequences of aberrant EMT and MET in the head.
In this study, we fabricated paclitaxel (PTX) and etoposide (ETP) loaded Poly (lactic-co-glycolic acid) (PLGA) microspheres with core–shell structures and particle sizes ranging from 1 to 4 µm by coaxial electrospraying. The microspheres were analyzed by scanning electron microscopy (SEM), transmission electron microscopy (TEM). The drug loading rate and entrapment efficiency of the microspheres were detected by high performance liquid chromatograph (HPLC). Moreover, the drug release profiles and degradation of drug-loaded PLGA microspheres in vitro were investigated, respectively. The distinct layered structure that existed in the manufactured core–shell microspheres can be observed by TEM. The in vitro release profiles indicated that the PLGA/PTX+ETP (PLGA/PE) microspheres exhibited the controlled release of two drugs in a sequential manner. Cell Counting Kit-8 was used to detect the toxic and side effects of the microspheres on bone tumor cells. PTX and ETP for combination drug therapy loaded microspheres had more cytotoxic effect on saos-2 osteosarcoma cells than the individual drugs. In conclusion, core–shell PLGA microspheres by electrospraying for combination drug therapy is promising for medicine applications, the PLGA/PE microspheres have some potential for osteosarcoma treatment.
