Xiaoyan Xuan

S100A4 mediated cell invasion and metastasis of esophageal squamous cell carcinoma via the regulation of MMP-2 and E-cadherin activity

It is well documented that S100A4 is upregulated in a large amount of invasive tumors and plays a pivotal role in tumor invasion and metastasis. However, the precise role and mechanism S100A4 exerts in the invasion and metastasis of esophageal squamous cell carcinoma (ESCC) have not been fully elucidated to date. Our data demonstrated that S100A4 was overexpressed in human ESCC tissues, especially in ESCC with poor differentiation, deep invasion and lymph node metastasis. Subsequently, the knockdown of S100A4 by RNAi in ESCC cell line (EC-1) could reduce cell invasion, metastasis and proliferation ability in vitro. Most importantly, S100A4 regulated MMP-2 positively and E-cadherin negatively in vivo and in vitro to some extent. Our results suggest that S100A4 is an important factor in the invasion, metastasis and proliferation of ESCC and may control invasion and metastasis at least in part through the regulation of MMP-2 and E-cadherin activity. S100A4 may serve as a biomarker for progression of ESCC and a potential molecular target for biotherapy of ESCC.

Astragalus saponins affect proliferation, invasion and apoptosis of gastric cancer BGC-823 cells

BackgroundAstragalus memebranaceus is a traditional Chinese herbal medicine used in treatment of common cold, diarrhea, fatigue, anorexia and cardiac diseases. Recently, there are growing evidences that Astragalus extract may be a potential anti-tumorigenic agent. Some research showed that the total saponins obtained from Astragalus membranaceus possess significant antitumorigenic activity. Gastric cancer is one of the most frequent cancers in the world, almost two-thirds of gastric cancer cases and deaths occur in less developed regions. But the effect of Astragalus membranaceus on proliferation, invasion and apoptosis of gastric cancer BGC-823 cells remains unclear.MethodsAstragalus saponins were extracted. Cells proliferation was determined by CCK-8 assay. Cell cycle and apoptosis were detected by the flow cytometry. Boyden chamber was used to evaluate the invasion and metastasis capabilities of BGC-823 cells. Tumor growth was assessed by subcutaneous inoculation of cells into BALB/c nude mice.ResultsThe results demonstrated that total Astragalus saponins could inhibit human gastric cancer cell growth both in vitro and in vivo, in additional, Astragalus saponins deceased the invasion ability and induced the apoptosis of gastric cancer BGC-823 cells.ConclusionsTotal Astragalus saponins inhibited human gastric cancer cell growth, decreased the invasion ability and induced the apoptosis. This suggested the possibility of further developing Astragalus as an alternative treatment option, or perhaps using it as adjuvant chemotherapeutic agent in gastric cancer therapy.

Myricetin exerts anti-proliferative, anti-invasive, and pro-apoptotic effects on esophageal carcinoma EC9706 and KYSE30 cells via RSK2

Myricetin, a common dietary flavonoid, is widely distributed in fruits and vegetables and is used as a health food supplement based on its anti-tumor properties. However, the effect and mechanisms of myricetin in esophageal carcinoma are not fully understood. Here, we demonstrated the effect of myricetin on the proliferation, apoptosis, and invasion of the esophageal carcinoma cell lines EC9706 and KYSE30 and explored the underlying mechanism and target protein(s) of myricetin. CCK-8 assay, transwell invasion assay, wound-healing assay, cell cycle analysis, and apoptosis assay were used to evaluate the effects of myricetin on cell proliferation, invasion, and apoptosis. Nude mouse tumor xenograft model was built to understand the interaction between myricetin and NTD RSK2. Pull-down assay was used to verify molecular mechanism. Myricetin inhibited proliferation and invasion and induced apoptosis of EC9706 and KYSE30 cells. Moreover, myricetin was shown to bind RSK2 through the NH2-terminal kinase domain. Finally, myricetin inhibited EC9706 and KYSE30 cell proliferation through Mad1 and induced cell apoptosis via Bad. Myricetin inhibits the proliferation and invasion and induces apoptosis in EC9706 and KYSE30 cells via RSK2. Myricetin exerts anti-proliferative, anti-invasive, and pro-apoptotic effects on esophageal carcinoma EC9706 and KYSE30 cells via RSK2. Our results provide novel insight into myricetin as a potential agent for the prevention and treatment of esophageal carcinoma.

Differential expression profiling of microRNAs and their potential involvement in esophageal squamous cell carcinoma

MicroRNAs are small, noncoding RNAs approximately 18–24 nucleotides in length that negatively regulate gene expression at the posttranscriptional and/or translational level by binding to complimentary sequences in the 3′-untranslated regions of target mRNAs. Growing evidence has indicated the important roles for different miRNA species in the development of different cancers. Therefore, miRNAs have the potential to become new biological markers for esophageal squamous cell carcinoma (ESCC) and to be applied in the diagnosis, prognosis, and targeted treatment of ESCC. In this study, we performed a miRNA microarray to analyze the miRNA expression profile in ESCC compared to normal tissues. Then, we made a preliminary analysis of the biological function for the most differentially expressed miRNAs and their potentially target genes regulated. Some microarray results were validated by performing quantitative RT-PCR. The study provided evidence that linked the biological role of miRNAs to ESCC and showed that miRNAs could undertake a variety of mechanisms. Additionally, we also found that altered miR-429 and miR-451 expression levels were associated with the occurrence of lymph node metastases and the differentiation status and TNM stage in ESCC. The study of miRNAs may lead to finding novel methods to diagnose, treat, and prevent ESCC.

RNAi targeting CXCR4 inhibits proliferation and invasion of esophageal carcinoma cells.

CXC chemokine receptor 4 was found to be expressed by many different types of human cancers and its expression has been correlated with tumor aggressiveness, poor prognosis and resistance to chemotherapy. However the effect of CXCR4 on the esophageal carcinoma cells remains unclear, the present study explored the effects of CXCR4 siRNA on proliferation and invasion of esophageal carcinoma KYSE-150 and TE-13 cells. Two siRNA sequence targeting CXCR4 gene were constructed and then were transfected into KYSE-150 and TE-13 cells by Lipofectamine™2000. Changes of CXCR4 mRNA and protein were analyzed by qRT-PCR and Western blot. Effect of CXCR4 siRNA on KYSE-150 and TE-13 cells proliferation was determined by MTT. Transwell invasion assay was used to evaluate the invasion and metastasis of KYSE-150 and TE-13 cells. Tumor growth was assessed by subcutaneous inoculation of cells into BALB/c nude mice. qRT-PCR and Western blot demonstrate that the expression level of CXCR4 gene were obviously decreased in KYSE-150 and TE-13 cells transfected with CXCR4 targeting siRNA expression vectors. The average amount of cells transfected with CXCR4 siRNA penetrating Matrigel was significantly decreased (p<0.05). Injection of CXCR4 siRNA transfected cells inhibited tumor growth in a xenograft model compared with blank and negative control groups (p <0.05). CXCR4 silenced by siRNA could suppress the proliferation, invasion and metastasis of esophageal carcinoma cell lines KYSE-150 and TE-13 in vitro and in vivo. The results provide a theoretical and experimental basis for the gene therapy of ESCC using RNAi technology based on CXCR4 target site.Virtual slidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/3502376691001138

Notch-1-mediated esophageal carcinoma EC-9706 cell invasion and metastasis by inducing epithelial–mesenchymal transition through Snail

Notch has recently been shown to promote epithelial-to-mesenchymal transition (EMT) by involving in the EMT process that occurs during tumor progression and converts polarized epithelial cells into motile, invasive cells. However, it is still unclear whether the Notch signaling pathway is associated with the regulation of EMT in esophageal carcinoma. The present study explored Notch-1-mediated esophageal carcinoma EC-9706 cell invasion and metastasis by inducing epithelial–mesenchymal transition through Snail. The results demonstrated that the inhibition of Notch-1 expression in the esophageal carcinoma cell line EC-9706 could suppress the occurrence of EMT and at the same time could decrease the invasion and metastasis ability of the EC-9706 cells, indicative of its role in EMT. Snail is a transcriptional repressor of E-cadherin. We found that with the inhibition of Notch-1 expression in the esophageal carcinoma cell line EC-9706, the expression of Snail also decreased. Mechanistic studies showed that the up-expression of Snail in the EC-9706 cells restored the suppression of EMT regulated by Notch-1 inhibition, suggesting the role of Snail in Notch-1-mediated EMT. At the same time, the up-expression of Snail in the EC-9706 cells could also rescue the invasion and metastasis ability inhibited by Notch-1 siRNA. Taken together, our results had revealed that Notch-1 could participate in the invasion and metastasis of esophageal carcinoma through EMT via Snail. This study indicated that Notch-1 might be a useful target for esophageal carcinoma prevention and therapy.

Stathmin is a Marker of Progression and Poor Prognosis in Esophageal Carcinoma

Stathmin, also called oncoprotein 18, is a founding member of the family of microtubule-destabilizing proteins that play a critical role in the regulation of mitosis. At the same time stathmin has been recognized as one of responsible factors in cancer cells. The aim of this study was to assess stathmin status, its correlations with clinicopathological parameters and its role as a progosnostic marker in EC patients. The protein and mRNA levels of stathmin were examined by immunohistochemistry (IHC) and in situ hybridization in 100EC tissues and adjacent noncancerous tissues. mRNA and protein expression of stathmin in three EC cell lines(EC9706, ECa109, EC1 commonly used in research) were also analyzed using immunocytochemistry, western blot and in situ hybridization. The prognostic value of Stathmin expression within the tumor tissues were assessed by Cox regression and Kaplan-Meier analysis. We showed that stathmin expression was significantly higher in EC tissues than in adjacent noncancerous tissues. High stathmin immunostaining score in the EC was positively correlated with tumor differentiation, Tumor invasion, Lymph node metastases, and TNM stage. In addition, we demonstrated that three EC cell lines examined, were constitutively expressing a high level of stathmin. Of those, EC-1 showed the strongest mRNA and protein expression for the stathmin analyzed. Kaplan-Meier analysis showed that significantly longer 5-year survival rate was seen in EC patients with high Stathmin expression, compared to those with low expression of Stathmin expression. Furthermore, multivariate Cox proportional hazard analyses revealed that Stathmin was an independent factors affecting the overall survival probability. In conclusion, our data provide a basis for the concept that stathmin might be associated with EC development and progression.. High levels of Stathmin expression in the tumor tissues may be a good prognostic marker for patients with EC.

Umbilical cord blood-derived dendritic cells loaded with BGC823 tumor antigens and DC-derived exosomes stimulate efficient cytotoxic T-lymphocyte responses and antitumor immunity in vitro and in vivo.

Background Umbilical cord blood (UCB) is a rich source of hematopoietic stem cells and from which a significant number of dendritic cells (DCs) can be produced. But the therapeutic role of DCs and exosomes (EXO) generated from DCs is not fully elucidated. Material and methods The UCB-derived DCs were loaded with tumor antigens generated from BGC823 cell line. Exosomes were derived from these DCs by ultracentrifugation. Dendritic cells and DCex were evaluated by light microscope, transmission electron microscope (TEM), flow cytometry, and western blot assay. The therapeutic role of DCs and EXO generated from DCs were then detected in vitro and in vivo. Results Dendritic cells isolated from umbilical cord blood after loading with tumor antigens generated from BGC823 cell line could express high levels of protein molecules: MHC-I, MHC-II, CD34, CD40, CD80, CD86, CD11c and CD54 and mediate a stronger promotion of T cells proliferation. And, they could also enhance the cytotoxicity effects of the generated CTL in vitro and in vivo. Exosomes isolated from these DCs were 40-90-nm round particles with a complete membrane structure and could also expressed molecules similar to DCs. Exosomes could stimulate T cell proliferation, produce effective cytotoxicity and induce more efficient in vivo antitumor immunity. Conclusions These results suggested that tumor antigens loaded DCs derived from unrelated umbilical cord blood or DCex can induce tumor specific cytotoxicity and this may represent a novel immunotherapy for tumors. Because of their advantage of stable, easy to store, DCex have a more brilliant prospects in the tumor immunity. Additional information We reported that exosomes derived from umbilical cord blood dendritic cell (UBDC), similar to DCs, can trigger activation of T cells significantly. These data demonstrate that DC-derived exosomes (DCex) can mediate essential adaptive immune functions.

Increased expression levels of S100A4 associated with hypoxia-induced invasion and metastasis in esophageal squamous cell cancer.

Here, we explored the expression of S100A4 in esophageal squamous cell cancer (ESCC) tissues and investigated its role in hypoxia-induced invasion and metastasis in ESCC cell lines EC-1 and EC-9706. Immunohistochemistry analysis demonstrated that S100A4 was overexpressed in human ESCC tissues especially in ESCC tissues with deep invasion and lymph node metastasis. Hypoxia-induced S100A4 overexpression was observed in EC-1 and EC-9706 cells, in which it was associated with invasion and metastasis. Furthermore, we used EC-1 and EC-9706 cells again to upregulate or knockdown the expression S100A4 to investigate the mechanism role of S100A4 in hypoxia-induced invasion and metastasis in ESCC cells. And the results showed that S100A4 played an important role in promoting the invasion and metastasis of EC-1 and EC-9706 cells under hypoxia. Therefore, S100A4 overexpression might be an important mechanism by which hypoxia induced invasion and metastasis, and S100A4 could also be a potential target for the treatment of ESCC.

CCR9 and CCR7 are over-expressed in CD4(-) CD8(-) thymocytes of myasthenia gravis patients.

Introduction: Chemokine CC motif receptors 9 and 7 (CCR9 and CCR7) play a major role in the migration of T‐cell precursors to the thymus to initiate T thymopoiesis. However, their role in development of T‐cells in myasthenia gravis (MG) patients has not been fully elucidated. Methods: Expression and distribution of CCR9+ and CCR7+ cells were detected by flow cytometry and immunofluorescence. Real‐time polymerase chain reaction was used to check the adhesion molecules on CD4–CD8– double‐negative (DN) thymocytes. Results: CCR9 and CCR7 expression by DN thymocytes increased in the MG thymus; the levels of CCR9, CCR7, interleukin‐7R mRNA increased, and CXCR4 levels decreased compared with levels in the non‐MG thymus. More CCR7 and CCR9 double‐positive (DP) thymocytes were gathered near the subcapsular region in MG thymus. Conclusions: Enhanced expression of CCR9 and CCR7 may complicate the differentiation of DP thymocytes from the DN stage in MG thymus. Muscle Nerve, 2016 Muscle Nerve 55: 84–90, 2017