Shi M.,Gastroenterology |
Li G.,Shanghai Changning Central Hospital |
Wang N.,Gastroenterology |
Wei J.,Gastroenterology |
And 3 more authors.
International Journal of Oncology | Year: 2013
We investigated the inhibition of apoptosis and proliferation of poorly differentiated AGS gastric cancer cells by epigallocatechin-3-gallate (EGCG), to establish target genes for regulation by EGCG. The proliferation and apoptosis of AGS gastric cancer cells treated with EGCG were observed by cell counting kit (CCK)-8 and flow cytometry. Differential gene expression in AGS cells treated with EGCG was screened by gene expression microarrays. Id1 gene and protein expression were determined by quantitative PCR and western blot analysis. The effect of Id1 on EGCG-induced apoptosis and cell cycle arrest of AGS cells was verified with RNAi. The proliferation and apoptosis of AGS cells treated with siRNA Id1 was observed by CCK-8 and flow cytometry. EGCG significantly promoted apoptosis and inhibited the proliferation of AGS cells. The Id1 gene was differentially expressed in AGS cells treated with EGCG, and Id1 mRNA and protein were downregulated in AGS cells treated with EGCG, confirmed by quantitative PCR and western blot analysis. Id1 mRNA and protein were also downregulated in AGS cells treated with siRNA-Id1. The apoptosis and proliferation of AGS cells treated with siRNA-Id1 were similar to those in cells treated with EGCG. EGCG induces apoptosis and inhibits proliferation of poorly differentiated AGS gastric cancer cells, and Id1 may be one of the target genes regulated by EGCG in cancer inhibition.
Li G.-M.,Shanghai JiaoTong University |
Wang Y.-G.,Shanghai Changning Central Hospital |
Pan Q.,Shanghai JiaoTong University |
Wang J.,Shanghai JiaoTong University |
And 2 more authors.
International Journal of Clinical and Experimental Pathology | Year: 2014
Sorafenib is the first drug currently approved to treat advanced hepatocellular carcinoma (HCC). However, very low response rate and acquired drug resistance makes rare patients benefit from sorafenib therapy, therefore it is urgent to find biomarkers for sorafenib sensitivity. Histone modifications, including histone methylation, have been demonstrated to influence the initiation and progression of HCC. It is of great interest to elicit the possibility whether histone methylation plays a role in regulation of sorafenib sensitivity. In present work, a high throughput RNAi screening with 176 shRNA pools against 88 histone methyltransferases (HMTs) and histone demethyltransferases genes was applied to HepG2 cells. Silencing of 3 genes (ASH1L, C17ORF49 and SETD4) was validated to specifically promote HepG2 cells sensitivity to sorafenib. Western blotting results showed that those 3 HMT genes knockdown alone or sorafenib treatments alone both induce AKT/ERK activation. However, combination treatment with sorafenib and silencing of C17ORF49 or SETD4 downregulated AKT phosphorylation and hence induced HCC cells death. Our work may provide potential biomarkers for sorafenib sensitivity and therapeutic combination for sorafenib treatment in HCC patients.
Shi J.,Tongji University |
Zhang M.,Shanghai Changning Central Hospital |
Zhang L.,First Peoples Hospital of Yunnan Province |
Wang P.,First Peoples Hospital of Yunnan Province |
And 2 more authors.
Microbial Biotechnology | Year: 2014
Xylose fermentation is necessary for the bioconversion of lignocellulose to ethanol as fuel, but wild-type Saccharomyces cerevisiae strains cannot fully metabolize xylose. Several efforts have been made to obtain microbial strains with enhanced xylose fermentation. However, xylose fermentation remains a serious challenge because of the complexity of lignocellulosic biomass hydrolysates. Genome shuffling has been widely used for the rapid improvement of industrially important microbial strains. After two rounds of genome shuffling, a genetically stable, high-ethanol-producing strain was obtained. Designated as TJ2-3, this strain could ferment xylose and produce 1.5 times more ethanol than wild-type Pichiastipitis after fermentation for 96h. The acridine orange and propidium iodide uptake assays showed that the maintenance of yeast cell membrane integrity is important for ethanol fermentation. This study highlights the importance of genome shuffling in P.stipitis as an effective method for enhancing the productivity of industrial strains. Designated as TJ2-3, this strain could ferment xylose and produce 1.5 times more ethanol than wild-type P. stipitis after fermentation for 96 h. The acridine orange and propidium iodide uptake assays showed that the maintenance of yeast cell membrane integrity is important for ethanol fermentation. This study highlights the importance of genome shuffling in P. stipitis as an effective method for enhancing the productivity of industrial strains. © 2013 The Authors.
Wang Y.,Shanghai Changning Central Hospital |
Shi M.,Shanghai Changning Central Hospital |
Fu H.,Shanghai Changning Central Hospital |
Xu H.,Shanghai Changning Central Hospital |
And 3 more authors.
Molecular Medicine Reports | Year: 2010
Non-alcoholic fatty liver disease (NAFLD) is a common liver disease associated with an increased risk of type 2 diabetes and cardiovascular disease. Many factors may contribute to NAFLD development and progression, but the exact mechanisms are still not fully understood. In this study, Sprague-Dawley rats were fed either a standard diet (control group), a high-fat diet for 8 weeks (the HFD-8 group) or a high-fat diet for 16 weeks (the HFD-16 group). The HFD animals showed high levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and insulin resistance index (Homa-IR). Mild and severe steatosis was found in both the HFD-8 and HFD-16 groups, respectively. Compared with the controls, mRNA levels of mTOR, S6K1, IL-1α, IL-6 and TNFα were significantly increased in the HFD-8 and HFD-16 groups. IRS-1 mRNA was significantly increased in the HFD-8 group, but not in the HFD-16 group. The protein levels of mTOR, pmTOR(Ser2448), S6K1, pIRS-1(Ser307), IL-1α and IL-6 were significantly increased in the HFD-8 and HFD-16 groups. The protein levels of pmTOR(Ser2448) and IL-1α were significantly higher in the HFD-16 group compared to those in the HFD-8 group. However, the protein expression level of mTOR did not differ significantly between the HFD-8 and HFD-16 groups. The pIRS-1(Tyr102) level was significantly lower in both the HFD-8 and HFD-16 groups when compared to that in the control group, and the pIRS-1(Tyr102) level was significantly lower in the HFD-16 group compared to that of the HFD-8 group. pmTOR(Ser2448) was positively correlated with the TNFα mRNA level, and pIRS-1(Ser307) was positively correlated with pmTOR(Ser2448), TNFα, S6K1 and mTOR. pIRS-1(Tyr102) was negatively correlated with pmTOR(Ser2448), TNFα, S6K1 and mTOR. These data indicate that mTOR contributes to insulin resistance and chronic liver inflammation, and may play an important role in the development and progression of NAFLD.
Wang Y.-G.,Shanghai Changning Central Hospital |
Wang N.,Shanghai Changning Central Hospital |
Li G.-M.,Shanghai University |
Fang W.-L.,Shanghai Changning Central Hospital |
And 4 more authors.
World Journal of Gastroenterology | Year: 2013
AIM: To explore the effect of lysine acetylation in related proteins on regulation of the proliferation of gastric cancer cells, and determine the lysine-acetylated proteins and the acetylated modified sites in AGS gastric cancer cells. METHODS: The CCK-8 experiment and flow cytometry were used to observe the changes in proliferation and cycle of AGS cells treated with trichostatin A (TSA). Real time polymerase chain reaction and Western blotting were used to observe expression changes in p21, p53, Bax, Bcl-2, CDK2, and CyclinD1 in gastric cancer cells exposed to TSA. Cytoplasmic proteins in gastric cancer cells before and after TSA treatment were immunoprecipitated with anti-acetylated lysine antibodies, separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis gel and silver-stained to detect the proteins by mass spectrometry after removal of the gel. The acetylated proteins in AGS cells were enriched with lysine-acetylated antibodies, and a high-resolution mass spectrometer was used to detect the acetylated proteins and modified sites. RESULTS: TSA significantly inhibited AGS cell proliferation, and promoted cell apoptosis, leading to AGS cell cycle arrest in G0/G1 and G2/M phases, especially G0/ G1 phase. p21, p53 and Bax gene expression levels in AGS cells were increased with TSA treatment duration; Bcl-2, CDK2, and CyclinD1 gene expression levels were decreased with TSA treatment duration. Two unknown protein bands, 72 kDa (before exposure to TSA) and 28 kDa (after exposure to TSA), were identified by silverstaining after immunoprecipitation of AGS cells with the lysine-acetylated monoclonal antibodies. Mass spectrometry showed that the 72 kDa protein band may be PKM2 and the 28 kDa protein band may be ATP5O. The acetylated proteins and modified sites in AGS cells were determined. CONCLUSION: TSA can inhibit gastric cancer cell proliferation, which possibly activated signaling pathways in a variety of tumor-associated factors. ATP5O was obviously acetylated in AGS cells following TSA treatment. © 2013 Baishideng. All rights reserved.