Human SGC-7901 Cells Research Articles

MiR-504 Promoted Gastric Cancer Cell Proliferation and Inhibited Cell Apoptosis by Targeting RBM4.

Background The purpose of this study was to explore the role and underlying mechanism of miR-504 and RBM4 in gastric cancer. Methods The qRT-PCR or Western blot was performed to determine the expressions of miR-504 and RBM4 in the gastric cancer tissues and normal tissues. Human SGC-7901 cells were transfected with miR-504 mimic/inhibitor or pcDNA-RBM4. Cell proliferation and cell apoptosis were assessed by colony formation assay and flow cytometry, respectively. Luciferase reporter gene assays were used to investigate interactions between miR-504 and RBM4 in SGC-7901 cells. Results The relative expression of miR-504 was significantly upregulated in the gastric cancer group (n = 25) than in the paired normal group (n = 25), but the relative RBM4 expression was remarkably downregulated in the gastric tumor group, compared with the normal group. Additionally, miR-504 overexpression increased the viability of gastric cancer cells. Moreover, RBM4 is a functional target of miR-504 in gastric cancer cells. miR-504 was further confirmed to promote SGC-7901 cell proliferation and inhibit cell apoptosis by downregulation RBM4 in vitro. Conclusions miR-504 promotes gastric cancer cell proliferation and inhibits cell apoptosis by targeting RBM4, and this provides a potential diagnostic biomarker and treatment for patients with gastric cancer.

Optimization of brewing conditions in epigallocatechin-3-gallate (EGCG) extraction from Jinxuan summer green tea by response surface methodology

The extraction conditions of epigallocatechin-3-gallate (EGCG) from Jinxuan summer green tea and antitumor activity against human gastric cancer SGC-7901 cells of the green tea extracts were investigated. On the basis of a single factor experiment, Box-Behnken design and response surface methodology were employed to optimize the hot water extraction conditions. The optimal extraction conditions for EGCG were determined as: extraction temperature of 85 °C, extraction time of 34 min, water-tea ratio of 41 mL/g, a solution of pH 6, and extraction twice. Under these conditions, the experimental extraction yield value of EGCG was 33.82 mg/g, which was not significantly different in comparison to predicted values. The results indicated that the regression models were suitable for the EGCG extraction from Jinxuan summer green tea. The summer green tea extract prepared under the optimal conditions had a higher antitumor activity against human gastric cancer SGC-7901 cells than the green tea extract made with traditional tea brewing method.

Increasing oxaliplatin sensitivity by down-regulation the expression of EZH2 in human gastric cancer SGC7901 cells

Objective To investigate the effect of enhancer of zeste homolog 2 (EZH2) knockdown on chemosensitivity to oxaliplatin in human gastric cancer cell line SGC7901. Methods Small interfering RNA (siRNA) fragments were designed and synthesized for EZH2 mRNA sequence and divided into siRNA-1 group, siRNA-2 group and siRNA-3 group. They were transfected into gastric cancer cell line SGC7901. The negative control group and blank control group were set up at the same time. The expressions of EZH2 mRNA and protein in the SGC7901 cells were detected by quantitative real-time polyme-rase chain reaction (qRT-PCR) and Western blotting. CCK-8 assay was used to detect cell proliferation inhibition rates of SGC7901 cells treated by different concentrations of oxaliplatin. Results After transfection of EZH2 siRNA, the relative expression levels of EZH2 mRNA in siRNA-1 group, siRNA-2 group, siRNA-3 group were 0.615±0.190, 0.241±0.152 and 0.450±0.097. The relative expression levels of EZH2 mRNA of the three groups were lower than that in the negative control group (1.165±0.376), and the differences were statistically significant (P=0.028; P=0.002; P=0.007). After transfection of the best siRNA fragment into SGC7901 gastric cancer cells, the relative expre-ssion levels of EZH2 protein in interference group, blank control group and negative control group were 0.036±0.017, 0.362±0.026 and 0.398±0.036, and the diffe-rence among the three groups was statistically significant (F=157.745, P<0.001). The difference between interference group and negative control group was statistically significant (P=0.001), as compared with the blank control group (P=0.002). When oxaliplatin concentration was 2, 4, 8 and 16 μg/ml, the differences of cell proliferation inhibition rate among interference group, negative control group and blank control group were statistically significant [(18.107±2.822)%, (5.867±2.272)%, (5.333±1.883)%, F=28.185, P=0.001; (54.953±2.550)%, (22.177±1.871)%, (20.077±6.032)%, F=74.206, P<0.001; (60.337±1.641)%, (34.597±3.592)%, (30.227±5.273)%, F=54.897, P<0.001; (78.340±2.081)%, (61.857±3.507)%, (63.077±8.473)%, F=8.586, P=0.017]. There was no significant difference among groups of oxaliplatin at the concentration of 32 μg/ml [(83.450±3.715)%, (72.190±3.948)%, (70.731±17.080)%, F=1.358, P=0.326]. The median inhibitory concentration (IC50) of oxaliplatin in siRNA group, negative control group and blank control group were 5.178, 12.643, 13.601 μg/ml. Conclusion Down-regulation of EZH2 gene expression can significantly inhibit the proliferation of gastric cancer cells, and effectively enhance the sensitivity of gastric cancer cells to oxaliplatin, which indicates EZH2 may play important roles in the development of gastric cancer chemotherapy. These results provide an important theoretical basis for gene therapy of gastric cancer. Key words: Stomach neoplasms; RNA, small interfering; Enhancer of zeste homolog 2; Oxaliplatin

The mechanism of stomatin-like protein 2 gene regulating Wnt/β-catenin signaling pathway on proliferation and apoptosis of gastric cancer cells

Objective To investigate the effect of stomatin-like protein 2 (SLP-2) gene on the proliferation and apoptosis of gastric cancer cells by Wnt/β-catenin signaling pathway. Methods The expression of SLP-2 gene in human gastric cancer tissues was detected by Western blotting; NC-siRNA, SLP-2-siRNA1 and SLP-2-siRNA2 were transfected into human gastric cancer SGC-7901 cells, and not transfected with any siRNA as the control group; the expression of SLP-2 protein after transfected cells was detected by Western blotting; cells transfected for 48 h, cell apoptosis was detected by flow cytometry; Cleaved Caspase-3, β-catenin, c-Myc, Cyclin D1protein expression was detected by Western blotting. Results The expression of SLP-2 protein in gastric carcinoma (0.283±0.016)was significantly higher than paracancerous tissues (0.082±0.016), (t=7.789, P=0.003); the inhibitory effect of SLP-2-siRNA2 group was more obvious, and was choiced as a follow-up study; the apoptosis rate (16.62±1.32)% of cells and Cleaved Caspase-3 (1.421±0.210) protein expression in SLP-2-siRNA group were significantly higher than the control group (1.56±0.56)%, (0.510±0.083), β-catenin (0.157±0.024), (0.039±0.016), Cyclin D1 (0.052±0.012) protein expression was significantly lower than the control group (0.311±0.036, 0.137±0.026, 0.183±0.021) (tapoptosis rate=9.324, Papoptosis rate=0.001; tCleaved Caspase-3=6.112, PCleaved Caspase-3=0.006; tβ-catenin=8.868, Pβ-catenin=0.003; tc-Myc=9.112, Pc-Myc=0.002; tCyclin D1=9.987, PCyclin D1=0.001). Conclusion SLP-2 is highly expressed in gastric cancer, Silencing SLP-2 expression can inhibit the proliferation and apoptosis of human gastric cancer cell line SGC-7901, and its mechanism is related to the regulation of Wnt/β-catenin signaling pathway. Key words: Stomatin-like protein 2; Wnt/β-catenin signaling pathway; Gastric cancer; Proliferation; Apoptosis

Mechansim of resveratrol inhibiting interleukin-6-induced invasion of human gastric cancer SGC7901 cells

Objective To elucidate the inhibitory effect of resveratrol on interleukin (IL)-6-induced invasive ability of SGC7901 cells and the molecular mechanism. Methods The cell viability of cells treated with resveratrol was evaluated by methyl thiazolyltetrazolium (MTT) assay. The SGC7901 cells were treated with IL-6 and non-cytotoxic concentration of resveratrol. The invasion of SGC7901 cells was detected by Boyden chamber assay. Matrix metalloproteinase (MMP)-2 activity was determined by gelatine zymography assay. The changes of the related proteins were detected by Western blotting. Results Resveratrol was not toxic to SGC7901 cells at the concentration under 30 μmol/L. The invasion ability of IL-6-induced SGC7901 cells was decreased about 20% after treatment with 20 μmol/L resveratrol for 24 h, accompanied by the inhibition of MMP-2 secretion for 50%, as well as the levels of phosphorylated extracellular regulated protein kinases 1/2 (p-ERK1/2). Conclusion Resveratrol of 20 μmol/L inhibits invasion of SGC7901 cells possibly through the suppression of the activation of ERK signaling pathway, subsequently exerting inhibitory effect on MMP-2. Key words: Resveratrol; Extracellular signal regulated kinase 1/2; Gastric carcinoma

Effect and mechanism of Vav3 on the proliferation of human gastric cancer SGC7901 cells

The purpose of this study was to investigate the effect and mechanism of Vav3 gene on the proliferation of human gastric cancer cell line SGC7901. The expressions of Vav3 proten in gastric cancer tissue, tumor-adjacent tissue, human gastric cancer cell line SGC7901 and gastric epithelial cell line GES-1 cells were tested by Western blot. Vav3-siRNA was transfected into the SGC7901 cells. The proliferation of SGC7901 cells in vitro was measured by MTT assay. Cell cycle of SGC7901 cells was determined by flow cytometry.The expressions of proliferation-related genes PCNA, p16, cyclin D1, Rb were determined by qPCR and Western blot assay. Orthotopic transplantation nude mouse models of gastric cancer were prepared, and the tumor growth and expressions of PCNA, P16, cyclin D1, and Rb proteins were examined. The relative expressions of Vav3 in the gastric cancer and peritumoral tissue were 0.910±0.242 and 0.243±0.045, respectively; the relative expressions of Vav3 in SGC7901 and GSE-1 cells were 0.925±0.127 and 0.277±0.038, respevtively (both P<0.05). The expression of Vav3 protein in SGC7901 cells was effectively inhibited by Vav3-siRNA. Proliferation of SGC7901 cells was inhibited by (83.43±10.17)% after 80 nmol/L Vav3-siRNA transfection (P<0.05). The ratio of SGC7901 cells in G0/G1 phase was increased, and in S phase decreased after Vav3-siRNA transfection (both P<0.05). The expressions of PCNA and cyclin D1 were decreased in cells after Vav3-siRNA transfection, and expressions of p16 and Rb were increased after Vav3-siRNA transfection (P<0.05 for all). The tumor growth in the Vav3-siRNA group was much slower than that in the other 2 control groups of nude mouse models. Compared with the two control groups, expressions of PCNA and cyclin D1 were significantly lower in the Vav3-siRNA group, while expressions of p16 and Rb were increased (P<0.05 for all). Vav3 can promote the proliferation of gastric cancer cells by regulating proliferation-related genes.

Tracing study of nanometer microspheres labeled with fluorescent dye on gastric cancer cells

To explore the tracing role of nanometer microspheres tagged by fluorescent dye NIR-797 in gastric cancer cells in order to guide precise resection for gastric cancer. Targeted polymer segment NH2-PEG-Pep-PCL and non-targeted polymer segment NH2-PEG- PCL(without Pep) were synthesized with NH2-PEG-NH2, gelatinase substrate peptides (Pep) and PCL-COOH by amidation method. Then nanometer microspheres were prepared by self-assemebly method. NIR-797 was linked into the end of PEG through chemical reaction. Molecular weight, particle size, polydispersity and surface potentials of the nanometer microspheres were detected. Uptake degree of human gastric SGC-7901 cells with targeted and non-targeted nanometer microspheres was observed under fluorescence microscope. Fluorescence intensity of above two nanometer microspheres in tumor sites of tumor-burdened nude mice after intravenous injection of nanometer microspheres at different time points was detected by multispectral imaging system. The molecular weight of NH2-PEG-Pep-PCL and NH2-PEG-PCL were similar to original design. The diameter of targeted nanometer microspheres before and after NIR-797-tagging was (124.1±2.1) nm and (129.7±2.8) nm, and the diameter of non-targeted ones before and after NIR-797-tagging was (120.6±2.6) nm and (124.3±2.9) nm (all P>0.05). Fluorescence intensity of SGC-7901 cells and tumor sites was significantly higher in the targeted nanometer microspheres group as compared to the non-targeted group. Targeted nanometer microspheres tagged by flourescent dye have higher targeted affinity to gastric cancer cells and better enrichment in tumor site, indicating a good tracing of tumor and a guidance for accurate resection of tumor.

Bcl-2 down-regulation by small interfering RNA induces Beclin1-dependent autophagy in human SGC-7901 cells.

While Bcl-2 protein is involved in the regulation of apoptosis, recent research showed that Beclin1, described as the essential autophagy effector and haploinsufficient tumor suppressor, was originally isolated as a Bcl-2 interacting protein. Beclin1 interacts with Bcl-2 through a BH3 domain; nevertheless, the function of the anti-apoptotic gene, Bcl-2, in autophagy is not well understood. We explored the role of Bcl-2 in autophagy in human SGC-7901 cells in which Bcl-2 is overexpressed. Knockdown of Bcl-2 by small interfering RNA in human SGC-7901 cells downregulated Bcl-2 protein levels ∼82% and induced autophagy. Beclin1 protein, the first identified autophagy gene product, was induced by as much as 58%. Transmission electron microscopy and DNA fragmentation assay showed that autophagy was enhanced, but not apoptosis, in Bcl-2 siRNA treated cells. The results provide evidence that knockout the anti-apoptotic gene Bcl-2 induces autophagy in SGC-7901 cells and Bcl-2 specific siRNA may be used as a potential therapeutic strategy in gastric cancer cells that overexpress Bcl-2.

Effect of antisense miR-224 on gastric cancer cell proliferation and apoptosis

To observe the effects of miR-224 antisense oligonucleotide (ASO) on the proliferation and apoptosis of gastric cancer cells in vitro and vivo. The expression of miR-224 in the cancer tissues and their adjacent tissues in 120 gastric cancer patients were detected by real-time quantitative PCR. The biological effects of miR-224 ASO on human gastric cancer SGC7901 cells was assessed by MTT assay, clone formation assay, flow cytometry and in vivo experiment in nude mice. Compared with the control group (0.50 ± 0.07), miR-224 ASO significantly reduced the miR-224 mRNA expression in the cancer patients (0.09 ± 0.01, P < 0.05). MTT assay results showed that the survival rate of gastric cells at 24 h, 48 h and 72 h was 53.6%, 59.1% and 70.1% in the miR-224 ASO group, and 12.3%, 17.4% and 24.7%, respectively, in the control group (P < 0.05 for all). Clone formation assay revealed that clone formation rate in the miR-224 ASO group was (5.33 ± 0.74)%, significantly lower than the (33.33 ± 8.38)% in the control group (P < 0.05). Flow cytometry indicated that the apoptotic index was (15.68 ± 1.46)% in the miR-224 ASO group and (3.36 ± 0.88)% in the control group (P < 0.01). In addition, the expressions of Bcl2 mRNA and protein were 1.05 ± 0.04 and 0.21 ± 0.03 in the miR-224 ASO group, significantly lower than that in the control group (4.87 ± 0.96 and 0.88 ± 0.09, P < 0.01). The in vivo study further showed that the tumor volume in the experimental group is significantly smaller than that in the control group (P = 0.01). MiR-224 is overexpressed in human gastric cancer. Reducing the expression of miR-224 can effectively inhibit the growth and promote apoptosis of gastric cancer cells. miR-224 may become a new target for the regulation of gene expression in gastric cancer.

Growth inhibition and induction of apoptosis in SGC-7901 human gastric cancer cells by evodiamine

Evodiamine is one of the major bioactive compounds isolated and purified from the fruit of Fructus Evodiae. Numerous studies have indicated that evodiamine exhibits activity against human tumor cells. In the present study, the effect of evodiamine on the proliferation and apoptosis of SGC-7901 human gastric cancer cells and the correlative mechanisms were investigated. This may provide further experimental evidence of the pharmacological actions of evodiamine and a strategy for its use as a novel chemotherapeutic drug. Following treatment with evodiamine, the typical morphological changes of apoptosis were observed in human SGC-7901 cells. Cell cycle analysis indicated that evodiamine induced G2/M phase arrest in SGC-7901 cells and flow cytometry revealed that evodiamine induced apoptosis. Analysis of the enzymatic activity demonstrated that evodiamine increased the activity of caspase-3, -8 and -9 in SGC-7901 cells. The protein expression of caspase-3, -8 and -9 and Bax increased, and the expression of Bcl-2 decreased following treatment with evodiamine. These results suggest that evodiamine is able to inhibit the proliferation of SGC-7901 cells by inhibiting the cell cycle at G2/M phase and inducing apoptosis in SGC-7901 cells by activating caspase-3, -8 and -9, and altering the expression of caspase-3, Bax and Bcl-2.

RNAi-mediated gene silencing of vascular endothelial growth factor-C inhibits tumor lymphangiogenesis and growth of gastric cancer in vivo in mice

Overexpression of vascular endothelial growth factor-C (VEGF-C) has been implicated as a critical molecular signal in tumor development by promoting intratumoral lymphangiogenesis. The aim of this study was to explore whether small hairpin RNA (shRNA) targeting VEGF-C could inhibit gastric cancer lymphangiogenesis and tumor growth. Plasmid-mediated expression of VEGF-C-shRNA was employed to silence VEGF-C gene expression in human SGC-7901 cell lines. The inhibition of the target gene expression was quantified by real-time quantitative polymerase chain reaction, Western blotting, and enzyme-linked immunosorbent assay. In vitro, the cell viability was determined by MTT assay, flow cytometry analysis, and migration assay. After VEGF-C knockdown was confirmed, the stable cells were inoculated into nude mice. Tumor growth, lymph vessel density (LVD), and microvascular density were compared for mice administered either VEGF-C-shRNA or control. VEGF-C-shRNA causes effective and specific downregulation of VEGF-C expression (P<0.05). The migration activity of SGC-7901 cells was attenuated in vitro (P<0.05). Tumor growth rate and LVD was suppressed in vivo (P<0.05). VEGF-C-shRNA effectively suppressed gastric cancer cell migration in vivo, retards tumorigenicity, and lymphangiogenesis in nude mice.

Effect of lobaplatin on proliferation, apoptosis and expression of Metadherin in human gastric cancer SGC-7901 cells

Objective To investigate The effect of lobaplatin (LBP) on proliferation,apoptosis and Metadherin (MTDH) expression in human gastric cancer SGC-7901 cells,and the possible mechanism.Methods Methyl thiazol tetrazolium (MTT) method was used to detect antiproliferative rate of LBP on SGC-7901 cells.Flow cytometry was used to analyze the gastric cancer cell apoptosis.Reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting were used to examine the expression of MTDH at mRNA and protein levels respectively.Results LBP significantly exhibited the cell growth inhibition in the SGC-7901 cells in a time-and concentration-dependent manner (P ＜ 0.05).LBP significantly induced apoptosis in SGC-7901 cells in a dose-dependent manner (P ＜ 0.05).MTDH expression was significantly decreased by LBP at mRNA levels,and MTDH protein levels in SGC-7901 cells treated with 0,5,10 and 20 mg/L LBP were 1.046 ±0.072,0.704 ±0.108,0.657 ±0.069 and 0.384 ±0.057 respectively (P ＜ 0.05).Conclusion LBP can inhibit proliferation of SGC-7901 ceils and induce their apoptosis in vitroprobably by suppressing the MTDH expression pathway. Key words: Lobaplatin; Gastric cancer; Proliferation ; Apoptosis ; Metadherin

Expression of Yes-associated protein in gastric adenocarcinoma and effects of Yes-associated protein knockdown on proliferation and metastasis of gastric cancer cells

Objective To examine the expression of Yes-associated protein (YAP) in human gastric adenocarcinomas,gastric adenomas and normal gastric mucosa tissues,and further explore the effects and molecular mechanisms of YAP on proliferation and metastasis of gastric cancer cells.Methods The expression of YAP in 78 cases of gastric adenocarcinomas,39 cases of gastric adenomas and 46 cases of normal gastric mucosa tissues was detected by using immunohistochemistry and real-time polymerase chain reaction (PCR).In vitro human gastric cancer SGC-7901 cells were stably transfected with lentivirus-mediated YAP small hairpin RNA (shRNA),and the expression levels of proliferating cell nuclear antigen (PCNA) and metalloproteinase-2 (MMP-2) were detected by using Western blotting.The proliferative activity and metastatic capability of gastric cancer cells were evaluated by methyl thiazol tetrazolium (MTT) and Transwell assays.Results The expression of YAP was observed in 69.23％ gastric adenocarcinoma tissues,and its mRNA expression level in gastric adenocarcinoma (1.74 ± 0.46) was significantly increased as compared with that in normal gastric mucosa (0.33 ± 0.15) and gastric adenoma (0.42 ±0.20).Knockdown of YAP by shRNA could down-regulate the expression of PCNA and MMP-2,and inhibit the proliferation and metastasis of gastric cancer cells.Conclusion YAP is strongly expressed in gastric adenocarcinoma,and knockdown of YAP inhibits proliferation and metastasis of gastric cancer cells. Key words: Gastric carcinoma; Yes-associated protein; Small hairpin RNA; Proliferation; Metastasis

Effects of serum containing Chinese medicine Sanpi Pingwei (散癖平胃) formula on proliferation and apoptosis of human SGC-7901 cells

To investigate the effects of serum containing Chinese medicine (CM) Sanpi Pingwei (, SPPW) formula on the proliferation and apoptosis of human SGC-7901 cells and the possible mechanism. Serum containing CM SPPW formula (SPPW serum) was prepared by a serum pharmacology method. Human SGC-7901 cells were incubated with SPPW serum at three different concentrations and with the anticancer drug 5-fluorouracil (5-FU), respectively. Cell proliferation was assessed by MTT assay, and cell apoptosis was detected by flow cytometry assay. Real-time quantitative polymerase chain reaction (RT-PCR) and Western blot assay were employed to confirm the expressions of Bcl-2, Bax and p53 in SGC-7901 cells at mRNA and protein levels, respectively. SPPW serum suppressed the proliferation of SGC-7901 cells in a time- and dose-dependent manner. The colony forming rate of negative control was 48.2%, while those in the three SPPW serum groups and the 5-FU group decreased significantly (P<0.01). The number of colony forming units in the SPPW high dosage group was significantly smaller than that in the 5-FU group (P<0.01). MTT assay showed that SPPW serum restrained the proliferation of SGC-7901 cells, and the inhibition rate increased significantly in a dose-dependent manner. Annexin V/PI Assay suggested that SPPW serum induced the apoptosis of SGC-7901 cells significantly. RT-PCR and western blot assay indicated that SPPW serum upregulated the protein and mRNA expression levels of Bax and p53 in SGC-7901 cells, but downregulated the protein and mRNA expressions of Bcl-2. SPPW formula inhibits the proliferation of SGC-7901 cells in vitro and induces the cell apoptosis. It plays an anticancer role by regulating the expressions of Bax, p53 and Bcl-2 in SGC-7901 cells.

Antitumor activity of Xiaoaiping injection on human gastric cancer SGC-7901 cells

Xiaoaiping, extracted from Marsdenia tenacissima (Asclepiadaceae), is a Chinese medicine used to treat cough and asthma. Previous studies have shown that its active ingredients possess anti-inflammatory and anticancer effects. The aim of this study was to investigate the antitumor activities of Xiaoaiping through apoptosis induction in human gastric cancer SGC-7901 cells. The MTT assay was used to assess the cell growth and cell viability. SGC-7901 cells were treated with Xiaoaiping injection [(20–40) mg·mL−1] for 24 and 48 h. The apoptotic cells and the cell cycle distribution were analyzed by flow cytometry. The in vivo activity of Xiaoaiping was determined by the growth inhibition of the established tumor xenografts in nude mice. Caspase-3 activity, Bax and Bcl-2 proteins in tumor tissue were measured by immunohistochemistry and apoptosis was assayed by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) method. Xiaoaiping inhibited SGC-7901 cell growth in a time and dose dependent-manner and the estimated IC50 was (38.20 ± 0.27) mg·mL−1 after 24 h of treatment. The body weight and the tumor volume were significantly reduced in nude mice bearing human gastric tumor treated with Xiaoaiping. The inhibition rate of tumor growth in the mid-dose (200 mg·kg−1) group and high dose (400 mg·kg−1) group were 61.19% and 69.07%, respectively. Immunohistochemical staining showed an increase in caspase-3 and Bax expression whereas Bcl-2 expression decreased gradually. Xiaoaiping exerted potent antitumor activity in vitro and in vivo against human SGC-7901 cells; induced apoptosis and G1 cell cycle arrest. These results suggest that Xiaoaiping is a promising antitumor agent for the treatment of human gastric cancer.

Inhibitory Effects of Ethyl Pyruvate on Invasion and Metastasis of Human Gastric Cancer SGC-7901 Cells viaDownregulation of Akt Pathway

Inhibitory Effects of Ethyl Pyruvate on Invasion and Metastasis of Human Gastric Cancer SGC-7901 Cells viaDownregulation of Akt Pathway

Dihydroartemisinin induces apoptosis of human gastric cancer cells through mitochondrial pathway

Objective To investigate the molecular mechanism of induction of apoptosis by dihydroartemisinin ( DHA ) in human gastric cancer SGC7901 cells in vitro. Methods Cultured SGC7901 cells were treated with DHA at different concentrations (6.25,12.5,25.0,50.0,and 100.0 μmoL/L)for different durations (24,48,and 72 h),and then the proliferation of cells was assayed by using MTT.SGC7901 cells were treated with DHA at different concentrations for 24 h,and the cell apoptosis rate was analyzed by using flow cytometry ( FCM ).The activities of Caspase-3 and Caspase-9 were assessed by colorimetric assay.The change of mitochondrial trans-membrane potential (△Ψm) was measured by JC-1 staining.The expression levels of bcl-2 and bax were detected by using Western blotting.Western blotting was also used to reveal the amount of Cyto C in cytosol.Results Various concentrations of DHA inhibited the growth of SGC7901 cells in a dose- and time-dependent manner.After SGC7901 cells were treated with DHA for 24 h,as compared with control group ( 1.88 ±0.25),the cell apoptosis rate [(4.21 ±0.83)％,( 6.92 ± 1.26 ) ％,( 9.78 ±2.55 ) ％,( 13.93 ± 2.94)％,(16.02±3.83)％] were increased significantly and the activities of Caspase-3 (0.094 ± 0.008,0.196±0.014,0.288±0.017,0.326 ± 0.024,0.491 ±0.042) and Caspase-9 (0.082 ±0.008,0.154 ±0.010,0.176 ±0.012,0.217 ±0.015,0.268 ±0.024)were increased dose-dependently ( P＜0.05 ). As compared with control group ( 100 ),△Ψm (74.18 ±7.84,70.08 ±6.53,58.66 ± 2.38,48.79 ± 1.31,31.24 ± 0.73 ) was declined significantly ( P ＜ 0.05 ).Western blotting showed that DHA increased bax expression and decreased bcl-2 expression ( P ＜ 0.05 ).The ratio of bcl-2/bax (2.37 ±0.24,1.51 ±0.21,0.82 ±0.16,0.64 ±0.14,0.37 ±0.07) was decreased as compared with control group (3.11±0.28).Western blotting also revealed that the amount of Cyto C (0.21 ± 0.05,0.25 ± 0.06,0.38 ± 0.08,0.39 ± 0.09 ) was increased in cytosol as compared with control group (0.18 ± 0.04 ) ( P ＜ 0.05 ).Conclusion DH A induces the apoptosis of human gastric cancer SGC7901 cells dose-dependently,which is probably mediated by the mitochondrial apoptosis pathway. Key words: Dihydroartemisinin; Gastric cancer; Apoptosis; Mitochondria

ERM stable knockdown by siRNA reduced in vitro migration and invasion of human SGC-7901 cells

Three closely related proteins, ezrin, radixin, and moesin (ERM), which primarily act as a linker between the plasma membrane and the cytoskeleton, are involved in many cellular functions, including regulation of actin cytoskeleton, control of cell shape, adhesion and motility, and modulation of signaling pathways. Although, ezrin is now recognized as a key component in tumor metastasis, the functional role of the radixin and moesin in tumor metastasis has not been established. In the present study, we chose highly metastatic human gastric carcinoma SGC-7901 cells, which express all of the ERM proteins as a model to examine the functional roles of these proteins in tumor metastasis. Ezrin, radixin or moesin stable knockdown SGC-7901 cell lines were established using siRNA methodology. In vitro cell migration and invasion studies showed that either ezrin, radixin or moesin specific deficiency in the cells caused the substantial reduction of the cell migration and invasion. Western blotting and immunofluorescence analysis showed that the expression of E-cadherin was also significantly increased when any member of ERM proteins was downregulated. Our results indicated that these three ERM proteins play similar roles in the SGC-7901 cell metastatic potential and their roles of upregulating the expression of E-cadherin may be important in tumor progression.

English

Recombinant retroviral vector containing human tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) gene was constructed and investigation of the in vitro invasion and metastasis of gastric cancer cells transfected with TIMP-2 was carried out. Human TlMP-2 was isolated from recombinant vector Bluescript 1/TIMP-2(+), and then inserted into the retroviral vector pL-MT. Correct orientation was verified by restriction endonuclease digestion. Human full length TIMP-2 gene was ligated into a plasmid, which was then transfected into PA317 cell line. G418-resistant individual clones were selected to transfect human SGC-7901 cell line. Cell proliferation, cell electrophoresis, soft agar colony formation and in vitro invasion were detected to analyze the bio-behavioral changes of cancer cells. The results from restriction endonuclease digestion were as theoretically expected. The cell electrophoresis rate, colony number and invasion ability in SGC-7901 cells and MFC cells transfected with TIMP-2 gene were significantly decreased when compared with control group. However, no significant changes were noted in the proliferation of cancer cells. We successfully construct a recombinant retroviral vector containing human TIMP-2. TIMP-2 transfection could markedly alter the membrane charge of cancer cells, resulting in decreased electrophoresis capacity, cell migration and invasion. However, cell growth was not affected by TIMP-2. These results suggested TIMP-2 transfection might exert effects on the malignant phenotype of cancer cells through affecting extracellular environment, which provided a new way to investigate gene regulation of in vitro collagen metabolism.

English

Recombinant retroviral vector containing human tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) gene was constructed and investigation of the in vitro invasion and metastasis of gastric cancer cells transfected with TIMP-2 was carried out. Human TlMP-2 was isolated from recombinant vector Bluescript 1/TIMP-2(+), and then inserted into the retroviral vector pL-MT. Correct orientation was verified by restriction endonuclease digestion. Human full length TIMP-2 gene was ligated into a plasmid, which was then transfected into PA317 cell line. G418-resistant individual clones were selected to transfect human SGC-7901 cell line. Cell proliferation, cell electrophoresis, soft agar colony formation and in vitro invasion were detected to analyze the bio-behavioral changes of cancer cells. The results from restriction endonuclease digestion were as theoretically expected. The cell electrophoresis rate, colony number and invasion ability in SGC-7901 cells and MFC cells transfected with TIMP-2 gene were significantly decreased when compared with control group. However, no significant changes were noted in the proliferation of cancer cells. We successfully construct a recombinant retroviral vector containing human TIMP-2. TIMP-2 transfection could markedly alter the membrane charge of cancer cells, resulting in decreased electrophoresis capacity, cell migration and invasion. However, cell growth was not affected by TIMP-2. These results suggested TIMP-2 transfection might exert effects on the malignant phenotype of cancer cells through affecting extracellular environment, which provided a new way to investigate gene regulation of in vitro collagen metabolism.