Establishment of BGC-823/pBaBb-puro-MACC1 gastric cancer cell line stably expressing MACC1 and its tumor-related gene expression profiles / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a gastric cancer cell line with stable expression of metastasis-associated in colon cancer 1 (MACC1) and detect the changes in tumor-related gene expression profiles for investigating the possible regulation mechanisms between MACC1 and the differentially expressed genes.</p><p><b>METHODS</b>The full-length MACC1 cDNA was amplified from human embryonic kidney 293FT cells and cloned into the pBaBb-puro vector. The recombinant pBaBb-puro-MACC1 expression vector, after identification with restriction enzyme digestion, was transfected into 293FT cells, and the expression of fluorescent reporter gene was observed. pBaBb-puro-MACC1 vector was transfected into human gastric cancer BGC-823 cell line to establish BGC-823/pBaBb-puro-MACC1 cell line stably expressing MACC1. Quantitative RT-PCR and Western blotting were used to detect MACC1 expression in both BGC-823/pBaBb-puro-MACC1 and control BGC-823 cells. High-throughout cDNA microarray was used to screen the effects of MACC1 on the gene expression profiles of gastric cancer cells.</p><p><b>RESULTS</b>The recombinant pBaBb-puro-MACC1 plasmid was successfully constructed and verified by PCR and sequencing. BGC-823/pBaBb-puro-MACC1 cells showed significantly increased MACC1 mRNA expression as compared with the control cells. The results of cDNA microarray identified 33 up-regulated and 24 down-regulated genes in the cells after MACC1 transfection involved were in various cellular functions.</p><p><b>CONCLUSION</b>The established BGC-823/pBaBb-puro-MACC1 gastric cancer cell line show some important molecular changes caused by MACC1.</p>

Effect of anti-EGFR monoclonal antibody on chemosensitivity of human colon cancer cells and the mechanism / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of anti-EGFR monoclonal antibody on the chemosensitivity of human colon cancer cells and explore the possible molecular mechanism.</p><p><b>METHODS</b>The inhibitory effect of irinotecan (CPT-11), oxaliplatin (L-OHP) and fluorouracil (5-Fu), used alone or in combination with anti-EGFR monoclonal antibody, on the proliferation of LoVo cells in vitro was assessed by MTT assay. The expressions of PI3K and Akt protein in the treated cells were examined by Western blotting, and their mRNA expressions were detected by RT-PCR.</p><p><b>RESULTS</b>Both h-R3 and C-225 treatments significantly increased the chemosensitivity of LoVo cells to irinotecan and oxaliplatin. 5-Fu and h-R3 coadministered showed a synergistic effect on the cells, but 5-Fu and C-225 had an antagonistic action. Treatment with C-225 or h-R3 resulted in lowered expressions of PI3K and Akt in LoVo cells.</p><p><b>CONCLUSION</b>Anti-EGFR monoclonal antibody can increase the chemosensitivity of human colon cancer cells to most chemotherapeutic drugs, and such effect might be attributed to the blocking of PI3K/Akt signaling pathway by these antibodies.</p>

Sorafenib reverses multidrug resistance of hepatoma cells in vitro / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the role of sorafenib in reversing multidrug resistance (MDR) in hepatoma BEL-7402/FU cells and its possible mechanisms.</p><p><b>METHODS</b>MTT colorimetric assay was used to obtain the dose-response curve of sorafenib in BEL-7402/FU cells, and flow cytometry performed to assess the effect of sorafenib on Rho123 concentration in the cells. The optimal dose of sorafenib for cell treatment was determined according to the results of MTT assay and flow cytometry. MTT assay was employed to evaluate the effect of sorfenib on the cytotoxicity of the antitumor drugs, flow cytometry performed to determine the expression of cell membrane transport protein (P-gp), and RT-PCR used to detect mdr1 gene expression in the cells treated with sorafenib at the optimal dose.</p><p><b>RESULTS</b>Sorafenib at the concentration of 4 micromol/L, efficiently reversed the MDR of the cells with minimal side effects. At the concentration of 4 micromol/L, sorafenib partially reversed the drug resistance of BEL-7402/FU cells to ADM, 5-FU, GEM and DDP, with reversal indexes of 2.98, 7.16, 1.99 and 10.08, respectively. Treatment of the cells with 4 micromol/L, sorafenib also partially down-regulated P-gp expression in BEL-7402/FU cells, and caused a reduction of mdr1 gene expression by 27.3% in comparison with the control cells.</p><p><b>CONCLUSION</b>Sorafenib can reverse MDR in human hepatoma cells probably in association with down-regulation of mdr1 gene expression and increased accumulation of the chemotherapeutic agents in the cells.</p>

Effects of bevacizumab and cisplatin on human lung adenocarcinoma A549/DDP xenografts in nude mice / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the effects of bevacizumab with or without cisplatin (DDP) on the growth of lung adenocarcinoma A549/DDP cell xenografts in mice.</p><p><b>METHODS</b>Human lung cancer A549/DDP cells was subcutaneously transplanted in to 25 nude mice, which were randomly divided into control group (group A), bevacizumab group (group B), DDP group (group C), combined treatment group (group D) and half-dose combined treatment group (group E). After corresponding treatments for 4 consecutive weeks, the tumor inhibition rate was evaluated, tumor microvessel density (MVD) measured with immunohistochemistry, and the mRNA expression of apoptosis-associated gene (bcl-2) and multidrug resistance genes (LRP and GST-pi) assessed by RT-PCR.</p><p><b>RESULTS</b>The tumor growth inhibition rates in groups B, D, and E with bevacizumab treatment were 20.96%, 51.67% and 50.95%, respectively, and the two combined treatment groups showed better effects. MVD in these 3 groups were 18.6-/+1.14, 13.6-/+1.14, and 14.4-/+0.55, respectively, and no significant difference was found in MVD between DDP group and the control group. Compared with the control group, the 3 bevacizumab-treated groups showed decreased expression of bcl-2 genes in A549/DDP tumors at a comparable amplitude, and LRP and GST-pi mRNA expression showed no significant differences between the 5 groups.</p><p><b>CONCLUSION</b>Bevacizumab has synergetic inhibitory effect with conventional chemotherapy against lung adenocarcinoma A549/DDP cell xenografts in mice by inhibiting angiogenesis of the tumor, and may enhance the sensitivity of A549/DDP cells to DDP by inducing cell apoptosis.</p>

Avastin combined with cisplan inhibits malignant ascites production in nude mice bearing transplanted ovary carcinoma with high VEGF expression / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a nude mouse model of malignant ascites with human ovarian carcinoma cell line OVCAR3 which highly expresses VEGF and evaluate the therapeutic of Avastin combined with cisplan.</p><p><b>METHODS</b>Forty-eight nude mice with malignant ascites resulting from intraperitoneal transplantation of human ovarian carcinoma cell line OVCAR3 were treated with intraperitoneal injection of Avastin, cisplan, their combination, and PBS, respectively, to observe the effect on ascites development, VEGF content in the ascites, peritoneal permeability, development of new vessels and number of tumor cells in the ascites.</p><p><b>RESULTS</b>Avastin obviously inhibited ascites accumulation and peritoneal capillary permeability, reduced VEGF protein level and microvascular density in the tumor tissues and the number of red cells and tumor cells in the malignant ascites, and prolonged the survival of the mice. The combination of Avastin and cisplan further enhanced the therapeutic efficacy of Avastin.</p><p><b>CONCLUSION</b>The bio-chemotherapeutic strategy with Avastin combined with cisplan can be a promising method for treatment of malignant ascites.</p>

Adsorption condition optimization for anti-HBsAg Fab fragment separation and purification from E. coli using Streamline SP / 南方医科大学学报

<p><b>OBJECTIVE</b>To optimize the adsorption condition of cation-exchange chromatographic media Streamline SP for separation and purification of anti-HBsAg Fab fragment from E. coli.</p><p><b>METHODS</b>The adsorption of the target protein for separation and purification by the cation-exchange chromatographic media Streamline SP was tested using test tube method in balanced buffer solution with different pH values and ion concentrations. The adsorption effect was then verified by cation-exchange chromatography using 1-ml Streamline SP prepacked column and 28-ml Streamline SP self-assembly column.</p><p><b>RESULTS</b>According to the experiment results of test tube method, the loading buffer with pH of 4.4 and ionic concentration of 100 to 600 mmol/L could achieve optimal target protein adsorption effect by cation-exchange chromatographic media Streamline SP, as verified by cation-exchange chromatography with 1-ml SP prepacked column and 28-ml Streamline SP self-assembly column.</p><p><b>CONCLUSION</b>The optimal condition of cation-exchange chromatography selected by test tube method can be applied for separation and purification of anti-HBsAg Fab fragment from E. coli.</p>