Decrease Expression Of TRIM29 In Proliferation Of Gastric Cancer Cell

BackgroundGastric cancer is the second leading cause of cancer deaths in the world. In order to obtain better diagnosis and treatment of gastric cancer, it is essential to determine the cause and mechanism in its gene and development of the disease. A series of studies have shown that, TRIM29 was a crucial oncogene in gastric cancer, and patients with TRIM29 mRNA overexpression had poor prognosis and a lower survival rate. However, the mechanism of action of TRIM29 in gastric cancer still remains unclear. A specific RNA interference technology in recent years found a high and simple operation in RNA gene transcription blocking technology. In this study, we will attempt to use Real time qRT-PCR, Western-blot and RNA interference to detect TRIM29 and its protein expression levels in human gastric cancer, so as to explain the relationship between TRIM29 expression in patients with gastric cancer and the clinical prognosis, provide the basis as well.Objective1. To investigate the relationship between TRIM29 expression and prognosis in patients with gastric cancer.2. To analyze the mechanism of action of TRIM29 by siRNAtransfected plasmid and decrease expression levels of TRIM29 mRNAin gastric cancer cell.3. By analyzing decrease overexpression TRIM29 mRNA level in tumor cellsto explore the effect of changing expression levels of TRIM29 in gastric cancer cell in nude mice experiments.Methods1. From March 2010 to January 2015,38 patients with gastric adenocarcinoma in Henan Cancer hospital were collected and divided into neoadjuvant chemotherapy group (16 cases) and direct surgery group (22 cases). Preoperative endoscopic biopsy specimens, standard D2 gastriectomy of primary gastric cancer specimens, lymph node specimens and normal mucosa specimens were collected. TRIM29 mRNA and protein levels in the above samples were compared and analyzed.2. We made the detection of the mRNA and protein expression levels of TRIM29 in different gastric cancer cell lines, and the overexpressed cell lines were selected. After transfection siRNA plasmid, real time qRT-PCR and Western-blot were used to detect the expression levels of TRIM29, P53 and B-catenin; MTT detection was used to detect cell doubling time and growth curve. Cell cycle distribution was used by flow cytometry analysis before and after the disturbance.3. One certain gastric cancer cell line which overexpressed TRIM29 was selected. After that, they were transfected with siRNA plasmid and transplanted subcutaneously into nude mice, in order to observe the size and growth of transplanted tumor. Finally, the expression levels of TRIM29 in tumors were detected.Results1. Through the Real-time qRT-PCR and Western blot, we found TRIM29 mRNA expression and protein levels were significantly higher in primary gastric cancer specimens, specimens of positive lymph nodes than normal gastric mucosa and negative lymph node specimens (P<0.001). In direct operation group and neoadjuvant chemotherapy group, mean survival time in patients with TRIM29 positive expression were significantly lower than TRIM29-negative patients (Log-rank X2= 4.068 expression; P= 0.044; Log-rank X2= 7.487; P= 0.006). TRIM29 mRNA expression level after neoadjuvant chemotherapy was significantly lower than that of chemotherapy patients before chemotherapy.2. TRIM29 mRNA and protein levels in the gastric cancer cell lines were found to have increased. P53 expression in different gastric cancer cell line with positive TRIM29 have shown a significant reduction, and B-catenin appeared abnormal expression. After RNA interference, in siRNA transfected plasmid gastric cancer cell lines, TRIM29 mRNA and protein expression levels had decreased, as well as significantly lower than siRNA plasmid blank plasmid transfected cells. Furthermore, p53 mRNA and protein expression level was conspicuously higher than that of RNA interference before. B-catenin mRNA and protein expression levels were significantly lower than before the disturbance. Compared with the previous RNA interference, after RNA interference, cell proliferation significantly decreased in TRIM29 over-expression gastric cancer cell lines.3. Within 60 days of metastases in five nude mice, gastric cancer cells with TRIM29 expression before siRNA plasmid transfection were shown tumor grown. Tumor diameter was 0.4-1.9cm, and after siRNA plasmid in gastric cancer cells, four nude body was found solid tumors, with diameter 0.1～0.5cm (P<0.001), and one was not found visible solid tumors. TRIM29 mRNA and protein levels before siRNA plasmid transfection in gastric cancer cells in the metastatic tumor formation in nude mice was significantly higher than after siRNA plasmid transfection.Conclusions1. TRIM29 mRNA and protein expression levels significantly improved in patients with gastric cancer; Survival time in positive TRIM29 expression patients is shorter and the prognosis is even worse. Neoadjuvant chemotherapy can affect the expression levels of TRIM29 to a certain extent.2. TRIM29 downregulate p53 and enhance β-catenin levels to achieve enhanced tumor cell proliferation biological effects.3. In vitro experiments, we further demonstrate TRIM29 can improve gastric cancer cell proliferation.