Research Of TCF21Regulate Tumor Metastasis Suppressor Gene Kiss-1Expression In Colorectal Cancer

Objective:1. Analyse the influence affected by tumor metastasis suppressor gene TCF21andKiss-1on Clinicopathological features of colorectal cancer; discusses the correlationbetween their expression.2. Construct an expression plasmid of siRNA against TCF21gene in human colorectalcancer cell line HT-29; to investigate its impact to the expression of Kiss-1gene, andobserve the changes to the cell biological behavior of proliferation, invasion andmigration after inhibited.Methods:1. The expression of TCF21and Kiss-1gene were examined by IHC two step methodin71specimens of colorectal primary cancer tissues together with normal tissuesadjacent to cancer.2. Adopt qRT-PCR method to choose the relatively high expression of TCF21andKiss-1mRNA from colorectal cancer cell lines among Lovo, SW480, HT-29.3. Design three specific siRNA molecule fragments for TCF21, and transfected intoHT-29cells by lipofectamineTM2000; adopt qRT-PCR method to test the expressionlevel of TCF21mRNA after48h, pick out the highest inhibition effective fragment;adopt Western-blot method to test the expression level of TCF21protein after72h,identified the silencing effect of TCF21.4. After TCF21gene successfully silenced, test the expression level of TCF21mRNAand protein at48h and72h after transfection. 5. The Cell Counting Kit-8were used to detect cell proliferation capacity, andTranswell assay were used to detect the invasion and migration capacity aftertransfection for72h.Results:1.The expression rate of TCF21and Kiss-1gene in colorectal cancer primary lesionwere66.20%and70.42%, both lower than the rate of the corresponding normal tissueadjacent to cancer were91.55%and94.37%(P＜0.001); the expression of TCF21andKiss-1gene both relevant to lymph node metastasis, distant metastasis and clinicalTNM stage(P＜0.05), but both had not relationships with patient’s sex, age, degreesof differentiation, pathology type, depth of invasion, cancer embolus in vas, tumorsize and necrotic(P＞0.05);There was a significant positive correlation between theexpression of TCF21and Kiss-1gene in colorectal cancer primary lesion(r=0.450, P＜0.05).2. The expression of TCF21and Kiss-1mRNA in HT-29cells is higher than inSW480and Lovo.3. There were three siRNA transfected groups, the si1group could silence TCF21gene expression successfully, the expression of TCF21gene mRNA and proteinsignificantly declined, with statistically difference between the control group(P＜0.05).4.AT the same time, the expression of Kiss-1gene mRNA and protein in si1groupalso declined, with statistically significant difference between the control group(P＜0.05).5. CCK-8test and Transwell assay showed that cells after transfection proliferatedmore rapidly than those before transfection and had an increased capability ofinvasion and migration(P＜0.01).Conclusions:1. There was a significant positive correlation between the expression of TCF21andKiss-1gene in colorectal cancer. The missing expression of TCF21and Kiss-1gene isclosely related to the occurrence and development of colorectal cancer, and it might increase the risk of distant metastasis.2. Silence the the expression of TCF21gene in colorectal cancer cell lines HT-29,could decline the expression level of Kiss-1gene mRNA and protein in together, andenhance the ability of cell proliferation, invasion and migration.3. Kiss-1gene might serve as a downstream gene accepted TCF21gene transcriptionregulation, and brought into play its effect of metastasis suppression in colorectalcancer...