Proliferation Level Variation Of Colon Cancer HCT-8Cells And Tumor By RNA Intefrerence And Overexpression Targeting PSME2Gene

Colon cancer is one of the high-risk human malignant tumors, which incidence hasbeen on the rise in recent years. The incidence has risen to the third, and mortality hasrisen to the second in the global malignant tumors. In our country, the mortality ofcolorectal cancer has been the fifth in malignant tumors mortality, and it seriouslyharms human health. In the process of the development of colon cancer, it ofteninvolves a variety of changes in protein expression, which can be used as tumormarkers of colon cancer occurrence, development and prognosis. Studies have foundthat Homo sapiens proteasome （prosome, macropain） activator subunit2（PSME2orPA28β） can lead to cell proliferation and change of malignant degree, its decline isassociated with the occurrence and development of gastric adenocarcinoma. At thesame time, studies have shown that PSME2gene expression in liver cancer, breastcancer and kidney cancer organizations can also be changed, there have been reportedthat PSME2low expression in colon cancer. This experiment make PSME2gene asthe breakthrough point, discussed its RNA interference and high expression impact onthe proliferation of colon cancer, lay a foundation for tumor immunotherapy.This experiment build RNA interference and overexpression vector through theway of gene cloning and prokaryotic expression, and transfect cells. mRNAexpression level of PSME2gene was detected by Real-time PCR in colon cancerHCT-8cells; transcriptional level of PSME2gene was detected by Western-blottingin colon cancer HCT-8cells; proliferation of RNA interference and overexpression ofcolon cancer HCT-8cells was determined by MTT; proliferation of RNA interferenceand overexpression of colon cancer HCT-8tumor by animal tumor model. Real-time PCR result display, in the RNA interference plasmid transfection groups （U6/GFP/Neo-PSME2-homo-1、U6/GFP/Neo-PSME2-homo-2、U6/GFP/Neo-PSME2-homo-3、U6/GFP/Neo-PSME2-homo-4） mRNA expression level of PSME2gene in coloncancer HCT-8cell decreased (2^－ΔΔCT<0.5); in the overexpression plasmid transfectiongroup （pEGFP-C1-PSME2）, mRNA expression level rised (2^－ΔΔCT>2); and themRNA expression level of the negative control plasmid transfection groups（pEGFP-C1、 U6/GFP/Neo-shNC、 U6/GFP/Neo-shhGAPDH） had no significantdifference with the blank control group (2^－ΔΔCT>0.5). The results of Western-blottingshowed that expression quantity of PA28β that is translated by PSME2gene of coloncancer HCT-8cells decreased in RNA interference plasmid transfection groups;expression quantity of PA28β rised in overexpression plasmid transfection group; andthere was no significant difference between negative control groups and blank controlgroup. At the same time, studied the changes of cell proliferation in each groups byMTT method, the results showed that cell proliferation of RNA interference groupwas obviously faster than the blank control group （P<0.05）, overexpression plasmidgroup cell proliferation was slower （P<0.05）, negative control group and blank controlgroup had no significant difference. In animal experiment, we successfully establishedcolon cancer HCT-8cells in nude mice transplantation tumor model. The results oftreatment experiment showed that the high expression of PSME2genes obviouslymade HCT-8cells in nude mice transplantation tumor reduced （P<0.05）, whichinhibition rate was56.77%; and HCT-8cells in nude mice transplantation tumorgrowth rate in PSME2gene RNA interference group were significantly faster thanblank control group.