Preliminary Identification Of Mirna Associated With Gastric Cancer And The Effect Of Mmc On The Mirna Expression Profile In Gastric Cancer Cells

Objective Through identification of the closely related expression of miRNAin human gastric cancer cells which have variant differentiation degree，this studydiscusses the possible pathogeny. Meanwhile，it preliminarily analyze the effect ofMMC on miRNA expressions profile in gastric cancer cells and has laid sometheoretical foundation for further studying the molecular mechanism that MMC cantreat gastric cancer.Methods Culture human gastric cancer cell line SGC-7901，MKN-45andnormal gastric epithelium cell GES-1in vitro; The inhibition rate of SGC-7901proliferation were detected with MTT assay; The cell line SGC-7901was intervenedwith MMC in concentration of IC50which is calculated through the MTT assay;Extract the total RNA of all these cells and Test the quality of total RNA; Differentialexpression of miRNA were filtered using miRNA microarray technoloygy in the cellline SGC-7901，MKN-45and SGC-7901was intervened with MMC. Some ofdifferentially expressed miRNA were verified by qRT-PCR; Potential target genes ofmiR-498were predicted by DIANAmT，MiRanda，MiRDB，MiRWalk，PICTAR4，PITA，RNA22，RNAhybrid and Targetscan softwares.Results1.Compared to GES-1，the results of miRNA microarray detectionshowed that there were119miRNA with twice and more different expression inSGC-7901and MKN-45. There were30miRNA significantly up-regulated and25significantly down-regulated in SGC-7901. And there were58miRNA significantlyup-regulated，and33significantly down-regulated in MKN-45. There is an obviouslydifference in the miRNA expression profile between SGC-7901and MKN-45. 2.MMC inhibited the proliferation of gastric cancer cellSGC-7901in a dosedependent manner. IC50of MMC is1.33μmol/L at48h;It was revealed by miRNAmicroarray detection that there were60miRNA with twice and more differentexpression in SGC-7901intervened with MMC at the concentration of IC50compared to SGC-7901，of which miR-301a，miR-498，miR-18b，miR-196b，miR-1247ect，were significantly up-regulate（these miRNA were down-regulated inSGC-7901)， and miR-205*， miR-3924were significantly down-regulated(thesemiRNA were up-regulate in SGC-7901).3.Selecting miR-33b，miR-498，miR-7，miR-185and miR-3924to a real-time quantitative RT-PCR validation，the results areconsistent with of miRNA microarray results，reveal that results of the microarraywere credible.4. DIANAmT，MiRanda，MiRDB，MiRWalk，PICTAR4，PITA，RNA22，RNAhybrid and Targetscan softwares analysis showed that the oncogeneMDTH is one of the miR-498’s target genes.Conclusions1.The research shows that there is different expression profile ofmiRNA in gastric cancer cells，and there is also different expression profiles ofmiRNA in gastric cancer cells with variant differentiated expression.2.2.MMC canchange part of the miRNAs’ expression level in SGC-7901cells..3.The regulation ofMMC on the expression of some miRNA including miR-498may be one of the wayon inhibiting cell growth of gastric cancer.