Studies On The Differential Expressions And Function Of MicroRNAs Associated With The Pathogeny Of AIMAH And Adrenal Adenoma

Purpose:To investigate the differential expressions of microRNAs which are involved both in the AIMAH and adernal adenoma through evaluating their levels in the tumor specimens,compared with the normal adrenal cortex. Furthermore, the function of specific microRNAs was probed in SW-13cells preliminarily and the potential targets were screened through bioinformatic database.Materials and methods:There were57cases in study group, including AIMAH40cases, adernal adenoma7cases, and normal adrenal gland10cases.In the first part, we found the differential expressions in5cases with AIMAH, campared with normal adrenal cortex in5cases, by using microRNAs microarrays. Then, We identified the same differential expressions in the AIMAH (40cases) and adernal adenomas(7cases), by using the fluorogenic quantitative PCR.In the second part, we studied the function of the microRNAs and the target genes preliminarily. On the base of the first part of the study, we made the microRNA mimics and inhibitors, and mediated the the mimics and inhibitors into the SW-13cell line,then identified the effects of the mediation by fluorogenic quantitative PCR. We watched the proliferating of the cells by MTT and the apoptosis by Annexin V-FITC/PI. Based on the results, we choosed the microRNAs and searched the potential target genes of those microRNAs, though the targetscan (http://www.targetscan.org/) and miRBASE databases (http://www.mirbase.org/)Result:The first part, we found there were12different microRNAs between normal adrenal gland and AIMAH, including7up-regulating and5down-regulating microRNAs. Up-regulating microRNAs included:hsa-miR-663a, hsa-miR-498, hsa-miR-638, hsa-miR-501-5p, hsa-miR-585, hsa-miR-557and hsa-miR-144.Down-regulating microRNAs included hsa-miR-744,hsa-miR-143,hsa-miR-26a,hsa-miR-22,hsa-miR-29a. Then indentified the microRNAs both in the AIMAH and the adernal adenomas,we found there were4differential expressions both in the two diseases,compared with normal adrenal glands, including3up-regulating and1down-regulating microRNAs. Up-regulating microRNAs included hsa-miR-663a, hsa-miR-498, hsa-miR-557. Down-regulating microRNAs included hsa-miR-744-5p.The second part,we overexpressed all the4microRNAs found in the first part,by mediating the4microRNAs mimics into the SW-13cell line and knocked down the microRNAs by mediating the4microRNAs inhibitors into the SW-13cell line. By fluorogenic quantitative PCR,we identified the effects of the cell interferences,we found the microRNAs overexpressed in the mimics group and got down in the inhibitor group in SW-13cell line. We found more cell prolifering by ovexpressing hsa-miR-663a and less prolifering by koncking down hsa-miR-663a and hsa-miR-557. By the Annexin V-FITC/PI,we found more cell apoptosis,when overexpressing hsa-miR-744; more cell apoptosis,when knocking down hsa-miR-663;less cell apoptosis,when knocking down hsa-miR-577and hsa-miR-498.We choosed the hsa-miR-663a as the target microRNAs, searched the possible target mRNA though databases. There were5possible target genes in both databases, including DPF1,GRIN2D,PRRT1,SLC17A7and ZNF385A. We used the NCBI to find the ralated reports of the5genes.All the datas were managed by SPSS13.0. and when p<0.05, the difference was signiacant in t-test.Conclusion:Compared with normal adrenal glands, there were the same differential expressions of the microRNAs in both AIMAH and the adrenal adenoma. When hsa-miR-663a exprssion overexpressed, the cells prolifered more quickly, and knocked down it,the cells prolifered slowed down.The possible target genes were DPF1、GRIN2D、 PRRT1、SLC17A7and ZNF385A. According to the reportes, SLC17A7gene played a role in the releasing hormone, but we need further study to confirm.