The Experimental Study Of MiRNA In Pituitary Adenomas

Pituitary adenomas is a high incidence and account for about 10% of all cranial tumors. It is very difficult to be completely evacuated. Some of patients with pituitary adenomas may be suffered recurrance. Exploring pituitary adenoma pathogenesis and new non-operative theraputic means play very important role in clinical practice.MicroRNAs (miRNAs) constitute a recently discovered class of small non-coding RNAs that regulate expression of target genes either by decreasing the stability of the target mRNA or by translational inhibition. They are involved in diverse processes, including cellular differentiation, proliferation and apoptosis. Recent evidence also suggests their importance for cancerogenesis.This dissertation consists of three parts: The study of miRNA gene expression in pituitary adenomas;The study of miRNA by RT-PCR in pituitary adenomas;The study of target genes of miRNA in the pituitary adenomas.Part one. The study of miRNA gene expression in pituitary adenomas Object To investigate on the miRNA in the pituitary adenoma,establish the expression map for non-functioning pituitary adenomas and gonadotropin-secreting pituitary adenomas. And try to find out the up-regulated or down-regulated genes. Method The total RNA was extracted from 4 non-functioning pituitary adenomas and gonadotropin-secreting pituitary adenomas, 1 normal pituitary tissue and then purified. The samples were analysised by gene chip of Capitalbio Corporation. Results Of the non-functioning pituitary adenomas, 25 miRNA genes wereup-regulated, and 15 were down-regulated. 5 genes changed over 5-fold: miR-124a, miR-146a, miR-523, miR-lOb, PREDICTED_MIR240, PREDICTED_MIR207, last two were suspected by computer. MiR-124a was the most up-regulated gene, 38.58 fold. 6 miRNA genes down-regulate more than 10-fold: miR-3K miR-514* miR-503. miR-506 > miR-513. miR-218, miR-31 was the most dowm-regulated gene, 21.5 fold;of the gonadotropin-secreting pituitary adenomas, 16 miRNA genes were up-regulated, and 13 were down-regulated. 6 genes changed over 4-fold: miR-lOb, miR-523, miR-222, miR-422b, PREDICTEDJ1IR189, PREDICTED_MIR240, last two were suspected by computer. MiR-lOb was the most up-regulated gene, 48.73 fold. 7 miRNA genes down-regulate more than 10-fold: miR-503, miR-508, miR-514, miR-31, miR-506, miR-513, miR-509, miR-503 was the most dowm-regulated gene, 39.8 fold. Of two subtypes, 5 genes were up-regulate: miR-523, miR—10b, miR-520b, miR-422a, miR-422b. Conclusions MiRNA expressing map of non-functioning pituitary adenomas and gonadotropin-secreting pituitary adenomas were established. MiR-124a and miR-31 were the most special genes in the non-functioning pituitary adenomas. MiR-lOb and miR-503 were the most special genes in the gonadotropin-secreting pituitary adenomas.Part two. The study of miRNA by RT-PCR in pituitary adenomas Object To Verify the result of miRNA chip of miR-124a and miR-10b, as the most up-regulated in the genes expressing map of the non-functioning pituitary adenomas and gonadotropin-secreting pituitary adenomas Methods Gene expression was measured using real time RT—PCR with SYBR GREEN I. U6 was used as internal control. The total RAN was origin of the last experiment. Results Standard curves were linear and showed highcorrelations. It is confirmed that results of RT-PCR was consistent with the gene chips. Conclusions MiR-124a and miR-10b were up-regulate obviously in the non-functioning pituitary adenomas and gonadotropin-secreting pituitary adenomas, they may be the special genesof these subtypes.Part three. The bioinformatic study of target genes of miRNA in the pituitary adenomas.Object To investigate on the target genes of the miRNA. Methods To enquiry the target genes of miRNA in the internet with the PicTar, TargetScan.. MiRanda. Results There were 542 target genes for miR-124a, some of them were related with the cancerogenesis of pituitary adenomas. They were histone, platelet activating factor acetylhydrolase , STAT5B, TOB2, RIN2, RAS, and so on. There were 468 target genes for miR--10b, some of them were related with the cancerogenesis of pituitary adenomas, they were HOXA3, H0XD10, PTEN, RB1CC1, USF, E2F3, HNRPK. Conclusions Some target genes of miR-124a and miR-10b were related with the cancerogenesis of pituitary adenomas, part of them had direct effect for cancerogenesis, another had indirect effect.