ABSTRACT
Objective To investigate the effect of has-miR-663 on proliferation and apoptosis of human adrenal cortical cell line SW-13.Methods Based on the first stage study,in adrenocorticotropinindependent macronodular adrenal hyperplasia (AIMAH),hsa-miR-663 was up-regulated.The synthetic hsa-miR-663 mimics and inhibitors were transfected the SW-13 cell line.Real-time quantitative PCR (RT-PCR) tested the effect of the transfection.According to the results of RT-PCR,cell line was divided into five groups,including blank group,mimics NC group,inhibitor NC group,mimics group,inhibitor group.Then proliferating of the cell was tested by MTT and the apoptosis by Annexin V-FITC/PI.Result By RT-PCR,mimics group showed that has-miR-663 was over-expressed,and inhibitor group were knocked down.MTTshowed that,compared with NC group and Blank group,SW-13 cell line proliferated faster in mimics group and slower in inhibitor group.Annexin V-FITC/PI showed that apoptosis of SW-13 cell line was not different in all the five groups.Conclusion Up-regulating has-miR-663 could accelerate proliferation of adrenal cortical cells.The microRNA might play some role in the pathogenesis of AIMAH.
ABSTRACT
Objective To investigate the different expressed microRNA in adrenocorticotropinindependent macronodular adrenal hyperplasia (AIMAH).Methods For the screening different expressed microRNA in AIMAH,5 AIMAH fresh tissues and 5 normal adrenal fresh tissues were collected.Total RNA was extracted by using Trizol reagents,then was purified and tested by denaturing agarose gel electrophoresis and NanoDrop 2000.Then the purified RNA was labeled by using Hy3TM fluorescent label and hybridized with miRCURY Array.MicroRNA array was scanned and analyzed by GenePix 4000B microarray scanner and GenePix proV6.0.The ratio acquired by divided AIMAH fluorescent correction value by normal adrenal fluorescent correction value.If the ratio was greater than 3.0,the microRNA was defined as up-regulating,or smaller than 0.3,as down-regulating.For the validation 40 AIMAH samples and 10 normal adrenal samples were used.The miRCURY array result was validated by real-time quantitative PCR.All the data was managed by SPSS 17.0.When P ＜ 0.05,the difference was significant in Student's t test.Results There were 12 different expressed microRNA between normal adrenal gland and AIMAH,including 7 up-regulating and 5 down-regulating microRNAs.Up-regulating microRNAs included hsa-miR-663,hsa-miR-498,hsa-miR-638,hsa-miR-501-5p,hsa-miR-585,hsa-miR-557 and hsa-miR-144.Down-regulating microRNA included hsa-miR-744,hsa-miR-143,hsa-miR-26a,hsa-miR-22,hsa-miR-29a.Four different expressed microRNAs were identified in validation test,including 3 up-regulating and 1 down-regulating microRNA.Upregulating microRNA included has-miR-663,has-miR-498,and has-miR-557.Down-regulating microRNA was has-miR-744.Conclusion Compared with normal adrenal gland,there might be some different expressed microRNAs in AIMAH.