ABSTRACT
Objective:To verify DNA methyltransferase 1(DNMT1) is the direct target of miR-148a and explore the internal mechanism by which miR-148a regulates the cardiac differentiation of mesenchymal stem cells(MSCs)by directly targeting DNMT1.Methods:miRNA microarray was used to screen out the abnormally expressed miRNAs in MSCs before and after 5′-azacytidine(5′-aza) treatment.Target scan was uesd to predict the target of miR-148a.miR-148a mimics and DNMT1-wt,scramble and DNMT1-wt,miR-148a and DNMT1-mut,scramble and DNMT1-mut were co-transfected in MSCs cells and luciferase activity were detected by single photon.MSCs cells were transfected with miR-148a lentivirus plasmid.Respectively extract DNA,RNA and protein 1,7,14 and 28 days after transfection.The CpG methylation level on DNA regulatory sequences of Gata-4 upstream gene was detected by methylation detection,the mRNA and protein expressions of myosin heavy chain(MHC),cardiac troponin T(cTnT),CD90,CD29,Nkx2.5 and Gata-4 were detected by qRT-PCR and Western blot.Results:miRNA Microarray screened out the abnormally expressed miRNAs in MSCs before and after 5′-aza-induced,including miR-146a-5p,miR-148a,miR-539,etc.Among which miR-148a was remarkably upregulated.Targetscan,Luciferase Reporter Gene and qRT-PCR verified that DNMT1 was the direct target of miR-148a.By infected MSCs cells with miR-148a lentivirus plasmid,we confirmed that the methylation level of Gata-4 gene upstream was changed,and with prolong of transfection,the methylation level of Gata-4 was decreased,the expression of MHC and cTnT were increased,while CD90 and CD2 were continually decreased,the mRNA expression of Nkx2.5 and Gata-4 were increased MSCs cells started myocardial differentiation.Conclusion:miR-148a can regulate the cardiac differentiation of MSCs by directly targeting DNMT1.
ABSTRACT
Objective To understand the mental health status between left‐behind children and un‐left‐behind children during primary school period in Yongchuan district of Chongqing city ,and to provide the theoretical basis for the study of the left‐behind children children′s mental problems and formulating the strategy and measures for preventing their psychological problems .Methods The Mental Health Rate Scale for Pupil (M HRSP) was used to evaluate the mental health condition in 4 987 primary school pu‐pils from 4 primary schools in Yongcuan discrict ,including 3 482 un‐left‐behind children(69 .82% ) and 1 505 left‐behind children (30 .18% ) .Results Totally 4 753 effective questionnaires were taken back with the effective rate of 95 .31% .The detection rate of total score deviation in left‐behind children was 13 .98% ,which was significantly higher than in un‐left‐behind children(6 .33% ) . The deviation detection rate of learning disabilities in left‐behind children was 14 .90% ,emotional disorder was 16 .81% ,character flaw was 12 .36% ,social adjustment disorder was 13 .91% ,moral defect was 14 .27% ,bad habits was 15 .61% ,behavior disorder rate was 16 .53% and the special obstacles was 11 .72% .Compared with un‐left‐behind children ,excluding the character flaw ,social adjustment and moral defect ,the deviation detection rate and scores of other 5 items in left‐behind children were higher than those in un‐left‐behind children .The scores of learning disabilities ,emotional disorder and bad habits in male pupils were higher than those in female pupils with statistical difference(P< 0 .05) .The scores of learning disabilities ,emotional disorder ,character flaw and behavior disorder in the high grade pupils were higher than those in the low grade pupils ,the differences were statistically sig‐nificant(P<0 .05) .Conclusion The mental health level of rural left‐behind children in Yongchuan district is low ,the problems are more serious compared with un‐left‐behind children .In carrying out the mental health education work in primary school pupils ,es‐pecially the mental health education should be paid attention to rural left‐behind children ,moreover the mental health education should be carried out aiming at different ages and different genders .