ABSTRACT
Objective To analyze the expression of miRNA-29a in U937 macrophages infected with Mycobacterium tuberculosis and its regulation of target genes.Methods The target genes of miRNA-29a were predicted with softwares miRnada,RNAhybrid and targetscan.The differentiation of U937 macrophages was induced by phorbol ester(PMA), the induced U937 cells were infected with bacilli calmette guerin (BCG).The expression levels of miRNA-29a and its target genes in U937 cells were detected with real-time fluorescence quantitative PCR(RT-fqPCR).The miRNA-29a over-and low-expression U937 macrophage cell lines were constructed and the levels of miRNA-29a and its garget genes were detected.The SPSS 18.0 software was used to analyze the data.Results As predicted by relevant softwares,the miRNA-29a regulate the expression of VEGFA,NFATC3 and TSC22D3 genes.After BCG infection,the expression of miRNA-29a increased to 1.33 fold(P <0.05), and the expression levels of VEGFA, NFATC3 and TSC22D3 were increased to 5.34,99.25 and 2.12 fold,respectively(P<0.01).In the miRNA-29a over-expressing U937 macrophages,the expression levels of VEGFA,NFATC3 and TSC22D3 were up-regulated to 1.35,1.29 and 3.38 fold,respectively(P<0.05 or <0.01).While in the U937 macrophages with miRNA-29a knock-down,the expression levels of VEGFA, NFATC3 and TSC22D3 were down-regulated to 0.07, 0.08 and 0.55 fold, respectively(P <0.01).Conclusion The results suggest that Mycobacterium tuberculosis infection can increase the expression of miRNA-29a in U937 macrophages,further targeting the regulation of VEGFA, NFATC3 and TSC22D3 gene expression, which may participate in the pathogenesis and development of tuberculosis.
ABSTRACT
Objective To detect the expression of miRNA‐29a in abdominal aortic aneurysm(AAA) patients and construct a recombinant lentiviral vector carrying miRNA‐29a ,transfection of human abdominal aortic vascular smooth muscle cells (HVSMCs) ,and to investigate the alteration on the proliferation in HVSMCs .Methods Collected 25 AAA patients′serum and 25 normal elderly serum specimens from March 2011 to March 2014 ,the mean age were 62 .9 ± 13 .6 and 61 .5 ± 11 .8 separately .Em‐ploy RT‐PCR analysis the miRNA‐29 expression in different group .The MCS‐CMV‐miRNA‐29a was transfected into HVSMCs cells ,the transfection efficiency was observed with fluorescence microscope ,the mRNA and COL1A1 protein expresstion was detec‐ted by RT‐PCR and Western blot respectively .MTS method was used to determine effects of cell inhibition rates of different groups ,cell apoptosis was analysised by Annexin V/7‐AAD double staining after 72 h .Results The expression of miRNA‐29a with AAA was lower than normal group .The transfection efficiency was (74 .25 ± 10 .10)% .miRNA‐29a mRNA and its target protein COL1A1 expression increased significantly compared with the control group after transfection ,MTS and Annexin V/7‐AAD results showed that cell apoptosis was decreased after miRNA‐29a overexpression ,cell proliferation was increased compared with the con‐trol group .Conclusion The miRNA‐29 expression decreased associated with the occurrence of abdominal aortic aneurysm ,but overexpress miRNA‐29a in HVSMC cells by lentivirus mediated ,which could promote the proliferation of cells ,inhibit its apoptosis .