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Analysis of Combinatorial miRNA Treatments to Regulate Cell Cycle and Angiogenesis
Published on: March 30, 2019
Roles of miRNA-24 in regulating endothelial nitric oxide synthase expression and vascular endothelial cell
Wenyu Zhang1, Limei Yan, Yumei Li
1College of Pharmacy, Guangxi Medical University, No. 22, Shuangyong Road, Nanning, 530021, People's Republic of China.
Abstract:
This study is to investigate the effect of miRNA-24 on endothelial nitric oxide synthase (eNOS) expression and vascular endothelial cell proliferation. Constructed high expression miRNA-24 plasmids were introduced into human umbilical vein endothelial cells (HUVECs) by liposome. Cell numbers were counted by a Hemocytometer, and cell proliferation was detected by MTT assay. The expression levels of eNOS and specificity protein 1 (Sp1) at mRNA and protein levels were, respectively, examined by real-time PCR, immunohistochemistry, and Western blotting. Compared with the control group, endothelial cell proliferation in miRNA-24 group significantly decreased by 64.24 % (6.53 ± 0.11 vs 18.26 ± 0.45, P < 0.01). The expression of eNOS at mRNA and protein levels in miRNA-24 group decreased by 64.21 % (0.34 ± 0.01 vs 0.95 ± 0.02, P < 0.05) and 82.86 % (0.072 ± 0.06 vs 0.42 ± 0.06, P < 0.05), respectively. Meanwhile, the expression of Sp1 at mRNA and protein levels in miRNA-24 group decreased by 64.64 % (0.35 ± 0.01 vs 0.99 ± 0.03, P < 0.05) and 60.34 % (0.23 ± 0.05 vs 0.58 ± 0.07, P < 0.05), respectively. In anti-miRNA-24 group, endothelial cell proliferation decreased by 33.46 % compared with the control group (12.15 ± 0.21 vs 18.26 ± 0.45, P < 0.01), while it increased by 46.25 % compared with the miRNA-24 group (12.15 ± 0.21 vs 6.53 ± 0.11, P < 0.01). The expression of eNOS at mRNA and protein levels in anti-miRNA-24 group decreased by 44.21 % (0.53 ± 0.04 vs 0.95 ± 0.02, P < 0.05) and by 30.95 %(0.29 ± 0.05 vs 0.42 ± 0.06, P < 0.05) compared with the control group, while it increased by 35.84 % (0.53 ± 0.04 vs 0.34 ± 0.01, P < 0.05) and by 75.17 % (0.29 ± 0.05 vs 0.072 ± 0.06, P < 0.05) compared with miRNA-24 group. The expression of Sp1 at mRNA and protein levels in ant-miRNA-24 group decreased by 36.36 % (0.63 ± 0.04 vs 0.99 ± 0.03, P < 0.05) and by 22.41 % (0.45 ± 0.06 vs 0.58 ± 0.07, P < 0.05) compared with the control group, while it increased by 44.44 % (0.63 ± 0.04 vs 0.35 ± 0.01, P < 0.05) and by 48.88 % (0.45 ± 0.06 vs 0.23 ± 0.05, P < 0.05) compared with miRNA-24 group. HUVECs proliferation and eNOS expression are significantly inhibited by miRNA-24. Sp1, which is regulated by miRNA-24, might act as one of the important factors involved in eNOS gene expression.
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