miR-509-3p promotes oxidized low-density lipoprotein-induced apoptosis in mouse aortic endothelial cells.
10.3760/cma.j.cn121430-20230806-00583
- Author:
Rui ZHANG
1
;
Yanqiu SONG
2
;
Fumei ZHAO
2
;
Ting LIU
2
;
Hongliang CONG
1
;
Hui ZHAO
3
,
4
Author Information
1. Department of Cardiology, Tianjin Chest Hospital, Tianjin 300222, China.
2. Tianjin Cardiovascular Diseases Institute, Tianjin Chest Hospital, Tianjin 300222, China.
3. Department of Cardiology, Tianjin Hospital, Tianjin 300211, China. Corresponding author: Cong Hongliang, Email: doctorchl@
4. com.
- Publication Type:Journal Article
- MeSH:
Animals;
Mice;
Humans;
Endothelial Cells;
MicroRNAs/metabolism*;
Signal Transduction;
Lipoproteins, LDL/metabolism*;
Apoptosis;
RNA, Messenger/metabolism*;
Proto-Oncogene Proteins c-bcl-2/pharmacology*;
Atherosclerosis/metabolism*;
Luciferases/pharmacology*;
Cell Proliferation;
Human Umbilical Vein Endothelial Cells
- From:
Chinese Critical Care Medicine
2023;35(12):1291-1297
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To investigate the effect of microRNA-509-3p (miR-509-3p) on the apoptosis of atherosclerotic vascular endothelial cells.
METHODS:Mouse aortic endothelial cells (MAECs) were divided into normal control group, oxidized low-density lipoprotein (ox-LDL) group, miR-509-3p overexpression group, miR-509-3p overexpression control group, miR-509-3p inhibitor + ox-LDL group, and miR-509-3p inhibitor control + ox-LDL group. MAEC were induced with 100 mg/L ox-LDL for 24 hours, and then transfected with miR-509-3p overexpression/inhibitor and corresponding control for 48 hours. The miR-509-3p expression in MAECs exposed to ox-LDL was detected using real-time fluorescence quantitative polymerase chain reaction (RT-qPCR). Flow cytometry was used to detect the level of apoptosis, and cell counting kit (CCK-8) was used to detect the proliferation activity of MAECs. The direct gene targets of miR-509-3p were predicted using bioinformatics analyses and confirmed using a dual luciferase reporter assay. The expression of Bcl-2 mRNA and protein was detected by RT-qPCR and Western blotting, respectively.
RESULTS:Compared with the normal control group, miR-509-3p was significantly upregulated in ox-LDL-stimulated MAECs (1.68±0.85 vs. 1.00±0.30, t = 2.398, P < 0.05). After transfection of MAECs with miR-509-3p overexpression, the luciferase activity of the BCL2 3'UTR WT reporter gene was significantly lower than that of miR-509-3p overexpression control group (0.83±0.06 vs. 1.00±0.07, t = 4.531, P = 0.001). The luciferase activity of the BCL2 3'-UTR mutant (MUT) reporter gene was not significantly different from that of miR-509-3p overexpression control group (0.94±0.05 vs. 1.00±0.08, t = 1.414, P = 0.188). Compared with the normal control group and miR-509-3p mimics control group, the cell proliferation activity was decreased [(0.60±0.06)% vs. (1.00±0.09)%, (0.89±0.04)%, both P < 0.01], the percentage of apoptotic cells were increased [(23.46±2.02)% vs. (7.66±1.52)%, (10.40±0.78)%, both P < 0.05], and the mRNA and protein expression of Bcl-2 were significantly downregulated (Bcl-2 mRNA: 0.52±0.13 vs. 1.00±0.36, 1.10±0.19, Bcl-2 protein: 0.42±0.07 vs. 1.00±0.11, 0.93±0.10, both P < 0.01) in miR-509-3p overexpression group. Compared with the ox-LDL group, inhibition of miR-509-3p expression could increase the proliferation activity of MAECs induced by ox-LDL [(0.64±0.35)% vs. (0.34±0.20%)%, P < 0.05], and reduce the apoptosis rate [(13.59±2.22)% vs. (29.84±5.19)%, P < 0.01], and up-regulated the expression of Bcl-2 mRNA and protein in MAECs induced by ox-LDL (Bcl-2 mRNA relative expression: 0.82±0.09 vs. 0.52±0.10, Bcl-2 protein relative expression: 0.83±0.17 vs. 0.40±0.07, both P < 0.05).
CONCLUSIONS:Bcl-2 was one of the target genes of miR-509-3p. miR-509-3p can reduce the proliferation activity of endothelial cells, reduce the expression of Bcl-2, and promote cell apoptosis, thereby promoting the occurrence and development of atherosclerosis. Inhibition of miR-509-3p expression may be a potential therapeutic target for atherosclerosis.
