CD137 signaling regulates the expression of nuclear factor of activated T cells c1 through miR-145a-5p in ApoE(-)/(-) mice.
- Author:
Wei ZHONG
1
;
Jinchuan YAN
2
;
Email: YANJINCHUAN@HOTMAIL.COM.
;
Zhongqun WANG
1
;
Yi LIANG
1
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Apolipoproteins E; Mice; Mice, Knockout; MicroRNAs; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; NFATC Transcription Factors; Plaque, Atherosclerotic; RNA, Messenger; Signal Transduction; T-Lymphocytes; Transfection; Tumor Necrosis Factor Receptor Superfamily, Member 9
- From: Chinese Journal of Cardiology 2015;43(10):887-893
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo investigate if miR-145a-5p participates the modulation process of CD137 signaling on the expression of nuclear factor of activated T cells c1 (NFATc1) in ApoE(-)/(-) mice.
METHODSAtherosclerotic plaque model was produced by perivascular carotid collar placement in ApoE(-)/(-) mice. After surgery, the mice were randomly divided into the following groups: CD137 activated group (CD137 group, n = 6), CD137 inhibited group (anti-CD137 group, n = 6) and control group (n = 6). The mRNA expression of miR-145a-5p in plaque and cells was measured by real-time quantitative PCR (RT-PCR). Immunofluorescence was used to observe the distribution of NFATc1 in plaque and the expression of NFATc1 at mRNA and protein levels were detected by qRT-PCR, Western blot, respectively. The mouse vascular smooth muscle cells (VSMCs) were isolated and transfected with miR-145a-5p mimics or inhibitors by Lipofectamine. The eukaryotic expression vector and luciferase vector including p3xFLAG-NFATc1, p3xFLAG-NFATc1-3'UTR, psicheck2-NFATc1, psicheck2-NFATc1-Mut were constructed through molecular cloning and homologous recombination techniques, 293T cells were transfected with the miR-145a-5p mimics or inhibitors and the protein level and fluorescence intensity were then measured, respectively.
RESULTSIn vivo or in vitro, the level of miR-145a-5p was significantly decreased (0.21 ± 0.06 vs. 1.00 ± 0.00, P < 0.05, 0.22 ± 0.07 vs. 0.50 ± 0.12, P < 0.05) while the opposite effects were observed in anti-CD137 group. NFATc1 expression was decreased or increased in VSMCs transfected with miR-145a-5p mimics or inhibitors, respectively (all P < 0.05). miR-145a-5p mimics decreased the expression of p3xFLAG-NFATc1-3'UTR and the fluorescence intensity (0.56 ± 0.08 vs. 1.00 ± 0.00, P < 0.05).
CONCLUSIONCD137 signaling participates the regulation process on the expression of NFATc1 through miR-145a-5p in ApoE(-)/(-) mice.