PRMT2 通过 ABCA1 介导的胆固醇流出抑制 ox-LDL 诱导的 RAW264.7 巨噬细胞形成泡沫细胞。
PRMT2 inhibits the formation of foam cell induced by ox-LDL in RAW 264.7 macrophage involving ABCA1 mediated cholesterol efflux.
机构信息
Institute of Clinical Medicine, First Affiliated Hospital of University of South China, Hengyang, 421001, Hunan, PR China; Department of Cardiovascular Medicine, First Affiliated Hospital of University of South China, Hengyang, 421001, Hunan, PR China.
Department of Physiology, Basic Medical College, Guilin Medical University, Guilin, 541199, Guangxi, China; Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin, 541199, Guangxi, PR China.
出版信息
Biochem Biophys Res Commun. 2020 Mar 26;524(1):77-82. doi: 10.1016/j.bbrc.2020.01.040. Epub 2020 Jan 21.
OBJECTIVES
Protein arginine methyltransferase 2 (PRMT2) is closely related to the occurrence and development of atherosclerosis. However, its underlying mechanisms remain to be elucidated. The purpose of this study is to observe the effect of overexpression of PRMT2 on the formation of foam cells and to explore its possible mechanism in RAW 264.7 macrophage.
METHODS
Lentivirus vector of overexpression PRMT2 (LV-PRMT2) was constructed. LV-PRMT2 and lentivirus vector GV492 were transfected into RAW 264.7 macrophages, positive clone cells were screened by treatment with 4.0 μg/mL puromycin for 4 weeks. The macrophages were treated with ox-LDL (50 μg/mL) for 48 h to induce foaming. The lipid accumulation of macrophages was observed by oil red O staining. The levels of cellular total cholesterol (TC), free cholesterol (FC) and cholesteryl ester (CE) were measured by high performance liquid chromatography (HPLC) assays. The cholesterol efflux of macrophages was tested by the [H] labeled cholesterol. The expressions of ATP binding cassette transporter A1 (ABCA1), ATP binding cassette transporter G1 (ABCG1), CD36 and scavenger receptor A1 (SR-A1) in macrophages were measured by Western Blot.
RESULTS
The results showed that LV-PRMT2 and lentivirus vector has been successfully transfected into RAW 264.7 macrophage. Compared with the Vector group, the mRNA and protein expressions of PRMT2 were significantly up-regulated (P < 0.05). Compared with Control group, the expression of PRMT2 was significantly down-regulated in ox-LDL group (P < 0.05). A large number of red lipid droplets appeared in the cells in Vector group. Compared with Vector group, lipid droplets, the levels of TC, FC and CE and CE/TC, cholesterol efflux rate and expression of ABCA1 in RAW 264.7 macrophage was significantly decreased in LV-PRMT2 group (all P < 0.05). There was no significant difference about the expressions of ABCG1, CD36 and SR-A1 between LV-PRMT2 group and Vector group (all P > 0.05).
CONCLUSIONS
Overexpression of PRMT2 inhibits the formation of foam cell induced by ox-LDL in RAW 264.7 macrophage, and the mechanism may be related to the increase of ABCA1 expression and ABCA1 mediated cholesterol efflux.
目的
蛋白质精氨酸甲基转移酶 2（PRMT2）与动脉粥样硬化的发生发展密切相关。但其潜在机制尚待阐明。本研究旨在观察过表达 PRMT2 对泡沫细胞形成的影响，并探讨其在 RAW 264.7 巨噬细胞中的可能机制。
方法
构建过表达 PRMT2 的慢病毒载体（LV-PRMT2）。用 4.0μg/mL 嘌呤霉素处理 4 周筛选转染 RAW 264.7 巨噬细胞的阳性克隆细胞。用 ox-LDL（50μg/mL）处理巨噬细胞 48 h 诱导泡沫化。油红 O 染色观察巨噬细胞内脂质蓄积。高效液相色谱（HPLC）法测定细胞总胆固醇（TC）、游离胆固醇（FC）和胆固醇酯（CE）水平。[H]标记胆固醇检测巨噬细胞胆固醇流出率。Western blot 法检测巨噬细胞中 ATP 结合盒转运体 A1（ABCA1）、ATP 结合盒转运体 G1（ABCG1）、CD36 和清道夫受体 A1（SR-A1）的表达。
结果
LV-PRMT2 和慢病毒载体已成功转染 RAW 264.7 巨噬细胞。与 Vector 组比较，PRMT2mRNA 和蛋白表达均显著上调（P<0.05）。与对照组比较，ox-LDL 组 PRMT2 表达显著下调（P<0.05）。Vector 组细胞内出现大量红色脂质滴。与 Vector 组比较，LV-PRMT2 组 RAW 264.7 巨噬细胞内脂质滴、TC、FC 和 CE 水平及 CE/TC、胆固醇流出率和 ABCA1 表达均显著降低（均 P<0.05）。LV-PRMT2 组与 Vector 组 ABCG1、CD36 和 SR-A1 表达比较差异均无统计学意义（均 P>0.05）。
结论
过表达 PRMT2 抑制 ox-LDL 诱导的 RAW 264.7 巨噬细胞泡沫细胞形成，其机制可能与 ABCA1 表达增加和 ABCA1 介导的胆固醇流出有关。
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