Role of RIP2 in macrophage inflammatory activation and polarization transformation
10.20039/j.cnki.1007-3949.2025.01.005
- VernacularTitle:RIP2对巨噬细胞炎性活化及极性转化的调控
- Author:
Liang HOU
1
;
Yanchun DING
Author Information
1. 长江航运总医院心内科,湖北省武汉市 430000
- Publication Type:Journal Article
- Keywords:
receptor interacting protein 2;
oxidized low density lipoprotein;
atherosclerosis;
macrophage
- From:
Chinese Journal of Arteriosclerosis
2025;33(1):30-37
- CountryChina
- Language:Chinese
-
Abstract:
Aim To observe the effects of receptor interacting protein 2(RIP2)on macrophage inflammatory ac-tivation and polarity transformation,and to explore the mechanism of RIP2 in macrophage phagocytosis of oxidized low den-sity lipoprotein(ox-LDL).Methods THP-1 derived macrophages were treated with different doses(10,25 and 50 mg/L)of ox-LDL for 24 hours,and treated with 50 mg/L ox-LDL for 8,16 and 24 hours.Real-time quantitative PCR and Western blot were used to detect the expression of RIP2 mRNA and protein in THP-1 derived macrophages,and ELISA was used to detect the secretion of tumor necrosis factor-α(TNF-α)and monocyte chemoattractant protein-1(MCP-1).Three pairs of RIP2 siRNA were designed,transfecting them into cells using hiperfict transfection reagent,real-time quanti-tative PCR and Western blot were used to detect the expression of RIP2 mRNA and protein in THP-1 derived macrophages after transfection,in order to screen for the optimal siRNA transfection concentration and the most effective pair of siRNA.After transfection with the most effective RIP2 siRNA,cells were treated with 50 mg/L ox-LDL for 24 hours,ELISA was used to detect the secretion of TNF-α,MCP-1,interleukin-10(IL-10)and interleukin-12(IL-12),real-time quantitative PCR was used to detect the expression of inducible nitric oxide synthase(iNOS)and arginase-1(Arg-1),flow cytometry was used to detect the expression of cell surface antigens CD86,CD80 and CD163.Results Ox-LDL induced the ex-pression of RIP2 in macrophages in a dose-dependent and time-dependent manner.With the increase of ox-LDL treatment dose and time,the expression of RIP2 mRNA and protein increased.Among them,the expression of RIP2 protein in the 50 mg/L group was 7.6 times of the control group,and the expression of RIP2 protein in the 24 h group was 17.9 times of the control group(P＜0.001).The ELISA results showed that with the increase of ox-LDL treatment dose and time,the secretion of TNF-α and MCP-1 increased(P＜0.05).After transfection of RIP2 siRNA into cells,ELISA results showed that the secretion of TNF-α,MCP-1 and IL-10 in the ox-LDL group was 2.4 times,2.9 times and 1.8 times of the control group(P＜0.01),and the secretion of IL-12 decreased by 34.2％compared to the control group(P＜0.01);the secretion of TNF-α,MCP-1 and IL-10 in the siRNA+ox-LDL group decreased by 37.4％,45.3％and 27.4％,respectively,com-pared to the ox-LDL group(P＜0.01),while the secretion of IL-12 increased by 31.4％(P＜0.05).The results of flow cytometry and real-time quantitative PCR showed that the expression of CD86,CD80 and iNOS mRNA in the ox-LDL group was 14.2 times,33.8 times and 4.5 times of those of the control group,respectively,while the expression of CD163 and Arg-1 mRNA decreased by 33.4％and 43.0％,respectively,compared with the control group(P＜0.05);the expression of CD86,CD80 and iNOS mRNA in the siRNA+ox-LDL group decreased by 27.6％,29.3％and 34.3％,respectively,compared to the ox-LDL group,while the expression of CD163 and Arg-1 mRNA increased by 30.3％and 38.6％,respec-tively(P＜0.05).Conclusion RIP2 expression in macrophages can be induced by ox-LDL in a dose-dependent and time-dependent manner.RIP2 gene silencing can inhibit ox-LDL induced M1 macrophage transformation.
