阻断连接子蛋白43(Cx43)抑制脂多糖诱导的小鼠RAW264.7巨噬细胞M1型极化

Blocking connexin 43 inhibits the M1 polarization of mouse 264.7 macrophages induced by lipopolysaccharide

目的研讨连接子蛋白43(Cx43)在脂多糖(LPS)诱导的小鼠RAW264.7巨噬细胞M1型极化中的作用。方法将体外培养的小鼠RAW264.7巨噬细胞分为对照组、LPS组、LPS联合Cx43阻断剂模拟肽19(Gap19)或模拟肽26(Gap26)预处理组(LPS联合Gap19组、LPS联合Gap26组)。Western blot法检测小鼠RAW264.7巨噬细胞Cx43和M1型极化标志物CD86、诱导型一氧化氮合酶(iNOS)的蛋白水平,免疫荧光细胞化学染色观察CD86在RAW264.7巨噬细胞的表达和定位,流式细胞术检测小鼠RAW264.7巨噬细胞M1型极化标志物CD86的表达频数。结果与对照组相比,LPS组中CD86、iNOS和Cx43的蛋白表达明显升高,CD86的表达频数明显升高;与LPS组相比,在LPS联合Gap19组和LPS联合Gap26组中CD86和iNOS的蛋白表达均显著降低,CD86的表达频数均显著降低。因此,LPS可以诱导小鼠RAW264.7巨噬细胞向M1型极化,而Gap19和Gap26能够降低M1型极化标志物的表达。结论通过阻断Cx43可以抑制巨噬细胞M1型极化。

Objective To investigate the effect of connexin 43(Cx43) on M1 polarization of mouse RAW264.7 macrophages induced by lipopolysaccharide(LPS). Methods RAW264.7 macrophages were cultured in vitro and randomly divided into four groups: control group, LPS group, LPS combined with Gap19 group, LPS combined with Gap26 group. The protein levels of Cx43 and M1 polarization marker CD86 and inducible nitric oxide synthase(iNOS) in mouse RAW264.7 macrophages were detected by Western blot analysis. The expression and localization of CD86 in RAW264.7 macrophages were observed by immunofluorescence cytochemistry, and the expression frequency of M1 polarization marker CD86 in mouse RAW264.7 macrophages was detected by flow cytometry. Results Compared with the control group, the protein expression of CD86, iNOS and Cx43, as well as the expression frequency of CD86 in LPS group showed a significant increase. However, compared with LPS group, the protein expression of CD86 and iNOS, and the expression frequency of CD86 decreased significantly in LPS combined with Gap19 group and LPS combined with Gap26 group. As such, LPS could induce M1 polarization of macrophage, while Gap19 and Gap26 can reduce the expression of M1 polarization markers. Conclusion M1 polarization of macrophages can be inhibited by blocking Cx43.