敲低Spectrinβ Ⅱ加重棕榈酸诱导的心肌细胞损伤

杨容金; 邢萍萍; 张顺; 张晓萌; 张飞宇; 王姗; 陶凌

doi:10.12125/j.chj.202109072

敲低Spectrinβ Ⅱ加重棕榈酸诱导的心肌细胞损伤

Spectrin β Ⅱ knockdown aggravates cardiomyocyte injury induced by palmitic acid

摘要

摘要:
目的研究血影蛋白β Ⅱ（Spectrin βⅡ）在棕榈酸诱导的小鼠原代心肌细胞DNA损伤和细胞凋亡中的作用。
方法 ①Western blot检测棕榈酸处理后小鼠原代心肌细胞Spectrin β Ⅱ表达量变化。②分别用Ad-sh-scramble（对照组）和Ad-sh-Spectrin β Ⅱ（Spectrin β Ⅱ干涉组）的腺病毒感染小鼠原代心肌细胞，荧光显微镜下观察病毒的感染效率。利用Western blot和qPCR检测Spectrin β Ⅱ的干涉效率。③Western blot检测棕榈酸处理后Ad-sh-scramble组和Spectrin β Ⅱ干涉组小鼠原代心肌细胞DNA损伤指标γ-H2AX及凋亡蛋白剪切型半胱天冬酶3(cleaved-caspase 3)的表达，采用流式细胞术检测各组细胞凋亡水平。
结果 ①给予棕榈酸处理的小鼠原代心肌细胞Spectrin β Ⅱ蛋白水平显著降低（P<0.05）。②与Ad-sh-scramble组相比，Ad-sh-Spectrin β Ⅱ组Spectrin β Ⅱ的蛋白和mRNA水平均显著降低（P<0.05）③给予棕榈酸处理后，与Ad-sh-scramble组相比，Ad-sh-Spectrin β Ⅱ组的γ-H2AX2和cleaved-caspase 3蛋白表达显著升高（P<0.05），并且细胞凋亡率显著升高（P<0.05）。
结论敲低Spectrin β Ⅱ加重棕榈酸诱导的小鼠原代心肌细胞DNA损伤和细胞凋亡。

Abstract:
AIM To investigate the effects of Spectrin β Ⅱ on cardiomyocyte DNA damage and apoptosis induced by palmitic acid (PA).
METHODS Western blot was used to detect the protein levels of Spectrin β Ⅱ in mouse primary cardiomyocytes (MCMs) treated with PA. MCMs were infected with adenovirus expressing independent hairpin shRNA sequences for Spectrin β Ⅱ (Ad-sh-Spectrin β Ⅱ) or control sh-scramble sequence (Ad-sh-scramble). The infection efficiency of the adenovirus was examined by fluorescence microscope and confirmed by Western blot and qPCR. The protein levels of γ-H2AX, an indicator of DNA damage, and apoptotic protein cleaved-caspase 3 were detected by Western blot. Flow cytometry was used to analyze the apoptotic rate.
RESULTS The protein levels of Spectrin β Ⅱ were significantly decreased in MCMs treated with PA (P<0.05). Compared with those in the Ad-sh-scramble group, the protein and mRNA levels of Spectrin β Ⅱ in the Ad-sh-Spectrin β Ⅱ group were significantly decreased (P< 0.05). Compared with those in the Ad-sh-scramble group, the protein levels of γ-H2AX and cleaved-caspase3 were significantly increased (P<0.05) and the apoptotic rate was significantly increased (P<0.05) in the Ad-sh- Spectrin β Ⅱ group after treated with PA.
CONCLUSION Spectrin β Ⅱ knockdown aggravates cardiomyocyte DNA damage and apoptosis induced by palmitic acid.

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