摘要
目的基于赖氨酸乙酰转移酶2A(GCN5)探讨暖心康对心力衰竭小鼠心肌组织DNA损伤的作用及机制。方法将60只小鼠随机分为假手术组、模型组、沙库巴曲缬沙坦组(25 mg·kg^(-1))及暖心康低、中、高剂量组(2.93、5.85、11.70 g·kg^(-1)),每组10只。采用主动脉弓缩窄术构建压力负荷导致的心力衰竭小鼠模型。各给药组按照设定剂量灌胃给药,假手术组及模型组灌胃生理盐水(10 mL·kg^(-1)),每日1次,持续6周。采用超声心动图检测小鼠心功能,主要评估指标:左室射血分数(LVEF)、左室短轴缩短率(LVFS)、左室收缩末期内径(LVIDs)、左室舒张末期内径(LVIDd);HE染色法观察心肌组织病理变化;免疫组织化学法检测心肌组织γH2AX、8-OHdG蛋白表达情况;TUNEL染色法检测心肌细胞凋亡水平;qRT-PCR法检测心肌组织PARP、53BP1、GCN5 mRNA表达水平;Western Blot法检测心肌组织GCN5、BAX、Bcl2蛋白表达水平。结果与假手术组比较,模型组小鼠的LVEF、LVFS水平显著降低(P<0.01),而LVIDs、LVIDd水平显著升高(P<0.01);心肌细胞显著增大,形态不规则,排列不整齐;心肌组织中γH2AX、8-OHdG阳性染色明显增强,呈棕黄色颗粒或斑块状弥散分布,γH2AX、8-OHdG蛋白表达及PARP、53BP1 mRNA表达均显著上调(P<0.01);心肌组织GCN5蛋白及mRNA表达水平显著升高(P<0.01);心肌组织TUNEL阳性细胞率显著升高(P<0.01);心肌组织抗凋亡蛋白Bcl2表达水平明显降低(P<0.05),促凋亡蛋白BAX表达水平明显升高(P<0.05)。与模型组比较,暖心康中、高剂量组小鼠的LVIDd水平显著降低(P<0.05,P<0.01);各给药组小鼠的LVEF、LVFS水平显著提高(P<0.01),LVIDs水平显著降低(P<0.05,P<0.01),心肌细胞形态及排列状况均有所改善,心肌组织中γH2AX、8-OHdG阳性染色强度明显减弱,γH2AX、8-OHdG蛋白表达及PARP、53BP1 mRNA表达均显著下调(P<0.01),心肌组织GCN5蛋白及mRNA表达水平显著降低(P<0.01),心肌组织TUNEL阳性细胞率显著降低(P<0.05,P<0.01);暖心康各剂量组小鼠心肌组织Bcl2蛋白表达水平显著升高(P<0.05,P<0.01),BAX蛋白表达水平显著降低(P<0.05,P<0.01)。结论暖心康能够改善压力负荷致心力衰竭小鼠的心功能,减轻病理损伤及心肌细胞凋亡,作用机制可能与抑制GCN5表达,减轻心肌组织DNA损伤有关。
Objective To investigate the effect and mechanism of Nuanxinkang(NXK)on myocardial DNA damage in mice with heart failure,focusing on lysine acetyltransferase 2A(GCN5).Methods Sixty mice were randomly divided into six groups(n=10 per group):sham-operated group,model group,sacubitril/valsartan group(25 mg·kg^(-1)),and low-(2.93 g·kg^(-1)),medium-(5.85 g·kg^(-1)),and high-dose(11.70 g·kg^(-1))NXK groups.A pressure overload-induced heart failure model was established by transverse aortic constriction(TAC).The treatment groups received their respective drugs by gavage,while the sham and model groups received an equal volume of saline(10 mL·kg^(-1)),once daily for 6 weeks.Cardiac function was assessed by echocardiography,measuring left ventricular ejection fraction(LVEF),left ventricular fractional shortening(LVFS),left ventricular internal diameter at end-systole(LVIDs),and left ventricular internal diameter at end-diastole(LVIDd).Myocardial histopathology was observed by hematoxylineosin(HE)staining.Protein expression ofγH2AX and 8-OHdG in myocardial tissue was detected by immunohistochemistry.Cardiomyocyte apoptosis was assessed by TUNEL staining.mRNA expression levels of PARP,53BP1,and GCN5 in myocardial tissue were measured by qRT-PCR.Protein expression levels of GCN5,BAX,and Bcl-2 were determined by Western Blot.Results Compared with the sham group,the model group showed significantly decreased LVEF and LVFS(P<0.01),and significantly increased LVIDs and LVIDd(P<0.01).Cardiomyocytes were significantly enlarged,irregular in shape,and disorganized.Positive staining forγH2AX and 8-OHdG in myocardial tissue was markedly enhanced,appearing as diffuse brownish-yellow granules or plaques;protein expression ofγH2AX and 8-OHdG and mRNA expression of PARP and 53BP1 were significantly upregulated(P<0.01).Both protein and mRNA expression levels of GCN5 in myocardial tissue were significantly increased(P<0.01).The TUNEL-positive cell rate in myocardial tissue was significantly elevated(P<0.01).The expression level of the anti-apoptotic protein Bcl-2 was significantly decreased(P<0.05),while the expression level of the pro-apoptotic protein BAX was significantly increased(P<0.05).Compared with the model group,the medium-and high-dose NXK groups showed significantly decreased LVIDd(P<0.05,P<0.01).All NXK treatment groups exhibited significantly improved LVEF and LVFS(P<0.01),significantly reduced LVIDs(P<0.05,P<0.01),and improved cardiomyocyte morphology and arrangement.The intensity of positive staining forγH2AX and 8-OHdG in myocardial tissue was markedly weakened;protein expression ofγH2AX and 8-OHdG and mRNA expression of PARP and 53BP1 were significantly downregulated(P<0.01).Protein and mRNA expression levels of GCN5 in myocardial tissue were significantly reduced(P<0.01).All NXK dose groups showed significantly increased Bcl-2 protein expression(P<0.05,P<0.01)and significantly decreased BAX protein expression(P<0.05,P<0.01)in myocardial tissue.Conclusion Nuanxinkang can improve cardiac function,alleviate pathological damage,and reduce cardiomyocyte apoptosis in mice with pressure overload-induced heart failure.Its mechanism of action may be related to the inhibition of GCN5 expression and the subsequent reduction of myocardial DNA damage.
作者
时恺芸
王陵军
梁碧容
吴辉
李志芳
方红城
SHI Kaiyun;WANG Lingjun;LIANG Birong;WU Hui;LI Zhifang;FANG Hongcheng(Shenzhen Clinical Medical College of Integrated Traditional Chinese and Western Medicine,Guangzhou University of Chinese Medicine/Shenzhen Hospital of Integrated Traditional Chinese and Western Medicine,Shenzhen 518104 Guangdong,China;The First Affiliated Hospital of Guangzhou University of Chinese Medicine/Guangdong Provincial Hospital of Chinese Medicine Clinical Research Institute,Guangzhou 510405 Guangdong,China;The First School of Clinical Medicine,Guangzhou University of Chinese Medicine,Guangzhou 510006 Guangdong,China)
出处
《中药新药与临床药理》
北大核心
2025年第12期2024-2033,共10页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
国家自然科学基金面上项目(82474403)
广东省自然科学基金项目(2023A1515010282,2024A15150121712)
国家中医药传承创新中心项目(2023QN19)
深圳市可持续发展科技专项(JCYJ20230807141402005)
深圳市“医疗卫生三名工程”项目(SZZYSM202106006)。
