精胺对高糖引起的大鼠心肌纤维化的影响及其机制

doi:10.12047/j.cjap.5952.2020.043

摘要/Abstract

摘要： 目的: 观察精胺对糖尿病心肌病(DCM)及高糖处理的心肌成纤维细胞(CFs)的保护作用并探讨其机制。方法: ①动物实验:24 只雄性Wistar大鼠随机分为正常组(Control),糖尿病组(T1D)和精胺组(T1D+Sp),每组8只。采用一次性腹腔注射链脲佐菌素(STZ,60 mg/kg)复制 1 型糖尿病大鼠模型,精胺组在 STZ 注射前两周每天腹腔注射精胺(Sp,5 mg/(kg·d)),随后隔天注射,饲养至 12 周。检测各组大鼠血糖、胰岛素水平、射血分数(EF)和缩短分数(FS),并对大鼠心脏组织进行 Masson 染色和 Sirius red 染色。②细胞实验:出生1～3 d的大鼠心脏提取原代 CFs,随机分为正常组(Control),高糖组(HG)和精胺组(HG+Sp,每组 n=6)。高糖(HG,40 mmol/L)处理 CFs 复制细胞模型,精胺组在高糖处理前给予Sp(5 μmol/L)预处理30 min。CCK8检测细胞活性,ELISA法检测培养基中胶原含量,Western blot 测定细胞周期相关蛋白(PCNA、CyclinD1 及 P27)的表达。结果: 与 Control 组相比,T1D 大鼠血糖显著上升,胰岛素水平和心脏功能降低;染色结果显示心肌胶原含量增加。同时,HG 组细胞活力与培养基中胶原含量明显增加,PCNA、CyclinD1 表达上调,而 P27 表达下调。精胺能减轻上述变化,表现为改善心脏功能,调节细胞周期蛋白表达和减轻心肌纤维化水平。结论: 精胺可减轻糖尿病心肌病心肌纤维化的发生,其机制可能与调节细胞周期有关。

关键词: 糖尿病心肌病, 心肌纤维化, 细胞周期, 精胺, 大鼠

Abstract: Objective: To investigate the protective effect of spermine (Sp) on diabetic cardiomyopathy (DCM) and high glucose-induced cardiac fibroblasts (CFs), and to explore its mechanism.Methods: ①Animal experiments: 24 male Wistar rats were randomly divided into control group, type 1 diabetes group (TID) and spermine group (TID+Sp, each group n=8). TID rats were induced by streptozocin (STZ, 60 mg/kg), and TID+Sp rat were pretreated with spermine (Sp, 5 mg/(kg·d)) for 2 weeks before STZ injection. After 12 weeks of modeling, blood glucose, insulin levels, ejection fraction (EF) and shortening fraction (FS) were measured, and Masson staining and Sirius red staining were performed in the rat cardiac tissues. ②Cell experiments: primary CFs were extracted from newborn (1-3 d) Wistar rat hearts, and were randomly divided into control group, high-glucose group (HG) and HG+Sp group (n=6 per group). HG group was treated with 40 mmol/L glucose, and the HG+Sp group was pretreated with 5 μmol/L Sp for 30 min before HG treatment. The cell viability of CFs was detected by CCK8, the content of collagen in culture medium was analyzed by ELISA, and protein expressions of cell cycle related proteins (PCNA, CyclinD1 and P27) were detected by Western blot. Results: Compared with control group, the blood glucose and collagen content were increased, and the insulin level and heart function were decreased in the T1D group. Meanwhile, HG induced an increasing of the cell viability, the collagen content in the medium and the expressions of PCNA and CyclinD1, while the expression of P27 was down-regulated. Spermine could reduce the above changes, manifested as improving the cardiac function, regulating the expression of cyclin and reducing the level of myocardial fibrosis. Conclusion: Spermine can alleviate myocardial fibrosis in diabetic cardiomyopathy, which mechanism is related to the regulation of cell cycle.

Key words: diabetic cardiomyopathy, myocardial fibrosis, cell cycle, spermine, rats

中图分类号:

R363

扈敬, 李法东, 邵小婷, 李思葳, 张欣颖, 赵冰冰, 徐长庆, 魏璨. 精胺对高糖引起的大鼠心肌纤维化的影响及其机制[J]. 中国应用生理学杂志, 2020, 36(3): 193-196.

HU Jing, LI Fa-dong, SHAO Xiao-ting, LI Si-wei, ZHANG Xin-ying, ZHAO Bing-bing, XU Chang-qing, WEI Can. Spermine attenuates high glucose-induced myocardial fibrosis by regulating the cell cycle[J]. CJAP, 2020, 36(3): 193-196.

参考文献

[1] Zatalia SR, Sanusi H. The role of antioxidants in the pathophysiology, complications, and management of diabetes mellitus[J]. Acta Med Indones, 2013, 45(2): 141-147.
[2] Ward ML, Crossman DJ. Mechanisms underlying the impaired contractility of diabetic cardiomyopathy[J]. World J Cardiol, 2014, 6(7): 577-584.
[3] Fan D, Takawale A, Lee J, et al. Cardiac fibroblasts, fibrosis and extracellular matrix remodeling in heart disease[J]. Fibrogenesis Tissue Repair, 2012, 5(1): 15.
[4] Mackenna D, Summerour SR, Villarreal FJ. Role of mechanical factors in modulating cardiac fibroblast function and extracellular matrix synthesis[J]. Cardiovasc Res, 2000, 46(2): 257-263.
[5] Liu Z, Zhang Y, Tang Z, et al. Matrine attenuates cardiac fibrosis by affecting ATF6 signaling pathway in diabetic cardiomyopathy[J]. Eur J Pharmacol, 2017, 804: 21-30.
[6] Wang Y, Chen J, Li S, et al. Exogenous spermine attenuates rat diabetic cardiomyopathy via suppressing ROS-p53 mediated downregulation of calcium-sensitive receptor[J]. Redox Biol, 2020, 32: 101514
[7] 袁迪, 王跃虹, 袁辉, 等. 外源性精胺对缺氧诱导乳鼠心肌细胞凋亡的影响及其机制[J]. 中国应用生理学杂志, 2018, 34(3): 193-196.
[8] Wei C, Li H, Wang Y, et al. Exogenous spermine inhibits hypoxia/ischemia-induced myocardial apoptosis via regulation of mitochondrial permeability transition pore and associated pathways[J]. Exp Biol Med (Maywood), 2016, 241(14): 1505-1515.
[9] 范玉琪, 王跃虹, 邵毅英, 等. 外源性精胺抑制高尔基体应激减轻高糖诱导的心肌损伤[J]. 中国应用生理学杂志, 2019, 35(5): 385-389.
[10]Shao X, Zhang X, Hu J, et al. Dopamine 1 receptor activation protects mouse diabetic podocytes injury via regulating the PKA/NOX-5/p38 MAPK axis[J]. Exp Cell Res, 2020, 388(2): 111849.
[11]Pitt B, Zannad F. The detection of myocardial fibrosis: an opportunity to reduce cardiovascular risk in patients with diabetes mellitus[J]? Circ Cardiovasc Imaging, 2012, 5(1): 9-11.
[12]Wei C, Li HZ, Wang YH, et al. Exogenous spermine inhibits the proliferation of human pulmonary artery smooth muscle cells caused by chemically-induced hypoxia via the suppression of the ERK1/2- and PI3K/AKT-associated pathways[J]. Int J Mol Med, 2016, 37: 39-46.
[13]Tsai WC, Cheng JW, Chen JL, et al. Low-level laser irradiation stimulates tenocyte proliferation in association with increased NO synthesis and upregulation of PCNA and cyclins[J]. Lasers Med Sci, 2014, 29(4): 1377-1384.
[14]Yang G, Wu L, Bryan S, et al. Cystathionine gamma-lyase deficiency and overproliferation of smooth muscle cells[J]. Cardiovasc Res, 2010, 86(3): 487-495.
[15]Sherr CJ, Roberts JM. Inhibitors of mammalian G1 cyclin-dependent kinases[J]. Genes Dev, 1995, 9(10): 1149-1163.
[16]Tsai YC, Lee YM, Hsu CH, et al. The effect of ferulic acid ethyl ester on leptin-induced proliferation and migration of aortic smooth muscle cells[J]. Exp Mol Med, 2015, 47(8): e180.