HSF1/AMPK信号通路在铁死亡参与糖尿病心肌病发病的机制研究

Role of the HSF1/AMPK signaling pathway in the pathogenesis of ferroptosis-involving diabetic cardiomyopathy

摘要

目的 探讨热休克因子1（heat shock factor 1，HSF1）/5'-单磷酸腺苷活化蛋白激酶（adenosine monophosphate activated protein kinase，AMPK）信号通路调控铁死亡对糖尿病心肌病（diabetic cardiomyopathy，DCM）发病的影响。 方法 本研究为实验研究，采用空白对照与多组实验对照。H9c2细胞随机分为4组：低葡萄糖组（control，Con）、高葡萄糖组（high glucose，HG）、HG+HSF1组、HG+HSF1+化合物C（compound C，CC）组。分别对细胞进行罗丹明胶质蛋白染色、细胞线粒体（reactive oxygen species，ROS）检测和细胞脂质ROS检测，并通过Western blot分析AMPK信号表达。雄性C57/BL6小鼠随机分为4组：NC组、NC+HSF1组、DM组和DM+HSF1组，每组12只。通过超声心动图评估了小鼠心血管功能参数。 结果 与Con组相比，HG组HSF1、pAMPK/AMPK水平明显下调（P=0.005、0.002），和相对细胞表面积、线粒体Fe²⁺水平、线粒体ROS水平、细胞脂质ROS水平明显增加（P=0.001、0.003、0.006、0.002）。与HG组相比，HG+HSF1组明显逆转了这些变化（P=0.001、0.001、0.002、0.006、0.007、0.003），但加入CC时HSF1的逆转作用明显减弱（P<0.05）。与NC组相比，DM组EF%、FS%、E/A、E'/A'和心脏组织中HSF1、pAMPK/AMPK表达明显降低（均P<0.01），和心脏组织中Fe²⁺、ROS、丙二醛（Malondialdehyde，MDA）水平和4-羟基壬烯酸（4-Hydroxynonenal，4-HNE）蛋白水平明显增加（P=0.004、0.003、0.001、0.004），DM+HSF1组明显逆转了这些变化。 结论 HSF1在DCM病理过程中发挥心脏保护作用，其抗铁死亡作用可能与AMPK依赖性的脂质代谢和线粒体稳态调节有关。

Abstract

Objective To investigate the effect of ferroptosis regulated by the heat shock factor 1（HSF1）/adenosine 5’-monophosphate-activated protein kinase（AMPK） signaling pathway on the pathogenesis of diabetic cardiomyopathy（DCM）. Methods This was an experimental study with a blank control and multiple experimental controls. H9c2 cells were randomly divided into 4 groups：low glucose group（control，Con），high glucose（HG） group，HG+HSF1 group，and HG+HSF1+compound C（CC） group. The cells were stained by rhodamine colloid protein，and tested for mitochondrial reactive oxygen species（ROS） and cellular lipid ROS. The AMPK signal expression was analyzed by Western blot. Male C57/BL6 mice were randomly divided into four groups： negative control（NC） group，NC+HSF1 group，DM group，and DM+HSF1 group，with 12 mice in each group. The cardiovascular function parameters of mice were evaluated by echocardiography. Results Compared with the Con group，the levels of HSF1 and pAMPK/AMPK in the HG group decreased significantly（P=0.005，0.002），and the relative cell surface area，mitochondrial Fe²⁺ level，mitochondrial ROS level，and cellular lipid ROS level increased significantly（P=0.001，0.003，0.006，0.002）. Compared with the HG group，the HG+HSF1 group significantly reversed these changes（P=0.001，0.001，0.002，0.006，0.007，0.003），but the reversal effect of HSF1 was obviously weakened when CC was added（P<0.05）. Compared with the NC group，the ejection fraction，fractional shortening，mitral ratio of peak early to late diastolic filling velocity（E/A），and E'/A' as well as the expression levels of HSF1 and pAMPK/AMPK in heart tissues in the DM group were significantly decreased（P<0.01），and the levels of Fe²⁺，ROS，malondialdehyde，and 4-hydroxynonenal protein in heart tissues were significantly increased（P=0.004，0.003，0.001，0.004）；the DM+HSF1 group significantly reversed these changes. Conclusion HSF1 plays a cardioprotective role in the pathological process of DCM，and its anti-ferroptosis effect may be related to AMPK-dependent lipid metabolism and mitochondrial homeostasis regulation.

关键词

热休克因子1 / 5'-单磷酸腺苷活化蛋白激酶 / 糖尿病心肌病 / 小鼠 / 铁死亡

Key words

heat shock factor 1 / adenosine 5’-monophosphate-activated protein kinase / diabetic cardiomyopathy / mouse / ferroptosis

基金资助

湖南省卫生健康委科研计划资助项目(202100236512)