基于RCS诱导RyR2功能异常探讨参松养心胶囊干预糖尿病心肌病的作用机制研究

Diabetic cardiomyopathy(DCM) is the importance of complication in diabetes mellitus, numbers of evidence based medicine concluded that clinical efficacy of Shensong yangxin capsule (SSYX) is fully established, however, its mechanisms needs to better understand; As our knowledge of the ryanodine receptor has advanced an appreciation that this key E-C coupling component may have a role in the pathogenesis of human cardiac disease has emerged. We demonstrated that carbonylation induces heterogeneity in cardiac ryanodine receptor (RyR2) function in diabetes mellitus and diabetes lung disease by reactive carbonyl species (RCS) is play a role in the pathogenesis of DCM. Therefore, we further hypothesized that (1) the target pathways of SSYX treatment DCM is normalizing the dysfunction RyR2 and scavengering RCS, (2) all of the mechanisms of both in those treatment SSYX and in the pathogenesis of DCM should be based on a Chinese medicine theory of “Fei zhu zhi jie”. We focused our attention on RyR2s and RCS, the proposal is in combination with RyR2 purification and reconstitution into proteoliposomes, and lipid bilayer was used to determine whether post translational modification by RCS represented a contributing cause, and the molecular and cellular mechanisms underlying on RyR2 and RCS of SSYX therapeutic in DCM; this study is a strong desire to identify and/or develop novel therapeutic targets of treatment DCM, this knowledge is critical to the development of novel, precision strategies to manipulate RyR2 function progression in DCM, and to help prevent, detect, and guide treatment represent the future of diabetics and cardiomyopathy management. Those research anther goals are to better understand molecular and cellular mechanisms responsible for “Fei zhu zhi jie”.

糖尿病心肌病为糖尿病主要并发症之一，多项循证医学显示参松养心胶囊疗效确切，其机制亟待深入研究。目前认为RyR2作为心肌E-C Coupling的关键分子在心肌病病理过程中起到重要作用，课题组研究提示糖尿病羰基应激以及糖尿病肺功能受限导致RCS积聚对RyR2蛋白翻译后修饰引起RyR2功能增强是糖尿病心肌病的重要机制；据此提出假说(1)参松养心胶囊可能通过抑制RyR2功能或（和）清除糖尿病体内RCS治疗糖尿病心肌病，(2)糖尿病心肌病的RCS修饰RyR2功能增强的机制及参松养心胶囊干预机制可能是中医“肺主治节”理论的体现。本项目拟以RCS、RyR2为切入点，采用RyR2蛋白重组、电生理技术研究RyR2属性；探讨糖尿病心肌RyR2增强机制，参松养心胶囊清除RCS、调控RyR2治疗机理，力图为中医药治疗糖尿病心肌病提供新的靶点和有益思路，同时尝试丰富中医“肺主治节”理论物质基础。

糖尿病心肌病、肺病是糖尿病主要并发症，其病理生理条件下心肺相关机制不清楚。参松养心胶囊是治疗心律失常的有效方剂，其治疗糖尿病心肌病机制需深入研究。中医肺主治节理论体现为肺通过宗气参与心跳、脉搏节律的调节，其分子基础不清。研究提示糖尿病羰基应激以及糖尿病肺功能受限导致RCS积聚对RyR2蛋白翻译后修饰引起RyR2功能增强是糖尿病心肌病的重要机制。RCS积聚参与修饰RyR2导致心率、心律失常可能是肺主治节理论体现。本项目探讨糖尿病心肺组织内RCS积聚、RyR2功能异常、免疫因子释放的相关机制及参松养心胶囊干预机制；探索中医肺主治节理论的分子基础。结果发现与正常组相比，糖尿病大鼠血糖及HbA1c升高、血清TBARS水平和SSAO活性均升高；心肺组织MGO、MDA含量升高。与糖尿病组相比，松养心胶囊治疗降低血清TBARS水平、SSAO活性、心组织MGO、MDA含量、肺组织MDA含量。Western blot显示与正常组相比，心脏和肺组织内NLRP3、IL-1、IL-18、IL-6、TGF-β、TNF-α表达上调；心肌p-RyR2、P-CaMKII明显上调；与糖尿病组相比，松养心胶囊治疗降低心肌NLRP3、IL-1、IL-18、IL-6、TGF-β、TNF-α，p-RyR2、P-CaMKII表达以及肺NLRP3、IL-1、IL-18、IL-6表达。电生理数据提示糖尿病体内RCS产物MGO、4-hydroxy-2-nonenal、Glyoxal增加RyR2开放几率，参松养心胶囊抑制RyR2开放几率。这些数据提示糖尿病RCS产物导致相关免疫因子释放、RyR2功能增强是糖尿病心肺并发症的分子基础；参松养心胶囊保护糖尿病心肌机制是抑制心组织内RCS的水平、下调p-RyR2, p-CaMKII及相关免疫因子表达；RCS和免疫相关分子可能是肺主治节的物质基础，仍需进一步探索。