Proteomics-guided Studies On Cardioprotective Effects Of Xuesaitong Injection And Shenmai Injection

The traditional Chinese medicine usually exhibited the therapeutic effects by multiple components, multiple targets and multiple mechanisms. Proteomics is a powerful tool for revealing the active compounds and mechanisms of traditional Chinese medicine. Herein, proteomics-based methods were used to investigate the cardioprotective effects and active compounds of Xuesaitong Injection and Shenmai Injection. The main action modes of these injections were explored in animal models, and the mechanisms for cardioprotective were further verified on the celluar models. The main contents of this dissertation are as follows:1. Xuesaitong Injection could reduce the serum contents of CK-MB and LDH in rat model of myocardial ischemia-reperfusion. Two-dimensional gel electrophoresis was used to measure the profiling of protein expression of ischemia heart tissues. Differentially expressed proteins among MI rats and Xuesaitong Injection-treated rats were identified using MALDI-TOF-MS/MS. The results showed that one protein was up-regulated in myocardial ischemia-reperfusion rats, whilst six proteins were down-regulated. They exhibited correction after treating with Xuesaitong Injection. These differentially expressed proteins were classified according to their biological function, which mainly related to improve the expression of pyruvate dehydrogenase, peroxiredoxin 3 and so on. Our findings suggest that improving energy metabolism as well as anti-oxidative stress were the main pathway of Xuesaitong Injection on myocardial ischemia-reperfusion.2. We mainly investigated protective mechanism of Xuesaitong Injection in H9c2 cardiomyocytes by using the hypoxia/reoxygenation and oxidative damaged model. The results showed that Xuesaitong Injection had a dose-dependent protection on H9c2 cardiomyocytes damaged by hypoxia/reoxygenation. Preincubation of the cardiomyocytes with Xuesaitong Injection considerably attenuated the decreased PDH activity, Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities, ATP content and acetyl-CoA level while prominently inhibited the increased pyruvate level induced by hypoxia/reoxygenation in a dose-dependent manner. By the western blotting method, we found that the treatment of H9c2 cardiomyocytes with hypoxia/reoxygenation caused decrease in the expression of Pyruvate dehydrogenase E1 alpha and ATP synthase. However, pretreatment of the cardiomyocytes with Xuesaitong Injection considerably attenuated the decrease of ATP5A and PDHA1 expressions. While in the oxidative damaged model, Xuesaitong Injection had a dose-dependent of increasing cell survival as well as decreasing LDH release and MDA contents. From the cellular level, it was suggested that Xuesaitong Injection could improve energy metabolism by restoring pyruvate dehydrogenase activity, as well as anti-oxidative stress by reducing the release of LDH and MDA contents.3. Preliminary results of proteomics suggest that the therapeutic mechanisms of Shenmai formula are mainly related to improving energy metabolism. Therefore, oxidative damaged of myocardial cell model was used for the identification of active compounds of Shenmai formula. Totally,12 components with moderate anti-oxidant activity on the model of tert-butyl hydroperoxide damaged H9c2 cardiomyocytes were identified. By using LC-MS technology,24 probable compounds were speculated. Studying the active compounds and mechanisms of Shenmai Injection on the cardioprotective activity, the results showed that preincubation of the H9c2 cardiomyocytes with Shenmai Injection considerably attenuated the decreased ATP content, PDH activity and acetyl-CoA level while prominently inhibited the increased pyruvate level damaged by oxidative in a dose-dependent manner. Ophiopogonin D and ginsenoside Ro have simultaneously increased ATP content, PDH activity as well as reduced pyruvate content, and thus improved energy metabolism and protected cardiomyocytes.In summary, a proteomics-based method was employed to investigate on the cardiopretective effects of traditional Chinese medicine. We found that Xuesaitong Injection could protect myocardial cells through the ways of improving the activity of PDH and the content of ATP, reducing the release of LDH and MDA content. Ophiopogonin D and ginsenoside Ro in Shenmai Injection may exhibitited potent cardioprotective effects through improving energy metabolism, including increasing ATP content, PDH activity and restoring pyruvate metabolic pathway. This research provided new evidences for further comprehending the therapeutic mechanisms of Xuesaitong Injection and Shenmai Injection for treating cardiovascular diseases.