Cardioprotective Effects of Combined Administration of Cryptotanshinone and Salvianolic Acids Against Myocardial Ischemia-Reperfusion Injury by Regulating eNOS/GRK2 Balance Based on Chinese Yin-Yang Theory

Ethnopharmacological relevance: Salvia miltiorrhiza Bunge (Danshen), is a traditional Chinese medicine commonly used to treat cardiovascular diseases. Cryptotanshinone (CT) and salvianolic acids (Danshensu, DSS and Protocatechuic acid, PCA) are the main active constituents in Salvia miltiorrhiza Bunge , both of which have cardioprotective effects through different molecular mechanisms.Aim of the study: This study aimed to investigate the optimal cardioprotective effect of CT combined with DSS or PCA against myocardial ischemia/reperfusion (MI/R) injury in vitro and in vivo based on Chinese Yin-Yang theory.Materials and methods: The hypoxia/reoxygenation (H/R) injury in H9c2 cells and the MI/R injury in rats were established, respectively. Cell viability, myocardial zymetology, oxidative stress status and the infarct size were detected. Protein expressions of eNOS, GRK2 and p-eNOS were measured by Western blot in experimental groups. Cellular thermal shift assay and molecular docking were used to confirm the interaction between PCA, CT and GRK2, eNOS proteins, respectively.Results: Our Results showed that cardioprotective effects of CT combined with salvianolic acids (SAs) were stronger than that of either agent alone. The optimized ratio of CT to SAs was 1:3 among all prescription co-administrations. Importantly, the ratio of endothelial nitric oxide synthase (eNOS) to G protein-coupled receptor kinase 2 (GRK2) was apparently higher in CT+SAs group than that of the single treatment group.Conclusion: The cardioprotective effect of such combined therapy was mainly achieved by regulating the balance of eNOS/GRK2 based on Chinese Yin-Yang theory.