Tingli Dazao Decoction pretreatment ameliorates mitochondrial damage induced by oxidative stress in cardiomyocytes

Tingli Dazao Decoction (TLDZD) recorded in "Synopsis of Prescriptions of the Golden Chamber" is a classical prescription used for the treatment of heart failure nowadays. The studies of TLDZD were mainly focused on clinical practice where the formula was usually combined with other medicinal herbs. Chemical composition and cardiovascular pharmacological research of TLDZD were still insufficient.This study aimed to investigate the chemical constituents of TLDZD, evaluate the effects of TLDZD on mitochondria of myocardial cells under oxidative stress, and identify its potential cardioprotective components.Chemical composition analysis of TLDZD was performed by ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry. An in vitro oxidative stress model of cardiomyocytes was established by treating H9c2 cells with tert-butyl hydroperoxide (tBHP). The impact of TLDZD and its components on the production of cellular reactive oxygen species (ROS) and mitochondrial ROS (mROS), the level of malonaldehyde as well as the structure and function of mitochondria were evaluated. The effect of TLDZD on AKT/Nrf2/HO-1 signaling pathway in cardiomyocytes under oxidative stress were observed.Seventy-eight compounds were characterized from TLDZD, among which flavonoids, glucosinolates and phenylpropanoids were abundant, and a small number of cardiac glycosides and alkaloids also existed in TLDZD. Pretreatment with TLDZD significantly attenuated cell death, accompanied by decreased ROS and mROS production, reduced malonaldehyde level, lower mitochondrial membrane potential and adenosine triphosphate content in H9c2 cells stimulated with tBHP. The active components were mainly flavonoids of TLZ represented by quercetin-3-O-β-D-glucose-7-O-β-D-gentiobioside. In mechanism, the cardioprotective effect of TLDZD was proved to be associated with the activation of the AKT/Nrf2/HO-1 signaling pathway.The chemical profile of TLDZD was comprehensively investigated. Flavonoids with quercetin-3-O-β-D-glucose-7-O-β-D-gentiobioside as the representative, were the main component in TLDZD responsible for attenuating mitochondrial oxidative damage in cardiomyocytes.