Pre‑treatment with a combination of Shenmai and Danshen injection protects cardiomyocytes against hypoxia/reoxygenation‑ and H2O2‑induced injury by inhibiting mitochondrial permeability transition pore opening

Abstract

Increasing evidence has indicated that opening of the mitochondrial permeability transition pore (mPTP) has a vital role in myocardial ischemia/reperfusion (I/R) injury. Shenmai injection (SMI) plus Danshen injection (DSI) combination, termed Yiqi Yangyin Huoxue (YYH) therapy is used in the clinic to treat cardiovascular diseases, including myocardial I/R injury. Previous studies by our group have demonstrated the protective effect of pretreatment with YYH against myocardial I/R injury in isolated rat hearts. The present study aimed to examine the protective effect of YYH against hypoxia/reoxygenation (H/R)-and H 2 O 2-induced cardiomyocyte injury, and to determine whether this effect is produced by inhibition of mPTP opening. Primary cardiomyocytes isolated from neonatal rats were cultured and randomly grouped into a control group, injury group and pretreatment group, with six duplicated wells in each group during each assay. Cardiomyocytes in the injury group were subjected to H/R to simulate I/R or exposed to H 2 O 2 for 2 h to induce oxidative injury. Cellular injury was assessed via release of creatine kinase (CK) and lactate dehydrogenase (LDH), and cell viability was measured by an MTT assay. The mitochondrial membrane potential (ΔΨm) and cytosolic reactive oxygen species (ROS) were detected using the fluorescent probes rhodamine123 (Rh123) and chloromethyl-2,7-dichlorodihydrofluorescein diacetate (CM-H 2 DCFDA), respectively. Intracellular Ca 2+ , mitochondrial Ca 2+ and mPTP opening were measured using fluo-4 acetoxymethyl (Fluo-4/AM), rhodamine-2 acetoxymethyl (Rhod-2/AM) and calcein acetoxymethyl (Calcein/AM) probes, respectively. The results indicated that pretreatment with YYH enhanced cell viability, increased ΔΨm, reduced CK and LDH release, and decreased intracellular ROS and Ca 2+ , thus reducing cardiomyocyte injury induced by H/R or H 2 O 2. LY294002, a specific phosphoinositide 3-kinase (PI3K) inhibitor, and PD98059, a specific inhibitor of the extracellular signal-regulated kinase 1/2 (Erk1/2) pathway, eliminated the protective effects of the combination therapy on cell viability and the change in the ΔΨm in cardiomyocytes. In conclusion, pre-treatment with YYH has cardioprotective effects against H/R injury and oxidative stress via activation of the PI3K/Akt and Erk1/2 signaling pathways, which reduces mPTP opening, overproduction of ROS and calcium overload.

Salvia miltiorrhiza in Treating Cardiovascular Diseases: A Review on Its Pharmacological and Clinical Applications

“…Danshen, Danhong, salvianolate, STS injections, and other SM preparations are widely used in China to treat stable AP (SAP) caused by CHD (Zhang et al, 2018a). Salvianolate injections are composed of water-soluble extract of SM (Han et al, 2011; Li et al, 2019). Danhong injection is a modern patented Chinese medicine extracted from SM and Flos Carthami (Zou et al, 2018; Feng et al, 2019).…”

Thyroid stimulating hormone aggravates diabetic retinopathy through the mitochondrial apoptotic pathway

“…Moreover, oxidative stress causes cellular lipid peroxidation, with an increased level of MDA, which further permeabilizes the mitochondrial outer membrane (Anuradha et al, 2001; Sandbach et al, 2001). This process facilitates the release of cytochrome c, and promotes the activation of caspase 9 and other effector caspases, ultimately contributing to endogenous apoptosis (Li et al, 2019; Raupach et al, 2019). Here, high glucose induced excessive ROS generation in mitochondria, which reduced ΔΨm and ATP concentrations, and increased MDA levels, suggesting a disruption of mitochondrial integrity and function.…”

PHLDA3 inhibition attenuates endoplasmic reticulum stress‑induced apoptosis in myocardial hypoxia/reoxygenation injury by activating the PI3K/AKT signaling pathway

“…Neonatal rat cardiomyocyte (NRCM) culture. NRCMs were isolated from the ventricles of 144 1-3-day-old male Sprague-Dawley rats (body weight, 15-20 g) as previously described (9,(14)(15)(16). Animals were purchased from the Animal Experimental Center of Hainan Medical University (Haikou, China).…”