Effects And Mechanisms Of Asiaticoside And Madecassoside On Myocardial Ischemia/reperfusion Injury

Centella asiatica (Hindi name Brahma-manduki, Family Umberlliferae ) occupies an important place in the traditional Chinese medicine, is used in folk medicine to treat a wide range of indications. Centella asiatica also appears to be effective in the treatment of wound healing disturbances. Asiaticoside(AC)and Madecassoside( MC), are the principle terpenoids in centella asiatica, have been shown to promote fibroblast proliferation and collagen synthesis. However, whether asiaticoside could exert cardiovascular biologic effects remains unclear. In the progress of screening for natural compounds with anti-ischemia activity, it was found that AC and MC can protect isolated rat hearts and cultured neonatal rat cardiomyocytes against reperfusion injury in our previous studies. Therefore, this study focused on the effects and mechanisms of AC and MC on prevention from the cardiomyocytes ischemia and the ischemia/reperfusion injury model in vivo and in vitro.1. Effects and its mechanisms of AC on myocardial ischemia injuryThe ischemia reperfusion model was established in isolated rat hearts. The infarcted areas were expressed as percentage. The coronary flow (CF) and heart rat (HR) were monitored. Cardiomyocytes ischemia model in rats and ischemia/reperfusion injury model in rabbits were established. Left ventricular function was monitored at the timepoints during the reperfusion period to assess the extent of postischemic recovery in comparison with baseline value. The infarcted areas were expressed as percentage. The levels of lactate dehydrogenase (LDH), creatinephosphokinase (CK), malondialdehyde (MDA), super-oxide dismutase(SOD) and C-reactive protein (CRP) in serum were determined. Cardiomyocytic apoptosis were measured by TUNEL staining. Treatment with AC (24, 12, 6mg/kg) during ischemia reperfusion injury from duodenum in vivo attenuated myocardial damage characteristic of decreasing infarct size, decreasing LDH and CK release. Activities of SOD were diminished and MDA level increased obviously in IR group whereas pretreatment with AC significantly blunted the decrease of SOD activity, markedly reduced the level of MDA and the activity of CRP, relieved myocardial cell apoptosis. These results showed that AC has the protective effect on myocardial ischemia/reperfusion injury possibly through its roles of anti-lipid peroxidation, enhancement of SOD activity, anti-inflammation and anti-apoptosis.2. Effects and its mechanisms of MC on myocardial ischemia/ reperfusion injuryThe ischemia reperfusion model was established in isolated rat hearts and in rabbits. The right carotid artery and femoral vein were cannulated to monitor left ventricular pressure as its first derivative (±dp/dt), and a vinyl catheter was inserted into femoral artery to monitor blood pressure via a transducer attached to a multi-channel recorder. Myocardial ischemia was confirmed by regional cyanosis of myocardial surface and electrocardiographic (ECG) change. The infarcted areas were expressed as percentage. The levels of LDH, CK, MDA, SOD and CRP in serum were determined. Cardiomyocytic apoptosis were measured by TUNEL staining. The monoclonal antibody of Rabbit anti-Goat Bcl-2 proteins was used as the primary antibody for Bcl-2 immunohistochemical staining. We also built the cardiac arrhythim model of ischemia/ reperfusion injury in rats to study the protection of MC on delaying the time of arrhythim.In cultured neonatal rat cardiomyocytes, mechanisms of MC on myocardial ischemia/ reperfusion injury were investigated. Release of LDH and CK in the medium was determined as injury index of cardiomyocytes. The contents of Ca²⁺, ROS and ATP in cardiomyocytic cells were determined by Fluostar luminometer.Treatment with MC (3.2, 1.6, 0.8 mg/kg) during ischemia reperfusion injury in vivo attenuated myocardial damage characteristic of decreasing infarct size. Pretreatment with MC significantly blunted the decrease of LDH, CK and SOD activity, markedly reduced the level of MDA and the activity of CRP, relieved myocardial cell apoptosis and upregulate with the expression of Bcl-2. The study of mechanisms findings demonstrate that MC has the protective effect on myocardial ischemia/reperfusion injury which possibly contributes to the inhibition of ROS and Calcium overload, antioxidation, anti-inflammation and anti-apoptosis by upregulated with the expression of Bcl-2 and ATP.