| BackgroundWith the increasing living standards and the aging population, the incidence of coronary heart disease (CHD) is gradually increasing. In the treatment of CHD, revascularization is currently the most common application; but the ischemia reperfusion (IR) injury can impair the systolic and diastolic functions of the heart. Therefore, how to effectively reduce the myocardial IR has important practical significance.The current view is that oxygen free radical (OFR), myocardial intracellular calcium overload, the neutral granular cell activation, reduce and fluidity of cell membrane, cell membrane permeability increased and in IR injury. Free radical is refers to the floorboard of a single unpaired electrons in atoms, groups of atoms or molecules in the outer electron orbitals, including nonfat free radical and lipid free radical, which is necessary for the maintenance of normal life activities. The former mainly refers to OFR, including superoxide anion and hydroxyl radical, etc. Free radicals as the normal metabolism of human body, has a positive role in defense of removal harmful substances under physiological conditions. Too much or too little radical will adversely affect the body. Excessive production of free radicals, especially OFR, has strong oxidation ability, through oxidation attack the large biological molecules, changing in composition of the cell membrane and the formation of lipid triad, promoting the lower cell membrane fluidity, and activating neutrophils, damaging mitochondrial, which lead to the destruction of the structure and function of organism. It is reported that in the literature OFR is one of the direct causes of myocardial IR injury, which is more obvious to the injury of cell membrane, which leads to intracellular calcium overload, and forms a vicious circle, eventually leads to cell death.There is still no effective drug for the treatment of myocardial IR damage. At present, there are two kinds of the scavengers for free radicals in the organism, namely enzymes and non-enzyme compounds. Enzyme scavengers has the following characteristics:high molecular weight, low stability, sensitive to physical and chemical factors, difficulty for transdermal or transdermal absorption. Non enzymatic scavenger has the following characteristics:strong reducibility, poor selectivity, may cause endogenous redox imbalance. Therefore, it has become a focus in the field of medicine and life science to study a clear and selective free radical scavenger. In 2007 the Japanese scholars found that hydrogen can play a protective effect on IR injury by selectively scavenging free radicals. The hydrogen dissolved in the liquid can produce selective neutralization of IR in the process of hydroxyl radicals and peroxynitrite. Then some scholars found that respiratory hydrogen also play an important role in the protection of the liver, retina IR damage.ObjectiveThis study intends to use a rat in vivo myocardial IR model and cardiac myocytes in vitro hypoxia-reoxygen model, to study hydrogen rich saline play an protective effect on myocardial cell membrane in IR injury and its mechanisms, hydrogen treatment of ischemic myocardium ischemia-reperfusion IR injury and provide relevant evidence theory basis, also for the clinical treatment IR myocardial ischemia reperfusion injury will provide a new way of thinking. To provide relevant theoretical basis for the hydrogen treatment for myocardial IR injury, and to provide new ideas for the clinical treatment of myocardial IR.MethodsIn vivo experimental study:Adult male SD rats were randomly divided into five groups: sham operation group (only the implementation of the open chest surgery, and needle through the front left down a margin, not ligation), the IR group (ligation of the left anterior descending caused myocardial ischemia, half an hour for ligation,2 hours for reperfusion), the saturated H2 saline treated groups (5min before myocardial reperfusion, given different concentrations (5, 10,20 ml/kg) saturated H2 saline pretreatment by intraperitoneal injection, the rest as the same as IR group). The changes of calcium channel flow in different groups were detected by patch clamp technique and laser scanning confocal microscopy. The fluidity of myocardial cell membrane was detected by electron spin resonance. The activity of Bcl-2 and Bax were determined by Western-blot and immunohistochemistry. Apoptosis rate of myocardial cells was measured by Tunel staining method.In vitro experimental study:In vitro cultured neonatal rat cardiomyocytes was used a model of hypoxia/reoxygenation to simulate the IR damage in vivo. The myocardial cells were divided into 3 groups:Control group:myocardial cells was cultured 2.5 h,5% carbon dioxide and 95% air,37°. IR group:myocardial cells was cultured 5% carbon dioxide and 95% nitrogen, 37°for 30 min minutes, then 2 h hours for 5% carbon dioxide and 95% air,37°. H2 treatment group:the H2 rich medium was given before hypoxia/reoxygenation, and the others were treated with IR group. The viability of cardiac muscle cells was compared by CCK8 method, and the metabolic function indexes (LDH, CK) were determined.Results1. Compared with the sham operation group, the total phospholipid content, cholesterol content, cell membrane fluidity and regional mobility in H2 group and IR group were significantly lower than those in sham group. In the H2 groups, the trend of this decrease was effectively alleviated, especially in the 20 ml/kg group. This indicates that H2 saline can affect the overall flow and regional mobility of the myocardial cell membrane after reperfusion.2. Calcium overload level in IR group of was significantly higher than that in sham group. While, in H2 groups the calcium overload level were lower than that in the IR group, especially the 20 ml/kg was the most significant. It is showed that the H2 saline can effectively reduce the calcium overload during the IR process.3. Compared the viability of cells between each groups using CCK8 and analysis myocardial metabolic function. The viability of cells in IR group were significantly lower than those in H2 group, and the metabolic function indexes (LDH, CK) is significantly higher in IR group than in H2 group. It is showed that the H2 saline can effectively reduce the cell damage during the IR process.4. Myocardial apoptosis and apoptosis related protein in IR group were significantly higher than those in the H2 groups. It is proved that the H2 saline has a significant anti apoptotic effect in the process of IR.Conclusions1. Saturated H2 saline have the protective effect on the myocyte membrane fluidity by persist the ratio of phospholipids and cholesterol.2. Saturated H2 saline provides stablisition effect on calcium homeostasis by persist the function of L type calcium channel.3. Saturated H2 saline can decrease the apoptosis rate of myocardial cells and inhibit the activity of apoptosis related proteins, so as to play a protective role in the myocardium. |
PDF Full Text Request