Research On Mechanism Of Cell Membranes' Immobilization And Protective Effect Of Yi Qi Fu Mai Powder Injection On Myocardial Injury

Objective:1.To investigate the mechanism of cell membrane immobilized on porous silica beads,and to promote the preparation of biomimic carriers for chromatographic methods.2.To explore the protective effect of Yiqi Fumai powder injection(YQFM)on doxorubicin(DOX)induced cardiotoxicity by established cell and animal models.Methods: Rabbit red blood cell membranes were prepared by hypotonic swelling method.Cell membrane modified carriers were prepared according to literature with modification.Batch immobilization studies were carried out under type of carriers,pore size of porous silica beads,enemy activity and kinetics of cell membrane immobilization.High performance liquid chromatography(HPLC)was used to investigate the binding ability and stability of porous silica spheres with positive drug.The distribution of the fluorescently labeled cell membranes on the prepared porous and nonporous silica spheres was further observed by laser scanning confocal microscope.The protective effect of YQFM on DOX-induced cardiotoxicity was firstly to establish DOX-induced H9c2 model of myocardial injury.Different concentrations of YQFM 2 h in advance,and then incubated with DOX for 48 h.LDH and ATP levels were measured by CCK-8 kits.Cell apoptosis was observed by Hoechst staining.The mitochondrial membrane potential was detected by JC-1 probe.The expression of caspase-3 was detected by western blot.Then,DOX-induced acute and chronic models of myocardial injury in mice were established.The effect of YQFM was determined by measured the changes of body weight and heart index,serum creatine kinase(CK)levels and HE staining in the heart.Results: According to type of carriers,pore size of porous silica beads,enemy activity and kinetics of cell membrane immobilization,we found that the silanol group(Si-OH)as a polar ligand displays a strong and irreversible adsorption of cell membranes,facilitating their immobilization onto the silica surface.Also,silica beads with the appropriate pore diameter would be benefit to the immobilization of cell membrane.The optimized column has successfully applied for the identification of a model drug nimodipine with high retention factor and good stability.Finally,theinsertion/self-fusion mechanism involved in the cell membrane immobilization had further been confirmed by laser scanning confocal microscope.The protective effect of YQFM on DOX-induced cardiotoxicity in vitro cell experiments,the results found that DOX decreased H9c2 cell viability in the case,the vlabllity rate of H9c2 cells was increased in a dose-dependent manner after pretreatment with YQFM.YQFM pretreatment group significantly reduced intracellular LDH activity,increased ATP content,compared with the DOX control group.YQFM pretreatment significantly reduced DOX-induced apoptosis and reduced mitochondrial membrane potential in DOX-induced H9c2 cells by Hoechst staining and JC-1 probe.YQFM down-regulated the expression of caspase-3 and reduced the apoptosis.DOX-induced acute models of myocardial injury in mice of DOX dose was determined to be 20 mg/kg,the chronic model of DOX dose of 3 mg/kg every other day,a total of 6 times.There was no significant difference between YQFM groups and DOX control group in the changes of body weight and heart index.The results of CK in serum showed that YQFM reduced the content of CK in acute experiment.The results of HE staining showed that YQFM groups had protective effects on DOX induced myocardial injury in acute and chronic models.Conclusion: In conclusion,an insertion/self-fusion mechanism of the cell membrane immobilization has been proved.Clarification of cellular membrane immobilization mechanism is to facilitate the membrane proteins immobilization with the enhanced loading and improved stability,and provide criteria for the preparation of other biomimic carriers.The protective effect of YQFM on DOX-induced cardiotoxicity confirmed that YQFM inhibited the apoptosis of H9c2 cells,and alleviated the damage of cardiac tissue.Through cell and animal model,detection of cell vlabllity,mitochondrial membrane potential,apoptosis,myocardial enzymes were proved.In order to provide some theoretical basis for clinical rational use of drugs.