| Objective:In this study,the Chinese medicine Astragaloside IV was loaded with a hyperbranched polymer of 4-vinylphenylboronic acid(PBA)-polyethylene glycol diacrylate 700(PEGDA).React with thiolated hyaluronic acid(HA-SH)and add gold nanoparticles to obtain HB(PEG-PBA)/ HA-SH / GNRs injectable conductive hydrogel loaded with stilbene glycoside.First detect the characterization of the composite conductive hydrogel and the release of drugs in vivo and in vitro;Then,the left anterior descending coronary artery was ligated to prepare myocardial infarction model in rats.The hydrogel system was injected into the rat myocardial tissue in situ and after 28 days,using high-resolution small animal ultrasound imaging,hemodynamics,pathological section staining,immunofluorescence,RT-PCR,Western Blot and other related experimental techniques,To investigate the effect of astragaloside-loaded HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogel on the specific effects of cardiac function,cardiomyocyte conduction and apoptosis signaling pathway in model rats,To explore the effect and mechanism of Astragaloside IV-loaded HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogel on prevention and treatment of myocardial infarction.Methods:1.Synthesis of hyperbranched polymer polyethylene glycol diacrylate-4-vinylbenzeneboronic acid(PEGDA-PBA)using Michael addition reaction,PBA successfully docked in hyperbranched polymer polyethylene glycol diacrylate-4-vinylphenylboronic acid(PEGDA-PBA)adsorbs insoluble water-soluble astragaloside nanospheres through hydrogen bonding between hydroxyl groups,Thereby increasing the drug loading of the hydrogel system;Further adding nano-gold particles to the composite hydrogel system to increase its conductivity;The thiolated hyaluronic acid(HA-SH)reacts with the residual double bond on the polymer to quickly form the hydrogel into an injectable solution.Structural characterization of hyperbranched polymer PBA-PEGDA by nuclear magnetic resonance and infrared spectroscopy;detection of the strength of gel material bymechanical property test;Observe the morphology of nano-scale particles using TEM transmission electron microscope;Measure the conductivity and drug release properties of different groups of hydrogels;And the conductive hydrogel was tested for cytotoxicity.2.The rat model of myocardial infarction was prepared by ligation of the left anterior descending coronary artery,and was randomly divided into a sham operation group,a model group,a hydrogel group loaded with high-dose Astragaloside,a nanogold hydrogel group loaded with low-dose Astragaloside,Nanogold hydrogel group and simple hydrogel group loaded with high-dose Astragaloside IV.The 7th,14 th and 28 th day after the operation were selected to perform echocardiography on the rats in each group to evaluate the changes of left ventricular function.On the 28 th day,samples were taken from each group of rats.(1)Detect the content of lactate dehydrogenase(LDH),creatine kinase(CK)and creatine kinase isoenzyme(CK-MB)in serum by myocardial enzymology;Hemodynamics and high-resolution small animal ultrasound imaging system to investigate the improvement of ventricular wall motion and changes in left ventricular systolic and diastolic function;(2)Through the pathological analysis of HE and MASSON,investigate the influence of drugs on the heart morphology and structure;(3)Through the influence on the expression of CX43(connexin 43),investigate the electrical conductivity and mechanical coupling effect of drugs on myocardial tissue;Through the influence on the expression of ANP(Atrial Natriuretic Peptide),investigate the specific effects of drugs on myocardial tissue;By analyzing the expression of angiogenesis on α-SMA(vascular smooth muscle cell marker)at the margin of infarction of myocardial tissue,the improvement effect of drugs on neovascularization was investigated;(4)Through the verification of the related genes CX43,α-SMA,TNF-α,Ang-1,VEGF,c Tn T,to investigate the effect of drugs on the inside of myocardial tissue.3 Through molecular biology techniques,Fluorescence staining of myocardial tissue TUNEL to detect cardiac apoptosis;Western Blot was used to detect the expression of Bax,Bcl-2,Caspase-3,Cyt-c and Cleaved caspase-3 in myocardial tissue;WesternBlot was used to detect the expression of JNK / MAPK pathway-related proteins in the myocardial ischemic injury tissue of each group,thus indicating the loading of astragaloside HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogel improves cardiac function by inhibiting myocardial fibrosis and reducing cardiomyocyte apoptosis,and explores the possible mechanism of action of drugs.Results:1 Synthesis of hyperbranched polymer HB(PEG-PBA)by chemical synthesis of traditional Chinese medicine,and then chemical reaction with 4-vinylphenylboronic acid HA-SH,and preparation of traditional Chinese medicine astragaloside IV gold nanoparticle solution,chemically these three parts Cross-linking reaction,through the bonding of chemical bonds,the hydrogen bonding of PBA and astragaloside IV was successfully docked to enhance the drug loading of the hydrogel,thereby preparing astragaloside-loaded HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogel.The release properties of the hydrogels were found by testing the release properties.The results show that the hydrogel has certain practicability,low toxicity and harmlessness,and can be applied to the heart.2 By injecting astragaloside-loaded HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogel into the marginal area of myocardial infarction model rats.The results showed that the left ventricular function was improved after drug intervention;And reduce the content of LDH,CK,CK-MB in ischemic rats;HE and MASSON staining results show that astragaloside-loaded HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogel can reduce inflammatory cell infiltration,inhibit myocardial fibrosis,reduce ventricular remodeling and delay ventricular remodeling.Further by immunofluorescence analysis of CX43,ANP and α-SMA,the results showed that myocardial electrical activity of myocardial tissue was improved after drug intervention and cardiac function was improved;The genetic studies of CX43,ANP,α-SMA,VEGF,Ang-1 and c Tn T have also been confirmed.3 TUNEL fluorescence staining and Western Blot were used to detect the expression of Bax,Bcl-2,Caspase-3,Cyt-c,Cleaved caspase-3 and JNK / MAPK pathway-related proteins in myocardial tissue,confirming that astragaloside glucosideloaded with HB(PEG-PBA)/ HA-SH / GNRs Gum has an anti-apoptotic effect.Analysis conclusion:Through the above studies,it was clarified that astragaloside IV-loaded HB(PEG-PBA)/ HA-SH / GNRs conductive hydrogels can improve cardiac function,inhibit myocardial fibrosis,delay ventricular remodeling,and cardiac function in myocardial infarction model rats Electrical conductivity may be related to JNK / MAPK apoptotic signaling pathway. |
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