Study On The Relationship Between Human Cytomegalovirus Infection And Myocardial Damage

Objective To investigate the human cytomegalovirus whether or not to damage cardiac myocytes in vitro and vivo and provide the theoretical basic for clinical study of viral etiology and diagnosis of myocarditis。Methods The study is divided into two parts :(一) In vitro: The first one is cultured primary rat myocardial cells. 15 newborn Sprague-Dawley rats of 1-3 days age were prepared. The ventricles muscle were take out to make into myocardial cells with no germ after they were digested with 0.25 per cent of trypsinase. The myocardial cells were moved into the culture bottle to culture them. The second one is the use of HCMV-infected new-born SD rat cultured myocardial cells. Monolayer grown from neonatal rat myocardial cells, divided into two groups:①cell matched control group,②virus matched control group. Growth medium with 100TCID50HCMV were add 0.5ml/hole in Virus infected cells, and the same time only solution were add in the cell matched control group. 37℃, after 2h incubation, the supernatant discarded in each group, and the growth of liquid were add 1ml / hole, we cultivate them in 37℃5% CO2 incubator and stop cultivation when CPE of the virus matched control group appear ++++ effect .We observe cell disease and pulse after infection, and take out each group of cell supernatant to metry myocardial aspartate aminotransferase (AST), lactate dehydrogenase (LDH), creatine kinase (CK), CK-MB and cTnI levels with ELISA method, and identification of HCMV infection in the state of myocardial cells.(二)In vivo: HCMV-induced BALB / C mice were divided into A, B, C three groups. Group A is one of HCMV infection, each mice were injected intraperitoneally 0.5ml with HCMV AD169 strain suspension; group B is one of the HF control group, each mice were injected intraperitoneally 0.5ml with HF suspension ; group C is one of the DMEM control group, each mice were injected intraperitoneally 0.5ml with DMEM suspension . 20 days after injection, we take the blood, myocardial specimens for virus isolation and myocardial pathology and electron microscopy. Results In vitro part : after SD rats cultured rat neonatal myocardial cells infected with HCMV , the cells appear to gradually lesions, cells stop pulse and in the 5 days and 7 days the AST, LDH, CK, CK-MB , cTnI were significantly higher than the control group cells in the culture medium (p <0.05), and the same time HCMV pp65 protein was also positive, by detecting immunofluorescence. In vivo part: The HCMV infection incidence rate of myocarditis in mice is80.0%, two mice were died at the first 5 to 10 days in the virus infection group, and the mortality is20.0% . After HCMV infection during 5 to 20 days, we can see that myocardial cells is swelling, degeneration, and there is substantial inflammatory cell surround blood vessels, and we find cytomegalovirus particles in the myocardial cells under electron microscope. Conclusion 1.the morphocytology and enzymology of cardiac myocyte were changed by infected with HCMV AD169 in vitro and inflammatory reaction were happened in vivo ,so HCMV AD169strain may have the ability of infected myocardial cells.2. HCMV may be one of the important pathogens of the viral myocarditis. In the clinical treatment of children with viral myocarditis, we should consider the anti-viral treatment of HCMV by combination with pathogen detection.