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BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) can grow in host myocardium, differentiate under myocardial condition, improve cardiac function. However, biological characteristics of BMSC differentiation are still unclear presently.OBJECTIVE: To study the expression and electrophysiological characteristics of BMSC/n vitro connexin-43 following 5-azacitidine (5-aza) treatment.DESIGN, TIME AND SETTING: The cytological in vitro controlled study was perormed at the Heart Center, Hebei Provincial People's Hospital from July 2007 to February 2009.MATERIALS: A total of 24 male pigs aged 2 months were purchased from Exparimental Animal Center, Hebei Medical University.METHODS: Bilateral femoral bone marrow was obtained from pigs under sterile condition. BMSCs were harvested by Percoll density gradient in vitro. At passage 2, BMSCs were treated with 10 μmol/L 5-aza, and incubated in DMeM without inductor 24 hours later. Indices were measured I, 2, 3 weeks following induction. A control group was set up, which was not treated with 5-azacitidine. Following bone marrow extraction, experimental pigs were anesthetized to obtain ventricular muscle. Normal ventricular muscle cells were isolated and cultured by tissue block enzyme digestion method.MAIN OUTCOME MEASURES: Expression of connexin-43 was measured by immunohistochemical staining (ABC method). Ito current density and action potential were determined by patch clamp technique.RESULTS: At 1 and 2 weeks following 5-aza induction, some BMSCs were positive for connexin-43, with the presence of brown particles surrounding nuclei. At 3 weeks, positive rate of connexin-43 was 95%. The area with large cell density was presented with similar structure to normal myocardium. At +80 mV, compared with normal myocardial cells, Ito current density was significantly reduced in BMSCs following 1 and 2 weeks and in the control group (P < 0.05). Ito current density was significantly increased to a normal levels in BMSCs 3 weeks following induction (P > 0.05). No action potential was detected in BMSCs following 1 and 2 weeks of 5-aza, and action potential could be determined 3 weeks following induction, which was identical to normal myocardial cells.CONCLUSION: Through induced by 5-aza for three weeks, BMSCs have the similar expression of connexin-43 and electrophysiological characteristics as normal myocardium.
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BACKGROUND:Bone marrow stem cell transplantation can improve heart function and prevent ventricle remodeling.At present,the adult bone marrow stem cells used for transplantation primarily included bone marrow mononuclear cells (BM-MNCs) and mesenchymal stem cells (MSCs),and endothelial progenitor cells.The curative effects and precise mechanisms of transplantation of various bone marrow stem cells remain unknown.OBJECTIVE:To compare the effects of transplantation of autologous BM-MNCs and MSCs via the coronary artery on ventricle remodeling subsequent to acute myocardial infarction (AMI). DESIGN,TIME AND SETTING:Randomized controlled animal experiment performed at the Center for Clinical Research,Hebei Provincial People's Hospital,Electron Microscope Room,Hebei Medical University between March 2005 and December 2006.MATERIALS:Thirty-six male Jizhong pigs,were randomly divided into 4 groups:control group (n = 6),infarct model group (n = 10),BM-MNC group (n = 10),and MSC group (n = 10).METHODS:Porcine autologous BM-MNCs were isolated by gradient density centrifugation,and MSCs were obtained by adherence method.Prior to transplantation,both BM-MNCs and MSCs were colloidal gold labeled.Except the infract model group,pigs in the other 3 groups were developed into AMI models by oppressing the left anterior descending branch with balloon catheter.Ninety minutes after modeling,(6.0±1.3)×107 autologous BM-MNCs and (4.5±2.1)x 107 MSCs were respectively transplanted into pigs in the BM-MNC group and the MSC group via the coronary artery and cultured for 28 days.MAIN OUTCOME MEASURES:Observation of pathological changes of cardiac muscle tissue by light and electron microscope;Examination of cardiac function by ultrasonograph;Detection of the number of blood vessels and apoptotic myocardial cells,and expression of nuclear factor-κB (NF-κB) and troponin Ⅰ and its correlation to cardiac function by immunohistochemistry;Detection of mRNA expression of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in the cardiac tissue as well as its correlation to cardiac function by reverse transcription-polymerase chain reaction (RT-PCR).RESULTS:In the MSC group,there was proliferation of a great deal of blood vessels as well as growth of abnormal cell masses around the coronary vessels,while the BM-MNC group exhibited the "budding" of many capillary vessels.Prior to transplantation,cardiac function indices were basically similar among each group (F = 1.550,P＞0.05).Twenty-eight days after transplantation,left ventricular ejection fraction was significantly lower in the control,BM-MNC,and MSC groups than in the infarct model group (F = 5.30,P＜0.05),while endocardial fractional shortening was significantly higher (F = 10.67,P＜0.01).Compared with the infarct model group,the number of blood vessels in the infarct zone and infarct border zone was increased in the BM-MNC group (F=29.56-34.87,P＜0.01) and had no apparent change in the MSC group.In the BM-MNC and MSC groups,apoptotic myocardial cells in the infarct zone and infarct border zone were significantly reduced (F=14.31-35.34,P＜0.01 ) and troponin I expression rate was significantly increased (F=19.05,P＜0.01 ),as compared with the infarct model group.In addition,NF-κB positive rate in the infarct border zone was significantly lower in the BM-MNC and MSC groups than in the infarct model group (F=19.05,P＜0.01).VEGF gene expression level in the infarct border zone was significandy higher in the BM-MNC group than in the infarct model group and MSC group (F = 49.41,P＜0.01).bFGF gene expression level in the infarct border zone was significantly higher in the MSC group than in the infarct model and BM-MNC groups (F=4.71,P＜0.01).LVEF was negatively correlated to myocardial cell apoptosis rate and NF-κB level (r=-0.441 1,P＜0.05;r=-0.579 6,P＜0.01 ).LVEF was positively correlated to number of blood vessels,VEGF and bFGF expression (r=0.775,P＜0.01;r=0.565 1,P＜0.05;r=0.573 5,P＜0.05).CONCLUSION:Transplantation of both autologous BM-MNC and MSC via coronary artery can improve the condition of left ventricular remodeling subsequent to myocardial infarction.The improvement of cardiac functions is related to the increase of blood vessels,VEGF and bFGF expression,the decrease of myocardial cell apoptosis and NF-κ B level in cardiac muscle tissues after stem cell transplantation.BM-MNC transplantation better promotes blood vessel proliferation and VEGF expression in the cardiac tissue but produces worse effects on bFGF gene expression than MSC transplantation.
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Objective To investigate the factors and rate of the forming of the collateral feeding arteries from inferior phrenic artery(IPA) in hepatocellular carinoma(HCC).Methods IPA was demonstrated on angiographs in 137 patients with HCC during transcatheter arterial chemoembolization(TACE),TACE was performed through diaphragmatic inferior artery(DIA) super setectively.Results Of 137 cases,21 cases underwent TACE through DIA(15.3%),of them 2 cases were recurrent after surgical operation(9.5%),3 cases had the tumor receive blood supply from IPA at first TACE(14.3%) and 16 cases were occured after TACE tow and more times(76.2%).Conclusion Collateral branches originated from IPA are important feeding arteries in HCC,TACE of IPA can be performed with a high success rate without major complications.