Research Of EPC Change With Coronary Heart Diseases Patients And Effect Of LPS And Rosiglitazion On EPC

Charpterâ… The reseach of isolation,culture,characterization of endothelia progenitor cells in human peripheral bloodTo determine the isolation and culture of endothelial progenitor cells from peripheral blood in healthy adults,and reseach cell shape,amount,activity and clusters.Methods:Mononuclear cells isolated from peripheral blood of healthy adults by Ficoll-density centrifugation.The isolated cells were cultured in 1640 medium supplemented with VEGF₁₆₅and bFGF.The EPC specific surface mark CD34⁺ and KDR were assessed by fluorescence activated cell sorter(FACS)analysis.After in vitro cultivation,diferentiating EPCs were identified asadherent cells double positive for Dil-acLDL and FITC-UEA-1 by direct fluorescentstaining under a laser scanning confocal microscope.,The RT-PCR were assessed to characterizate the market of EPC,KDR,CD31,VE-Cadherin.EPC migration was assayed by MTT assay.,and reseach cell shape,amount, activity and clustersConclusion:1.Mononuclear cells isolated from peripheral blood will be induced endothelial progenitors cells by VEGF₁₆₅and bFGF. 2.Endothelial progenitors cells assessed by fluorescence activated cell sorter(FACS)analysis,laser scanning confocal microscope.and RT-PCR.3.Epc have good migration capacity by VEGF₁₆₅and bFGF induced.Charpter 2 Reseach of endothelial projenitor cells from prepheral bloode in patients with coronary heart diseasesPartâ… .The study on changes in the number and function of endothelial progenitor cells from coronary heart diseasesObjeeive To investigate the isolation and culture of endothelial progenitor cells from peripheral blood in patients with coronary heart diseases,,and to determine cell shape,amount,activity and clusters.And study the correlation of EPC number and the degree of coronary artery stenosis and risk factors aggregation.Methods:Mononuclear cells isolated from peripheral blood of patients with coronary heart diseases(n=57)and control(n=30)by Ficoll-density centrifugation.The isolated cells were cultured in 1640 medium supplemented with VEGF and bFGF.The EPC specific surface mark CD34 and KDR were assessed by fluorescence activated cell sorter (FACS)analysis.EPC were characterized as adherent cells double positive for DiL-acLDL uptake and lectin binding by direct fluorescent staining under a laser scanning confocal microscope.EPC migration were assayed by MTT assay.And study the correlation of EPC number and the degree of coronary artery stenosis and risk factors aggregation.Results:1.The number of EPC was significantly reduced in patients with CAD compared with control subjects.In addition,the number of cell clusters,migration capacity w as also impaired in patients with CAD.2.EPC number and function were decresed accompaniment with the degree and extent of coronary artery stenosis,and risk factors aggregation.3.the number of EPC have significant correlation with aging, hypertension,smoking,high-LDL.Conclusion The number,the number of cell clusters,migration capacity of EPC was significantly reduced in patients with CAD compared with control subjects.The number of EPC have nagative correlation with the degree and extent of coronary artery stenosis,the number of EPC have significant correlation with aging,hypertension, smoking,high-LDL. Partâ… .The study on changes in the tPA and PAI and inflammatory factor of EPC with coronary heart diseasesObjecive To investigate the change of number and function of EPC with coronary heart diseases patients;To study the change of EPC with tPA and PAI and inflammatory factor.Methods:EPC isolated from peripheral blood of patients with coronary heart diseases(n=57)and control(n=30)and cultured.There are determing the concentration and activity of tPA and PAI by ELISA. There are determing the expression of tPAmRNA and PAImRNA,VCAM-1 mRNA and ICAM-1mRNA,and PPARγmRNA by RT-PCR.There are determing the high erpression VCAM-1 and ICAM-1 on cell membrane by immunohistochemical.Results:1 The concentration and activity of tPA have decresed on prepheral bloode-serum in patients with coronary heart diseases,and the tPA mRNA expression have degression tendency by RT-PCR.2 The concentration and activity of PAI have incresed on prepheral bloode-serum in patients with coronary heart diseases,and the PAI mRNA expression have upgrade tendency by RT-PCR.3 The PPARγmRNA expression have degression tendency by RT-PCR.in patients with coronary heart diseases4 There are determing the high erpression VCAM-1 and ICAM-1 on cell membrane by immunohistochemical,the VCAM-1 mRNA and ICAM-1mRNA expression have upgrade tendency by RT-PCR.Conclusion The fuction of tPA and PAI have be inhibited,and inflammatory factor excreted have increased,and PPARγmRNA expression have degressed in patients with coronary heart diseasesCharpter 3 Effect of LPS and Rosiglitazion on tPA and PAI and inflammatory factorObjeeive To investigate the EPC migration capcity change by LPS and Rosiglitazion effect.And study the function change of tPA and PAI and inflammatory factorMethods:EPC isolated from peripheral blood of health adult.After LPS and Rosiglitazion incubation EPC to determine their fuction.There are determing the concentration and activity of tPA and PAI by ELISA. There are determing the expression of tPAmRNA and PAImRNA,VCAM-1 mRNA and ICAM-1mRNA,and PPARγmRNA by RT-PCR.Results:1.The Rosiglitazion increased EPC migration captivity and LPS have no influence to EPC migration captivity2.The Rosiglitazion increased the concentration and activity of tPA upgrade the tPA mRNA expression.The Rosiglitazion decresed the concentration and activity of PAI,and decresed PAImRNA expression.And the Rosiglitazion inhabited inflammatory factor mRNA expression3.LPS inhabited the concentration and activity and mRNA expression of tPA,and increased the concentration and activity and mRNA expression of PAI.LPS precipitate inflammatory factor mRNA expression4.Rosiglitazion can inhabit LPS effect to EPC.Conclusion The LPS can inhabit the fuction of tPA and increase the fuction of PAI,and precipitate inflammatory factor expression. Rosiglitazion can inhabit the effect of LPS to EPC,and increased the migretion capcity of EPC.