The Research Of Vascular Umbilical Vein Endothelial Cells Function And Its Regulation Mechanism In Preeclampsia

Part I The research of umbilical vein endothelial cell function in patients of preeclampsiaObjective:Damage of endothelial cells in the pathogenesis of preeclampsia is an apparent factor.Through detecting endothelial function of umbilical vein endothelial cells(HUVEC),our experiment verified that the damage of endothelial function in patients of preeclampsia is an important mechanism.Methods:Patients of preeclampsia(PE)and normal pregnant ones(NP)are chosen as two teams respectively.Collecting HUVEC and serum of both groups,methods of PCR and Western blot were used to detect gene and protein expression of eNOS in both groups,and nitrate reduction method was used to detect the NO content of endothelial cell medium.Results:Compared with NP,the NO content of HUVEC of PE group was obviously lower(P < 0.05).Gene and protein expression of HUVEC eNOS in the PE group was decreased obviously(P < 0.05).Conclusion:Through NO content and expression of eNOS of HUVEC in the two groups,endothelial function of PE is significantly weaker than NP.Part II The change of potassium channel current impact on umbilical vein endothelial cell function in patients of preeclampsiaObjective: Based on damage of endothelial cells in the pathogenesis of preeclampsia,we assumed that potassium channels currents are connected with umbilical vein endothelial cells of preeclampsia.All above verified that the change of preeclampsia is caused by K+ channel and endothelial cell damage.Methods: We measured potassium channel current of the two groups(PE and NP ones)respectively with patch clamp technique.After adding IBTX(BKa channel inhibitor)and apamin(SKa channel inhibitor)to endothelial cell medium,potassium channel current of the two groups was measured again with the same technique.Results: Compared with NP,potassium channel current of HUVEC of PE group was obviously lower,(NP vs.PE: 12.28±1.25 p A/p F vs.5.45±0.51 p A/p F;TP=+60 m V,P<0.05).After adding IBTX and apamin to endothelial cell medium.Potassium channel current of HUVEC of NP team decreased obviously(from 12.08±1.16 p A/p F at baseline to 7.24±0.80 p A/p F,TP=+60 m V,P<0.05,Figure 2C),while that change of HUVEC of PE team is little.Conclusion: Through two groups data of potassium channel current of HUVEC,The function of ion channel of NP is normal,but that of PE is be destroyed.It showed that damage of potassium channel current impact on umbilical vein endothelial cell function in patients of preeclampsia.Part III Oxidative stress related to potassium channel current of endothelial cell and NO synthesis in preeclampsiaObjective: Oxidative stress of preeclampsia has been unbalance.Our experiment tested the content of superoxide and protein expression t of the umbilical vein cells,which verified that the oxidative stress of preeclampsia damaged potassium channels currents of endothelial cells and endothelial function.Methods: We compared two groups of HUVEC fluorescence color with DHE technique respectively.The content of the two groups of HUVEC superoxide was detected by ELISA;NOX1,NOX2,NOX4 protein and gene expression of the two groups of HUVEC is inspected with Q-PCR and Western technique.After adding Apocynin(inhibitors of NADPH oxidase)to HUVEC culture of two groups,all above are inspected again.KCNMB1,SK3,as well as the expression of e NOS were compared.After collection of the dosing of HUVEC culture,the NO content was detected.Results: Compared with NP,the HUVEC fluorescence color OF PE group was obvious.The content of superoxide and protein expression of HUVEC of PE group were higher than those of NP ones.Potassium channels currents is higher after adding Apocynin to HUVEC culture of PE.Besides,KCNMB1,SK3 and e NOS protein and gene expression of PE raised markedly.Conclusion: NOX-mediated ROS(reactive oxygen species)inhibited BKCa ? SKCa and damaged endothelial NO system.