KCa3.1通道在内皮祖细胞生物学功能改变及分化中的作用
|
摘要
目的探讨KCa3.1通道对内皮祖细胞(endothelial progenitor cell,EPCs)生物学功能及分化的影响。方法应用qRTPCR检测EPCs上KCa3.1、vWF、CD31基因表达;采用CCK-8法、细胞贴壁法及Matrigel基质胶分别检测细胞增殖、黏附及体外血管形成能力的变化;应用细胞免疫荧光、Western blot及流式细胞术分别检测KCa3.1通道蛋白、内皮分化标志v WF和CD31、整合素β_1、β_3的表达。结果阻断KCa3.1功能或干扰KCa3.1的表达能够降低EPCs的增殖、黏附及血管形成能力,但是却可以促进EPCs向内皮分化;激活或过表达KCa3.1通道能够增强EPCs的增殖、黏附及血管形成能力,但是阻碍EPCs向内皮分化。阻断KCa3.1离子通道后,细胞膜整合素β_1表达下调,而激活KCa3.1离子通道整合素β_1表达上调。结论 KCa3.1通道功能变化及表达变化能够改变EPCs的生物学特性及向内皮分化水平。
OBJECTIVE To investigate the effect of KCa3. 1 channel on the function of EPCs. METHODS The gene expression of KCa3. 1,v WF and CD31 on EPCs were detected by qRT-PCR. CCK-8 kit,cell adherent method and Matrigel were used to detect the changes of cell proliferation,adhesion and in vitro angiogenesis; cell immunofluorescence or fluorescence activated cell sorter( FACS) was used to detect the protein expression of KCa3. 1,v WF,CD31,integrin β1,integrin β3 separately. RESULTS Blocking the function of KCa3. 1 or interfering with the expression of KCa3. 1 can attenuated EPC function of proliferation,adhesion and angiogenesis,but it can promote the differentiation of EPCs. Overexpression or activation of KCa3. 1 channel can enhance EPCs proliferation,adhesion and angiogenesis but decrease the level of differentiation. The expression of integrin β1 on EPCs was attenuated with blocking KCa3. 1 channel,but the expression effect was reversible by the activator. CONCLUSION The alteration of KCa3. 1 channel function or expression affect the biological characteristics and differentiation of EPCs.
OBJECTIVE To investigate the effect of KCa3. 1 channel on the function of EPCs. METHODS The gene expression of KCa3. 1,v WF and CD31 on EPCs were detected by qRT-PCR. CCK-8 kit,cell adherent method and Matrigel were used to detect the changes of cell proliferation,adhesion and in vitro angiogenesis; cell immunofluorescence or fluorescence activated cell sorter( FACS) was used to detect the protein expression of KCa3. 1,v WF,CD31,integrin β1,integrin β3 separately. RESULTS Blocking the function of KCa3. 1 or interfering with the expression of KCa3. 1 can attenuated EPC function of proliferation,adhesion and angiogenesis,but it can promote the differentiation of EPCs. Overexpression or activation of KCa3. 1 channel can enhance EPCs proliferation,adhesion and angiogenesis but decrease the level of differentiation. The expression of integrin β1 on EPCs was attenuated with blocking KCa3. 1 channel,but the expression effect was reversible by the activator. CONCLUSION The alteration of KCa3. 1 channel function or expression affect the biological characteristics and differentiation of EPCs.
引文
[1]ASAHARA T,MUROHARA T,SULLIVAN A,et al.Isolation of putative progenitor endothelial cells for angiogenesis[J].Science,1997,275(5302):964-967.
[2]TAYLOR D A,RESENDE M,SAMPAIO L C.Mobilizing EPCs:it is not just an acute issue[J].Int J Cardiol,2018,257:272-273.
[3]ZHANG L C,FAN W,BAO H F,et al.Neuroprotective role of statins in acute phase of ischemic stroke[J].Chin Pharm J(中国药学杂志),2014,49(22):2018-2022.
[4]GARDOS G.The function of calcium in the potassium permeability of human erythrocytes[J].Biochim Biophys Acta,1958,30(3):653-654.
[5]CUI X,ZHANG X,GUAN X,et al.Shear stress augments the endothelial cell differentiation marker expression in late epcs by upregulating integrins[J].Biochem Biophys Res Commun,2012,425(2):419-425.
[6]TAO R,LAU C P,TSE H F,et al.Regulation of cell proliferation by intermediate-conductance Ca2+-activated potassium and volume-sensitive chloride channels in mouse mesenchymal stem cells[J].Am J Physiol Cell Physiol,2008,295(5):1409-1416.
[7]COLEMAN N,BROWN B M,OLIVAN-VIGUERA A,et al.New positive Ca2+-activated K+channel gating modulators with selectivity for KCa3.1[J].Mol Pharmacol,2014,86(3):342-357.
[8]ADELMAN J P,MAYLIE J,SAH P.Small-conductance Ca2+-activated K+channels:form and function[J].Annu Rev Physiol,2012,74(1):245-269.
[9]LI H,ZHAO J L,ZHANG Y M,et al.Inhibitoryeffects of candesartan on KCa3.1 potassium channel expression and cell culture and proliferation in peripheral blood CD4(+)T lymphocytes in kazakh patients with hypertension from the Xinjiang region[J].Clin Exp Hypertens,2018,40(4):303-311.
[10]ANUMANTHAN G,GUPTA S,FINK M K,et al.KCa3.1 ion channel:a novel therapeutic target for corneal fibrosis[J].PLo S One,2018,13(3):e192145.
[11]BUSSE R,EDWARDS G,FELETOU M,et al.EDHF:bringing the concepts together[J].Trends Pharmacol Sci,2002,23(8):374-380.
[12]DENG X L.KCa3.1:a potential therapeutic target for cardiovascular disease[J].J Xi'an Jiaotong Univ(西安交通大学学报),2013,34(2):139-143.
[13]LIU C,ZHAO F,LI Q Z.Biological function of integrin in cell adhesion[J].J Biol(生物学杂志),2012,29(1):75-78.
[14]MA X L,LIU H Q.Effect of calcium on the proliferation and differentiation of murine corneal epithelial cells in vitro[J].Int J Ophthalmol,2011,4(3):247-249.
[15]YU Y.The biological role of Ca2+-activated K+channel(KCa3.1)on hemodynamic induced cerebral aneurysm initiation mechanism[D].Shanghai:Second Milary Medical University,2015.
[16]YANG H,LI X,LIU Y,et al.Crocinimproves the endothelial function regulated by KCa3.1 through ERK and Akt signaling pathways[J].Cell Physiol Biochem,2 0 1 8,4 6(2):7 6 5-7 8 0.
[17]BONITO B,SAUTER D R,SCHWAB A,et al.KCa3.1(IK)modulates pancreatic cancer cell migration,invasion and proliferation:anomalous effects on TRAM-34[J].Pflugers Arch,2016,468(11-12):1865-1875.
[18]SONG P,DU Y,SONG W,et al.KCa3.1 as an effective target for inhibition of growth and progression of intrahepatic cholangiocarcinoma[J].J Cancer,2017,8(9):1568-1578.
[2]TAYLOR D A,RESENDE M,SAMPAIO L C.Mobilizing EPCs:it is not just an acute issue[J].Int J Cardiol,2018,257:272-273.
[3]ZHANG L C,FAN W,BAO H F,et al.Neuroprotective role of statins in acute phase of ischemic stroke[J].Chin Pharm J(中国药学杂志),2014,49(22):2018-2022.
[4]GARDOS G.The function of calcium in the potassium permeability of human erythrocytes[J].Biochim Biophys Acta,1958,30(3):653-654.
[5]CUI X,ZHANG X,GUAN X,et al.Shear stress augments the endothelial cell differentiation marker expression in late epcs by upregulating integrins[J].Biochem Biophys Res Commun,2012,425(2):419-425.
[6]TAO R,LAU C P,TSE H F,et al.Regulation of cell proliferation by intermediate-conductance Ca2+-activated potassium and volume-sensitive chloride channels in mouse mesenchymal stem cells[J].Am J Physiol Cell Physiol,2008,295(5):1409-1416.
[7]COLEMAN N,BROWN B M,OLIVAN-VIGUERA A,et al.New positive Ca2+-activated K+channel gating modulators with selectivity for KCa3.1[J].Mol Pharmacol,2014,86(3):342-357.
[8]ADELMAN J P,MAYLIE J,SAH P.Small-conductance Ca2+-activated K+channels:form and function[J].Annu Rev Physiol,2012,74(1):245-269.
[9]LI H,ZHAO J L,ZHANG Y M,et al.Inhibitoryeffects of candesartan on KCa3.1 potassium channel expression and cell culture and proliferation in peripheral blood CD4(+)T lymphocytes in kazakh patients with hypertension from the Xinjiang region[J].Clin Exp Hypertens,2018,40(4):303-311.
[10]ANUMANTHAN G,GUPTA S,FINK M K,et al.KCa3.1 ion channel:a novel therapeutic target for corneal fibrosis[J].PLo S One,2018,13(3):e192145.
[11]BUSSE R,EDWARDS G,FELETOU M,et al.EDHF:bringing the concepts together[J].Trends Pharmacol Sci,2002,23(8):374-380.
[12]DENG X L.KCa3.1:a potential therapeutic target for cardiovascular disease[J].J Xi'an Jiaotong Univ(西安交通大学学报),2013,34(2):139-143.
[13]LIU C,ZHAO F,LI Q Z.Biological function of integrin in cell adhesion[J].J Biol(生物学杂志),2012,29(1):75-78.
[14]MA X L,LIU H Q.Effect of calcium on the proliferation and differentiation of murine corneal epithelial cells in vitro[J].Int J Ophthalmol,2011,4(3):247-249.
[15]YU Y.The biological role of Ca2+-activated K+channel(KCa3.1)on hemodynamic induced cerebral aneurysm initiation mechanism[D].Shanghai:Second Milary Medical University,2015.
[16]YANG H,LI X,LIU Y,et al.Crocinimproves the endothelial function regulated by KCa3.1 through ERK and Akt signaling pathways[J].Cell Physiol Biochem,2 0 1 8,4 6(2):7 6 5-7 8 0.
[17]BONITO B,SAUTER D R,SCHWAB A,et al.KCa3.1(IK)modulates pancreatic cancer cell migration,invasion and proliferation:anomalous effects on TRAM-34[J].Pflugers Arch,2016,468(11-12):1865-1875.
[18]SONG P,DU Y,SONG W,et al.KCa3.1 as an effective target for inhibition of growth and progression of intrahepatic cholangiocarcinoma[J].J Cancer,2017,8(9):1568-1578.