微电场通过非整合素途径活化粘着斑激酶促进滋养细胞迁移/侵袭研究
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摘要
目的探讨生理性微电场促进体外培养的人胎盘滋养细胞迁移/侵袭功能是否与滋养细胞表面整合素(integrin)表达有关。方法用150 m V/mm的直流微电场刺激滋养细胞,时间分别为5、10和15 h,测定其迁移情况并观察细胞形态变化。Western blot检测刺激前后1 h内粘着斑激酶FAK活化情况和细胞表面integrinα1、integrinα5、integrinαV和integrinα1蛋白表达水平。结果在含有10%胎牛血清的培养基中,150 m V/mm电场刺激下滋养细胞向负极定向迁移,迁移速度和距离较对照组明显增加(P=0.021),胞体拉长,垂直于电场方向排列;胞内FAKTyr397位点于刺激后5、10、30、60 min内迅速活化并逐渐加强(P<0.05);刺激前后滋养细胞表面integrinα1、integrinα5、integrinαV、和integrinα1蛋白表达水平无明显改变(P>0.05)。结论生理性直流微电场可能通过非整合素途径活化FAK从而促进滋养细胞迁移/侵袭功能,但其详细机制仍需进一步研究。
Objective To investigate whether the effect of physiological micro-electrical field on migration/invasion of trophoblast cells in vitro was associated with integrins expression of the cells. Methods The trophoblast cells were exposed to the direct current electrical field(DC-EF) at 150 m V/mm for 5,10 and 15 hours. Cell images and migration distance were recorded with continuous photographing and analyzed by image analyzer. The activation of FAK at 5,10,30 and 60 min of DC-EF stimulation were measured and the expression levels of integrinα1,integrinα5,integrinαV and integrinα1 were detected by using Western blot assay. Results The application of 150 m V/mm DC electrical stimulation,trophoblast cells cultured in media containing 10% calf serum showed a cathode migration. The migration velocity and distance were obviously increased when compared to the control group without EF stimulation(P =0. 021). The cells exposed to the EF also showed elongation and perpendicular orientation,while control cells that were not subjected to EF showed no such responsiveness. Compared with the non-EF stimulation controls,trophoblasts under EF stimulation had a quickly activation of FAKTyr397 location within 60 min(P < 0. 05),while no significant changes in expressions of integrinα1,integrinα5,integrinαV and integrinα1 were observed(P > 0. 05). Conclusion The physiological direct current electrical field may activate FAK by non-integrin pathway to promote trophoblast cell migration/invasion function. However,its detailed mechanism needs further studies.
Objective To investigate whether the effect of physiological micro-electrical field on migration/invasion of trophoblast cells in vitro was associated with integrins expression of the cells. Methods The trophoblast cells were exposed to the direct current electrical field(DC-EF) at 150 m V/mm for 5,10 and 15 hours. Cell images and migration distance were recorded with continuous photographing and analyzed by image analyzer. The activation of FAK at 5,10,30 and 60 min of DC-EF stimulation were measured and the expression levels of integrinα1,integrinα5,integrinαV and integrinα1 were detected by using Western blot assay. Results The application of 150 m V/mm DC electrical stimulation,trophoblast cells cultured in media containing 10% calf serum showed a cathode migration. The migration velocity and distance were obviously increased when compared to the control group without EF stimulation(P =0. 021). The cells exposed to the EF also showed elongation and perpendicular orientation,while control cells that were not subjected to EF showed no such responsiveness. Compared with the non-EF stimulation controls,trophoblasts under EF stimulation had a quickly activation of FAKTyr397 location within 60 min(P < 0. 05),while no significant changes in expressions of integrinα1,integrinα5,integrinαV and integrinα1 were observed(P > 0. 05). Conclusion The physiological direct current electrical field may activate FAK by non-integrin pathway to promote trophoblast cell migration/invasion function. However,its detailed mechanism needs further studies.
引文
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[13]Forrester JV,Lois N,Zhao M,et al.The Spark of Life:the role of electric fields in regulating cell behaviour using the eye as a model system[J].Ophthalmic Res,2007,39(1):4-16.
[14]Tang CL,Zhao HB,Li MQ,et al.Focal adhesion kinase signaling is necessary for the Cyclosporin A-enhanced migration and invasion of human trophoblast cells[J].Placenta,2012,33(9):704-711.
[15]张曦倩,陈士岭,邢福祺.黏着斑激酶对ERK介导的滋养层细胞侵袭行为的调节作用[J].中南大学学报(医学版),2011,36(6):559-564.
[2]Fei L,Tunan C,Shengli H,et al.Superoxide Mediates Direct Current Electric Field Induced Directional Migration of Glioma Cells through the Activation of AKT and ERK[J].PLOS ONE,2013,8(4):e61195.
[3]Bai H,Forrester JV,Zhao M.DC electric stimulation up regulates angiogenic factors in endothelial cells through activation of VEGF receptors[J].Cytokine,2011,55(1):110-115.
[4]Zhao M,Chalmers L,Cao L,et al.Electrical signaling in control of ocular cell behaviors[J].Prog Retin Eye Res,2012,31(1):65-88.
[5]张娟,李明勇,贺元,等.微电场对滋养细胞迁移/侵袭相关MMPs/TIMPs表达的影响[J].重庆医学杂志,2016,45(7):869-872.
[6]张娟,李明勇,贺元,等.微电场通过活化粘着斑激酶信号通路促进缺氧培养滋养细胞的迁移/侵袭[J].临床检验杂志,2015,33(11):813-817.
[7]Zhang J,Ren RM,Luo XF,et al.A Small Physiological Electric Field Mediated Responses of Extravillous Trophoblasts Derived from HTR8/SVneo Cells:Involvement of Activation of Focal Adhesion Kinase Signaling[J].PLOS ONE,2014,3(9):e92252.
[8]Furmento VA,Marino J,Blank VC,et al.Granulocyte colony-stimulating factor(G-CSF)upregulatesβ1 integrin and increases migration of human trophoblast Swan 71 cells via PI3K and MAPK activation[J].Exp Cell Res,2016,342(2):125-134.
[9]Lim SM,Jang HY,Lee JE,et al.Alteration of Pituitary Tumor Transforming Gene-1 Regulates Trophoblast Invasion via the Integrin/RhoFamily Signaling Pathway[J].PLo S One,2016,11(2):e0149371.
[10]Liao WC,Liu CH,Chen CH,et al.α-1,4-Galactosyltransferase III suppresses extravillous trophoblast invasion through modifyingβ1-integrin glycosylation[J].Placenta,2015,36(4):357,364.
[11]刘万钱,管章委,邓小燕,等.整合素β1对人体滋养层细胞黏附和迁移行为的影响[J].生物医学工程学杂志,2010,27(1):67-70.
[12]Zhao M,Pu J,Forrester JV,et al.Membrane lipids,EGF receptors,and intracellular signals colocalize and are polarized in epithelial cells moving directionally in a physiological electric field[J].Faseb J,2002,16(8):857-859.
[13]Forrester JV,Lois N,Zhao M,et al.The Spark of Life:the role of electric fields in regulating cell behaviour using the eye as a model system[J].Ophthalmic Res,2007,39(1):4-16.
[14]Tang CL,Zhao HB,Li MQ,et al.Focal adhesion kinase signaling is necessary for the Cyclosporin A-enhanced migration and invasion of human trophoblast cells[J].Placenta,2012,33(9):704-711.
[15]张曦倩,陈士岭,邢福祺.黏着斑激酶对ERK介导的滋养层细胞侵袭行为的调节作用[J].中南大学学报(医学版),2011,36(6):559-564.