南方医科大学学报
南方醫科大學學報
남방의과대학학보
Journal of Southern Medical University
2015年
8期
1128-1132,1142
,共6页
张建成%林琨%魏芝雄%陈茜%刘丽%赵晓静%赵颖%徐斌%陈曦
張建成%林琨%魏芝雄%陳茜%劉麗%趙曉靜%趙穎%徐斌%陳晞
장건성%림곤%위지웅%진천%류려%조효정%조영%서빈%진희
大蒜素%HEK293细胞%HERG电流%膜片钳技术
大蒜素%HEK293細胞%HERG電流%膜片鉗技術
대산소%HEK293세포%HERG전류%막편겸기술
allitridum%HEK293 cell line%rapidly delayed rectifier potassium current%patch clamp technique
目的 研究大蒜素对HEK293细胞HERG电流的作用,探讨其抗心律失常的可能机制.方法 采用瞬时转染的方法,将HERG通道质粒转入HEK293细胞上,应用细胞外局部灌流法于膜片钳高阻抗封接形成后给予大蒜素,使其终浓度为30μmol/L.室温下,采用全细胞膜片钳技术在电压钳形模式下记录电流和门控动力学,观察大蒜素对HERG电流的作用.结果 30μmol/L大蒜素对正常大鼠心室肌细胞HERG电流有显著的阻滞效应,使其尾电流密度由73.5±4.3 pA/pF降低至42.1±3.6 pA/pF(P<0.01,n=14).其作用呈浓度依赖性.半数抑制浓度为34.74μmol/L,Hill系数为1.01.大蒜素可使HERG的电流-电压曲线降低,且随着去极化电位的增加,作用更加明显,提示其作用具有电压依赖性,门控机制研究发现大蒜素可以使通道激活曲线向更正的方向移动,进而延迟激活;使通道稳态失活更负的方向移动,导致失活加速.同时,使通道灭活的慢时间常数缩短,从而加速通道的灭活.结论 大蒜素抑制HEK293细胞上HERG电流,提示这可能是其治疗心律失常的细胞电生理基础.
目的 研究大蒜素對HEK293細胞HERG電流的作用,探討其抗心律失常的可能機製.方法 採用瞬時轉染的方法,將HERG通道質粒轉入HEK293細胞上,應用細胞外跼部灌流法于膜片鉗高阻抗封接形成後給予大蒜素,使其終濃度為30μmol/L.室溫下,採用全細胞膜片鉗技術在電壓鉗形模式下記錄電流和門控動力學,觀察大蒜素對HERG電流的作用.結果 30μmol/L大蒜素對正常大鼠心室肌細胞HERG電流有顯著的阻滯效應,使其尾電流密度由73.5±4.3 pA/pF降低至42.1±3.6 pA/pF(P<0.01,n=14).其作用呈濃度依賴性.半數抑製濃度為34.74μmol/L,Hill繫數為1.01.大蒜素可使HERG的電流-電壓麯線降低,且隨著去極化電位的增加,作用更加明顯,提示其作用具有電壓依賴性,門控機製研究髮現大蒜素可以使通道激活麯線嚮更正的方嚮移動,進而延遲激活;使通道穩態失活更負的方嚮移動,導緻失活加速.同時,使通道滅活的慢時間常數縮短,從而加速通道的滅活.結論 大蒜素抑製HEK293細胞上HERG電流,提示這可能是其治療心律失常的細胞電生理基礎.
목적 연구대산소대HEK293세포HERG전류적작용,탐토기항심률실상적가능궤제.방법 채용순시전염적방법,장HERG통도질립전입HEK293세포상,응용세포외국부관류법우막편겸고조항봉접형성후급여대산소,사기종농도위30μmol/L.실온하,채용전세포막편겸기술재전압겸형모식하기록전류화문공동역학,관찰대산소대HERG전류적작용.결과 30μmol/L대산소대정상대서심실기세포HERG전류유현저적조체효응,사기미전류밀도유73.5±4.3 pA/pF강저지42.1±3.6 pA/pF(P<0.01,n=14).기작용정농도의뢰성.반수억제농도위34.74μmol/L,Hill계수위1.01.대산소가사HERG적전류-전압곡선강저,차수착거겁화전위적증가,작용경가명현,제시기작용구유전압의뢰성,문공궤제연구발현대산소가이사통도격활곡선향경정적방향이동,진이연지격활;사통도은태실활경부적방향이동,도치실활가속.동시,사통도멸활적만시간상수축단,종이가속통도적멸활.결론 대산소억제HEK293세포상HERG전류,제시저가능시기치료심률실상적세포전생리기출.
Objective To study the effect of allitridum on rapidly delayed rectifier potassium current (IKr) in HEK293 cell line. Methods HEK293 cells were transiently transfected with HERG channel cDNA plasmid pcDNA3.1 via Lipofectamine. Allitridum was added to the extracellular solution by partial perfusion after giga seal at the final concentration of 30μmol/L. Whole-cell patch clamp technique was used to record the HERG currents and gating kinetics before and after allitridum exposure at room temperature. Results The amplitude and density of IHERG were both suppressed by allitridum in a voltage-dependent manner. In the presence of allitridum, the peak current of IHERG was reduced from 73.5±4.3 pA/pF to 42.1±3.6 pA/pF at the test potential of +50 mV (P<0.01). Allitridum also concentration-dependently decreased the density of the IHERG. The IC50 of allitridum was 34.74 μmol/L with a Hill coefficient of 1.01. Allitridum at 30 μmol/L caused a significant positive shift of the steady-state activation curve of IHERG and a markedly negative shift of the steady-state inactivation of IHERG, and significantly shortened the slow time constants of IHERG deactivation. Conclusion Allitridum can potently block IHERG in HEK293 cells, which might be the electrophysiological basis for its anti-arrhythmic action.