【摘要】：目的:探討不同濃度二十二碳六烯酸(DHA)對正常大鼠冠狀動脈平滑肌細(xì)胞大電導(dǎo)鈣激活鉀離子通道(BK通道)的激活作用及其機制。方法:酶消化法分離正常大鼠冠狀動脈平滑肌細(xì)胞。采用全細(xì)胞膜片鉗實驗技術(shù)記錄在未孵育與孵育細(xì)胞色素P450環(huán)氧化酶抑制劑SKF525A并灌流不同濃度DHA條件下BK通道電流密度變化;采用單通道膜片鉗實驗技術(shù)記錄灌流不同濃度DHA時BK單通道開放概率的變化。結(jié)果:0.01~1.00μmol/L(定義為低濃度)DHA激活BK通道的半效濃度(EC50)為(0.24±0.05)μmol/L,但經(jīng)SKF525A孵育后激活作用消失;3.00~10.00μmol/L(定義為高濃度)DHA激活BK通道的EC50為(2.38±0.22)μmol/L,經(jīng)SKF525A孵育后,激活作用仍存在,且EC50無明顯變化。在電極外液鈣離子濃度為1μmol/L和刺激電位60 m V條件下,DHA濃度為0、0.01、0.03、0.10、0.30、1.00μmol/L時,BK通道開放概率分別為(0.095 2±0.009 5)、(0.093 9±0.012 6)、(0.098 6±0.016 9)、(0.099 5±0.014 7)、(0.097 5±0.010 4)和(0.102 3±0.020 6)(P0.05,n=5),提示低濃度DHA不能激活BK單通道;繼續(xù)增加DHA濃度,當(dāng)濃度為3、10μmol/L時,BK單通道開放概率分別為(0.700 3±0.013 2)和(0.892 7±0.052 3)(P0.05,n=5),提示高濃度DHA呈濃度依賴性激活BK單通道,EC50為(3.37±0.10)μmol/L。結(jié)論:不同濃度DHA激活BK通道的機制不同,低濃度DHA通過細(xì)胞色素P450環(huán)氧化酶途徑激活BK通道,高濃度DHA可能通過與BK通道結(jié)合后直接激活。
[Abstract]:Aim: to investigate the effect and mechanism of 22 carbohexaenoic acid (DHA) on calcium activated potassium channel (BK channel) in normal rat coronary artery smooth muscle cells. Methods: normal rat coronary artery smooth muscle cells were isolated by enzyme digestion. Whole cell patch clamp technique was used to record the changes of current density of BK channel in unincubated and incubated cytochrome P450 cyclooxygenase inhibitor (SKF525A) and perfused with different concentrations of DHA. The single channel patch clamp technique was used to record the change of BK single channel open probability when perfused with different concentrations of DHA. Results: the half effect concentration (EC50) of DHA activated BK channel was (0. 24 鹵0. 05) 渭 mol/L, (defined as low concentration) by 0. 01 渭 mol/L (defined as low concentration), but the activation effect disappeared after incubation with SKF525A (0. 24 鹵0. 05) 渭 mol/L,. The EC50 of 3.00 渭 mol/L (defined as high concentration) DHA activated BK channel was (2.38 鹵0.22) 渭 mol/L,. After incubation with SKF525A, the activation still existed, and EC50 did not change significantly. The opening probability of BK channel was (0.095 2 鹵0.009 5), (0.093 9 鹵0.012 6) when the concentration of calcium ion was 1 渭 mol/L and the stimulation potential was 60 MV, and the concentration of DHA was 0 0. 01 渭 mol/L, 0. 10 and 0. 30 渭 mol/L, respectively, when the concentration of Ca2 + was 1 渭 mol/L and the stimulation potential was 60 MV, the opening probability of BK channel was (0.095 2 鹵0.009 5), (0.093 9 鹵0.012 6). (0.098 6 鹵0.016 9), (, 0.099 5 鹵0.014 7), (, 0.097 5 鹵0.010 4) and (0.102 3 鹵0.020 6) (P 0.05 鹵0.020 6), indicating that low concentration DHA could not activate BK single channel. The open probability of single channel of BK was (0.700 3 鹵0.013 2) and (0.892 7 鹵0.052 3) (P 0.05), respectively, when the concentration of BK was 3 渭 mol/L and 10 渭 mol/L, suggesting that high concentration of DHA activated BK single channel in a concentration-dependent manner. EC50 = (3.37 鹵0.10) 渭 mol/L. Conclusion: the mechanism of activation of BK channel by different concentration of DHA is different. Low concentration of DHA activates BK channel through cytochrome P450 cyclooxygenase pathway, and high concentration of DHA may be activated directly by binding to BK channel.
【作者單位】： 南京醫(yī)科大學(xué)附屬無錫市人民醫(yī)院心內(nèi)科;山東省醫(yī)學(xué)科學(xué)院基礎(chǔ)醫(yī)學(xué)研究所心血管病研究室;Mayo
【基金】：國家自然科學(xué)基金(81370303,81500249,81500323) 江蘇省自然科學(xué)基金(BK20151110) 江蘇省人事廳“六大人才高峰”第七批高層次項目(006) 江蘇省醫(yī)學(xué)重點人才資助項目(RC201134)
【分類號】：R54

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