【摘要】 目的　研究 17β -雌二醇 (E2 )对新生小牛主动脉血管内皮细胞 (BAECs)中内皮型一氧化氮合酶 (eNOS)活性的长期基因效应 ,并进一步探讨胞内钙信号途径在其中的作用。方法　采用培养的BAECs,同位素法测定eNOS的活性 ;Westernblot检测eNOS蛋白表达 ;荧光分光光度计法检测E2 对细胞内钙的影响。结果　 (1)E2 (0 .0 0 1,0 0 1,0 .1,1μmol/L)作用于BAECs 4 8h ,使eNOS活性分别增加 130 %± 2 7% ,178%± 19% ,14 2 %± 2 0 % ,14 4 %± 2 7%。E2 激活eNOS活性的作用被雌激素受体拮抗剂Tamoxifen(0 .1μmol/L)所阻断 ;(2 ) 0 .0 1μmol/LE2 作用于BAECs 1h对eNOS蛋白表达无明显影响 ,而作用 2 4h和 4 8h分别使eNOS蛋白表达增加了 2 76 %± 36 %和374 %± 2 0 % ;Tamoxifen明显抑制E2 处理 4 8h后诱导的eNOS蛋白表达 ,抑制率为 6 6 5 %± 7 4 % ;(3) 0 .0 1μmol/LE2 作用BAECs 4 8h ,分别使静息 [Ca2 + ]i增加了 70 .3± 2 1.5nmol/L ,使 10mmol/LATP诱导的BAECs[Ca2 + ]i峰值增加了 2 4 0 .2± 6 8 2nmol/L ,ATP诱导的 [Ca2 + ]i增加值 (峰值与静息值之差 )增加了 170 .5± 4 8.9nmol/L。结论　雌激素可通过长期基因效应而激活eNOS ,至少部分通过核ER介导 ,并与胞内钙动员有关更多还原

【Abstract】 Objective To study the activation of endothelial nitric oxide synthase(eNOS) induced by 17β-estradiol(E 2) through genomic pathway and the influence of intracellular free calcium concentration([Ca 2+ ]i) on these effects. Methods Confluent bovine aortic endothelial cells were cultured. eNOS activity was assessed in whole cells by measuring [ 3H]L-arginine conversion to [ 3H]L-citrulline by NOS Assay Kit. The expression of eNOS protein was measured by Western blot. Calcium concentration was measured by fluorometry. Results (1) E 2(0.001,0.01,0.1,1 μmol/L) resulted in increase of eNOS activity in BAECs(130%±27%;178%±19%;142%±20%;144%±27%) within 48 h. The effect of E 2 on eNOS activation was obviously inhibited by Tamoxifen(0.1 μmol/L); (2) The abundant expression of eNOS protein increased obviously after treatment with 0.01 μmol/L E 2 for 24 h and 48 h(276%±36%;374%±20%), the effects of E 2 on eNOS protein was inhibited by Tamoxifen(66.5%±7 4%);(3) Treatment with E 2 for 48 h significantly increased the resting [Ca 2+ ]:(70.3±21.5nmol/L) and the rise of[Ca 2+ ]i(240 2±68.2 nmol/L) induced by ATP in BAECs. Conclusion E 2 activate eNOS activity through genomic pathway,which may be mediated by nuclear estrogen receptor and related to calcium mobilization.更多还原