EFFECT OF EXTRACELLULAR ATP LEVEL ON FLOW-INDUCED CA++ RESPONSE IN CULTURED VASCULAR ENDOTHELIAL-CELLS
EFFECT OF EXTRACELLULAR ATP LEVEL ON FLOW-INDUCED CA++ RESPONSE IN CULTURED VASCULAR ENDOTHELIAL-CELLS
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DOI：
10.1016/0006-291x(91)91698-c
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发表时间：
1991-09-30
影响因子：
3.1
通讯作者：
KAMIYA, A
中科院分区：
文献类型：
--
作者：
ANDO, J;OHTSUKA, A;KAMIYA, A
培养的血管内皮细胞负载高荧光钙敏感染料Fura-2，暴露于含有不同浓度三磷酸腺苷(0，0.5，1，5μM)的流体中，在基于流体动力学的装置中，用荧光显微镜监测细胞内游离钙离子浓度的同步变化。灌流液流量从0~6.3~22.8~39.0 cm/s，细胞表面产生的剪应力分别为0、2.9、10.4和17.9dynes/cm2。当ATP浓度低于100 nM时，细胞内Ca~(2+)水平无明显变化，而在ATP浓度为500 nM时，细胞内Ca~(++)浓度随着灌流液流量的增加而升高。在此ATP水平下，0、6.3、22.8和39.0 cm/s流速下的细胞内钙离子水平分别为44.8±7.3、60.3±10.7、74.0±5.8和89.4±6.4 nM(平均值±SD；n=8)。当ATP水平超过1μM时，Ca++反应的流量依赖性变得不如在ATP500 nM时观察到的那样明显。当在灌流液中加入2 mM的EGTA来螯合细胞外钙离子时，这些钙离子对流量变化的反应消失。这些结果提示，在适当的ATP浓度下，血管内皮细胞具有与流量相一致的升高细胞内钙离子水平的机制，而这种机制下细胞内钙离子水平的变化似乎主要是由细胞外钙离子内流引起的。
Cultured vascular endothelial cells loaded with the highly fluorescent Ca++-sensitive dye Fura-2 were exposed to the flow of a fluid containing various concentrations of ATP (0, 0.5, 1,5 μM) in an apparatus designed on the basis of fluid dynamics, and simultaneous changes in intracellular free Ca++concentration were monitored by photometric fluorescence microscopy. The flow rate of the perfusate was altered from 0 to 6.3 to 22.8 to 39.0 cm/sec, inducing shear stress on the cell surface of 0, 2.9, 10.4, and 17.9 dynes/cm2, respectively. Although no significant change in intracellular Ca++level was observed at ATP levels below 100 nM, at an ATP level of 500 nM, the intracellular Ca++level increased together with an increase in the flow rate of the perfusate. At this level of ATP, the intracellular Ca++levels at flow rates of 0, 6.3, 22.8, and 39.0 cm/sec were 44.8 ± 7.3, 60.3 ± 10.7, 74.0 ± 5.8 and 89.4 ± 6.4 nM (mean ± SD; n=8), respectively. At ATP levels over 1 μM, the flow-rate dependency of Ca++response became less clear than that observed at the ATP level of 500 nM. These Ca++responses to changes in flow rate disappeared when extracellular Ca++was chelated by adding 2 mM of EGTA to the perfusate. These results suggest that the vascular endothelial cell has a mechanism that elevates the intracellular Ca++level in accord with the flow rate at appropriate ATP concentrations, and that changes in intracellular Ca++level under this mechanism seem to be chiefly caused by the influx of extracellular Ca++into cells.