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Modulation of intracellular Ca ion concentrations by endothelium-derived hyperpolarizing factor in vascular smooth muscle

血管平滑肌内皮源性超极化因子对细胞内钙离子浓度的调节

基本信息

批准号：
04454150
负责人：
SUZUKI Hikaru
金额：
$ 4.03万
依托单位：
Nagoya City University Medical School
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454150/
关键词：
Endothelium-derived hyperpolarizing factao Vascular smooth muscle Calcium

项目摘要

(1) Vascular smooth muscles for the measurement of intracellular Ca ion concentrations and also for mechanical responses were successfully prepared in the guinea-pig aorta. Smooth muscle of this artery was confirmed to be relaxd by EDHF.(2) Endothelial cells isolated from bovine thoracic aorta using enzymatic dispersion were cultured on microcarrier beeds. Preliminary experiments showed that these cells (3x10^7 cells/each experiment) can produce several types of vasoactive substance in response to bradikinine (Bk).(3) Production of EDHF from the cultured endothelial cells was estimated by using isolated thoracic aortic ring of the guinea-pig. Superfusates of the cultured endothelial cells with Bk relaxd the Bk-contracted aortic ring preparations, and the relaxation was resistant to nitroarginine and indomethacin, but sensitive to high-K solution containing nitroaginine. The results suggested that Bk stimulates the cultured endothelial cells to release vesorelaxtant factors which may be … More different from EDRF or PG12. The effects of these unidentified factors reduced in high-K solution, indicated an involvement of membrane hyperpolarization.4. Intracellular Ca ion concentrations modulated by EDHF were estimated from the fluorescent intensities of Ca-sensitive dye, Fura-2 loaded in primary cultured smooth muscle cells of the guinea-pig aorta (Fura-2 was loaded into the cell in a form of Fura-2 AM). Bk elevated the fluorescent transiently to level equivalent to 300-500nM Ca from the resting level of about 50nM, and them reached the steady level of about 200nM.Bk-containing superfusate elluted from the cultured endothelial cells reduced the Fura-2 fluorescent to the resting level, irrespective to the absence and presence of nitroarginine and indonethacin. The reduction of fluorescent intensity by the endothelial effluents was not clearly detected in high-K solution. The results suggested that hyperpolarization by endothelial factor may be responsible to the reduction of fluorescent. Thus, EDHF relax arterial smooth muscles with reduced Ca concentrations. Less

(1)在豚鼠主动脉中成功地制备了用于测量细胞内钙离子浓度和机械响应的血管平滑肌。EDHF证实该动脉平滑肌松弛。(2)采用酶法分离牛胸主动脉内皮细胞，并将其置于微载体上培养。初步实验表明，这些细胞（3 × 10^7个细胞/每次实验）可以产生几种类型的血管活性物质，以响应布拉迪基宁（BK）。(3)用离体豚鼠胸主动脉环测定培养的内皮细胞产生EDHF的能力。用BK超灌注培养的内皮细胞可舒张BK收缩的主动脉环，这种舒张作用对硝基精氨酸和吲哚美辛不敏感，但对含硝基精氨酸的高钾溶液敏感。结果提示，BK刺激培养的内皮细胞释放血管舒张因子，可能是其作用机制之一。 ...更多信息不同于EDRF或PG 12。在高钾溶液中，这些未知因子的作用减弱，表明膜超极化参与其中.由EDHF调节的细胞内Ca离子浓度由加载在原代培养的豚鼠主动脉平滑肌细胞中的Ca敏感染料Fura-2的荧光强度估计（Fura-2以Fura-2AM的形式加载到细胞中）。Bk使Fura-2的荧光强度由静息时的约50 nM升高至300- 500 nM Ca，并达到稳定水平（约200 nM），含Bk的灌流液使Fura-2的荧光强度降低至静息时的水平，而不管有无nitroarginine和indonethacin。在高钾溶液中，没有清楚地检测到内皮流出物对荧光强度的降低。结果提示，内皮细胞因子引起的超极化可能是荧光减弱的原因。因此，EDHF松弛动脉平滑肌，降低Ca浓度。少