PHORBOL ESTER STIMULATES ACETYLCHOLINE SYNTHESIS IN CULTURED ENDOTHELIAL-CELLS ISOLATED FROM PORCINE CEREBRAL MICROVESSELS
PHORBOL ESTER STIMULATES ACETYLCHOLINE SYNTHESIS IN CULTURED ENDOTHELIAL-CELLS ISOLATED FROM PORCINE CEREBRAL MICROVESSELS
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DOI：
10.1016/0006-8993(94)91608-x
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发表时间：
1994-08-29
期刊：
影响因子：
2.9
通讯作者：
MUROTA, SI
中科院分区：
文献类型：
--
作者：
IKEDA, C;MORITA, I;MUROTA, SI
乙酰胆碱（Acetylcholine，ACh）是通过释放内皮源性舒张因子而引起血管舒张的因子之一。本研究的目的是阐明内皮细胞是否可以合成ACh以及调节内皮细胞合成ACh的物质类型。我们测定了从猪脑微血管中分离的内皮细胞和培养液的ACh含量。ACh在培养12 h后在培养基中检测到二异丙基氟磷酸盐，一种非特异性胆碱酯酶抑制剂的存在下，并线性增加至24 h。佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA，10（-7）M）以剂量依赖方式增加培养液中ACh含量。PMA的作用在处理后12~24 h最为明显，放线菌酮可抑制PMA的作用。蛋白激酶C（PKC）的特异性抑制剂Calphostin C不抑制PMA的作用。二辛酰甘油，PKC的特异性激活剂，不增加细胞内ACh含量或释放到培养基中的量。乙酰胆碱合成不抑制溴乙酰胆碱，胆碱乙酰转移酶（ChAT）的特异性抑制剂。PMA处理不影响内皮细胞合成ACh的比活性。这些数据表明，内皮细胞能够合成ACh，并且PMA通过蛋白诱导的PKC非依赖性机制上调ACh合成。在内皮细胞中合成ACh的酶不是ChAT。PMA诱导的ACh合成增加可能不是由于ACh合成酶的诱导。
Acetylcholine (ACh) is one of the factor which induces vasodilation through the release of endothelium-derived relaxing factor. The aim of this study was to clarify whether endothelial cells can synthesize ACh and the types of substance which regulate the synthesis of ACh in endothelial cells. We determined the ACh content of endothelial cells isolated from porcine cerebral microvessels and of the culture medium. ACh was detected in the medium after 12 h incubation in the presence of diisopropylfluorophosphate, a non-specific cholinesterase inhibitor, and increased linearly up to 24 h. Phorbol 12-myristate 13-acetate (PMA, 10(-7) M) increased the ACh content of the medium in a dose-dependent manner. The effect of PMA was most apparent between 12 and 24 h after treatment, and was inhibited by cycloheximide. Calphostin C, a specific inhibitor of protein kinase C (PKC), did not inhibit the effect of PMA. Dioctanoyl glycerol, a specific activator of PKC, did not increase the intracellular ACh content or the amount released into the culture medium. ACh synthesis was not inhibited by bromoacetyl-choline, a specific inhibitor of choline acetyltransferase (ChAT). PMA treatment did not affect the specific activity of ACh synthesis in endothelial cells. These data show that endothelial cells are able to synthesize ACh, and that ACh synthesis is up-regulated by PMA through the PKC independent mechanism via protein induction. The enzyme which synthesizes ACh in endothelial cells is not ChAT. The increase in ACh synthesis induced by PMA may not be due to induction of the ACh synthetic enzyme.