Anandamide uptake by human endothelial cells and its regulation by nitric oxide
Anandamide uptake by human endothelial cells and its regulation by nitric oxide
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DOI：
10.1074/jbc.275.18.13484
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发表时间：
2000-05-05
影响因子：
4.8
通讯作者：
Finazzi-Agrò, A
中科院分区：
文献类型：
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作者：
Maccarrone, M;Bari, M;Finazzi-Agrò, A
大麻素（AEA）具有血管扩张活性，可通过细胞再摄取和降解而终止。我们研究了人脐静脉内皮细胞（HUVECs）中AEA转运蛋白的存在和调节，HUVECs通过易化转运（表观Km = 190 +/- 10 nM和V-max = 45 +/- 3 pmol min（-1））摄取AEA。mg（-1）蛋白质），其被α-亚麻酰基-香草基-酰胺和N-（4-羟基苯基）花生四烯酸酰胺抑制，并被一氧化氮（NO）供体刺激高达2.2倍。NO清除剂羟钴胺素废除后者的效果，而不是增强超氧阴离子，但抑制超氧化物歧化酶和N-乙酰半胱氨酸，谷胱甘肽合成的前体。过氧亚硝酸盐（ONOO-）导致AEA转运到细胞中的4倍活化。HUVEC AEA转运蛋白有助于终止AEA的典型1型大麻素受体（CB 1）介导的作用，即抑制毛喉素刺激的腺苷酸环化酶，因为发现NO/ONOO-供体和α-亚麻酰-香草酰-酰胺/N-（4-羟基苯基）-花生四烯酸酰胺分别减弱和增强AEA的这种作用。一致地，通过AEA或大麻素HU-210激活CB、大麻素受体引起HUVEC诱导型NO合酶活性和表达的刺激分别高达2.9倍和2.6倍。HU-210以对NO合成酶抑制剂N ω-硝基-L-精氨酸甲酯敏感的方式增强HOVEC对AEA的摄取。这些发现表明CB 1大麻素受体和AEA转运蛋白之间存在NO介导的调节环。
Anandamide (AEA) has vasodilator activity, which can be terminated by cellular re-uptake and degradation. Here we investigated the presence and regulation of the AEA transporter in human umbelical vein endothelial cells (HUVECs), HUVECs take up AEA by facilitated transport (apparent K-m = 190 +/- 10 nM and V-max = 45 +/- 3 pmol min(-1) . mg(-1) protein), which is inhibited by alpha-linolenoyl-vanillyl-amide and N-(4-hydroxyphenyl) arachidonoylamide, and stimulated up to 2.2-fold by nitric oxide (NO) donors. The NO scavenger hydroxocobalamin abolishes the latter effect, which is instead enhanced by superoxide anions but inhibited by superoxide dismutase and N-acetylcysteine, a precursor of glutathione synthesis. Peroxynitrite (ONOO-) causes a 4-fold activation of AEA transport into cells. The HUVEC AEA transporter contributes to the termination of a typical type 1 cannabinoid receptor (CB1) -mediated action of AEA i.e. the inhibition of forskolin-stimulated adenylyl cyclase, because NO/ONOO- donors and alpha-linolenoyl-vanillyl-amide/N-(4-hydroxyphenyl)-arachidonoylamide were found to attenuate and enhance, respectively, this effect of AEA. Consistently, activation of CB, cannabinoid receptors by either AEA or the cannabinoid HU-210 caused a stimulation of HUVEC inducible NO synthase activity and expression up to 2.9- and 2.6-fold, respectively. Also these effects are regulated by the AEA transporter, HU-210 enhanced AEA uptake by HOVECs in a fashion sensitive to the NO synthase inhibitor N omega-nitro-L-arginine methyl ester, These findings suggest a NO-mediated regulatory loop between CB1 cannabinoid receptors and AEA transporter.