Nitric oxide synthase activity in tissues of the bovine eye
Nitric oxide synthase activity in tissues of the bovine eye
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DOI：
10.1007/bf02332864
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发表时间：
1997-12-01
影响因子：
2.7
通讯作者：
Mittag, T
中科院分区：
文献类型：
--
作者：
Geyer, O;Podos, SM;Mittag, T
背景：一氧化氮合酶（NOS）广泛存在于眼的多种组织中，具有不同的生理功能。方法：用免疫组织化学方法检测视网膜、脉络膜、睫状突（CP）和小梁网（TM）中NOS的分布及其生化特性。需要NADPH的视网膜NOS（二苯基碘鎓，黄素蛋白抑制剂，其抑制酶活性的IC 50为0.36 μ M，FAD（40 μ M），FMN（40 μ M），和BH 4（4 μ M）作为最佳活性的辅因子。眼部NOS似乎受游离二价阳离子调节，因为其活性被EDTA抑制（斜率>3.0，IC 50值分别为12.8、19.7和53 μ M）。眼部NOS需要钙调素，因为NOS活性被三氟拉嗪（钙调素抑制剂，IC 50 =41 μ M）抑制。NOS活性广泛分布于眼内（脉络膜>视网膜>CP >TM），并且主要是细胞溶质（70-95%）。L-精氨酸类似物抑制视网膜、脉络膜和TM中的NOS。在所有三种组织中，N-G-甲基-L-精氨酸显示出最高的抑制亲和力（IC 50 =0.2-0.7 μ M），其次是刀豆氨酸（IC 50 =13-33 μ M），而氨基胍仅微弱地抑制NOS（IC 50 =93-179 μ M）。此外，TM可能含有不止一种形式的NOS。
Background: Nitric oxide synthase (NOS) is present in many ocular tissues where it may have different physiological functions. This warrants a thorough characterization of NOS activity in the eye.Methods: NOS distribution and its biochemical properties were determined in the retina, choroid, ciliary processes (CP), and trabecular meshwork (TM),Results: Retinal NOS required NADPH (diphenylene-iodonium, a flavoprotein inhibitor, which inhibited enzyme activity with an IC50 of 0.36 mu M, FAD (40 mu M), FMN (40 mu M), and BH4 (4 mu M) as cofactors for optimal activity. Ocular NOS appeared to be regulated by free divalent cations, since its activity was inhibited by EDTA (slopes >3.0 and IC50 values of 12.8, 19.7, and 53 mu M, respectively). Ocular NOS required calmodulin, since NOS activity was inhibited by trifluoperazine (calmodulin inhibitor, IC50=41 mu M). NOS activity is widely distributed in the eye, (choroid >reinta >CP >TM) and is mainly cytosolic (70-95%). L-Arginine analogs inhibited NOS in the retina, choroid, and TM. In all three tissues, N-G-methyl-L-arginine displayed the highest affinity for inhibition (IC50=0.2-0.7 mu M) followed by canavanine (IC50=13-33 mu M), while aminoguanidine only weakly inhibited NOS (IC50=93-179 mu M).Conclusion: In all tissues, the order of potency of inhibition points to the presence of constitutive rather than inducible NOS. Moreover, it is possible that TM contains more than a single form of NOS.