Böger R H, Bode-Böger S M, Kienke S, Stan A C, Nafe R, Frölich J C
Institute of Clinical Pharmacology, Hannover Medical School, Germany.
Atherosclerosis. 1998 Jan;136(1):67-77. doi: 10.1016/s0021-9150(97)00183-4.
L-arginine, the precursor of endogenous nitric oxide (NO), has been shown to enhance endothelial function and to reduce the progression of atherosclerosis in cholesterol-fed rabbits. In the present study, we investigated whether myointimal cell proliferation is enhanced in hypercholesterolaemic rabbit aorta and whether chronic treatment of the rabbits with L-arginine or with the NO synthase inhibitor L-NAME influences this proliferative response and vascular monocyte accumulation. Rabbits were fed 1% cholesterol or normal rabbit chow for 12 weeks. Subgroups of cholesterol-fed rabbits were treated with oral L-arginine (2.25%) or L-NAME (3 mg/dl) in drinking water. Myointimal cell proliferation was quantified in aortic segments by immunohistochemical detection of bromodeoxyuridine (BrdU) incorporation into nuclear DNA; vascular monocyte accumulation was assessed by immunohistochemistry using a monoclonal anti-macrophage/monocyte antibody (RAM-11). Plasma levels of L-arginine and the endogenous NO synthase inhibitor, ADMA, were quantified by high-performance liquid chromatography (HPLC). Cholesterol feeding increased the aortic intima/media (I/M) ratio, which was not measurable in the control group, to 1.9 +/- 0.3. This was paralleled by enhanced cell proliferation (cholesterol, 2.4 +/- 0.2%; P < 0.05; control, 0.02 +/- 0.001% BrdU-positive cells per 72 h) and vascular monocyte accumulation. Double immunostaining for BrdU and alpha-actin showed that about two thirds of the proliferating cells were smooth muscle cells. ADMA levels increased from 0.8 +/- 0.1 micromol/l to 2.2 +/- 0.2 micromol/l in cholesterol-fed rabbits, but were unchanged by L-arginine or L-NAME treatment. Myointimal proliferation and intima/media ratios were correlated with ADMA plasma levels. Dietary L-arginine reduced monocyte accumulation by 85 +/- 2% (P < 0.05 vs cholesterol), myointimal cell proliferation (1.8 +/- 0.3% per 72 h; P < 0.05) and intimal thickening (I/M ratio: 0.7 +/- 0.2), whereas the inhibitor of NO synthase, L-NAME, further increased cell proliferation to 3.1 +/- 0.4% per 72 h (P < 0.05). No significant difference was observed in vascular monocyte infiltration between the cholesterol and L-NAME groups. We conclude that cell proliferation and vascular monocyte accumulation are enhanced in hypercholesterolaemic rabbit aorta. These atherogenic effects can be attenuated by dietary L-arginine. Decreased NO formation might underlie the enhanced monocyte accumulation and cell proliferation in hypercholesterolaemic rabbit aorta. The observed inhibition of cell proliferation adds to our understanding of the antiatherosclerotic effects of L-arginine in vivo.
L-精氨酸是内源性一氧化氮（NO）的前体，已被证明可增强内皮功能，并减缓胆固醇喂养兔的动脉粥样硬化进展。在本研究中，我们调查了高胆固醇血症兔主动脉中肌内膜细胞增殖是否增强，以及用L-精氨酸或一氧化氮合酶抑制剂L-NAME长期治疗兔是否会影响这种增殖反应和血管单核细胞积聚。给兔喂食1%胆固醇或正常兔饲料12周。给喂食胆固醇的兔亚组口服L-精氨酸（2.25%）或在饮用水中加入L-NAME（3mg/dl）进行治疗。通过免疫组织化学检测溴脱氧尿苷（BrdU）掺入核DNA来定量主动脉段中的肌内膜细胞增殖；使用单克隆抗巨噬细胞/单核细胞抗体（RAM-11）通过免疫组织化学评估血管单核细胞积聚。通过高效液相色谱（HPLC）定量血浆中L-精氨酸和内源性一氧化氮合酶抑制剂非对称二甲基精氨酸（ADMA）的水平。喂食胆固醇使主动脉内膜/中膜（I/M）比值增加至1.9±0.3，而对照组无法测量该比值。这与细胞增殖增强（胆固醇组，2.4±0.2%；P<0.05；对照组，每72小时0.02±0.001%BrdU阳性细胞）和血管单核细胞积聚同时出现。对BrdU和α-肌动蛋白的双重免疫染色显示，约三分之二的增殖细胞为平滑肌细胞。喂食胆固醇的兔中ADMA水平从0.8±0.1μmol/l增加至2.2±0.2μmol/l，但L-精氨酸或L-NAME治疗对其无影响。肌内膜增殖和内膜/中膜比值与血浆ADMA水平相关。饮食中的L-精氨酸使单核细胞积聚减少85±2%（与胆固醇组相比，P<0.05），肌内膜细胞增殖减少（每72小时1.8±0.3%；P<0.05），内膜增厚减少（I/M比值：0.7±0.2），而一氧化氮合酶抑制剂L-NAME使细胞增殖进一步增加至每72小时3.1±0.4%（P<0.05）。胆固醇组和L-NAME组之间在血管单核细胞浸润方面未观察到显著差异。我们得出结论，高胆固醇血症兔主动脉中的细胞增殖和血管单核细胞积聚增强。这些致动脉粥样硬化作用可通过饮食中的L-精氨酸减轻。一氧化氮生成减少可能是高胆固醇血症兔主动脉中单核细胞积聚和细胞增殖增强的基础。观察到的细胞增殖抑制增加了我们对L-精氨酸体内抗动脉粥样硬化作用的理解。
