Department of Physiology and Biophysics, University of Louisville School of Medicine, Louisville, Kentucky 40202, USA.
Am J Physiol Lung Cell Mol Physiol. 2010 Sep;299(3):L301-11. doi: 10.1152/ajplung.00065.2010. Epub 2010 Jun 25.
A decrease in vascular elasticity and an increase in pulse wave velocity in hyperhomocysteinemic (HHcy) cystathionine-beta-synthase heterozygote knockout (CBS(-/+)) mice has been observed. Nitric oxide (NO) is a potential regulator of matrix metalloproteinase (MMP) activity in MMP-NO-tissue inhibitor of metalloproteinase (TIMP) inhibitory tertiary complex. However, the contribution of the nitric oxide synthase (NOS) isoforms eNOS and iNOS in the activation of latent MMP is unclear. We hypothesize that the differential production of NO contributes to oxidative stress and increased oxidative/nitrative activation of MMP, resulting in vascular remodeling in response to HHcy. The overall goal is to elucidate the contribution of the NOS isoforms, endothelial and inducible, in the collagen/elastin switch. Experiments were performed on six groups of animals [wild-type (WT), eNOS(-/-), and iNOS(-/-) with and without homocysteine (Hcy) treatment (0.67 g/l) for 8-12 wk]. In vivo echograph was performed to assess aortic timed flow velocity for indirect compliance measurement. Histological determination of collagen and elastin with trichrome and van Gieson stains, respectively, was performed. In situ measurement of superoxide generation using dihydroethidium was used. Differential expression of eNOS, iNOS, nitrotyrosine, MMP-2 and -9, and elastin were measured by quantitative PCR and Western blot analyses. The 2% gelatin zymography was used to assess MMP activity. The increase in O(2)(-) and robust activity of MMP-9 in eNOS(-/-), WT+Hcy, and eNOS(-/-)+Hcy was accompanied by the gross disorganization and thickening of the ECM along with extensive collagen deposition and elastin degradation (collagen/elastin switch) resulting in a decrease in aortic timed flow velocity. Results show that an increase in iNOS activity is a key contributor to HHcy-mediated collagen/elastin switch and resulting decline in aortic compliance.
在高同型半胱氨酸血症（HHcy）胱硫醚-β-合酶杂合子敲除（CBS（- / +））小鼠中观察到血管弹性降低和脉搏波速度增加。一氧化氮（NO）是基质金属蛋白酶（MMP）活性在 MMP-NO-金属蛋白酶组织抑制剂（TIMP）抑制性三元复合物中的潜在调节剂。然而，内皮型一氧化氮合酶（eNOS）和诱导型一氧化氮合酶（iNOS）在潜在 MMP 激活中的作用尚不清楚。我们假设，NO 的差异产生有助于氧化应激和 MMP 的氧化/硝化激活增加，从而导致对 HHcy 的血管重塑。总体目标是阐明内皮型和诱导型一氧化氮合酶同工酶在胶原/弹性蛋白转换中的贡献。在六组动物[野生型（WT），eNOS（- / -）和 iNOS（- / -）]中进行了实验，并且有和没有同型半胱氨酸（Hcy）处理（0.67 g / l）8-12 周。进行体内超声心动图以评估主动脉时间流速以进行间接顺应性测量。分别用三原色和凡戈尔氏染色法进行胶原和弹性蛋白的组织学测定。使用二氢乙啶原位测量超氧化物的产生。通过定量 PCR 和 Western blot 分析测量 eNOS、iNOS、硝基酪氨酸、MMP-2 和 -9 以及弹性蛋白的差异表达。使用 2％明胶酶谱法评估 MMP 活性。在 eNOS（- / -），WT+Hcy 和 eNOS（- / -）+Hcy 中，O2（-）的增加和 MMP-9 的强活性伴随着 ECM 的严重紊乱和增厚，以及广泛的胶原沉积和弹性蛋白降解（胶原/弹性蛋白转换），导致主动脉时间流速降低。结果表明，iNOS 活性的增加是 HHcy 介导的胶原/弹性蛋白转换和主动脉顺应性下降的关键因素。
