Kundu Soumi, Kumar Munish, Sen Utpal, Mishra Paras K, Tyagi Neetu, Metreveli Naira, Lominadze David, Rodriguez Walter, Tyagi Suresh C
Department of Physiology & Biophysics, University of Louisville School of Medicine, Louisville, Kentucky 40202, USA.
J Cell Biochem. 2009 Jan 1;106(1):119-26. doi: 10.1002/jcb.21982.
Increased levels of homocysteine (Hcy), recognized as hyperhomocysteinemia (HHcy), were associated with cardiovascular diseases. There was controversy regarding the detrimental versus cardio protective role of inducible nitric oxide synthase (iNOS) in ischemic heart disease. The aim of this study was to test the hypothesis that the Hcy generated nitrotyrosine by inducing the endothelial nitric oxide synthase, causing endothelial-myocyte (E-M) coupling. To differentiate the role of iNOS versus constitutive nitric oxide synthase (eNOS and nNOS) in Hcy-mediated nitrotyrosine generation and matrix remodeling in cardiac dysfunction, left ventricular (LV) tissue was analyzed from cystathionine beta synthase (CBS) heterozygote knockout, iNOS homozygote knockout, CBS-/+/iNOS-/- double knockout, and wild-type (WT) mice. The levels of nitrotyrosine, MMP-2 and -9 (zymographic analysis), and fibrosis (by trichrome stain) were measured. The endothelial-myocyte function was determined in cardiac rings. In CBS-/+ mice, homocysteine was elevated and in iNOS-/- mice, nitric oxide was significantly reduced. The nitrotyrosine and matrix metalloproteinase-9 (MMP-9) levels were elevated in double knockout and CBS-/+ as compared to WT mice. Although MMP-2 levels were similar in CBS-/+, iNOS-/-, and CBS-/+/iNOS-/-, the levels were three- to fourfold higher than WT. The levels of collagen were similar in CBS-/+ and iNOS-/-, but they were threefold higher than WT. Interesting, the levels of collagen increased sixfold in double knockouts, compared to WT, suggesting synergism between high Hcy and lack of iNOS. Left ventricular hypertrophy was exaggerated in the iNOS-/- and double knockout, and mildly increased in the CBS-/+, compared to WT mice. The endothelial-dependent relaxation was attenuated to the same extent in the CBS-/+ and iNOS-/-, compared to WT, but it was robustly blunted in double knockouts. The results concluded that homocysteine generated nitrotyrosine in the vicinity of endothelium, caused MMP activation and endothelium-myocyte uncoupling. The generation of nitrotyrosine was independent of iNOS.
同型半胱氨酸（Hcy）水平升高，即高同型半胱氨酸血症（HHcy），与心血管疾病相关。关于诱导型一氧化氮合酶（iNOS）在缺血性心脏病中有害还是具有心脏保护作用存在争议。本研究的目的是检验以下假设：Hcy通过诱导内皮型一氧化氮合酶产生硝基酪氨酸，从而导致内皮-心肌（E-M）偶联。为了区分iNOS与组成型一氧化氮合酶（eNOS和nNOS）在Hcy介导的硝基酪氨酸生成及心脏功能障碍时基质重塑中的作用，对胱硫醚β合酶（CBS）杂合子敲除、iNOS纯合子敲除、CBS-/+ /iNOS-/- 双敲除及野生型（WT）小鼠的左心室（LV）组织进行了分析。检测了硝基酪氨酸、MMP-2和-9水平（酶谱分析）以及纤维化程度（三色染色法）。在心脏环中测定了内皮-心肌功能。在CBS-/+ 小鼠中，同型半胱氨酸升高，而在iNOS-/- 小鼠中，一氧化氮显著减少。与WT小鼠相比，双敲除和CBS-/+ 小鼠中硝基酪氨酸和基质金属蛋白酶-9（MMP-9）水平升高。虽然CBS-/+、iNOS-/- 及CBS-/+ /iNOS-/- 小鼠中MMP-2水平相似，但比WT小鼠高3至4倍。CBS-/+ 和iNOS-/- 小鼠中胶原蛋白水平相似，但比WT小鼠高3倍。有趣的是，与WT小鼠相比，双敲除小鼠中胶原蛋白水平增加了6倍，表明高Hcy与缺乏iNOS之间存在协同作用。与WT小鼠相比，iNOS-/- 和双敲除小鼠的左心室肥厚更为明显，CBS-/+ 小鼠轻度增加。与WT小鼠相比，CBS-/+ 和iNOS-/- 小鼠中内皮依赖性舒张功能减弱程度相同，但双敲除小鼠中则显著减弱。结果表明，同型半胱氨酸在内皮附近产生硝基酪氨酸，导致MMP激活及内皮-心肌解偶联。硝基酪氨酸的产生与iNOS无关。
