Himmelfarb J, McMonagle E, McMenamin E
Maine Medical Center, Portland, and Maine Medical Center Research Institute, South Portland, Maine 04102, USA.
Kidney Int. 2000 Dec;58(6):2571-8. doi: 10.1046/j.1523-1755.2000.00443.x.
Myeloperoxidase-catalyzed oxidative pathways have recently been identified as an important cause of oxidant stress in uremia and hemodialysis (HD), and can lead to plasma protein oxidation. We have examined patterns of plasma protein oxidation in vitro in response to hydrogen peroxide (H2O2) and hypochlorous acid (HOCl). We measured thiol oxidation, amine oxidation, and carbonyl concentrations in patients on chronic maintenance HD compared with patients with chronic renal failure (CRF) and normal volunteers. We have also examined the effect of the dialysis procedure on plasma protein oxidation using biocompatible and bioincompatible membranes.
Plasma proteins were assayed for the level of free thiol groups using spectrophotometry, protein-associated carbonyl groups by enzyme-linked immunosorbent assay, and oxidation of free amine groups using a fluorescent spectrophotometer.
In vitro experiments demonstrate HOCl oxidation of thiol groups and increased carbonyl formation. In vivo, there are significant differences in plasma-free thiol groups between normal volunteers (279 +/- 12 micromol/L), CRF patients (202 +/- 20 micromol/L, P = 0.005) and HD patients (178 +/- 18 micromol/L, P = 0.0001). There are also significant differences in plasma protein carbonyl groups between normal volunteers (0.76 +/- 0.51 micromol/L), CRF patients (13.73 +/- 4.45 micromol/L, P = 0.015), and HD patients (16.95 +/- 2.62 micromol/L, P = 0.0001). There are no significant differences in amine group oxidation. HD with both biocompatible and bioincompatible membranes restored plasma protein thiol groups to normal levels, while minimally affecting plasma protein carbonyl expression.
First, both CRF and HD patients have increased plasma protein oxidation manifested by oxidation of thiol groups and formation of carbonyl groups. Second, HD with biocompatible and bioincompatible membranes restored plasma protein thiol groups to normal levels. Third, these experiments suggest that there is a dialyzable low molecular weight toxin found in uremia that is responsible for plasma protein oxidation.
髓过氧化物酶催化的氧化途径最近被确定为尿毒症和血液透析（HD）中氧化应激的一个重要原因，并且可导致血浆蛋白氧化。我们已经在体外研究了血浆蛋白对过氧化氢（H2O2）和次氯酸（HOCl）的氧化反应模式。我们测量了慢性维持性血液透析患者与慢性肾衰竭（CRF）患者及正常志愿者相比的血浆蛋白巯基氧化、胺氧化和羰基浓度。我们还使用生物相容性和生物不相容性膜研究了透析过程对血浆蛋白氧化的影响。
使用分光光度法检测血浆蛋白中游离巯基的水平，通过酶联免疫吸附测定法检测与蛋白结合的羰基，并用荧光分光光度计检测游离胺基的氧化。
体外实验表明HOCl可氧化巯基并增加羰基形成。在体内，正常志愿者（279±12μmol/L）、CRF患者（202±20μmol/L，P = 0.005）和HD患者（178±18μmol/L，P = 0.0001）之间的血浆游离巯基存在显著差异。正常志愿者（0.76±0.51μmol/L）、CRF患者（13.73±4.45μmol/L，P = 0.015）和HD患者（16.95±2.62μmol/L，P = 0.0001）之间的血浆蛋白羰基也存在显著差异。胺基氧化无显著差异。使用生物相容性和生物不相容性膜进行血液透析均可将血浆蛋白巯基恢复至正常水平，同时对血浆蛋白羰基表达影响最小。
第一，CRF患者和HD患者均存在血浆蛋白氧化增加，表现为巯基氧化和羰基形成。第二，使用生物相容性和生物不相容性膜进行血液透析均可将血浆蛋白巯基恢复至正常水平。第三，这些实验表明尿毒症中存在一种可透析的低分子量毒素，它是血浆蛋白氧化的原因。
