血液透析病人外周血单核细胞糖基化终产物受体的表达及其与细胞因子水平的关系

Expression of receptors for advanced glycation end products at surface of peripheral blood monocytes and relationship thereof to plasma proinflammatory cytokines in patients undergoing hemodialysis

目的 探讨慢性肾功能衰竭长期血液透析病人外周血单核细胞表面晚期糖基化终产物(AGE)受体的表达及其与循环肿瘤坏死因子α(TNFα)和白细胞介素 1β(IL 1β)水平之间的关系 ,以及血透病人循环中高水平促炎症细胞因子的发生机制。方法 选择慢性肾功能衰竭病人 5 9例 (其中非透析组 2 2例 ,维持性血透组 37例 )和正常人 30例 ,观察单核细胞表面AGE结合蛋白的数量和亲和力 ,检测单核细胞表面AGE受体 (RAGE)的表达及单核细胞培养上清中的TNFα和IL 1β的水平。 结果 血透病人单核细胞与12 5I AGE 白蛋白的特异性结合率 (41 4fmol/ 10 5细胞± 2 2fmol/ 10 5细胞 )较正常人 (31 6fmol/ 10 5细胞± 4 1fmol/ 10 5细胞 ,P <0 0 5 )明显升高 ,细胞表面AGE结合蛋白的数目(2 4 2± 0 2 1× 10 5/细胞 )高于正常人 (1 74± 0 2 9× 10 5/细胞 ,P <0 0 5 ) ,但亲和常数 (Kd)与正常人单核细胞相比无明显差异 ;血透病人单核细胞表面RAGE表达亦高于正常人 ;在相同剂量AGE 白蛋白刺激下 ,血透病人单核细胞分泌的TNFα和IL 1β水平较正常人明显升高 ,这种升高作用可被抗RAGE抗体所阻断 ;单核细胞AGE结合蛋白表达上调的血透病人 ,其循环TNFα和IL 1β水平亦显著升高。结论 血透病人单核细胞AGE受体表达上调 。

Objective To elucidate the relationship between the expression of receptors for AGE (RAGE) at the surface of monocyte and the plasma levels of tumor nocrosis factor α (TNFα) and interleukin 1β (IL 1β), proinflammatory cytokines in patients with chronic renal failure (CRF) undergoing hemodialysis (HD). Methods Human serum albumn modified with AGE (AGE HSA) was prepared in vitro and was radiolabeled with 125 I. Human peripheral blood monocytes were isolated from 59 CRF patients (37 of which underwent hemodialysis) and 30 normal volunteers by Ficoll hypaque centrifugation technique. Specific binding of AGE HSA to monocytes was analyzed by radioactive ligand receptor binding assay. The degradation to AGE HSA by monocytes was measured by trichloroacetic acid precipitation method. The expression of REGE in monocytes was determined by flow cytometry. The levels of TNFα and IL 1β were detected by a sandwich enzyme immunoassay. Results The specific binding of 125 I AGE HSA to monocytes was 41.4 fmol/10 5 cells±2.2 fmol/10 5 cells, significantly higher than that in normal controls (31.6 fmol/10 5 cells±4.1 fmol/10 5 cells, P <0.05). The number of AGE conjugated protein at the monocytes′ surface of CRF patients undergoing HD was 2.42±0.21×10 5/cells, significantly higher than that in normal controls (1.74±0 29×10 5/cells, P <0.001), and the affinity constant (Kd) of the monocytes of CRF patients undergoing HD was 228 nmol/L±100 nmol/L, not significantly different from that in normal controls (262 nmol/L±108 nmol/L, P >0.05). The degration rate of AGE HAS by monocytes in CRF patients undergoing HD was 25.24%±4.35%, significantly higher than that in normal controls (18%±0.6%, P <0.05). The fluorescent intensity of RAGE at the surface of monocytes in CRF patients undergoing HD was 3.68±0.42, significantly higher than that in normal controls (1.09±0.37, P <0.05), When the monocytes were incubated in vitro with 50 μg/ml of AGE HSA, the levels of TNFα and IL 1β in the supernatants of CRF patients undergoing HD were 197±98 ng/L and 357.0±140.1 ng/L respectively, both significantly higher than those in the normal controls (111±77 ng/L and 184±118 ng/L respectively, both P <0.05). The secretion of proinflammatory cytokines was inhibited when the monocytes were preincubated with anti RAGE. Increased RAGE level was accompanied by higher plasma levels of TNFα and IL 1β. Conclusion The expression of AGE receptors in monocytes is up regulated in patients with CRF. This may contribute to the enhanced plasma level of proinflammatory cytokines seen in HD patients.