体外培养Wilson病患者肝细胞铜转运研究

Cu^(2+) -transporting in Cultured Hepatocytes of Wilson Disease Patients

【目的】探讨铜转运P型ATP酶在离体肝细胞铜代谢及Wilson病 (WD)发病机制中的作用。【方法】WD患者和对照者离体培养肝细胞模型 ,经单用铜 (15mg/L)及铜加P型ATP酶阻滞剂矾酸钠 (18 39mg/L)、铜加P型ATP酶激动剂长春新碱 (0 5mg/L)孵育 ,差速离心分离肝细胞胞浆、溶酶体、线粒体和微粒体 ,在不同孵育条件下WD患者及对照组各亚细胞组分的铜含量分别对照分析。【结果】WD患者肝细胞各组分的铜含量均显著高于对照组 ,P型ATP酶阻滞剂与激动剂对WD患者肝细胞微粒体、质膜铜转运的作用均出现异常。【结论】WD患者肝细胞亚组分的铜转运P型ATP酶的功能障碍 ,在WD的发病机制中有重要作用。

Objective To evaluate the effect of copper transporting P-type ATPase on hepatocyte′s copper metabolism and pathogenesis of Wilson disease (WD). Methods Human cultured hepatocytes from WD patients and controls were incubated in media of copper 15 mg/L only, copper 15 mg/L with vincristine (agonist of P-type ATPase) 0 5 mg/L, or copper 15 mg/L with vanadate (antagonist of P-type ATPase) 18 39 mg/L. Microsome (endoplasmic veticulum), lysosome, mitochondria, and cytosol were isolated by differential centrifugation. The influence on copper transport of these organelles by vanadate and vincristine were comparatively analyzed between WD patients and controls under different culturing conditions. Results Significant disorders of copper transport in these organelles of WD patients were found comparing with controls in each group. The antagonist and agonist of P-type ATPase obviously changed copper transporting of microsomes and plasma membranes. Conclusion Copper transporting P-type ATPase plays an important role in hepatocytic copper metabolism. Dysfunction of hepatocytic copper transporting P-type ATPase might be one of the most important pathogenesis of WD.