外周血淋巴细胞LDL受体活性测定新法

A simple assay for LDL receptor activities in human periphral lymphocytes

混悬于RPMI-1640中的淋巴细胞分两组培养96小时,一组加入0.6μM mevinolin(实验组),另一组加入等量的二甲亚砜溶媒(对照组),两组均加入50μg/ml PHA及不同浓度的LDL-ch(0/10μg/ml)。细胞收获前6小时各孔加入0.5μCi ~3H-TdR,收获细胞于4.9型纤维滤膜上测cpm值。LDL受体活性以淋巴细胞分裂抑制率表示,后者=(1-cpm实验/cpm对照)×100%。结果,当LDL-ch浓度为5μg/ml时,正常人分裂抑制率小于20%,FH杂合子分裂抑制率为25%～55%,FH纯合子分裂抑制率大于60%。由此可将FH与正常人以及普通高脂血症病人区别开来。

Periphral lymphocytes suspended in RPMI-1640 were incubated in plates of 96 wells for 96 hours with 50 μg/ml of PHA, various concentrations of LDL-ch and0.6 μM of mevinolin (experimental) or equal volume of its solvent-dimethyl sulfoxide (control). 6 hours before harvesting cells, 0.5μCi of ~3H-TdR was added to each well, then cells were harvested and their cpm counted. The LDL receptor activity was expressed as mevinolin-mediated lymphocyte proliferation inhibition rate which can be calculated by the following equation:Percent Inhibition Rate=(1-cpm exp./cpm con.)100%.It was found that the lymphocyte proliferation inhibition rate at 5μg/ml of LDL-ch was less than 20% for normal and non-FH hyperlipidemic subjects, 25-55% for FH heterozygotes and over 60% for FH homozygotes. Thus, FH patients were distinguished from normal subjects and non-FH hyperlipidemic patients.