扩张型心肌病新型抗钙通道抗体的发现

A novel autoantibody against Ca^(2+) channel in patients with idiopathic dilated cardiomyopathy

目的:观察扩张型心肌病(DCM)患者血清中是否独立存在抗心肌细胞L型钙离子通道(Human Cav1.2)自身抗体(简称:抗钙通道抗体)。方法:以合成多肽片段作为抗原,用ELISA法检测DCM患者(DCM组)及正常人(正常对照组)(各60例)血清中的抗钙通道抗体和抗心肌细胞线粒体内膜ADP/ATP载体蛋白(ANT)抗体。用亲合层析法提纯DCM患者(DCM组)血清中的抗钙通道抗体,以制备的大鼠抗Human Cav1.2α1亚单位多克隆抗体作为阳性对照,用免疫印记技术,免疫荧光组织化学技术检测DCM患者血清中抗钙通道抗体。结果:ELISA检测60例DCM患者中29例抗钙通道抗体阳性(29/60,阳性率48.33%),明显高于正常对照组(4/60,6.67%,P<0.01)。抗钙通道抗体和抗ANT抗体检出结果无一致性(P>0.05)。免疫印记中亲合层析法提纯的DCM患者抗钙通道抗体可特异地识别240000的多肽(Human Cav1.2α1c亚单位),结果与阳性对照相符。抗钙通道抗体经抗原充分吸附后特异性识别被阻断。免疫荧光实验显示,抗钙通道抗体能特异性地结合在大鼠心室肌细胞膜上。结论:DCM患者血清中存在新型的抗钙通道抗体。

Objective： In this study we sought to investigate whether there is an independent serum autoantibody against the cardiac L-type Ca^2＋ channel （human Cav1. 2） in patients with dilated cardiomyopathy （DCM）. Method：Peptide corresponding to the sequence （residues 2- 15） of the S1 segment, domain Ⅰ of 1 subunit, human Cavl. 2 was detected as an antigen to screen sera from patients with DCM （n= 60）, and healthy blood donors （HBD, control group, n 60） in ELISA while the autoantibody against the adenine nucleotide translocator（ANT） was monitored. Affinity-purified autoantibodies from positive sera of patients with DCM and rat antipeptide polyclonal antibodies were deteceted by immunoblotting assay and immunofluorescence technique. Result：Sera from 29 patients with DCM （48. 33%） and 4 HBD （6.67%） recognized the peptide of human Cavl. 2 in ELISA and the difference was significant （P〈0.01）. Autoantibody against the ANT and the autoantibody against the human Cavl. 2 were not in concordance. Affinity-purified autoantibody from positive sera of patients with DCM recognized a major band of about 240kD on the electrotransferred protein of rat ventricular cell membrane. This band was monospecific since its staining was blocked after preincubation with the human Cav1. 2 peptide and it coincided well with the band revealed by rat antipeptide polyclonal antibodies. In immunofluorescence, the stain was specially located with membranes of rat ventricular cardiocytes. Conclusion：These data confirme that there is a novel type of autoantibody against the human Cav1. 2 in the sera of patients with DCM.