红斑狼疮患者皮损免疫复合物中DNA抗原性质的研究

Characterization of DNA Antigen from Immune Complexes Deposited in the Skin Lesions of Patients with Lupus Erythematosus

目的探讨红斑狼疮(LE)患者皮损免疫复合物中DNA分子的大小和来源.方法取32例LE患者的皮肤标本,用冷沉淀法提取皮肤中免疫复合物,再用蛋白酶消化、酚氯仿抽提其中的DNA分子,经琼脂糖凝胶电泳分析其大小,斑点杂交法检测其可能来源.结果LE患者皮损区标本有27例提取到DNA,经琼脂糖电泳分析显示4种不同大小的DNA片段:Ⅰ(约20 000bp)、Ⅱ(约1 300bp)、Ⅲ(800～900bp)、Ⅳ(100～200bp).15例标本显示以20000 bp的大片段为主;2例以条带Ⅰ、Ⅱ、Ⅲ为主;10例出现所有的4个条带.低相对分子质量DNA片段(100～200bp)的表达与病情成正相关(X2p=4.98,P＜0.05,r=0.43).比患者皮肤免疫复合物中DNA与人类基因组DNA有高度同源性,患者DNA样本之间也有同源片段.结论DNA(核抗原)可能参与了LE皮肤损害中免疫沉积的构成,以内源性的可能性大,并具有一定的相对分子质量范围,从100～20 000 bp不等.其中低相对分子质量的DNA片段具有特异性,与疾病活动性相关.

Objective To explore the characteristics of DNA antigen deposited in the skin lesions of patients with lupus erythematosus (LE). Methods Thirty-two LE skin specimens were collected. The deposited immune complexes were obtained by cryoprecipitation methods. Then the samples were digested by protease. Finally DNA was extracted with phenol and chloroform. The size of DNA fragments was detected on agarose gel electrophoresis. Ten kinds of probes were used to analyze the origin of these DNA molecules by dot hybridization. Results DNA fragments were successfully isolated from 27 skin specimens with four kinds of different sizes including band-I (20 000 bp), band-Ⅱ (1 300 bp), band-Ⅲ (800-900 bp) and band-Ⅳ (100-120 bp). In 15 specimens most of DNA was identified as band-I. In 2 specimens band-Ⅰ, -Ⅱ and -Ⅲ were all noticed, while all four bands were detected in 10 specimens. There was a positive correlation between small-sized fragments (100-200 bp) and disease activity (P < 0.05, r = 0.429). The hybridization showed a significant homology between the isolated DNA fragments and human genomic DNA. There was homology between the DNA fragments isolated from different individuals. Conclusions DNA and its immune complexes might contribute to the pathogenesis of skin lesion of LE. The sizes of these DNAs are 100 bp to 20 000 bp, which may be endogenous. Small sized DNA fragments may be more pathogenic and related to disease activity.