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目的获得hnRNPA2/B1cDNA序列,研究其在滑膜组织中的表达水平,探讨它在类风湿滑膜炎中可能的致病作用。方法从人外周血单个核细胞中提取总RNA进行逆转录反应,经PCR扩增目的片段,将其克隆于PUC-T1质粒上,并经酶切电泳、DNA测序鉴定分析。采用免疫组化和原位杂交方法,分别利用单克隆抗体和特异的cDNA探针,检测滑膜组织中hnRNPA2/B1的含量及表达水平。结果从人外周血单个核细胞总RNA中扩增得到目的片段,并经测序证实该片段为hnRNPA2/B1的编码序列。免疫组化和原位杂交显示hnRNPA2/B1在类风湿关节炎(RA)滑膜中的表达水平整体上较骨关节炎(OA)和正常对照滑膜组高(P<0.05)。结论成功克隆hnRNPA2/B1cDNA;初步证明hnRNPA2/B1水平在RA关节局部增高,推测它可能参与了类风湿滑膜炎的发病。
OBJECTIVE: To obtain the hnRNPA2 / B1 cDNA sequence and investigate the expression of hnRNPA2 / B1 in synovial tissue and to explore its possible pathogenic role in rheumatoid synovitis. Methods Total RNA was extracted from human peripheral blood mononuclear cells and subjected to reverse transcription reaction. The target fragment was amplified by PCR, cloned into PUC-T1 plasmid, and identified by DNA sequencing. Immunohistochemistry and in situ hybridization methods were used to detect the content and expression of hnRNPA2 / B1 in synovial tissue by using monoclonal antibodies and specific cDNA probes respectively. Results The target fragment was amplified from total RNA of human peripheral blood mononuclear cells and confirmed by sequencing to be the coding sequence of hnRNPA2 / B1. Immunohistochemistry and in situ hybridization showed that the expression level of hnRNPA2 / B1 in rheumatoid arthritis (RA) synovium was overall higher than that in osteoarthritis (OA) and normal control synovial group (P <0.05). Conclusion The hnRNPA2 / B1 cDNA was cloned successfully. It was initially proved that the level of hnRNPA2 / B1 was locally increased in RA joints, suggesting that it may be involved in the pathogenesis of rheumatoid synovitis.