应用基因芯片研究肾移植急性排斥发生时外周血免疫基因变化

Immunological gene change in peripheral blood of the renal transplant recipients during the acute rejection phase by gene chips

目的借助基因芯片研究肾移植受者外周血淋巴细胞免疫相关基因在急性排斥反应中的作用。方法8例肾移植术后发生急性排斥反应患者,于手术和肾穿刺当日收集外周血作为对照血样和实验血样。用Ficoll技术分离外周血淋巴细胞,并采用一步法提取总RNA,逆转录合成cDNA探针,荧光染料标记后,采用免疫相关基因芯片进行芯片杂交,扫描后筛选出差异表达基因。结果急性排斥发生时外周血淋巴细胞免疫相关基因差异表达49条,其中上调25条,下调24条。结论从基因水平上,外周血淋巴细胞在肾移植急性排斥的免疫应答中涉及不同环节,免疫抑制剂在这些环节上对基因的表达存在不同程度的影响。

Objective:To explore the role of peripheral blood lymphocyte (PBL) of the renal transplant recipients during the acute rejection phase by gene chips.Methods:The 8 patients with acute rejection (AR) after renal transplantation were collected peripheral blood before operation (as control samples) and renal biopsy (as experimental samples).By Ficoll method,PBL was collected.Total RNA were extracted by one-step technique and purified.The total RNA were labeled with Cy5-dUTP (experimental samples) or Cy3-dUTP (control samples),then to label the cDNA probe by reverse transcript way.The gene chip (419 genes) was hybridized and scanned.Then fluorescent signal value of gene expressing was obtained,and differential expression genes were sifted.Results:There were differential expression 49 immunological genes in peripheral blood lymphocyte (PBL) of the renal transplant recipients during the acute rejection phase,including up-regulated 25 and down-regulated 24.Conclusion:Peripheral blood lymphocyte was involved in various stages during the acute rejection,and immunosuppressants influenced on these stages in various degrees. [