急性早幼粒细胞白血病患者血肿瘤细胞NLS-RARα蛋白的表达与定位

Location of NLS-RARαprotein in peripheral blood tumor cells of patients with acute promyeolic leukemia

目的验证急性早幼粒细胞白血病(APL)患者血肿瘤细胞中带核定位信号的维甲酸受体α(nuclear localization signal-retinoic acid receptor alpha,NLS-RARα)蛋白的存在及定位。方法采用蛋白质印迹法验证患者血肿瘤细胞中性粒细胞弹性蛋白酶(neutrophil elastase,NE)的存在;提取患者血肿瘤细胞核蛋白,蛋白质印迹法检测细胞核中NLS-RARα蛋白的表达;FITC/DAPI双染色免疫荧光法检测患者血肿瘤细胞中NLS-RARα的表达及定位;FITC/PI双染色激光共聚焦法检测患者血肿瘤细胞中NLS-RARα的表达及定位。以重组腺病毒Ad-NE感染的NB4细胞中NLS-RARα蛋白的表达及定位作阳性对照,以正常人血中性粒细胞中野生型RARα的表达和定位作阴性对照。结果阳性对照组设置成功。APL患者血肿瘤细胞中存在NE且有NLS-RARα蛋白表达。细胞免疫荧光法、激光共聚焦法检测结果提示APL患者血肿瘤细胞NLSRARα蛋白的表达主要位于胞核。结论成功用3种方法检测出APL患者血肿瘤细胞中NLS-RARα蛋白的存在并推测其定位,为进一步研究APL的早期诊断及复发监测提供了新思路。

Objective To verify the presence and location of the NLS-RARa protein in the peripheral blood tumor cells of patients with acute promyeolic leukemia （APL）. Methods Western blotting analysis was used to identify the NE enzyme in the peripheral blood tumor cells of APL patients. The nucleoprotein in tumor cells was prepared and NLS-RARa protein was detected by Western blotting analysis. The expression and location of NLS-RARa protein in peripheral blood tumor cells of APL patients were examined by FITC/DAPI double immunofluorescence staining and FITC/PI double staining laser confocal microscopy. The expression and location of NLS-RARa protein in NB4 cells infected with recombinant adenovirus Ad-NE was used as positive control and those of wildtype RARa in the neutrophils of healthy controls were taken as negative control. Results Positive control was successfully established. NE enzyme and NLS-RARa protein were expressed in peripheral blood tumor cells of APL patients. Imrnunofluorescence and laser confocai findings indicated that NLS-RARa protein was mainly located at the nuclei of peripheral blood tumors cells in APL patients. Conclusion NLS-RARa protein has been successfully detected by 3 different methods in the peripheral blood tumor cells of APL patients and its intracellular location has also been proposed, which can contribute to the early diagnosis and recurrence monitoring of APL.