AQP1在维甲酸诱导红白血病细胞红系分化中的作用

目的建立稳定抑制水通道蛋白1（AQP1）表达的K562细胞株，探讨AQP1在维甲酸（RA）诱导红白血病细胞红系分化中
的作用。方法RA处理K562细胞，通过real-time PCR法检测γ-珠蛋白表达和分光光度法检测血红蛋白的含量研究RA对K562
细胞红系分化的影响。Real-time PCR法、蛋白质印迹法检测RA诱导后K562细胞AQP1转录和蛋白表达水平的变化。设计特
异AQP1shRNA序列，构建pSUPER-retro-puro重组质粒转染K562细胞，筛选建立稳定抑制AQP1基因表达的K562细胞株
（K562-shAQP1）；通过比较K562-shAQP1细胞株与对照组细胞在维甲酸诱导后γ-珠蛋白和血红蛋白的表达，研究AQP1在维甲
酸诱导K562细胞红系分化中的作用。结果RA处理K562细胞后红系分化指标γ-珠蛋白和血红蛋白的表达均明显增加，同时
AQP1mRNA及蛋白表达随RA作用时间显著增加（P<0.01）。pSUPER-retro-puro-shAQP1干扰载体转染K562细胞后AQP1基
因在转录和蛋白水平上的表达均被抑制，差异有统计学意义（P<0.01）；K562-shAQP1细胞与对照K562细胞相比，在RA诱导后
γ-珠蛋白和血红蛋白表达受到不同程度的抑制，差异有统计学意义（P<0.01）。结论RA诱导K562细胞红系分化后AQP1表达
显著增加，而抑制AQP1基因的表达可部分阻断RA诱导红系分化的作用，说明AQP1在RA诱导K562细胞红系分化过程中发
挥重要作用。

Role of aquaporin-1 gene in erythroid differentiation of erythroleukemia K562 cells induced by retinoic acid

ObjectiveTo explore the role of aquaporin-1(AQP1) gene in erythroid differentiation of erythroleukemia K562cells
induced by retinoic acid (RA). MethodsK562cells were cultured in the presence of1μmol/L RA for varying lengths of time,
andγ-globin mRNA expression and hemoglobin content in the cells were detected by real-time PCR (RT-PCR) and ultraviolet
spectrophotometry, respectively, to evaluate the erythroid differentiation of K562cells. RT-PCR and Western blotting were
used to examine AQP1expression in the cells following RA treatment. A retroviral expression vector of AQP1small interfering
RNA (pSUPER-retro-puro-shAQP1) was constructed and transfected into K562cells to establish a K562cell line with stable
AQP1down-regulation (K562-shAQP1), in which the changes in γ-globin and hemoglobin expressions after RA treatment
were detected.ResultsRA treatment significantly increasedγ-globin and hemoglobin expressions in K562cells (P<0.01),
causing also significantly enhanced AQP1 mRNA and protein expressions over time (P<0.01). Transfection with the
recombinant plasmids pSuper-retro-puro-shAQP1resulted in stable AQP1suppression in K562cells (P<0.01), which showed
markedly reducedγ-globin and hemoglobin expressions after RA induction as compared to the control K562cells (P<0.01).
ConclusionsK562 cells show a significant increase of AQP1 expression after RA-induced erythroid differentiation, and
suppression of AQP1expression can partially block the effect of RA, suggesting the important role of AQP1in RA-induced
erythroid differentiation of K562cells.