ZEB2和PTEN在先天性巨结肠发生中的意义分析

目的研究ZEB2和PTEN在先天性巨结肠中的表达情况,探讨两者在先天性巨结肠发生中可能的调控关系。方法采用实时定量PCR和蛋白电泳技术检测64例先天性巨结肠狭窄段和扩张段中ZEB2和PTEN mRNA及蛋白表达情况。采用Pearson相关性检验分析ZEB2和PTEN在先天性巨结肠狭窄段和扩张段中表达的相关性。在体外细胞SH-SY5Y中应用ZEB2 siRNA干扰技术降低ZEB2表达,检测其对PTEN表达的影响,利用Transwell实验、CCK-8实验和流式细胞仪技术检测ZEB2对细胞迁移、增殖、周期和凋亡功能。结果 ZEB2和PTEN mRNA在先天性巨结肠狭窄段的表达均比扩张段显著增高(ZEB2:1.2 823±0.1 323 vs 0.987 7±0.124 9,P=0.007 3;PTEN:0.113 2±0.010 9 vs 0.045 9±0.005 8,P0.001)。ZEB2和PTEN在先天性巨结肠狭窄段和扩张段组织中蛋白表达与mRNA表达一致(ZEB2:0.709±0.035 vs 0.531±0.027,P=0.016 6;PTEN:0.466±0.047 vs 0.234±0.052,P=0.029 3)。ZEB2与PTEN mRNA表达在巨结肠狭窄段(r=0.48,P0.001)和扩张段(r=0.54,P0.001)中均呈显著正相关。干扰ZEB2mRNA表达后,体外细胞SH-SY5Y的PTENmRNA和蛋白表达显著下降,细胞增殖和迁移能力受到显著抑制。结论 ZEB2和PTEN在先天性巨结肠狭窄段中表达显著增加;ZEB2表达增加可能是PTEN通过竞争性内源性RNA机制调控后的继发性的改变。

Functions of ZEB2 and PTEN in Hirschsprung disease

Objective To explore the expressions and regulations of ZEB2 and PTEN in Hirschsprung disease (HSCR). Methods The relative expression levels of mRNAs and proteins of ZEB2 and PTEN were measured in stenotic and expanded segments of 64 HSCR patients by quantitative real-time polymerase chain re-action (PCR)and Western blot.The correlation of ZEB2 and PTEN mRNAs levels was measured by Pearson’ s test.Small RNA interference transfection was used for examining the functions and regulations of ZEB2 and PTEN in SH-SY5 Y cell line.Cell migration,cell proliferation,cell cycle and cell apoptosis of ZEB2 in SH-SY5 Y cells were detected by Transwell assay,CCK8 assay and flow cytometry. Results ZEB2 and PTEN mR-NA (ZEB2∶1 .282 3 ±0.1 32 3 vs 0.987 7 ±0.1 24 9,P=0.007 3;PTEN∶0.1 1 3 2 ±0.01 0 9 vs 0.045 9 ± 0.005 8,P<0.001 )and protein expression levels(ZEB2∶0.709 ±0.035 vs 0.531 ±0.027,P=0.01 6 6;PTEN∶0.466 ±0.047 vs 0.234 ±0.052,P=0.029 3)were significantly elevated in HSCR stenotic segments, and small interfering RNA (siRNA)-mediated knock-down of ZEB2 inhibited cell migration and proliferation without affecting cell apoptosis or cell cycle.Both mRNA and protein of PTEN decreased after a knockdown of ZEB2. Conclusions ZEB2 and PTEN are overexpressed in HSCR.And an up-regulation of ZEB2 may occur through a competitive mechanism of endogenous RNA regulation of PTEN.