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目的通过检测肛门直肠畸形(ARM)胎鼠直肠末端神经生长因子受体TrkA、p75NTR mRNA的表达水平,探讨ARM直肠末端肠神经系统发育情况。方法成年SD孕鼠16只。其中实验组10只,对照组6只。实验组孕鼠在孕11 d、13 d 2次给予10 g.L-1乙烯硫脲(125 mg.kg-1)灌胃,制作先天性ARM动物模型;对照组灌胃注入等量9 g.L-1盐水。2组孕鼠在孕20 d时行剖宫手术取出胎鼠,取其直肠末端并保存于-80℃冰箱;采用反转录/实时定量PCR(RT-PCR)方法检测正常胎鼠和ARM胎鼠直肠末端TrkA、p75NTR mRNA表达情况。结果 TrkA、p75NTR mRNA在正常胎鼠直肠末端的相对表达量分别是162.221±104.675,129.778±51.976,在ARM胎鼠直肠末端中表达分别是78.937±33.425,87.145±25.812,两组比较差异均有统计学意义(Pa<0.05);琼脂糖凝胶电泳显示TrkA、p75NTR基因扩增后产物cDNA片段与引物设计的扩增片段完全吻合,TrkA、p75NTR基因条带亮度对照组均高于实验组。结论 TrkA、p75NTR mRNA在ARM胎鼠直肠末端的表达异常,提示神经生长因子及受体TrkA、p75NTR可能参与ARM肠神经系统的发育。
Objective To investigate the expression of TrkA and p75NTR mRNA in the rectum of the rectum of the anorectal malformation (ARM) in rats and to explore the development of the end-rectal neuroendocrine system in the rectum. Methods 16 adult SD pregnant rats. Including 10 experimental group and 6 control group. Pregnant rats in experiment group were given gavage of 10 gL-1 ethylenethiourea (125 mg.kg-1) twice daily on the 11th and 13th day of pregnancy to make congenital animal model of ARM. The control group was given intragastric administration of 9 gL-1 brine. The pregnant rats in 2 groups were removed by cesarean section during the first trimester of pregnancy, and the rectum was taken at the end of the rectum and stored at -80 ℃. The normal fetus and ARM embryos were detected by reverse transcription / real-time PCR (RT-PCR) The expression of TrkA and p75NTR mRNA at the end of mouse rectum. Results The relative expression levels of TrkA and p75NTR mRNA in the rectum of normal fetuses were 162.221 ± 104.675 and 129.778 ± 51.976, respectively, and 78.937 ± 33.425 and 87.145 ± 25.812 respectively in the rectum of the fetus of ARM. The differences between the two groups were statistically significant (P <0.05). The results of agarose gel electrophoresis showed that the cDNA fragments amplified by TrkA and p75NTR genes were completely consistent with those of primers. The brightness of TrkA and p75NTR gene bands in control group were higher than those in experimental group. Conclusion The abnormal expression of TrkA and p75NTR mRNA in the rectum of ARM fetus suggests that NGF and TrkA and p75NTR may be involved in the development of the ARM enteric nervous system.