先天性肛门直肠畸形支配盆底肌的脊髓神经元定量研究

Quantitative analysis of motoneurons innervating the muscle levator ani in rats with anorectal malformation

目的 探讨先天性肛门直肠畸形支配盆底肌肉脊髓运动神经元的发育情况。方法 将胎儿外科与显微注射技术相结合 ,对乙烯硫脲致畸的先天性无肛畸形胎鼠的支配肛提肌 (相当于人的耻骨直肠肌 )脊髓神经元进行荧光金 (FG)神经逆向示踪研究 ,并对脊髓连续切片定量计数标记神经元数量。根据畸形类型分为肛门直肠畸形组、神经管畸形组和并发肛门直肠与神经管畸形组 ,并与正常组进行比较。结果 FG标记支配肛提肌的脊髓神经元总数在正常组为 135± 2 9个 ,先天性肛门直肠畸形组为 5 5± 2 6个 ,先天性神经管畸形组为 4 8± 30个 ,合并先天性肛门直肠与神经管联合畸形组为 5 4± 2 9个。结论 先天性肛门直肠畸形和神经管畸形鼠均存在脊髓神经元发育异常。

Objective To quantify the motorneurons innervating the muscle levator ani (rat equivalent of puborectalis in human) by retrograde tract-tracing in a rat model of anorectal malformation. Methods Anorectal malformation is induced in fetuses by gavage feeding of ethylenethiourea (ETU; 125 mg/kg body weigh) to pregnant outbreed Wistar rats at day 11 of gestation. At E20, fetal surgery was performed to expose the muscle levator ani and fluorogold (FG) was injected. Fetuses were collected 24 hours later, fixed by paraformaldehyde infusion cardially. The lumbosacral spinal cord was removed for cryosection. Serial longitudinal sections (60 μm) encompassing the whole width of lumbosacral spinal cord were studied with fluorescent microscopy. The number of FG-labelled motoneurons were scored and compared between fetuses with or without malformation in the ETU-injection group, and normal controls. Results In the ETU-injection group, 55.6% (95/171) of fetuses displayed anorectal malformation, 56.7% of fetuses displayed anomalies of neural tube. FG was injected to the muscle levator ani of 8 fetuses without defects of anorectum or neural tube, 5 fetuses with imperforate anus only, 10 fetuses with anomalies of neural tube, and 15 fetuses with combined imperforate anus and anomalies of neural tube. Fetuses (n=9) from non-injected with ETU group were injected with FG as normal controls. The total number of FG-labelled motoneurons in the fetuses without defects of anorectum and neural tube, with imperforate anus, with anomalies of neural tube, with combined imperforate anus and anomalies of neural tube, and normal controls were 109±38, 55±26, 48±30, 54±29 and 135±29 respectively. FG-labelled motoneurons in the fetuses with imperforate anus, anomalies of neural tube, and combined imperforate anus and anomalies of neural tube are significantly fewer than that in fetuses without defects of anorectum or neural tube(P<005), and in normal controls (P<0.005). The number of FG-labelled motoneurons did not differ between the fetuses without defects of anorectum or neural tube and normal controls (P>0.05). Conclusion Fewer motoneurons that innervates the puborectalis may contribute to the poor postoperative anorectal function in patients with anorectal malformation.