Suzuki Ryota, Miyahara Katsumi, Murakami Hiroshi, Doi Takashi, Lane Geoffrey J, Mabuchi Yo, Suzuki Nobuharu, Yamataka Atsuyuki, Akazawa Chihiro
Department of Biochemistry and Biophysics, Graduate School of Health Care Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, 113-8510, Tokyo, Japan.
Pediatr Surg Int. 2014 Feb;30(2):189-95. doi: 10.1007/s00383-013-3452-z.
BACKGROUND/PURPOSE: Despite technical advances in the surgical/medical care of anorectal malformation (ARM), persistent unsatisfactory postoperative bowel habit has been attributed to histopathologic abnormalities of the distal rectum/pouch (DRP) and hypoplasia of anal sphincter muscles (ASM). We used Sox10-Venus mice with ARM induced by all-trans retinoic acid (ATRA) to investigate neural crest cell (NCC) innervation in the DRP and ASM.
Pregnant Sox10-Venus mice were administered single doses of 50, 70, or 100 mg/kg of ATRA on embryonic day 8.5 (E8.5) then sacrificed on either E16.5 or E19.5. Bowel specimens comprising the anorectum were examined using fluorescence microscopy without immunohistochemical staining (FMIS). Anti-PGP9.5 was used to delineate ganglion cells and anti-SMA for smooth muscles.
The appropriate dose of ATRA for inducing ARM was 50 mg/kg. Under FMIS, all ARM embryos (n = 5; all high type; 3 male:2 female) had less NCC innervation with thick Venus-positive nerve fibers in the DRP compared with normal embryos (n = 8); there was abnormal NCC innervation in the DRP and absent ASM in ARM mice.
We are the first to delineate abnormal enteric nervous system innervation in the DRP of ARM mice without using immunohistochemical staining techniques thus allowing specimens to be examined without any distortion.
