Shh/BMP4Involved In Enteric Nervous System Development In Anorectal Malformations

Objective1To study ENS in the rectal end with different types of ARTA-induced fetal rats withARM, we performed immunohistochemistry for NSE neurotransmitters and BMP4in theterminal bowel, explore the possible pathological mechanisms of postoperative defecationdisorders.2By observing the expression of Shh/BMP4signaling pathway and to explore the possibleregulatory mechanism about the development of ENS in anorectal malformations.Material and Methods1Animal modle: The timed-pregants rats were divided into two groups(the ATRA treatedgroup and the control group)in random. The ATRA treated group females (n=10)were gavagefed a single dose of100mg/kg of ARTA on embryonic day10(E10), the control group(n=6)received the same volume of olive oil on the same day.2NSE and BMP4immunohistochemistry staining: Embryos were harvested via caesareansection on E20, then embryos divided into normal(N) group, low ARM (l) group and highARM(H) group.15embryos of each group were fixed in4%formaldehyde for24hours,then theembryos were dehydrated, embedded in paraffin and sectioned serially sagittally at4Pmthickness, and then stained immunohistochemically.3RT-PCR: The full thickness of the terminal rectum of residuary embryos on E20wasdissected free of surrounding tissues undermagnification and immediately frozen in liquidnitrogen. RT-PCR analyses the quantify expression of Shh/BMP4mRNA in teminal rectum. Theexpression of Shh/BMP4mRNA in teminal rectum were studied in the fetal rats withhigh(n=12) èlow(n=12)anorectal malformation and12fetal rats with no anorectal malfomationformed the control group êResults1The expression of NSE and BMP4were mainly located in the myentreic plexus andsubmucous plexus of control group by immunohistochemistry. Statistically significantdifferences were found between groups high, low and control regarding the expression of NSEand BMP4.The expression of NSE and BMP4in the high ARM was obviously lower than thatin the low ARM and control group, and there were markedly differences between low ARM andcontrol group (P<0.05). 2There was positive correlation between Shh and BMP4(P<0.01èr=0.884).Shh and BMP4expression significantly decreased in the terminal rectum of high group compared with thecontrol and low group (P<0.05).There was significant difference between the low group andcontrol group (P<0.05).Conclusions1The developmental extent of ENS in the rectums of different types of ARM wasdiscrepant èwhich related to cleidoic position and the expression of BMP4.2This study demonstrates downregulation of Shh/BMP4in ARM (ATRA-exposed)fetalrats,which may disturb the ENS development of the distal hindgut.