Myogenic Pathogenesis Of Hirschsprung Disease

Hirschsprung disease,also called HD,is the most common disease involving the deadly obstruction of the colon of children,with an incidence of 1/3000 to 1/5000.This disorder is markedly characterized by a narrow distal segment,a funnel-shaped dilatation at the transition zone,and a dilated proximal colon.It is generally considered that the absence of ganglions caused by the failure migration of neural crest cells was the mechanism of this disease.However,this mechanism of aganglions cannot well explain the clinical phenomenon with contradictions.For instance,the aganglion in colon can lead to weakened contraction rather than spastic contraction in colon.According to our former researches,we consider that the myogenic changed contraction,as a new mechanism,may also play a significant role in HD.To confirm our hypothesis,we first analyzed the contractile characteristics of smooth muscle in HD.We selected the samples of dilated and narrow colon from HD patients respectively,and then compared the contractile properties among different smooth muscle layers.The circular smooth muscle from narrow part displayed hyper-responsiveness to KCl-induced depolarization,but displayed hypo-responsiveness,even non-responsiveness to acetylcholine.Interestingly,both the longitudinal smooth muscles and the circular smooth muscles of the proximal dilated segments displayed normal responses.Meanwhile,to exclude the interference of neuron cells,we separated the single cells of smooth muscle from circular smooth muscle,and detected the responsiveness to ACh.The results showed that the contractile property of single cells was consistent with that of muscle strips.Therefore,we speculated that the changed contractile property of smooth muscle was a myogenic reason for HD development.To prove the myogenic mechanism,we did a series of experiments.First,we analyzed the signaling pathway of smooth muscle contraction in HD,and found a sustained enhanced phosphosrylation of RLC(RLCp)in circular narrow smooth muscle;meanwhile,the smooth muscle tonic-like contraction could not be relaxed when the RhoA/ROCK/MYPT1 signaling pathway was inhibited.It was supposed that the abnormal contraction was caused by the enhanced Ca2+ sensitivity.According to these results,we detected the protein levels in the Ca2+ sensitivity signaling pathway.Although the expressions of ROCK,PKC,RLC and RhoA had no changed in circular narrow,the expression of MYPT1 and PPlc was significant decreased and even lost in some samples.Moreover,we found the MYPT1SMKO mice displayed the similar phenotype with the HD,including hyper-responsiveness to KCl-induced depolarization and hypo-responsiveness to acetylcholine,the decrease of ganglion and so on.Thus,we considered the decreased expression of MYPT1 was the myogenic mechanism of HD.MYPT1 expression can be regulated by multiple of factors.We have proved that LPS and some microbial metabolites can alter the expression of MYPT1.Therefore,we concluded that the infection of newborn may be one of crucial reasons for HD.To relieve the sustained contraction of smooth muscle and improve the obstructive phenotype of spastic contraction in HD,we tested a series of drugs that potentially relax the smooth muscle contractions.We found that NO and Nifedipine(an inhibitor of L-type channel)can relax the tonic-like contractions of circular narrow colon.Therefore,we suppose the enemas by Nifedipine can release the symptom of HD;meanwhile,we also consider that we may minimize the extent of surgery,maybe by only cutting off the circular smooth muscle of narrow part to research the purpose of treatment.In conclusion,our study revealed that the aberrant expression of MYPT1 could change the contractile properties of smooth muscle,suggesting a myogenic mechanism of HD,and providing a possible improved treatment.