A Study Of The Association Of SOX2,SOX11,β-catenin And TCF/LEF Family With Hirschsprung’s Disease

Objective: To study the expression of SOX2,SOX11,β-catenin and TCF / LEF family in the enteric plexus of Hirschprung’s Disease(HD),and explore their relationship with the development of Enteric nervous system and the pathogenesis of HD.Methods: SP immunohistochemistry was used to detect the expression of SOX2 and SOX11 protein in the resected intestine of proximal margin,transitional segment and narrow segment in 50 patients with HD;RT-qPCR was used to detect the expression of SOX2 and SOX11 mRNA in the proximal margin and narrow segment of 20 HD cases;Also,the above 50 cases of HD resection of bowel were stained with hematoxylin-eosin staining,and micro measurement equipment was applied to gauge their development state of myenteric plexus.SP immunohistochemistry was used to detect the expression of β-catenin,TCF / LEF family protein in the resected intestine of proximal margin and narrow segment in 44 patients with HD;RT-qPCR was used to detect the expression of β-catenin,TCF1 and TCF4 mRNA in the proximal margin and narrow segment of 19 HD cases;And the above 44 cases of HD resection of bowel were stained with hematoxylin-eosin staining,then micro measurement equipment was applied to gauge their development condition of myenteric plexus.Results:(1)In proximal margin,transitional segment and narrow segment of 44 patients with HD,the average optical density of SOX2 protein in myenteric plexus were 0.1325 ± 0.0469,0.0538 ± 0.0143 and 0.0244 ± 0.0122 respectively,(P <0.001);The average optical density of SOX11 protein in myenteric plexus were 0.0696 ± 0.0187,0.0304 ± 0.0106 and 0.0054 ± 0.0044 respectively,(P <0.001).The results of RT-qPCR showed that the expression of SOX2 mRNA in the proximal margin was 3.05 times of that in the narrow segment(P <0.001);The expression of SOX11 mRNA in the proximal margin was 2.79 times of that in the narrow segment(P <0.001).In 50 cases,the microscopic measurement results showed that the myenteric plexus diameter of proximal margin,transitional segment and narrow segment were 57.42 ± 11.42 um,33.16 ± 4.76 um and 30,53 ± 6.50 um respectively,(p <0.001);The number of ganglion cells in the single microscope field(×400)of myenteric plexus of proximal margin and transitional segment were 5.53±1.45 and 1.97±0.80,(p <0.001);The ratio of cytoplasm to nucleus of ganglion cells in myenteric plexus of proximal margin and transitional segment were 2.18 ± 0.20 and 1.63 ± 0.14,(p<0.05);In proximal margijn and transitional segment,the average diameter of ganglion cells in myenteric plexus were 16.09 ± 1.75 um and 11.91 ± 1.30 um,(p<0.05).(2)The expression of β-catenin,TCF1 and TCF4 protein in myenteric plexus of proximal margin were significantly higher than that in the myenteric plexus of narrow segment in 44 patients with HD(P < 0.001,P <0.001,P <0.001).However,LEF1 and TCF3 protein had no positive expression in myenteric plexus of proximal margin and narrow segment in 44 patients with HD.The result of RT-qPCR showed that the expression of β-catenin mRNA in the proximal margin was 1.39 times of that in the narrow segment(P <0.05);The expression of TCF1 mRNA in the proximal margin was 1.96 times of that in the narrow segment(P <0.001);And the expression of TCF4 mRNA in the proximal margin was 1.62 times of that in the narrow segment(P <0.001).In 44 cases,the microscopic measurement results showed that the myenteric plexus diameter of proximal margin and narrow segment were 57.44±11.95 um and 30.71±6.64 um respectively,(p <0.001);The number of ganglion cells in the single microscope field(×400)of myenteric plexus of proximal margin was 5.60±1.38.Conclusion:(1)The expression levels of SOX2 and SOX11 in glial cells or ganglion cells of the enteric plexus may affect the development of enteric plexus and ganglion cells,and thus participate in the pathogenesis of HD.(2)In the Wnt canonical pathway,the expression of β-catenin,TCF1 and TCF4 in the resected bowel segment may be related to the development of ENS and the pathogenesis of HD.