Effect Of Notch-Pax9 Signaling Pathway In Ethanol-associated Oral Cancer

Oral cancer is the sixth most common malignancies.In recent years,the incidence of upper gastrointestinal malignancies,including oral cancer in developing countries is on the rise,especially in young men.Meanwhile,many epidemiologicl studies have consistently shown that alcohol drinking is an etiological factor of human oral cancer.The risk and mortality of oral cancer are associated with alcohol drinking in dose dependent.But until now,molecular mechanisms of ethanol-associated oral squamous cell carcinomas are still not well understood.In this study,firstly we found that the expression of Pax9 and Notch target gene were decreased in samples from human oral cancer and esophagus cancer,and there was a same trend in vivo and in vitro expreiments.Secondly,Pax9 was likely to be the downstream of Notch signaling pathway.Thirdly,Notch-Pax9 signaling pathway may play a role in pathogenesis of oral cancer.1.The expression of Pax9 and Notch pathway in alcohol associatd oral cancerImmunohistochemical staining was applied to detect Pax9 expression in tissue array of oral squamous cell carcinomas(OSCC)and esophageal squamous cell carcinoma(ESCC).Through testing the samples that collected from Duke University by immunohistochemical staining,the expressions of Pax9 and Notch signaling pathway were compared between normal tissue and ESCC.Based on 4 microarray data sets,we analyzed the expressions of Pax9 and Notch signaling pathway in normal tissue and OSCC/ESCC.All the experimental results showed that compared with matched normal tissue,Pax9 expression was decreased or missing in OSCC and ESCC,as well as the downstream effector of Notch,Hes1.And compared with non-drinkers,Pax9 expression in ESCC of drinkers was further decreased.In vitro study,KYSE510 cells were treated with different concentrations of alcohol for 72 hr.The results showed that alcohol inhibited Pax9 and Notch signaling pathway in concentration-dependent manner.2.The relationship between Pax9 and Notch signaling pathwayChIP test was applied to explore the relationship between Pax9 and Notch signaling pathway in mice tongue,esophagus and stomach.We established RBPJ knockout mice and found down-regulation of Pax9 expression in squamous epithelial cells of tongue,esophagus and stomach.The findings showed that Pax9 is likely to be downstream of Notch signaling pathway.KYSE510 cells were treated with DBZ(Notch inhibitor)or Jagged1(Notch activator).The results showed that the alteration of Pax9 expression was consistent with the target gene of Notch signaling pathway.3.The effect of Pax9 in NMBA-induced SCCThe activity of cell proliferation was detected in Pax9 knockout mice model by the fluorescent immunohistochemical staining and immunohistochemical staining.The results showed that Pax9-knockout promoted proliferation and inhibited differenciation of esophageal epithelial cells.Pax9 knockout mice were treated with NMBA.Pax9 deficiency promoted NMBA-induced tumors in mouse forestomach and tongue.In conclusion,our data suggest Pax9 is likely to be the downstream gene of Notch signaling pathway,and Notch-Pax9 inhibition may be involved in the development of alcohol associated oral cancer.