The Dynamic Changes Of Hedgehog Signaling Pathway Molecules During Murine HCC Carcinogenesis And Development

Objective: To explore the expression and changes of Hedgehog signalingpathway members Shh, Ptch and Smo in the chemical induced live cancer inmice at various stages and preliminary explore Hedgehog signaling pathwayregulatory mechanism.Methods:95male C57BL/6J mice was selected according with a standard,and divided into chemical-induced group(50mice) and normal control group(45mice). Then the normal control group mice was fed with the convention standardand the other group mice was induced HCC by DEN/CCl4/Ethanol. To surveythe mice growth conditions. Since the4th week, both two group mice was killedto get its liver tissue specimens, and repeated this operation every two weeks toobserve the change of pathology by HE stain. At the same time, the protein andmRNA dynamic changes of Hedgehog signaling pathway members Shh, Ptchand Smo expression was respectively detected by immunohistochemical method,Western blotting method and real-time fluorescent quantitative PCR method.Result: HE stain showned the induced carcinoma group mice had beenexperienced a series of changes, such as from the liver cell degeneration necrosis, fibrosis, atypical hyperplasia to the formation of hepatocellularcarcinoma. So the chemical-induced experiment model had been successfulinduced after20weeks. Immunohistochemical showned in the chemical-inducedgroup, the expression of Shh protein was weakly positive at the6th week,moderately positive at the16th week and strongly positive at the20th week;Ptch protein expression was detected weakly positive, moderately positive andstrongly positive respectively at the8th week, the16th week and the20th week;And Smo protein expression was discovered weakly, moderately and stronglypositive respectively at the10th week, the16th week and the20th week. Whilethe normal control group was not monitored by immunohistochemical method.Western blotting result showed that the Shh, Ptch and Smo protein expressionwas found weakly positive stripe differently from the6th week, the8th weekand the10th week in the chemical-induced group. As the extension of time,three proteins expression had increased and the stripe had broadened gradually.At the20th week,all proteins expression enhanced significantly,the proteinexpression quantity achieved the most. While the three proteins expression ofthe normal control group was not detected by Western blotting. The result ofreal-time fluorescent quantitative PCR showed that the differences of ShhmRNA, Ptch mRNA and Smo mRNA expression at the same gene normalcontrol group was not statistical significance from the4th week to the20th week(P>0.05); And at the same gene of the chemical-induced group, the expressionlevel of three continued to rise with time progressed, the expression of ShhmRNA, Ptch mRNA and Smo mRNA at later stages was higher than at earlierstages, and at the20th week all of three expression had reached the highest level,such as Shh mRNA was15.986±1.784, Ptch mRNA was11.272±0.295, SmomRNA was10.185±0.716; Furthermore Shh mRNA, Ptch mRNA and Smo mRNA expression in the chemical-induced group was obviously higher than inthe same period normal control group since from the6th, the6th and the8thweek, this difference had statistically significant (P<0.05).Conclusion: Shh, Ptch and Smo was the important part of the Hedgehogsignaling pathway, the significant change of all the expression quantity duringthe inducing cancer process caused the disorder of the mice liver tissue internalenvironment. The Hedgehog signaling pathway had been activation, and led tothe formation of the mice hepatocellular carcinoma. Hedgehog signalingpathway gene Shh, Ptch and Smo expression level had been found its increasedgradually with the extension of time, to involve in the development andprogression of liver cancer. So Hedgehog signaling pathway could be consideredto play a regulatory role in the occurrence and development of the hepatocellularcarcinoma.