Relationship Between The Occurring Of Marek’s Disease And Meq, Pp38, PRNP, SPRN, Akt Gene Expression In Marek’s Disease Tissues

[Objective] In order to investigate the relationship between the expression of meq, pp38, PRNP, SPRN, akt and the pathogenesis of Marek’s disease. This study used MDV-MD5 to established chicken tumor case, through the methods of Real-time PCR to detect the relative expression of meq, pp38, PRNP, SPRN and akt in m RNA levels and analyze the regulation expression in chickens infected MDV. Explore the correlation of these genes expression and MDV infection and MD tumor formation, occurrence and development, in order to provide a theoretical basis of further studying their role in the embryology of cancer.[Methods](1)We used agar diffusion, molecular biology and plaque experiment to identify MDV strain. Establish chicken MD tumor case by artificial attack MDV. After 60 days, autopsy positive infected chickens and control chickens every 30 days, after 150 days, slaughter all chickens. Record infection rates, mortality rates and tumor rates.(2)By pathological histology observation, 150 days infected chickens can be divided into tumor lesions, no tumor lesions and negative control chickens.(3)Randomly selected three chickens from each groups and respectively collected 14 tissue samples from per chicken. Total RNA was extracted from every samples and reverse transcription. Use c DNA as template for Real-time PCR of meq, pp38, PRNP, SPRN, akt and β-actin genes, and analyze the gene relative expression differences in m RNA levels of 14 different tissue samples from three experimental groups.[Results](1)The results of agar diffusion, amplification meq and pp38 by RT-PCR and plaque experiment were all positive.TCID50=10-5.6363/0.1 m L, use MD5 to copy MD tumor case. We began to detect until 30 days, the infection rate was 85.42%. After 60 days, infected chickens began to die. After 90 days, we found the earliest tumor. The tumor incidence was 17.07%, the mortality rate was 24.39%. Tumors usually were found in liver, spleen and kidney.(2)There were many visceral cancer lesions in MD tumor disease chickens, hard and gray nodules of different sizes on surface and section. Microscopic observation found that normal tissue structures were destroyed and there were a large number of lymphocyte infiltration. There were no apparent gross lesions at MD no tumor lesions and negative control chickens.(3)Detect the expression differences in m RNA levels of meq,pp38, PRNP, SPRN and akt gene in 14 tissue samples from three experimental groups. meq gene only expressed in MDV infected chickens and didn’t express in negative control chickens. Except heart and ovary, the expressions of meq gene in the rest 12 tumor lesions were higher than those without lesions; Compared with neoplastic lesions chickens, pp38 gene expression in eyelid, ovary and pancreas were not significant(P > 0.05). The expression of pp38 gene in other 11 tumor lesions were higher than those without tumor lesion chickens; Except muscular stomach and intestine, the expression of PRNP gene in the remaining 12 tumor lesions tissues were higher than non-pathological chickens and contrast chickens, the expression in spleen and kidney were the highest. The expression difference of 14 tissues from no tumor lesion chickens compared with control chickens were not significant(P > 0.05); SPRN gene was not expressed in muscular stomach, and was weakly detected in glandular stomach, intestinal and eyelid. Except skin and sciatic nerve, the expressions of SPRN gene in the rest 8 tumor lesions were higher than control chickens. The SPRN gene expressions of 14 organizations from no neoplastic lesions chickens compared with control chickens were not significant(P > 0.05); The akt expression difference in liver, spleen, lung, kidney, brain and skin of tumor chickens compared to control chickens were significantly different(P < 0.05), The rest 8 tumor lesions tissues compared with control tissues were not significant(P > 0.05). The expressions of akt gene in 14 organizations from no neoplastic lesions chickens compared with control chickens were no significant difference(P > 0.05). In MD tumor tissue, liver, spleen, lung, kidney and skin,meq, pp38, PRNP, akt simultaneously over expression. And there was no significant change in the expression of SPRN in addition to lung.[Conclusions](1)This test successfully established MD tumor case.(2)Through Real-time PCR we found that meq and pp38 which was the marker gene of MDV significantly expressed in MD tumor lesions chickens, and the expression of meq was related to the formation of tumor. The expression of pp38 was nothing to do with the formation of tumor.(3)PRNP gene expression was related to the infection of MDV, PRNP overexpression in tumor tissues, and the differential expression of PRNP gene in various neoplastic tissues was related to the lesion degree. The more severe the tumor changes, the greater the increase of PRNP gene expression.(4)The SPRN gene expression quantities in tissues had nothing to do with the infection of MDV and the formation of MD tumor, and had nothing to do with the expression quantity of PRNP gene.(5)The infection of MDV did not cause the expression quantity increase of akt gene, akt overexpression in tumor tissues.(6)meq, pp38, PRNP, akt in MD tumor tissues over expression at the same time, and the expression of SPRN has no obvious change.