Periodontal Ligament Cell Line Establishment By Adenovirus Mediated HTERT And Its Osteogenic Ability

Objective: 1.Establishment of a periodontal ligament cell line overexpressing human telomerase reverse transcriptase gene by adenovirus method;2.The osteogenic capacity was compared with the periodontal ligament cell lines overexpressing the hTERT gene established by the lentivirus method,to explore the construction of a highly effective and stable periodontal ligament cell line that is more conducive to periodontal regeneration.Methods: 1.PCR was firstly used to amplify the full length of hTERT gene to construct recombinant adenovirus plasmid pAd-pshuttle-cmv-hTERT.Packaging of adenovirus particles and infection of human periodontal ligament cells.2.The expression level of hTERT mRNA was detected by qRT-PCR;3.The osteogenic genes like ALP,OPN,OC,BSP,Runx2,Col?mRNA was detected by qRT-PCR;4.After 21 days of culture of periodontal ligament cells with osteogenic induction medium,alizarin red staining solution at pH 4.1 was stained for 10 minutes;5.And cell proliferation was determined by CCK-8.Results: The adenovirus particles containing hTERT gene were successfully constructed and infected with periodontal ligament cells.The infected cells were similar to normal periodontal ligament cells and showed fibrous growth.The results showed that hTERT and osteogenesis-associated genes were highly expressed in the periodontal ligament cell lines constructed by adenoviruses,and were significantly different from normal cells.Alizarin red staining showed that the periodontal ligament cell line had strong osteogenic differentiation ability.Conclusion: The human periodontal ligament cell line with high efficiency and stable expression of hTERT was successfully established by adenovirus method,which laid the experimental foundation for the treatment of periodontal disease.