Method: pulp cells cultures from impacted third molars were established and maintained in standard medium. Primary cells in passage 3 were transfected with SV40 (ATCC) using Lipofectramine 2000 Transfection Reagent (Invitrogen Life Technologies). Detection of transformation by immunohistochemistry was done to confirm virus presence in transfected pulp cells nuclei. Cells were incubated with a primary monoclonal mouse antibody anti-SV40 T-Ag (BD Pharmingen). After immortalization success the experiments were performed to characterize the immortalized cells compared to primary cells. Cell viability of primary cells (passage 3) and immortalized cells (passage 10) were analyzed by MTT assay. Then, the analysis of possible changes in the DNA of primary and immortalized cells was performed by the micronucleus assay. Statistical analysis used ANOVA and Tukey test (5%).
Result: it was observed that primary cells showed higher cell viability at 24 h, 4 and 6 days in culture (p <0.05) compare to immortalized cells. At 48h both cell cultures presented similar cell viability. There were no statistical differences between micronucleus formation of primary and immortalized cells (p> 0.05).
Conclusion: it is concluded that primary and immortalized cells presented different viability pattern, and, for each cell line, the best time of culture must be respect. Micronuclei data showed that immortalization, inserting viral DNA in pulp-derived cell, did not increase genotypic sensibility to genotoxic agent.