Transfection of stem cells derived from rat dental pulp with green fluorescent protein infection by lentiviral vector

doi:10.3969/j.issn.2095-4344.2014.45.016

Abstract

Abstract:

BACKGROUND: Stable and efficient labeling of dental pulp stem cells in vitro is most important in tracer technique, which is also the basis of tooth regeneration in vivo.

OBJECTIVE: To determine the optimal condition and method for transfection of stem cells derived from rat dental pulp with green fluorescent protein infection by lentiviral vector and to determine whether green fluorescent protein-labeled dental pulp stem cells maintain their stem cell properties.

METHODS: Rat dental pulp stem cells were obtained by modified enzyme digestion method, to identify the immune phenotype and differentiation potential. Dental pulp stem cells were infected with green fluorescent protein by lentiviral vector for 24 and 48 hours at different multiplicity of infection (MOI) (5, 10, 25, 50 and 100). The infection efficiency and fluorescence intensity were analyzed by inverted fluorescent microscopy. The clonal and proliferation ability, cell cycle and the mineralization potential were compared before and after transfection. Based on those mentioned above, we could evaluate the influence of infection on their biological characteristics.

RESULTS AND CONCLUSION: Flow cytometry results showed that rat dental pulp stem cells expressed STRO-1 and CD146 but not CD34 or CD45. The dental pulp stem cells could differentiate into osteoblasts and adipocytes when cultured in specific medium for each lineage differentiation. The highest efficiency of infection and strongest fluorescence expression appeared at 48 hours of infection and MOI 50. There were no significant differences in growth ability, cell colony formation rate and cell cycle before and after transfection (P > 0.05). And the alkaline phosphatase expressed positively. Infection for 48 hours at MOI 50 is optimal for transfecting dental pulp stem cells with green fluorescent protein by a lentiviral vector, thereby providing reliable tracer method for the study of rat dental pulp stem cells in vivo.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

全文链接：

Key words: stem cells, dental pulp, lentivirus infections, green fluorescent proteins

CLC Number:

R394.2

Zhang Rui-han, Liu Jia, Nie Shan-shan, Wang Xuan, Li Bo-qi, Sun Da-lei, Refukati Dilimaolati, Liu Yi-shan. Transfection of stem cells derived from rat dental pulp with green fluorescent protein infection by lentiviral vector[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(45): 7299-7305.

Figures/Tables 9

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