SV40Tag基因片段转染SD大鼠牙囊细胞后相关生物学特性初步研究

The preliminary study on biological characteristics of the SD rat dental follicle cells infected with SV40Tag

目的转染猿肾病毒SV40Tag基因片段至SD大鼠牙囊细胞,获取具有无限增殖能力和稳定生物学性状的牙囊细胞用于牙周组织工程的研究。方法利用293细胞包装病毒构建含SV40Tag的逆转录病毒载体,制备重组逆转录病毒并转染至SD大鼠牙囊细胞。以正常牙囊细胞为对照,倒置显微镜下观察细胞形态及活力,分析细胞端粒酶活性,并检测其成骨分化及增殖特性。结果转染SV40Tag基因后,SD大鼠牙囊细胞传代至60代,生长依然旺盛,存在较强活力;牙囊细胞端粒酶活性显著增强,与对照组有统计学差异(P=0.033);牙囊细胞成骨分化相关基因(碱性磷酸酶、骨钙素、骨形态发生蛋白、Runx2基因)、促有丝分裂相关基因(碱性成纤维细胞生长因子、胰岛素样生长因子)的电泳条带与对照组均无统计学差异(P>0.05)。结论 SV40Tag基因片段转染SD大鼠牙囊细胞后具有无限传代增殖和抗衰老的潜在能力,其细胞生物学特性相对稳定,可为牙周组织工程提供较理想的种子细胞来源。

Objective To provide reliable seed cells with unlimited passage proliferation and stable biological cha- racteristics for periodontal tissue engineering research through infecting a retrovirus carrying SV40Tag into SD rat den- tal follicle cells. Methods Retroviral virus vector containing SV40Tag by 293 cells was packaged and infected into SD rat dental follicle cells. Normal dental capsule cells were used as control group. The cell morphology and vitality were observed by inverted microscope, telomerase activity, osteogenic differentiation and proliferation of dental follicle cells were analyzed. Results The SD rat dental follicle cells infected with SV40Tag could be passaged for 60 gene- rations in vitro culture with strong growth activity. The telomerase activity was significantly enhanced compared with the control group （P--0.033）. The expression of alkaline phosphatase, osteocalcin, bone morphogenesis protein-2, Runx2, basic fibroblast growth factor, insulin-like growth factors had no statistical difference compared with control cells （P〉 0.05）. Conclusion The SD rat dental follicle cells infected with SV40Tag not only have a strong growth activity and infinite passaged capacity, but also have a stable biological property as normal dental follicle cells, so it can be re- garded as the excellent seed ceils in periodontal tissue engineering.