慢病毒介导TGFβ3和TIMP1体外联合感染对人退变椎间盘髓核细胞的影响

EFFECTS OF IN VITRO CO-INFECTION WITH LENTIVIRAL-MEDIATED TGFβ3 AND TIMP1 ON HUMAN DEGENERATIVE NUCLEUS PULPOSUS CELLS

目的探讨慢病毒载体介导的转化生长因子β3（TGFβ3）和基质金属蛋白酶抑制剂1（TIMP1）联合感染对人退变椎间盘髓核细胞的影响。方法应用构建的TGFβ3-P2A-TIMP1慢病毒载体感染人髓核细胞,通过实时荧光定量PCR（Real-time qPCR）技术、Western blot技术检测其对人髓核细胞代谢的影响。结果Real-time qPCR检测结果显示,目的基因感染组的TGFβ3、TIMP1、Ⅱ型胶原及蛋白多糖mRNA表达量均明显高于其他各组（F=16.350~74.378,q=2.010~10.620,P〈0.05）。Western blot检测结果显示,目的基因感染组Ⅱ型胶原、蛋白多糖的蛋白表达量均明显高于对照组（F=50.495~66.110,q=8.061~16.624,P〈0.05）。结论慢病毒载体介导的TGFβ3和TIMP1双基因联合感染人髓核细胞可促进Ⅱ型胶原和蛋白多糖的合成,改善髓核细胞外基质代谢。

Objective To study the effects of in-vitro co-infection with lentivirus-mediated TGFβ3and TIMP1 on human degenerative nucleus pulposus cells（NPCs）. Methods Using constructed TGFβ3-P2A-TIMP1 lentiviral vector to infect human NPCs,its impacts on metabolism of human nucleus pulposus cells were detected by Real-time qPCR and Western blot. Results The Real-time qPCR demonstrated that the expressions of TGFβ3,TIMP1,Collagen Ⅱ and Aggrecan mRNA in positive group（carrying target genes）were higher than that in the other groups（F=16.350-74.378,q=2.010-10.620,P0.05）.Western blot indicated that the protein levels of Collagen Ⅱ and Aggrecan were higher than that in the control groups（F=50.495-66.110,q=8.061-16.624,P0.05）. Conclusion Combined Lenti-TGFβ3-P2A-TIMP1 can promote the synthesis of Collagen Ⅱand Aggrecan and improve the metabolism of extracellular matrix of human nucleus pulposus cells.