Effect of double gene recombinant vector in inhibiting adipogenic and promoting osteogenic gene expression in rabbit model of ethanol-induced osteonecrosis of the femoral head

Objective To investigate the effect of double gene recombinant vector in inhibiting adipogenic and promoting osteogenic gene expression in rabbit model of ethanol-induced osteonecrosis of the femoral head. Methods Take 80 healthy rabbits, and randomly divided into 5 groups: (1) double gene group: double gene recombinant vector transfected bone marrow mesenchymal stem cells (BMSCs) injected into one side femoral head of the rabbits and 10 ml/(kg·d) of alcohol gavage after horse serum sensitization. (2) unrelated sequence group: rabbits were treated with unrelated sequence transfected BMSCs and 10 ml/(kg·d) of alcohol gavage after horse serum sensitization. (3) model group: rabbits were treated with 10 ml/(kg·d) of alcohol gavage after horse serum sensitization. (4) sensitization group: rabbits were treated with horse serum sensitization only. (5) normal groups: rabbits with no treatment served as controls. The expression of the peroxisome proliferator-activated receptor-γ (PPARγ), CGRP, steogenesis-associated genes 2 (Runx2), osteocalcin mRNA and protein were determined through TaqMan real-time quantitative polymerase chain reaction (Real-time PCR) and Western blotting at 4w and 8w of the experiment. Results At 4 weeks, the expression levels of PPARγ mRNA and protein in double gene group (0.502±0.054 and 0.302±0.035) were similar to the normal group (0.486±0.054 and 0.315±0.041), with no significant differences observed (P>0.05), and were significantly lower than that in model group and unrelated sequence group (0.821±0.092 and 0.703±0.081), (0.875±0.103 and 0.664±0.077), with the difference statistically significant (P<0.05). The expression levels of CGRP mRNA and protein were high (0.772±0.109 and 0.608±0.059), significantly higher than that in groups normal, model and unrelated sequence (0.467±0.062 and 0.296±0.027), (0.118±0.015 and 0.113±0.010), (0.123±0.016 and 0.122±0.013), and the difference was statistically significant (P<0.05). The expression levels of Runx2 and osteocalcin mRNA and protein in double gene group were high (0.733±0.092 and 0.613±0.069), (0.628±0.073 and 0.673±0.081), and significantly higher than that in groups normal, model and unrelated sequence (0.436±0.057 and 0.325±0.036), (0.367±0.041 and 0.374±0.046), (0.133±0.014 and 0.096±0.012), (0.096±0.010 and 0.126±0.015), (0.113±0.016 and 0.104±0.011), (0.087±0.011 and 0.132±0.015), and the difference was statistically significant (P<0.05). At 8 weeks, the expressions in all groups were consistent with that at the 4 weeks. Conclusion The two-gene recombinant vector can effectively target the expression of adipogenic gene in the femoral head cells of ethanol-induced ONFH model of rabbits in vivo, and promote the expression of osteogenic gene in the cells at the same time. Which is significantly better than that of single gene effect. Key words: Double gene recombinant vector; Ethanol; Stem cells; Gene expression; Rabbit