Responsiveness of Human Intervertebral Disc Cells in Matrix Synthesis To Adenovirus-Mediated Gene Transfer(IGF-1, TGF-beta1 Encoding Genes) / 대한척추외과학회지

ABSTRACT

BACKGROUND: Genetic modification of cells through gene transfer gains popularity as a sophisticated delivery system in the management of musculoskeletal disease. Anabolic growth factors like IGF-1 BMP-2 OP-1, and TGF-beta1 were candidate for therapeutic purposus for regenerating matrix of intervertebral disc. TGF-beta1, OP-1 and BMP-2 share same pathway i.e. Smad while IGF-1 utilize P13K pathway. Accordingly it is logical to use two cytokine encoding genes which using different signal transduction pathway to upregulate matrix synthesis. PURPOSE: To elucidate the anabolic effect of the combination gene transfer(IGF-1 and TGF-beta1 encoding gene) to human disc cells, cultured in alginate beads. MATERIALS AND METHODS: Lumbar and cervical intervertebral disc tissue was obtained from surgical disc procedure from fifteen patients. After isolation and culture of the cells, cultures were transduced with first, Adenovirus-TGFbeta1 construct(Ad/TGF-beta1) and Ad/IGF-1 respectively, second, with combination of two viral constructs(Ad/TGF-beta1+Ad/IGF-1). Cultures treated with saline and Ad/luciferase served as control. Then cultures were incorporated into alginate beads. Exogenous TGF-beta1(2ng/ml) and IGF-1(100ng/ml) were administered also. Newly synthesized proteoglycan was assessed using S35 incorporation using chromatography on Sephadex G-25 in PD-10 column. RESULTS: In cultures transduced with single therapeutic gene construct, there were statistically significant 2.9 fold(Ad/TGF-beta1 and 1.8 fold(Ad/IGF-1) increase in newly synthesized proteoglycan comparing control(p<0.05). In culture transduced with double combination of therapeutic gene construct, there were 3.9 fold(Ad/TGF-beta1+Ad/IGF-1) increase in newly synthesized proteo-glycan comparing control(p<0.05). Culture treated with TGF-beta1(2ng/ml) showed 3.9 fold increase and IGF-1(100ng/ml) 2.9 fold increase, TGF-beta1+IGF-1 4.2 fold increase in proteoglycan synthesis compared to control. CONCLUSIONS: Combination gene therapy provided efficient mechanism of upregulating matrix regeneration of the human inter-vertebral disc cells.