Adenovirus-mediated gene transfer of glutamine: Fructose-6-phosphate amidotransferase antagonizes the effects of interluekin-1β on rat chondrocytes

Objective: To determine whether overexpression of glutamine: fructose-6-phosphate amidotransferase (GFAT) in synoviocytes will antagonize the response to interleukin-1β (IL-1β) of chondrocytes and synovial fibroblasts in co-culture. Methods: Synovial fibroblasts from the rat were transduced by an adenovirus carrying the cDNA for GFAT and then co-cultured with rat chondrocytes encapsulated in alginate beads. Following challenge with 1, 5, or 10 ng/ml of IL-1β for 24 h, proteoglycan synthesis by the chondrocytes was determined by incorporation of Na₂³⁵SO₄. Production of nitric oxide (NO) and prostaglandin E₂ (PGE₂) were monitored by assay of conditioned medium from the co-culture. Results: IL-1β treatment of untransduced-synoviocyte/chondrocyte co-cultures resulted in markedly decreased proteoglycan synthesis by the chondrocytes, and increased NO and PGE₂ levels in the culture medium. In contrast, adenovirus-mediated transfer of GFAT in synoviocytes prevented both the decrease in chondrocyte proteoglycan synthesis and increases in NO and PGE₂ provoked by IL-1β. Conclusions: Our study suggests that in a synoviocyte/chondrocyte co-culture system, overexpression of GFAT by synoviocytes significantly inhibits subsequent stimulation by IL-1β in vitro. Since GFAT is the rate limiting enzyme in the synthesis of intracellular glucosamine and its derivatives, these results may open new possibilities for osteoarthritis treatment.