This study investigates the oxidative effects of tumour necrosis factor alpha (TNF) and glucose (G) in a cell culture model of osteoblasts and the modifying role of minocycline (M), using a steroid marker of wound healing (5 alpha- dihydrotestosterone: DHT). This model simulates chronic inflammation and hyperglycaemia, relevant to chronic inflammatory periodontal disease in uncontrolled diabetics and prospects of adjunctive management, using minocycline.
Monolayer cultures of MG63 human osteoblasts were established in Eagles MEM in multiwell plates. They were incubated with 14C- testosterone in the presence or absence of optimal concentrations of TNF20 ng, G1000, M25 micrograms/ml, alone and in combination for 24 h in a humidified tissue culture incubator. The medium was then eluted, solvent extracted for formed metabolites and separated by thin layer chromatography in a benzene acetone solvent system 4:1 v/v. The metabolites were then quantified using a radioisotope scanner.
The metabolites formed were diols, DHT and 4-androstenedione. There were 63% reductions in the yields of DHT in response to TNF and G alone, while M stimulated yields by 71% compared with controls (n=8; P<0.0001). The combination of TNF+G resulted in 27% less DHT than either alone. When M was added to this combination, there was a 3-fold increase in the yield of DHT, over TNF+G (n=8; P<0.0001). Yields of 4-androstenedione were inversely proportional to those of DHT, less so with the combination of TNF+G. The yields of diols were significantly decreased in response to agents compared with controls, again less so with the combination TNF+G.
TNF and G induced oxidative responses in cultured osteoblasts which were overcome by minocycline in this hyperglycaemic model. This can be extrapolated to the in vivo situation, reinforcing therapeutic efficacy of minocycline for adjunctive management of uncontrolled periodontal disease in diabetic patients, in keeping with previously reported antioxidant properties of minocycline.
06 - 07 Nov 2006
Society for Endocrinology