Does Vitamin D3 Prevent the Inhibitory Effect of Vancomycin on Osteoblasts?

Background The utility of vancomycin powder to prevent surgical site infection, mainly in spinal surgery, has been widely examined, and the local administration of vancomycin powder to wounds has been reported to be effective in preventing surgical site infections after spine surgery. However, in vitro studies have shown that high local concentrations of vancomycin may inhibit osteogenesis, although it remains unclear how these high concentrations influence osteoblasts. No candidate drug has been reported to recover cytotoxicity with high concentrations of vancomycin, but we suggest that vitamin D3, which induces osteoblast proliferation, may be administrated concomitantly with vancomycin in these situations. Questions/purposes (1) Does a high concentration of vancomycin reduce viable osteoblast numbers in cell culture compared with controls? (2) Does vitamin D3 administration confer a protective effect on osteoblasts when administered with continuous vancomycin? (3) Does vitamin D3 administration confer a protective effect on osteoblasts when administered with pulsed vancomycin (24 hours of administration)? (4) Does vitamin D3 administration confer alkaline phosphatase, mineralization, and gene expression when administered with pulsed vancomycin? Methods MC3T3-E1 cells were cultured at 37 degrees C in an alpha-minimum essential medium supplemented with 10% fetal bovine serum in a humidified incubator containing 5% CO2. The experimental concentrations of vancomycin (2500 mu g/mL, 5000 mu g/mL, and 7500 mu g/mL) were determined based on previous reports and preliminary experiments. We concomitantly administered vitamin D3 (0.01 nM) to prevent cytotoxicity in osteoblasts, using two different treatments: continuous vancomycin administration (measured at 6 hours, 12 hours, 24 hours, and 72 hours) and pulsed vancomycin for 24 hours (measured at 1 days, 3 days, and 7 days). We analyzed cell numbers and morphologic changes in cells treated with vancomycin or vancomycin plus 0.01 nM vitamin D3. Osteoblast differentiation was assessed with alkaline phosphatase staining, alkaline phosphatase activity, and Alizarin red S staining. Results The number of cells was reduced at 6 hours, 24 hours, 48 hours, and 72 hours in response to continuous vancomycin administration at 7500 mu g/mL (at 72 hours, control 14.6 x 10(4) cells/mL 0.260 x 10(4) cells/mL, vancomycin at 0.917 x 10(4) cells/mL 0.288 x 10(4) cells/mL, mean difference -13.7 x 10(4) cells/mL 0.388 x 10(4) cells/m