RESUMEN
Prior studies have shown that implant surface roughness affects osteoblast proliferation, differentiation, matrix synthesis, and local factor production. Further, cell response is modulated by systemic factors, such as 1,25(OH)2D3 and estrogen as well as mechanical forces. Based on the fact that peri-implant bone healing occurs in a site containing elevated amounts of prostaglandin E2 (PGE2), the hypothesis of the current study is that PGE2 and arachidonic acid (AA), the substrate used by cyclooxygenase to form PGE2, influence osteoblast response to implant surface roughness. To test this hypothesis, 4 different types of commercially pure titanium (cpTi) disks with surfaces of varying roughness (smooth Ti, R(a) 0.30 microm; smooth and acid etched Ti [SAE Ti], R(a) 0.40 microm; rough Ti, R(a) 4.3 microm; rough and acid etched Ti [RAE Ti], R(a) 4.15 (microm) were prepared. MG63 osteoblasts were seeded onto the surfaces, cultured to confluence, and then treated for the last 24 hours of culture with AA (0, 0.1, 1, and 10 nM), PGE2 (0, 1, 10, 25, and 100 nM), or the general cyclooxygenase inhibitor indomethacin (0 or 100 nM). At harvest, the effect of treatment on cell proliferation was assessed by measuring cell number and [3H]-thymidine incorporation, and the effect on cell differentiation was determined by measuring alkaline phosphatase (ALP) specific activity. The effect of AA and PGE2 on cell number was somewhat variable but showed a general decrease on plastic and smooth surfaces and an increase on rough surfaces. In contrast, [3H]-thymidine incorporation was uniformly decreased with treatment on all surfaces. ALP demonstrated the most prominent effect of treatment. On smooth surfaces, AA and PGE2 dose-dependently increased ALP, while on rough surfaces, treatment dose-dependently decreased enzyme specific activity. Indomethacin treatment had either no effect or a slightly inhibitory effect on [3H]-thymidine incorporation on all surfaces. In contrast, indomethacin inhibited ALP on smooth surfaces and stimulated ALP on rough. Taken together, the results indicate that both AA and PGE2 influence osteoblast response by promoting osteoblast differentiation on smooth surfaces, while inhibiting it on rough surfaces. Because implants with rough surfaces are acknowledged to be superior to those with smooth surfaces, these results suggest that use of nonsterioidal anti-inflammatory drugs to block PGE2 production and reduce inflammation may be beneficial in the postoperative period after implant placement. They also indicate that manipulation of the AA metabolic pathway may offer a new therapeutic approach for modulating bone healing after implant placement. Because peri-implant healing takes place in a complex cellular environment quite different from the one used in the present study, additional work will be necessary to substantiate these possibilities.
Asunto(s)
Ácido Araquidónico/farmacología , Materiales Dentales/química , Dinoprostona/farmacología , Osteoblastos/efectos de los fármacos , Titanio/química , Grabado Ácido Dental , Fosfatasa Alcalina/análisis , Remodelación Ósea/efectos de los fármacos , Recuento de Células , Diferenciación Celular/efectos de los fármacos , Línea Celular , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Inhibidores de la Ciclooxigenasa/farmacología , ADN/biosíntesis , Relación Dosis-Respuesta a Droga , Humanos , Indometacina/farmacología , Osteoblastos/citología , Radiofármacos , Propiedades de Superficie , Timidina/metabolismo , Factores de Tiempo , TritioRESUMEN
BACKGROUND: Chronic, nonhealing skin wounds are a common ailment in uncontrolled diabetes and are associated with significant morbidity. The nonhealing diabetic foot wound displays pathologically elevated matrix metalloproteinase (MMP) activity. In contrast, the concentration of nitric oxide (NO) is significantly reduced in these chronic ulcers. Addition of NO to diabetic wounds improves wound healing, but the mechanism for this effect is poorly understood. MATERIALS AND METHODS: Diabetic and nondiabetic human skin fibroblasts were cultured to confluence and then treated with 0, 1, 10, and 100 nm concentrations of three NO donors (NOR-3, SNAP, and SNOG) with varying half-lives for 1, 3, and 7 days. At harvest, the cultures were analyzed for their production of NO and the effect of NO donor treatment on cell proliferation (cell number) and MMP expression (MMP-1, -2, -8, -9, and -13). RESULTS: The NO donor with the shortest half-life (NOR-3) produced a rise in NO on day 1 in both normal and diabetic fibroblasts at the highest concentration used; there was a corresponding decrease in both MMP-8 and MMP-9 expression in the diabetic fibroblasts and a decrease in only MMP-9 expression in the normal fibroblasts. After longer times in culture or at lower concentrations, NOR-3 was without effect on NO production or MMP expression. Further, NOR-3 had no effect on cell proliferation. In contrast to NOR-3, NO donors with longer half-lives (SNAP and SNOG) significantly (P < 0.05) increased NO production by both normal and diabetic fibroblasts during the entire course of the experiment and even after a media change lacking additional NO donor at day 3. SNAP and SNOG dose-dependently reduced MMP-8 and -9 mRNA expression in both normal and diabetic fibroblasts through day 7. The expression of MMP-1, -2, and -13 was not significantly affected by any of the NO donor treatments. CONCLUSIONS: These experiments show distinct deficits in NO production and elevations in MMP-8 and -9 expression in diabetic human skin fibroblasts compared to normal. In addition, treating these cultures with NO donor compounds with half-lives greater than 5 h selectively raised NO production by the cells, increased cell proliferation, and decreased MMP-8 and -9 expression in a dose-dependent manner. There was no effect of the NO donor compounds on MMP-1, -2, or -13 expression. One possible mechanism to account for NO's beneficial effect on wound healing may involve stimulation of cell proliferation and down-regulation of MMP expression.