Phenylephrine effects on the morphology of osteoblasts and expression of nicotinamide phosphoribosyltransferase under oxidative stress / 中国组织工程研究

ABSTRACT

BACKGROUND: Phenylephrine has been proved to exert a protective effect on radiant-induced salivary gland and epithelial cell injuries, but its effect on hydrogen peroxide (H2O2)-induced oxidative stress in osteoblasts are not fully understood. OBJECTIVE: To explore the effect of phenylephrine on H2O2-induced oxidative stress in osteoblasts, and to explore the mechanism underlying the regulation by the expression level of nicotinamide phosphoribosyltransferase (Nampt). METHODS: Primary osteoblasts were cultured and randomly divided into four groups blank control group, H2O2group, phenylephrine group, and combination group (0.5 hour pretreatment of 1×10-5mol/L phenylephrine, and then given 300 μmol/L H2O2). The morphology of osteoblasts was observed at different time points. Osteoblasts were collected after 24-hour culture, and total RNA and protein were then extracted to detect the mRNA and protein expression levels of Nampt by RT-PCR and western blot assay, respectively. RESULTS AND CONCLUSION: Compared with the blank control group, reduced osteoblasts and evident cell shrinks were observed in the H2O2group, while the number of osteoblasts significantly increased in the combined group compared with the H2O2group at 12, 24 and 48 hours of culture. RT-PCR results showed that the mRNA level of Nampt in the H2O2group was reduced by 31.23% of that in the blank control group, while the mRNA level of Nampt in the combination group was dramatically increased by 206.20% of that in the H2O2group at 24 hours of culture (both P < 0.05). Furthermore, western blot assay findings revealed that the protein level of Nampt in the H2O2group was reduced by 67.98% of that in the blank control group, while the protein level of Nampt in the combination group was increased by 152.25% of that in the H2O2group at 24 hours of culture (both P < 0.05). Our results indicate that phenylephrine can alleviate the shrink and atrophy of osteoblasts caused by H2O2, thereby exerting protective effect by up-regulating the mRNA and protein levels of Nampt that may be a regulatory gene.