ABSTRACT
Objective To investigate the effect of 1,25-(OH)2-vitamin D3 (VD3) or mechanical strain alone and their combined treatment on proliferation and differentiation of pre-osteoblast MC3T3-E1 cells in vitro, as well as gene and protein expression of osteoprotegerin (OPG) and receptor activator of nuclear factor-кB ligand (RANKL) in those cells. Methods MC3T3-E1 cells were treated with 10 nmol/L VD3, intermitted mechanical strain or with a combination of these two factors. Cell proliferation was assessed with flow cytometry, and alkaline phosphatase (ALP) activity was measured using a fluorometric detection kit. The mRNA expression of ALP, runt-related transcriptional factor 2 (Runx2), OPG, and RANKL genes was determined by real-time PCR. The proteins expression of Runx2, OPG, and RANKL was determined by Western blotting. ResultsVD3 inhibited the proliferation of MC3T3-E1 cells, but the mechanical strain had no effect on cell proliferation. Mechanical strain, VD3, and the combined treatment enhanced the ALP activity of MC3T3-E1 cells as well as the protein expression of Runx2. The effect of combined treatment was less pronounced than the effect of VD3 or mechanical strain alone. Mechanical strain promoted the gene and protein expression of osteoprotegerin (OPG) and increased the ratio of OPG/RANKL. However, the combination of VD3 and mechanical strain led to a decrease in ratio of OPG/RANKL. Conclusions Mechanical strain might be effective in inducing osteogenic differentiation and increasing bone formation. A joint stimulation with VD3 and strain can decrease proliferation and osteogenic differentiation and increase RANKL expression, which might affect bone remodeling. This study supplies some new data, which might be important in theoretical and clinical research of osteoporosis (OP) and other related bone diseases.
ABSTRACT
OBJECTIVE: To develop a sensitive and specific reverse-phase high-performance liquid chromatography (RP-HPLC) method to determine propofol in follicular fluid. METHODS: Thymol was added into follicular fluid samples as the internal standard, and the samples were then precipitated with methanol. After centrifugation at 20000 r·min-1 for 15 min, 20 μL of supernatant was directly injected into an Agilent Extend-C18 column (4.6 mm×250 mm, 5 μm). The mobile phase was composed of methanol-water (77:23). Analysis was run at a flow rate of 1.0 mL ·min-1. Fluorescence detection was applied with excitation and emission wavelengths of 276 and 310 nm, respectively. RESULTS: The calibration curves had good linearity in the range of 0.108-5.4 mg·L-1. The linear regression equation was A=0.2074ρ-0.0012(r=0.9993). The limits of detection and quantity proved to be 54 and 108 μg·L-1, respectively. The method recoveries for low, intermediate and high concentrations were 117.9%, 101.7% and 99.4%, respectively. The RSDs of inter-day assay were 4.10%, 1.28% and 0.20%(n=5), and intra-day assay were 4.34%, 2.33% and 0.61%(n=5), respectively. CONCLUSION: The method is accurate and convenient. It can be used to determine propofol for studying its clinical pharmacokinetics.