| Objective:1.To explore the effect of alendronate on osteoblast proliferation and intracellular alkaline phosphatase activity in vitro.2.To explore the effect of alendronate on bone metabolism in vivo and provide a reference for clinical treatment.Methods:1.Effect of alendronate on the proliferation of osteoblasts:using the thiazole blue colorimetric test（MTT）,after inducing the generation of osteoblasts in the culture plate,observe its growth,and then use different concentrations of alendronate for osteoblasts,the enzyme-linked immunoassay detector measures its absorbance value.2.Effect of alendronate on the activity of osteoblasts to synthesize alkaline phosphatase:after inducing osteoblast formation in culture flasks,and then use different concentrations of alendronate for osteoblasts,using p-nitrophenyl phosphate the disodium method was used to determine the alkaline phosphatase activity of cells.3.Effect of alendronate on bone metabolism in ovariectomized SD rats:6-month-old SD rats will be treated with ovariectomized treatment and treated with alendronate.At three time points of ovariectomy 4W,8W,and 12W,peripheral blood was collected from rats in the period of rapid bone loss.After centrifugation,the serum OPG and RANKL were detected by ELISA,and then the ratio was calculated.Next,the dual energy X-ray absorptiometry（DXA）was used to measure bone density of the hip,lumbar spine L2-4,femoral neck area,and femoral trochanter at three time points of alendronate treatment 0,26 and 52 weeks.Results:1.Most osteoblasts adhere to the wall.At 24h,the shapes are mostly triangular,polygonal,elongated and irregular.With the prolongation of the experimental time,the density of osteoblasts gradually increased,becoming long spindle-shaped,straight-tempered and fused into pieces.When cultured for 72 hours,rat osteoblasts gradually changed from polygonal,triangular and elongated shapes to oval and cobblestone shapes with unclear boundaries.2.MTT test results showed that alendronate(1.0×10-5mol/L and 1.0×10-4mol/L)with a certain concentration could inhibit the proliferation of osteoblasts at 24h,36h and 48h.On the contrary,a certain concentration of alendronate(1.0×10-8mol/L,1.0×10-7mol/L,1.0×10-6mol/L)could promote the proliferation of osteoblasts,and1.0×10-8mol/L alendronate has the strongest effect on promoting the proliferation of osteoblasts.In addition,alendronate at concentrations of(1.0×10-9mol/L,1.0×10-10mol/L)had no obvious effect on the proliferation promotion of osteoblasts.3.After different concentrations of alendronate were used to treat osteoblasts for72 hours,their effects on osteoblast alkaline phosphatase activity were analyzed and compared.The results showed that the activity of alkaline phosphatase in alendronate at a concentration of 1.0×10-5 mol/L and 1.0×10-6 mol/L in rat osteoblasts increased.4.Compared with 4 weeks of treatment,treatment with alendronate for 12 weeks could significantly increase the OPG and OPG/RANKL ratio after treatment.5.Alendronate could significantly increase the bone density of ovariectomized SD rats.Conclusion:1.Different concentrations of alendronate have different effects on osteoblast proliferation and intracellular alkaline phosphatase activity.2.Alendronate could promote the osteogenic differentiation and increase the bone density of rats. |
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