Expression Of BMP-2,Smad1/5 And Smad6 In The Femur Of Osteoporosis Rats Treated By Tonifying Kidney And Strengthening Bone Method

ObjectiveWith the aging of the global population,the number of patients with osteoporosis is increasing,which brings heavy pressure and burden to society and families.Therefore,it is necessary to strengthen the research on osteoporosis and develop effective prevention measures as soon as possible.The purpose of this study is to explore the mechanisms of bushenzhuanggufang on osteoporosis,in order to provide experimental data and clinical basis for prevention and treatment of OP in clinic.Methods60 female 12–month-old SD rats with SPF grade,fed 7 days adaptively,were randomly divided into sham operation group,model group,bushenzhuanggufang high,middle,low-dose group and estradiol group,with 10 in each group.Bilateral ovaries were removed by surgery except sham operation group.7 days after the operation,the rats of treatment group gave proper drugs according to bushenzhuanggufang and estradiol suspension for 12 weeks,while sham operation group and model group gave the same dose of ddH₂O for 12 weeks.The common condition of rats were observed?spirit,weight,etc?for 12 weeks,and the BMD in left femur was detected by bone densitometer.The morphology of bone and uterine were tested by HE staining.The content of Serum Ca was detected using micro-plate method,serum P using phosphomolybdate method,and serum E₂ by Elisa.The expressions of BMP-2,Smad1/5 and Smad6 protein in bones were detected by immunohistochemical SABC method.Results1.The rats in model group had symptoms like a relatively weak spirit,slow reaction,sleepiness,less movement after bilateral ovariectomy.They were in good spirit,had hair flush and reaction agility after drug administration.2.BMD of the left lower extremities in rats:the BMD in the model group decreased significantly compared with sham group.BMD in high and middle dose group and estradiol group increased significantly after the drug administration in treatment groups.HE staining showed that the cavity of the bone marrow was smaller than that in the model group,and the number of bone trabeculae was increased.The arrangement was more homogeneous and the structure was closer than those in the model group.3.Uterus index of rats:the uterus coefficient in the model group reduced significantly compared with the sham group.The uterus index in the high and middle dose group and the estradiol group increased compared with the model group.Uterine coefficient in low dose group increased significantly.HE staining in uterus showed that endometrium is slightly thicker,dilatation of gland cystic atrophy with more gland than those in model group.4.The changes of serum Ca?P?E₂ levels in rats:the serum Ca and E₂ content in model group decreased significantly compared with sham group.The level of serum P in the model group was increased?P<0.01?.Serum Ca?E₂ were increased,and serum P was increased after twelve weeks.5.Expression of BMP-2,Smad1/5,and Smad6 proteins:The positive expression of BMP-2 and Smad1/5 were lower,but the positive expression of Smad6 was higher than those in the model group.The expression of BMP-2?Smad1/5 in each drug group was increased significantly,and Smad6 expression was decreased compared with the model group after twelve weeks.Conclusion1.The OP model was successfully replicated using bilateral ovariectomy in female rats.2.Tonifying kidney and strengthening bone method can significantly improve the BMD of OP model rats,promote the integrity of trabecular bone,and reduce the cavity of marrow cavity.Bushenzhuanggufang can improve the uterus coefficient of OP-model rats significantly and stimulate the growth of uterus,thicken endometrium,improve glandular cystic atrophy and increase number of glands,and regulate the content of serum Ca,P and E₂.3.Tonifying kidney and strengthening bone method can improve the BMD in OP model rats and improve bone metabolism,which may be related to the increasing the level of BMP-2 and Smad1/5 protein and the decreasing the level of Smad6 protein.Therefore tonifying kidney and strengthening bone method could promote the proliferation and differentiation of OB and promote the formation of new bone.