Objective:To explore the mechanism of Gubi Tongxiao granule in treating osteoblast injury induced by glucocorticoid.Methods:Mouse osteogenic precursor cell MC3T3-E1 was cultured in vitro,the control group was conventi...Objective:To explore the mechanism of Gubi Tongxiao granule in treating osteoblast injury induced by glucocorticoid.Methods:Mouse osteogenic precursor cell MC3T3-E1 was cultured in vitro,the control group was conventionally cultured,the model group was treated with dexamethasone(10^(-5) M,10^(-6) M,10^(-7) M)solution with gradient concentration to induce injury,and the experimental group was added with Gubi Tongxiao granule drug-containing serum to intervene culture on the basis of the model group.Cell viability was detected by CCK-8 method,autophagy and apoptosis were detected by MDC/PI staining,Beclin-1,Bcl-2 and Bax expression levels were detected by Western-blot,and the expression of target genes in each group was analyzed by RT-qPCR.Results:Compared with the control group,the activity of osteoblasts in the model decreased significantly.The effect of dexamethasone on autophagy and apoptosis of MC3T3-E1 cells was time-dependent and dose-dependent(P<0.05).The autophagy marker Beclin-1 increased at low doses of dexamethasone(10^(-7) or 10^(-6) M)and decreased at high doses(10^(-5) M).Compared with model group,Beclin-1 and Bax mRNA expression in experimental group decreased(P<0.01),and Bcl-2 mRNA expression was significantly up-regulated(P<0.01).In addition,the results of semi-quantitative analysis of apoptosis staining showed that autophagy and apoptosis were inhibited in different degrees(P<0.01).Conclusion:Gubi Tongxiao granule has a protective effect on osteoblast injury caused by dexamethasone,and can effectively reduce autophagy and apoptosis induced by glucocorticoid,which may be one of the mechanisms of maintaining bone mass and delaying the occurrence and development of femoral head necrosis.展开更多
Objective:To investigate the therapeutic effect of osteopathic osteopathy on hormonal osteonecrosis in rabbits by regulating the Wnt/PI3K-AKT signaling pathway.Methods:Thirty clean-grade New Zealand rabbits were rando...Objective:To investigate the therapeutic effect of osteopathic osteopathy on hormonal osteonecrosis in rabbits by regulating the Wnt/PI3K-AKT signaling pathway.Methods:Thirty clean-grade New Zealand rabbits were randomly divided into control group,model group and experimental group,and all except for the control group,allogeneic serum combined with glucocorticoids were used to establish animal models of simulated hormonal osteonecrosis of the femoral head.After successful modeling,the experimental group was given the model group and the control group the same amount of normal saline,and the experimental group was given bone palsy granule concentrate according to the equivalent dose of humans and rabbits,once a day for 8 weeks,and after intraperitoneal anesthesia,blood was collected from the extracorporeal heart and the serum ALP and TGF-β1 levels were detected by immunoenzyme-linked adsorption.The experimental animals were sacrificed,,and the bilateral femoral heads of rabbits were removed for hematoxylin-eosin staining,and the apoptosis of cells in the femoral head area was observed by TUNEL staining.RT-qPCR analyzed the expression of Bax and Bcl-2 genes,and Western blot detected the expression levels of pathway proteins Wnt,β-catenin,GSK-3β,AKT and p-AKT.Results:Compared with the control group,the expression levels of ALP and TGF-β1 in rabbit bone tissue in the model group decreased significantly(P<0.05),the bone trabeculae were sparse and thinned,the number of hollow bone fosses and the number of apoptosis-positive cells increased significantly(P<0.05),the expression of pro-apoptotic gene Bax was increased,and the expression of anti-apoptotic gene Bcl-2 was inhibited(P<0.05).After the intervention and treatment of bone palsy granules,the hollow bone depression and apoptosis of rabbit bone tissue in the experimental group were effectively improved(P<0.05).In addition,compared with the model group,the osteopathic elimination particles could upregulate the expression of ALP and TGF-β1(P<0.05),promote the expression of pathway proteins Wnt andβ-catenin,increase the phosphorylation level of AKT,and downregulate the expression of GSK-3β(P<0.05).Conclusion:Under glucocorticoids,osteoblast development dysfunction and apoptosis increase.Bone palsy granules can regulate the two signaling pathways of Wnt/PI3K-AKT,promote osteoblast development and inhibit apoptosis,effectively maintain bone metabolic balance,prolong the collapse time of the femoral head,and further treat osteonecrosis of the femoral head.展开更多
Objective:To investigate the effect of Gubi Tongxiao granules on vascular formation and cell apoptosis in the process of glucocorticoid-induced femoral head necrosis.Methods:Thirty experimental New Zealand rabbits wer...Objective:To investigate the effect of Gubi Tongxiao granules on vascular formation and cell apoptosis in the process of glucocorticoid-induced femoral head necrosis.Methods:Thirty experimental New Zealand rabbits were randomly divided into groups.Except the control group,the animal model of steroid-induced femoral head necrosis was established by lipopolysaccharide combined with glucocorticoids.After successful modeling,the experimental group was given Gubi Tongxiao granules intragastric treatment,the model and control group were given the same amount of normal saline intragastric treatment,once a day,for 8 weeks,the experimental animals were sacrificed,the bilateral femoral head of each group was taken out for hematoxylin-eosin staining,and protein CD34,CYR61 and VEGF were immunohistochemical staining localization.TUNEL staining was used to observe the apoptosis of femoral head cells.The expression levels of apoptosis proteins Bax and Bcl-2 were detected by WB.The expression of VEGF eNOs Bax and Bcl-2 genes was analyzed by RT-qPCR.Results:Gubi Tongxiao granules reduced the number of hollow bone lacunae and apoptotic positive cells(P<0.01),and up-regulated the expressions of CD34,CYR61 and VEGF in femoral head tissue.Compared with control group,the expression of pro-apoptotic protein Bax increased and the expression of anti-apoptotic protein Bcl-2,VEGF and eNOs mRAN decreased significantly in the model group(P<0.01),the opposite results were obtained in the experimental group after Gubi Tongxiao granule intervention(P<0.05).Conclusion:Gubi Tongxiao granules can improve the expression of angiogenic genes and regulate apoptosis-related proteins in the bone tissue of rabbit model with steroid-induced necrosis of femoral head,promote angiogenic differentiation and inhibit apoptosis,and thus achieve the effect of treating steroid-induced necrosis of femoral head.展开更多
基金General Program of Anhui Natural Science Foundation(2008085MH281)University Natural Science Research Project of Anhui Provincial Department of Education(KJ2020A0403)。
文摘Objective:To explore the mechanism of Gubi Tongxiao granule in treating osteoblast injury induced by glucocorticoid.Methods:Mouse osteogenic precursor cell MC3T3-E1 was cultured in vitro,the control group was conventionally cultured,the model group was treated with dexamethasone(10^(-5) M,10^(-6) M,10^(-7) M)solution with gradient concentration to induce injury,and the experimental group was added with Gubi Tongxiao granule drug-containing serum to intervene culture on the basis of the model group.Cell viability was detected by CCK-8 method,autophagy and apoptosis were detected by MDC/PI staining,Beclin-1,Bcl-2 and Bax expression levels were detected by Western-blot,and the expression of target genes in each group was analyzed by RT-qPCR.Results:Compared with the control group,the activity of osteoblasts in the model decreased significantly.The effect of dexamethasone on autophagy and apoptosis of MC3T3-E1 cells was time-dependent and dose-dependent(P<0.05).The autophagy marker Beclin-1 increased at low doses of dexamethasone(10^(-7) or 10^(-6) M)and decreased at high doses(10^(-5) M).Compared with model group,Beclin-1 and Bax mRNA expression in experimental group decreased(P<0.01),and Bcl-2 mRNA expression was significantly up-regulated(P<0.01).In addition,the results of semi-quantitative analysis of apoptosis staining showed that autophagy and apoptosis were inhibited in different degrees(P<0.01).Conclusion:Gubi Tongxiao granule has a protective effect on osteoblast injury caused by dexamethasone,and can effectively reduce autophagy and apoptosis induced by glucocorticoid,which may be one of the mechanisms of maintaining bone mass and delaying the occurrence and development of femoral head necrosis.
基金supported by Fund Information:Postgraduate Research Project of Universities in Anhui Province(YJS20210477)Natural Science Foundation of Anhui Province(2008085MH281)Key Project of Natural Science Research in Colleges and Universities in Anhui Province(KJ2020A0403).
文摘Objective:To investigate the therapeutic effect of osteopathic osteopathy on hormonal osteonecrosis in rabbits by regulating the Wnt/PI3K-AKT signaling pathway.Methods:Thirty clean-grade New Zealand rabbits were randomly divided into control group,model group and experimental group,and all except for the control group,allogeneic serum combined with glucocorticoids were used to establish animal models of simulated hormonal osteonecrosis of the femoral head.After successful modeling,the experimental group was given the model group and the control group the same amount of normal saline,and the experimental group was given bone palsy granule concentrate according to the equivalent dose of humans and rabbits,once a day for 8 weeks,and after intraperitoneal anesthesia,blood was collected from the extracorporeal heart and the serum ALP and TGF-β1 levels were detected by immunoenzyme-linked adsorption.The experimental animals were sacrificed,,and the bilateral femoral heads of rabbits were removed for hematoxylin-eosin staining,and the apoptosis of cells in the femoral head area was observed by TUNEL staining.RT-qPCR analyzed the expression of Bax and Bcl-2 genes,and Western blot detected the expression levels of pathway proteins Wnt,β-catenin,GSK-3β,AKT and p-AKT.Results:Compared with the control group,the expression levels of ALP and TGF-β1 in rabbit bone tissue in the model group decreased significantly(P<0.05),the bone trabeculae were sparse and thinned,the number of hollow bone fosses and the number of apoptosis-positive cells increased significantly(P<0.05),the expression of pro-apoptotic gene Bax was increased,and the expression of anti-apoptotic gene Bcl-2 was inhibited(P<0.05).After the intervention and treatment of bone palsy granules,the hollow bone depression and apoptosis of rabbit bone tissue in the experimental group were effectively improved(P<0.05).In addition,compared with the model group,the osteopathic elimination particles could upregulate the expression of ALP and TGF-β1(P<0.05),promote the expression of pathway proteins Wnt andβ-catenin,increase the phosphorylation level of AKT,and downregulate the expression of GSK-3β(P<0.05).Conclusion:Under glucocorticoids,osteoblast development dysfunction and apoptosis increase.Bone palsy granules can regulate the two signaling pathways of Wnt/PI3K-AKT,promote osteoblast development and inhibit apoptosis,effectively maintain bone metabolic balance,prolong the collapse time of the femoral head,and further treat osteonecrosis of the femoral head.
基金Natural Science Research Project of Higher Education Department of Anhui Province(No.KJ2020A0403)Natural Science Foundation of Anhui Province(No.2008085MH281)。
文摘Objective:To investigate the effect of Gubi Tongxiao granules on vascular formation and cell apoptosis in the process of glucocorticoid-induced femoral head necrosis.Methods:Thirty experimental New Zealand rabbits were randomly divided into groups.Except the control group,the animal model of steroid-induced femoral head necrosis was established by lipopolysaccharide combined with glucocorticoids.After successful modeling,the experimental group was given Gubi Tongxiao granules intragastric treatment,the model and control group were given the same amount of normal saline intragastric treatment,once a day,for 8 weeks,the experimental animals were sacrificed,the bilateral femoral head of each group was taken out for hematoxylin-eosin staining,and protein CD34,CYR61 and VEGF were immunohistochemical staining localization.TUNEL staining was used to observe the apoptosis of femoral head cells.The expression levels of apoptosis proteins Bax and Bcl-2 were detected by WB.The expression of VEGF eNOs Bax and Bcl-2 genes was analyzed by RT-qPCR.Results:Gubi Tongxiao granules reduced the number of hollow bone lacunae and apoptotic positive cells(P<0.01),and up-regulated the expressions of CD34,CYR61 and VEGF in femoral head tissue.Compared with control group,the expression of pro-apoptotic protein Bax increased and the expression of anti-apoptotic protein Bcl-2,VEGF and eNOs mRAN decreased significantly in the model group(P<0.01),the opposite results were obtained in the experimental group after Gubi Tongxiao granule intervention(P<0.05).Conclusion:Gubi Tongxiao granules can improve the expression of angiogenic genes and regulate apoptosis-related proteins in the bone tissue of rabbit model with steroid-induced necrosis of femoral head,promote angiogenic differentiation and inhibit apoptosis,and thus achieve the effect of treating steroid-induced necrosis of femoral head.
