Study On The Expression Mechanism Of MTOR Pathway In Steroid-induced Femoral Head Necrosis

Background: With the widespread using of hormone drugs in clinical,femoral head necrosis(osteonecrosis of femoral head,ONFH)had been reported is gradually increasing around the world.Femoral head necrosis is a serious complication recognized by hormones treatment in clinical applications,and the disability rate is high,which seriously affects the physical and mental health in patients.In China,long-term and high-dose corticosteroids are the most common pathogenic factors for nontraumatic femoral head necrosis.However,the pathogenesis remains unclear.But there are some studies have shown that the change of epigenetic genome is closely related to its occurrence and the signaling pathways.In Currently studies,which have found that the mTOR pathway plays an important role in the development of cartilage degenerative disease and osteosarcoma.However,the specific role of the mTOR pathway in steroidinduced femoral head necrosis is still unknow,we need more studies and further confirmation.Objective: The purpose of this study was to investigate the role of mTOR pathway signaling pathway in steroid-induced femoral head osteonecrosis in rats.In order to provide protease mTOR pathway and the changing factors in the regulation mechanism between hormone osteonecrosis,providing a certain hormone osteonecrosis in the clinical practice.Materials and Methods: Forty eight healthy were injected intramus-cularly with 20 mg/kg methylprednisolone(MP)for 8 weeks,twice per week.The animals were sacrificed at 2,4 and 8 weeks after the last MP injection,respectively,and then allocated to the 2-,4-and 8-week model groups(n=24Each).Rats in the control group(n=24)were not given any treatment,Histopathological analysis was performed and the concentration of tartrate-resistant acid phosphatase(TRAP)in plasma was determined.The activation of osteoclasts in the femoral head was assessed by TRAP staining.Western-blot analysis: RANK,VEGF,OPG,RUNX2 and other proteins in rabbit femoral head were detected.TOR pathway protein analysis: S6 protein,4E binding protein 1(4EBP1),PI3 K,AKT and other proteins in rabbit femoral head cells were detected.The expression levels of apoptosis-related proteins Bax,bcl-2 and caspase-3 were observed by Western-blot analysis.Results: The results showed that the osteonecrosis in the femoral head was clearly observed and the concentration of TRAP in the plasma was increased in the mode l rats.The femoral head tissues in MP-treated rats were positive for TRAP and the intensity of TRAP staining was greater in MP-treated rats than in control rats.HE dyeing analysis: the control group: overall bone trabecular structure is complete,the spatial configuration,the bone cells can be clearly seen,full of rich hematopoietic cells in bone marrow,fat cells is relatively small,normal cells form.Model group: the cartilaginous surface showed a certain degree of fold,the bone trabeculae was sparse on the whole,and there was local fracture.The nucleus of bone nucleus appeared to be fixed to a certain extent,and the fat cells in bone marrow were abundant.Analysis of hollow bone fossa rate: the number of empty bone fossa over the control group was small,and its fossa rate was(8.33±1.21)%.In the model group,a large number of empty bone dimples appeared,and the fossa rate was(23.58±3.38)%.The data of the two groups were compared,P<0.05,and the difference was statistically significant comparison of Cspase-3 content: control group < model group;Comparison of Bcl-2 content: model group < control group,suggesting a certain degree of apoptosis in bone cells.Western-blot protein analysis: femoral head protein: RANK,RUNX2 increased expression after the model establishment,P<0.05,the difference was statistically significant.VEGF,OPG decreased after the model was established,P<0.05,and the difference was statistically significant.TOR pathway proteins: the key proteins S6,4EBP1,PI3 K,and AKT were all increased to a certain extent after the establishment of the model,P<0.05,the difference was statistically significant.Apoptosis protein analysis: The expression levels of pro-apoptotic proteins Bax and caspase-3 increased after the establishment of the model.The data of the two groups were statistically significant(P<0.05).The expression of apoptotic protein Bcl-2 decreased,P< 0.05,and the difference was statistically significant.Conclusion: Steroid-induced femoral head necrosis can cause substantial damage to bone cells.It can effectively promote the apoptosis of bone cells.The mTOR signaling pathway molecules S6,4Ebp1,PI3 K,and AKT in the upstream and downstream was obvious activation in bone tissue after the model establishment,prompting the mTOR signaling pathways involved in the process of the hormone osteonecrosis.