The Role Of SIRT1 Gene In The Pathogenesis Of Aseptic Prosthesis Loosening And Related Mechanisms

Background:Aseptic prosthesis loosening is one of the major reasons for the failure of primary total hip arthroplasty and total knee arthroplasty,and it is highly relevant to the disturbance in bone metabolism.To date,researches on the mechanisms of aseptic loosening are focus on interaction between wear particles and multiple cell types,the characteristics of which are chronic inflammatory responses and osteolysis around prosthesis.However,plenty of the studies in these years do not clearly discover the pathogenesis of aseptic loosening,and an effective treatment to prevent and cure aseptic loosening has not been found.On the other hand,Sirtuin 1(SIRT1)has recently been reported to be involved in agerelated diseases,such as neurodegenerative diseases,metabolic diseases,cancer,cardiovascular diseases and osteoporosis.In female mice that were SIRT1 knockdown,researchers have reported that significant reduction in bone mass is exhibited in those mice,the characteristic of which is decreased bone formation.On the other hand,in the pathogenesis of multiple inflammatory disorders,such as rheumatoid arthritis,SIRT1 gene also plays an key role in anti-inflammatory effect.However,it is not clear that whether SIRT1 gene plays a role in the pathogenesis of aseptic prosthesis loosening and what are the related mechanisms.Furthermore,the evidences that targeted SIRT1 gene for the therapy of aseptic prosthesis loosening still need to be explored.Methods:we collected the clinical data and tissues around prosthesis in patients with aseptic prosthesis loosening after the primary total hip arthroplasty,and the clinical data and tissues in patient without aseptic loosening was used as control.The SIRT1 protein expression was detected in patients with aseptic prosthesis loosening and control by western blots and immunofluorescence staining.The expression of inflammatory cytokines was detected by ELISA,including IL-6,TNF-α,and IL-β.Two metal wear particles which were commonly used for prosthesis were prepared,including TiAlV4 particles and CoCrMo particles.The expression of inflammatory cytokines was detected by ELISA,the number of osteoblast was detected by MTT assay,and the osteoblast apoptosis was detected by flow cytometry.Resveratrol,a SIRT1 activator and EX527,a SIRT1 inhibitor were used to investigate the relation among SIRT1 expression,particle-induced chronic inflammatory responses and particle-induced osteoblast apoptosis.To provide the direct evidence that SIRT1 regulated particle-induced inflammatory responses and osteoblast apoptosis,we further overexpressed SIRT1 in macrophages and osteoblasts by recombinant lentivirus to further explore the downstream genes of SIRT1 and the signal network related to aseptic prosthesis loosening.A murine calvarial osteolysis model was constructed and treated with resveratrol to explore whether SIRT1 could be a potential target for the treatment to aseptic loosening.The severity of osteolysis was observed by micro-CT and the toluidine blue staining.The expression of inflammatory cytokines in the periosteum and calvarias were detected by ELISA and qPCR.The amount of osteoblasts was observed by Hematoxylin-eosin(HE)staining.The alkaline phosphatase(ALP)activity and the genes expression related to bone formation were also detected.Results:.the SIRT1 protein expression was significantly decreased in patients with aseptic prosthesis loosening after primary total hip arthroplasty compared with the patients without aseptic prosthesis loosening.Wear particles could induce the downregulation of SIRT1 in macrophages and osteoblast.At the same time,wear particles could significantly increase the expression of inflammatory cytokines in macrophages and osteoblasts.On the other hand,wear particles could reduce the number of osteoblasts and promote osteoblast apoptosis.Resveratrol,a SIRT1 activator,could significantly suppress particleinduced inflammatory responses in macrophages and osteoblasts through NF-κB signal pathway.And resveratrol could also significantly suppress osteoblast apoptosis through p53 signal pathway.SIRT1 overexpression in macrophages and osteoblasts by using recombine lentivirus could significantly suppress particle-induced inflammatory responses,and suppress particle-induced osteoblast apoptosis.Resveratrol could significant reduce the severity of osteolysis in murine calvarial osteolysis model.On the other hand,resveratrol could also significantly decrease the inflammatory responses in the calvarias and periosteum,and significantly increased the amount of osteoblasts,the activity of ALP and the expression of bone formationrelated genes(Runx2、Ocn、Dkk1、Sost、Osx).Conclusion:SIRT1 gene plays an important role in the pathogenesis of aseptic prosthesis loosening.Our study demonstrated that SIRT1 could suppress chronic inflammatory responses and osteoblast apoptosis and disclosure a new strategy for aseptic loosening treatment.SIRT1 gene may be a potential effective target for the treatment of aseptic prosthesis loosening.