| Part One Experimental study of the effects of hypoxia simulator onosteointegration of titanium prosthesis in osteoporotic ratsObjective:Poor osseointegration is an important cause of implant failure after joint replacement.As a hypoxia simulator,deferoxamine(DFO)can stabilize hypoxia inducible factor 1α(HIF-1α)under normoxia.Then a large number of cytokines,such as vascular endothelial growth factor(VEGF)etc,are generated downstream to promote bone formation through the coupling pathway of angiogenesis and osteogenesis.The purpose of this study was to establish a knee arthroplasty model in osteoporotic rats and to observe the effect of local administration of DFO on osteogenesis and osseointegration with knee prosthesis in osteoporotic bone of rats.Methods:In this study,90 female rats were used.The osteoporosis model was established by ovariectomy.The titanium prosthesis of rat knee was made by 3D printing technology and implanted into the upper tibia of osteoporotic rats.Then the rats were randomly divided into three groups with30 rats in each group.The first group was treated with 100umol DFO locally as the low-dose experimental group,the second group was treated with200umol DFO locally as the high-dose experimental group,and the third group was treated with normal saline locally as the control group.Two weeks after operation,the expressions of HIF-1α,VEGF and CD31 in periprosthetic bone were detected by real-time quantitative PCR,immunohistochemistry and Western blot.At 12 weeks after operation,the maximum pull-out force of the prosthesis in each group was measured by biomechanics,the bone formation and bone trabecular parameters around the prosthesis were detected by micro CT,and the pathological tissue sections were made to observe the integration of the prosthesis and the surrounding bone.Results:q PCR results showed that the m RNA expressions of VEGF and CD31 in high-dose and low-dose DFO groups were higher than those in the control group.The m RNA expressions of VEGF and CD31 in high-dose DFO group were higher than those in low-dose DFO group.Immunohistochemical staining showed that the positive expressions of HIF-1α,VEGF and CD31 in the two DFO groups were higher than those in the control group.CD31showed obvious cluster expression in the DFO group.At the same time,the positive expression of HIF-1α,VEGF and CD31 in the high-dose DFO group was higher than that in the low-dose DFO group.Western blot showed that the protein expression levels of HIF-1α,VEGF and CD31 in high-dose and low-dose DFO groups were significantly higher than those in the control group.The protein expression levels of HIF-1α,VEGF and CD31 in high-dose DFO group were higher than those in low-dose group.DFO caused obvious angiogenesis.At 12 weeks after operation,the biomechanical results showed that the maximum pull-out force of prosthesis in the two DFO groups was higher than that in the control group,and there was no significant difference between the high-dose DFO group and the low-dose group.Then micro CT examination was performed.The results showed that the bone formation around the implant in DFO group was significantly higher than that in control group,BMD,BV/TV,Tb.N,Tb.Th was higher than that in the control group,TB.SP was lower than that in the control group,while BMD,BV/TV,Tb.N,Tb.Th in the high-dose DFO group was higher than that in low dose group,TB.SP was lower than that in the low dose group.The results of histological sections showed that Bone-to-implant contact(BIC)value in DFO group was higher than that in control group,and the BIC in high-dose DFO group was higher than that in low-dose group.Conclusion:The animal model of osteoporotic knee arthroplasty in rats was successfully established.Local administration of DFO to activate HIF-1αsignaling pathway can promote periprosthesis osteogenesis and osteointegration in osteoporotic bone of rats.With the increase of dose,periprosthesis osteogenesis and osteointegration rate increase accordingly,which provides a new solution for preventing prosthesis loosening after arthroplasty in patients with osteoporosis.Part Two Effect of deferoxamine on osseointegration of knee prosthesisin rats through different administration methods and comparative studyObjective:Iron overload is closely related to osteoporosis.In addition to activating HIF-1αsignal pathway,deferoxamine,as an iron chelator,can also reduce iron overload,prevent bone loss and improve osteoporosis.The purpose of this study is to observe the effects of systemic administration of deferoxamine on bone mass and bone integration of knee prosthesis in osteoporotic rats,and further compare the effects of DFO with different ways of medication on osseointegration,so as to explore the optimal method.Methods:In this study,110 female osteoporosis rats were used for knee prosthesis implantation,and then the rats were randomly divided into 5 groups with 22 rats in each group.They were systemic administration of DFO group(DFO-S group),local administration of DFO group(DFO-L group),local and systemic administration of DFO group(DFO-LS group),alendronate group(ALN group),and control group(OVX group).In addition,22 female rats with normal bone were selected as the sham operation group(Sham group),which also underwent prosthesis implantation,a total of 6 groups were studied.The rats in the DFO-S group were injected with 100mg/kg DFO intraperitoneally,the rats in the DFO-L group were injected with 100mg/kg DFO intra-articular,and the rats in the DFO-LS group were injected with100mg/kg DFO intra-articular and intraperitoneally,once every other day for one month.Rats in the ALN group were given 0.5mg/kg alendronate by gavage once every other day for 1 month.The rats in the control group were given normal saline.One month after prosthesis implantation,the m RNA expression of VEGF,CD31,osteocalcin and Runx-2 in periprosthetic bone was detected by real-time quantitative PCR,the expression of HIF-1α,VEGF and CD31 in bone tissue was detected by immunohistochemistry.The contents of serum ferritin,type I procollagen amino-terminal peptide(PINP)and?-type I collagen carboxy-terminal peptide(?-CTX)were detected by ELISA.Two months after operation,the maximum pull-out force of the prosthesis in each group was measured by biomechanics,the bone formation around the prosthesis and bone trabecular parameters were detected by micro CT,and the pathological tissue sections were made to observe the integration of the prosthesis and the surrounding bone.Results:q PCR results showed that the m RNA expression levels of VEGF and CD31 in DFO-S group were higher than those in OVX group and lower than those in DFO-L and DFO-LS groups(P<0.05),but there was no significant difference between DFO-L and DFO-LS groups(P>0.05).The m RNA expression levels of osteocalcin and Runx-2 in DFO-S group were not significantly different from those in OVX group(P>0.05).The m RNA expression levels of osteocalcin and Runx-2 in DFO-L and DFO-LS groups were significantly higher than those in OVX group and DFO-S group(P<0.05).Immunohistochemical results showed that the expression level of HIF-1αin DFO-L,DFO-S and DFO-LS groups was significantly higher than that in OVX group(P<0.05).The expression levels of VEGF and CD31 in DFO-L group and DFO-LS group were significantly higher than those in OVX group(P<0.05).There was no significant difference in the expression levels of VEGF and CD31 between DFO-S group and OVX group(P>0.05).The results of ELISA showed that the level of serum ferritin in DFO-S group and DFO-LS group was significantly lower than that in OVX group(P<0.05).There was no significant difference in serum ferritin level between DFO-L group and OVX group(P>0.05).The levels of serum PINP and CTX in DFO-S group and DFO-LS group were lower than those in OVX group(P<0.05).There was no significant difference in the levels of PINP and CTX between DFO-S group and DFO-LS group(P>0.05).There was no significant difference in PINP and CTX levels between DFO-L group and OVX group(P>0.05).The level of CTX in DFO-LS group was significantly lower than that in DFO-L group(P>0.05).The biomechanical results showed that the maximum pull-out force of prosthesis of the DFO-S,DFO-L,DFO-LS and ALN groups was significantly higher than that of the OVX group(P<0.05).Micro CT results showed that the bone mass around the prosthesis in DFO-S,DFO-L,DFO-LS and ALN groups was significantly higher than that in OVX group,BMD,BV/TV,TB.Th,Tb.N was significantly higher than that in the OVX group,TB.SP was lower than that in the OVX group.DFO-LS group had the most bone mass,thick bone trabeculae and regular arrangement(P<0.05).The BIC in the DFO-S group was 54.8±7.8%,the BIC of the DFO-L was 62.2±7.7%,and the BIC of the DFO-LS group was 71.0±5.5%.However,the BIC of the OVX group was only 38.6±10.3%.The BIC of the DFO-S,DFO-L,DFO-LS groups was significantly higher than that of the OVX group,and the BIC of the DFO-LS group was the highest.The BIC of DFO-L group was slightly higher than that of DFO-S group,but slightly lower than that of DFO-LS group(P>0.05).BIC of rats in ALN group was slightly lower than that in DFO-L and DFO-S groups(P>0.05).Conclusion:Systemic administration of deferoxamine can significantly increase the bone mass around the knee prosthesis,improve the bone integration and increase the stability of the prosthesis in osteoporotic rats.Its mechanism is mainly related to DFO reducing iron load,inhibiting bone resorption and reducing bone turnover.The effect of systemic administration of DFO on bone integration was weaker than that of local administration,and there was no significant difference between the systemic administration group and the alendronate group.The effect of bone integration in the combined group was stronger than that in the local group,systemic group and alendronate group.Part Three Evaluation of bone mineral density,bone turnover index andrisk factors before knee arthroplastyObjective:Low bone mineral density(BMD)is a risk factor for implant osseointegration,long-term survival and prosthesis complications in arthroplasty.Less attention was paid to bone mineral density and bone metabolism before operation.The purpose of this study is to observe the level of bone mineral density and bone turnover markers before knee arthroplasty,evaluate the incidence of osteoporosis,analyze the risk factors of preoperative osteoporosis and high turnover,and help find patients with osteoporosis and high turnover.Methods:From May 2018 to May 2020,104 patients with osteoarthritis who met the criteria and planned to undergo knee replacement were included.The BMD of lumbar spine and total hip joint were measured by dual energy X-ray absorptiometry,and the incidence of osteoporosis was analyzed according to T value;The level of bone turnover markers was detected,the type I procollagen amino terminal peptide(PINP),a marker of bone formation,was detected by ELISA,and the cross-linked carboxy terminal peptide(?-CTX),a marker of bone resorption,was detected by immunoluminescence.Pearson correlation coefficient was used to analyze the correlation between bone mineral density and various factors.Logistic regression analysis was used to assess the risk factors of osteoporosis and high bone bone turnover rate.Results:In this cohort study,the BMD of lumbar spine was0.773±0.160g/cm~2and that of hip joint was 0.737±0.157g/cm~2.The incidence of lumbar osteoporosis and osteopenia were 22.1%and 35.6%respectively,and the prevalence of osteoporosis or osteopenia was 57.7%.The incidence of osteoporosis and osteopenia of hip joint were 26.0%and 43.3%respectively,and the incidence of osteoporosis or osteopenia was 69.2%.In terms of gender,the incidence rate of lumbar osteoporosis in men and women was 6.3%and25%respectively.The incidence of lumbar vertebral osteopenia in men and women was 12.5%and 38.6%respectively,and the incidence of osteoporosis in women was significantly higher than that in men(P<0.05).There was no significant difference in the incidence of osteoporosis or osteopenia measured at the lumbar spine and hip(P>0.05).Lumbar BMD before knee arthroplasty was negatively correlated with age and knee stiffness,and positively correlated with body mass index(P<0.05).Female,old age,low body mass index and knee stiffness are the risk factors of osteoporosis in patients with knee osteoarthritis.Before knee arthroplasty,the bone turnover markers PINP and?-CTX were 40.26±8.11ug/L and 0.470±0.176ug/L,which meant high turnover state.The bone turnover rate of women is higher than that of men.There was a positive correlation between preoperative plasma?-CTX level and knee stiffness score(P<0.05).Female,high K-L grade and knee stiffness are risk factors for high bone turnover rate in patients with knee osteoarthritis.Conclusion:the incidence of osteoporosis and osteopenia in patients with osteoarthritis before knee arthroplasty is high,especially in postmenopausal women.The overall bone turnover of patients before operation was at a high level,and the elevated?-CTX level indicated that there was systemic or local abnormal bone turnover.Paying attention to the risk factors of patients with osteoporosis and high turnover and strengthening the preoperative bone mineral density and bone turnover examination can help clinicians do a good job in perioperative bone evaluation.This information is of great significance to improve the bone conditions of patients and optimize the effect of surgery. |
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