Effects And Its Mechanism Of SR On Medial Meniscectomy Induced Knee Degeneration Of DH Guinea Pigs

Part one Experimental study on medial menisectomized guinea pigs withosteoarthritisObjective: To evaluate the histological changes of medial meniscectomy(MNX)induced cartilage degeneration in Dunkin Hartley(DH)guinea pigs and provide a scientific model of osteoarthritis.Methods:1 Twenty-four 3-month-old male DH guinea pigs were randomly divided into two groups(n=12),Sham group and MNX group,animals were received sham operation or medial meniscectomy,respectively.2 All animals were sccrificed 12 weeks post operation,then the joints were evaluated by OARSI histological scoring system,OA-related proteins were assessed by immunohistochemical techniques,and micro-architecture of subchondral trabecular bone were scanned by micro-CT.Results:1 Bodyweights for animals from both groups were growing through time,there were no statistical significance were observed between groups at any time points.2 Joints from MNX group were severely degenerated post operation,the cartilage thicknesses were significantly reduced compared to Sham group(P<0.05).The results indicate OARSI total scores with CS,PG,CL and OST subsets in MNX group were siginificantly higher than Sham group(P< 0.05).3 The expression level of AGG in cartilage of MNX group was significantly reduced compared to Sham group,however,the expression level of MMP-13,ADAMTS4 and Caspase-3 were significantly up-regulated(P<0.05).The ratio of AGG/ADAMTS4 and Col-II/MMP-13 were both significantly lower in MNX group,however,the ratio of Col-II/AGG wassignificantly higher than Sham group(P< 0.05).4 Micro CT results showed that BMD?BV/TV?Tb.Th were significantly less in the MNX group than in the sham groups,and Other subchondral bone micro-architecture parameters(Tb.Sp and SMI)indicated subchondral bone higher in the MNX group.Part two Effects of strontium strontium on the arthritis of MNX guineapigObjective: To study the effects of SR on cartilage degeneration in MNX induced OA model of DH guinea pigs.Methods:1 Thirty-six 3-month-old male DH albino guinea pigs were randomly divided into three groups as follows: MNX group,CLX group and SR group,twelve animals per group.Each group was then randomly divided into two subgroups for macroscopic scoring(n=6)or histological scoring(n=6).2 Animals were anesthetized with sodium pentobarbital(30 mg/kg,ip),and a medial meniscectomy was performed on the right knee.Meniscectomized animals began 12 weeks of treatment by oral administration of CLX(20 mg/kg,daily)and SR(625 mg/kg,daily),or normal saline for MNX group.3 All animals were sccrificed,then the joints were evaluated by OARSI histological scoring system.Results:1 Bodyweights for animals from each group were growing through time,there were no statistical significance were observed between any two groups at any time points.The operation significantly impaired articular cartilage of medial tibial plateau.Medial tibial plateau cartilage surface of animals undergoing MNX operation became rougher and larger with cleft and chondral hyperplasia forming on the medial edge of the cartilage.2 The OARSI macroscopic scores of SR or CLX groups are slightly lower than MNX + NS group,but no significant difference between the three groups.3 OARSI histological scores showed: total scores in MNX + CLXgroup were significantly lower than MNX + NS group(MNX + CLX group vs.MNX + NS group,10.78 ± 1.91 vs 17.36 ± 1.92,P <0.05.),no significant difference between MNX + SR group and MNX + NS group.Cartilage thickness measurements showed:MNX + CLX animals cartilage thickness was significantly higher than MNX + NS group(MNX + CLX group vs.MNX +NS group,360.19 ± 38.93?mvs.264.73 ± 58.01?m,P <0.05),and MNX + SR group cartilage thickness MNX + NS group compared with no significant difference.In addition,MNX + SR group was significantly higher than the height of osteophytes MNX + NS group(MNX + SR group vs.MNX + NS group,198.18 ± 48.31?mvs.106.86 ± 57.11,P <0.05),MNX + CLX group osteophytes height slightly less than MNX + NS group,but no significant difference.4 There are 7 samples in SR group showed collapsed osteophytes,while there is 1 sample indicates the same appearance in MNX group,this change was found significant differences between SR group and MNX group.Part three The mechanism of SR on MNX guinea pigsObjective: By the previous experimental observation,Strontium Ranelate(625 mg/kg`day)fail to protect cartilage from degeneration in menisectomized guinea pigs,SR administration results in chondral part of osteophyte collapsed.Therefore,in this study,we explore the mechanism of this phenomenon through the further research on cartilage subchondral bone after the treatment of SR.Methods:1 Thirty-six 3-month-old male DH albino guinea pigs were randomly divided into three groups as follows: MNX group,CLX group and SR group,twelve animals per group.Each group was then randomly divided into two subgroups for micro CT testing(n=6)or histological scoring(n=6).MNX were performed on right knees of animals in each group.2 Meniscectomized animals began 12 weeks of treatment by oral administration of CLX(20 mg/kg,daily)and SR(625 mg/kg,daily),or normal saline for MNX group.3 All animals were sccrificed,then the joints were evaluated by immunohistochemistry with primary anti-body of AGG(1 : 200),ADAMTS-4(1:200),Caspase-3(1:200),Col-II(1:50),and MMP-13(1:200),respectively.The proximal tibiae were scanned using a micro-CT to quantify micro-architecture of subchondral trabecular bone.Results:1 Immunohistochemical analysis:Among the three groups AGG,Col-II,Caspase3 and MMP13 in weight-bearing area on the medial tibial plateau expression results showed no significant difference,and MNX+CLX group of ADAMTS4 expression levels in the ipsilateral medial tibial plateau weight-bearing area was significantly lower than that of MNX+NS group(MNX+CLX group vs.MNX+NS group,5243.96 + 2507.38 vs.11153.19 +1958.72,P < 0.05).In addition,medial tibial plateau weight-bearing area AGG/ADAMTS4 expression ratio in group MNX+CLX significantly higher than MNX+NS group(MNX+CLX group vs.MNX+NS group and 0.45 + 128+ 0.63 vs.0.14,P < 0.05),and Col-II/MMP13 and Col-II/AGG two indicators among the three groups no significant difference.2 MicroCT results showed that the CLX caused no significant effect on subchondral bone of MNX guinea pig whereas SR enhanced greatly BMD of subchondral bone under medial tibial plateau cartilage in the weight-bearing region,and SR significantly improve the BV / TV,TB.Th,TB.SP and SMI morphological parameters.Conclusions:1 Medial meniscectomy could result in rapid degeneration of cartilage in medial tibial plateau from operated knee of DH guinea pigs in 12 weeks post operation,the cartilage and subchondral bone changes of this model reflect the pathological changes in early phase of OA.2 SR enhanced greatly BMD of subchondral bone under medial tibial plateau cartilage in the weight-bearing region,and SR leads to the subchondral bone morphogenetic changes significantly inhibited.But SR could not delay joint degeneration,whereas the clinical guidelines recommendmedication-celecoxib exhibits delayed effect of joint degeneration to some extent.3 Strontium Ranelate(625 mg/kg`day)fail to protect cartilage from degeneration in menisectomized guinea pigs,the reason for this result may be due to SR failed to inhibit the activity of ADAMT4 and MMP3,and then failed to stop the degradation of extracellular matrix agg.Another reason may be SR failed to inhibit chondrocyte apoptosis and subchondral bone sclerosis significantly.As a consequence of strontium accumulation,chondrocytes in osteophytes could be specifically stimulated and the chondral part of osteophyte grows and eventually forming larger osteophytes.SR intake leads to osteophytes hyperplasia,or even collapse.When osteoarthritis future research strontium ranelate when this adverse effect should take it seriously,because it may mask its therapeutic effect.