Effect Of Raloxifene On Osteoporosis-related Joint Degeneration In Ovariectomized Rats

Objectives To explore the feasibility of using bilateral ovariectomy(OVX)to mimic human menopausal changes in mature Sprague-Dawley(SD)rats,eventually inducing the prevalence of postmenopausal osteoarthritis(OA);With the early administration of raloxifene in this postmenopausal OA model,to study the effects of raloxifene on articular cartilage and subchondral bone;And to study the age-related changes of SD rats.Methods Twenty-four 6-month-old SD rats were randomly divided into four groups: Baseline group,Sham+V group,OVX+V group and OVX+RAL group.Baseline group was euthanized at the beginning of experiment.Bilateral OVX was performed in OVX+V group and OVX+RAL group,Controlled surgery with no substantive ovariectomy was performed in Sham+V group.Raloxifene was given to the rats in the OVX+RAL group at the dose of 6.2 mg/kg·day by gavage,and OVX+V group and Sham+V group were given the same amount of normal saline by gavage.Three months after administration,all rats were euthanized by 7% chloral hydrate solution,and then the right knee was harvested and fixed in 4% formaldehyde.The proximal tibial bone was prepared to evaluate subchondral bone histomorphometry by Micro-CT.After Micro-CT measurement,the right knee was decalcified for 8 weeks.Then all specimen were paraffin-embedded and sectioned.The paraffin-embedded sections were stained with toluidine blue and COL-II,Caspase-3,MMP-13 immunohistochemical staining respectively.Toluidine blue stained sections were analyzed according to OARSI histological score.The expression levels of COL-II,Caspase-3,MMP-13 were assessed respectively by immunoradioactive score.The data of each group were analyzed statistically.OVX+V group were compared with Sham+V group to verify the modeling success,and OVX+V group were compared with OVX+RAL group to observe the effects of raloxifene on the cartilage and subchondral bone of postmenopausal OA.Results 1 Almost all scores of OARSI histological score in addition to total cartilage degeneration width and medial joint capsule repair were higher in OVX+V group than in Sham+V group and OVX+RAL group,especially cartilage matrix loss width and significant cartilage degeneration width(P <0.05,respectively),indicating the significant pathological OA changes in joint in OVX+V group.All scores of OARSI histological score except the sames of cartilage matrix loss width and significant cartilage degeneration width were higher in Sham+V group than in Baseline group,but these did not reach statistical significances.2 The expression of COL-II was significantly lower in OVX+V group than in Sham+V group or OVX+RAL group(P <0.05,respectively).The expression of Caspase-3 were increased in OVX+V group than in Sham+V group or OVX+RAL group and in Sham+V group than in Baseline group,but with nonsignificant differences.The expression of MMP-13 in OVX+V group was significantly higher than that in Sham group or RAL group(P <0.05,respectively).The ratios of COL-II/Caspase-3 and COLII/MMP-13 were significantly lower in OVX+V group than in Sham group or RAL group(P <0.05,respectively).The ratio of COL-II/MMP-13 was higher in Baseline group than Sham+V group(P <0.05).The COL-II/Caspase-3 ratio was not statistical significance between Baseline group and Sham+V group.3 Micro-CT analysis found that compared with Sham+V group,BV/TV,Tb.N,I.S,and BS/TV decreased and Tb.Sp and SMI increased in OVX+V group;I.S in Sham+V group were statistically higher than that in OVX+V group(P = 0.022);Tb.Sp was significantly higher in OVX+V group than in Baseline group(P= 0.034).BV/TV,Tb.N,I.S and BS/TV,however,were significantly increased in OVX+RAL group than in OVX+V group(P = 0.001,0.000,0.000 and 0.002,respectively);Tb.Sp and SMI values were significantly lower in OVX+RAL group than in OVX+V group(P = 0.012 and 0.002,respectively).BV/TV,Tb.N,I.S and BS/TV were increased in Sham+V group than in Baseline group,Tb.Sp and SMI values were lower,but with nonsignificant differences.Conclusions 1 OVX can accelerate the degenerations of subchondral bone and articular cartilage,which proves that 6-month-old OVX rats are mild animal model to mimic the occurrence and development of postmenopausal OA.2 Raloxifene that were administrated by gavage for 3 months can delay the degenerations of subchondral bone and articular cartilage of postmenopausal OA,and may improve OVX-induced joint capsule changes,and eventually may delay the disease progression of OA from the joint organs as a whole.3 With the increase of age in adult rats,the degenerations of subchondral bone and articular cartilage increase gradually.