| Objective: The purpose of this study was to observe the changes in the local morphology and repair-related factors of the patella-patella tendon junction(PPTJ)with the accumulation of jump load,and to clarify the characteristics of changes in adaptation-related factors caused by load accumulation.To provide scientific basis for preventing the injury of the bone-tendon junction by revealing the adaptation process of the bone-tendon junction to load.Methods: Forty female New Zealand white rabbits(18 weeks old),both two hindlimb patellar-patella tendon complexes(n=80)were involved.They were randomly divided into a control(CON)group(n=10)and a single time jump training(S-JUMP)group(n=10)and multiple time jump training(M-JUMP)group(n=60).The M-JUMP group was divided into a 3-day jump(3D-JUMP)group(n=10),a 5-day jump(5D-JUMP)group(n=10),and a 10-day jump(10D-JUMP)group(n=10),20-day jump(20D-JUMP)group(n=10),30-day jump(30D-JUMP)group(n=10),and40-day jump(40D-JUMP)group(n=10).The first study involved the CON group and the S-JUMP group,and the second study involved the CON group and the M-JUMP group.The rabbits were sacrificed after 24 hours after completing the last training,then the PPTJ was fixed,decalcified,dehydrated,and embedded,and then made into paraffin sections for H?E,immunohistochemistry,and immunofluorescence staining.Observation indicators include tissue morphology,CD,FZT;the expression of TGF-?1,Smad2,Tgif-1,Sno N,SCXA,SOX-9,TNF?,COX-2,PTGES-2,IL-6,MMP-1,MMP-9,TIMP-1,TIMP-2,COL-I,COL-II,and COL-III;the ratio of SCXA/SOX-9,TIMP-1/MMP-1,TIMP-1/MMP-9,TIMP-2/MMP-9,and COL-I/COL-III.Statistical method: two-way anova was used for analysis the difference in groups,the Pearson correlation coefficient was used for the relationship between eachindex,and the significance level was p<0.05.Results:(1)Histological results.Compared with the CON group,the cells in PPTJ of the S-JUMP group were unevenly distributed,distributed in bands,local cells were aggregated,the tide line was blurred,the collagen fibers became thick,and some collagen fibers appeared micro-fracture;3D-JUMP group,5D-JUMP group,10D-JUMP group and 20D-JUMP group PPTJ showed uneven cell distribution,abnormal aggregation,increased CD,blurred to disappeared tide lines,and damaged collagen structure;30D-JUMP group PPTJ showed a repair period characterized by the repair of collagen structure;40D-JUMP group PPTJ showed some changes in adapt to load,accompanied by the characteristics of poor healing,indicating failure of adaptation;Compared with the CON group,CD and FZT in all jumping groups increased significantly(all p<0.05).(2)Effect on the expression of PPTJ cartilage proliferation and differentiation-related factors.Compared with the CON group,the expressions of Tgif-1 and Sno N were up-regulated in the S-JUMP group(all p<0.05);the expressions of TGF-?1 and Tgif-1were up-regulated in the 3D-JUMP group(all p<0.05);the expression of Sno N was up-regulated(p<0.05)in the 5D-JUMP group;the expression of TGF-?1 and Sno N was up-regulated in the 10D-JUMP group(all p<0.05);the expression of TGF-?1 and Sno N was up-regulated in the 20D-JUMP group(all p<0.05);the expression of TGF-?1 and Sno N was up-regulated in the 30D-JUMP group(all p<0.05);the expression of TGF-?1 was up-regulated in the 40D-JUMP group(p<0.05).Compared with the S-JUMP group,TGF-?1 was up-regulated in the 20D-JUMP group and30D-JUMP group(all p<0.05).The comparison results within the M-JUMP group showed that TGF-?1 reached the peak in the 30D-JUMP group,and showed a trend of first increasing and then decreasing during the 40-days jump load accumulation.(3)Effect on the expression of transcription factors related to PPTJ cartilage proliferation and differentiation.Compared with the CON group,SCXA expression was up-regulated in the S-JUMP group,3D-JUMP group,5D-JUMP group,10D-JUMP group,30D-JUMP group and 40D-JUMP group(all p<0.05);SOX-9expression in S-JUMP group,3D-JUMP group,10D-JUMP group,30D-JUMP group and 40D-JUMP group was up-regulated(all p<0.05).Compared with the S-JUMP group,SOX-9 was down-regulated in the 5D-JUMP group(p<0.05);SCXA was down-regulated in the 20D-JUMP group(p<0.05).(4)Effect on the expression of factors related to ECM metabolism.Compared with CON group,TNF?,IL-6 expression in the 3D-JUMP group was up-regulated(both p<0.05);TNF?,IL-6 expression in the 5D-JUMP group was up-regulated(both p<0.05);the expression of TNF?,COX-2,PTGES-2 and IL-6 was up-regulated(all p<0.05)in the 10D-JUMP group;the expression of TNF?,COX-2,PTGES-2,and IL-6 in the 20D-JUMP group was up-regulated(all p<0.05);the expression of COX-2in the 30D-JUMP group was up-regulated(p<0.05);the expression of COX-2 and PTGES-2 in the 40D-JUMP group was up-regulated(both p<0.05).Compared with the S-JUMP group,COX-2 was down-regulated in the 3D-JUMP and the 5D-JUMP groups(both p<0.05);TNF? and PTGES-2 were up-regulated in the 10D-JUMP group(both p<0.05).The comparison results within the M-JUMP group showed that the expression of TNF? in the 10D-JUMP group was significantly higher than that in the 5D-JUMP,30D-JUMP and 40D-JUMP groups(all p<0.05);the expression of COX-2 in the 10D-JUMP group was significantly higher than that in the 3D-JUMP group,5D-JUMP group and 40D-JUMP group(all p<0.05);the expression of PTGES-2 in 10D-JUMP group,20D-JUMP group,30D-JUMP group and 40D-JUMP group was significantly higher than that in 3D-JUMP group and 5D-JUMP group(all p<0.05);the expression of IL-6 in the 3D-JUMP group was significantly higher thanthat in the 20D-JUMP group,30D-JUMP group and 40D-JUMP group(all p<0.05).(5)Effect on the expression of enzymes related to ECM metabolism.Compared with the CON group,TIMP-1/MMP-1 and TIMP-1/MMP-9 increased in the S-JUMP group(both p<0.05);TIMP-1/MMP-1,TIMP-1/MMP-9 increased(all p<0.05)in the3D-JUMP group;TIMP-1/MMP-1,TIMP-1/MMP-9 increased(all p<0.05)in the5D-JUMP group;MMP-1,TIMP-2 expression was inhibited in the 10D-JUMP group(all p<0.05);20D-JUMP group TIMP-2/MMP-9 decreased(all p<0.05);MMP-1expression was inhibited,and TIMP-1 was activated(all p<0.05)in the 30D-JUMP group.Compared with the S-JUMP group,TIMP-1/MMP-1 and TIMP-1/MMP-9decreased in the 10D-JUMP group,20D-JUMP group,30D-JUMP group and40D-JUMP group(all p<0.05).The comparison results within the M-JUMP group showed that TIMP-1/MMP-1 and TIMP-1/MMP-9 in the 3D-JUMP group and5D-JUMP group were significantly higher than those in the 10D-JUMP group,20D-JUMP group,30D-JUMP group,and 40D-JUMP group(both p<0.05).(6)Effect on the mainly collagen expression of ECM.Compared with the CON group,the proportion of COL-I and COL-I/COL-III in the S-JUMP group increased(both p<0.05);the expression of COL-I and COL-II in the 3D-JUMP group increased(both p< 0.05),the proportion of COL-III decreased(p<0.05);the proportion of COL-I and COL-II in the 5D-JUMP group increased(both p<0.05),and the proportion of COL-I/COL-III increased(p<0.05),the expression of COL-I and COL-III was up-regulated in the 10D-JUMP group(both p<0.05);the expression of COL-I and COL-II in the 20D-JUMP group was up-regulated(both p<0.05);the expression of the COL-I in the 30D-JUMP group was up-regulated(p<0.05),and COL-I/COL-III increased(p<0.05);the proportion of COL-II,and COL-I/COL-III increased(p<0.05)in the 40D-JUMP group.Conclusion:(1)The accumulation of jump load caused the imbalance betweensynthesis and degradation of PPTJ collagen in rabbits,then the expression level of inflammatory factors increased,but the histology did not show corresponding inflammatory response characteristics,which called para-inflammatory change,and that's also the local adaptation process to load accumulation.(2)Factors affecting the local adaptation process included increasing the expression level of TIMPs,decreasing the expression level of MMPs,and increasing the expression of SCXA and/or SOX-9 to increase type I collagen and/or type II collagen expression.(3)During the accumulation of jump loads,the promotion of MMPs by TNF?,COX-2,and PTGES-2 was insufficient to resist the promotion of TIMPs by TGF-?1 and/or IL-6 during 3?30 days jump load accumulation,ECM degradation metabolism suppressed;with the adaptation to the load,ECM degradation metabolism increased,and ECM synthesis and degradation metabolism come to a new balance during 30?40days jump load accumulation. |
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