| Purpose:To investigate the effect of compound bone graft tablets on tendon-bone interface healing of ACL tendon reconstruction in rabbits,and to observe and analyze the possible mechanism of compound bone graft tablet.Materail and Method:In this experiment,36 Japanese white rabbits were selected,and10 other rabbits were selected as tendon donors.According to the random number table method,9 were selected as D-d group(sham operation group),the other 27 rabbits were treated by the same surgeon with anterior cruciate ligament dissection and modified anterior cruciate ligament reconstruction with treated allogeneic semitendinosus.After the rabbit was fully awake after anesthesia,three drugs a,b and c were given to the rabbit by gavage.The three drugs a,b and c were all configured by the third person not related to the experiment,so as to implement double blindness.At the same time,they were divided into groups a,b and c,and labeled as groups A-a,B-b and C-c.At 4,8 and 12 weeks after the gavage,3 rabbits in each group were randomly selected and sacrificed in batches,rabbit venous blood was used for PINP,TRACP-5b,C-RP and WBC counts.Rabbit knee specimens were taken to observe the bone tunnel expansion rate through Micro-CT,and the biomechanics test was performed to detect the ultimate tensile load of the post-transplant ligament.Then HE and Masson histological staining were performed on the samples,and the tendon-bone tunnel healing at each time point in each group.Results:1.General Observation This research adopts the factorial design method to observe the number of cases in each group to reduce to a minimum,save the sample size,although there are 3 rabbits dead after building models,but does not affect the factorial design analysis results,overall,each with 4 weeks,8weeks,12 weeks time,postoperative Lachman drawer before the experiment all have different degrees of improvement.2.Blood Indicators(Table 4,5,6,7 for details)At week 4,the relative expressions of PINP,TRACP-5b,C-RP and WBC in the compound bone graft group were(64.38±0.57ng/ml,8.95±1.01ng/ml,30.01±4.43ug/ml,9.75±0.22~*10^9/L),and those in the bone peptide group(66.92±0.25ng/ml,8.56±0.44ng/ml,31.04±1.40ug/ml,11.52±0.76~*10^9/L).Placebo group(66.92±0.25ng/ml,8.48±0.25ng/ml,25.87±2.10ug/ml,13.19±1.16~*10^9/L)and sham group(62.01±0.68ng/ml,10.05±0.28ng/ml,27.11±2.00ug/ml,11.15±0.28~*10^9/L).F=2.70,p=0.07>0.05;F=2.63,p=0.076 > 0.05;F=14.13,p=0.000 < 0.01)At week 8,the relative expression levels of PINP,TRACP-5b,C-RP and WBC in the compound bone graft group were(95.23±2.26ng/ml,10.07±0.19ng/ml,40.07±1.97ug/ml,7.67±1.13~*10^9/L),and those in the bone peptide group(89.94±0.48ng/ml,13.13±0.76ng/ml,37.75±1.68ug/ml,8.94±0.47~*10^9/L).Placebo group(76.30±3.14ng/ml,12.76±0.56ng/ml,40.24±1.25ug/ml,12.21±0.49~*10^9/L),sham group(63.73±1.10ng/ml,10.48±0.79ng/ml,37.25±1.50ug/ml,8.52±0.45~*10^9/L),the relative expression of various indicators between groups(F=252.55,p=0.000<0.01).F=12.62,p=0.000>0.05;F=1.108,p=0.379 > 0.05;F=48.269,p=0.000 < 0.01)At week 12,the relative expressions of PINP,TRACP-5b,C-RP and WBC in the compound bone graft group were(73.16±0.93ng/ml,9.84±2.02ng/ml,21.47±1.28ug/ml,4.36±0.38~*10^9/L),and those in the bone peptide group(67.78±0.78ng/ml,10.79±0.04ng/ml,23.01±3.19ug/ml,4.49±0.09~*10^9/L).Placebo group(69.82±0.70ng/ml,10.47±0.27ng/ml,27.21±5.32ug/ml,8.15±0.77~*10^9/L)and sham group(62.99±1.46ng/ml,10.59±1.02ng/ml,25.66±3.26ug/ml,4.42±0.25~*10^9/L).F=0.73,p=0.546 >F=2.080,p=0.134 > 0.05;F=16.883,p=0.000 < 0.01)3.Imaging and biomechanics(Table 8 and 9 for details)By Micro-CT detection,according to the results of each bone tunnel diameter were experiencing the process of first enlarged to reduce bone tunnel group differences,expanding rate by calculation of compound bone piece group at 4,8,12 weeks compared with other groups of bone tunnel expanding rate is minimal,compared with the rest of the two groups of F values were 42.82,16.71,13.34,the difference is statistically significant;The ultimate tensile load of ligaments in each group was detected by the universal tensile instrument.As time extended,the ultimate tensile load of ligaments after transplantation increased in eachgroup,reaching the ultimate tensile level of normal ligaments at week 12.The F values of ligaments in each group at weeks 4,8 and 12 were 31.36,15.12 and 11.52,respectively,showing statistically significant differences.4.Histomorphological observation(Attached figures 7,8 for details)By HE and Masson staining showed that each group at 4,8,12 weeks,the tendon to bone interface were experiencing-the process of reshaping tissue necrosis,because this experiment for the tendon to bone at the junction of a transverse,cannot see the tendon contorts the complete,but the number of cells proliferation can observe tendon bone interface to determine level of tendon to bone healing,4 weeks,groups of tendon to bone interface inflammatory cell proliferation is more apparent,lower bone density of cell proliferation,cell arrangement(fiber contorts)is irregular,the interface between tendon and bone gap exists,can be scattered collagen fiber formation;At 8 weeks,the proliferation density of osteocytes at the tendon-bone interface of each group increased,the cell arrangement tended to be regular,and the blank area at the tendon-bone interface decreased,indicating that more collagen fibers were generated.At 12 weeks,the cells at the tendon-bone interface of each group produced a fusion phenomenon,indicating sharpey-like structure,regular arrangement of cells,and significantly increased production of collagen fibers at the tendon-bone junction.The results of the compound bone graft group were better than those of the other two groups.Conclusion:1.Compound bone graft tablets and osteopeptide tablets can promote the increase of PINP in blood and promote bone formation.At the same time,the compound bone graft tablets can also reduce the blood TRACP-5b content in the 8th week and reduce bone destruction.In general,the compound bone graft tablets have a more stable effect.2.The application of compound bone graft tablets after anterior cruciate ligament reconstruction can reduce the expansion rate of bone tunnel after reconstruction in rabbits,make the tendon-bone interface interwoven closely and firmly,and thus increase the ultimate tensile load of the transplanted tendon.3.Compound bone graft tablets can promote the formation of bone cells and the formation of sharpey-like fibers at the tendon-bone interface to promote tendon-bone healing. |
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