低强度脉冲超声促进腱骨界面愈合的机制

The mechanism of low-intensity pulsed ultrasound accelerating tendon bone interface healing

目的探索低强度脉冲超声(LIPUS)促进腱骨界面损伤修复的作用及相关机制。方法建立大鼠(中南大学动物学部提供)肩袖损伤模型并按照随机数字表法分为LIPUS治疗组和对照组,并在术后3周和5周收取冈上肌肌腱-肱骨头复合体标本,进行组织学、免疫组织化学及生物力学检测,相关统计采用双因素方差分析。体外利用LIPUS刺激巨噬细胞并向M2型诱导,检测M2型巨噬细胞占比,收集巨噬细胞上清液干预骨髓间充质干细胞(BMSCs),检测其成骨和成软骨分化效果,组间比较采用t检验。结果在大鼠肩袖损伤模型中,术后第5周时LIPUS治疗组的腱骨界面成熟度(13.50±2.47比17.33±1.20,t=2.605,P<0.05)和生物学力性能均优于对照组(拉断负荷:49.60±6.12比34.27±5.85,t=4.950;拉断刚度:20.85±2.70比13.82±3.01,t=3.974,P<0.01)。免疫组织化学提示LIPUS治疗组大鼠肩袖损伤局部M2型巨噬细胞比例高于对照组(0.77±0.11比0.59±0.10,t=2.582,P<0.05)。体外实验也证实LIPUS组M2型巨噬细胞占比高于对照组(65.77±4.72%比42.50±3.20%t=7.068,P<0.01),其上清液促进BMSCs成骨(3.92±0.47比1.0±0.12,t=10.43,P<0.01)和成软骨(2.25±0.35比1.0±0.25,t=5.034,P<0.01)分化能力更强。结论LIPUS可能通过调节巨噬细胞向M2极化,并影响BMSCs分化,从而有效提高大鼠肩袖损伤模型腱骨界面愈合质量。

Objective To explore the effect of Low-intensity pulsed ultrasound(LIPUS)on the repair of tendon-bone interface(TBI)injury and the potential mechanism.Methods Rat rotator cuff tear models were established and randomly assigned to the LIPUS and control group.At postoperative 3 and 5 weeks,TBI specimens were harvested for histological,immunohistochemical,and biomechanical tests,and 2-way ANOVA was analyzed.In vitro,macrophages were induced for M2 macrophages,with or without LIPUS stimulation,and liquid supernatant was collected to intervene in the differentiation of bone mesenchymal stem cells(BMSCs).T tests were analyzed for these data.Results The LIPUS group had better TBI maturity than control group(13.50±2.47 vs.17.33±1.20,t=2.605,P<0.05)and biological performance at postoperative 5 week(failure load:49.60±6.12 vs.34.27±5.85,t=4.950;stiffness:20.85±2.70 vs.13.82±3.01,t=3.974,P<0.01).Immunohistochemical results showed there were more local M2-type macrophages around the TBI area in the LIPUS group(0.77±0.11 vs.0.59±0.10,t=2.582,P<0.05).The vitro experiments showed,in LIPUS group,more M2 polarization of macrophages[(65.77±4.72)%vs.(42.50±3.20)%,t=7.068,P<0.01],and better osteogenic(3.92±0.47 vs.1.0±0.12,t=10.43,P<0.01)and chondrogenic differentiation(2.25±0.35 vs.1.0±0.25,t=5.034,P<0.01)ability for BMSCs.Conclusion LIPUS was able to promote the differentiation of BMSCs by regulating the polarization of macrophages to M2,and effectively improve the quality of TBI healing in the rat rotator cuff injury model.