Early treadmill running delays rotator cuff healing via Neuropeptide Y mediated inactivation of the Wnt/β-catenin signaling

Abstract

Background Defining the optimal rehabilitation programs for rotator cuff healing remains a challenge. Early treadmill running may have negative effects on tendon-bone interface (TBI) healing with increased expression of Neuropeptide Y (NPY). However, the underlying mechanism is still unknown. Methods The mice were randomly assigned to four groups: control group, treadmill group, treadmill ​+ ​BIBO3304 group and BIBO3304 group alone. Specifically, the control group was allowed free cage activity without any treatment after surgery. The treadmill group received early treadmill running initiated from postoperative day 2. The treadmill ​+ ​BIBO3304 group received treadmill running combined with intra-articular injection of BIBO3304 postoperatively. The BIBO3304 group only received type 1 NPY receptor (Y1 receptor, Y1R) antagonist BIBO3304 postoperatively. Healing outcomes of the rotator cuff were evaluated by histological analysis, synchrotron radiation micro-computed tomography (SR-μCT) scanning, and biomechanical testing at 4 and 8 weeks after surgery. The expression of NPY and its Y1 receptor during the treadmill running were tested by immunofluorescence. In addition, the related signaling pathway of Neuropeptide Y among all groups was detected by immunohistochemistry and western-blot. Results Immunofluorescence results show that early treadmill training could lead to a significant increase in the expression of NPY at the healing site, and Y1R was widely expressed in both normal or injured rotator cuff without statistical difference. At the same time, early treadmill running delayed the healing of rotator cuff, as indicated with unsatisfactory outcomes, including a significantly lower histological score, decreased bone formation and inferior biomechanical properties at postoperative week 4 and 8. Moreover, the use of BIBO3304 could partly alleviate the negative effects of early treadmill running on the healing of rotator cuff and promote the natural healing process of rotator cuff, as evidenced by significant differences observed between the treadmill and treadmill ​+ ​BIBO3304 groups, as well as observed between the control and BIBO3304 groups. On the other hand, the expressions of Wnt3a and β-catenin in the treadmill group were significantly lower compared with the other groups, while the expression in the BIBO3304 group was the highest, as evaluated by immunohistochemistry and western-blot. Conclusions Early treadmill running increased the expression of NPY at the RC healing site, which might burden the expression of Wnt3a/β-catenin and delay the healing process, inhibition of Y1 receptor with BIBO3304 could promote bone-tendon healing through the Wnt/β-catenin signaling. The translational potential of this article: This is the first study to evaluate the specific role of the NPY-Y1R axis and its underlying mechanism by which early treadmill running delays bone-tendon healing. Further, our study...

Mechanical stimulation improves rotator cuff tendon-bone healing via activating IL-4/JAK/STAT signaling pathway mediated macrophage M2 polarization

“…Nowadays, many researchers regard loading exercise as an important part of postoperative rehabilitation following a tendon injury, while to what extent the mechanical stimulation would benefit the tendon-bone healing is controversial [ 6 , 12 , 32 ]. Recent progress confirms that short-term immobilization and delayed application of moderate mechanical loading could improve the healing quality of TBI [ 26 , 33 ], however, premature and excessive loading exercise is detrimental to T-B healing [ 25 , 34 ]. This study shows the same result as the previous one, we illustrate that appropriate mechanical stimulation loaded by treadmill running promotes the T-B healing in the RC repair model, as evidenced by enhanced osteogenesis, chondrogenesis, and biomechanical property.…”

“…The 8-week-old mature male C57BL6 mice were used to establish the rotator cuff (RC) repair model as previously reported [ [23] , [24] , [25] ]. As shown in Figure S1 , animals were firstly anesthetized with pentobarbital, and then a skin incision was made over the anterolateral side of the left shoulder.…”

Rotator cuff healing is regulated by the lymphatic vasculature

“…The C57BL/6 mice underwent left RC repair surgery as reported in previous studies [ 25 , 26 ]. After the mice were anesthetized with 0.3% pentobarbital (0.6 ml/20 ​g; Sigma–Aldrich, St. Louis, MO), a longitudinal incision was made on the skin surface of the left shoulder.…”

“…Tissue sections were observed using a light microscope (CX31, Olympus, Germany). Healing of the RC insertion was assessed by two blinded observers according to a previously reported bone-tendon healing scoring system [ 26 ], which includes: cellularity, vascularity, continuity, fibrocartilage cells, tidemark and ranked on a scale from 1 to 4 ( Table S1 ) [ 26 ]. A higher histology score indicates improved or more mature tendon-to-bone healing.…”

Intermittent fasting promotes repair of rotator cuff injury in the early postoperative period by regulating the gut microbiota

“…The scoring process was carried out in a blinded manner. The area and thickness of fibrocartilage layer and the mean optical density of proteoglycans (PG) were quantitatively measured according to a previously established protocol [ [31] , [32] , [33] , [34] ]. Briefly, the area of the fibrocartilage layer refers to the region between the proximal boundary of humerus and the distal boundary of SST.…”