Cuff Injury Model Research Articles

Effect of Exosomes From Bone Marrow-Derived Mesenchymal Stromal Cells and Adipose-Derived Stromal Cells on Bone-Tendon Healing in a Murine Rotator Cuff Injury Model.

Bone-tendon injury is characterized by poor self-healing. It is established that exosomes are favorable for tissue repair and regeneration. However, their effect on bone-tendon healing has not yet been determined. To compare the effectiveness of exosomes derived from adipose-derived mesenchymal stromal cells (ADSC-Exos) and bone marrow-derived mesenchymal stromal cells (BMSC-Exos) on bone-tendon interface healing in murine rotator cuff injury model and explore the underlying mechanisms thereof. Controlled laboratory study. A total of 63 male C57BL6 mice with rotator cuff injuries underwent surgery and were randomly assigned to a control group treated without exosomes (n = 21), an ADSC-Exos group (n = 21), or a BMSC-Exos group (n = 21). The mice were sacrificed 4 or 8 weeks after surgery, and tissues were collected for histologic examination and radiographic and biomechanical testing. For exosome tracing in vivo, mice were sacrificed 7 days after surgery. A series of functional assays (radiographic evaluation, proliferation assay, Alizarin Red staining, alkaline phosphatase staining and activity, Alcian blue staining, quantitative polymerase chain reaction analyses, and glycosaminoglycans quantification) were conducted to evaluate the effect of exosomes on the cellular behaviors of the BMSCs in vitro. A statistical analysis of multiple-group comparisons was performed by 1-way analysis of variance, followed by the Bonferroni post hoc test to assess the differences between the 2 groups. The ADSCs and BMSCs were positive for surface markers CD29 and CD90 and negative for surface markers CD34 and CD45 and could differentiate into osteoblasts, chondrocytes, and adipocytes. Exosomes showed a cup- or sphere-shaped morphology and were positive for CD63 and TGS101. Local injection of ADSC-Exos and BMSC-Exos could recruit BMSCs and promote osteogenesis, chondrogenesis, and bone-tendon healing. In vitro, ADSC-Exos and BMSC-Exos could significantly promote the proliferation, migration, osteogenic differentiation, and chondrogenic differentiation ability of BMSCs. In vivo, ADSC-Exos and BMSC-Exos significantly accelerated bone-tendon injury healing, with no significant statistical difference between them. ADSC-Exos and BMSC-Exos exhibited similar therapeutic effects on bone-tendon healing in our murine animal model. ADSC-Exos and BMSC-Exos may be used to develop a new cell-free therapy method for promoting rotator cuff injury repair.

Development of a Rabbit Chronic-Like Rotator Cuff Injury Model for Study of Fibrosis and Muscular Fatty Degeneration

Rabbit rotator cuff (RC) pathophysiology can lead to progressive and highly degenerative changes in its associated musculature and tendons, which negatively influences clinically relevant parameters, such as strength and retraction of the muscle-tendon/myotendinous unit, ultimately causing loss of shoulder function and negatively affecting RC repair outcomes. Animal models that mimic aspects of human RC anatomy and pathophysiology are crucial for advancing the conceptual understanding of injury progression and developing effective tissue engineering and regenerative medicine-based therapeutics. Within this context, a rabbit subscapularis (SSC) model is suitable due to (i) its anatomical similarity to the human supraspinatus (SSP) bone-tendon-muscle unit, which is the most frequently injured RC site; (ii) its pathophysiological similarity to humans in terms of fibrosis and muscle fatty degeneration (FD); and (iii) its amenability to surgical procedures. Therefore, the goal of this study is to describe the surgical techniques for inducing SSC RC injury. Briefly, the procedure involves the isolation of the SSC by identifying the coracobrachialis muscle followed by a full-thickness transection at the muscle-tendon junction and wrapping the free end of the muscle-tendon junction with a silicone-based penrose tubing to prevent spontaneous reattachment. Histologic evaluations are performed to monitor the progression of muscle FD at 4 weeks post-surgery using hematoxylin and eosin (H&E) as well as Masson's trichrome staining. Loss of muscle and FD were evident 4 weeks after transection of the SSC muscle-tendon junction, similar to human RC pathophysiological conditions. This protocol demonstrates the steps for successfully establishing a chronic-like rabbit SSC RC injury model, which can serve as a powerful tool to study skeletal muscle changes associated with RC pathophysiology and aid the development of novel therapeutic strategies for chronic-like RC tears.

Assessment of a Synergistic Effect of Platelet-Rich Plasma and Stem Cell-Seeded Hydrogel for Healing of Rat Chronic Rotator Cuff Injuries

Outcomes after repair of chronic rotator cuff injuries remain suboptimal. Type-1 collagen-rich tendon hydrogel was previously reported to improve healing in a rat chronic rotator cuff injury model. Stem cell seeding of the tendon hydrogel improved bone quality in the same model. This study aimed to examine whether there was a synergistic and dose-dependent effect of platelet-rich plasma (PRP) on tendon–bone interface healing by combining PRP with stem cell–seeded tendon hydrogel. Human cadaveric tendons were processed into a hydrogel. PRP was prepared at two different platelet concentrations: an initial concentration (initial PRP group) and a higher concentration (concentrated PRP group). Tendon hydrogel was mixed with adipose-derived stem cells and one of the platelet concentrations. Methylcellulose, as opposed to saline, was used as a negative control due to comparable viscosity. The supraspinatus tendon was detached bilaterally in 33 Sprague-Dawley rats (66 shoulders). Eight weeks later, each detached tendon was repaired, and a hydrogel mixture or control was injected at the repair site. Eight weeks after repair, shoulder samples were harvested and assigned for biomechanical testing (n = 42 shoulders) or a combination of bone morphological and histological assessment (n = 24 shoulders). Biomechanical testing showed significantly higher failure load and stiffness in the concentrated PRP group than in control. Yield load in the initial and concentrated PRP groups were significantly higher than that in the control. There were no statistically significant differences between the initial and concentrated PRP groups. The addition of the highly concentrated PRP to stem cells–seeded tendon hydrogel improved healing biomechanically after chronic rotator cuff injury in rats compared to control. However, synergistic and dose-dependent effects were not seen.

Poster 130: Tendon-to-bone Healing in a CCR2 Knockout Mouse Model of Delayed Rotator Cuff Repair

Poster 130: Tendon-to-bone Healing in a CCR2 Knockout Mouse Model of Delayed Rotator Cuff Repair

Enhanced Repaired Enthesis Using Tenogenically Differentiated Adipose-Derived Stem Cells in a Murine Rotator Cuff Injury Model.

Rotator cuff tear (RCT) is among the most common shoulder injuries and is prone to rerupture after surgery. Selecting suitable subpopulations of stem cells as a new specific cell type of mesenchymal stem cells has been increasingly used as a potential therapeutic tool in regenerative medicine. In this study, murine adipose-derived SSEA-4+CD90+PDGFRA+ subpopulation cells were successfully sorted, extracted, and identified. These cells showed good proliferation and differentiation potential, especially in the direction of tendon differentiation, as evidenced by qRT-PCR and immunofluorescence. Subsequently, we established a murine rotator cuff injury model and repaired it with subpopulation cells. Our results showed that the subpopulation cells embedded in a fibrin sealant significantly improved the histological score, as well as the biomechanical strength of the repaired tendon enthesis at four weeks after surgery, compared with the other groups. Hence, these findings indicated that the subpopulation of cells could augment the repaired enthesis and lead to better outcomes, thereby reducing the retear rate after rotator cuff repair. Our study provides a potential therapeutic strategy for rotator cuff healing in the future.

3D bioprinting of multilayered scaffolds with spatially differentiated ADMSCs for rotator cuff tendon-to-bone interface regeneration

Regeneration of the gradient structure of the tendon-to-bone interface is still a significant clinical challenge. This study reports a novel therapeutic method combining three-dimensional (3D) bioprinting and melt electrospinning writing techniques to regenerate a functional tendon-to-bone interface. We generated biomimetic multilayered scaffolds with 3D-bioprinted pre-differentiated autologous adipose-derived mesenchymal stem cells (ADMSC), which recapitulated compositional and cellular structures of the interface. The hydrogel-based bioinks offered high cell viability and proliferative capability for rabbit ADMSCs. The hydrogels with pre-differentiated (into tenogenic, chondrogenic, and osteogenic lineages) or undifferentiated rabbit ADMSCs were 3D-bioprinted into zonal-specific constructs to mimic the structure of the tendon-to-bone interface. These scaffolds were tested in a rabbit rotator cuff injury model and the histological, radiological, and biomechanical changes were analyzed. The in vivo studies demonstrated that the scaffold with spatially differentiated autologous ADMSCs had a superior histological score and improved collagen organization when compared to acellular scaffolds and similar T2 value as the normal interface tissue. The biomechanical characterization demonstrated that the application of multilayered scaffolds improved the biomechanical properties of the tendon-to-bone interface at 12 weeks after rotator cuff reconstruction surgery, but the incorporation of autologous ADMSCs within the multilayered scaffolds showed a limited contribution. Thus, our work provides a 3D-bioprinting-based strategy with the application of autologous ADMSCs to reconstruct massive rotator cuff tendon tears.

A Dual-Factor Releasing Hydrogel for Rotator Cuff Injury Repair

Rotator cuff injury causes pain in the shoulder and is a challenge to be repaired even after surgical reconstruction. Here, we developed a dual-factor releasing hydrogel based on sulfhydrylated chitosan to deliver KGN and FGF-2 to the injured area to enable fast healing of the tendon–bone interface, which is essential for the repair of rotator cuff injury. We found that the two factors could be easily loaded into the hydrogel, which could in turn continuously release the factors in physiological conditions. The hydrogel was found to be a porous structure through a scanning electron microscope (SEM). The micropores in the hydrogel structure enable the loading and releasing of these molecules. This study showed that KGN and FGF-2 could play a synergistic effect by recruiting and promoting stem cell proliferation and chondrogenesis, thus accelerating the healing of the tendon–bone interface. An in vivo study based on a rabbit rotator cuff injury model demonstrated that the dual-factor releasing hydrogel possesses superior repair capacity than a single-factor releasing hydrogel and the untreated groups. In conclusion, the KGN and FGF-2 dual-factor releasing hydrogel could be a promising biomaterial for the regeneration of the tendon–bone interface and rotator cuff injury repair.

Relationship of superoxide dismutase to rotator cuff injury/tear in a rat model.

Rotator cuff degeneration is one of the several factors that lead to rotator cuff tears. Oxidative stress and superoxide dismutase have been reported to be related to rotator cuff degeneration; however, the precise mechanism still remains unclear. In this study, we investigated the relationship of oxidative stress and superoxide dismutase to the degeneration of the rotator cuff using rat models. Eighty-four rats were used to create a collagenase-induced rotator cuff injury model (injury model) and a rotator cuff tear model (tear model). The controls were administered saline and had only a deltoid incision, respectively. We evaluated degeneration morphology of the rotator cuff using a degeneration score; dihydroethidium fluorescence intensity, which detects oxidative stress; gene expression; and superoxide dismutase activity. The rotator cuffs in the injury and tear models significantly increased degeneration scores and dihydroethidium fluorescence intensity. On the other hand, gene expression of superoxide dismutase isoform, superoxide dismutase 1, and superoxide dismutase activity were significantly decreased in the injury model but showed no significant difference in the tear model. These findings suggested that superoxide dismutase might not be associated with rotator cuff degeneration after tear but may be involved in degenerative rotator cuff without tear. However, we found that rotator cuff degeneration involves oxidative stress both with and without tear. Based on these findings, it is presumed that different treatments may be appropriate, depending on the state of rotator cuff degeneration, because the mechanisms of the degeneration may be different.

Effect of Exercise Intensity on the Healing of the Bone-Tendon Interface: A Mouse Rotator Cuff Injury Model Study

Background: Injuries at the bone-tendon interface (BTI) are common findings in clinical practice. Rehabilitation procedures after BTI surgery are important but are controversial. Purpose: To investigate the effects of different exercise intensities on BTI healing by means of an established mouse rotator cuff injury model. Study Design: Controlled laboratory study. Methods: A total of 150 specific pathogen free male C57BL/6 mice, with supraspinatus insertion injury, were randomly assigned to 1 of 5 groups according to postoperative rehabilitation of different exercise intensities: (1) control group, (2) low-intensity exercise group, (3) moderate-intensity exercise group, (4) high-intensity exercise group, and (5) increasing-intensity exercise group (IG). The specimens were harvested 4 or 8 weeks postoperatively for microarchitectural, histological, molecular biological, and mechanical evaluations. Results: Histological test results showed that the degrees of tissue fusion and polysaccharide protein distribution at the healing interface at 4 and 8 weeks after surgery were significantly better in the IG than in the other 4 groups. Synchrotron radiation micro–computed tomography showed that the quantity of subchondral bone at the enthesis (bone volume/total volume fraction, trabecular thickness, trabecular number) was higher and trabecular separation was lower in the IG than in the other 4 groups. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis revealed that the healing interface in the IG expressed more transcription factors, such as sox 9, runx 2, and scleraxis, than the interfaces in the other groups. Although no significant difference was seen in the cross-sectional area between the groups at postoperative weeks 4 and 8 (P > .05), the tensile load, ultimate strength, and stiffness of the specimens in the IG were significantly better than those in the other 4 groups (P < .05). Conclusion: The rehabilitation program with increasing-intensity exercise was beneficial for BTI healing. Clinical Relevance: The results of this study provide evidence supporting the use of a simple and progressive exercise rehabilitation program after rotator cuff surgery.

Adipose-derived stem cellexosomes facilitate rotator cuff repair by mediating tendon-derived stem cells.

Aim: To evaluate the potential capability of adipose-derived stem cellexosomes (ADSC-exos) on rotator cuff repair by mediating the tendon-derived stem cells (TDSCs) and explored the mechanism. Methods: First, we investigated the growth, survivaland migration of TDSCs in the presence of ADSC-exos in vitro. Using a rat rotator cuff injury model to analyze the ability of the ADSC-exos to promote rotator cuff healing in vivo. Results: The hydrogel with ADSC-exos significantly improved the osteogenic and adipogenesis differentiation and enhanced the expression of RUNX2, Sox-9, TNMD, TNCand Scxand the mechanical properties of the articular portion. Conclusion: The ADSC-exos have the potential to promote the rotator cuffrepair by mediating the TDSCs.

Dual-modal magnetic resonance and photoacoustic tracking and outcome of transplanted tendon stem cells in the rat rotator cuff injury model

Stem cells have been used to promote the repair of rotator cuff injury, but their fate after transplantation is not clear. Therefore, contrast agents with good biocompatibility for labeling cell and a reliable technique to track cell are necessary. Here, we developed a micron-sized PLGA/IO MPs to label tendon stem cells (TSCs) and demonstrated that PLGA/IO MPs were safe and efficient for long-term tracking of TSCs by using dual-modal MR and Photoacoustic (PA) imaging both in vitro and in rat rotator cuff injury. Moreover, TSCs improved the repair of injury and the therapeutic effect was not affected by PLGA/IO MPs labeling. We concluded that PLGA/IO particle was a promising dual-modal MR/PA contrast for noninvasive long-term stem cell tracking.

Localized delivery of ibuprofen via a bilayer delivery system (BiLDS) for supraspinatus tendon healing in a rat model

The high prevalence of tendon retear following rotator cuff repair motivates the development of new therapeutics to promote improved tendon healing. Controlled delivery of non-steroidal anti-inflammatory drugs to the repair site via an implanted scaffold is a promising option for modulating inflammation in the healing environment. Furthermore, biodegradable nanofibrous delivery systems offer an optimized architecture and surface area for cellular attachment, proliferation, and infiltration while releasing soluble factors to promote tendon regeneration. To this end, we developed a bilayer delivery system (BiLDS) for localized and controlled release of ibuprofen (IBP) to temporally mitigate inflammation and enhance tendon remodeling following surgical repair by promoting organized tissue formation. In vitro evaluation confirmed the delayed and sustained release of IBP from Labrafil-modified poly(lactic-co-glycolic) acid microspheres within sintered poly(ε-caprolactone) electrospun scaffolds. Biocompatibility of the BiLDS was demonstrated with primary Achilles tendon cells in vitro. Implantation of the IBP-releasing BiLDS at the repair site in a rat rotator cuff injury and repair model led to decreased expression of proinflammatory cytokine, tumor necrotic factor-α, and increased anti-inflammatory cytokine, transforming growth factor-β1. The BiLDS remained intact for mechanical reinforcement and recovered the tendon structural properties by 8 weeks. These results suggest the therapeutic potential of a novel biocompatible nanofibrous BiLDS for localized and tailored delivery of IBP to mitigate tendon inflammation and improve repair outcomes. Future studies are required to define the mechanical implications of an optimized BiLDS in a rat model beyond 8 weeks or in a larger animal model.

Cowpea-like bi-lineage nanofiber mat for repairing chronic rotator cuff tear and inhibiting fatty infiltration

Abstract The repair of chronic rotator cuff injury primarily depends on surgery to biomechanically strengthen the connection between the tendon and bone. There are numerous studies on tendon-to-bone healing to promote bone regeneration or collagen reconstruction in tendons. However, chronic rotator cuff injuries are often associated with fat infiltration, and few studies have taken this aspect into account, let alone overcome the two challenges of bone healing and fat infiltration at the same time. Here, inspired by the structure of cowpea (Vigna unguiculata), we have developed lithium-containing mesoporous silica combined with electrospun PEUU nanofibers (Li+@MSNs/PEUU), which can slowly release lithium ions, inhibiting rotator cuff fat infiltration, and promoting bone regeneration to enhance tendon-to-bone healing on the basis of surgical repair. Using in vitro experiments we studied the mechanism of lithium ions playing a two-way role, which was found to be closely related to the Wnt pathway. In vivo, the system showed significant success in the treatment of chronic rotator cuff injury model in rats. Both histological and micro-CT results confirmed that the healing condition of the Li+@MSNs/PEUU nanofibrous mat group was better than that of the other control groups after 8 weeks. Overall, this work shows that lithium-containing mesoporous silica-bound electrospun nanofibrous mat have a long-term effect on promoting postoperative healing of rotator cuff injury with fat infiltration, and the regulation of the two lineages is achieved using only a single medium, leading to good potential for application.

Enhancement of tendon-bone healing after rotator cuff injuries using combined therapy with mesenchymal stem cells and platelet rich plasma.

The injuries of rotator cuff will cause the shoulder dysfunctions. Due to limited self-regeneration abilities of the tendon-bone part, rotator cuff injuries remain a clinical challenge. Previous studies have proposed many strategies for treating this disease. In this work, we aimed to combine different strategies to achieve better beneficial effects on tendon-bone repair. We isolated mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) and tested the effects of PRP on the gene expression, cell death resistance, and osteogenic differentiation of MSCs. Then, we utilized multiple strategies to treat rotator cuff injuries. We evaluated the expression of genes that related to tissue repair, bone formation, and tendon regeneration. We also tested the biomechanical property of repair tissues. We found that the in-vitro co-culture with PRP endowed MSCs with enhanced production of growth factors, better osteogenic differentiation ability, and stronger ability to resist cell death. Next, we applied MSCs, PRP, and MSCs-PRP combined therapies in rat rotator cuff injury model to compare their therapeutic effects in vivo. Through biomechanical testing, we found that the combined therapy was most efficient to promote tissue regeneration and enhance the biomechanical property of the newly generated bone. The combined treatment induced strongest signals related to angiogenesis, bone formation, and tendon generation in-situ. We demonstrated that the combination of MSCs and PRP synergistically promotes tendon-bone healing and holds great promise for the treatment of rotator cuff injuries.

Human Tendon–Derived Collagen Hydrogel Significantly Improves Biomechanical Properties of the Tendon-Bone Interface in a Chronic Rotator Cuff Injury Model

Poor healing of the tendon-bone interface (TBI) after rotator cuff (RTC) tears leads to high rates of recurrent tear following repair. Previously, we demonstrated that an injectable, thermoresponsive, type I collagen-rich, decellularized human tendon-derived hydrogel (tHG) improved healing in an acute rat Achilles tendon injury model. The purpose of this study was to investigate whether tHG enhances the biomechanical properties of the regenerated TBI in a rat model of chronic RTC injury and repair. Tendon hydrogel was prepared from chemically decellularized human cadaveric flexor tendons. Eight weeks after bilateral resection of supraspinatus tendons, repair of both shoulders was performed. One shoulder was treated with a transosseous suture (control group) and the other was treated with a transosseous suture plus tHG injection at the repair site (tHG group). Eight weeks after repair, the TBIs were evaluated biomechanically, histologically, and via micro-computed tomography (CT). Biomechanical testing revealed a larger load to failure, higher stiffness, higher energy to failure, larger strain at failure, and higher toughness in the tHG group versus control. The area of new cartilage formation was significantly larger in the tHG group. Micro-CT revealed no significant difference between groups in bone morphometry at the supraspinatus tendon insertion, although the tHG group was superior to the control. Injection of tHG at the RTC repair site enhanced biomechanical properties and increased fibrocartilage formation at the TBI in a chronic injury model. Treatment of chronic RTC injuries with tHG at the time of surgical treatment may improve outcomes after surgical repair.

Dual-modal magnetic resonance and photoacoustic tracking of tendon stem cell labeled with PLGA/IO MPs

Objective To detect the efficiency of the newly developed PLGA/IO MPs in tracking tendon stem cells (TSCs) by magnetic resonance (MR) and photoacoustic (PA) imaging. Methods Both PLAG/IO MPs and TSCs were prepared and acquired according to the previous study, and TSCs were incubated with PLGA/IO MPs for labeling.TSCs were collected for MR and PA imaging, prussian blue staining was performed, and the iron concentration of labeled TSCs was determined using inductively coupled plasma optical emission spectrometry (ICP-OES) at 3, 7, 14, 21 and 28 days after labeling respectively. The rotator cuff injury model was built on the right side of SD rats by surgery and the labeled TSCs were implanted instantly. Dual-modal MR/PA imaging was performed to observe the implanted labeled TSCs at day 3, 7, 14, 21 and 28 after implantation respectively. Results Along with the increase of labeling time, both MR and PA signal of labeled TSCs decreased gradually, and the amount of intracellular Fe loading was gradually decreased. At day 28, the difference of Fe concentration per cell between labeled TSCs and non-labeled TSCs was not significant (1.45 pg Fe/cell vs 1.17 pg Fe/cell, P>0.05). MR and PA imaging allowed a long-term tracking of labeled TSCs for 21 and 7 days respectively in the rat rotator cuff injury model. Conclusions PLGA/IO MPs are able to label TSCs for up to 21 days, and dual-modal MR/PA imaging could be used to track the labeled TSCs in the rat rotator cuff injury model. Key words: Tendon stem cells; Cell tracking; Magnetic resonance imaging; Photoacoustic imaging

Human Tendon-Derived Hydrogel Improves Healing of Chronic Tendon–Bone Interface Injuries

Grant support received from: 2016 AFSH Basic Science Grant There is no financial information to disclose. Previously our laboratory reported the positive impact of thermoresponsive, type 1 collagen–rich, decellularized human tendon-derived hydrogel (tHG) on tendon healing using a rat Achilles tendon acute injury model. We hypothesized tHG would facilitate healing of chronic injuries at tendon–bone interface (TBI) in a rotator cuff model. Rotator cuff tears are important causes of pain and disability of upper extremity and involve injury at TBI. In this study a rat chronic rotator cuff injury model was used to evaluate TBI healing after repair. Twenty Sprague–Dawley rats underwent bilateral detachment of supraspinatus (SSp) tendons. Eight weeks later, both SSp tendons were anatomically reattached to humerus. One shoulder underwent a suture to bone repair using bone tunnels (control group, n = 20) and the other underwent the same repair plus tHG injection at repair site (tHG group, n = 20). Eight weeks after repair, SSp tendon–muscle units with humeral bone were harvested after animal death. Biomechanical tests were performed on 15 samples in each group. Five additional samples in each group were evaluated via micro CT, followed by histological examinations. Paired t tests were performed for statistical analyses. Biomechanical testing revealed significantly larger load to failure (27.5 ± 11.8N vs 19.7 ± 5.4N, P = .03) and higher stiffness (15.7 ± 6.3 N/mm vs 11.5 ± 3.5 N/mm, P = .02) in the tHG group versus control. Safranin O staining showed the significantly larger area of new cartilage formation in the tHG group (0.48 ± 0.1 mm2 vs 0.32 ± 0.06 mm2, P = .03). Based on birefringence under polarized microscopy in Picosirius red–stained slides collagen fiber organization was greater in tHG group compared with the control (40.4 ± 9.8 gray scale vs 31.1 ± 4.2 gray scale, P = .07), although it did not significantly differ. Micro CT demonstrated higher bone mineral density (296.1 ± 21.9 mg/cm3 vs 274.6 ± 17.2 mg/cm3, P = .05) and larger trabecular number (1.7 ± 0.1/mm vs 1.5 ± 0.2, P = .07) at the supraspinatus insertion area of the humerus in tHG group than those in the control, although the difference did not reach statistical significance. •Injection of collagen-rich tHG at the rotator cuff repair site statistically significantly enhanced biomechanical properties and increased fibrocartilage formation at the TBI in a chronic injury model.•An improvement collagen fiber organization, higher bone mineral density, and larger trabecular number were also seen in the tHG group versus control.•This human-derived collagen gel is a promising scaffold to augment healing of chronic TBI injuries during surgical intervention. This research was supported by a Basic Science Grant from the American Foundation for Surgery of the Hand.

Human Amniotic Membrane Improves Healing in a Chronic, Massive Rotator Cuff Repair Model

Objectives:Rotator cuff tendon heals by fibrovascular scar that is weaker than native tissue leading to repairs that are prone to failure. Our objective was to investigate the ability of an amniotic membrane-derived human allograft to improve rotator cuff tendon-bone healing and skeletal muscle architecture in a chronic massive rotator cuff injury and repair model in rats. We hypothesized that application of human amniotic membrane to the tendon-bone interface at the time of repair will result in increased attachment strength and rotator cuff muscle fiber force secondary to improved bone formation and nthesis histomorphometry, less plasminogen activator inhibitor-1 (PAI-1) expression, and induced muscle repair gene expression.Methods:Eighty-five male Sprague-Dawley rats were allocated to one of four groups: (1) uninjured (U), (2) injury only (IO), (3) chronic injury and repair (CR), or (4) chronic injury and augmented repair (ER). The IO, CR, and ER groups underwent unilateral detachment of the supraspinatus and infraspinatus tendons with injection of the corresponding muscle with 3 u/kg Onabotulinum toxin A. Four weeks following injury, a surgical repair was performed in the CR and ER groups using transosseous suture fixation with human amniotic membrane applied to the tendon-bone repair site in the ER group. Animals were euthanized at four weeks following the repair. Biomechanical testing of the supraspinatus and infraspinatus tendon-bone complex and single fiber contractility testing of the supraspinatus muscle were performed. Microcomputed tomography was utilized to quantitate bone microstructure at the repair site. The healing tendon-bone interface was evaluated with histomorphometric quantification of fibrocartilage formation and collagen organization, and immunostaining with PAI-1. The rotator cuff muscle was evaluated with immunohistochemical localization for lipid deposition and total cross-sectional area. Real-time polymerase chain reaction evaluated relative expression of genes relevant to muscle repair. Statistical analysis was performed using a nonparametric Kruskal-Wallis test with significance set at p<0.05.Results:Augmentation with amniotic membrane did not lead to increased load-to-failure or stiffness compared to CR. The cross-sectional area, maximum isometric force, and specific force of individual supraspinatus muscle fibers were significantly greater with ER compared to CR (Figure 1). Microcomputed tomography revealed a larger volume of newly formed bone present at the bony footprint with ER compared to CR. Histomorphometric analysis demonstrated significantly greater fibrocartilage area and collagen organization at the healing tendon-bone insertion site with ER compared to CR at 4 week. Qualitatively, there was less PAI-1 immunostaining noted at the tendon-bone interface in the ER group. There was less lipid deposition and greater cross-sectional area of the rotator cuff muscle fibers in the ER group. Rotator cuff muscle gene expression with ER demonstrated a downregulation of apoptosis regulatory genes BCL2 and Caspase-3, adipogenesis regulatory genes FITM1 and FITM2, TNF-alpha and TGF-beta 3 compared to CR.Conclusion:Augmentation with human amniotic membrane in a chronic, massive rotator cuff injury and repair model led to greater bone formation, more organized tendon enthesis, induced muscle repair gene expression, and greater rotator cuff muscle fiber force secondary to less PAI-1 expression.

Poly-N-Acetyl Glucosamine (sNAG) Enhances Early Rotator Cuff Tendon Healing in a Rat Model

Rotator cuff injuries frequently require surgical repairs which have a high failure rate. Biological augmentation has been utilized in an attempt to improve tendon repair. Poly-N-acetyl glucosamine (sNAG) polymer containing nanofibers has been shown to increase the rate for healing of venous leg ulcers. The purpose of this study was to investigate the healing and analgesic properties of sNAG in a rat rotator cuff injury and repair model. 144 adult male Sprague-Dawley rats underwent a transection and repair of their left supraspinatus tendons. Half of the animals received a sNAG membrane on the tendon-to-bone insertion site. Animals were further subdivided, receiving 1 or 3days of analgesics. Animals were sacrificed 2, 4, or 8weeks post-injury. Animals sacrificed at 4 and 8weeks underwent longitudinal in vivo ambulatory assessment. Histological properties were assessed at 2, 4, and 8weeks, and mechanical properties at 4 and 8weeks. In the presence of analgesics, tendons receiving the sNAG polymer had significantly increased max load and max stress at 4weeks, but not at 8weeks. Ambulatory improvements were observed at 14days in stride length and speed. Therefore, sNAG improves tendon-to-bone healing in a rat rotator cuff detachment and repair model.

Icariin Promotes Tendon-Bone Healing during Repair of Rotator Cuff Tears: A Biomechanical and Histological Study.

To investigate whether the systematic administration of icariin (ICA) promotes tendon-bone healing after rotator cuff reconstruction in vivo, a total of 64 male Sprague Dawley rats were used in a rotator cuff injury model and underwent rotator cuff reconstruction (bone tunnel suture fixation). Rats from the ICA group (n = 32) were gavage-fed daily with ICA at 0.125 mg/g, while rats in the control group (n = 32) received saline only. Micro-computed tomography, biomechanical tests, serum ELISA (calcium; Ca, alkaline phosphatase; AP, osteocalcin; OCN) and histological examinations (Safranin O and Fast Green staining, type I, II and III collagen (Col1, Col2, and Col3), CD31, and vascular endothelial growth factor (VEGF)) were analyzed two and four weeks after surgery. In the ICA group, the serum levels of AP and OCN were higher than in the control group. More Col1-, Col2-, CD31-, and VEGF-positive cells, together with a greater degree of osteogenesis, were detected in the ICA group compared with the control group. During mechanical testing, the ICA group showed a significantly higher ultimate failure load than the control group at both two and four weeks. Our results indicate that the systematic administration of ICA could promote angiogenesis and tendon-bone healing after rotator cuff reconstruction, with superior mechanical strength compared with the controls. Treatment for rotator cuff injury using systematically-administered ICA could be a promising strategy.