Rat Rotator Cuff Repair Model Research Articles

Effect of magnetic microbeads on sustained and targeted delivery of transforming growth factor-beta-1 for rotator cuff healing in a rat rotator cuff repair model

Structural failure is a well-established complication of rotator cuff repair procedures. To evaluate the effect of magnetic microbeads, designed for precise drug delivery via magnetic force, on sustained transforming growth factor-beta-1 (TGF-β1) release and rotator cuff healing in a rat rotator cuff repair model. TGF-β1 laden microbeads were prepared, and baseline in vitro experiments included the magnetization of the microbeads and TGF-β1 release tests. In an in vivo experiment using a rat rotator cuff repair model on both shoulders, 72 rats were randomly assigned to three groups (24 per group): group A, conventional repair; group B, repair with and simple TGF-β1 injection; and group C, repair with magnet insertion into the humeral head and TGF-β1 laden microbead injection. Delivery of TGF-β1 was evaluated at 1 and 7 days after the intervention using PCR, Western blot, and immunohistochemistry. At 6 weeks post-intervention, rotator cuff healing was assessed using biomechanical and histological analysis. The in vitro experiments confirmed the magnetization property of the microbeads and sustained delivery of TGF-β1 for up to 10 days. No difference in the TGF-β1 expression was found at day 1 in vivo. However, at day 7, group C exhibited a significantly elevated expression of TGF-β1 in both PCR and Western blot analyses compared to groups A and B (all P < 0.05). Immunohistochemical analysis revealed a higher expression of TGF-β1 at the repair site in group C on day 7. At 6 weeks, biomechanical analysis demonstrated a significantly higher ultimate failure load in group C than in groups A and B (P < 0.05) and greater stiffness than in group A (P = 0.045). In addition, histological analysis showed denser and more regular collagen fibers with complete continuity to the bone in group C than in groups A and B, a statistically significant difference according to the semi-quantitative scoring system (all P < 0.05). The use of the TGF-β1 laden magnetic microbeads demonstrated sustained delivery of TGF-β1 to the repair site, improving rotator cuff healing.

Effect of aging on recruitment of tenogenic progenitor during tendon-to-bone healing in a rat rotator cuff repair model

Effect of aging on recruitment of tenogenic progenitor during tendon-to-bone healing in a rat rotator cuff repair model

Three-Dimensional Cultured Human Rotator Cuff Spheroids: tendon-related Marker Expression and Efficacy in Rat Rotator Cuff Repair Model

Three-Dimensional Cultured Human Rotator Cuff Spheroids: tendon-related Marker Expression and Efficacy in Rat Rotator Cuff Repair Model

The Effect of Vitamin C and N-Acetylcysteine on Tendon-to-Bone Healing in a Rodent Model of Rotator Cuff Repair

Background: Oxidative stress inhibits tendon-to-bone healing after rotator cuff repair. Regulation of oxidative stress has the potential to accelerate this healing, but its mechanism remains unclear. Purpose: To investigate the effects of reducing oxidative stress by applying antioxidants, such as N-acetylcysteine (NAC) and vitamin C (VC), on rotator cuff repair in a rat rotator cuff repair model. Study design: Controlled laboratory study. Methods: A total of 48 Sprague Dawley rats underwent bilateral surgery to repair the infraspinatus tendon to its insertion site 1 week after detachment. Rats were assigned to either the NAC group, the VC group, or a control group. Histological evaluation was performed via hematoxylin-eosin or toluidine blue staining, and oxidative stress was assessed via dihydroethidium intensity and protein carbonyl concentration at 3 and 6 weeks. Superoxide dismutase 1 (SOD1), SOD2, SOD3, peroxiredoxin 5, collagen type I (COL1), COL3, matrix metalloproteinase 1 (MMP-1), MMP-3, and MMP-13 expression and SOD activity were determined at 3 and 6 weeks. Biomechanical tests were performed at 6 and 12 weeks. Results: Histological evaluation showed that the number of chondrocytes in the NAC group at 6 weeks and in the VC group at 3 and 6 weeks, the area of fibrocartilage at 6 weeks in the VC group, and collagen fibers at 6 weeks in the NAC and VC groups were significantly increased compared with those in the control group. Dihydroethidium intensity at 3 and 6 weeks and protein carbonyls at 6 weeks in the NAC and VC groups were significantly decreased. SOD1 expression and SOD activity at 3 weeks in the VC group and peroxiredoxin 5 expression at 6 weeks in the NAC group were significantly upregulated compared with that in the control group. COL3 expression was significantly upregulated at 6 weeks in the VC group, and MMP-13 expression was significantly decreased at 6 weeks in the NAC and VC groups. The biomechanical strength showed no significant difference. Conclusion: Antioxidant treatment, via NAC or VC administration, reduced oxidative stress in the rotator cuff repair site and accelerated healing. Clinical Relevance: These findings provide essential indications to develop clinical strategies for improved healing after rotator cuff surgical repair in patients.

OBESITY IMPAIRS ENTHESIS HEALING AFTER ROTATOR CUFF REPAIR IN A RAT MODEL

Obesity is associated with poor outcomes and increased risk of failure after rotator cuff (RC) repair surgery. The effect of diet-induced obesity (DIO) on enthesis healing has not been well characterised and whether its effects can be reversed with dietary intervention is unknown. We hypothesised that DIO would result in inferior enthesis healing in a rat model of RC repair and that dietary intervention in the peri-operative period would improve enthesis healing.A total of 78 male Sprague-Dawley rats were divided into three weight-matched groups from weaning and fed either: control diet (CD), high-fat diet (HFD), or HFD until surgery, then CD thereafter (HF-CD). After 12 weeks the left supraspinatus tendon was detached, followed by immediate surgical repair. At 2 and 12 weeks post-surgery, animals were cullers and RCs harvested for biomechanical and histological evaluation. Body composition and metabolic markers were assessed via DEXA and plasma analyses, respectively.DIO was established in the HFD and HF-CD groups prior to surgery, and subsequently reversed in the HF-CD group after surgery. At 12 weeks post-surgery, plasma leptin concentrations were higher in the HFD group compared to the CD group (5.28 vs. 2.91ng/ml, P=0.003). Histologically, the appearance of the repaired entheses was poorer in both the HFD and HF-CD compared to the CD group at 12 weeks (overall histological score 6.20 (P=0.008), 4.98 (P=0.001) and 8.68 out of 15, respectively). The repaired entheses in the HF-CD group had significantly lower (26.4 N, P=0.028) load-at-failure 12 weeks post-surgery compared to the CD group (34.4 N); while the HFD group was low, but not significantly different (28.1 N, P=0.096). Body mass at the time of surgery, plasma leptin and body fat percentage were negatively correlated with histological scores and plasma leptin with load-at-failure 12 weeks post-surgery.DIO impaired enthesis healing in this rat RC repair model, with inferior biomechanical and histological outcomes. Restoring normal weight with dietary change after surgery did not improve healing outcomes. Exploring interventions that improve the metabolic state of obese patients and counselling patients appropriately about their modest expectations after repair should be considered.

OBESITY IMPAIRS ENTHESIS HEALING AFTER ROTATOR CUFF REPAIR IN A RAT MODEL

We hypothesised that diet-induced obesity (DIO) would result in inferior enthesis healing in a rat model of rotator cuff (RC) repair and that dietary intervention in the peri-operative period would improve enthesis healing.A total of 78 male Sprague-Dawley rats were divided into three weight-matched groups from weaning and fed either: control diet (CD), high-fat diet (HFD), or HFD until surgery, then CD thereafter (HF-CD). After 12 weeks, the left supraspinatus tendon was detached, followed by immediate surgical repair. At 2 and 12 weeks post-surgery, animals were culled, and RCs harvested for biomechanical and histological evaluation. Body composition and metabolic markers were assessed via DEXA and plasma analyses, respectively.DIO was established in the HFD and HF-CD groups before surgery and subsequently reversed in the HF-CD group after surgery. Histologically, the appearance of the repaired entheses was poorer in both the HFD and HF-CD groups compared with the CD group at 12 weeks after surgery, with semiquantitative scores of 6.2 (p&lt;0.01), 4.98 (p&lt;0.01), and 8.7 of 15, respectively. The repaired entheses in the HF-CD group had a significantly lower load to failure (P=0.03) at 12 weeks after surgery compared with the CD group, while the load to failure in the HFD group was low but not significantly different (P=0.10). Plasma leptin were negatively correlated with histology scores and load to failure at 12 weeks after surgery.DIO impaired enthesis healing in this rat RC repair model, with inferior biomechanical and histological outcomes. Restoring normal weight with dietary change after surgery did not improve healing outcomes. Circulating levels of leptin significantly correlated with poor healing outcomes. This pre-clinical rodent model demonstrates that obesity is a potentially modifiable factor that impairs RC healing and increases the risk of failure after RC surgery.

The recombinant human fibroblast growth factor-18 (sprifermin) improves tendon-to-bone healing by promoting chondrogenesis in a rat rotator cuff repair model.

Rotator cuff healing is improved by reconstructing the fibrocartilaginous structure of the tendon-to-bone enthesis. Fibroblast growth factor (FGF)-18 (sprifermin) is a well-known growth factor that improves articular cartilage repair via its anabolic effect. This study aimed to investigate the effect of recombinant human FGF-18 (rhFGF-18) on the chondrogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) invitro and tendon-to-bone healing in a rat model of rotator cuff repair. Histological and reverse transcription-quantitative real-time polymerase chain reaction analyses of chondral pellets cultured with different concentrations of rhFGF-18 were performed. Bilateral detachment and repair of the supraspinatus tendon were performed on rats. The rats were administered 0.2mL of sodium alginate (SA) hydrogel with (rhFGF-18/SA group, n=12) or without (SA group, n=12) 20μg of rhFGF-18 into the repaired side. The simple repair group (n=12) served as a control. At 4 and 8weeks after surgery, histological analysis and biomechanical tests were performed. After chondrogenesis induction, compared with the control group, 10ng/mL of rhFGF-18 increased pellet volume significantly (P=.002), with improved histological staining. It was noted that 10ng/mL of rhFGF-18 upregulated the mRNA expression (relative ratio to control) of aggrecan (2.59±0.29, P<.001), SRY-box transcription factor 9 (1.88±0.05, P<.001), and type II collagen (1.46±0.18, P=.009). At 4 and 8weeks after surgery, more fibrocartilage and cartilaginous extracellular matrix was observed in rhFGF-18/SA-treated rats. The semiquantitative data from picrosirius red staining test were 31.1±4.5 vs. 61.2±4.1 at 4weeks (P<.001) and 61.5±2.8 vs. 80.5±10.5 at 8weeks (P=.002) (control vs. rhFGF-18/SA). Ultimate failure load (25.42±3.61N vs. 18.87±2.71N at 4weeks and 28.63±5.22N vs. 22.15±3.11N at 8weeks; P=.006 and P=.03, respectively) and stiffness (18.49±1.38N/mm vs. 14.48±2.01N/mm at 8weeks, P=.01) were higher in the rhFGF-18/SA group than in the control group. rhFGF-18 promoted chondrogenesis in the hBMSCs invitro. rhFGF-18/SA improved tendon-to-bone healing in the rats by promoting regeneration of the fibrocartilage enthesis. rhFGF-18 (sprifermin) may be beneficial in improving tendon-to-bone healing after rotator cuff repair.

3D cell-printing of gradient multi-tissue interfaces for rotator cuff regeneration.

Owing to the prevalence of rotator cuff (RC) injuries and suboptimal healing outcome, rapid and functional regeneration of the tendon–bone interface (TBI) after RC repair continues to be a major clinical challenge. Given the essential role of the RC in shoulder movement, the engineering of biomimetic multi-tissue constructs presents an opportunity for complex TBI reconstruction after RC repair. Here, we propose a gradient cell-laden multi-tissue construct combined with compositional gradient TBI-specific bioinks via 3D cell-printing technology. In vitro studies demonstrated the capability of a gradient scaffold system in zone-specific inducibility and multi-tissue formation mimicking TBI. The regenerative performance of the gradient scaffold on RC regeneration was determined using a rat RC repair model. In particular, we adopted nondestructive, consecutive, and tissue-targeted near-infrared fluorescence imaging to visualize the direct anatomical change and the intricate RC regeneration progression in real time in vivo. Furthermore, the 3D cell-printed implant promotes effective restoration of shoulder locomotion function and accelerates TBI healing in vivo. In summary, this study identifies the therapeutic contribution of cell-printed constructs towards functional RC regeneration, demonstrating the translational potential of biomimetic gradient constructs for the clinical repair of multi-tissue interfaces.

Caffeine intake does not appear to impair tendon-to-bone healing strength in a rat rotator cuff repair model.

BackgroundTendon-to-bone (TtB) healing is essential for successful rotator cuff repair (RCR). This study aimed to investigate if caffeine intake impaired TtB healing in a rat RCR model.MethodsSeventy-two rats were randomized into a caffeinated group or a noncaffeinated group. Specimens received one week of oral caffeine solution or normal saline before RCR. All rats then underwent bilateral RCR. Caffeination or saline gavages continued until rats were sacrificed at 2, 4, and 8 weeks postoperatively. Load-to-failure (primary outcomes measure), maximum stress, and stiffness of the TtB interface were measured for one shoulder of each specimen. Six random shoulders from each group underwent histological assessment of TtB healing.ResultsLoad-to-failure and maximum stress of RCR did not appear to differ between groups at any time point. No difference in RCR stiffness was found between groups at 2 and 4 weeks; however, stiffness in the caffeinated group did appear to lower at 8 weeks (P = .04).ConclusionPerioperative caffeine intake did not appear to affect load-to-failure strength of RCR in an animal model. Although our secondary outcome measures of maximum stress and stiffness also did not appear to be influenced by perioperative caffeine intake, there did appear to be a trend toward decreased RCR stiffness at 8 weeks postoperatively in specimens that received caffeine.

Obesity Impairs Enthesis Healing After Rotator Cuff Repair in a Rat Model

Background: Being overweight or obese is associated with poor outcomes and an increased risk of failure after rotator cuff (RC) surgery. However, the effect of obesity on enthesis healing has not been well characterized. Hypotheses: Diet-induced obesity (DIO) would result in inferior enthesis healing in a rat model of RC repair, and a dietary intervention in the perioperative period would improve enthesis healing. Study Design: Controlled laboratory study. Methods: Male Sprague-Dawley rats were divided into 3 weight-matched groups (n = 26 per group): control diet (CD), high-fat diet (HFD), or HFD until surgery and then CD thereafter (HF-CD). After 12 weeks, the left supraspinatus tendon was detached, followed by immediate repair. Animals were sacrificed, and RCs were harvested at 2 and 12 weeks after surgery for biomechanical and histological evaluations. Metabolic end points were assessed using dual-energy X-ray absorptiometry and plasma analyses. Results: DIO was established in the HFD and HF-CD groups before surgery and subsequently reversed in the HF-CD group after surgery. At 12 weeks after surgery, the body fat percentage (P = .0021) and plasma leptin concentration (P = .0025) were higher in the HFD group compared with the CD group. Histologically, the appearance of the repaired entheses was poorer in both the HFD and HF-CD groups compared with the CD group at 12 weeks after surgery, with semiquantitative scores of 6.20 (P = .0078), 4.98 (P = .0003), and 8.68 of 15, respectively. The repaired entheses in the HF-CD group had a significantly lower load to failure (P = .0278) at 12 weeks after surgery compared with the CD group, while the load to failure in the HFD group was low but not significantly different (P = .0960). There were no differences in the biomechanical and histological results between the groups at 2 weeks after surgery. Body mass at the time of surgery, plasma leptin concentration, and body fat percentage were negatively correlated with histology scores and plasma leptin concentration was correlated with load to failure at 12 weeks after surgery. Conclusion: DIO impaired enthesis healing in this rat RC repair model, with inferior biomechanical and histological outcomes. Restoring a normal weight with dietary changes after surgery did not improve healing outcomes. Clinical Relevance: Obesity is a potentially modifiable factor that impairs RC healing and increases the risk of failure after surgery. Exploring interventions that improve the metabolic state of obese patients and counseling patients appropriately about their modest expectations after repair should be considered.

Enhanced Tendon-to-Bone Healing via IKKβ Inhibition in a Rat Rotator Cuff Model

Background: More than 450,000 rotator cuff repairs are performed annually, yet healing of tendon to bone often fails. This failure is rooted in the fibrovascular healing response, which does not regenerate the native attachment site. Better healing outcomes may be achieved by targeting inflammation during the early period after repair. Rather than broad inhibition of inflammation, which may impair healing, the current study utilized a molecularly targeted approach to suppress IKKβ, shutting down only the inflammatory arm of the nuclear factor κB (NF-κB) signaling pathway. Purpose: To evaluate the therapeutic potential of IKKβ inhibition in a clinically relevant model of rat rotator cuff repair. Study Design: Controlled laboratory study. Methods: After validating the efficacy of the IKKβ inhibitor in vitro, it was administered orally once a day for 7 days after surgery in a rat rotator cuff repair model. The effect of treatment on reducing inflammation and improving repair quality was evaluated after 3 days and 2, 4, and 8 weeks of healing, using gene expression, biomechanics, bone morphometry, and histology. Results: Inhibition of IKKβ attenuated cytokine and chemokine production in vitro, demonstrating the potential for this inhibitor to reduce inflammation in vivo. Oral treatment with IKKβ inhibitor reduced NF-κB target gene expression by up to 80% compared with a nontreated group at day 3, with a subset of these genes suppressed through 14 days. Furthermore, the IKKβ inhibitor led to enhanced tenogenesis and extracellular matrix production, as demonstrated by gene expression and histological analyses. At 4 weeks, inhibitor treatment led to increased toughness, no effects on failure load and strength, and decreases in stiffness and modulus when compared with vehicle control. At 8 weeks, IKKβ inhibitor treatment led to increased toughness, failure load, and strength compared with control animals. IKKβ inhibitor treatment prevented the bone loss near the tendon attachment that occurred in repairs in control. Conclusion: Pharmacological inhibition of IKKβ successfully suppressed excessive inflammation and enhanced tendon-to-bone healing after rotator cuff repair in a rat model. Clinical Relevance: The NF-κB pathway is a promising target for enhancing outcomes after rotator cuff repair.

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.

In situ tissue engineering of the tendon-to-bone interface by endogenous stem/progenitor cells

The long-term success of surgical repair of rotator cuff tears is largely dependent on restoration of a functional tendon-to-bone interface. We implemented micro-precise spatiotemporal delivery of growth factors in three-dimensional printed scaffolds for integrative regeneration of a fibrocartilaginous tendon-to-bone interface. Sustained and spatially controlled release of tenogenic, chondrogenic and osteogenic growth factors was achieved using microsphere-based delivery carriers embedded in thin membrane-like scaffolds. In vitro, the scaffolds embedded with spatiotemporal delivery of growth factors successfully guided regional differentiation of mesenchymal progenitor cells, forming multiphase tissues with tendon-like, cartilage-like and bone-like regions. In vivo, when implanted at the interface between the supraspinatus tendon and the humeral head in a rat rotator cuff repair model, these scaffolds promoted recruitment of endogenous tendon progenitor cells followed by integrative healing of tendon and bone via re-formation of strong fibrocartilaginous interfaces. Our findings demonstrate the potential of in situ tissue engineering of tendon-to-bone interfaces by endogenous progenitor cells. The in situ tissue engineering approach shows translational potential for improving outcomes after rotator cuff repair.

Is a Local Administration of Parathyroid Hormone Effective to Tendon-to-Bone Healing in a Rat Rotator Cuff Repair Model?

To evaluate the effect of local parathyroid hormone (PTH) administration on rotator cuff tendon-to-bone healing in a rat model compared with systemic PTH injection and untreated controls. PTH-alginate scaffold was prepared and sustained release of PTH was confirmed. Bilateral supraspinatus tendon repairs were performed in 39 rats (group 1, supraspinatus repair only; group 2, supraspinatus repair with systemic PTH injection; group 3, supraspinatus repair with local PTH administration via an absorbable scaffold; n = 13 each). Biomechanical (cross-sectional area, mode of failure, load to failure, and ultimate stress: right side) and histological analyses (hematoxylin and eosin stain, Masson's Trichrome stain Picrosirius red stain, Immunohistochemistry for BMP2, PTH1R, ColI, and ColIII: Left side) were performed to evaluate tendon-to-bone healing quality at 8 weeks after repair, and blood test (osteocalcin and procollagen type I N-terminal pro-peptide [PINP] levels) was performed in all rats. There was no intergroup difference in the healing failure rate (p = 0.910) or failure mode (p = 0.585). Biomechanically, subjects in groups 2 and 3 exhibited significantly larger cross-sectional areas and higher ultimate failure loads and ultimate stress than those in group 1 (all p < 0.05); however, no differences were noted between groups 2 and 3 (all p > 0.05). Histologically, groups 2 and 3 exhibited more organized tendon-to-bone interface structures with higher density, parallel orientation, and collagen fiber continuity than group 1 (all p < 0.05 except collagen fiber continuity in group 1 vs. 2); however, no differences in histological parameters between groups 2 and 3 (all p > 0.05). The protein levels of bone morphogenic protein 2, PTH 1 receptor, and collagen I and III and the serum level of PINP were increased in groups 2 and 3 versus group 1 (all p < 0.05) without showing differences between groups 2 and 3 (all p > 0.05). Local PTH administration using an absorbable scaffold improved the biomechanical and histological outcomes of rotator cuff tendon-to-bone healing comparable with systemic PTH injection at 8 weeks after repair in a rat model. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:82-91, 2020.

The effect of erythropoietin on rat rotator cuff repair model: An experimental study.

The aim of this study was to determine whether erythropoietin (EPO) can enhance rotator cuff healing in rats as measured by histological analysis and biomechanical testing. A total of 72 rats were included in this study. In the control group (n = 24), repair was performed without EPO injection. In the local group (n = 24) EPO was injected in the repair site. In the systemic group (n = 24) EPO was administered as an intraperitoneal injection every day for 10 days after repair. Rats were euthanized on day 10 (n = 12 from each group) and day 28 (n = 12 from each group). Histopathological (n = 6) and biomechanical examinations (n = 6) were done. Biomechanical results reveal that the maximum load to failure values of the early control group were statistically lower than those of the early systemic group (p = 0.006). Comparing the the total Bonar values histopathologically reveal that the early systemic group was statistically higher than those of the early local group (p = 0.043). The late control group was statistically higher than those of the late local group (p = 0.003) and the late systemic group (p = 0.034). The late systemic group was statistically higher than those of the late local group (p = 0.003). EPO application had a positive effect biomechanically in the early euthanized group and histopathologically in the late euthanized group.

Overlay repair with a synthetic collagen scaffold improves the quality of healing in a rat rotator cuff repair model

Augmenting repairs with extracellular matrix-based scaffolds is a common option for rotator cuff tears. In this study, a new collagen scaffold was assessed for its efficacy in augmenting rotator cuff repair. The collagen scaffold was assessed in vitro for cytocompatibility and retention of tenocyte phenotype using alamarBlue assays, fluorescent imaging, and real-time polymerase chain reaction. Immunogenicity was assessed in vitro by the activation of human monocytes. In vivo, by use of a modified rat rotator cuff defect model, supraspinatus tendon repairs were carried out in 40 animals. Overlay augmentation with the collagen scaffold was compared with unaugmented repairs. At 6 and 12 weeks postoperatively, the repairs were tested biomechanically to evaluate repair strength, as well as histologically to assess quality of healing. The collagen scaffold supported human tendon-derived cell growth in vitro, with cells demonstrating proliferation and appearing morphologically tenocytic over the experimental period. No immunogenic responses were provoked compared with suture material control. In vivo, augmentation with the scaffold improved the histologic scores at 12 weeks (8.4 of 15 vs 6.4 of 15, P = .032). However, no significant difference was detected with mechanical testing. The new collagen scaffold was supportive of cell growth in vitro and generated a minimal acute inflammatory response. In vivo, we observed an improvement in the histologic appearance of the repair at 12 weeks. However, a meaningful increase in biomechanical strength was not achieved. Further modification and improvement of the scaffold are required prior to consideration for clinical use.

Raman Spectroscopic Measurement of Bone–Tendon Insertion in Rat Rotator Cuff Repair Model

Raman Spectroscopic Measurement of Bone–Tendon Insertion in Rat Rotator Cuff Repair Model

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.

Effect of Footprint Preparation on Tendon-to-Bone Healing: A Histologic and Biomechanical Study in a Rat Rotator Cuff Repair Model

To compare the histologic and biomechanical effects of 3 different footprint preparations for repair of tendon-to-bone insertions and to assess the behavior of bone marrow-derived cells in each method of insertion repair. We randomized 81 male Sprague-Dawley rats and green fluorescent protein-bone marrow chimeric rats into 3 groups. In group A, we performed rotator cuff repair after separating the supraspinatus tendon from the greater tuberosity and removing the residual tendon tissue. In group B, we also drilled 3 holes into the footprint. The native fibrocartilage was preserved in groups A and B. In group C, we excavated the footprint until the cancellous bone was exposed. Histologic repair of the tendon-to-bone insertion, behavior of the bone marrow-derived cells, and ultimate force to failure were examined postoperatively. The areas of metachromasia in groups A, B, and C were 0.033 ± 0.019, 0.089 ± 0.022, and 0.002 ± 0.001mm2/mm2, respectively, at 4weeks and 0.029 ± 0.022, 0.090 ± 0.039, and 0.003 ± 0.001mm2/mm2, respectively, at 8weeks. At 4 and 8weeks postoperatively, significantly higher cartilage matrix production was observed in group B than in group C (4weeks, P= .002; 8weeks, P < .001). In green fluorescent protein-bone marrow chimeric rats in group B, bone marrow-derived chondrogenic cells infiltrated the fibrocartilage layer. Ultimate force to failure was significantly higher in group B (19.7 ± 3.4N) than in group C (16.7 ± 2.0N) at 8weeks (P= .031). Drilling into the footprint and preserving the fibrocartilage improved the quality of repair tissue and biomechanical strength at the tendon-to-bone insertion after rotator cuff repair in an animal model. Drilling into the footprint and preserving the fibrocartilage can enhance repair of tendon-to-bone insertions. This method may be clinically useful in rotator cuff repair.

TGF-β1 Improves Biomechanical Strength by Extracellular Matrix Accumulation Without Increasing the Number of Tenogenic Lineage Cells in a Rat Rotator Cuff Repair Model

Background: Transforming growth factor β1 (TGF-β1) positively regulates the tenogenic marker genes scleraxis (Scx) and tenomodulin (Tnmd) in mesenchymal progenitors in vitro. However, little is known about the effect of TGF-β1 on the expression of tenogenic markers during rotator cuff (RC) healing in rats. Hypothesis: TGF-β1 improves the biomechanical properties and histological maturity of reparative tissue in a rat RC repair model by stimulating the growth of tenogenic cells. Study Design: Controlled laboratory study. Methods: Adult male Sprague-Dawley rats (N = 180) underwent unilateral supraspinatus tendon-to-bone surgical repair and were randomly treated with a gelatin hydrogel presoaked in TGF-β1 (100 ng) or phosphate-buffered saline. The effects of TGF-β1 on RC healing were investigated at 2, 4, 6, 8, and 12 weeks postoperatively by immunostaining for proliferating cell nuclear antigen, by real-time reverse transcription polymerase chain reaction and in situ hybridization or immunostaining for enthesis-related markers (SRY-box containing gene 9 [Sox9], Scx, and Tnmd), and by real-time reverse transcription polymerase chain reaction and immunostaining for type I and III collagen. At 6 and 12 weeks postoperatively, biomechanical testing, micro–computed tomography, and biochemical analysis were also performed. At 2 and 4 weeks postoperatively, mesenchymal stem cell–related markers, phospho-Smad2, and matrix metalloproteinase 9 (MMP-9) and MMP-13 were assessed by immunostaining. Results: The TGF-β1-treated group had significantly higher ultimate load to failure and tissue volume at 6 and 12 weeks postoperatively and a higher collagen content at 12 weeks compared with the saline group. Tendon-related gene expression, histological maturity, cell proliferation, and mesenchymal stem cell–related marker immunoreactivity were not affected by exogenously administrated TGF-β1 at all time points. In the TGF-β1-treated group, the percentage of phospho-Smad2-positive cells within the healing tissue increased, whereas the expression of MMP-9 and MMP-13 significantly decreased at 2 and 4 weeks postoperatively. Conclusion: TGF-β1 enhances formation of tough fibrous tissues at the healing site by inhibiting MMP-9 and MMP-13 expression to increase collagen accumulation but without the growth of tenogenic lineage cells. Clinical Relevance: These findings suggest that TGF-β1 could be used for enhancing biomechanical strength after RC surgical repair.