Enhancing Functional Recovery after Segmental Nerve Defect Using Nerve Allograft Treated with Plasma-Derived Exosome.

BACKGROUND: Nerve injuries can result in detrimental functional outcomes. Currently, autologous nerve graft offers the best outcome for segmental peripheral nerve injury. Allografts are alternatives, but do not have comparable results. This study evaluated whether plasma-derived exosome can improve nerve regeneration and functional recovery when combined with decellularized nerve allografts. METHODS: The effect of exosomes on Schwann cell proliferation and migration were evaluated. A rat model of sciatic nerve repair was used to evaluate the effect on nerve regeneration and functional recovery. A fibrin sealant was used as the scaffold for exosome. Eighty-four Lewis rats were divided into autograft, allograft, and allograft with exosome groups. Gene expression of nerve regeneration factors was analyzed on postoperative day 7. At 12 and 16 weeks, rats were subjected to maximum isometric tetanic force and compound muscle action potential. Nerve specimens were then analyzed by means of histology and immunohistochemistry. RESULTS: Exosomes were readily taken up by Schwann cells that resulted in improved Schwann cell viability and migration. The treated allograft group had functional recovery (compound muscle action potential, isometric tetanic force) comparable to that of the autograft group. Similar results were observed in gene expression analysis of nerve regenerating factors. Histologic analysis showed no statistically significant differences between treated allograft and autograft groups in terms of axonal density, fascicular area, and myelin sheath thickness. CONCLUSIONS: Plasma-derived exosome treatment of decellularized nerve allograft may provide comparable clinical outcomes to that of an autograft. This can be a promising strategy in the future as an alternative for segmental peripheral nerve repair. CLINICAL RELEVANCE STATEMENT: Off-the-shelf exosomes may improve recovery in nerve allografts.

Carbodiimide-Derivatized Synovial Fluid for Tendon Graft Coating Improves Long-Term Functional Outcomes of Flexor Tendon Reconstruction.

BACKGROUND: Flexor digitorum profundus (FDP) tendon injury is common in hand trauma, and flexor tendon reconstruction is one of the most challenging procedures in hand surgery because of severe adhesion that exceeds 25% and hinders hand function. The surface properties of a graft from extrasynovial tendons are inferior to those of the native intrasynovial FDP tendons, which has been reported as one of the major causations. Improved surface gliding ability of the extrasynovial graft is needed. Thus, this study used carbodiimide-derivatized synovial fluid and gelatin (cd-SF-gel) to modify the surface of the graft, thus improving functional outcomes using a dog in vivo model. METHODS: Forty FDP tendons from the second and fifth digits of 20 adult women underwent reconstruction with a peroneus longus (PL) autograft after creation of a tendon repair failure model for 6 weeks. Graft tendons were either coated with cd-SF-gel ( n = 20) or not. Animals were euthanized 24 weeks after reconstruction, and digits were collected after the animals were euthanized for biomechanical and histologic analyses. RESULTS: Adhesion score (cd-SF-gel, 3.15 ± 1.53; control, 5 ± 1.26; P < 0.00017), normalized work of flexion (cd-SF-gel, 0.47 ± 0.28 N-mm/degree; control, 1.4 ± 1.45 N-mm/degree; P < 0.014), and distal interphalangeal joint motion (cd-SF-gel, 17.63 ± 6.77 degrees; control, 7.07 ± 12.99 degrees; P < 0.0015) in treated grafts all showed significant differences compared with nontreated grafts. However, there was no significant difference in repair conjunction strength between the two groups. CONCLUSION: Autograft tendon surface modification with cd-SF-gel improves tendon gliding ability, reduces adhesion formation, and enhances digit function without interfering with graft-host healing. CLINICAL RELEVANCE STATEMENT: The authors demonstrate a clinically relevant and translational technology by using the patient's own synovial fluid to "synovialize" an autologous extrasynovial tendon graft to improve functional outcomes following flexor tendon reconstruction.

The Effects of the TSOL Knot on the Repair Strength and Gliding Resistance Following Flexor Tendon Repair.

BACKGROUND: The stability of a suture knot construct has been realized as an important parameter that affects the strength of flexor tendon repairs. A novel 2-strand-overhand-locking (TSOL) knot, which is not commonly used in the clinical setting, recently was reported to increase repair strength and to decrease tendon gliding resistance in a 2-strand repair technique. The purpose of the present study was to investigate the effect of the TSOL knot on tendon repair strength and gliding resistance compared with a typical surgical knot in both 2-strand and 4-strand repair techniques using an in vitro turkey flexor tendon model. METHODS: Sixty flexor digitorum profundus tendons from the long digit of the turkey foot were divided evenly into 4 groups and repaired with the following techniques: (1) a 2-strand modified Pennington repair with a square knot, (2) a 2-strand modified Pennington repair with a TSOL knot, (3) a 4-strand grasping cruciate repair with a square knot, and (4) a 4-strand grasping cruciate repair with a TSOL knot. Repaired tendons were tested for failure mode, gliding resistance, and repair strength at failure. RESULTS: The repair strength and stiffness of the 4-strand repairs were significantly higher than those of the 2-strand repairs, regardless of knot type (p < 0.05). The repair strength at failure of the TSOL knot was significantly greater than that of the square knot in 2-strand repairs (p < 0.05) but not in 4-strand repairs. The gliding resistance of the TSOL knot was significantly decreased compared with that of the square knot in both 2-strand and 4-stand repairs (p < 0.05). With regard to failure mode, the TSOL knot was less likely to fail due to knot unravelling. CONCLUSIONS: In this in vitro biomechanical study involving the use of turkey flexor tendons to compare gliding resistance and repair strength characteristics for knot-inside 2 and 4-strand repairs, the TSOL knot was associated with decreased repaired tendon gliding resistance, regardless of the number of strands used. Although the TSOL knot also increased the repair strength, the difference was only significant when 2-strand repairs were used. The results of our study support the use of the TSOL knot in the clinical setting of flexor tendon repair using 2 or 4-strand, knot-inside methods. CLINICAL RELEVANCE: In surgical repair of flexor tendons, there is substantial interest in maximizing strength while minimizing friction. This study shows the potential utility of the TSOL knot to increase repair strength while decreasing gliding resistance, particularly in 2-strand repairs.

Effects of purified exosome product on rotator cuff tendon-bone healing in vitro and in vivo.

Exosomes have multiple therapeutic targets, but the effects on healing rotator cuff tear (RCT) remain unclear. As a circulating exosome, purified exosome product (PEP) has the potential to lead to biomechanical improvement in RCT. Here, we have established a simple and efficient approach that identifies the function and underlying mechanisms of PEP on cell-cell interaction using a co-culture model in vitro. In the in vivo trial, adult female Sprague-Dawley rats underwent unilateral surgery to transect and repair the supraspinatus tendon to its insertion site with or without PEP. PEP promoted the migration and confluence of osteoblast cells and tenocytes, especially during direct cell-cell contact. Expression of potential genes for RCT in vitro and in vivo models were consistent with biomechanical tests and semiquantitative histologic scores, indicating accelerated strength and healing of the RC in response to PEP. Our observations suggest that circulating exosomes provide an effective option to improve the healing speed of RCT after surgical repair. The regeneration of enthesis following PEP treatment appears to be related to a mutually reinforcing relationship between direct cell-cell contact and PEP activity, suggesting a dual approach to the healing process.

Engineered tendon-fibrocartilage-bone composite and bone marrow-derived mesenchymal stem cell sheet augmentation promotes rotator cuff healing in a non-weight-bearing canine model.

Reducing rotator cuff failure after repair remains a challenge due to suboptimal tendon-to-bone healing. In this study we report a novel biomaterial with engineered tendon-fibrocartilage-bone composite (TFBC) and bone marrow-derived mesenchymal stem cell sheet (BMSCS); this construct was tested for augmentation of rotator cuff repair using a canine non-weight-bearing (NWB) model. A total of 42 mixed-breed dogs were randomly allocated to 3 groups (n = 14 each). Unilateral infraspinatus tendon underwent suture repair only (control); augmentation with engineered TFBC alone (TFBC), or augmentation with engineered TFBC and BMSCS (TFBC + BMSCS). Histomorphometric analysis and biomechanical testing were performed at 6 weeks after surgery. The TFBC + BMSCS augmented repairs demonstrated superior histological scores, greater new fibrocartilage formation and collagen fiber organization at the tendon-bone interface compared with the controls. The ultimate failure load and ultimate stress were 286.80 ± 45.02 N and 4.50 ± 1.11 MPa for TFBC + BMSCS group, 163.20 ± 61.21 N and 2.60 ± 0.97 MPa for control group (TFBC + BMSCS vs control, P = 1.12E-04 and 0.003, respectively), 206.10 ± 60.99 N and 3.20 ± 1.31 MPa for TFBC group (TFBC + BMSCS vs TFBC, P = 0.009 and 0.045, respectively). In conclusion, application of an engineered TFBC and BMSCS can enhance rotator cuff healing in terms of anatomic structure, collagen organization and biomechanical strength in a canine NWB model. Combined TFBC and BMSCS augmentation is a promising strategy for rotator cuff tears and has a high potential impact on clinical practice.

Comparison of Autograft and Allograft with Surface Modification for Flexor Tendon Reconstruction: A Canine in Vivo Model.

BACKGROUND: Flexor tendon injury is common, and tendon reconstruction is indicated clinically if the primary repair fails or cannot be performed immediately after tendon injury. The purpose of the current study was to compare clinically standard extrasynovial autologous graft (EAG) tendon and intrasynovial allogeneic graft (IAG) that had both undergone biolubricant surface modification in a canine in vivo model. METHODS: Twenty-four flexor digitorum profundus (FDP) tendons from the second and fifth digits of 12 dogs were used for this study. In the first phase, a model of failed FDP tendon repair was created. After 6 weeks, the ruptured FDP tendons with a scarred digit were reconstructed with the use of either EAG or IAG tendons treated with carbodiimide-derivatized hyaluronic acid and lubricin. At 12 weeks after tendon reconstruction, the digits were harvested for functional, biomechanical, and histologic evaluations. RESULTS: The tendon failure model was a clinically relevant and reproducible model for tendon reconstruction. The IAG group demonstrated improved digit function with decreased adhesion formation, lower digit work of flexion, and improved graft gliding ability compared with the EAG group. However, the IAG group had decreased healing at the distal tendon-bone junction. Our histologic findings verified the biomechanical evaluations and, further, showed that cellular repopulation of allograft at 12 weeks after reconstruction is still challenging. CONCLUSIONS: FDP tendon reconstruction using IAG with surface modification has some beneficial effects for reducing adhesions but demonstrated inferior healing at the distal tendon-bone junction compared with EAG. These mixed results indicate that vitalization and turnover acceleration are crucial to reducing failure of reconstruction with allograft. CLINICAL RELEVANCE: Flexor tendon reconstruction is a common surgical procedure. However, postoperative adhesion formation may lead to unsatisfactory clinical outcomes. In this study, we developed a potential flexor tendon allograft using chemical and tissue-engineering approaches. This technology could improve function following tendon reconstruction.

Kinetic analysis of canine gait on the effect of failure tendon repair and tendon graft.

Kinetic analysis of canine gait has been extensively studied, including normal and abnormal gait. However, no research has looked into how flexor tendon injury and further treatment would affect the walking pattern comparing to the uninjured state. Therefore, this study was aimed to utilize a portable pressure walkway system, which has been commonly used for pedobarographic and kinetic analysis in the veterinary field, to examine the effect of a failed tendon repair and tendon graft reconstruction on canine digit kinetics during gait. 12 mixed breed (mongrel) hound-type female dogs were included in this study and 2nd and 5th digits were chosen to undergo flexor tendon repair and graft surgeries. Kinetic parameters from the surgery leg in stance phase were calculated. From the results, after tendon failure repair, decrease of weight bearing was seen in the affected digits and weight bearing was shifted to the metacarpal pad. After tendon graft reconstruction, weight bearing returned to the affected digits and metacarpal pads. Slight alteration in peak pressure and instant of peak force were identified, but it was estimated to have little influence on post-reconstruction gait. This study could serve as a reference in evaluating canine digit function in flexor tendon injury for future studies.

Effects of lubricant and autologous bone marrow stromal cell augmentation on immobilized flexor tendon repairs.

The purpose of the study was to test a novel treatment that carbodiimide-derivatized-hyaluronic acid-lubricin (cd-HA-lubricin) combined cell-based therapy in an immobilized flexor tendon repair in a canine model. Seventy-eight flexor tendons from 39 dogs were transected. One tendon was treated with cd-HA-lubricin plus an interpositional graft of 8 × 10(5) BMSCs and GDF-5. The other tendon was repaired without treatment. After 21 day of immobilization, 19 dogs were sacrificed; the remaining 20 dogs underwent a 21-day rehabilitation protocol before euthanasia. The work of flexion, tendon gliding resistance, and adhesion score in treated tendons were significantly less than the untreated tendons (p < 0.05). The failure strength of the untreated tendons was higher than the treated tendons at 21 and 42 days (p < 0.05). However, there is no significant difference in stiffness between two groups at day 42. Histologic analysis of treated tendons showed a smooth surface and viable transplanted cells 42 days after the repair, whereas untreated tendons showed severe adhesion formation around the repair site. The combination of lubricant and cell treatment resulted in significantly improved digit function, reduced adhesion formation. This novel treatment can address the unmet needs of patients who are unable to commence an early mobilization protocol after flexor tendon repair.

Surface Modification with Chemically Modified Synovial Fluid for Flexor Tendon Reconstruction in a Canine Model in Vivo.

BACKGROUND: Functional restoration is the major concern after flexor tendon reconstruction in the hand. The purpose of the present study was to investigate the effects of modifying the surface of extrasynovial tendon autografts with carbodiimide-derivatized synovial fluid with gelatin (cd-SF-G) on functional outcomes of flexor tendon reconstruction using a canine model. METHODS: The second and fifth flexor digitorum profundus tendons from eleven dogs were transected and repaired in zone II. The dogs then had six weeks of free activity leading to tendon rupture and scar formation (the repair-failure phase). In the reconstruction phase, two autologous peroneus longus tendons from each dog were harvested; one tendon was coated with cd-SF-G and the other, with saline solution, as a control. A non-weight-bearing rehabilitation protocol was followed for six weeks after reconstruction. The digits were then harvested and evaluations of function, adhesion status, gliding resistance, attachment strength, cell viability, and histology were performed. RESULTS: The tendons coated with cd-SF-G demonstrated significantly lower values (mean and standard deviation) compared with the saline-solution group for work of flexion (0.63 ± 0.24 versus 1.34 ± 0.42 N-mm/deg), adhesion score (3.5 ± 1.6 versus 6.1 ± 1.3), proximal adhesion breaking force (8.6 ± 3.2 versus 20.2 ± 10.2 N), and gliding resistance (0.26 ± 0.08 versus 0.46 ± 0.22 N) (p < 0.05). There was no significant difference between the cd-SF-G and saline-solution groups (p > 0.05) in distal attachment-site strength (56.9 ± 28.4 versus 77.2 ± 36.2 N), stiffness (19 ± 7.5 versus 24.5 ± 14.5 N/mm), and compressive modulus from indentation testing (4.37 ± 1.26 versus 3.98 ± 1.24 N/mm). Histological analysis showed that tendons coated with cd-SF-G had smoother surfaces and demonstrated tendon-to-bone and tendon-to-tendon incorporation. No significant difference in viable cell count between the two groups was observed on tendon culture. CONCLUSIONS: Modification of the flexor tendon surface with cd-SF-G significantly improved digital function and reduced adhesion formation without affecting graft healing and stiffness. CLINICAL RELEVANCE: This study used native synovial fluid as a basic lubricating reagent to treat a tendon graft in vivo, a novel avenue for improving clinical outcomes of flexor tendon reconstruction. This methodology may also apply to other surgical procedures where postoperative adhesions impair function.

Biomechanical evaluation of flexor tendon graft with different repair techniques and graft surface modification.

The purpose of this study was to investigate the biomechanical properties of modified repair techniques for flexor tendon reconstruction and the effects of surface modification using carbodiimide-derivatized synovial fluid plus gelatin (cd-SF-G), compared to the traditional repair techniques. The second and fifth digits from 16 canine forepaws were randomly divided into 4 groups: (1) traditional graft repairs (TGR group) including distal Bunnell repair and proximal Pulvertaft weave repair; (2) modified graft repairs (MGR group) including distal graft bony attachment repair and proximal step-cut repair; (3) group TGR coated with cd-SF-G (TGR-C group); and (4) group MGR coated with cd-SF-G (MGR-C group). Digit normalized work of flexion (nWOF), ultimate failure strength, and stiffness were measured. The nWOF in MGR group was significantly less than TGR group (p < 0.05). The nWOF in groups treated with cd-SF-G was significantly less than their untreated counterparts (p < 0.05). Ultimate load to failure of the MGR-C group was significantly greater than the TGR-C group (p < 0.05), but no significant difference in stiffness was found between these two groups. The modified techniques cannot only improve tendon gliding abilities but can also improve breaking strength. Additionally, surface modification with cd-SF-G significantly decreased the work of flexion.

CORR® ORS Richard A. Brand Award for Outstanding Orthopaedic Research: Engineering flexor tendon repair with lubricant, cells, and cytokines in a canine model.

BACKGROUND: Adhesions and poor healing are complications of flexor tendon repair. QUESTIONS/PURPOSES: The purpose of this study was to investigate a tissue engineering approach to improve functional outcomes after flexor tendon repair in a canine model. METHODS: Flexor digitorum profundus tendons were lacerated and repaired in 60 dogs that were followed for 10, 21, or 42 days. One randomly selected repair from either the second or fifth digit in one paw in each dog was treated with carbodiimide-derivatized hyaluronic acid, gelatin, and lubricin plus autologous bone marrow stromal cells stimulated with growth and differentiation factor 5; control repair tendons were not treated. Digits were analyzed by adhesion score, work of flexion, tendon-pulley friction, failure force, and histology. RESULTS: In the control group, 35 of 52 control tendons had adhesions, whereas 19 of 49 treated tendons had adhesions. The number of repaired tendons with adhesions in the control group was greater than the number in the treated group at all three times (p = 0.005). The normalized work of flexion in treated tendons was 0.28 (± 0.08), 0.29 (± 0.19), and 0.32 (± 0.22) N/mm/° at Day 10, Day 21, and Day 42 respectively, compared with the untreated tendons of 0.46 (± 0.19) at Day 10 (effect size, 1.5; p = 0.01), 0.77 (± 0.49) at Day 21 (effect size, 1.4; p < 0.001), and 1.17 (± 0.82) N/mm/° at Day 42 (effect size, 1.6; p < 0.001). The friction data were comparable to the work of flexion data at all times. The repaired tendon failure force in the untreated group at 42 days was 70.2 N (± 8.77), which was greater than the treated tendons 44.7 N (± 8.53) (effect size, 1.9; p < 0.001). Histologically, treated repairs had a smooth surface with intrinsic healing, whereas control repairs had surface adhesions and extrinsic healing. CONCLUSIONS: Our study provides evidence that tissue engineering coupled with restoration of tendon gliding can improve the quality of tendon healing in a large animal in vivo model. CLINICAL RELEVANCE: Tissue engineering may enhance intrinsic tendon healing and thus improve the functional outcomes of flexor tendon repair.

The effects of biological lubricating molecules on flexor tendon reconstruction in a canine allograft model in vivo.

BACKGROUND: Using allograft is an attractive alternative for flexor tendon reconstruction because of the lack of donor-site morbidity, and better matching to the intrasynovial environment. The purpose of this study was to use biological lubricant molecules to modify the graft surface to decrease adhesions and improve digit function. METHODS: Twenty-eight flexor digitorum profundus tendons from the second and fifth digits of 14 dogs were lacerated and repaired to create a model with repair failure and scar digit for tendon reconstruction. Six weeks after the initial operation, the tendons were reconstructed with flexor digitorum profundus allograft tendons obtained from canine cadavers. One graft tendon in each dog was treated with saline as a control and the other was treated with carbodiimide-derivatized hyaluronic acid and gelatin plus lubricin. Six weeks postoperatively, digit function, graft mechanics, and biology were analyzed. RESULTS: Allograft tendons treated with carbodiimide-derivatized hyaluronic acid-lubricin had decreased adhesions at the proximal tendon/graft repair and within the flexor sheath, improved digit function, and increased graft gliding ability. The treatment also reduced the strength at the distal tendon-to-bone repair, but the distal attachment rupture rate was similar for both graft types. Histologic evaluation showed that viable cells migrated to the allograft, but these were limited to the tendon surface. CONCLUSIONS: Carbodiimide-derivatized hyaluronic acid-lubricin treatment of tendon allograft improves digit functional outcomes after flexor tendon reconstruction. However, delayed bone-to-tendon healing should be a caution. Furthermore, the cell infiltration into the allograft tendon substance should be a target for future studies, to shorten the allograft self-regeneration period.