Insulin-like growth factor-I improves cellular and molecular aspects of healing in a collagenase-induced model of flexor tendinitis.

Flexor tendinitis is a common and debilitating injury of elite and recreational athletes. Healing may be improved through intratendinous injection of insulin-like growth factor-I (IGF-I), which has been shown in vitro to stimulate mitogenesis and enhance tendon matrix production. This study investigated the effects of intratendinous injection of IGF-I on tendon healing in an equine model of flexor tendinitis. Collagenase-induced lesions were created in the tensile region of theflexor digitorum superficialis tendon of both forelimbs of eight horses. Treated tendons were injected with 2 microg rhlGF-I intralesionally every other day for 10 injections, while controls received 0.9% NaCl. Tendon fiber deposition and organization were evaluated serially using ultrasonography throughout the 8 week trial period. Following euthanasia, the tendons were harvested and DNA, hydroxyproline, and glycosaminoglycan content determined, mechanical strength and stiffness evaluated, gene expression and spatial arrangement of collagen types I and III assessed by northern blot and in situ hybridization, and tendon fiber architecture assessed by polarized light microscopy. Local soft tissue swelling was reduced in the IGF-I treated limbs. Similarly, lesion size in IGF-I treated tendons was smaller 3 and 4 weeks after initiation of treatment. Cell proliferation and collagen content of the IGF-I treated tendons were increased compared to controls. Mechanically, IGF-I treated tendons showed a trend toward increased stiffness compared to saline treated controls. Considered together with the decreased soft tissue swelling and improved sonographic healing, these data support the potential use of intralesional IGF-I for treatment of debilitating tendon injuries.     

Temporal and spatial expression of bone morphogenetic proteins in extracorporeal shock wave-promoted healing of segmental defect

Extracorporeal shock wave (ESW) is a noninvasive acoustic wave, which has recently been demonstrated to promote bone repair. The actual healing mechanism triggered by ESW has not yet been identified. Bone morphogenetic proteins (BMP) have been implicated as playing an important role in bone development and fracture healing. In this study, we aimed to examine the involvement of BMP-2, BMP-3, BMP-4, and BMP-7 expression in ESW promotion of fracture healing. Rats with a 5-mm segmental femoral defect were given ESW treatment using 500 impulses at 0.16 mJ/mm(2). Femurs and calluses were subjected to immunohistochemistry and RT-PCR assay 1, 2, 4, and 8 weeks after treatment. Histological observation demonstrated that fractured femurs received ESW treatment underwent intensive mesenchymal cell aggregation, hypertrophic chondrogenesis, and endochondral/intramembrane ossification, resulting in the healing of segmental defect. Aggregated mesenchymal cells at the defect, chondrocytes at the hypertrophic cartilage, and osteoblasts adjunct to newly formed woven bone showed intensive proliferating cell nuclear antigen expression. ESW treatment significantly promoted BMP-2, BMP-3, BMP-4, and BMP-7 mRNA expression of callus as determined by RT-PCR, and BMP immunoreactivity appeared throughout the bone regeneration period. Mesenchymal cells and immature chondrocytes showed intensive BMP-2, BMP-3, and BMP-4 immunoreactivity. BMP-7 expression was evident on osteoblasts located at endochondral ossification junction. Our findings suggest that BMP play an important role in signaling ESW-activated cell proliferation and bone regeneration of segmental defect.     

Shock wave application enhances pertussis toxin protein-sensitive bone formation of segmental femoral defect in rats.

Extracorporeal shock waves (ESWs) elicit a dose-dependent effect on the healing of segmental femoral defects in rats. After ESW treatment, the segmental defect underwent progressive mesenchymal aggregation, endochondral ossification, and hard callus formation. Along with the intensive bone formation, there was a persistent increase in TGF-beta1 and BMP-2 expression. Pretreatment with pertussis toxin reduced ESW-promoted callus formation and gap healing, which presumably suggests that Gi proteins mediate osteogenic signaling. INTRODUCTION: Extracorporeal shock waves (ESWs) have previously been used to promote bone repair. In our previous report, we found that ESWs promoted osteogenic differentiation of mesenchymal cells through membrane perturbation and activation of Ras protein. In this report, we show that ESWs elicit a dose-dependent effect on the healing of segmental defects and that Gi proteins play an important role in mediating ESW stimulation. MATERIALS AND METHODS: Rats with segmental femoral defects were subjected to ESW treatment at different energy flux densities (EFD) and impulses. Bone mass (mineral density and calcium content), osteogenic activities (bone alkaline phosphatase activity and osteocalcin content), and immunohistochemistry were assessed. RESULTS: An optimal ESW energy (500 impulses at 0.16 mJ/mm2 EFD) stimulated complete bone healing without complications. ESW-augmented healing was characterized by significant increases (p < 0.01) in callus size, bone mineral density, and bone tissue formation. With exposure to ESW, alkaline phosphatase activity and osteocalcin production in calluses were found to be significantly enhanced (p < 0.05). After ESW treatment, the histological changes we noted included progressive mesenchymal aggregation, endochondral ossification, and hard callus formation. Intensive bone formation was associated with a persistent increase in transforming growth factor-beta 1 (TGF-beta1) and bone morphogenetic protein-2 (BMP-2) expression, suggesting both growth factors were active in ESW-promoted bone formation. We also found that pertussis toxin, an inhibitor of membrane-bound Gi proteins, significantly reduced (p < 0.01) ESW promotion of callus formation and fracture healing. CONCLUSION: ESW treatments enhanced bone formation and the healing of segmental femoral defects in rats. It also seems likely that TGF-beta1 and BMP-2 are important osteogenic factors for ESW promotion of fracture healing, presumably through Gi protein-mediated osteogenic signaling.     

Recruitment of mesenchymal stem cells and expression of TGF-beta 1 and VEGF in the early stage of shock wave-promoted bone regeneration of segmental defect in rats.

Extracorporeal shock wave (ESW) treatment has recently been established as a method to enhance bone repair. Here, we reported that ESW-promoted healing of segmental defect via stimulation of mesenchymal stem cell recruitment and differentiation into bone forming cells. Rats with a segmental femoral defect were exposed to a single ESW treatment (0.16 mJ/mm(2), 1 Hz, 500 impulses). Cell morphology and histological changes in the defect region were assessed 3, 7, 14, and 28 days post-treatment. Presence of mesenchymal stem cell was assayed by immuno-staining for RP59, a recently discovered marker, and also production of TGF-beta 1 and VEGF was monitored. ESW treatment increased total cell density and the proportion of RP59 positive cells in the defect region. High numbers of round- and cuboidal-shaped cells strongly expressing RP59 were initially found. Later, the predominant cell type was spindle-shaped fibroblastic cells, subsequently, aggregates of osteogenic and chondrogenic cells were observed. Histological observation suggested that bone marrow stem cells were progressively differentiated into osteoblasts and chondrocytes. RP59 staining was initially intense and decreased with the appearance of expression depended on the differentiation states of osteogenic and chondrogenic cells during the regeneration phase. Mature chondrocytes and osteoblasts exhibited only slight RP59 immuno-reactivity. Expression of TGF-beta 1 and VEGF-A mRNA in the defect tissues was also significantly increased (P<0.05) after ESW treatment as determined by RT-PCR. Intensive TGF-beta 1 immuno-reactivity was induced immediately, whereas a lag period was observed for VEGF-A. Chondrocytes and osteoblasts at the junction of ossified cartilage clearly exhibited VEGF-A expression. Our findings suggest that recruitment of meseoblasts at the junction of ossified cartilage clearly exhibited mesenchymal stem cells is a critical step in bone reparation that is enhanced by ESW treatment. TGF-beta 1 and VEGF-A are proposed to play a chemotactic and mitogenic role in recruitment and differentiation of mesenchymal stem cells.     

Structural and biomechanical characteristics after early mobilization in an achilles tendon rupture model: operative versus nonoperative treatment

Acute Achilles tendon ruptures are common sports injuries; however, treatment remains a clinical challenge. Studies show a superior effect of early mobilization and full weight bearing on tendon healing and clinical outcome; however, few data exist on structural and biomechanical characteristics in the early healing phase. This study investigated the histological and biomechanical characteristics of early mobilization and full weight bearing in an Achilles tendon rupture model. Eighty rats underwent dissection of a hindpaw Achilles tendon; 40 rats were treated conservatively and 40 underwent open repair of the transected Achilles tendon by suturing. Early mobilization and full weight bearing were allowed in both groups. At 1, 2, 4, and 8 weeks after tenotomy, tensile strength, stiffness, thickness, tissue characteristics (histological analysis), and length were determined. Dissected Achilles tendons healed in all animals during full weight-bearing early mobilization. One and 2 weeks after tenotomy, rats in the operative group showed increased tensile strength and stiffness compared with the nonoperative group. Repair-site diameters were increased at 1, 2, and 8 weeks after tenotomy. Tendon length was decreased in the operative group throughout observation, whereas the nonoperative group showed increased structural characteristics on the cellular level and a more homogeneous collagen distribution. Surgical treatment of dissected rat Achilles tendons showed superior biomechanical characteristics within the first 2 weeks. Conservative treatment resulted in superior histological findings but significant lengthening of the tendon in the early healing phase (weeks 1-8).     

Activation of extracellular signal-regulated kinase (ERK) and p38 kinase in shock wave-promoted bone formation of segmental defect in rats

Extracorporeal shock waves (ESW) have recently been used in bone repair. Extracellular signal-regulated kinase (ERK) and p38 kinase are found to act as important mediators for osteogenic factor and mechanical-stimulated proliferation and differentiation of bone-forming cells. A previous study reported that ESW promoted healing of segmental defects in rats by inducing bone morphogenetic proteins (Bone 32 (2003) 387-396) and stimulating osteogenic differentiation of mesenchymal stem cells. In this study, we found that ERK and p38 activation was involved in ESW-augmented bone regeneration of segmental defects. ESW treatment (0.16 mJ/mm2, 1 Hz, 500 impulses) rapidly promoted [3H]-thymidine uptake in 1 day and progressively increased alkaline phosphatase activity, collagen I, II, and osteocalcin synthesis in callus organ culture within 14 days after treatment. Results of [gamma-32P]-phosphotransferase activity assay showed that ERK and p38 in calluses were rapidly activated 1 day and 7 days after ESW treatment, respectively. Histological observation showed that segmental defects subjected to ESW treatment underwent typical bone formation (mesenchymal cell aggregation, hypertrophic cartilage, and endochondral/intramembrane ossification). Intensive bone formation coincided with evident expression of phosphorylated ERK and p38. Moreover, expression of phosphorylated ERK persisted in mesenchymal, chondral, and osteoblastic cells at newly developed bone and cartilage, and the expression of activated p38 was evident on chondral cells located at hypertrophic cartilage. Our findings suggest that mitogen-activated protein kinases (MAPK) regulate the stimulation of biophysical ESW, triggering mitogenic and osteogenic responses in the defects. ERK phosphorylation is active throughout the period of ESW-induced bone regeneration. p38 activation most likely plays an important role in signaling cartilage formation in callus.     

GDF-5 deficiency in mice delays Achilles tendon healing.

The aim of this study was to examine the role of one of the growth/differentiation factors, GDF-5, in the process of tendon healing. Specifically, we tested the hypothesis that GDF-5 deficiency in mice would result in delayed Achilles tendon repair. Using histologic, biochemical, and ultrastructural analyses, we demonstrate that Achilles tendons from 8-week-old male GDF-5 -/- mice exhibit a short-term delay of 1-2 weeks in the healing process compared to phenotypically normal control littermates. Mutant animals took longer to achieve peak cell density, glycosaminoglycan content, and collagen content in the repair tissue, and the time course of changes in collagen fibril size was also delayed. Revascularization was delayed in the mutant mice by 1 week. GDF-5 deficient Achilles tendons also contained significantly more fat within the repair tissue at all time points examined, and was significantly weaker than control tissue at 5 weeks after surgery, but strength differences were no longer detectable by 12-weeks. Together, these data support the hypothesis that GDF-5 may play an important role in modulating tendon repair, and are consistent with previously posited roles for GDF-5 in cell recruitment, migration/adhesion, differentiation, proliferation, and angiogenesis.     

The effects of extracorporeal shock waves on the rat Achilles tendon: is there a critical dose for tissue injury?

Introduction Extracorporeal shock waves (ESW) have been extensively studied in the field of orthopedics. Experimental and focused, well-designed clinical studies have suggested the clinical utilisation of ESW in several pathologies including delayed bone union, tennis elbow, and plantar fasciitis. However, the unwanted detrimental effects of ESW on various tissues have been questioned by some authors. In this experimental study, the effects of ESW were investigated at different intensity applications on the Achilles tendons of rat.Materials and methods A total of 32 adult Wistar albino rats was divided into four groups. The first three groups received 1000 impulses of 0.15 mJ/mm², 1500 impulses of 0.15 mJ/mm², and 2000 impulses of 0.20 mJ/mm², respectively. The last group was kept as the control group. Subsequently, Achilles tendons were harvested for histological studies from all rats at the 3rd week after a single application of ESW.Results There were no histological abnormalities observed in the Achilles tendons of the first two groups compared with the control group. No alteration in the histological configuration was observed, and consequently the pathologist who had been blinded could not differentiate these rats from the control group by light microscopy. However, in the high intensity group (2000 impulses of 0.20 mJ/mm²), grade II and III disorganisation of collagen fibers was noticed in 7 out of 8 rats, which was not detected in any of the rats from the first two groups (p<0.05). Consequently, the pathologist could distinguish the majority of the rats (7 out of 8) of this group from the remaining ones. Meanwhile, grade I lymphocyte infiltration was observed in some sections of the rats receiving the highest ESW dose.Conclusion This study confirms that ESW application at high intensity is associated with detrimental tissue effects. Additionally, it was suggested that the extent of tissue injury caused by ESW is dose-related.     

Matrix remodeling in healing rabbit Achilles tendon

Biochemical, biomechanical and ultrastructural properties of the connective tissue matrix were investigated during the early remodeling phase of tissue repair in experimentally tenotomized and repaired rabbit Achilles tendons. Sterile surgical tenotomy was performed on the right Achilles tendons of 14 rabbits and allowed to heal for 15 days. The animals were euthanized and the Achilles tendons excised from both limbs. The left contralateral Achilles tendon of each rabbit was used as a control in the experiments. Prior to biochemical analysis, both intact and healing tendons were tested for their biomechanical integrity. The results revealed that the healing tendons had regained some of their physicochemical characteristics, but differed significantly from the intact left tendons. The healing tendons regained 48% tensile strength, 30% energy absorption, 20% tensile stress, and 14% Young's modulus of elasticity of intact tendons. In contrast, biochemical analysis showed that the healing tendons had 80% of the collagen and 60% of the collagen crosslinks (hydroxypyridinium) of normal tendons. Sequential extraction of collagen from the tissues yielded more soluble collagen in the healing tendons than intact tendons, suggesting either an increase in collagen synthesis and/or enhanced resorption of mature collagen in healing tendons compared to intact tendons. Electron microscopic studies revealed remarkable differences in the ultrastructure between intact and healing tendons. These observations could explain, in part, the connective tissue response to healing during the early phases of tissue remodeling.     

Effects of transforming growth factor-beta 1 on the early stages of healing of the Achilles tendon in a rat model.

We studied the effects of transforming growth factor-beta 1 (TGF-beta 1) on the genetic expression of procollagen type I and III and its effects on structural properties in the early stages of healing in rat Achilles tendon. The Achilles tendons in 90 rats were transsected and repaired immediately. TGF-beta 1 dissolved in phosphate-buffered saline was injected locally at the repair site using two different doses, and outcomes in both groups were compared to that in the control group given phosphate-buffered saline only. Five animals in each group were killed at one, two, and four weeks postoperatively, and the healing tendon was evaluated. A dose-dependent increase in the expression of procollagen type I and III mRNA was found one week postoperatively. The failure load and stiffness of the healing tendon were increased by TGF-beta 1 at two and four weeks.     

An in-vivo experimental evaluation of He–Ne laser photostimulation in healing Achilles tendons

There is no method of treatment that has been proven to accelerate the rate of tendon healing or to improve the quality of the regenerating tendon. Low level laser photostimulation has gained a considerable attention for enhancing tissue repair in a wide spectrum of applications. However, there is controversy regarding the effectiveness of laser photostimulation for improvement of the healing process of surgically repaired tendons. Accordingly, the present study was conducted to evaluate the role of helium–neon (He–Ne) laser photostimulation on the process of healing of surgically repaired Achilles tendons. Thirty unilateral Achilles tendons of 30 Raex rabbits were transected and immediately repaired. Operated Achilles tendons were randomly divided into two equal groups. Tendons at group A were subjected to He–Ne laser (632.8 nm) photostimulation, while tendons at group B served as a control group. Two weeks later, the repaired Achilles tendons were histopathologically and biomechanically evaluated. The histopathological findings suggest the favorable qualitative pattern of the newly synthesized collagen of the regenerating tendons after He–Ne laser photostimulation. The biomechanical results support the same favorable findings from the functional point of view as denoted by the better biomechanical properties of the regenerating tendons after He–Ne laser photostimulation with statistical significance (p ≤ 0.01) at most of the biomechanical parameters. He–Ne laser photostimulation reported a great value after surgical repair of ruptured and injured tendons for a better functional outcome. It could be applied safely and effectively in humans, especially with respect to the proposed long-term clinical outcome.     

Achilles tendon healing: A correlation between functional and mechanical performance in the rat

The pathogenesis and treatment of rupture of the Achilles tendon remain a source of controversy. This study presents the results of a biomechanical, functional, and morphological evaluation of a group of rats that had division and repair of the Achilles tendon. A total of 46 rats were used: 18 for biomechanical testing, 18 for functional evaluation, and 10 for histology. Morphological examination revealed an early inflammatory response with loose connective tissue formation that was replaced gradually by fibroblasts and a collagenous matrix. The functional evaluation (Achilles functional index [AFI]) was made from measurements of the hind pawprints of walking rats. Division and repair of the Achilles tendon produced a significant functional impairment (mean [±SEM] AFI = ?87 ± 8; p < 0.001), which gradually improved with healing time. The load to failure for the repaired tendons consistently improved with healing time, in a manner similar to the functional recovery. The average deformation (repair/control) varied considerably and was not related to healing time. The stiffness of the repaired tendons increased with healing time and was 60% of the corresponding control side by day 15. The major finding of this study was a strong correlation between the AFI and the failure load of the healing tendon-bone constructs (250–300 g group, r = 0.97, p < 0.001; 325–375 g group, r = 0.96, p < 0.001).     

Nonsurgical treatment and early return to activity leads to improved Achilles tendon fatigue mechanics and functional outcomes during early healing in an animal model

Achilles tendon ruptures are common and devastating injuries; however, an optimized treatment and rehabilitation protocol has yet to be defined. Therefore, the objective of this study was to investigate the effects of surgical repair and return to activity on joint function and Achilles tendon properties after 3 weeks of healing. Sprague–Dawley rats (N = 100) received unilateral blunt transection of their Achilles tendon. Animals were then randomized into repaired or non‐repaired treatments, and further randomized into groups that returned to activity after 1 week (RTA1) or after 3 weeks (RTA3) of limb casting in plantarflexion. Limb function, passive joint mechanics, and tendon properties (mechanical, organizational using high frequency ultrasound, histological, and compositional) were evaluated. Results showed that both treatment and return to activity collectively affected limb function, passive joint mechanics, and tendon properties. Functionally, RTA1 animals had increased dorsiflexion ROM and weight bearing of the injured limb compared to RTA3 animals 3‐weeks post‐injury. Such functional improvements in RTA1 tendons were evidenced in their mechanical fatigue properties and increased cross sectional area compared to RTA3 tendons. When RTA1 was coupled with nonsurgical treatment, superior fatigue properties were achieved compared to repaired tendons. No differences in cell shape, cellularity, GAG, collagen type I, or TGF‐β staining were identified between groups, but collagen type III was elevated in RTA3 repaired tendons. The larger tissue area and increased fatigue resistance created in RTA1 tendons may prove critical for optimized outcomes in early Achilles tendon healing following complete rupture. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:2172–2180, 2016.     

Smad8/BMP2‐engineered mesenchymal stem cells induce accelerated recovery of the biomechanical properties of the achilles tendon

Tendon tissue regeneration is an important goal for orthopedic medicine. We hypothesized that implantation of Smad8/BMP2‐engineered MSCs in a full‐thickness defect of the Achilles tendon (AT) would induce regeneration of tissue with improved biomechanical properties. A 2 mm defect was created in the distal region of murine ATs. The injured tendons were then sutured together or given implants of genetically engineered MSCs (GE group), non‐engineered MSCs (CH3 group), or fibrin gel containing no cells (FG group). Three weeks later the mice were killed, and their healing tendons were excised and processed for histological or biomechanical analysis. A biomechanical analysis showed that tendons that received implants of genetically engineered MSCs had the highest effective stiffness (>70% greater than natural healing, p < 0.001) and elastic modulus. There were no significant differences in either ultimate load or maximum stress among the treatment groups. Histological analysis revealed a tendon‐like structure with elongated cells mainly in the GE group. ATs that had been implanted with Smad8/BMP2‐engineered stem cells displayed a better material distribution and functional recovery than control groups. While additional study is required to determine long‐term effects of GE MSCs on tendon healing, we conclude that genetically engineered MSCs may be a promising therapeutic tool for accelerating short‐term functional recovery in the treatment of tendon injuries. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 30:1932–1939, 2012     

Thrombocytopenia alters early but not late repair in a mouse model of Achilles tendon injury

Besides their hemostatic function, platelets can express key factors involved in tissue healing. However, the role of platelets in tendon healing following acute injury is poorly understood. We investigated this role by injecting male C57BL/6 mice with an antiplatelet antibody to induce thrombocytopenia. Placebo animals received serum only. The right Achilles tendon was sectioned and sutured using the 8-strand technique that allows immediate weight bearing. Platelet depletion did not alter the accumulation of neutrophils and macrophages or cell proliferation. A slight increase in vascularization was observed 7 days postinjury in tendons from thrombocytopenic mice relative to placebo animals, but the effect had disappeared by day 14. Furthermore, collagen content had a tendency to decrease in Achilles tendons under thrombocytopenia when compared with placebo treatment at 7 days posttrauma. This was correlated with a decline in maximal stress sustained by tendons at day 14 but not after 28 days. The impact of thrombocytopenia was otherwise negligible, as force relaxation and stiffness were similar in the two groups. Our findings demonstrate that platelets modulate early tendon repair following rupture, although the effect is limited over time. Nevertheless, platelets are not essential for the recruitment of inflammatory cells, proliferation, angiogenesis, and tendon maturation.     

Early nerve regeneration after Achilles tendon rupture — a prerequisite for healing? A study in the rat

Nerve regeneration during healing of Achilles tendon rupture in the rat was studied by immunohistochemistry including semi-quantitative assessment. Neuronal markers for regenerating and mature fibers, ie., growth associated protein 43 (GAP-43) and protein gene product 9.5 (PGP 9.5), respectively, were analyzed at different time points (1-16 weeks) post-rupture. In the paratenon, both the ruptured and intact contralateral tendon (control) consistently exhibited immunoreactivity to the two neuronal markers. However, in the proper tendinous tissue only the ruptured tendon showed immunoreactivity to GAP-43 and PGP 9.5. This expression was seen already at week 1 post-rupture to reach a peak at week 6 followed by a successive drop till week 16. Also the occurrence of sensory and autonomic fibers according to immunoreactivity for calcitonin gene-related peptide (CGRP) and neuropeptide Y (NPY), respectively, was analyzed. CGRP-positivity was abundantly seen from weeks 2-6 in both perivascular and sprouting free nerve endings in the proper tendon tissue undergoing healing. NPY appeared later, at weeks 6-8 post-rupture around blood vessels mainly located in the surrounding loose connective tissue. Apart from a role in vasoaction (CGRP, vasodilatory; NPY, vasoconstrictory). both neuropeptides have been implicated in fibroblast and endothelial cell proliferation required for angiogenesis. The present study shows that early healing of ruptured tendons is characterized by an orchestrated, temporal appearance of nerve fibers expressing peptides with different actions. The observed pattern of neuronal regeneration and neuropeptide expression may prove to be important for normal connective tissue healing.     

Unfavorable effect of knee immobilization on Achilles tendon healing in rabbits

This study was undertaken to assess the effect of knee immobilization on the treatment of Achilles tendon rupture. After their Achilles tendons were severed, rabbits were divided into 2 groups. In Group A, only the ankle joint was immobilized. In Group B, both the knee and ankle joints were immobilized. At 4 weeks after surgery, both the ultimate tensile force and stiffness of the severed tendons were significantly greater in Group A than in Group B. In Group A, dense collagen fibers were seen in the repaired tendons, and the bundles of collagen fibers were parallel to one another along the axis of the tendons. In contrast, in Group B, dilated veins and capillaries were seen in the repaired tendons, and the proliferation of connective tissue containing collagen fibers was severely reduced around these veins and capillaries and was in general irregular and uneven. These results suggest that knee immobilization retards the healing of a ruptured Achilles tendon without suture, due to congestion and tension deprivation produced by keeping the tendon static.     

In-Vivo Efficacy of Recombinant Human Hyaluronidase (rHuPH20) Injection for Accelerated Healing of Murine Retrocalcaneal Bursitis and Tendinopathy

The deposition of aggrecan/hyaluronan (HA)-rich matrix within the tendon body and surrounding peritenon impede tendon healing and result in compromised biomechanical properties. Hence, the development of novel strategies to achieve targeted removal of the aggrecan-HA pericellular matrix may be effective in treating tendinopathy. The current study examined the therapeutic potential of a recombinant human hyaluronidase, rHuPH20 (FDA approved for reducing HA accumulation in tumors) for treating murine Achilles tendinopathy. The 12-week-old C57Bl/6 male mice were injected with two doses of rHuTGF-β1 into the retrocalcaneal bursa (RCB) to induce a combined bursitis and tendinopathy. Twenty-four hours following induction of injury, treatment groups were administered rHuPH20 Hyaluronidase (rHuPH20; Halozyme Therapeutics) into the RCB. At either 6 h (acute), 9 days, or 25 days following hyaluronidase treatment, Achilles tendons were analyzed for gene expression, histology and immunohistochemistry, fluorophore-assisted carbohydrate electrophoresis, and biomechanical properties. The rHuPH20 treatment was effective, particularly at the acute and 9-day time points, in (a) removing HA deposits from the Achilles tendon and surrounding tissues, (b) improving biomechanical properties of the healing tendon, and (c) eliciting targeted increases in expression of specific cell fate, extracellular matrix metabolism, and inflammatory genes. The potential of rHuPH20 to effectively clear the pro-inflammatory, HA-rich matrix within the RCB and tendon strongly supports the future refinement of injectable glycosidase preparations as potential treatments to protect or regenerate tendon tissue by reducing inflammation and scarring in the presence of bursitis or other inducers of damage such as mechanical overuse. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:59–69, 2020     

Tendon-derived stem cells (TDSCs) promote tendon repair in a rat patellar tendon window defect model

Injured tendons heal slowly and often result in the formation of mechanically and functionally inferior fibrotic scar tissue or fibrous adhesions. This study investigated the use of tendon-derived stem cells (TDSCs) for tendon repair in a rat patellar tendon window defect model. Fibrin glue constructs with or without GFP-TDSCs were transplanted into the window defect. The patellar tendons were harvested for histology, ex vivo fluorescent imaging and biomechanical test at various time points up to week 4. Our results showed that TDSCs significantly enhanced tendon healing as indicated by the increase in collagen production as shown by hematolxylin stain-ability of the tissue, improvement of cell alignment, collagen fiber alignment and collagen birefringence typical of tendon. The labeled cells were observed at weeks 1 and 2 and became almost undetectable at week 4. Both the ultimate stress and Young's modulus were significantly higher in the TDSCs group compared to those in the fibrin glue group at week 4. In conclusion, TDSCs promoted earlier and better repair in a rat patellar tendon window defect model.     

Unfavorable effect of knee immobilization on Achilles tendon healing in rabbits

This study was undertaken to assess the effect of knee immobilization on the treatment of Achilles tendon rupture. After their Achilles tendons were severed, rabbits were divided into 2 groups. In Group A, only the ankle joint was immobilized. In Group B, both the knee and ankle joints were immobilized. At 4 weeks after surgery, both the ultimate tensile force and stiffness of the severed tendons were significantly greater in Group A than in Group B. In Group A, dense collagen fibers were seen in the repaired tendons, and the bundles of collagen fibers were parallel to one another along the axis of the tendons. In contrast, in Group B, dilated veins and capillaries were seen in the repaired tendons, and the proliferation of connective tissue containing collagen fibers was severely reduced around these veins and capillaries and was in general irregular and uneven. These results suggest that knee immobilization retards the healing of a ruptured Achilles tendon without suture, due to congestion and tension deprivation produced by keeping the tendon static.