George A. C. Murrell

The roles of growth factors in tendon and ligament healing.

AbstractTendon healing is a complex and highly-regulated process that is initiated, sustained and eventually terminated by a large number and variety of molecules. Growth factors represent one of the most important of the molecular families involved in healing, and a considerable number of studies have been undertaken in an effort to elucidate their many functions. This review covers some of the recent investigations into the roles of five growth factors whose activities have been best characterised during tendon healing: insulin-like growth factor-I (IGF-I), transforming growth factor β (TGFβ), vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), and basic fibroblast growth factor (bFGF). All five are markedly up-regulated following tendon injury and are active at multiple stages of the healing process. IGF-I has been shown to be highly expressed during the early inflammatory phase in a number of animal tendon healing models, and appears to aid in the proliferation and migration of fibroblasts and to subsequently increase collagen production. TGFβ is also active during inflammation, and has a variety of effects including the regulation of cellular migration and proliferation, and fibronectin binding interactions. VEGF is produced at its highest levels only after the inflammatory phase, at which time it is a powerful stimulator of angiogenesis. PDGF is produced shortly after tendon damage and helps to stimulate the production of other growth factors, including IGF-I, and has roles in tissue remodelling. In vitro and in vivo studies have shown that bFGF is both a powerful stimulator of angiogenesis and a regulator of cellular migration and proliferation. This review also covers some of the most recent studies into the use of these molecules as therapeutic agents to increase the efficacy and efficiency of tendon and ligament healing. Studies into the effects of the exogenous application of TGFβ, IGF-I, PDGF and bFGF into the wound site singly and in combination have shown promise, significantly decreasing a number of parameters used to define the functional deficit of a healing tendon. Application of IGF-I has been shown to increase in the Achilles Functional Index and the breaking energy of injured rat tendon. TGFβ and PDGF have been shown separately to increase the breaking energy of healing tendon. Finally, application of bFGF has been shown to promote cellular proliferation and collagen synthesis in vivo.

Osteoporosis influences the early period of fracture healing in a rat osteoporotic model

Osteoporotic fractures commonly occur in the elderly. Although current therapies are aimed at the prevention and treatment of osteoporotic fractures, studies examing the fracture healing process in osteoporotic bone are limited. We produced an osteoporotic rat model by ovariectomy (ovx) and maintained a low calcium diet (LCD) in order to evaluate the influence of osteoporosis on fracture healing. Callus formation and strength was monitored over a 3 week period by histological and biomechanical assessment. Data collected simultaneously on a group of rats undergoing sham surgery (sx) were used for comparison. A 40% reduction in fracture callus cross-sectional area and a 23% reduction in bone mineral density in the healing femur of the ovx rats was observed on day 21 following fracture as compared with the sx group (p < 0.01). Biomechanical data from the healing femur of the ovx rats revealed a fivefold decrease in the energy required to break the fracture callus, a threefold decrease in peak failure load, a twofold decrease in stiffness and a threefold decrease in stress as compared with the sx group (p < 0.01, respectively). Histomorphological analysis revealed a delay in fracture callus healing with poor development of mature bone in the ovx rats. This study provides physical evidence of altered fracture healing in osteoporotic bone, which may have important implications in evaluating the effects of new treatments for osteoporosis on fracture healing.

Treatment of Tendinopathy: What Works, What Does Not, and What is on the Horizon

Tendinopathy is a broad term encompassing painful conditions occurring in and around tendons in response to overuse. Recent basic science research suggests little or no inflammation is present in these conditions. Thus, traditional treatment modalities aimed at controlling inflammation such as corticosteroid injections and nonsteroidal antiinflammatory medications (NSAIDS) may not be the most effective options. We performed a systematic review of the literature to determine the best treatment options for tendinopathy. We evaluated the effectiveness of NSAIDS, corticosteroid injections, exercise-based physical therapy, physical therapy modalities, shock wave therapy, sclerotherapy, nitric oxide patches, surgery, growth factors, and stem cell treatment. NSAIDS and corticosteroids appear to provide pain relief in the short term, but their effectiveness in the long term has not been demonstrated. We identified inconsistent results with shock wave therapy and physical therapy modalities such as ultrasound, iontophoresis and low-level laser therapy. Current data support the use of eccentric strengthening protocols, sclerotherapy, and nitric oxide patches, but larger, multicenter trials are needed to confirm the early results with these treatments. Preliminary work with growth factors and stem cells is promising, but further study is required in these fields. Surgery remains the last option due to the morbidity and inconsistent outcomes. The ideal treatment for tendinopathy remains unclear.Level of Evidence: Level II, systematic review. See the Guidelines for Authors for a complete description of levels of evidence.

Genotype dependent and cigarette specific effects on endothelial nitric oxide synthase gene expression and enzyme activity

We explored the interactive effects of endothelial nitric oxide synthase (eNOS) genotypes and cigarette smoking on protein levels and enzyme activity in 33 postpartum placentas. Whilst the eNOS protein levels were lower in the rare allele (0.48±0.11, n=9 vs. 1.05±0.10, n=24, P<0.01), the eNOS enzyme activity was about 7‐fold higher in the rare allele (4556.2±255.4 vs. 621.8±180.5 cpm/mg/min, P<0.01). Smokers had lower eNOS protein levels (1.07±0.09 vs. 0.50±0.19, P<0.05) in both alleles. It reduced the eNOS activities only in the rare allele (non‐smokers: 6143.8±251.2, n=5, smokers: 2968.5±259.4, n=4, 52% reduction, P<0.01). We conclude that associations between eNOS polymorphism and protein levels and enzyme activities are modifiable by smoking, the effects of smoking are dependent on the eNOS genotypes.

Apoptosis in rotator cuff tendonopathy.

The aim of this study was to investigate the involvement of apoptosis (programmed cell death) in the pathogenesis of rotator cuff disorders. The edges of torn supraspinatus rotator cuff tendons were collected from patients with rotator cuff tear (n = 25). Samples of the intra‐articular portion of subscapularis tendons were collected from patients without rotator cuff tear as control (n = 6). To minimize individual variance, we also collected six pairs of supraspinatus tendon and subscapularis tendon from six patients with rotator cuff tears. Apoptosis was detected by in situ DNA end labelling assay and DNA laddering assay. Immunohistochemical staining was performed to identify cells undergoing apoptosis. Control subscapularis tendon had normal morphology. Tendon from torn supraspinatus rotator cuff showed significant mucoid degeneration. Within the areas of degeneration, there were large numbers of apoptotic cells. The percentage of apoptotic cells in the degenerative rotator cuff (34%) was significantly higher than that in controls (13%) (p < 0.001). The excessive apoptosis detected in degenerative rotator cuff tissue was confirmed by DNA laddering assays. This is the first report of excessive apoptosis in degenerating rotator cuff tendon. Cells undergoing apoptosis in rotator cuff were mainly fibroblast‐like cells. These finding indicate that apoptosis may play an important role in the pathogenesis of rotator cuff degeneration.

Restore Orthobiologic Implant: Not Recommended for Augmentation of Rotator Cuff Repairs

BACKGROUND Following repairs of large-to-massive tears of the rotator cuff, the rates of tendon retears are high and often involve tissue deficiency. Animal studies of the Restore Orthobiologic Implant, a collagen-based material derived from the small intestine mucosa of pigs, have indicated that it might be used to help overcome such problems. We carried out a study to determine whether patients who received this xenograft to augment a rotator cuff repair exhibited greater shoulder strength, shoulder function, and/or resistance to retearing. METHODS We compared data from a group of patients who had undergone conventional rotator cuff repair with xenograft augmentation (the xenograft group) with data from a group in whom a repair had been done by the same surgeon without augmentation (the controls). The groups were matched for gender, mean age, and mean size of the rotator cuff tear. All subjects completed a pain and function questionnaire and were given a systematic clinical shoulder examination preoperatively and at three, six, and twenty-four months postoperatively. The twenty-four-month visit included magnetic resonance imaging to determine whether a retear had occurred. RESULTS Four patients who had received a xenograft had a severe postoperative reaction requiring surgical treatment. At two years after the surgery, six of the ten tendons repaired with a xenograft and seven of the twelve control tendons had retorn, as documented by magnetic resonance imaging. The patients with a xenograft had significantly less lift-off strength, as measured with a dynamometer, and significantly less strength in internal rotation and adduction than the controls at two years after the surgery (all p < 0.05). Also, the xenograft group had significantly more impingement in external rotation, a slower rate of resolution of pain during activities, more difficulty with hand-behind-the-back activities, and less sports participation (all p < 0.05). CONCLUSIONS Two years after surgical repair of a large rotator cuff defect supplemented with a xenograft, patients had several persisting deficits and no recognizable benefit as compared with the results in a control group. In view of these findings, together with the unsatisfactorily high proportion of patients with a severe inflammatory reaction to the xenograft, we do not recommend use of the Restore Orthobiologic Implant in its present form. LEVEL OF EVIDENCE Therapeutic Level III. See Instructions to Authors for a complete description of levels of evidence.

The Basic Science of Tendinopathy

Tendinopathy is a common clinical problem with athletes and in many occupational settings. Tendinopathy can occur in any tendon, often near its insertion or enthesis where there is an area of stress concentration, and is directly related to the volume of repetitive load to which the tendon is exposed. Recent studies indicate tendinopathy is more likely to occur in situations that increase the “dose” of load to the tendon enthesis – including increased activity, weight, advancing age, and genetic factors. The cells in tendinopathic tendon are rounder, more numerous, and show evidence of oxidative damage and more apoptosis. These cells also produce a matrix that is thicker and weaker with more water, more immature and cartilage-like matrix proteins, and less organization. There is now evidence of a population of regenerating stem cells within tendon. These studies suggest prevention of tendinopathy should be directed at reducing the volume of repetitive loads to below that which induces oxidative-induced apoptosis and cartilage-like genes. The management strategies might involve agents or cells that induce tendon stem cell proliferation, repair and restoration of matrix integrity.

Topical glyceryl trinitrate treatment of chronic noninsertional achilles tendinopathy: A randomized, double-blind, placebo-controlled trial

BACKGROUND Noninsertional Achilles tendinopathy is a degenerative overuse disorder. No method has been universally successful in treating this condition. Topically applied nitric oxide has been shown, in animal models, to be effective for the treatment of fractures and cutaneous wounds through mechanisms that may include stimulation of collagen synthesis in fibroblasts. The goal of the present study was to determine if topical glyceryl trinitrate improves clinical outcome measures in patients with Achilles tendinopathy. METHODS A prospective, randomized, double-blind, placebo-controlled trial involving a total of sixty-five patients (eighty-four Achilles tendons) was performed to compare continuous application of topical glyceryl trinitrate (at a dosage of 1.25 mg per twenty-four hours) with rehabilitation alone for the treatment of noninsertional Achilles tendinopathy. RESULTS Compared with the control group, the glyceryl trinitrate group showed reduced pain with activity at twelve weeks (p = 0.02) and twenty-four weeks (p = 0.03), reduced night pain at twelve weeks (p = 0.04), reduced tenderness at twelve weeks (p = 0.02), decreased pain scores after the hop test at twenty-four weeks (p = 0.005), and increased ankle plantar flexor mean total work compared with the baseline level at twenty-four weeks (p = 0.04). Twenty-eight (78%) of thirty-six tendons in the glyceryl trinitrate group were asymptomatic with activities of daily living at six months, compared with twenty (49%) of forty-one tendons in the placebo group (p = 0.001, chi-square analysis). The mean effect size for all outcome measures was 0.14. CONCLUSIONS Topical glyceryl trinitrate significantly reduced pain with activity and at night, improved functional measures, and improved outcomes in patients with Achilles tendinopathy.

Early Inflammatory Reaction After Rotator Cuff Repair With a Porcine Small Intestine Submucosal Implant A Report of 4 Cases

Background Porcine small intestine submucosal grafts have been successful in enhancing soft tissue repair, as demonstrated by animal studies. Currently, there are no reports of the use of such implants in human rotator cuff repair. Study Design Case series; Level of evidence, 4. Methods Over a 6-month period, 25 patients underwent rotator cuff repair by one surgeon using the Restore Orthobiologic Implant to augment the repaired tendon or fill a defect. Results Four of 25 patients experienced an overt inflammatory reaction at a mean of 13 days postoperatively. All patients underwent open irrigation and debridement of the rotator cuff and porcine small intestine submucosal implant. Conclusion Porcine small intestine submucosal implants should be used in rotator cuff surgery with the awareness that a non-specific inflammatory reaction can occur in the early postoperative period. This inflammatory reaction may cause breakdown of the repair. Further studies are needed to further characterize the reaction and determine which patients are susceptible.

Diagnosis of rotator cuff tears

Rotator cuff tears account for almost 50% of major shoulder injuries but are sometimes difficult to diagnose. To aid diagnosis, we did a prospective study, comparing results of 23 clinical tests from 400 patients with and without rotator cuff tears. Three simple tests were predictive for rotator cuff tear: supraspinatus weakness, weakness in external rotation, and impingement. When all three were positive, or if two tests were positive and the patient was aged 60 or older, the individual had a 98% chance of having a rotator cuff tear; combined absence of these features excluded this diagnosis.