Differential MMP 1 and MMP 13 expression in proliferation and ligamentization phases of graft remodeling in anterior cruciate ligament reconstruction.

Purpose: Graft remodeling in anterior cruciate ligament reconstruction (ACLR) demonstrates three distinct phases: necrosis, proliferation and ligamentization. Biological enhancement involves modulating these processes, but the cellular activities related to extracellular matrix remodeling have not been investigated. We hypothesized that changes in matrix metalloproteinases (MMPs) 1 and 13 expression are involved in the transition of proliferation phase to ligamentization phase of graft remodeling.Materials and methods: Thirty-three rats underwent ACLR. Tendon grafts were harvested at week 1 (necrosis), 2 (proliferation), or 12 (ligamentization) post-operation for histological examination (n = 3), or for isolation of graft-derived cells (n = 8) for flow cytometry, proliferation assay, cell invasion assay, measurement of gene expression related to matrix remodeling (Col1A1, Col3A1, MMP1, tissue inhibitor of marix metalloproteinase 1 (TIMP1), and MMP13) and total MMP activities.Results: Increased cellularity in tendon graft was contributed by active cell proliferation and migration at week 2 post-operation, while decreased cellularity were paralleled by increased apoptosis at week 12. All genes measured (Col1A1, Col3A1, MMP1, TIMP1, and MMP13) increased significantly in week 2 cells compared to week 1 cells. MMP1 expression subsided at week 12, while MMP13 expression kept increasing till 12 weeks post-operation. Total MMP activities was 3-fold higher in cultured graft-derived cells from week 2 as compared to cells from week 12. Two distinct processes of graft remodeling were identified, characterized by increased MMP1 expression with cell proliferation and increased MMP13 expression with cell apoptosis.Conclusions: Unfavorable matrix remodeling during the proliferation phase is found with increased MMP1, while remodeling leading to ligamentization is associated with increased MMP13 expression.

Hydrogen peroxide induced tendinopathic changes in a rat model of patellar tendon injury.

Tendinopathy includes cases with chronic tendon pain and spontaneous tendon ruptures, which is putatively resulted from failed tendon healing. Overuse is a major risk factor of tendinopathy, which can impose mechanical and oxidative stress to tendons. Previous studies investigated the influences of mechanical stress, but the direct impact of oxidative stress on tendon healing remains unclear. We hypothesized that imposed oxidative stress can impair tendon healing and lead to tendinopathic changes. Thirty-nine rats were operated for patellar tendon window injury. From weeks 3-5 post-operation, the rats received three weekly subcutaneous injections of saline, 50 or 500 µM H2 O2 (n = 13) over patellar tendon. Gait analysis for pain assessment and 3D ultrasound imaging for detection of tendinopathic changes were performed at pre-injury and 6-week post-operation. At week 6, knee specimens were harvested for histology or tensile mechanical test. Elastic modulus of the healing patellar tendons was significantly lower in 50 µM but not 500 µM H2 O2 group, while ultimate mechanical stress was not significantly different across groups. Similarly, only the 50 µM H2 O2 group exhibited pain-associated gait asymmetry. Significant tendon swelling with increased tendon volume was observed in the 50 µM H2 O2 group. There were hypoechogenic changes in the tendon wound, but there was no significant difference in percentage vascularity. H2 O2 impaired tendon healing and elicited tendinopathic changes, with respect to pain and structural abnormalities. Oxidative stress plays a role in the failed tendon healing of tendinopathies, and H2 O2 -induced failed tendon healing may serve as a good animal model to study tendinopathy. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:3268-3274, 2018.

Anterior internal impingement of the shoulder in rugby players and other overhead athletes.

BACKGROUND/OBJECTIVE: Impingement syndromes are a common cause of shoulder pain in overhead athletes. Anterior internal impingement is a recently suggested mechanism for activity-related pain of the shoulder. Impingement syndromes were initially described to occur due to repetitive or excessive contact between the rotator cuff and other structures in the shoulder. METHODS: This is a retrospective, clinical case study reporting 54 consecutive cases of anterior internal impingement in overhead athletes, of which 28 (51.2%) cases were of rugby players. All had undergone physiotherapy without relief of the symptoms. Of 54, there were 45 male and 9 female patients with an average age of 27 years (range, 17-51). The mean duration from injury to surgery was 40.7 weeks (range, 5-364). Of the 54 patients, 29 (53.7%) were full-time professional and 25 (46.3%) were semiprofessional or recreational athletes. The players associated the onset of pain occurred following an injury in 29/54 cases (53.4%), whereas in the remaining 25 cases (46.2%), a gradual onset of symptoms was described. All 54 patients could demonstrate a "functional impingement sign" in positioning their arm and provoke pain. RESULT: On examination, the examiner could reproduce the same pain in 38/54 (70.3%) patients only. Of the 54 patients, "SLAP tests" including O'Brien's test, Palm up test, and compression rotation test were positive in shoulders of 39 (72.2 %) patients, Jobe's test in 27 (50%), Gerber's lift off test in 6 (11%), and Hawkin's test in 6 (11%) patients. During arthroscopic assessment, impinging flap tears were found in 44 (81.4%) patients from the SLAP, whereas undersurface rotator cuff flap tears were found in 24 (44.4%), flap tears from the anterior or inferior labrum were found in 16 (29.6%), and distal subscapularis flap tears were found in 10 (18.5%) patients. Only in 12/54 patients (22.2%) was an isolated pathology found, in all cases SLAP tears. Treatment included vaporisation and excision of the impinging flaps. In 15/54 (27.7%) patients, repair of an unstable SLAP tear was undertaken using absorbable suture anchors and fibre wires. All athletes returned to their previous activity level within 17.2 weeks (range, 6-36) from surgery and were discharged when they claimed that they were symptom free. CONCLUSION: This series of anterior internal impingement, which we believe is the largest in the literature to date, demonstrates the value of an to assess and successfully treat overhead athletes with anterior impingement syndrome.

Presence of Bacteria in Spontaneous Achilles Tendon Ruptures.

BACKGROUND: The structural pathology of Achilles tendon (AT) ruptures resembles tendinopathy, but the causes remain unknown. Recently, a number of diseases were found to be attributed to bacterial infections, resulting in low-grade inflammation and progressive matrix disturbance. The authors speculate that spontaneous AT ruptures may also be influenced by the presence of bacteria. HYPOTHESIS: Bacteria are present in ruptured ATs but not in healthy tendons. STUDY DESIGN: Cross-sectional study; Level of evidence, 3. METHODS: Patients with spontaneous AT ruptures and patients undergoing anterior cruciate ligament (ACL) reconstruction were recruited for this study. During AT surgical repair, excised tendinopathic tissue was collected, and healthy tendon samples were obtained as controls from hamstring tendon grafts used in ACL reconstruction. Half of every sample was reserved for DNA extraction and the other half for histology. Polymerase chain reaction (PCR) was conducted using 16S rRNA gene universal primers, and the PCR products were sequenced for the identification of bacterial species. A histological examination was performed to compare tendinopathic changes in the case and control samples. RESULTS: Five of 20 AT rupture samples were positive for the presence of bacterial DNA, while none of the 23 hamstring tendon samples were positive. Sterile operating and experimental conditions and tests on samples, controlling for harvesting and processing procedures, ruled out the chance of postoperative bacterial contamination. The species identified predominantly belonged to the Staphylococcus genus. AT rupture samples exhibited histopathological features characteristic of tendinopathy, and most healthy hamstring tendon samples displayed normal tendon features. There were no apparent differences in histopathology between the bacterial DNA-positive and bacterial DNA-negative AT rupture samples. CONCLUSION: The authors have demonstrated the presence of bacterial DNA in ruptured AT samples. It may suggest the potential involvement of bacteria in spontaneous AT ruptures.

Tripeptide-copper complex GHK-Cu (II) transiently improved healing outcome in a rat model of ACL reconstruction.

After anterior cruciate ligament reconstruction (ACLR), the biological healing of the graft is a rate-limiting step which can contribute to graft failure. The tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu(II) (GHK-Cu) is a well-known activator of tissue remodeling. We investigated whether GHK-Cu can improve graft healing following ACLR. Seventy-two rats underwent unilateral ACLR were randomized to saline, 0.3 or 3 mg/ml GHK-Cu groups (n = 24). Post-operational intra-articular injections were given from week 2, once a week, for 4 weeks. Gait analysis was performed pre-injury and at harvesting time. At 6 or 12 weeks post-operation, knee specimens were harvested for knee laxity test, graft pull-out test, and histology. At 6 weeks post-ACLR, GHK-Cu groups resulted in a smaller side-to-side difference in knee laxity as compared to the saline group (p = 0.009), but there was no significant difference at 12 weeks post-operation. The graft complex in the 0.3 mg/ml GHK-Cu group had higher stiffness than saline group at 6 weeks post-operation (p = 0.026), but there was no significant difference in ultimate load, gait parameters, and histological scores among treatment groups. All grafts failed mid-substance during pull-out test. Intra-articular supplementation with a bioactive small molecule GHK-Cu improved graft healing following ACLR in rat, but the beneficial effects could not last as treatment discontinued.

Shoulder injuries in professional rugby: a retrospective analysis.

BACKGROUND: In the literature, little is known about the level and pattern of rugby injuries. Of the shoulder injuries reported, 51% of these are caused during a tackle, and 65% of all match injuries affected the shoulder. OBJECTIVE: The study aims to describe a sport-specific unique intra-articular shoulder pathology of professional rugby players, who presented with persistent pain and dysfunction despite physiotherapeutic treatment and rest. METHOD: This study is a retrospective analysis set at a university sports medicine clinic. Eighty-seven professional rugby players, referred by their professional medical team since they could no longer play, underwent shoulder arthroscopy between June 2001 and October 2007 due to persistent shoulder pain and dysfunction. All were full-time professional male rugby union and rugby league players. They all had failed conservative treatment for their complaint, and the diagnosis was unclear. Arthroscopic findings were used as a measure of main outcome. RESULTS: The primary mechanism of injury was reported as direct tackling (56%; n = 49) followed in succession by falling onto the arm (10%; n = 8). However, in 30% of the cases, no definite injury could be recalled. The main operative finding was that most patients exhibited multiple shoulder pathologies, with 75% of cases presenting with two or more pathologies. A superior labrum anterior to posterior (SLAP) lesion was evident at arthroscopy in 72 of the 87 cases (83%), while rotator cuff tears were evident in 43% of cases (n = 37). One-third of all cases had a Bankart tear (n = 29), despite none of them reporting previous dislocations, while other labral tears, excluding SLAP tears, to the inferior or posterior labrum were present in 34% (n = 30) of the cohort. CONCLUSIONS: Repeated tackling, which is clearly rugby specific, is most likely to be responsible for most of these shoulder injuries, which upon arthroscopic examination, showed signs of mixed pathology. We suggest that an early arthroscopic investigation is valuable in this population in order to confirm treatable diagnosis on the painful shoulder and expedite a safe return to play.

Effect of graft tensioning on mechanical restoration in a rat model of anterior cruciate ligament reconstruction using free tendon graft.

PURPOSE: Initial graft tensioning is important in anterior cruciate ligament reconstruction (ACLR), but its effect on graft healing is still not clear. Since all previous animal studies on graft tensioning used bone-patellar tendon-bone, this study aimed to investigate the effect of initial graft tensioning on ACLR using tendon graft. METHODS: Fifty-five Sprague-Dawley rats underwent ACLR using flexor digitorum longus tendon graft. A constant force of 2 or 4 N was applied during graft fixation. At 0, 2, and 6 weeks, knee samples were harvested (n = 6) for static knee laxity test and graft pull-out test. Histological examination was performed at 2 and 6 weeks post-injury (n = 4). RESULTS: At time zero, knee laxity was restored by ACLR with 2 or 4 N tensioning as compared to ACL-deficient group (p < 0.001), and the 4 N group exhibited a better restoration as compared to 2 N group (p = 0.031). At week 2 post-operation, the 4 N group still exhibited a better restoration in knee laxity (p = 0.001) and knee stiffness (p = 0.002) than the 2 N group; the graft pull-out force (p = 0.032) and stiffness (p = 0.010) were also higher. At week 6 post-operation, there was no significant difference between the 2 and 4 N group in knee laxity and graft pull-out strength. Histological examination showed that the beneficial effect of higher initial graft tension may be contributed by maintenance of graft integrity at mid-substance and reduction in adverse peri-graft bone changes in the femoral tunnel region. CONCLUSIONS: A higher initial graft tension favours the restoration of knee laxity and promotes graft healing in ACLR using free tendon graft in the rat model.

Clinical and arthroscopic findings in recreationally active patients.

OBJECTIVE: To examine the diagnostic accuracy of standard clinical tests for the shoulder in recreational athletes with activity related pain. DESIGN: Cohort study with index test of clinical examination and reference standard of arthroscopy. SETTING: Sports Medicine clinic in Sheffield, U.K. PARTICIPANTS: 101 recreational athletes (82 male, 19 female; mean age 40.8 +/- 14.6 years) over a six year period. INTERVENTIONS: Bilateral evaluation of movements of the shoulder followed by standardized shoulder tests, formulation of clinical diagnosis and shoulder arthroscopy conducted by the same surgeon. MAIN OUTCOME MEASUREMENTS: Sensitivity, specificity, likelihood ratio for a positive test and over-all accuracy of clinical examination was examined retrospectively and compared with arthroscopy. RESULTS: Isolated pathology was rare, most patients (72%) having more than one injury recorded. O'Brien's clinical test had a mediocre sensitivity (64%) and over-all accuracy (54%) for diagnosing SLAP lesions. Hawkins test and Jobe's test had the highest but still not impressive over-all accuracy (67%) and sensitivity (67%) for rotator cuff pathology respectively. External and internal impingement tests showed similar levels of accuracy. When a positive test was observed in one of a combination of shoulder tests used for diagnosing SLAP lesions or rotator cuff disease, sensitivity increased substantially whilst specificity decreased. CONCLUSIONS: The diagnostic accuracy of isolated standard shoulder tests in recreational athletes was over-all very poor, potentially due to the majority of athletes (71%) having concomitant shoulder injuries. Most likely, this means that many of these injuries are missed in general practice and treatment is therefore delayed. Clinical examination of the shoulder should involve a combination of clinical tests in order to identify likely intra articular pathology which may warrant referral to specialist for surgery.

The importance of early arthroscopy in athletes with painful cartilage lesions of the ankle: a prospective study of 61 consecutive cases.

BACKGROUND: Ankle sprains are common in sports and can sometimes result in a persistent pain condition. PURPOSE: Primarily to evaluate clinical symptoms, signs, diagnostics and outcomes of surgery for symptomatic chondral injuries of the talo crural joint in athletes. Secondly, in applicable cases, to evaluate the accuracy of MRI in detecting these injuries. TYPE OF STUDY: Prospective consecutive series. METHODS: Over around 4 years we studied 61 consecutive athletes with symptomatic chondral lesions to the talocrural joint causing persistent exertion ankle pain. RESULTS: 43% were professional full time athletes and 67% were semi-professional, elite or amateur athletes, main sports being soccer (49%) and rugby (14%). The main subjective complaint was exertion ankle pain (93%). Effusion (75%) and joint line tenderness on palpation (92%) were the most common clinical findings. The duration from injury to arthroscopy for 58/61 cases was 7 months (5.7-7.9). 3/61 cases were referred within 3 weeks from injury. There were in total 75 cartilage lesions. Of these, 52 were located on the Talus dome, 17 on the medial malleolus and 6 on the Tibia plafond. Of the Talus dome injuries 18 were anteromedial, 14 anterolateral, 9 posteromedial, 3 posterolateral and 8 affecting mid talus. 50% were grade 4 lesions, 13.3% grade 3, 16.7% grade 2 and 20% grade 1. MRI had been performed pre operatively in 26/61 (39%) and 59% of these had been interpreted as normal. Detection rate of cartilage lesions was only 19%, but subchondral oedema was present in 55%. At clinical follow up average 24 months after surgery (10-48 months), 73% were playing at pre-injury level. The average return to that level of sports after surgery was 16 weeks (3-32 weeks). However 43% still suffered minor symptoms. CONCLUSION: Arthroscopy should be considered early when an athlete presents with exertion ankle pain, effusion and joint line tenderness on palpation after a previous sprain. Conventional MRI is not reliable for detecting isolated cartilage lesions, but the presence of subchondral oedema should raise such suspicion.

Glucocorticoids inhibit tenocyte proliferation and Tendon progenitor cell recruitment.

Corticosteroid injection is commonly used to treat tendon injuries but is often associated with tendon rupture and impaired tendon healing. The effects of dexamethasone on tenocytes have been studied in vitro but only using high concentrations of dexamethasone in monolayer cultures of tenocytes over short periods of time. We have therefore investigated the effects of physiological and pharmacological concentrations of dexamethasone on monolayer cultures of tenocytes over extended time periods. We have also used fibroblastic-colony forming unit cultures to examine the effects of dexamethasone on a progenitor cell population located in tendons. Culturing tenocytes in the presence of dexamethasone for a period of 24 days resulted in a concentration-related decrease in cell number and collagen synthesis as compared to control cultures. This effect was time dependent with cell number in both dexamethasone-treated and control cultures leveling off after 14 days with the control cultures reaching higher cell densities. In contrast in control cultures, collagen accumulation continued to increase until week 4, whereas in the presence of dexamethasone, this tended to level off after 14 days. To study the role of progenitor cell recruitment, the effects of dexamethasone were investigated using the fibroblastic-colony forming unit assay. Treatment with dexamethasone at concentrations of 0.1 nM to 10 microM leads to a progressive reduction in mean colony size as compared to control cultures. Colony number remained constant at concentrations below 10 nM but fell progressively at concentrations above this. In conclusion, dexamethasone reduces both cell number and collagen synthesis in tenocyte cultures in a concentration-dependent manner by both direct effects on tenocyte proliferation and collagen accumulation, and also by modulating the recruitment of tendon progenitor cells.

Medial collateral ligament bursitis in a 12-year-old girl.

A 12-year-old female athlete, training for an international career in pentathlon, was referred to our clinic because of 2 years of recurrent localized swelling and activity-related pain in the medial aspect of her right knee, since falling from a horse and hitting her knees on the ground. She had been examined by a number of doctors over this period and treated with physiotherapy without a proper diagnosis and with no improvement. She could run only very limited distances before excruciating pain and swelling, and she had to abandon participation in competitions. During rest, the swelling decreased but never disappeared entirely. Clinical examination revealed minor effusion and localized tenderness on palpation around the medial joint line. Magnetic resonance imaging showed fluid between the medial capsule and medial collateral ligament, but could not identify any connection to the joint. Arthroscopy revealed a cleavage in the posterior medial joint capsule, superior and close to the medial meniscus, that was connected to the medial collateral ligament bursa. The cleavage was simply expanded, which emptied the bursa, followed by compression bandage and ice for 2 weeks. An immediate positive effect was observed and she could run pain free without swelling within 3 weeks of surgery. Six months after surgery, there has been no recurrence. To our knowledge, this is the first time this injury has been described in a child.

Increased expression of matrix metalloproteinase 1 (MMP1) in 11 patients with patellar tendinosis.

We studied the expression of procollagen type I, matrix metalloproteinase 1 (MMP1) and tissue inhibitor of metalloproteinase 1 (TIMP-1) by immunohistochemistry in human patellar tendinosis tissues and healthy patellar tendons. In situ gelatin zymography was used to detect collagenolytic activities. The productions of MMP1, TIMP1 and gelatinolytic activities were compared in cell cultures from tendinosis samples and controls. Tendinosis tissues and cultures showed an increase in the expression level of MMP1 and a decrease in that of TIMP1, a condition favoring collagen degradation. Gelatinolytic activities in tendinosis tissues and cultures were elevated. Collagenolysis is a striking feature in patellar tendinosis.