Exosomes derived from inflammatory myoblasts promote M1 polarization and break the balance of myoblast proliferation/differentiation.

BACKGROUND: Acute muscle injuries are one of the most common injuries in sports. Severely injured muscles are prone to re-injury due to fibrotic scar formation caused by prolonged inflammation. How to regulate inflammation and suppress fibrosis is the focus of promoting muscle healing. Recent studies have found that myoblasts and macrophages play important roles in the inflammatory phase following muscle injury; however, the crosstalk between these two types of cells in the inflammatory environment, particularly the exosome-related mechanisms, had not been well studied. AIM: To evaluate the effects of exosomes from inflammatory C2C12 myoblasts (IF-C2C12-Exos) on macrophage polarization and myoblast proliferation/differentiation. METHODS: A model of inflammation was established in vitro by lipopolysaccharide stimulation of myoblasts. C2C12-Exos were isolated and purified from the supernatant of myoblasts by gradient centrifugation. Multiple methods were used to identify the exosomes. Gradient concentrations of IF-C2C12-Exos were added to normal macrophages and myoblasts. PKH67 fluorescence tracing was used to identify the interaction between exosomes and cells. Microscopic morphology, Giemsa stain, and immunofluorescence were carried out for histological analysis. Additionally, ELISA assays, flow cytometry, and western blot were conducted to analyze molecular changes. Moreover, myogenic proliferation was assessed by the BrdU test, scratch assay, and CCK-8 assay. RESULTS: We found that the PKH-67-marked C2C12-Exos can be endocytosed by both macrophages and myoblasts. IF-C2C12-Exos induced M1 macrophage polarization and suppressed the M2 phenotype in vitro. In addition, these exosomes also stimulated the inflammatory reactions of macrophages. Furthermore, we demonstrated that IF-C2C12-Exos disrupted the balance of myoblast proliferation/differentiation, leading to enhanced proliferation and suppressed fibrogenic/myogenic differentiation. CONCLUSION: IF-C2C12-Exos can induce M1 polarization, resulting in a sustained and aggravated inflammatory environment that impairs myoblast differentiation, and leads to enhanced myogenic proliferation. These results demonstrate why prolonged inflammation occurs after acute muscle injury and provide a new target for the regulation of muscle regeneration.

MRI-based tendon bone healing is related to the clinical functional scores at the first year after anterior cruciate ligament reconstruction with hamstring tendon autograft.

PURPOSE: The correlation between tendon bone healing and clinical functional scores after anterior cruciate ligament reconstruction (ACLR) using four-stranded hamstring tendon autograft has rarely being reported. The purpose of this study was to determine the correlation between magnetic resonance imaging (MRI)-based tendon bone healing and clinical functional scores after ACLR using hamstring tendon. METHODS: Thirty-eight patients with ACLR using four-stranded hamstring tendon autograft were included in this prospective study in the authors' hospital from 2013 to 2014. All patients were performed Tegner, Lysholm, International Knee Documentation Committee (IKDC) subjective scores, KT-1000 and MRI examinations in 3, 6, 12 months after the operation, respectively. According to MRI, the healing degree of tendon bone was divided into five grades, and the healing degree of the tendon at different time points was evaluated. Moreover, the correlations between the clinical scores and tendon bone healing level at 12 months after the operation were determined. RESULTS: The Tegner, Lysholm, and IKDC scores of all patients were gradually improved over time after ACLR, and the degree of tendon bone healing was gradually increased. Moreover, there were significantly positive correlations between the level of tendon bone healing and the clinical functional scores at 12 months after the operation. CONCLUSION: The clinical functional scores and the degree of tendon bone healing were gradually improved over time after ACLR. Moreover, there were significant positive correlations between the level of tendon bone healing and clinical functional scores of knee joint at the first year after the operation. LEVEL OF EVIDENCE: III.

Features of acetabular labral tears on X-ray, magnetic resonance imaging and hip arthroscopy - the observational pilot study.

INTRODUCTION: To analyze the X-ray and magnetic resonance imaging (MRI) features of acetabular labral tears, so as to improve the recognition of the disease. MATERIAL AND METHODS: Imaging features of 19 patients with acetabular labral tears confirmed by arthroscopy were analyzed retrospectively. All cases were examined by X-ray. Fourteen patients underwent MRI examinations. RESULTS: Seventeen of the 19 patients had at least one osseous abnormality consistent with femoroacetabular impingement (FAI) under arthroscopy; among them 7 cases were cam impingement, 3 cases were pincer impingement and 7 cases were mixed impingement. X-ray manifestations: 12 of the 19 patients had FAI; among them 6 cases were cam impingement, 2 cases were pincer impingement and 4 cases were mixed impingement. Five cases showed various degrees of degenerations of hip joint. Two patients had no identifiable structural abnormalities. Magnetic resonance imaging showed various degrees of injury of labrum in 6 patients, joint effusion in 5 cases, and bone marrow edema of femoral head in 3 cases. CONCLUSIONS: The FAI has much to do with acetabular labral tears. X-ray examination is important for FAI diagnosis. Given its high sensitivity and accuracy, MRI is an effective preoperative tool for defining the location and extent of labral tears. Combined with X-ray imaging and clinical manifestation, MRI can provide a reliable basis for clinical diagnosis and treatment of acetabular labral tears.

[Clinical observation of anterior cruciate ligament reconstruction with tibial-remnant preserving technique using hamstring autograft].

OBJECTIVE: To observe the outcome of ACL reconstruction with tibial-remnant preserving technique using hamstring autograft. METHODS: From January 2007 to June 2009,18 patients, including 11 male and 7 female with an average age of 28.2 years (from 18 to 42 years), received ACL reconstruction in Kunshan traditional Chinese medicine hospital, Jiangsu province. All cases were confirmed that the remnant of torn ACL were more than 1/3 by arthroscopic observation. The gracillis and semitendinosus tendons harvested from the patient were folded and weaved as a graft, which was pulled into the femoral and tibia tunnel through the sheath formed by the remnant preserved carefully, then fixed by Rigidfix at femoral side and Intrafix at tibial side respectively. The Lysholm scores were recorded at the last follow-up to evaluate the postoperative knee function. The knee stability, range of motion (ROM), and proprioception were evaluated by physical examination. RESULTS: The average duration of follow-up of all 18 patients was 17.3 months (ranged from 15 to 24 months), no synovitis, graft tear, and ROM limitation were found. The average postoperative Lysholm score was (91.56 +/- 4.60) improved from preoperative (49.43 +/- 2.85) (P < 0.05). There were no differences between bilateral knees on the knee flexion (P > 0.05). CONCLUSION: To preserve the tibial remnant of torn ACL during ACL reconstruction had good outcomes, and contributed to recovery of knee proprioceptive function.

[Effect of exogenous interferon gamma on the healing of injured skeletal muscle following injury].

OBJECTIVE: To observe the influence of combined injection with human interferon (hlFNgamma) and human insulin-link growth factor-1 (hIGF-1) on regeneration and fibrosis of skeletal muscle after acute contusion. METHODS: A standard contusion model was reproduced at the right gastrocnemius in 64 male mice of 7 to 12 weeks. All the mice were randomly divided into 4 groups, such as group A (injection with hIFNgamma), group B (injection with hIGF-1), group C (injection with hIGF-1 and hIFNgamma), and group D (injection with physiological saline as control). All injections were introduced on day 10 after injury at local injured gastrocnemius. Before intervention (7 d following injury), and 4 d, 18 d, 32 d after intervention, the local injured gastrocnemius were harvested from 4 mice of each group. Then the expression of MHC- II b and vimentin was detected by real time PCR and immunohistochemistry techniques. RESULTS: (1) At the each time following intervention, the expression of MHC-II b mRNA and protein in local injured muscle of group B and C were significantly higher than those of group A and D. (2) After intervention,the expression of vimentin mRNA and protein in local injured muscle of group A, group B, and group C were more inhibited than those of group D. The inhibition of vimentin expression in group A and C was significant. CONCLUSION: It is indicated that injection of hIGF-1 into the injured skeletal muscle following acute contusion could enhance muscle regeneration,and inhibit fibrosis to some extent. (2) It is identified that hIFNgamma injected into injured muscle has the effect of anti-fibrosis, which is more significant than that of hIGF-1. (3) Combined injection with hIGF-1 and hIFNgamma could improve muscle regeneration and inhibit fibrosis simultaneously, and promote the healing of injured muscle.

[A study of the bacteriophage-based assay for the detection of pyrazinamide resistance in Mycobacterium tuberculosis].

OBJECTIVE: To set up and evaluate the method of phage amplified biologically assay (PhaB) in rapid detection of pyrazinamide (PZA) resistance. METHODS: The PhaB assay was developed and applied in detecting PZA resistance in 108 clinical isolates of Mycobacterium tuberculosis and the results were compared with those of the absolute concentration method. The minimum inhibitory concentration (MIC) was detected for all discrepancy isolates. RESULTS: The results showed that the optimal detecting condition was pH 5.5, PZA 200 microg/ml and 48 h. Of the 108 strains of Mycobacterium tuberculosis, 28 strains were PZA-susceptible and 80 strains were PZA-resistant detected by PhaB; while 32 strains were PZA-susceptible and 76 strains were PZA-resistant by absolute concentration method. Twenty-eight of the 108 strains were PZA-susceptible and 71 were PZA-resistant by the two methods. The concordant isolates of determination of PZA resistance were 99 by the two methods and the concordance rates was 91.7%. There were 9 strains in discordant isolates, of which 7 were the same with MIC method and gene chip in drug susceptibility. If the results of absolute concentration method was the gold standard, the sensitivity, specificity, positive and negative predictive values, as well as accuracy of PhaB assay was 94.7%, 84.8%, 93.4%, 87.5% and 91.7% respectively. CONCLUSION: The PhaB assay can be used as a rapid screening method for detection of drug susceptibility of PZA in clinical isolates of Mycobacterium tuberculosis.

Structure-activity relationship of natural flavonoids in hydroxyl radical-scavenging effects.

AIM: To study the relationship between the structure and hydroxyl radical (*OH)-scavenging activity of twelve natural flavonoids. METHODS: The hydroxyl radical-generating chemiluminescence system with ascorbate-CuSO4-yeast-H2O2 was used to determine the hydroxyl radical-scavenging activity of twelve natural flavonoids. RESULTS: Guercetin, heliosin, hyperoside, kaempferol, baicalin, corylifolin, lysionotin, matteucinol, corylifolinin, and genistein could effectively scavenge. OH and inhibit the chemiluminescence of the system. The IC50 values (95 % confidence limits) of the flavonoids were 12.1 (9.9-14.5) g/L, 15.8(14.0-19.2) g/L, 19.5 (16.8-27.4) g/L, 20.1 (13.6-29.0) g/L, 34.6 (28.4-43.4) g/L, 66.8 (63.2-74.4) g/L, 187 (147-235) g/L, 211 (165-284) g/L, 262 (190-346) g/L, and 708 (498-994) g/L, respectively; whereas nobilelin and corylifolin-Ac could not scavenge *OH. CONCLUSION: (1) Phenolic hydroxyls in flavonoids were the main active groups capable of scavenging *OH; (2) Hydroxyl groups in ring B and A were important *OH-scavenging active groups; (3) The ortho-dihydroxyl groups in ring A and/or B could greatly enhance the *OH-scavenging activity of the rings; (4) Comparing the IC50 values of guercetin, heliosin, hyperoside, baicalin, lysionotin, and matteucinol, it was suggested that the hydroxyl groups on 3',4' position of ring B possessed high *OH-scavenging activity and the scavenging activity of hydroxyl groups in ring B was higher than that of hydroxyl groups in ring A. The hydroxyl group or glucoside on 3 position of ring C of the above mentioned 6 flavonoids was also related to the. OH-scavenging ability. (5) The structural types of flavonoids themselves could influence their *OH-scavenging activity.