Hanako Nishimoto

Platelet-rich plasma protects rotator cuff-derived cells from the deleterious effects of triamcinolone acetonide.

Triamcinolone acetonide (TA) injections are widely used to treat enthesopathy, but they may induce adverse effects such as tendon impairment and rupture. Platelet‐rich plasma (PRP) is a blood fraction containing high platelet concentrations and various growth factors that play a role in tissue repair processes. The purpose of this study is to investigate whether TA has deleterious effects on human rotator cuff‐derived cells, and if PRP can protect these cells from the effects of TA. Human rotator cuff‐derived cells were cultured with and without TA and PRP, and the culture without any additive served as the control. Cell morphology was assessed at days 7 and 21. Cell viability was evaluated at days 1, 7, 14, and 21 by a water‐soluble tetrazolium salt assay. Induction of apoptosis was measured by immunofluorescence staining and flow cytometry at day 7. Induction of cleaved caspase‐3 was measured by immunofluorescence staining at day 7. The cells cultured with TA had a flattened and polygonal shape at day 7. The cells cultured with both TA and PRP were similar in appearance to control cells. Exposure to TA also significantly decreased cell viability, but cell viability did not decrease when PRP was added along with TA. The number of apoptotic cells increased with TA exposure, while addition of PRP prevented cell apoptosis. In conclusion, the deleterious effect of TA was prevented by PRP, which can be used as a protective agent for patients receiving local TA injections.

Regeneration of rotator cuff tear using electrospun poly(d,l-Lactide-Co-Glycolide) scaffolds in a rabbit model.

PURPOSE The purpose of this study was to evaluate an application of poly(d,l-lactide-co-glycolide) (PLG) scaffold created by electrospinning in a rabbit rotator cuff defect model. METHODS Forty-two Japanese white rabbits were used in this study. Defects of the infraspinatus tendon were created, and the PLG scaffolds were implanted. Contralateral infraspinatus tendons were reattached without creating defects. Histologic analyses were performed 4, 8, and 16 weeks after the operation, and mechanical evaluations were performed 0, 4, 8, and 16 weeks after the operation. RESULTS Scaffold fibers remained without dissolution and spindle-shaped cells were observed inside of the scaffold at 4 weeks postoperatively. At 8 weeks, the PLG scaffold had dissolved and bone formation was observed at the scaffold-bone interface. At 16 weeks, the scaffold-bone interface matured and expression of type II collagen was observed. A statistical difference in ultimate failure load was not seen between the scaffold group and reattachment group or normal tendon after 8 weeks postoperatively. The stiffness in the scaffold group was not significantly different from that in the reattachment group at each time point. However, it was significantly weaker than normal tendon at each time point. CONCLUSIONS Transplantation of cell-free PLG scaffold showed cell migration and type II collagen and proteoglycan expression at the scaffold-bone junction by 16 weeks postoperatively with a sufficient ultimate failure load in a rabbit rotator cuff defect model. CLINICAL RELEVANCE The PLG scaffold could be applied to bridge rotator cuff defects. The results showed that bridging with scaffold can be equivalent to reattachment.

Application of layered poly (L-lactic acid) cell free scaffold in a rabbit rotator cuff defect model

BackgroundThis study evaluated the application of a layered cell free poly (L-lactic acid) (PLLA) scaffold to regenerate an infraspinatus tendon defect in a rabbit model. We hypothesized that PLLA scaffold without cultivated cells would lead to regeneration of tissue with mechanical properties similar to reattached infraspinatus without tendon defects.MethodsLayered PLLA fabric with a smooth surface on one side and a pile-finished surface on the other side was used. Novel form of layered PLLA scaffold was created by superimposing 2 PLLA fabrics. Defects of the infraspinatus tendon were created in 32 rabbits and the PLLA scaffolds were transplanted, four rabbits were used as normal control. Contralateral infraspinatus tendons were reattached to humeral head without scaffold implantation. Histological and mechanical evaluations were performed at 4, 8, and 16 weeks after operation.ResultsAt 4 weeks postoperatively, cell migration was observed in the interstice of the PLLA fibers. Regenerated tissue was directly connected to the bone composed mainly of type III collagen, at 16 weeks postoperatively. The ultimate failure load increased in a time-dependent manner and no statistical difference was seen between normal infraspinatus tendon and scaffold group at 8 and 16 weeks postoperatively. There were no differences between scaffold group and reattach group at each time of point. The stiffness did not improve significantly in both groups.ConclusionsA novel form of layered PLLA scaffold has the potential to induce cell migration into the scaffold and to bridge the tendon defect with mechanical properties similar to reattached infraspinatus tendon model.

A new bioabsorbable cotton-textured synthetic polymer scaffold for osteochondral repair

PurposeWe have previously reported that a cylindrical bioabsorbable synthetic polymer scaffold made of poly (DL-lactide-co-glycolide) (PLG) can be used to repair osteochondral defects without using cultured cells in a rabbit model. This cylindrical scaffold has a solid and pre-formed design, which limits its widespread application. Therefore, we created a cotton-textured PLG scaffold, which would be superior to other scaffolds in terms of plastic property and operability. The purpose of the present study was to examine the efficacy of the cotton-textured PLG scaffold in the repair of osteochondral defects.MethodsCotton-textured PLG scaffolds were prepared using the electrospinning method and used to repair osteochondral defects produced on the right femoral condyle in 36 rabbits. As a control, the defect was left untreated. The outcomes of repair were examined histologically at postoperative weeks four, eight, and 12.ResultsIn the untreated control group, the surface of the defect remained concave and the regenerated cartilaginous tissue partially covered the articular surface even at postoperative week 12. In the scaffold group, cartilaginous tissue covered the surface of the defect at postoperative week four, and the surface was smooth and the cartilaginous tissue was well regenerated and integrated with the native cartilage at postoperative week 12.ConclusionsThe cotton-textured PLG scaffold could repair the osteochondral defect with good outcomes similar to those previously reported for the cylindrical scaffold, with its characteristic advantages of better plasticity and operability. We conclude that the cotton-textured PLG scaffold has potential for clinical application in comminuted osteochondral injury.

Increase in carbon dioxide accelerates the performance of endurance exercise in rats

Endurance exercise generates CO2 via aerobic metabolism; however, its role remains unclear. Exogenous CO2 by transcutaneous delivery promotes muscle fibre-type switching to increase endurance power in skeletal muscles. Here we determined the performance of rats running in activity wheels with/without transcutaneous CO2 exposure to clarify its effect on endurance exercise and recovery from muscle fatigue. Rats were randomised to control, training and CO2 groups. Endurance exercise included activity-wheel running with/without transcutaneous CO2 delivery. Running performance was measured after exercise initiation. We also analysed changes in muscle weight and muscle fibres in the tibialis anterior muscle. Running performance improved over the treatment period in the CO2 group, with a concomitant switch in muscle fibres to slow-type. The mitochondrial DNA content and capillary density in the CO2 group increased. CO2 was beneficial for performance and muscle development during endurance exercise: it may enhance recovery from fatigue and support anabolic metabolism in skeletal muscles.

Association of Young Age with Bone Mineral Density Improvement with Denosumab Treatment in Patients with Osteoporosis

Introduction: Analysis of predictive factors for the improvement of bone mineral density (BMD) in response to osteoporosis treatment is critical. Several studies reported on the analysis of bone turnover markers as a predictive factor for the improvement of BMD; however, few studies reported on predictive factors other than bone turnover markers. Thus, this study aimed to analyze other predictive factors for distal radial BMD improvement in response to denosumab treatment among patients with osteoporosis.Methods: We evaluated 133 patients with osteoporosis over a 24 month period. All patients received denosumab (60 mg) subcutaneously every 6 months. The BMD of the distal radius was assessed in all patients and serum concentrations of PINP and TRACP5b determined.Results: Denosumab treatment resulted in a 3.3% increase, from baseline, in distal radius BMD at 24 months. The average BMD change at 24 months was not significantly changed in relation to gender or prevalent medication. The average BMD change at 24 months was negatively correlated with patient’s age. Almost all the points of PINP and TRACP5b were not correlated with BMD except TRACP5b value at 24 months.Conclusions: We found that improvement of bone mineral density with denosumab treatment for osteoporosis is associated with young age of the patients. We recommend denosumab treatment for younger patients with osteoporosis.

Transcutaneous carbon dioxide application with hydrogel prevents muscle atrophy in a rat sciatic nerve crush model: TRANSCUTANEOUS CO2 PREVENTS MUSCLE ATROPHY

The acceleration of nerve regeneration remains a clinical challenge. We previously demonstrated that transcutaneous CO2 application using a novel hydrogel increases the oxygen concentration in local tissue via an “artificial Bohr effect” with the potential to prevent muscle atrophy. In this study, we investigated the effect of transcutaneous CO2 administration on limb function after peripheral nerve injury in a rat sciatic nerve injury model. In total, 73 Sprague–Dawley rats were divided into a sham group, a control group (crush injury to sciatic nerve and no treatment) or a CO2 group (crush injury with transcutaneous CO2 application). CO2 was administered percutaneously for 20 min five times per week. Scores for the sciatic function index and pinprick test were significantly higher in the CO2 group than control group. The muscle wet weight ratios of the tibialis anterior and soleus muscles were higher in the CO2 group than control group. Electrophysiological examination showed that the CO2 group had higher compound motor action potential amplitudes and shorter distal motor latency than the control group. Histological examination of the soleus muscle sections at postoperative week 2 showed shorter fiber diameter in the control group than in the CO2 group. The mRNA expression of Atrogin‐1 and MuRF‐1 was lower, mRNA expression of VEGF and myogenin and MyoD was higher in CO2 group at postoperative week 2 compared to the control group. Clinical significance: Transcutaneous CO2 application has the therapeutic potential to accelerate the recovery of muscle atrophy in peripheral nerve injury.

AB0075 AICAR Induced Mitochondrial Apoptosis and Inhibited Cell Proliferation and MMP-3/RANKL Secretion via Enhancement of Mitochondrial Biogenesis in Rheumatoid Arthritis Fibroblast-Like Synovial Cells

Background Joint destruction in rheumatoid arthritis (RA) progresses via the hyperproliferation of the synovium and secretion of MMP-3/RANKL from fibroblast-like synoviocytes (FLS). [1] In tumors, we previously reported that the expression of mitochondrial biogenesis-related genes is low and increased numbers of mitochondria enhance cell apoptosis. [2] However, the relationship between mitochondrial biogenesis and cell apoptosis in RA-FLS remains unclear. Objectives This research was investigated the relationship between mitochondrial biogenesis and cell apoptosis in RA-FLS. Methods RA and Osteoarthritis (OA)-FLS were obtained during total knee replacement surgery from patients with RA. All experiments were conducted using cells from passages 3–7. Quantitative PCR was used to assess the expression of mitochondria-related mRNA (PGC-1α, NRF-1, and TFAM) and number of mitochondrial DNA (mtDNA) in RA- and OA-FLS with/without 5-aminoimidazole-4-carboxamide riboside (AICAR) (2 mM). Apoptosis was evaluated immunoblot analysis for cleaved caspase-3, -8, -9, and PARP 48 hours after an AICAR treatment in RA-FLS. Cell viability was assessed using WST-8 assay, and MMP-3/RANKL secretion was measured using an immunoassay (immunoblot and ELISA) with/without IL-1β or TNFα stimulation in RA-FLS. Inhibitory experiments were used AMPK inhibitor (20 μM Compound C) and mitochondrial biogenesis- inhibitor (10 μM TFAM-siRNA) by Lipofectamine® RNAiMAX Reagent in RA-FLS. Results The expression of NRF-1 and TFAM and the levels of mtDNA were lower in RA-FLS than in OA-FLS. In RA-FLS, the levels of mtDNA and mRNA expression of mitochondria-related genes were enhanced by AICAR. Additionally, AICAR enhanced mitochondrial apoptosis (Increased the protein of cleaved caspase-3, -9, and PARP), and Treatment with an AMPK inhibitor (compound C) and a mitochondrial biogenesis- inhibitor (TFAM-siRNA) suppressed the inhibitory effect of AICAR on apoptosis. Moreover, AICAR inhibited cell viability and IL-1β- or TNFα-induced MMP-3/RANKL secretion in inflammation-induced RA-FLS. Conclusions Mitochondrial biogenesis in RA-FLS was down-regulation. Enhancement of mitochondrial biogenesis resulted in the suppression of disease activities of RA, such as increasing mitochondrial apoptosis and reducing RA-FLS viability and secretion of MMP-3/RANKL from RA-FLS. A novel therapeutic approach by changing mitochondrial biogenesis is proposed. References Bottini N, Firestein GS, Duality of fibroblast-like synoviocytes in RA: passive responders and imprinted aggressors. Nat Rev Rheumatol. 2013 Onishi Y, Ueha T, Kawamoto T, et al, Regulation of mitochondrial proliferation by PGC-1α induces cellular apoptosis in musculoskeletal malignancies. Sci Rep. 2014 Acknowledgement The authors wish to express sincere thanks to Ms. Kyoko Tanaka, Ms. Maya Yasuda, and Ms. Minako Nagata (Department ofOrthopaedic Surgery, Kobe University Graduate School of Medicine) for their technical assistance. Disclosure of Interest None declared