Ryo Oda

In vivo assessment of peripheral nerve regeneration by diffusion tensor imaging

To evaluate the sensitivity of diffusion tensor imaging (DTI) in assessing peripheral nerve regeneration in vivo. We assessed the changes in the DTI parameters and histological analyses after nerve injury to examine degeneration and regeneration in the rat sciatic nerves.

Prevalence and Clinical Characteristics of Osteochondritis Dissecans of the Humeral Capitellum Among Adolescent Baseball Players

Background: The prevalence and clinical characteristics of osteochondritis dissecans (OCD) of the humeral capitellum among adolescent baseball players are unknown. Purpose: To determine the OCD prevalence in adolescent competitive baseball players and to investigate the clinical characteristics of these patients. Study Design: Cross-sectional and case-control study; Level of evidence, 3. Methods: A total of 2433 baseball players (mean age, 14.5 ± 1.5 years) belonging to junior high school and high school baseball clubs were enrolled. Players completed a questionnaire, and the elbow of each player’s throwing arm was assessed by ultrasonography. Participants with abnormal results on ultrasonography were further examined through radiographic study. The OCD lesions were classified into stages based on radiographic results, and demographic data were compared between players with and without OCD lesions. Results: Osteochondritis dissecans of the humeral capitellum was found in 82 (3.4%) elbows by ultrasonography. Players with an OCD lesion began playing baseball at an earlier age (P = .016), had a longer duration of competitive play (P = .0013), and had experienced more present (P = .0025) and past (P < .0001) elbow pain compared with players without a lesion. There were no differences between the 2 groups in the position played (P = .26). Sixty-eight patients underwent further radiographic examination for OCD (radiography, computed tomography, magnetic resonance imaging). Of these players, 10 (14.7%) were classified as having stage I OCD (radiolucent stage); 26 (38.2%), stage II (fragmentation stage); 9 (13.2%), stage III (loose body stage); 9 (13.2%), stage IV (residual stage); and 14 (23.5%), stage V (postoperative stage). Conclusion: The prevalence of OCD of the humeral capitellum, including latent cases, was 3.4% among adolescent baseball players. Players with OCD lesions began playing baseball at earlier ages, had played for longer periods, and had experienced more elbow pain. The player’s current baseball position may not be related to the existence of OCD lesions in adolescents.

Endogenous glucocorticoids improve myelination via Schwann cells after peripheral nerve injury: An in vivo study using a crush injury model

Glucocorticoids improve the symptoms of peripheral nerve disorders, such as carpal tunnel syndrome and peripheral neuropathy. The effects of glucocorticoids are mainly anti‐inflammatory, but the mechanisms of their effects in peripheral nerve disorders remain unclear. Schwann cells of the peripheral nerves express glucocorticoid receptors (GR), and glucocorticoids enhance the rate of myelin formation in vitro. Therefore, it is possible that the clinical improvement of peripheral nerve disorders by glucocorticoids is due, at least in part, to the modulation of myelination. In this study, an adrenalectomy (ADX) was performed, and followed by a daily injection of either low dose (1 mg/kg) or high dose (10 mg/kg) corticosterone (CORT). We then simulated a crush injury of the sciatic nerves. A sham ADX operation, followed by a simulated crush injury, was conducted as a control. Immunohistochemistry showed that the nuclei of in vivo Schwann cells expressed GR and that glucocorticoids impacted the GR immunoreactivity of the Schwann cells. The mRNA and protein expression of myelin basic protein was significantly lower in the animals given ADX with vehicle than in the sham operation group. However, the expression was restored in the low‐dose CORT replacement group. Morphological analyses showed that the ADX with vehicle group had a significantly lower myelin thickness than did the low‐dose CORT replacement group and the sham operation group. These results suggest that endogenous glucocorticoids have an important role in myelination through the GR in Schwann cells after an in vivo peripheral nerve injury.

Silencing the expression of connexin 43 decreases inflammation and joint destruction in experimental arthritis.

The objective of the present study was to determine whether the expression of connexin 43 (Cx43) effected on inflammatory conditions in rat fibroblast‐like synoviocytes (FLS) and on rat model of rheumatoid arthritis (RA). The expression of Cx43 in rat FLS stimulated with lipopolysaccharide (LPS) was confirmed by real‐time reverse transcriptase polymerase chain reaction (RT‐PCR). The effects of small‐interfering RNA targeting Cx43 (siCx43) on pro‐inflammatory cytokines and chemokine were assessed by real‐time RT‐PCR and enzyme‐linked immunosorbent assay (ELISA). The therapeutic and side effects of siCx43 in a rat model of collagen‐induced arthritis (CIA) were examined by in vivo electroporation method. LPS markedly enhanced Cx43 gene expression in rat FLS, with transfection of siCx43 suppressing the over‐expression of pro‐inflammatory cytokines and the chemokine. Treatment of CIA rats with siCx43 significantly ameliorated paw swelling, and significantly reduced histological arthritis scores and radiographic scores. In histological appearance of rat ankle joints, siCx43 treatment significantly decreased the number of tartrate‐resistant acid phosphatase (TRAP)‐positive (osteoclast‐like) cells. These findings indicated that siCx43 had anti‐inflammatory effects in rat FLS and efficiently inhibited the development of CIA. Cx43 may play an important role in the pathophysiology of RA, and may be a potential target molecule for novel RA therapies.

Prolonged bioluminescence monitoring in mouse ex vivo bone culture revealed persistent circadian rhythms in articular cartilages and growth plates.

The bone is a metabolically active organ which undergoes repeated remodeling cycles of bone resorption and formation. In this study, we revealed a robust and extremely long-lasting circadian rhythm in ex vivo culture maintained for over six months from the femoral bone of a PERIOD2Luciferase mouse. Furthermore, we also identified robust circadian clocks in flat bones. High- or low-magnification real-time bioluminescence microscopic imaging revealed that the robust circadian rhythms emanated from the articular cartilage and the epiphyseal cartilage within the growth plate of juvenile animals. Stimulation by forskolin or dexamethasone treatment caused type 0 phase resetting, indicating canonical entraining properties of the bone clock. Together, our findings from long-term ex vivo culture revealed that “tissue-autonomous” circadian rhythm in the articular cartilage and the growth plate of femoral bone functions for several months even in an organ culture condition, and provided a useful in vitro assay system investigating the role of the biological clock in bone formation or development.

In vivo evaluation of rabbit sciatic nerve regeneration with diffusion tensor imaging (DTI): correlations with histology and behavior

Diffusion tensor imaging (DTI) is widely used in the study of the central nervous system. DTI represents a potential diagnostic tool for the peripheral nerve. However, more detailed information is needed for application of DTI in the clinical setting. In this study, peripheral degeneration and regeneration were evaluated using DTI-based analyses in a rabbit model. The changes in DTI parameters were compared to histological and functional changes after nerve injury. We used a high magnetic field (7.04T) MRI system. Japanese white male rabbits were used as the model of sciatic nerve crush injury. MR images were obtained before injury and at 2, 4, 6 and 8 weeks post-injury. The DTI parameters of fractional anisotropy (FA), axial diffusivity (λ||), and radial diffusivity (λ⊥) were calculated. Our results showed decreased FA and increased λ⊥ during the degenerative phase after sciatic nerve injury. In contrast, increased FA and decreased λ⊥ were observed during the regenerative phase. FA changes were correlated with axon number and with motor function recovery, assessed with the toe-spreading index. This study clearly demonstrates the validity of applying DTI parameters to the in vivo evaluation of peripheral nerve regeneration. Furthermore, results suggest that DTI can be a potent tool for predicting the extent of functional recovery after peripheral nerve injury.

Long-Term Results of Vascularized Bone Graft for Stage III Kienböck Disease

PURPOSE Vascularized bone grafting (VBG) is one of the therapeutic approaches for treating advanced Kienböck disease; however, few reports on long-term outcomes are available for this technique. The purpose of this study is to evaluate long-term results by following up patients with stage III Kienböck disease for more than 10 years after VBG. METHODS The study included 18 patients with advanced Kienböck disease (Lichtman stage IIIA, n = 10; stage IIIB, n = 8) who received VBG between 1996 and 2001 and were followed up for at least 10 years. Eleven patients received transplantation from the metacarpal base and 7 patients from the distal radius. Radial shortening and capitate shortening were performed in 5 and 2 stage IIIB patients, respectively. RESULTS The mean follow-up period was 12 years, 3 months. Based on the Mayo Modified Wrist Score, clinical results were excellent in 8 patients, good in 7 patients, and fair in 3 patients. The Stahl index and carpal height ratio were not improved in stage IIIA patients who received bone graft alone, whereas significant improvement was observed in stage IIIB patients who received shortening, as well. CONCLUSIONS Vascularized bone grafting for stage III Kienböck disease demonstrated favorable long-term results and is recommended as a surgical treatment. TYPE OF STUDY/LEVEL OF EVIDENCE Therapeutic III.

Lysophospholipid Receptors Are Differentially Expressed in Rat Terminal Schwann Cells, As Revealed by a Single Cell RT-PCR and In Situ Hybridization

Terminal Schwann cells (TSCs) that cover motor neuron terminals, are known to play an important role in maintaining neuromuscular junctions, as well as in the repair process after nerve injury. However, the molecular characteristics of TSCs remain unknown, because of the difficulties in analyzing them due to their paucity. By using our previously reported method of selectively and efficiently collecting TSCs, we have analyzed the difference in expression patterns of lysophospholipid (LPL) receptor genes (LPA1, LPA2, LPA3, S1P1, S1P2, S1P3, S1P4, and S1P5) between TSCs and myelinating Schwann cells (MSCs). LPL, which includes lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P), is the bioactive lipid that induces a myriad of cellular responses through specific members of G-protein coupled receptors for LPA. It turned out that LPA3 was expressed only in TSCs, whereas S1P1 was expressed in TSCs and skeletal muscle, but not in MSCs. Other types of LPL receptor genes, including LPA1, S1P2, S1P3, S1P4, were expressed in both types of Schwann cells. None of the LPL receptor gene family showed MSCs-specific expression.

Knock out of S1P3 receptor signaling attenuates inflammation and fibrosis in bleomycin-induced lung injury mice model.

Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid metabolite involved in many critical cellular processes, including proliferation, migration, and angiogenesis, through interaction with a family of five G protein–coupled receptors (S1P1–5). Some reports have implicated S1P as an important inflammatory mediator of the pathogenesis of airway inflammation, but the role of S1P3 in the pathogenesis of lung diseases is not completely understood. We used S1P3-deficient (knockout (KO)) mice to clarify the role of S1P3 receptor signaling in the pathogenesis of pulmonary inflammation and fibrosis using a bleomycin-induced model of lung injury. On the seventh day after bleomycin administration, S1P3 KO mice exhibited significantly less body weight loss and pulmonary inflammation than wild-type (WT) mice. On the 28th day, there was less pulmonary fibrosis in S1P3 KO mice than in WT mice. S1P3 KO mice demonstrated a 56% reduction in total cell count in bronchoalveolar lavage fluid (BALF) collected on the seventh day compared with WT mice; however, the differential white blood cell profiles were similar. BALF analysis on the seventh day showed that connective tissue growth factor (CTGF) levels were significantly decreased in S1P3 KO mice compared with WT mice, although no differences were observed in monocyte chemotactic protein-1 (MCP-1) or transforming growth factor β1 (TGF-β1) levels. Finally, S1P levels in BALF collected on the 7th day after treatment were not significantly different between WT and S1P3 KO mice. Our results indicate that S1P3 receptor signaling plays an important role in pulmonary inflammation and fibrosis and that this signaling occurs via CTGF expression. This suggests that this pathway might be a therapeutic target for pulmonary fibrosis.

Monocarboxylate Transporter 4, Associated With the Acidification of Synovial Fluid, Is a Novel Therapeutic Target for Inflammatory Arthritis

Synovial fluid pH is decreased in patients with rheumatoid arthritis (RA); however, the underlying mechanisms are unclear. We undertook this study to examine the mechanism by which synovial fluid pH is regulated and to explore the possibility of a therapeutic strategy by manipulating this mechanism.