Publications by authors named "Shigeru Miyaki"

48 Publications

Pharmacological Targeting of Heme Oxygenase-1 in Osteoarthritis.

Antioxidants (Basel) 2021 Mar 9;10(3). Epub 2021 Mar 9.

Medical Center for Translational and Clinical Research, Hiroshima University Hospital, Hiroshima 7348551, Japan.

Osteoarthritis (OA) is a common aging-associated disease that clinically manifests as joint pain, mobility limitations, and compromised quality of life. Today, OA treatment is limited to pain management and joint arthroplasty at the later stages of disease progression. OA pathogenesis is predominantly mediated by oxidative damage to joint cartilage extracellular matrix and local cells such as chondrocytes, osteoclasts, osteoblasts, and synovial fibroblasts. Under normal conditions, cells prevent the accumulation of reactive oxygen species (ROS) under oxidatively stressful conditions through their adaptive cytoprotective mechanisms. Heme oxygenase-1 (HO-1) is an iron-dependent cytoprotective enzyme that functions as the inducible form of HO. HO-1 and its metabolites carbon monoxide and biliverdin contribute towards the maintenance of redox homeostasis. HO-1 expression is primarily regulated at the transcriptional level through transcriptional factor nuclear factor erythroid 2 (NF-E2)-related factor 2 (Nrf2), specificity protein 1 (Sp1), transcriptional repressor BTB-and-CNC homology 1 (Bach1), and epigenetic regulation. Several studies report that HO-1 expression can be regulated using various antioxidative factors and chemical compounds, suggesting therapeutic implications in OA pathogenesis as well as in the wider context of joint disease. Here, we review the protective role of HO-1 in OA with a focus on the regulatory mechanisms that mediate HO-1 activity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/antiox10030419DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001640PMC
March 2021

Histological scoring system for subchondral bone changes in murine models of joint aging and osteoarthritis.

Sci Rep 2020 06 22;10(1):10077. Epub 2020 Jun 22.

Department of Molecular Medicine, Scripps Research, La Jolla, California, USA.

To establish a histopathological scoring system for changes in subchondral bone in murine models of knee osteoarthritis (OA), three key parameters, subchondral bone plate (Subcho.BP) consisting of the combination of Subcho.BP.thickness (Subcho.BP.Th) and angiogenesis, bone volume (BV/TV) and osteophytes, were selected. The new grading system was tested in two mouse OA models, (1) senescence accelerated mouse (SAM)-prone 8 (SAMP8) as spontaneous OA model with SAM-resistant 1 (SAMR1) as control; (2) destabilization of the medial meniscus in C57BL/6 mice as surgical OA model. Results of the spontaneous OA model showed that Subcho.BP.Th was significantly wider, angiogenesis was greater, and BV/TV was higher in SAMP8 than SAMR1. Notably, subchondral bone score was dramatically higher in SAMP8 at 6 weeks than SAMR1, while OARSI cartilage scores became higher only at 14 weeks. In the surgical OA model, the results were similar to the spontaneous OA model, but osteophytes appeared earlier. There were strong correlations both in Subcho.BP.Th and BV/TV between this scoring system and µCT (r = 0.89, 0.84, respectively). Inter-rater reliabilities for each parameter using this system were more than 0.943. We conclude that this new histopathological scoring system is readily applicable for evaluating the early changes in aging and OA-affected murine subchondral bone.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-66979-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7308327PMC
June 2020

Role of vasoactive intestinal peptide in the progression of osteoarthritis through bone sclerosis and angiogenesis in subchondral bone.

J Orthop Sci 2020 Sep 9;25(5):897-906. Epub 2020 Jan 9.

Department of Orthopaedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3, Kasumi, Minami-ku, Hiroshima, 734-8551, Japan.

Objective: Osteoarthritis (OA) is a progressive joint disorder, with abnormal remodeling of subchondral bone linked to the disruption of cartilage metabolism. Nerves also play an important role in bone remodeling in OA progression, and vasoactive intestinal peptide (VIP), one of the neuropeptides, plays an important role in bone metabolism. The aim of this study was to analyze the expression pattern of VIP in subchondral bone, and its potential as a therapeutic target for OA progression.

Design: The pattern of VIP expression in the human tibia was histologically evaluated. The effect of VIP on angiogenesis was investigated using human umbilical vein endothelial cells (HUVECs). Knee OA was induced by the resection of the medial meniscotibial ligament in C57BL/6 mice. A VIP receptor antagonist was intraperitoneally administered postoperatively, and therapeutic effects were analyzed at 4 and 8 weeks.

Results: VIP expression in the subchondral bone increased as OA progressed in human tibia. VIP was also expressed in the vascular channels into the cartilage layer. The total length and branch points were significantly increased, due to the VIP receptor agonist in HUVECs. In OA mice, the VIP receptor antagonist could prevent cartilage degeneration and subchondral bone sclerosis. The Osteoarthritis Research Society International score in the VIP receptor antagonist group was significantly lower than in the control group.

Conclusion: VIP is involved in the progression of OA through its effect on subchondral bone sclerosis and angiogenesis. Inhibition of VIP signaling has the potential to be a therapeutic target to prevent OA progression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jos.2019.11.010DOI Listing
September 2020

A Novel Mucidosphaerium sp. Downregulates Inflammatory Gene Expression in Skin and Articular Cells.

Altern Ther Health Med 2021 Jan;27(1):28-34

Context: Hot-spring therapy is occasionally used for the treatment of inflammatory diseases. Microorganisms might contribute to the anti-inflammatory functions seen in thermal mud therapies. Natural microorganisms, derived from traditional spa resorts, could be useful as a preventive strategy for alternative medical applications.

Objective: The aim of the study was to find effective microalgae from prominent hot springs to use for the treatment of inflammatory diseases.

Design: The research team performed an in-vitro study. Microalgae, derived from Beppu hot springs, were isolated and homogeneously cultured.

Setting: The study took place at the Saravio Central Institute at Saravio Cosmetics in Oita, Japan and the Department of Bioscience and Biotechnology in the Graduate School of Agriculture at Shinshu University in Nagano, Japan.

Intervention: For identification, the 18S ribosomal RNA genes of microalgae were investigated by DNA sequencing and homology search, together with microscopic observation.

Outcome Measures: To examine the pharmacological activities of the algal extracts, real-time polymerase chain reactions were performed, using either primary dermal fibroblasts (DFs), dermal papilla cells (DPCs), or fibroblast-like synoviocytes (FLSs). To test the antioxidant activity, both the oxygen radical absorbance capacity and the generation of intracellular reactive oxygen species (ROS) were evaluated.

Results: A novel strain of green algae, Mucidosphaerium sp., was isolated from a Beppu hot spring. The algal extract downregulated gene-expression levels of pro-inflammatory cytokines, such as interleukin-1β (IL-1β), IL-6, and tumor necrosis factor- alpha (TNF-α), in various primary cells pre-exposed to IL-1β. The protein level of the risk factors was concomitantly reduced. In addition, the algal extract suppressed the IL-1β-induced upregulation of cyclooxygenase-2, nerve growth factor, and matrix metalloproteinase-1 (MMP-1) and MMP-3 in DFs. It also inhibited that of MMP-1, -3, and -9 in FLSs. Moreover, the extract inhibited total MMP protease activities. The microalgae decreased the intracellular reactive oxygen species (ROS) level in FLSs with an antioxidant activity of 178.3 ± 0.9 μmol of trolox equivalent/g.

Conclusions: The present study showed that the novel Mucidosphaerium sp., derived from a Beppu hot spring, suppressed inflammatory reactions in both cutaneous and articular cells, partly due to its antioxidative properties. The novel algal strain may be a useful tool as an alternative medicine for skin and joint inflammatory disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
January 2021

Repair of an Osteochondral Defect With Minced Cartilage Embedded in Atelocollagen Gel: A Rabbit Model.

Am J Sports Med 2019 07 17;47(9):2216-2224. Epub 2019 Jun 17.

Department of Orthopaedic Surgery, Graduate School of Biomedical & Health Sciences, Hiroshima University, Kasumi, Minami-ku, Hirosima-shi, Hiroshima, Japan.

Background: Autologous chondrocyte implantation (ACI) is often performed for large cartilage defects. Because this technique has several disadvantages, including the need for second-stage surgery, cartilage repair using minced cartilage has been suggested. However, this technique could be improved using 3-dimensional scaffolds.

Purpose: To examine the ability of chondrocyte migration and proliferation from minced cartilage in atelocollagen gel in vitro and evaluate the repairable potential of minced cartilage embedded in atelocollagen gel covered with a periosteal flap in a rabbit model.

Study Design: Controlled laboratory study.

Methods: Minced cartilage or isolated chondrocytes from rabbits were embedded in atelocollagen gel and cultured for 3 weeks. Chondrocyte proliferation and matrix production were evaluated in vitro. An osteochondral defect at the trochlear groove was created in 56 rabbits, which were divided into 4 groups. The defect was left empty (defect group), filled with allogenic minced cartilage (minced cartilage group), filled with isolated allogenic chondrocytes embedded in atelocollagen gel (ACI group), or filled with atelocollagen gel (atelocollagen with periosteal flap group). At 4, 12, and 24 weeks after surgery, repair of the defect was evaluated in these 4 groups.

Results: In vitro, the number of chondrocytes and abundant matrix on the surface of the gel significantly increased in the minced cartilage group compared with the ACI group ( < .05). In vivo, the minced cartilage and ACI groups showed good cartilage repair compared with the empty defect and atelocollagen/periosteal flap groups ( < .05); there was no significant difference in the Pineda score between the minced cartilage and ACI groups.

Conclusion: Minced cartilage in atelocollagen gel had good chondrocyte migration and proliferation abilities in vitro, and osteochondral defects were well repaired by implanting minced cartilage embedded in the atelocollagen gel in vivo. Implantation of minced cartilage embedded in atelocollagen gel showed good cartilage repair equivalent to ACI.

Clinical Relevance: Implantation of minced cartilage embedded in atelocollagen gel as a 1-step procedure has outcomes similar to those of ACI but is cheaper and more convenient than ACI.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/0363546519854372DOI Listing
July 2019

The Benefit of Minced Cartilage Over Isolated Chondrocytes in Atelocollagen Gel on Chondrocyte Proliferation and Migration.

Cartilage 2021 Jan 12;12(1):93-101. Epub 2018 Oct 12.

Department of Orthopaedic Surgery, Graduate School of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan.

Objective: Autologous chondrocyte implantation is a necessary procedure for the repair of articular cartilage defects; however, isolated chondrocyte implantation requires a 2-step procedure (for harvesting and implantation) and is limited by cytotoxicity due to enzymatic digestion. Therefore, in this study, we evaluated the possible benefit of using minced cartilage embedded in a 3-dimensional culture scaffold and fixed with fibrin glue, in comparison with isolated chondrocytes in atelocollagen, to induce cell migration, proliferation, and matrix production, using cartilage from patients with knee joint osteoarthritis.

Design: Cartilage fragments were obtained from 7 female patients with knee osteoarthritis (OA) and embedded in atelocollagen gels. As a control, chondrocytes were isolated and embedded in gels in the same manner. These composites were cultured for 3 weeks, and cell proliferation and matrix production were evaluated using histology and immunochemistry.

Results: Histologically, minced cartilage showed cell migration from the cartilage fragments into the gel, with the Bern score and cell count in the minced cartilage group being significantly higher than those in the control group. Immunohistochemistry revealed that the number of Ki67-positive cells, the expression of LECT-1 and TGF-β, and the glycosaminoglycan content were significantly higher in the minced cartilage than in the control group. Minced cartilage exhibited superior cell migration, proliferation, and glycosaminoglycan content than isolated chondrocytes.

Conclusion: Our findings support that minced cartilage has a favorable potential for cell proliferation and matrix production compared with the isolated chondrocytes after enzymatic treatment.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/1947603518805205DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7755964PMC
January 2021

Histological Evaluation of Early-Phase Changes in the Osteochondral Unit After Microfracture in a Full-Thickness Cartilage Defect Rat Model.

Am J Sports Med 2018 10 1;46(12):3032-3039. Epub 2018 Aug 1.

Department of Orthopaedic Surgery, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

Background: The microfracture (MF) technique is an established surgical treatment for cartilage injury. However, the early-phase histological changes in full-thickness cartilage defects (FTCDs) after MF and the concomitant changes in the subchondral bone are still unknown.

Purpose: To determine the early-phase histological changes in FTCD associated with subchondral bone remodeling after MF in rat model.

Study Design: Controlled laboratory study.

Methods: Rats were subjected to FTCD, followed by MF at the trochlear groove. For histological analysis, experiment and untreated control rats were sacrificed at 0, 1, 3, 7, 14, 28, and 56 days postoperatively (n = 6 knees per time point). Cartilage healing response was evaluated with the Pineda score. Osteoclast activity was evaluated by counting and locating the number of tartrate-resistant acid phosphatase (TRAP)-positive cells in the subchondral bone. MF hole diameter and bone mineral density in the subchondral bone were measured sequentially in 3 rats (n = 6 knees) by 3-dimensional μ-computed tomography.

Results: Pineda score showed no difference in cartilage response from day 0 to day 3 but a significant improvement from day 7 to day 56. Although the MF holes were filled with blood clots in all specimens, the defect sites were not. The number of TRAP-positive cells peaked at day 3, mostly accumulating around the deeper zone of the MF holes. Over time, the number of TRAP-positive cells decreased to preoperative levels, localizing around the aperture of the MF holes where there was active remodeling of the osteochondral unit. The MF hole diameter was largest at day 14, and most holes disappeared by day 28. Bone mineral density was also highest at day 14 and decreased to preoperative levels by day 56.

Conclusion: Histological changes in the FTCD after MF were derived from endochondral ossification within the deeper zone of the MF holes.

Clinical Relevance: The absence of healing by blood clot in the FTCD should be noted by surgeons performing MF, and indications for MF should be carefully considered not only for maximizing the postoperative clinical outcome but also minimizing potential complications, such as formation of subchondral bone cysts or intralesional osteophytes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/0363546518787287DOI Listing
October 2018

IL-1β Enhances Wnt Signal by Inhibiting DKK1.

Inflammation 2018 Oct;41(5):1945-1954

Department of Clinical Immunology and Rheumatology, Hiroshima University Hospital, 1-2-3 Kasumi, Minami, Hiroshima, 734-8551, Japan.

Aberrant endochondral bone formation in the physis is a unique bone lesion in neonatal-onset multisystem inflammatory disease (NOMID), also called chronic infantile neurologic cutaneous articular (CINCA), the most severe of the cryopyrin-associated periodic syndrome (CAPS) diseases, which are interleukin-1β (IL-1β)-related monogenic autoinflammatory diseases. The wingless (Wnt) pathway plays an important role in osteoblast differentiation. In this study, we explored the potential role of IL-1β on the expression of WNT genes and the Wnt antagonist Dickkopf-1 (DKK1). The expression of WNT and DKK1 in fibroblast-like synoviocytes (FLS), which are articular resident cells, was quantified by quantitative PCR and enzyme-linked immunosorbent assay. Additionally, we used T cell factor (TCF) reporter assays to evaluate the activity of the canonical Wnt signal pathway in the presence or absence of the supernatant of cultured FLS treated with or without IL-1β and IL-6. Anti-DKK1 antibodies were used to neutralize DKK1. The expression of both canonical and non-canonical WNT genes as well as DKK1 was observed in FLS. The supernatant of cultured FLS suppressed the luciferase activity of the TCF reporter, and this effect was reduced by its pre-treatment with an anti-DKK1 antibody. Both IL-1β and IL-6 significantly reduced DKK1 production. Furthermore, the supernatant of FLS cultured with IL-1β or IL-6 showed a reduced inhibitory effect on Wnt signaling, compared with the supernatant of untreated FLS. These data suggest that IL-1β, like IL-6, dampens DKK1 production, and thereby promotes Wnt signal activation. Therefore, increased levels of IL-1β may contribute to the dysregulation of endochondral ossification in NOMID/CINCA.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10753-018-0838-zDOI Listing
October 2018

[Cartilage/chondrocyte research and osteoarthritis. The role of microRNAs and extracellular vesicles in osteoarthritis pathogenesis.]

Authors:
Shigeru Miyaki

Clin Calcium 2018;28(6):783-788

Medical Center for Translational and Clinical Research, Hiroshima University Hospital, Japan.

Extracellular vesicles(EV)have been emerged as carrier for the exchange of microRNAs and bioactive factors within cartilage and between joint tissues to promote joint homeostasis and osteoarthritis(OA)pathogenesis. The purpose of this review is to describe how microRNAs and EV regulate functions of chondrocytes in OA pathogenesis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/CliCa1806783788DOI Listing
June 2019

Carnosic acid attenuates cartilage degeneration through induction of heme oxygenase-1 in human articular chondrocytes.

Eur J Pharmacol 2018 Jul 17;830:1-8. Epub 2018 Apr 17.

Department of Orthopaedic Surgery, Hiroshima University, 1-2-3 Kasumi Minami-ku, Hiroshima 734-8551, Japan.

Osteoarthritis (OA) is common age-associated disease, and associated with joint pain, mobility limitations and compromised overall quality of life. OA treatment is currently limited to pain management and joint arthroplasty at end stage disease. Oxidative damage to cartilage extracellular matrix and cells is an important mechanism in joint aging and OA pathogenesis. Evidence from in vitro and in vivo models of OA suggests that pharmaceuticals and natural compounds with antioxidant properties reduce expression of mediators of OA pathogenesis and OA severity in animal models. Among the signaling pathways that control cellular protective mechanisms against oxygen radical damage is heme oxygenase-1 (HO-1). We recently report HO-1 reduced OA severity in a mouse model. This led to the hypothesis that compounds that increase HO-1 expression have therapeutic potential in OA. Carnosic acid (CA), a natural diterpene with oxidant activity, is prevents cartilage degeneration though induction of HO-1. CA induced HO-1 and miR-140 expression in human articular chondrocytes, and cartilage degeneration was attenuated by CA treatment. Induced HO-1 by CA was in part associated with downregulation via miR-140 binding to 3'UTR of BTB and CNC homology 1 (BACH1). These findings suggest that CA attenuates cartilage degradation through HO-1 upregulation and has potential as a supplement for OA prevention.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejphar.2018.04.018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6240351PMC
July 2018

The delaying effect of alpha-glycerophosphocholine on senescence, transthyretin deposition, and osteoarthritis in senescence-accelerated mouse prone 8 mice.

Biosci Biotechnol Biochem 2018 Apr 1;82(4):647-653. Epub 2017 Dec 1.

e National Research Institute of Brewing , Higashi-Hiroshima , Japan.

Administration of alpha-glycerophosphocholine (GPC), a choline compound in food, is expected to contribute to human health. In this study, we evaluated its effect on aging in senescence-accelerated mouse prone 8 (SAMP8) mice. Male SAMP8 mice had free access to a commercial stock diet and drinking water with or without GPC (0.07 mg/ml). Mice in the GPC group had significantly lower total senescence grading score than that of the control group at 36 weeks of age. Administration of GPC decreased the deposition of transthyretin (TTR), an amyloidogenic protein, in the brain. Aggregated TTR activated microglia and led to neuroinflammation. Thus, GPC would protect the brain by reducing TTR deposition and preventing neuroinflammation. In a histological study of knee joints, it was found that SAMP8 mice administered GPC showed decreased joint degeneration. These results suggest that GPC delays the aging process and may be a useful compound in anti-aging functional food development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/09168451.2017.1403883DOI Listing
April 2018

Extracellular vesicles in cartilage homeostasis and osteoarthritis.

Curr Opin Rheumatol 2018 Jan;30(1):129-135

Department of Molecular Medicine, The Scripps Research Institute, La Jolla, California, USA.

Purpose Of Review: Extracellular vesicles carry bioactive molecules that can be transferred between cells and tissues. The purpose of this review is to describe how extracellular vesicles regulate functions of cells in cartilage and other joint tissues. The potential application of extracellular vesicles in the treatment of osteoarthritis and as biomarkers will also be discussed.

Recent Findings: Extracellular vesicles are found in synovial fluid, in articular cartilage and in the supernatants of synoviocytes and chondrocytes. Extracellular vesicles in cartilage have been proposed to be involved in cross talk between cells in joint tissues and to affect extracellular matrix turnover and inflammation. Extracellular vesicles from arthritic joints can promote abnormal gene expression and changes in cartilage extracellular matrix, including abnormal mineralization. Promising results were obtained in the therapeutic application of mesenchymal stem cell-derived extracellular vesicles for cartilage repair and experimental osteoarthritis.

Summary: Extracellular vesicles have emerged as vehicles for the exchange of bioactive signaling molecules within cartilage and between joint tissues to promote joint homeostasis and arthritis pathogenesis. As the molecular content of extracellular vesicles can be customized, they offer utility in therapeutic applications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/BOR.0000000000000454DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5810945PMC
January 2018

The role of tetraspanin CD9 in osteoarthritis using three different mouse models.

Biomed Res 2016 ;37(5):283-291

Department of Orthopaedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University.

Although osteoarthritis (OA) is the most prevalent aging-related joint disease, the understanding of mechanisms of OA pathogenesis remains limited. Key features include the progressive degradation of articular cartilage, synovial hyperplasia, and angiogenesis in joint tissues. CD9, a member of the tetraspanin family, is localized in the cell membranes and partly in the endosomes of all mammalian cell types. CD9 is associated with inflammation and angiogenesis through cell adhesion, migration, and signal transduction. This study examined the role of CD9 in OA development in three different mouse models: an aging model, a surgical model and antigen-induced arthritis (AIA) model, using CD9 deficient mice. Our study showed that CD9 deficiency reduced the severity of hallmarks of OA including cartilage degradation and soft tissue inflammation in aged mice. In the AIA model, cartilage damage and inflammation were also reduced in CD9 mice. This was in contrast to the surgical OA model where disease severity was similar in wild-type and CD9 mice. Col2a1 and Aggrecan expression was increased in chondrocytes of CD9 mice compared with those of wild-type mice. Our results indicate that the suppression of cartilage degradation in CD9 could be in part related to an increase in the expression of the two main cartilage extracellular matrix proteins aggrecan and type II collagen.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2220/biomedres.37.283DOI Listing
January 2017

Inhibition of microRNA-222 expression accelerates bone healing with enhancement of osteogenesis, chondrogenesis, and angiogenesis in a rat refractory fracture model.

J Orthop Sci 2016 Nov 18;21(6):852-858. Epub 2016 Aug 18.

Department of Orthopedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima, Japan.

Background: It is difficult to achieve bone union in case of non-union with non-invasive techniques. MicroRNAs (miRNAs) are short, non-coding RNAs that act as repressors of gene expression at the level of post-transcriptional regulation. This study focuses on microRNA (miR)-222 as it is known to be a negative modulator of angiogenesis, an essential component of fracture healing. The purpose of this study was to analyze the effects of miR-222 on osteogenic and chondrogenic differentiation in human mesenchymal stromal cell (MSC)s in vitro, and to determine whether local administration of miR-222 inhibitor into the fracture site could achieve bone union in vivo.

Method: miR-222 expression in human bone marrow mesenchymal stem cells (hMSCs), and osteogenic differentiation in hMSCs, were investigated. The gain or loss of miR-222 function was examined, in order to assess the effects of miR-222 on osteogenic and chondrogenic differentiation in hMSCs. A femoral transverse fracture was completed in rats, and the periosteum at the fracture site was cauterized. Then, either an miR-222 inhibitor or an miR-222 mimics, mixed with atelocollagen, was administered into the fracture site. A non-functional inhibitor negative control was administered to the control group. At 2, 4, 6, and 8 weeks, radiographs of the fractured femurs were obtained. Immunohistochemistry was performed at 2 weeks to evaluate the capillary density. At 8 weeks, micro-computed tomography (μCT) imaging analysis and histological evaluations were performed.

Results: The expression of miR-222 significantly decreased as osteogenic differentiation of hMSCs proceeded. Inhibition of miR-222 promoted osteogenic differentiation, and over expression of miR-222 inhibited osteogenic differentiation in hMSCs, which was confirmed by measuring expression of Runx2, collagen type 1A1 (COL1A1), and osteocalcin. Inhibition of miR-222 promoted chondrogenic differentiation in hMSCs, which was confirmed by measuring expression of collagen type II (COL2A1), aggrican, and SOX9. Bone union at the fracture site was achieved in only the groups treated with the miR-222 inhibitor, confirmed by radiographic, μCT and histological evaluation at 8 weeks after administration. Immunohistochemistry showed that capillary density in the miR-222 inhibitor group was significantly higher than that in the control group and in the miR-222 mimics group.

Conclusion: Local administration of miR-222 inhibitor can accelerate bone healing by enhancing osteogenesis, chondrogenesis, and angiogenesis in the rat refractory model.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jos.2016.07.021DOI Listing
November 2016

Mesenchymal Stem Cell-Derived Exosomes Promote Fracture Healing in a Mouse Model.

Stem Cells Transl Med 2016 12 26;5(12):1620-1630. Epub 2016 Jul 26.

Department of Orthopaedics Surgery, Integrated Health Sciences, Institute of Biomedical and Health Science, Hiroshima University, Hiroshima, Japan.

: Paracrine signaling by bone-marrow-derived mesenchymal stem cells (MSCs) plays a major role in tissue repair. Although the production of regulatory cytokines by MSC transplantation is a critical modulator of tissue regeneration, we focused on exosomes, which are extracellular vesicles that contain proteins and nucleic acids, as a novel additional modulator of cell-to-cell communication and tissue regeneration. To address this, we used radiologic imaging, histological examination, and immunohistochemical analysis to evaluate the role of exosomes isolated from MSC-conditioned medium (CM) in the healing process in a femur fracture model of CD9 mice, a strain that is known to produce reduced levels of exosomes. We found that the bone union rate in CD9 mice was significantly lower than wild-type mice because of the retardation of callus formation. The retardation of fracture healing in CD9 mice was rescued by the injection of exosomes, but this was not the case after the injection of exosomes-free conditioned medium (CM-Exo). The levels of the bone repair-related cytokines, monocyte chemotactic protein-1 (MCP-1), MCP-3, and stromal cell-derived factor-1 in exosomes were low compared with levels in CM and CM-Exo, suggesting that bone repair may be in part mediated by other exosome components, such as microRNAs. These results suggest that exosomes in CM facilitate the acceleration of fracture healing, and we conclude that exosomes are a novel factor of MSC paracrine signaling with an important role in the tissue repair process.

Significance: This work focuses on exosomes, which are extracellular vesicles, as a novel additional modulator of cell-to-cell communication. This study evaluated the role of exosomes isolated from mesenchymal stem cell (MSC)-conditioned medium (MSC-CM) in the fracture-healing process of CD9 mice, a strain that is known to produce reduced levels of exosomes. Retardation of fracture healing in CD9 mice was rescued by the injection of MSC exosomes, but this was not the case after the injection of exosome-free CM. This study finds that MSC exosomes are a novel factor of MSC paracrine signaling, with an important role in the tissue repair process.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.5966/sctm.2015-0285DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5189643PMC
December 2016

Carnosic acid protects starvation-induced SH-SY5Y cell death through Erk1/2 and Akt pathways, autophagy, and FoxO3a.

Int J Food Sci Nutr 2016 Dec 19;67(8):977-82. Epub 2016 Jul 19.

a Department of Human Life Science Education, Graduate School of Education , Hiroshima University , Hiroshima , Japan ;

Carnosic acid (CA) is recognized as a unique neuroprotective compound in the herb rosemary, since it induces expression of antioxidant enzymes including heme oxygenase-1 (HO-1), γ-glutamylcysteine synthase (γ-GCS), and glutathione S-transferase (GST) via activation of nuclear factor erythroid 2-related factor 2 (Nrf2), which is a nuclear transcription factor. In this study, we examined the cytoprotective effects of CA against starvation. We found that CA protected starvation-induced SH-SY5Y cell death by activating Akt and extracellular signal-regulated kinase 1/2 (Erk1/2). Interestingly, CA induced moderate autophagy and dephosphorylation of a transcriptional factor, the forkhead box protein O3a (FoxO3a). These effects of CA play an important role in cytoprotection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/09637486.2016.1208734DOI Listing
December 2016

Attenuation of cartilage degeneration by calcitonin gene-related paptide receptor antagonist via inhibition of subchondral bone sclerosis in osteoarthritis mice.

J Orthop Res 2016 07 12;34(7):1177-84. Epub 2016 Jan 12.

Department of Orthopaedic Surgery, Integrated Health Sciences, Institute of Biomedical and Health Science, Hiroshima University, Hiroshima, Japan.

Osteoarthritis (OA) is a progressive joint disorder which affects cartilage and subchondral bone. Calcitonin gene-related peptide (CGRP) plays a role in bone metabolism. The purpose of this study is to examine the therapeutic effect of the blocking CGRP on OA progression in mice by inhibition of subchondral bone sclerosis. OA was induced by the resection of the medial meniscotibial ligament of the knee in C57/BL6 mice. An intraperitoneal injection of the CGRP receptor antagonist (BIBN4096) was administered after OA surgery. At 1, 4, and 8 weeks after injection, histological analysis were performed. In vitro, the effect of CGRP and BIBN4096 on osteogenesis and osteoclastogenesis was analyzed. BIBN4096 could prevent cartilage degeneration and subchondral bone sclerosis. The OARSI score in the BIBN4096 group was significantly lower than that in the control. In vitro, CGRP up regulated osteocalcin expression, but its expression was down regulated by BIBN4096. CGRP inhibited osteoclastogenesis of raw 267.4 cells, but its effect was reduced by the addition of BIBN4096.The current study showed that subchondral bone sclerosis and increasing expression of CGRP occurs in the early phase of OA in relation to cartilage degeneration, and that BIBN4096 could effectively attenuate OA progression. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1177-1184, 2016.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jor.23132DOI Listing
July 2016

Bach1 deficiency reduces severity of osteoarthritis through upregulation of heme oxygenase-1.

Arthritis Res Ther 2015 Oct 13;17:285. Epub 2015 Oct 13.

Department of Orthopedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3 Kasumi, Hiroshima, 734-8551, Japan.

Introduction: BTB and CNC homology 1 (Bach1) is a transcriptional repressor of Heme oxygenase-1 (HO-1), which is cytoprotective through its antioxidant effects. The objective of this study was to define the role of Bach1 in cartilage homeostasis and osteoarthritis (OA) development using in vitro models and Bach1 (-/-) mice.

Methods: HO-1 expression in Bach1 (-/-) mice was analyzed by real-time PCR, immunohistochemistry and immunoblotting. Knee joints from Bach1 (-/-) and wild-type mice with age-related OA and surgically-induced OA were evaluated by OA scoring systems. Levels of autophagy proteins and superoxide dismutase 2 (SOD2) were determined by immunohistochemistry. The relationship between HO-1 and the protective effects for OA was determined in chondrocytes treated with small interfering RNA (siRNA) targeting HO-1 gene.

Results: HO-1 expression decreased with aging in articular cartilages and menisci of mouse knees. Bach1 (-/-) mice showed reduced severity of age-related OA and surgically-induced OA compared with wild-type mice. Microtubule-associated protein 1 light chain 3 (LC3), autophagy marker, and SOD2 were increased in articular cartilage of Bach1 (-/-) mice compared with wild-type mice. Interleukin-1β (IL-1β) induced a significant increase in Adamts-5 in wild-type chondrocytes but not in Bach1(-/-) chondrocytes. The expression of SOD2 and the suppression of apoptosis in Bach1 (-/-) chondrocytes were mediated by HO-1.

Conclusions: Bach1 deficiency reduces the severity of OA-like changes. This may be due to maintenance of cartilage homeostasis and joint health by antioxidant effects through HO-1 and downregulation of extracellular matrix degrading enzymes. These results suggest that inactivation of Bach1 is a novel target and signaling pathway in OA prevention.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13075-015-0792-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4603301PMC
October 2015

Mesenchymal-stem-cell-derived exosomes accelerate skeletal muscle regeneration.

FEBS Lett 2015 May 8;589(11):1257-65. Epub 2015 Apr 8.

Department of Orthopedic Surgery, Hiroshima University, Hiroshima, Japan.

Mesenchymal stem cell (MSC) transplantation is used for treatment of many diseases. The paracrine role of MSCs in tissue regeneration is attracting particular attention. We investigate the role of MSC exosomes in skeletal muscle regeneration. MSC exosomes promote myogenesis and angiogenesis in vitro, and muscle regeneration in an in vivo model of muscle injury. Although MSC exosomes had low concentrations of muscle-repair-related cytokines, a number of repair-related miRNAs were identified. This study suggests that the MSC-derived exosomes promote muscle regeneration by enhancing myogenesis and angiogenesis, which is at least in part mediated by miRNAs such as miR-494.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.febslet.2015.03.031DOI Listing
May 2015

Exosomes from IL-1β stimulated synovial fibroblasts induce osteoarthritic changes in articular chondrocytes.

Arthritis Res Ther 2014 Aug 4;16(4):R163. Epub 2014 Aug 4.

Introduction: Osteoarthritis (OA) is a whole joint disease, and characterized by progressive degradation of articular cartilage, synovial hyperplasia, bone remodeling and angiogenesis in various joint tissues. Exosomes are a type of microvesicles (MVs) that may play a role in tissue-tissue and cell-cell communication in homeostasis and diseases. We hypothesized that exosomes function in a novel regulatory network that contributes to OA pathogenesis and examined the function of exosomes in communication among joint tissue cells.

Methods: Human synovial fibroblasts (SFB) and articular chondrocytes were obtained from normal knee joints. Exosomes isolated from conditioned medium of SFB were analyzed for size, numbers, markers and function. Normal articular chondrocytes were treated with exosomes from SFB, and Interleukin-1β (IL-1β) stimulated SFB. OA-related genes expression was quantified using real-time PCR. To analyze exosome effects on cartilage tissue, we performed glycosaminoglycan release assay. Angiogenic activity of these exosomes was tested in migration and tube formation assays. Cytokines and miRNAs in exosomes were analyzed by Bio-Plex multiplex assay and NanoString analysis.

Results: Exosomes from IL-1β stimulated SFB significantly up-regulated MMP-13 and ADAMTS-5 expression in articular chondrocytes, and down-regulated COL2A1 and ACAN compared with SFB derived exosomes. Migration and tube formation activity were significantly higher in human umbilical vein endothelial cells (HUVECs) treated with the exosomes from IL-1β stimulated SFB, which also induced significantly more proteoglycan release from cartilage explants. Inflammatory cytokines, IL-6, MMP-3 and VEGF in exosomes were only detectable at low level. IL-1β, TNFα MMP-9 and MMP-13 were not detectable in exosomes. NanoString analysis showed that levels of 50 miRNAs were differentially expressed in exosomes from IL-1β stimulated SFB compared to non-stimulated SFB.

Conclusions: Exosomes from IL-1β stimulated SFB induce OA-like changes both in vitro and in ex vivo models. Exosomes represent a novel mechanism by which pathogenic signals are communicated among different cell types in OA-affected joints.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/ar4679DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4261911PMC
August 2014

Administration of microRNA-210 promotes spinal cord regeneration in mice.

Spine (Phila Pa 1976) 2014 Jun;39(14):1099-107

*Department of Orthopaedic Surgery, Graduate School of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan †Translational Research Medical Center, Hiroshima University Hospital, Hiroshima, Japan ‡Department of Analysis and Control of Upper Extremity Function, Graduate School of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan; and §Department of Pediatric gastroenterology, University of California San Diego, CA.

Study Design: Experimental animal study of treatment of spinal cord injury (SCI).

Objective: To investigate the therapeutic effects of administering microRNA-210 (miR-210) to promote angiogenesis in a mouse SCI model.

Summary Of Background Data: Despite many previous studies regarding SCI, there is no established treatment in clinical practice. miRNAs have attracted immense attention because of their crucial role in human disease, and they have been proposed as potential new therapeutic targets for SCI.

Methods: At specific times after administration, mice were analyzed by several methods to examine the distribution of miR-210, histological angiogenesis and neurogenesis, functional recovery from SCI, and the expression levels of target genes of miR-210.

Results: After injection of miR-210 into the lesion of the injured spinal cord, expression of endogenous miR-210 increased until 6 days after injection. The administration of miR-210 promoted angiogenesis and astrogliosis, and improved functional recovery after SCI compared with the noninjected controls. Furthermore, the area made up of axons and myelin in the spinal cord tissues caudal to the injury site was larger in mice injected with miR-210 than those of the controls. Apoptotic cell death was lower in mice administered miR-210. After administration of miR-210, the expressions of protein-tyrosine phosphate 1B and ephrin-A3, both gene targets of miR-210, were downregulated at the protein level and protein-tyrosine phosphate 1B expression was also downregulated at the transcriptional level.

Conclusion: MiR-210 might contribute to spinal cord repair by promoting angiogenesis via the inhibition of protein-tyrosine phosphate 1B and ephrin-A3.

Level Of Evidence: N/A.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/BRS.0000000000000356DOI Listing
June 2014

Exosome-formed synthetic microRNA-143 is transferred to osteosarcoma cells and inhibits their migration.

Biochem Biophys Res Commun 2014 Mar 10;445(2):381-7. Epub 2014 Feb 10.

Department of Orthopedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima, Japan.

MicroRNAs (miRNAs) have emerged as potential anticancer agents, but their clinical application is limited by the lack of an effective delivery system to tumors. Exosomes are small vesicles that play important roles in intercellular communication. Here, we show that synthetic miR-143 introduced into cells is released enveloped in exosomes and that the secreted exosome-formed miR-143 is transferred to osteosarcoma cells. The delivery of exosome-formed miR-143 significantly reduced the migration of osteosarcoma cells. The delivery efficiency of exosome-formed miR-143 was less than that achieved with lipofection, but the migratory potential of osteosarcoma cells was similarly inhibited after both strategies. Our results suggest that exosomes can deliver synthetic miR-143 and are a potentially efficient and functional delivery system.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bbrc.2014.02.007DOI Listing
March 2014

Analysis of microRNAs expressions in chondrosarcoma.

J Orthop Res 2013 Dec 12;31(12):1992-8. Epub 2013 Aug 12.

Department of Systems Biomedicine, National Research Institute for Child Health and Development, Tokyo, Japan.

MicroRNAs (miRNAs) are small non-coding RNAs capable of inhibiting gene expression post-transcriptionally and expression profiling can provide therapeutic targets and tools for cancer diagnosis. Chondrosarcoma is a mesenchymal tumor with unknown cause and differentiation status. Here, we profiled miRNA expression of chondrosarcoma, namely clinical samples from human conventional chondrosarcoma tissue, established chondrosarcoma cell lines, and primary non-tumorous adult articular chondrocytes, by miRNA array and quantitative real-time PCR. A wide variety of miRNAs were differently downregulated in chondrosarcoma compared to non-tumorous articular chondrocytes; 27 miRNAs: miR-10b, 23b, 24-1*, 27b, 100, 134, 136, 136*, 138, 181d, 186, 193b, 221*, 222, 335, 337-5p, 376a, 376a*, 376b, 376c, 377, 454, 495, 497, 505, 574-3p, and 660, were significantly downregulated in chondrosarcoma and only 2: miR-96 and 183, were upregulated. We further validated the expression levels of miRNAs by quantitative real-time PCR for miR-181a, let-7a, 100, 222, 136, 376a, and 335 in extended number of chondrosarcoma clinical samples. Among them, all except miR-181a were found to be significantly downregulated in chondrosarcoma derived samples. The findings provide potential diagnostic value and new molecular understanding of chondrosarcoma.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jor.22457DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3835201PMC
December 2013

The effect of intra-articular injection of microRNA-210 on ligament healing in a rat model.

Am J Sports Med 2012 Nov 17;40(11):2470-8. Epub 2012 Sep 17.

Department of Orthopaedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University, Japan.

Background: It is known from clinical and experimental studies that the healing potential of the anterior cruciate ligament (ACL) is extremely poor and that early phases of ligament healing require an augmented blood supply. MicroRNA (miRNA) is a type of small, noncoding RNA that negatively regulates gene expression, and miRNA (miR)-210 is reported to be crucial for cell response to hypoxia, vascular endothelial growth factor (VEGF)-driven endothelial cell migration, and formation of capillary-like structures.

Purpose: The purpose of this study was to examine the effect of intra-articular injection of miRNA miR-210 on acceleration of ACL healing.

Study Design: Controlled laboratory study.

Methods: Two experiments were performed in this study. The ACLs of 12-week-old male LEW/CrlCrlj rats were partially transected. First, the temporal expression change of miR-210 after ACL injury was analyzed using real-time polymerase chain reaction (PCR) on day zero, and 1, 2, and 4 weeks after injury (n = 5 at each time point). Next, intra-articular injection of double-stranded (ds) miR-210 with atelocollagen was performed soon after injury. The control group was injected with control small interfering RNA (siRNA). Four weeks after injection, biomechanical and histological assessments of samples stained with H&E as well as Masson trichrome, and immunohistochemistry for VEGF, fibroblast growth factor 2 (FGF2), isolectin B4, and collagen type I, were performed. Real-time PCR analysis was also performed for quantitative evaluation of miR-210, VEGF-A, and collagen type I.

Results: Real-time PCR analysis revealed that miR-210 expression was decreased soon after injury but gradually increased thereafter. Histological analysis confirmed that the transected area was covered with healing tissue in the miR-210 group but remained devoid of any tissue in the control group 4 weeks after injury. Biomechanical analysis confirmed the improvement of biomechanical properties in the miR-210 group; the ultimate failure loads 4 weeks after injection were 30.5 ± 3.1 N in the miR-210 group and 22.8 ± 3.1 N in the control group (P < .05). Real-time PCR analysis showed that endogenous miR-210, VEGF, and collagen type I were highly expressed compared with controls, and immunohistochemistry for VEGF, FGF2, isolectin B4, and collagen type I showed that VEGF and FGF2 were highly upregulated, and there were abundant blood vessels and fibrotic deposition in the miR-210 group.

Conclusion: Injection of ds miR-210 was effective in promoting the healing of partially torn ACLs through enhancement of angiogenesis via upregulation of VEGF and FGF2.

Clinical Relevance: It might represent a potential therapeutic approach for treatment of ACL injury.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/0363546512458894DOI Listing
November 2012

Macro view of microRNA function in osteoarthritis.

Nat Rev Rheumatol 2012 Sep 14;8(9):543-52. Epub 2012 Aug 14.

Department of Regenerative Medicine, Hiroshima University Hospital, 1-2-3 Kasumi, Hiroshima 734-8551, Japan.

Osteoarthritis (OA), the most common musculoskeletal disorder, is complex, multifaceted, and characterized by degradation of articular cartilage and alterations in other joint tissues. Although some pathogenic pathways have been characterized, current knowledge is incomplete and effective approaches to the prevention or treatment of OA are lacking. Understanding novel molecular mechanisms that are involved in the maintenance and destruction of articular cartilage, including extracellular regulators and intracellular signalling mechanisms in joint cells that control cartilage homeostasis, has the potential to identify new therapeutic targets in OA. MicroRNAs control tissue development and homeostasis by fine-tuning gene expression, with expression patterns specific to tissues and developmental stages, and are increasingly implicated in the pathogenesis of complex diseases such as cancer and cardiovascular disorders. The emergent roles of microRNAs in cartilage homeostasis and OA pathogenesis are summarized in this Review, alongside potential clinical applications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nrrheum.2012.128DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3572197PMC
September 2012

L-Sox5 and Sox6 proteins enhance chondrogenic miR-140 microRNA expression by strengthening dimeric Sox9 activity.

J Biol Chem 2012 Jun 30;287(26):22206-15. Epub 2012 Apr 30.

Department of Systems Biomedicine, National Research Institute for Child Health and Development, 2-10-1 Okura, Setagaya-ku, Tokyo 157-8535, Japan.

Sox9 plays a critical role in early chondrocyte initiation and promotion as well as repression of later maturation. Fellow Sox family members L-Sox5 and Sox6 also function as regulators of cartilage development by boosting Sox9 activation of chondrocyte-specific genes such as Col2a1 and Agc1; however, the regulatory mechanism and other target genes are largely unknown. MicroRNAs are a class of short, non-coding RNAs that act as negative regulators of gene expression by promoting target mRNA degradation and/or repressing translation. Analysis of genetically modified mice identified miR-140 as a cartilage-specific microRNA that could be a critical regulator of cartilage development and homeostasis. Recent findings suggest Sox9 promotes miR-140 expression, although the detailed mechanisms are not fully understood. In this study we demonstrate that the proximal upstream region of pri-miR-140 has chondrogenic promoter activity in vivo. We found an L-Sox5/Sox6/Sox9 (Sox trio) response element and detailed binding site in the promoter region. Furthermore, detailed analysis suggests the DNA binding and/or transactivation ability of Sox9 as a homodimer is boosted by L-Sox5 and Sox6. These findings provide new insight into cartilage-specific gene regulation by the Sox trio.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M112.343194DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3381182PMC
June 2012

Angiogenic microRNA-210 is present in cells surrounding osteonecrosis.

J Orthop Res 2012 Aug 27;30(8):1263-70. Epub 2012 Jan 27.

Department of Orthopaedic Surgery, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8551, Japan.

A role of microRNAs (miRNAs), which are ≈ 22-nucleotide non-coding RNAs, has recently been recognized in human diseases. The objective of this study was to identify the expression pattern of miRNA (miR)-210, known to be associated with angiogenesis, in bone from patients with osteonecrosis (ON) of the femoral head. The expression of miR-210 in bone from 10 patients with osteoarthritis (OA) of the hip and ten with ON was analyzed by quantitative reverse transcription-polymerase chain reaction (RT-PCR) and by in situ hybridization. In addition, immunohistochemical staining for von Willebrand factor (vWF) and vascular endothelial growth factor (VEGF) was performed to identify the miR-210 expressing cells. We found that in ON samples, the expression of mature, primary miR-210, VEGF, matrix metalloproteinase (MMP)-2, and MMP-7 was significantly higher than that of OA samples. Section in situ hybridization of mature miR-210 revealed that mature miR-210 is expressed around the necrotic area. vWF and VEGF were also strongly expressed in the miR-210 expressing cells. This study shows that miR-210 is intensely expressed in ON, and might play a role in ON pathogenesis. The present study provides a solid basis for further functional analyses of miRNAs in ON.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jor.22079DOI Listing
August 2012

Autophagy activation by rapamycin reduces severity of experimental osteoarthritis.

Ann Rheum Dis 2012 Apr 14;71(4):575-81. Epub 2011 Nov 14.

Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California, USA.

Objectives: Osteoarthritis is associated with cell death and extracellular matrix degradation in articular cartilage. Autophagy is an essential cellular homeostasis mechanism that was found to be deficient in ageing and osteoarthritic cartilage. This study determined whether pharmacological inhibition of the mammalian target of rapamycin (mTOR), a key inhibitor of autophagy, has disease-modifying activity in experimental osteoarthritis.

Methods: Experimental osteoarthritis was induced by transection of the medial meniscotibial ligament and the medial collateral ligament in 2-month-old C57Bl/6 mice (n=36). Rapamycin (1 mg/kg weight/day) (n=18 mice) or dimethyl sulphoxide vehicle control (n=18 mice) was administered intraperitoneally for 10 weeks. Histopathological changes in articular cartilage and synovium were examined by using semiquantitative scoring systems. Rapamycin effects on mTOR signalling, autophagy, cartilage homeostasis and inflammation were analysed by immunohistochemistry and immunofluorescence staining.

Results: Rapamycin affected the mTOR signalling pathway in mouse knee joints as indicated by the inhibition of ribosomal protein S6 phosphorylation, a target of mTOR and activation of LC3, a main marker of autophagy. The severity of cartilage degradation was significantly (p<0.01) reduced in the rapamycin-treated group compared with the control group and this was associated with a significant (p<0.05) decrease in synovitis. Rapamycin treatment also maintained cartilage cellularity and decreased ADAMTS-5 and interleukin-1β expression in articular cartilage.

Conclusions: These results suggest that rapamycin, at least in part by autophagy activation, reduces the severity of experimental osteoarthritis. Pharmacological activation of autophagy may be an effective therapeutic approach for osteoarthritis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/annrheumdis-2011-200557DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3294168PMC
April 2012

Anterior cruciate ligament changes in the human knee joint in aging and osteoarthritis.

Arthritis Rheum 2012 Mar;64(3):696-704

Department of Molecular and Experimental Medicine,The Scripps Research Institute, La Jolla, CA 92037, USA.

Objective: The development and patterns of spontaneous age-related changes in the anterior cruciate ligament (ACL) and their relationship to articular cartilage degeneration are not well characterized. This study was undertaken to investigate the types and temporal sequence of age-related ACL changes and to determine their correlation with cartilage lesion patterns at all stages of osteoarthritis (OA) development in human knee joints without prior joint trauma.

Methods: Human knee joints (n = 120 from 65 donors ages 23-92) were obtained at autopsy, and ACLs and cartilage were graded macroscopically and histologically. Inflammation surrounding the ACL was assessed separately.

Results: Histologic ACL substance scores and ligament sheath inflammation scores increased with age. Collagen fiber disorganization was the earliest and most prevalent change. The severity of mucoid degeneration and chondroid metaplasia in the ACL increased with the development of cartilage lesions. A correlation between ACL degeneration and cartilage degeneration was observed, especially in the medial compartment of the knee joint.

Conclusion: Our findings indicate that ACL degeneration is highly prevalent in knees with cartilage defects and may even precede cartilage changes. Hence, ACL deficiencies may not only be important in posttraumatic OA, but may also be a feature associated with knee OA pathogenesis in general.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/art.33417DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3266452PMC
March 2012

Translational suppression of atrophic regulators by microRNA-23a integrates resistance to skeletal muscle atrophy.

J Biol Chem 2011 Nov 18;286(44):38456-38465. Epub 2011 Sep 18.

Division of Regenerative Medical Engineering, Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo, Bunkyo, Tokyo 113-0033, Japan; Institute for Biomedical Engineering Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku, Tokyo 162-0041, Japan; Venetian Institute of Molecular Medicine, 35129 Padova, Italy. Electronic address:

Muscle atrophy is caused by accelerated protein degradation and occurs in many pathological states. Two muscle-specific ubiquitin ligases, MAFbx/atrogin-1 and muscle RING-finger 1 (MuRF1), are prominently induced during muscle atrophy and mediate atrophy-associated protein degradation. Blocking the expression of these two ubiquitin ligases provides protection against muscle atrophy. Here we report that miR-23a suppresses the translation of both MAFbx/atrogin-1 and MuRF1 in a 3'-UTR-dependent manner. Ectopic expression of miR-23a is sufficient to protect muscles from atrophy in vitro and in vivo. Furthermore, miR-23a transgenic mice showed resistance against glucocorticoid-induced skeletal muscle atrophy. These data suggest that suppression of multiple regulators by a single miRNA can have significant consequences in adult tissues.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M111.271270DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3207415PMC
November 2011