Publications by authors named "Kumi Orita"

23 Publications

  • Page 1 of 1

Biglycan expression and its function in human ligamentum flavum.

Sci Rep 2021 Mar 1;11(1):4867. Epub 2021 Mar 1.

Department of Orthopaedic Surgery, Osaka City University Graduate School of Medicine, 1-4-3 Asahi Machi, Abeno-Ku, Osaka, 545-8585, Japan.

Hypertrophy of the ligamentum flavum (LF) is a major cause of lumbar spinal stenosis (LSS), and the pathology involves disruption of elastic fibers, fibrosis with increased cellularity and collagens, and/or calcification. Previous studies have implicated the increased expression of the proteoglycan family in hypertrophied LF. Furthermore, the gene expression profile in a rabbit experimental model of LF hypertrophy revealed that biglycan (BGN) is upregulated in hypertrophied LF by mechanical stress. However, the expression and function of BGN in human LF has not been well elucidated. To investigate the involvement of BGN in the pathomechanism of human ligamentum hypertrophy, first we confirmed increased expression of BGN by immunohistochemistry in the extracellular matrix of hypertrophied LF of LSS patients compared to LF without hypertrophy. Experiments using primary cell cultures revealed that BGN promoted cell proliferation. Furthermore, BGN induces changes in cell morphology and promotes myofibroblastic differentiation and cell migration. These effects are observed for both cells from hypertrophied and non-hypertrophied LF. The present study revealed hyper-expression of BGN in hypertrophied LF and function of increased proteoglycan in LF cells. BGN may play a crucial role in the pathophysiology of LF hypertrophy through cell proliferation, myofibroblastic differentiation, and cell migration.
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http://dx.doi.org/10.1038/s41598-021-84363-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7921587PMC
March 2021

Expression and function of fibroblast growth factor 1 in the hypertrophied ligamentum flavum of lumbar spinal stenosis.

J Orthop Sci 2021 Feb 23. Epub 2021 Feb 23.

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Background: Fibrosis is one of the main pathologies caused by hypertrophy of the ligamentum flavum (LF), which leads to lumbar spinal stenosis (LSS). The fibroblast growth factor (FGF) family is a key mediator of fibrosis. However, acidic fibroblast growth factor (FGF-1) expression and function are not well understood in LF. This study sought to evaluate FGF-1 expression in the hypertrophied and non-hypertrophied human LF, and to investigate its function using primary human LF cell cultures.

Methods: We obtained hypertrophied lumbar LF from LSS patients and non-hypertrophied lumbar LF from control patients during surgery. Immunohistochemistry and qPCR were performed to evaluate FGF-1 expression in LF tissue. The function of FGF-1 and transforming growth factor beta 1 (TGF-β1) was also investigated using primary LF cell culture. The effects on cell morphology and cell proliferation were examined using a crystal violet staining assay and MTT assay, respectively. Immunocytochemistry, western blotting, and qPCR were performed to evaluate the effect of FGF-1 on TGF-β1-induced myofibroblast differentiation and fibrosis.

Results: Immunohistochemistry and qPCR showed higher FGF-1 expression in hypertrophied LF compared to control LF. Crystal violet staining and MTT assay revealed that FGF-1 decreases LF cell size and inhibits their proliferation in a dose-dependent manner, whereas TGF-β1 increases cell size and promotes proliferation. Immunocytochemistry and western blotting further demonstrated that TGF-β1 increases, while FGF-1 decreases, α-SMA expression in LF cells. Moreover, FGF-1 also caused downregulation of collagen type 1 and type 3 expression in LF cells.

Conclusion: FGF-1 is highly upregulated in the LF of LSS patients. Meanwhile, in vitro, FGF-1 exhibits antagonistic effects to TGF-β1 by inhibiting cell proliferation and decreasing LF cell size as well as the expression of fibrosis markers. These results suggest that FGF-1 has an anti-fibrotic role in the pathophysiology of LF hypertrophy.
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http://dx.doi.org/10.1016/j.jos.2021.01.004DOI Listing
February 2021

Expression and function of FGF9 in the hypertrophied ligamentum flavum of lumbar spinal stenosis patients.

Spine J 2021 Feb 10. Epub 2021 Feb 10.

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Background Context: Ligamentum flavum (LF) hypertrophy plays a dominant role in lumbar spinal stenosis (LSS). A previous study found that fibroblast growth factor 9 (FGF9) was upregulated with mechanical stress in rabbit LF. However, the expression and function of FGF9 are not well understood in human LF.

Purpose: To evaluate FGF9 expression and function in human LF with and without hypertrophy.

Study Design: This study employed a basic research study design utilizing human LF tissue for histological analyses.

Patient Samples: Hypertrophied LF tissue sample from patients with LSS, and nonhypertrophied (control) LFs from patients with lumbar disc herniation or other diseases were obtained during surgery.

Methods: LF specimens were histologically analyzed for FGF9 and vascular endothelial growth factor A (VEGF-A) by immunohistochemistry. The number of total and FGF9 immuno-positive cells and blood vessels were counted and compared between LF with and without hypertrophy. For functional analysis, the effect of FGF9 on cell proliferation and migration was examined using a primary cell culture of human LF.

Results: Histological studies revealed that the total cell number was significantly higher in the LF of patients with LSS than in the LF of control patients. Immunohistochemistry showed that the percentage of FGF9-positive cells was significantly higher in the LF of patients with LSS than in the controls, and it positively correlated with patients' age, regardless of disease. Double immune-positive cells for FGF9 and VEGF-A were often observed in vascular endothelial cells and fibroblasts in the fibrotic area of hypertrophied LF, and the number of double positive vessels was significantly higher in LF of LSS patients than in the LF of controls. Primary cell culture of human LF revealed that FGF9 promoted the proliferation and migration of LF cells.

Conclusion: The present study demonstrated that FGF9 expression is highly upregulated in hypertrophied human LF. FGF9 potentially plays a pivotal role in the process of hypertrophy of LF, which is associated with mechanical stress, through cell proliferation and migration.

Clinical Significance: The results from this study partially reveal the molecular mechanisms of LF hypertrophy and suggest that FGF9 may be involved in the process of LF degeneration in elderly patients.
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http://dx.doi.org/10.1016/j.spinee.2021.02.004DOI Listing
February 2021

Risk Assessment for Pathological Fracture After Bone Tumour Biopsy.

Anticancer Res 2021 Feb;41(2):679-686

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Aim: This study aimed to identify the risk of pathological fractures after bone tumour biopsy.

Materials And Methods: Fifty rabbit femurs were divided into groups according to defect size: Control (no defect), type 1 (10% width), type 2 (20% width), type 3 (30% width), and type 4 (40% width). Another 20 were also divided into control, type A (27% length), type B (40% length), and type C (53% length) groups. Performing femoral head compression testing allowed each parameter (maximum load, displacement, elastic modulus, and fracture energy) to be calculated individually.

Results: Compressive maximum load was significantly higher for type 1 than for the other types when testing rectangular defects of different widths, while there were no significant differences between the three types when testing by defect length.

Conclusion: It may be useful for orthopaedic oncologists to make a rectangular biopsy hole with a width measuring less than 10% of the circumference and to enlarge the hole longitudinally to avoid pathological fracture.
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http://dx.doi.org/10.21873/anticanres.14819DOI Listing
February 2021

Vitamin E-infused highly cross-linked polyethylene did not reduce the number of in vivo wear particles in total knee arthroplasty.

Bone Joint J 2020 Nov;102-B(11):1527-1534

Department of Orthopaedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Aims: Vitamin E-infused highly cross-linked polyethylene (E1) has recently been introduced in total knee arthroplasty (TKA). An in vitro wear simulator study showed that E1 reduced polyethylene wear. However there is no published information regarding in vivo wear. Previous reports suggest that newly introduced materials which reduce in vitro polyethylene wear do not necessarily reduce in vivo polyethylene wear. To assist in the evaluation of the newly introduced material before widespread use, we established an in vivo polyethylene wear particle analysis for TKA. The aim of this study was to compare in vivo polyethylene wear particle generation between E1 and conventional polyethylene (ArCom) in TKA.

Methods: A total of 34 knees undergoing TKA (17 each with ArCom or E1) were investigated. Except for the polyethylene insert material, the prostheses used for both groups were identical. Synovial fluid was obtained at a mean of 3.4 years (SD 1.3) postoperatively. The in vivo polyethylene wear particles were isolated from the synovial fluid using a previously validated method and examined by scanning electron microscopy.

Results: The total number of polyethylene wear particles obtained from the knees with E1 (mean 6.9, SD 4.0 × 10 counts/knee) was greater than that obtained from those with ArCom (mean 2.2, SD 2.6 × 10 counts/knee) (p = 0.001). The particle size (equivalent circle of diameter) from the knees with E1 was smaller (mean 0.5 μm, SD 0.1) than that of knees with ArCom (mean 1.5, SD 0.3 μm) (p = 0.001). The aspect ratio of particles from the knees with E1 (mean 1.3, SD 0.1) was smaller than that with ArCom (mean 1.4, SD 0.1) (p < 0.001 ).

Conclusion: This is the first report of in vivo wear particle analysis of E1. E1 polyethylene did not reduce the number of in vivo polyethylene wear particles compared with ArCom in early clinical stage. Further careful follow-up of newly introduced E1 for TKA should be carried out. Cite this article: 2020;102-B(11):1527-1534.
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http://dx.doi.org/10.1302/0301-620X.102B11.BJJ-2020-0413.R1DOI Listing
November 2020

Role of Momordica charantia in preventing the natural aging process of skin and sexual organs in mice.

Dermatol Ther 2020 11 18;33(6):e14243. Epub 2020 Sep 18.

Kobayashi Dermatology Clinic, Nishinomiya, Japan.

Although various methods for improving the natural aging of skin have been examined, an effective method is currently unavailable. Therefore, in this study, we investigated the effects of Momordica charantia on the natural aging of skin of mice and how sex differences influenced these effects. To this end, we bred female and male hairless mice without ultraviolet ray irradiation and physical stress for 2 years. During the study period, mice were orally administered 50 mg/kg/day Momordica charantia fruit extract, three times per week. The characteristics of naturally aging skin, in terms of moisture retention, hydration, thickness, and reduced wrinkle score, improved after Momordica charantia treatment in both male and female mice. Furthermore, reduced cell apoptosis was observed in the female ovaries and male testes, and the levels of testosterone and 17β-estradiol in blood were maintained. After treatment with Momordica charantia, the expression of matrix metalloprotease (MMP)-1 and hyaluronidase (HAYL)2 decreased in the skin of female mice, whereas the serum levels of interleukin (IL)-33 increased in the male mice. These results indicated that the natural aging of the skin was decelerated by Momordica charantia via regulation of the 17β-estradiol/mast cell/MMP-1/HAYL2 and testosterone/mast cell/IL-33 signaling pathways in female and male mice, respectively.
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http://dx.doi.org/10.1111/dth.14243DOI Listing
November 2020

Intra-Articular Injection of Stromal Cell-Derived Factor 1α Promotes Meniscal Healing via Macrophage and Mesenchymal Stem Cell Accumulation in a Rat Meniscal Defect Model.

Int J Mol Sci 2020 Jul 30;21(15). Epub 2020 Jul 30.

Department of Orthopaedic Surgery, Osaka City University Graduate School of Medicine, Osaka 545-8585, Japan.

The stromal-cell-derived factor-1α (SDF-1) is well-known for playing important roles in the regeneration of tissue by enhancing cell migration. However, the effect of SDF-1 in meniscal healing remains unknown. The purpose of this study is to investigate the effects of intra-articular injection of SDF-1 on meniscus healing in a rat meniscal defect model. The intra-articular SDF-1 injection was performed at meniscectomy and one week later. Macroscopic and histological assessments of the reparative meniscus were conducted at one, two and six weeks after meniscectomy in rats. In the macroscopic evaluation, the SDF-1 group showed an increase in the size of the reparative meniscus at six weeks after meniscectomy compared to the phosphate-buffered saline (PBS) injection (no-treatment) group. Histological findings showed that intra-articular injection of SDF-1 enhanced the migration of macrophages to the site of the regenerative meniscus at one and two weeks after meniscectomy. CD68- and CD163-positive cells in the SDF-1 group at one week after meniscectomy were significantly higher than in the no-treatment group. CD163-positive cells in the SDF-1 group at two weeks were significantly higher than in the no-treatment group. At one week after meniscectomy, there were cells expressing mesenchymal-stem-cell-related markers in the SDF-1 group. These results indicate the potential of regenerative healing of the meniscus by SDF-1 injection via macrophage and mesenchymal stem cell accumulation. In the present study, intra-articular administration of SDF-1 contributed to meniscal healing via macrophage, CD90-positive cell and CD105-positive cell accumulation in a rat meniscal defect model. The SDF-1-CXCR4 pathway plays an important role in the meniscal healing process. For potential clinical translation, SDF-1 injection therapy seems to be a promising approach for the biological augmentation in meniscal injury areas to enhance healing capacity.
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http://dx.doi.org/10.3390/ijms21155454DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7432222PMC
July 2020

Ameliorative effect of green odor against UVB-induced immunosuppression of contact hypersensitivity.

Dermatol Ther 2020 11 8;33(6):e13848. Epub 2020 Jul 8.

Department of Pharmaceutical Sciences, Suzuka University of Medical Science, Suzuka, Japan.

Ultraviolet (UV) irradiation to the eye induces photoimmunosuppression. In here, we examined the effect of green odor against immunosuppression of contact hypersensitivity in the eye induced by ultraviolet B (UVB) irradiation. Systemic immunosuppression was induced in ICR mice sensitized with 0.5% oxazolone through the skin by a single exposure to UVB. Consecutive green odor treatment significantly counteracted UVB irradiation-induced immunosuppression of the contact hypersensitivity (CHS) response. The green odor treatment increased dopamine and β-endorphin levels in the brain and the plasma, respectively, and decreased the plasma corticosterone concentration in the oxazolone-sensitized mice after UVB irradiation to the eye, in contrast with that in acetone-treated mice (treatment negative control). Green odor prevented UVB irradiation-induced photoimmunosuppression of the CHS response by regulating the dopamine/β-endorphin/corticosterone pathway.
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http://dx.doi.org/10.1111/dth.13848DOI Listing
November 2020

Promising abscopal effect of combination therapy with thermal tumour ablation and intratumoural OK-432 injection in the rat osteosarcoma model.

Sci Rep 2020 06 15;10(1):9679. Epub 2020 Jun 15.

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-Machi, Abeno-Ku, Osaka, 545-8585, Japan.

Treatment options for metastatic osteosarcoma are limited. The present study aimed to evaluate whether radiofrequency ablation (RFA) combined with intratumoural OK-432 injection induces systemic anti-tumour immunity in rat osteosarcoma model. Eighty of 145 rats were assigned to four groups to evaluate overall survival and tumour size: control (no treatment), RFA-only, OK-432, and RFA-OK-432. The remaining 65 were assigned for histological examination. Maximum diameters of tibial and lung tumours were determined. Tumour samples were histologically examined using haematoxylin-eosin and immunohistochemical staining. Overall survival was significantly prolonged in the RFA-OK-432 group compared to the RFA-only and OK-432 groups. Only rats in the RFA-OK-432 group exhibited significant decreases in maximum tumour diameter after treatment. Ki-67-positive tumour cells in the RFA-OK-432 group were significantly stained negative on immunohistochemical analysis as opposed to those in the RFA-only and OK-432 groups. The number of CD11c+, OX-62+, CD4+, and CD8 + cells significantly increased in the RFA-OK-432 group compared to the RFA-only group. RFA with intratumoural OK-432 injection resulted in distant tumour suppression, prolonged survival, and increased dendritic cells, cytotoxic T cells, IFN-γ, and TNF-α, whereas RFA or OK-432 alone did not produce this effect. This combination may induce an abscopal effect in human osteosarcoma.
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http://dx.doi.org/10.1038/s41598-020-66934-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7296025PMC
June 2020

Local transplantation of adipose-derived stem cells has a significant therapeutic effect in a mouse model of rheumatoid arthritis.

Sci Rep 2020 02 20;10(1):3076. Epub 2020 Feb 20.

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Adipose-derived stem cells (ADSCs) have anti-inflammatory and regenerative properties. The purpose of this study was to investigate the effect of locally administered ADSCs in a rheumatoid arthritis (RA) mouse model. In an in vivo experiment, single-cell ADSCs and three dimensionally-cultured ADSC spheroids were injected intra-articularly into the knees of RA model mice and histologically assessed. Marked improvement of synovial inflammation and articular cartilage regeneration was found in ADSC-treated mice. Proliferation, migration, and apoptosis assays of synovial fibroblasts incubated with single-cell and spheroid ADSCs were performed. The expression levels of total cytokine RNA in ADSC single cells, spheroids, and ADSC-treated inflammatory synovial fibroblasts were also evaluated by quantitative reverse transcription PCR. ADSCs suppressed the proliferation and migration of activated inflammatory cells and downregulated inflammatory cytokines. TSG-6 and TGFβ1 were significantly upregulated in ADSCs compared to controls and TGFβ1 was significantly upregulated in ADSC spheroids compared to single cells. The apoptosis rate of ADSC spheroids was significantly lower than that of single-cell ADSCs. These results indicated that intra-articular administration of ADSC single cells and spheroids was effective in an RA mouse model, offering a novel approach for the development of effective localized treatments for patients with RA.
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http://dx.doi.org/10.1038/s41598-020-60041-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7033196PMC
February 2020

Ameliorative Effect of Hochu-ekki-to on Natural Skin Aging.

Pharmacology 2020 6;105(7-8):454-460. Epub 2019 Dec 6.

Department of Pharmaceutical Sciences, Suzuka University of Medical Science, Suzuka, Japan,

Introduction: Although it is beneficial to protect the skin from natural aging, especially in an aging society, the approach by which this can be achieved is still not well known. Hochu-ekki-to, a Chinese natural medicine, has various advantageous effects; however, there is no report about its influence on skin aging.

Objective: Therefore, we examined the effect of hochu-ekki-to against natural aging.

Methods: Hairless mice, bred without ultraviolet ray irradiation and physical stress, were orally administered huchu-ekki-to 3 times per week for 2 years. After that period, degree of skin hydration and permeability were measured. Furthermore, hematoxylin and eosin histochemistry was performed to determine the morphology and condition of the tissues. Lastly, levels of vitamin A, vitamin C, and reactive oxygen species (ROS) in plasma and skin, as well as concentration of hyaluronic acid in the skin, were measured.

Results: Signs of skin aging were ameliorated by administration of hochu-ekki-to, such as moisture retention, skin hydration, and the generation of wrinkles. Furthermore, vitamin A, vitamin C, collagen type I, collagen type III, fibroblasts, and hyaluronic acid levels in the skin increased, while levels of ROS decreased after hochu-ekki-to treatment.

Conclusion: These results indicated that natural skin aging was ameliorated by treatment with hochu-ekki-to, specifically moisture retention, and skin hydration, and thickening, via the regulation of the vitamin C/fibroblast, collagen type III/collagen type I, and vitamin A/hyaluronic acid signaling pathways.
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http://dx.doi.org/10.1159/000504985DOI Listing
February 2021

Changes in viscoelastic properties of articular cartilage in early stage of osteoarthritis, as determined by optical coherence tomography-based strain rate tomography.

BMC Musculoskelet Disord 2019 Sep 6;20(1):417. Epub 2019 Sep 6.

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi Abeno-ku, Osaka, 545-8585, Japan.

Background: Biomechanical changes in articular cartilage are associated with the onset of osteoarthritis. We developed an optical coherence tomography-based strain rate tomography method: stress relaxation optical coherence straingraphy (SR-OCSA). The purpose of this study was to establish an approach for measuring mechanical properties of articular cartilage using SR-OCSA, and to investigate the distribution of viscoelastic properties of articular cartilage in early osteoarthritis.

Methods: Anterior cruciate ligament transection surgery was performed on the left knees of 8-9-month-old New Zealand white rabbits. SR-OCSA was used to visualize and measure the viscoelastic properties of articular cartilage via attenuation coefficient of strain rate (ACSR). Using the same conditions as in the SR-OCSA test, an indentation test was conducted, and relaxation time was measured to evaluate the relationship between ACSR and relaxation time.

Results: SR-OCSA could nondestructively detect and visualize changes in the distribution of viscoelastic properties of articular cartilage in early osteoarthritis. SR-OCSA captured significant increases in ACSR in cartilage at 2 weeks after surgery, when a histologically slight osteoarthritis sign was present. As cartilage degeneration progressed, ACSR increased, whereas relaxation time decreased in a time-dependent manner. Moreover, ACSR negatively correlated with relaxation time. In particular, ACSR was elevated around the tidemark and the elevation tended to move as cartilage degeneration progressed.

Conclusions: SR-OCSA could tomographically and nondestructively detect and visualize changes in the distribution of viscoelastic properties of articular cartilage in early osteoarthritis. The mechanical properties around the tidemark were degraded as cartilage degeneration progressed. Thus, SR-OCSA provides important data needed to understand the biomechanics of early osteoarthritis.
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http://dx.doi.org/10.1186/s12891-019-2789-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6731561PMC
September 2019

Exploration of the Optimal Shape for Bone Tumour Biopsy.

Anticancer Res 2019 Aug;39(8):4191-4197

Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Background/aim: Biopsy hole for bone tumour biopsy may cause pathological fractures. This study aimed to identify the optimal shape of bone tumour biopsy hole using the rabbit femoral head compression test to avoid pathological fracture.

Materials And Methods: A compression test with no defect was performed to identify bone fracture location. Three shapes of biopsy holes (same size) were made artificially. Sixty rabbit femurs were randomly divided (n=15 each) into control (no defect), Shape 1 (round), Shape 2 (square), and Shape 3 (rectangular) groups.

Results: Twelve out of fifteen femurs were fractured on the femoral shaft; the femoral shaft was targeted for the compression test. Compressive maximum load and fracture energy were significantly higher for Shape 3 than for the other Shapes.

Conclusion: A rectangular biopsy hole helps minimise reduction in bone strength. The defect width may be related to fragility of the affected bone.
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http://dx.doi.org/10.21873/anticanres.13579DOI Listing
August 2019

The Clock Genes Are Involved in The Deterioration of Atopic Dermatitis after Day-and-Night Reversed Physical Stress in NC/Nga Mice.

Open Biochem J 2018 29;12:87-102. Epub 2018 Jun 29.

Department of Pharmaceutical Science, Suzuka University of Medical Science, 3500-3 Minamitamagakicho, Suzuka, Mie 513-8670, Japan.

Background: In modern society, irregular lifestyles are a problem. It is well known that Atopic Dermatitis (AD) occurs during physical stress in people with an irregular lifestyle. We evaluated the influence that day-and-night reversal physical stress has on AD.

Methods: Six-week-old specific-pathogen-free and conventional NC/Nga male mice were used. For the day-and-night reversal procedure, the mice ran on a treadmill at a slow speed of 10 m/min for 12 h (between 8:00 and 20:00). Then, between 20:00 and 8:00, we put the mice in a dark place. This treatment was repeated every day for two weeks. The behavioral circadian rhythm of the mice was evaluated with the open field test. Then, the mice were sacrificed and histological examinations of the tissues, the expression of peptide hormones, corticosterone, Immunoglobulin E, histamine, and cytokines was performed using an enzyme-linked immunosorbent assay.

Results: In the treadmill-treated conventional NC/Nga mice, AD symptoms were deteriorated compared with the non-treated conventional NC/Nga mice. The levels of Period (Per) 2, Clock, and brain and muscle arnt-like protein 1 (Bmal1) in the skin were increased constantly in the treadmill-treated conventional mice. Furthermore, the expression of Retinoic Acid-related Orphan Receptor (ROR)α, which activates Bmal1, was increased in the treadmill-treated conventional mice compared with the non-treated conventional mice. In addition, when non-treated conventional mice were administrated by the agonist of RORα, AD symptoms were deteriorated similar to treadmill-treated conventional mice.

Conclusion: In the day-and-night reversal mice, the clock genes were increased constantly, indicating that this is a factor that deteriorated AD.
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http://dx.doi.org/10.2174/1874091X01812010087DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6048832PMC
June 2018

The accelerated effect of recombinant human bone morphogenetic protein 2 delivered by β-tricalcium phosphate on tendon-to-bone repair process in rabbit models.

J Shoulder Elbow Surg 2018 May 12;27(5):894-902. Epub 2018 Feb 12.

Department of Orthopaedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

Background: Bone morphogenetic protein 2 (BMP-2) plays an important role in the tendon-to-bone repair process. However, there is no previous literature on acceleration of the tendon-to-bone repair process by BMP-2 delivered by β-tricalcium phosphate (β-TCP). The aim of this study was to investigate the accelerated effect of recombinant human BMP-2 (rhBMP-2) delivered by β-TCP on the tendon-to-bone repair process.

Methods: The infraspinatus tendon of elderly female Japanese white rabbits was detached from its insertion site on the humerus. A bone tunnel (4.2 mm) was created at the original insertion site of the tendon, which was repaired using the McLaughlin procedure after filling in β-TCP (porosity 75%) without BMP-2 (control group) or with 10 µg rhBMP-2 (BMP group). The rabbits were sacrificed at the second, fourth, and eighth weeks after surgery for histologic analysis and biomechanical testing. We also evaluated the maturity of the tendon-to-bone junction using the tendon-to-bone maturity score.

Results: Histologic analysis revealed no significant difference between the groups at 2 and 8 weeks but a more abundant organized fibrocartilage at the tendon-to-bone junction in the BMP group at 4 weeks. The tendon-to-bone maturity score improved sequentially. The interface of the BMP group at 4 weeks had significantly improved biomechanical properties than that of the control group.

Conclusion: The tendon-to-bone repair process was facilitated by the use of rhBMP-2 delivered by β-TCP at 4 weeks.
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http://dx.doi.org/10.1016/j.jse.2017.11.025DOI Listing
May 2018

The origin and distribution of CD68, CD163, and αSMA cells in the early phase after meniscal resection in a parabiotic rat model.

Connect Tissue Res 2017 Nov 6;58(6):562-572. Epub 2017 Feb 6.

a Department of Orthopaedic Surgery , Osaka City University Graduate School of Medicine , Osaka , Japan.

We previously reported that circulating peripheral blood-borne cells (PBCs) contribute to early-phase meniscal reparative change. Because macrophages and myofibroblasts are important contributors of tissue regeneration, we examined their origin and distribution in the reparative meniscus. Reparative menisci were evaluated at 1, 2, and 4 weeks post-meniscectomy by immunohistochemistry to locate monocytes and macrophages (stained positive for CD68 and CD163), and myofibroblasts (stained positive for αSMA). Of the total number of cells, 13% were CD68 at 1 week post-meniscectomy, which decreased to 1% by 4 weeks post-meniscectomy; of these, almost half of CD68 cells (49.4%: 98.8% as PBCs) were green fluorescent protein (GFP)-positive post-meniscectomy (1, 2, and 4 weeks), indicating that the majority of CD68 cells were derived from PBCs. Of the total cells, 6% were CD163 at 1 week post-meniscectomy, which decreased to 1% by week 4. Of the CD163 cells, the majority were GFP-positive (42.5%: 85.0% as PBCs) after 1 week; however, this decreased significantly over time, which indicates that the majority of CD163 cells are derived from PBCs during the early phase of meniscal reparative change, but are derived from resident cells at later time points. Of the total cells, 38% were αSMA at 1 week post-meniscectomy, which decreased to 3% by 4 weeks. The proportion of GFP-positive αSMA cells was 2.8% after 1 week, with no significant change over time, which indicates that the majority of αSMA cells originated from resident cells. Here, we describe the origin and distribution of macrophages and myofibroblasts during meniscal reparative change.
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http://dx.doi.org/10.1080/03008207.2017.1284825DOI Listing
November 2017

Inducible nitric oxide synthase plays important roles in allergic reactions of pollinosis in mice sensitized with pollen allergy.

J Clin Biochem Nutr 2013 Jan 24;52(1):17-21. Epub 2012 Oct 24.

Department of Dermatology, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno, Osaka 545-8585, Japan ; Brain Science Institute, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno, Osaka 545-8585, Japan.

To elucidate the possible involvement of nitric oxide (NO) derived from inducible NO-synthase (iNOS) in the pathogenesis of patients with allergic rhinitis, we analyzed changes in the frequency of sneezing, plasma levels of NO metabolites, α-melanocyte-stimulating hormone (MSH) and immunoglobulin E and tracheal expression of IgA and mast cell tryptase in control and iNOS(-/-) mice. Eight-week-old control and iNOS(-/-) male C57BL/6j mice were sensitized with Cry j I antigen. After the last intranasal challenge of antigen, changes in the frequency of sneezing and plasma levels of IgE, α-MSH and NO metabolites and tracheal expression of iNOS, IgA and mast cell tryptase were analyzed by ELISA and immunohistochemistry using specific antibodies. The sensitization of mice with Cry j I antigen increased plasma levels of NO metabolites, α-MSH and IgE and tracheal expression of iNOS, IgA and mast cell tryptase in control not but in iNOS(-/-) mice. Administration of N(G)-nitro-L-arginine methyl ester strongly inhibited all these changes occurred in control mice. These results indicate that the symptom of pollinosis including sneezing is enhanced by iNOS derived NO through activation of α-MSH-receptor containing mast cells enriched with tryptase.
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http://dx.doi.org/10.3164/jcbn.12-52DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3541413PMC
January 2013

Inducible nitric oxide synthase (iNOS) and α-melanocyte-stimulating hormones of iNOS origin play important roles in the allergic reactions of atopic dermatitis in mice.

Exp Dermatol 2011 Nov 6;20(11):911-4. Epub 2011 Sep 6.

Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, Osaka, Japan.

To elucidate the possible involvement of nitric oxide (NO) derived from inducible NO synthase (iNOS) in the pathogenesis of patients with allergic rhinitis, we used an animal model of atopic dermatitis (AD) induced by epicutaneous sensitization and analysed the differences in ear thickness, the frequency of scratching and plasma levels of ovalbumin-specific immunoglobulin E (OVA-IgE), transforming growth factor (TGF)-β, tumor necrosis factor (TNF)-α, adrenocorticotropic hormone (ACTH) and α-melanocyte-stimulating hormone (α-MSH) between control and iNOS(-/-) mice. Eight-week-old control and iNOS(-/-) male C57BL/6j mice were sensitized three times with OVA antigen. Before and after the last skin sensitization, the number of scratching incidents and the thickness of the ear were examined, and the plasma levels of OVA-IgE, α-MSH, ACTH, TGF-β and TNF-α were analysed by ELISA. Sensitization of mice with OVA resulted in increased plasma levels of OVA-IgE, α-MSH, ACTH, TGF-β and TNF-α in control, but not in iNOS(-/-) mice. The administration of l-nitro-arginine-methyl ester (l-NAME) abolished all the above changes that occurred in the control mice. In addition, iNOS(-/-) mice given α-MSH exhibited a change similar to that seen in the control, whereas iNOS(-/-) mice given ACTH, TGF-β or TNF-α did not demonstrate any changes. These results indicate that symptoms of AD such as scratching can be exacerbated by α-MSH, which is induced by iNOS-derived NO.
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http://dx.doi.org/10.1111/j.1600-0625.2011.01360.xDOI Listing
November 2011

Strong exercise stress exacerbates dermatitis in atopic model mice, NC/Nga mice, while proper exercise reduces it.

Exp Dermatol 2010 Dec 7;19(12):1067-72. Epub 2010 Sep 7.

Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, Abeno, Osaka, Japan.

Atopic dermatitis is well known to exacerbate by stress. How the influence of exercise stress on the skin symptoms in patients with atopic dermatitis has not been clarified. The purpose of our research is to investigate how different strength of exercise stress acts on atopic dermatitis. Specific pathogen-free (SPF) and conventional NC/Nga male mice were used for the experiments. Conventional mice but not SPF group spontaneously develop dermal symptom similar to that of patients with atopic dermatitis at their age of 7 weeks. They were given two types of stress, mild (20 m/min for 60 min) or strong exercise (25 m/min for 90 min), using a treadmill four times per day. The dermal symptom of the conventional group was strongly exacerbated by strong exercise but ameliorated by mild exercise. Under the standard experimental conditions, plasma concentrations of α-melanocyte-stimulating hormone (α-MSH), transforming growth factor-β (TGF-β) and substance P in conventional mice increased markedly with concomitant exacerbation of the symptom. The plasma concentrations of these proteins elevated after strong exercise but decreased after mild exercise. Under the conventional conditions, plasma levels of β-endorphin increased with time by some mechanisms enhanced by the mild exercise. These observations suggested that exercise-induced stress significantly affect the symptom of atopic dermatitis in a pivotal manner depending on the plasma levels of TGF-β, α-MSH, substance P and β-endorphin.
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http://dx.doi.org/10.1111/j.1600-0625.2010.01130.xDOI Listing
December 2010

Plasma cluster ions decrease the antigenicity of mite allergens and suppress atopic dermatitis in NC/Nga mice.

Arch Dermatol Res 2011 Jul 17;303(5):367-70. Epub 2010 Nov 17.

Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, Asahimachi, Abeno, Japan.

Mite antigens play important roles in the pathogenesis of atopic dermatitis (AD). We recently developed a novel air cleaner (KC-850U) using charged plasma cluster ions to eliminate a variety of allergens from house environments. The present work demonstrates the ability of KC-850U to decrease the symptoms of AD induced by mite allergens. Pooled sera from the conventional NC/Nga mice, and AD model animals, were incubated with varying concentrations of the control and KC-850U-pretreated allergens extracted from mite. The incubated mixtures were transferred to wells coated with intact allergens and subjected to ELISA to measure the amounts of immunoglobulin E (IgE) bound to the wells. Kinetic analysis revealed that exposure of mite extracts to plasma cluster ions destructed about 95% of the epitopes of the allergens. The specific pathogen-free and conventional mice were housed in rooms equipped with either KC-850U or a standard air cleaner and observed their dermal symptom for 2 weeks. Dermatological examination revealed the AD symptom of the conventional mice housed in a room equipped with an air cleaner. In contrast, the symptoms which became apparent during the experiments were suppressed remarkably exposing mice to plasma cluster ions. These observations suggested that plasma cluster ions generated by KC-850U destroyed the epitopes of mite allergens and suppressed the symptoms of AD in the mice.
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http://dx.doi.org/10.1007/s00403-010-1095-7DOI Listing
July 2011

Prevention of scattered light-induced asthenopia and fatigue by a polarized filter.

Photodermatol Photoimmunol Photomed 2010 Apr;26(2):89-92

Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, Abeno, Osaka, Japan.

Background: It has been well documented that a long-time irradiation of the eye by a strong light elicits eyestrain and fatigue. To elucidate the mechanism for the induction of light-induced fatigue and asthenopia, changes in the mouse were analyzed after white light-irradiation to the eye.

Methods: C57BL/6j male mice were irradiated with white light in a specially designed room equipped with four mirrors covering all areas of its four walls to elicit diffused reflected light, and changes in their plasma levels of cortisol, INF-gamma, interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta) were analyzed.

Results: Irradiation of mice with scattered white light significantly decreased the motional activity of animals, suggesting the occurrence of fatigue. Biochemical analysis and enzyme-immunoassay revealed that the irradiation of mice significantly elevated the plasma levels of cortisol, IFN-gamma, IL-10 and TGF-beta. All these changes were not observed with mice irradiated with the light in a similar room not equipped with mirrors. These changes were successfully inhibited by a polarized glass filter but not by a non-polarized filter with a similar absorbance.

Conclusions: These observations suggest that irradiation of the eye by scattered reflected light stimulated a stress response via hypothalamo-pituitary-adrenal axis to enhance the secretion of cortisol from the adrenal grand and increase the plasma levels of cytokines.
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http://dx.doi.org/10.1111/j.1600-0781.2010.00497.xDOI Listing
April 2010

Alpha-melanocyte-stimulating hormone plays an important role in the onset of pollinosis in a pollen allergy mouse model.

Int Arch Allergy Immunol 2010 30;153(1):13-8. Epub 2010 Mar 30.

Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, 1-4-3 Asahimachi, Abeno, Osaka, Japan.

Background: alpha-Melanocyte-stimulating hormone (alpha-MSH) is a neuropeptide that controls melanogenesis in pigmentary cells. In addition, its potent immunomodulatory activity has been recently described in cutaneous inflammatory disorders. However the mechanism of such pollen allergies remains to be elucidated. The purpose of this study was to investigate the role of alpha-MSH in a murine model of pollen allergy.

Methods: Eight-week-old male BDF-1 mice were sensitized with Cry j I. After the last intranasal antigen, the number of sneezes was counted for 5 min. In addition, the serum levels of IgE and neuronal hormones were measured by ELISA. The expression of IgA, melanocortin receptor 1 (MC1R) and MC5R in the trachea were also observed by immunohistochemistry.

Results: Both the concentration of alpha-MSH and adrenocorticotropin in plasma increase in pollen allergy model mice. Furthermore, MC5R increased in the trachea; however, MC1R did not increase in the trachea. In addition, the changes in sneezing and IgA expression in the pollen allergy model mice were suppressed by alpha-MSH antibody treatment, but they remained unchanged after MC1R antagonist (agouti) treatment.

Conclusions: These results indicate that sneezing due to pollen allergy is associated with an increased concentration of alpha-MSH and the expression of MC5R.
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http://dx.doi.org/10.1159/000301574DOI Listing
September 2010

Dynamic aspects of ascorbic acid metabolism in the circulation: analysis by ascorbate oxidase with a prolonged in vivo half-life.

Biochem J 2009 Jun 26;421(2):293-9. Epub 2009 Jun 26.

Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, 1-4-3 Asahimachi, Abeno, Osaka 545-8585, Japan.

Because AA (L-ascorbic acid) scavenges various types of free radicals to form MDAA (monodehydroascorbic acid) and DAA (dehydroascorbic acid), its regeneration from the oxidized metabolites is critically important for humans and other animals that lack the ability to synthesize this antioxidant. To study the dynamic aspects of AA metabolism in the circulation, a long acting AOase (ascorbate oxidase) derivative was synthesized by covalently linking PEG [poly(ethylene glycol)] to the enzyme. Fairly low concentrations of the modified enzyme (PEG-AOase) rapidly decreased AA levels in isolated fresh plasma and blood samples with a concomitant increase in their levels of MDAA and DAA. In contrast, relatively high doses of PEG-AOase were required to decrease the circulating plasma AA levels of both normal rats and ODS (osteogenic disorder Shionogi) rats that lack the ability to synthesize AA. Administration of 50 units of PEG-AOase/kg of body weight rapidly decreased AA levels in plasma and the kidney without affecting the levels in other tissues, such as the liver, brain, lung, adrenal grand and skeletal muscles. PEG-AOase slightly, but significantly, decreased glutathione (GSH) levels in the liver without affecting those in other tissues. Suppression of hepatic synthesis of GSH by administration of BSO [L-buthionin-(S,R)-sulfoximine] enhanced the PEG-AOase-induced decrease in plasma AA levels. These and other results suggest that the circulating AA is reductively regenerated from MDAA extremely rapidly and that hepatic GSH plays important roles in the regeneration of this antioxidant.
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http://dx.doi.org/10.1042/BJ20090286DOI Listing
June 2009