Epstein-barr virus infections induce aberrant osteoclastogenesis in immune system-humanized NOD/Shi-scid/IL-2RγCnull mice.

Epstein-Barr virus (EBV) and other viral infections are possible triggers of autoimmune diseases, such as rheumatoid arthritis (RA). To analyze the causative relationship between EBV infections and RA development, we performed experiment on humanized NOD/Shi-scid/IL-2RγCnull (hu-NOG) mice reconstituted human immune system components and infected with EBV. In EBV-infected hu-NOG mice, breakdown of knee joint bones was found to be accompanied by the accumulation of receptor activator of nuclear factor-κB (NF-κB) (RANK) ligand (RANKL), a key factor in osteoclastogenesis, human CD19 and EBV-encoded small RNA (EBER)-bearing cells. Accumulation of these cells expanded in the bone marrow adjacent to the bone breakage, showing a histological feature like to that in bone marrow edema. On the other hand, human RANK/human matrix metalloprotease-9 (MMP-9) positive, osteoclast-like cells were found at broken bone portion of EBV-infected mouse knee joint. In addition, human macrophage-colony stimulating factor (M-CSF), an essential factor in development of osteoclasts, evidently expressed in spleen and bone marrow of EBV-infected humanized mice. Furthermore, RANKL and M-CSF were identified at certain period of EBV-transformed B lymphoblastoid cells (BLBCs) derived from umbilical cord blood lymphocytes. Co-culturing bone marrow cells of hu-NOG mice with EBV-transformed BLBCs resulted in the induction of a multinucleated cell population positive for tartrate-resistant acid phosphatase and human MMP-9 which indicating human osteoclast-like cells. These findings suggest that EBV-infected BLBCs induce human aberrant osteoclastogenesis, which cause erosive arthritis in the joints.

Are Viral Infections Key Inducers of Autoimmune Diseases? Focus on Epstein-Barr Virus.

It is generally accepted that certain viral infections can trigger the development of autoimmune diseases. However, the exact mechanisms by which these viruses induce autoimmunity are still not understood. In this review, we first describe hypothetical mechanisms by which viruses induce some representative autoimmune diseases. Then, we focus on Epstein-Barr virus (EBV) and discuss its role in the pathogenesis of rheumatoid arthritis (RA). The discussion is mainly based on our own previous findings that (A) EBV DNA and its products EBV-encoded small RNA (EBER) and latent membrane protein 1 (LMP1) are present in the synovial lesions of RA, (B) mRNA expression of the signaling lymphocytic activation molecule-associated protein (SAP)/SH2D1A gene that plays a critical role in cellular immune responses to EBV is reduced in the peripheral T cells of patients with RA, and (C) EBV infection of mice reconstituted with human immune system components (humanized mice) induced erosive arthritis that is pathologically similar to RA. Additionally, environmental factors may contribute to EBV reactivation as follows: Porphyromonas gingivalis peptidylarginine deiminase (PAD), an enzyme required for citrullination, engenders antigens leading to the production of citrullinated peptides both in the gingiva and synovium. Anti-citrullinated peptides autoantibody is an important marker for diagnosis and disease activity of RA. These findings, as well as various results obtained by other researchers, strongly suggest that EBV is directly involved in the pathogenesis of RA, a typical autoimmune disease.

Antinuclear antibodies produced in HLA-DR transgenic humanized mice developed chronic graft-versus-host disease.

BACKGROUND: Chronic graft versus host disease (GVHD) has been reported in humanized mice after the implantation of human hematopoietic stem cells (hu-HSC). As such, humanized mice have been applied to a mouse model of chronic GVHD; however, B-cell activation and autoantibody production did not occur, and the clinical features of chronic GVHD were not sufficiently reproduced. The purpose of this study was to establish an improved humanized mouse model with chronic GVHD using HLA-DR transgenic NOD/Shi-scid, IL-2RγKO (NOG) mice. METHODS: CD34-positive cells were isolated from blood extracted from HLA-DRB1∗0405-positive umbilical cords using magnetic cell isolation. Then these were transplanted into NOG-Iab KO, HLA-DR 0405 Tg mice aged 8-16 weeks. GVHD symptoms were observed 26 weeks after transplantation. Histological findings of the skin, lung, liver, and spleen were compared with those of non-humanized mice. Antinuclear antibodies (ANA) were measured by indirect immunofluorescence using sera isolated 26 weeks after transplantation. RESULTS: Although GVHD symptoms were not observed in humanized (hu-HSC) NOG-Iab KO, HLA-DR 0405 Tg mice during the observation period, histological findings of human T-cell infiltration were observed in the skin, liver, and lung, suggesting that GVDH was present; human tingible body macrophages or clusters of BCL-6-positive human B-cells were observed in the spleen. Furthermore, human IgG ANA with peripheral or homogeneous staining patterns were also detected in the sera. CONCLUSION: Hu-HSC NOG-Iab KO, HLA-DR 0405 Tg mice differed from conventional models in terms of B-cell activation and ANA production. This study is the first to report on B-cell activation and autoantibody production in humanized mice with chronic GVHD, suggesting that hu-HSC NOG-Iab KO, HLA-DR 0405 Tg mice could be applied to a new humanized mouse model of chronic GVHD.

Comprehensive treatment for severe periodontitis with pathologic tooth migration-related bimaxillary protrusion: A case report with 3-year follow-up.

BACKGROUND AND OVERVIEW: Patients with severe periodontitis often experience pathologic tooth migration (PTM), which impairs esthetics and leads to occlusal disharmony (for example, premature contacts and traumatic occlusion) that can further exacerbate periodontitis. The authors describe a patient who exhibited severe periodontitis with PTM-related bimaxillary protrusion. This report includes 3-year clinical outcomes after periodontal regenerative therapy, implant-anchored orthodontic therapy, and implant prosthodontics intended to achieve both functional and esthetic improvements. CASE DESCRIPTION: A 63-year-old woman sought treatment with the chief complaint of maxillary anterior tooth mobility. Clinical examination revealed excessive tooth mobility, deep periodontal pockets, and infrabony defects in all teeth. All teeth exhibited PTM; the mandibular anterior teeth exhibited marked protrusion caused by the progression of periodontitis. After initial periodontal therapy, periodontal regenerative therapy was performed in all molar regions. At 6 and 9 months postoperatively, comprehensive orthodontic treatment was initiated for the mandible and maxilla, respectively, using orthodontic anchorage devices to achieve acceptable functional occlusion. After orthodontic treatment, staged guided bone regeneration was performed and dental implants were placed in the severely resorbed maxillary anterior ridge. This comprehensive treatment yielded favorable periodontal conditions, stable occlusion, and good esthetic outcomes. CONCLUSIONS AND PRACTICAL IMPLICATIONS: Favorable esthetics, stable occlusion, and highly cleansable periodontal tissues were achieved with well-planned interdisciplinary and comprehensive treatment, although the patient had severe periodontitis and PTM-related bimaxillary protrusion.

Human osteoclastogenesis in Epstein-Barr virus-induced erosive arthritis in humanized NOD/Shi-scid/IL-2Rγnull mice.

Many human viruses, including Epstein-Barr virus (EBV), do not infect mice, which is challenging for biomedical research. We have previously reported that EBV infection induces erosive arthritis, which histologically resembles rheumatoid arthritis, in humanized NOD/Shi-scid/IL-2Rγnull (hu-NOG) mice; however, the underlying mechanisms are not known. Osteoclast-like multinucleated cells were observed during bone erosion in this mouse model, and therefore, we aimed to determine whether the human or mouse immune system activated bone erosion and analyzed the characteristics and origin of the multinucleated cells in hu-NOG mice. Sections of the mice knee joint tissues were immunostained with anti-human antibodies against certain osteoclast markers, including cathepsin K and matrix metalloproteinase-9 (MMP-9). Multinucleated cells observed during bone erosion stained positively for human cathepsin K and MMP-9. These results indicate that human osteoclasts primarily induce erosive arthritis during EBV infections. Human osteoclast development from hematopoietic stem cells transplanted in hu-NOG mice remains unclear. To confirm their differentiation potential into human osteoclasts, we cultured bone marrow cells of EBV-infected hu-NOG mice and analyzed their characteristics. Multinucleated cells cultured from the bone marrow cells stained positive for human cathepsin K and human MMP-9, indicating that bone marrow cells of hu-NOG mice could differentiate from human osteoclast progenitor cells into human osteoclasts. These results indicate that the human immune response to EBV infection may induce human osteoclast activation and cause erosive arthritis in this mouse model. Moreover, this study is the first, to our knowledge, to demonstrate human osteoclastogenesis in humanized mice. We consider that this model is useful for studying associations of EBV infections with rheumatoid arthritis and human bone metabolism.

Interdisciplinary therapy for severe periodontitis with Angle Class II Division 1 malocclusion: A case report with 7-year follow-up.

BACKGROUND AND OVERVIEW: Previous studies have suggested that occlusal discrepancy is a risk factor contributing to periodontal disease. Occlusal discrepancy could increase the risk of developing infrabony defects. The authors present a case of a patient with severe periodontitis who exhibited many infrabony defects in the molar region due to malocclusion-induced trauma. They report the 7-year treatment outcomes of the patient after periodontal regenerative and comprehensive orthodontic therapies for functional recovery with implant prosthodontics. CASE DESCRIPTION: A 56-year-old woman sought treatment with the chief symptom of masticatory disturbance. In the molar region, excessive tooth mobility, deep periodontal pockets, and infrabony defects were observed. She had excessive overjet, resulting in collapse of anterior guidance. Malocclusion was considered to be an exacerbating factor of the infrabony defects. After initial periodontal therapy, the authors performed periodontal regenerative therapy in the mandibular molar regions. The authors carefully placed implants in a position in the maxillary molar region that would ensure an appropriate anterior dental relationship after orthodontic treatment. Comprehensive orthodontic treatment was subsequently performed, using implants as anchoring units. Definitive surgery was then performed on the mandibular molars before placing the final prosthesis. Favorable periodontal condition and stable occlusion have been maintained for the 7-year posttreatment period. CONCLUSIONS AND PRACTICAL IMPLICATIONS: Comprehensive and interdisciplinary treatment enables stable occlusion and establishment of periodontal and peri-implant tissues with high cleansability, even in patients with severe periodontitis and malocclusion. In this case, a favorable long-term treatment outcome can be expected.

Efficacy and safety of minodronic acid hydrate in patients with steroid-induced osteoporosis.

OBJECTIVES: Minodronic acid hydrate, an oral bisphosphonate, has a greater inhibitory effect on bone resorption than do other approved drugs; however, this has been studied only in patients with primary osteoporosis. Here, we administered minodronic acid hydrate to patients with steroid-induced osteoporosis who have been treated with steroids for rheumatoid arthritis or other collagen diseases, and the efficacy and safety of minodronic acid hydrate were prospectively investigated. METHODS: Twenty-five patients treated in our rheumatology clinic received minodronic acid hydrate 1 mg/day. The changes in bone mineral density (BMD) and bone turnover markers were investigated at 3 and 6 months, and adverse events, including the presence or absence of an incident osteoporotic fracture, were examined over a period of 6 months. RESULTS: Percent changes in BMD of the lumbar spine and femur significantly increased. The values of bone turnover markers significantly decreased. There were no patients with a radiographically apparent incident fracture. Adverse events included toothache for which the patient discontinued the treatment and three cases of gastrointestinal disorder that did not lead to discontinuation, and thus minodronic acid hydrate was well tolerated. CONCLUSIONS: Here, we show that minodronic acid hydrate is effectively and safely used for treatment of steroid-induced osteoporosis.

Ocular Behçet's disease is less complicated with allergic disorders. A nationwide survey in Japan.

OBJECTIVES: Behçet's disease (BD) is a systemic inflammatory disorder polarised to the Th1 and Th17 immune systems. Allergic diseases are polarised to the Th2 immune system. The aim of the present study is to investigate the prevalence of allergic diseases in patients who have BD. METHODS: The study involved a large-scale interview survey of Japanese patients with BD at 21 institutes of ophthalmology; 353 patients (255 males and 98 females) were recruited for this study. We analysed the history of allergic diseases such as atopic dermatitis (AD), allergic rhinitis (AR), bronchial asthma (BA) and drug/food allergies (FA). RESULTS: Oral aphthous ulcers, ocular lesions, skin lesions, genital ulcers, arthritis, neurological lesions, intestinal lesions, deep vein thrombosis and epididymitis were reported in 95.8%, 98.6%, 72.5%, 44.8%, 13.9%, 6.8%, 6.2%, 3.7% and 1.4% of the patients, respectively. It was also reported that 73 patients (20.7%) had histories of allergic diseases: AD (5 cases, 1.4%), AR (36 cases, 10.2%), BA (19 cases, 5.4%) and FA (30 cases, 8.5%). This percentage was significantly lower than in a survey that Japan's Ministry of Health, Labour and Welfare conducted for healthy population (47.6%) (odds ratio = 0.29, 95% confidence interval = 0.22-0.38, p=4.9×10-22). Frequencies of posterior/pan-uveitis, relatively severe ocular findings, and visual prognosis were not affected by a history of allergic diseases in BD. CONCLUSIONS: Patients with BD had fewer complications from allergic diseases than did the entire population of Japan.

Long-term stability of maxillary group distalization with interradicular miniscrews in a patient with a Class II Division 2 malocclusion.

We successfully treated a Class II Division 2 patient with maxillary group distalization using interradicular miniscrews. A woman, aged 28 years 11 months, had a convex profile and an excessive overjet caused by a skeletal Class II jaw-base relationship. After leveling and alignment, titanium miniscrews were obliquely implanted between the maxillary second premolar and first molar. To distalize the maxillary dentition, nickel-titanium closing coil springs with a 2-N load were placed between the screws and the hooks on the archwire. After 28 months of active orthodontic treatment, a proper facial profile and an acceptable occlusion were achieved with a 4-mm distalization of the maxillary dentition. The resultant occlusion was stable throughout a 5-year retention period. Interradicular miniscrews were useful to distalize the maxillary dentition for correcting a Class II malocclusion. This new strategy, group distalization with miniscrews, can make the treatment simpler with greater predictability.

Interdisciplinary treatment of a nonsyndromic oligodontia patient with implant-anchored orthodontics.

We successfully treated a nonsyndromic oligodontia patient with implant-anchored orthodontics and prosthetic restorations. A woman, age 18 years 11 months, had a straight profile and a skeletal Class I jaw-base relationship but had spaced arches because of 7 congenitally missing teeth. After leveling and alignment of the dentition, a titanium miniscrew was temporarily placed at the distal alveolus of the mandibular right first premolar, and the posterior teeth were mesialized to reduce the restorative spaces. After determination of the incisor positions, 3 dental implants were respectively inserted at the sites of the maxillary canines and the mandibular left lateral incisor with guided bone regeneration procedures. Then, screw-retained temporary prostheses were delivered after subepithelial connective tissue grafting and used for molar mesialization as absolute anchorage. After 36 months of active orthodontic treatment, an acceptable occlusion was achieved, both functionally and esthetically, with the 3 dental implants. The maxillary and mandibular molars were mesialized, but the changes of incisor position were minimal. As a result, a proper facial profile was maintained, and an attractive smile was achieved. The resultant occlusion was stable throughout a 3-year retention period. In conclusion, interdisciplinary treatment combined with orthodontics, implant surgery, and prosthodontics was useful for a nonsyndromic oligodontia patient. Especially, the new strategy-implant-anchored orthodontics-can facilitate the treatment more simply with greater predictability.

[Tumor necrosis factor-alpha in tears of patients with Sjögren syndrome].

PURPOSE: To determine the pathological role of tumor necrosis factor alpha (TNF-alpha) in ocular surface disorder of patients with Sjögren syndrome (SS), TNF-alpha in the tears of patients with SS were analyzed. In addition, the relationship between TNF-alpha levels in the tear samples and the severity of the corneal epithelial disorder was evaluated. MATERIAL AND METHODS: Tear samples were obtained from the 60 eyes of 30 patients with primary SS and the 60 eyes of 30 normal subjects. Tear samples were analyzed using a sandwich enzyme-linked immunosorbent assay to detect TNF-alpha. Corneal epithelial cell damage was examined with a slit-lamp biomicroscope and scored by the van Bijsterveld scoring system. RESULTS: TNF-alpha was detected in the tears of patients with SS at concentrations ranging from 8 to 1,500 pg/ml in 33 of the 60 eyes of the patients. In contrast, TNF-alpha was not detected in the tears of normal controls. The concentration of TNF-alpha was significantly higher in the tear samples from patients whose clinical scores were two or three points compared with patients whose scores were zero or one point (Mann-Whitney U test: p < 0.03). CONCLUSIONS: TNF-alpha was detected in the tears of patients with SS, and tear TNF-alpha levels showed a significant correlation with the grade of corneal epithelial damage, suggesting that TNF-alpha is a potent mediator in keratoconjunctivitis sicca.

[A case of multiple sclerosis with incongruous homonymous hemianopia due to lateral geniculate body lesion].

We report a rare case of a 31-year-old woman who was diagnosed as multiple sclerosis with homonymous hemianopia due to a demyelinative lesion in the lateral geniculate body. Paresthesia appeared in the distal parts of both legs from September, 2002. She complained of loss of visual field in both eyes 2 months later. Since a left asymmetric wedge-shaped homonymous hemianopia was detected by Octopus automatic perimetry, she was referred to our hospital on December 2, 2002. Neurological examination showed subjective paresthesia in the distal parts of both legs in addition to Lhermitte's sign. Brain MRI showed demyelinated lesions in the lateral geniculate body on the right and periventricular areas of the lateral ventricles. Spinal MRI revealed a demyelinated lesion in the upper cervical cord. She was diagnosed as multiple sclerosis and underwent a steroid pulse therapy. Homonymous hemianopia resolved a few days later.

Apoptosis of murine cultured biliary epithelial cells induced by glycochenodeoxycholic acid involves Fas receptor and its ligand.

Fas-mediated apoptosis of biliary epithelial cells (BECs) is suggested as a main effector process in immune-mediated biliary diseases. Glycochenodeoxycholic acid (GCDCA), a conjugated hydrophobic bile acid, is known to cause apoptosis of hepatocytes via the direct activation of Fas receptor (FasR). In this study, we investigated the apoptotic effect of GCDCA on cultured murine intrahepatic BECs. It was found that GCDCA induced apoptosis of BECs derived from BALB/c mice in a dose- and incubation time-dependent manner. The morphology and TUNEL positivity for the apoptotic process induced by GCDCA were similar to those induced by beauvericin (BV) which is known to cause apoptosis by a direct activation of caspase-3, a member of the caspase family, at its downstream. The GCDCA-induced apoptosis of BECs was accompanied by an up-regulation of FasR, Fas ligand (FasL), and also caspase-3 expression. GCDCA did not induce the apoptosis of BECs derived from C3H.MRL-Fas(lpr) mice possessing a non-functioning FasR, supporting that the GCDCA-induced apoptosis of BECs in BALB/c involves the Fas system. Furthermore, GCDCA induced an increase in interleukin-18 (IL-18) mRNA in BECs and secretion of activated IL-18 from BECs of BALB/c, which might have led to an up-regulation of FasL mRNA and protein expression in BECs followed by FasR/FasL interaction. These results suggest that GCDCA induces apoptosis of BECs through FasR/FasL interaction via an autocrine/paracrine effect, IL-18 is responsible for the expression of FasL in BEC, and the up-regulation of caspase-3 expression is involved in this model. This model could be useful for molecular and genetic studies of Fas-mediated apoptosis of BECs.

CD40 expression in normal human cornea and regulation of CD40 in cultured human corneal epithelial and stromal cells.

PURPOSE: To determine whether CD40-CD40 ligand (CD40L) interaction plays a role in corneal inflammatory responses, the expression of CD40 and CD40L on normal human cornea was investigated. In addition, using cultured human corneal epithelial (HCE) and human corneal stromal (HCS) cells, the regulation of CD40 expression in human corneal cells investigated, including that induced by proinflammatory cytokines such as interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha. METHODS: Frozen optimal cutting temperature (OCT) compound-embedded sections of corneal tissues obtained from 18 normal human corneas were examined by an immunoperoxidase staining technique with anti-CD40 and anti-CD40L monoclonal antibodies (mAbs). Also, cultured HCE and HCS cells, with IFN-gamma (250-1000 U/mL) or TNF-alpha (500-4000 U/mL) treatment for 1 to 4 days and with no treatment, were stained by the immunofluorescence technique with mAbs and analyzed by flow cytometry. RESULTS. The area of positive staining for CD40 showed a topographical difference. The limbal epithelial cells were predominantly positive for CD40. Positive staining was also found to a lesser extent on the cells in the basal layer of peripheral corneal epithelium. Epithelial cells of the central cornea showed no immunoreactivity for CD40. Corneal stromal cells were negative for CD40 in most of the donor tissues (positive: 5 of the 18 corneas). Endothelial cells were distinctly negative for CD40. Cultured HCE cells were also positive but decreased in positive cell number with lengthening culture period. None or less than 5% of the cultured HCS cells were CD40 positive. IFN-gamma enhanced CD40 expression on both cell types. In contrast, TNF-alpha enhanced CD40 on HCE but not on HCS cells. No component cells of normal human cornea or cultured HCE and HCS cells showed immunoreactivity for CD40L. CONCLUSIONS: In the human cornea, CD40 is expressed predominantly on limbal epithelial cells and also on cultured HCE cells with high proliferative potential. In addition, the expression of CD40 is induced on cultured HCE and HCS cells differentially by proinflammatory cytokines, such as IFN-gamma and TNF-alpha.