RESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Bone destruction plays a key role in damaging the joint function of rheumatoid arthritis (RA). Fengshi Qutong capsule (FSQTC) consisting of 19 traditional Chinese medicines has been used for treating RA in China for many years. Preliminary studies show that FSQTC has analgesic activity and inhibits synovial angiogenesis of collagen-induced arthritis (CIA), but its role on bone destruction of RA is still unclear. AIM OF THE STUDY: To explore the effect of FSQTC on bone destruction of RA and the possible mechanism of osteoclastogenesis in vivo and in vitro. MATERIALS AND METHODS: LC-MS system was used to detect the quality control components of FSQTC. The anti-arthritic effect of FSQTC on CIA rats was evaluated by arthritis score, arthritis incidence and histopathology evaluation of inflamed joints. The effect of treatment with FSQTC on bone destruction of joint tissues was determined with X-ray and micro-CT quantification, and on bone resorption marker CTX-I and formation marker osteocalcin in sera were detected by ELISA. Then, osteoclast differentiation and mature were evaluated by TRAP staining, actin ring immunofluorescence and bone resorption assay both in joints and RANKL-induced RAW264.7 cells. In addition, RANKL, OPG, IL-1ß and TNFα in sera were evaluated by ELISA. The molecular mechanisms of the inhibitions were elucidated by analyzing the protein and gene expression of osteoclastic markers CTSK, MMP-9 and ß3-Integrin, transcriptional factors c-Fos and NFATc1, as well as phosphorylation of ERK1/2, JNK and P38 in joints and in RANKL-induced RAW264.7 cells using western blot and/or qPCR. RESULTS: In this study, 12 major quality control components were identified. Our data showed that FSQTC significantly increased bone mineral density, volume fraction, trabecular thickness, and decreased trabecular separation of inflamed joints both at periarticular and extra-articular locations in CIA rats. FSQTC also diminished the level of CTX-I and simultaneously increased osteocalcin in sera of CIA rats. The effects were accompanied by reductions of osteoclast differentiation, bone resorption, and expression of osteoclastic markers (CTSK, MMP-9 and ß3-Integrin) in joints. Interestingly, FSQTC treatment could reduce the protein level of RANKL, increase the expression of OPG, and decrease the ratio of RANKL to OPG in inflamed joints and sera of CIA rats. In addition, FSQTC inhibited the levels of pro-inflammatory cytokines implicated in bone resorption, such as IL-1ß and TNFα in sera. When RAW264.7 cells were treated with RANKL, FSQTC inhibited the formation of TRAP + multinucleated cells, actin ring and the bone-resorbing activity in dose-dependent manners. Furthermore, FSQTC reduced the RANKL-induced expression of osteoclastic genes and proteins and transcriptional factors (c-Fos and NFATc1), as well as phosphorylation of mitogen-activated protein kinases (MAPKs). CONCLUSION: FSQTC may inhibit bone destruction of RA by its anti-osteoclastogenic activity both in vivo and in vitro.
Assuntos
Antirreumáticos/farmacologia , Artrite Reumatoide , Densidade Óssea/efeitos dos fármacos , Reabsorção Óssea , Medicamentos de Ervas Chinesas/farmacologia , Ligante RANK/análise , Animais , Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/imunologia , Colágeno Tipo I/sangue , Citocinas/análise , Citocinas/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Medicina Tradicional Chinesa/métodos , Camundongos , Osteocalcina , Osteogênese/efeitos dos fármacos , Fragmentos de Peptídeos/sangue , Células RAW 264.7 , RatosRESUMO
BACKGROUND: Vitamin D (VitD) and calcium (Ca) supplementation attenuates antiretroviral therapy (ART)-associated bone loss, but it is unclear whether this effect is mediated through immunomodulation. METHODS: In this exploratory analysis of A5280, a 48-week, randomized, double-blind, placebo-controlled study of VitD/Ca supplementation with ART initiation, we characterized lymphocyte phenotypes and receptor activator of nuclear factor kappa-B ligand (RANKL) expression by median fluorescence intensity (MFI) at baseline and 48 weeks. Changes were evaluated within and between treatment groups by Wilcoxon signed rank and rank sum tests, respectively. Spearman correlations estimated relationships between cellular phenotypes and bone mineral density (BMD). RESULTS: Of 165 participants enrolled, 138 had samples for cellular phenotypes (64 VitD/Ca, 74 placebo). Markers of CD4, CD8 activation (CD38+HLA-DR+) declined (all P<0.001), but did not differ between arms. There was no decline in either %T-cells (CD4 and CD8) expressing RANKL or expression of RANKL by MFI. CD4 and CD8 activation markers were not correlated with BMD at baseline (r<0.15 and P>0.09 for all), but greater declines in CD4 activation correlated with greater declines in hip and spine BMD in both arms (0.25 ≤r ≤0.37, all P<0.05). A greater decline in CD8 activation was correlated with greater declines in both hip and spine BMD in the placebo arm only (hip r=0.31, P=0.009; spine r=0.25, P=0.035). CONCLUSIONS: Reductions in T-cell activation are characteristic of ART initiation, but only correlated modestly with bone loss. VitD/Ca supplementation does not appear to mitigate bone loss through modulation of immune activation or expression of RANKL. TRIAL REGISTRATION NUMBER: NCT01403051.
Assuntos
Fármacos Anti-HIV/farmacologia , Imunomodulação , Vitamina D/metabolismo , Adulto , Fármacos Anti-HIV/uso terapêutico , Terapia Antirretroviral de Alta Atividade , Biomarcadores , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/imunologia , Contagem de Linfócito CD4 , Feminino , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , Infecções por HIV/virologia , Humanos , Imunomodulação/efeitos dos fármacos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Fatores de Tempo , Resultado do Tratamento , Vitamina D/farmacologiaRESUMO
This study evaluated the participation of CB1 and CB2 receptors in the antiresorptive effect of electroacupuncture (EA) on an experimental model of inflammatory bone loss in rats. 30 rats were divided into five groups: C (control); EP (experimental periodontitis); EA (C+ EA); EP-EA (EP+ EA in the acupoints LI4, LG11, ST36, ST44); EP - EA-sham (EP+ EA in sham acupoints). For the EP groups, a ligature was placed around the right mandibular first molars at day 1. Sessions of EA or EA-sham were assigned every other day. Animals were euthanized at day 11. Histometric analysis was performed to evaluate the percentage of bone area in the furcation area. Immunolabeling patterns in the periodontal tissues and immunofluorescent staining in the trigeminal ganglia and in the trigeminal spinal tract for CB1 and CB2 receptors were performed. It was observed increased bone loss in the furcation in the EP and EP-EA-sham groups, in comparison to the other groups (pâ¯<â¯0.05). Enhanced CB2 immunolabeling was observed in the periodontal tissues in the EP-EA group, when compared to the EP and EP-EA-sham groups (pâ¯<â¯0.05). Increased CB1 immunofluorescent staining was observed in the neural tissues in the EA treated group in comparison with the other groups (pâ¯<â¯0.05), while no expression of CB2 was observed in those regions. Our study showed that in the presence of inflammatory bone disease, EA treatment reduced bone erosion and increased the immunoexpression of CB1 in the neural tissues and CB2 in the periodontal tissues.
Assuntos
Reabsorção Óssea/imunologia , Reabsorção Óssea/terapia , Eletroacupuntura , Inflamação/patologia , Receptor CB1 de Canabinoide/imunologia , Receptor CB2 de Canabinoide/imunologia , Animais , Masculino , Periodonto/metabolismo , Ratos Wistar , Gânglio Trigeminal/metabolismoRESUMO
Cardiovascular (CV) disease and osteoporosis (OP) have become increasing challenges in the aging population and even more in patients with inflammatory rheumatic diseases, such as rheumatoid arthritis, spondyloarthropathies, and systemic lupus erythematosus. In this review, we discuss how the epidemiology and pathogenesis of CV events and OP are overlapping. Smoking, diabetes mellitus, physical inactivity as conventional risk factors as well as systemic inflammation are among the modifiable risk factors for both CV events and bone loss. In rheumatic patients, systemic "high-grade" inflammation may be the primary driver of accelerated atherogenesis and bone resorption. In the general population, in which some individuals might have low-grade systemic inflammation, a holistic approach to drug treatment and lifestyle modifications may have beneficial effects on the bone as well as the vasculature. In rheumatic patients with accelerated inflammatory atherosclerosis and bone loss, the rapid and effective suppression of inflammation in a treat-to-target regime, aiming at clinical remission, is necessary to effectively control comorbidities.
Assuntos
Aterosclerose/epidemiologia , Aterosclerose/terapia , Saúde Holística , Osteoporose/epidemiologia , Osteoporose/terapia , Artrite Reumatoide/epidemiologia , Artrite Reumatoide/imunologia , Artrite Reumatoide/terapia , Aterosclerose/imunologia , Reabsorção Óssea/epidemiologia , Reabsorção Óssea/imunologia , Reabsorção Óssea/terapia , Saúde Holística/tendências , Humanos , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/imunologia , Osteoporose/imunologiaRESUMO
Skeletal health consequences associated with inflammatory diseases of the airways significantly contribute to morbidity. Sex differences have been described independently for lung and bone diseases. Repetitive inhalant exposure to lipopolysaccharide (LPS) induces bone loss and deterioration in male mice, but comparison effects in females are unknown. Using an intranasal inhalation exposure model, 8-week-old C57BL/6 male and female mice were treated daily with LPS (100 ng) or saline for 3 weeks. Bronchoalveolar lavage fluids, lung tissues, tibias, bone marrow cells, and blood were collected. LPS-induced airway neutrophil influx, interleukin (IL)-6 and neutrophil chemoattractant levels, and bronchiolar inflammation were exaggerated in male animals as compared to female mice. Trabecular bone micro-CT imaging and analysis of the proximal tibia were conducted. Inhalant LPS exposures lead to deterioration of bone quality only in male mice (not females) marked by decreased bone mineral density, bone volume/tissue volume ratio, trabecular thickness and number, and increased bone surface-to-bone volume ratio. Serum pentraxin-2 levels were modulated by sex differences and LPS exposure. In proof-of-concept studies, ovarectomized female mice demonstrated LPS-induced bone deterioration, and estradiol supplementation of ovarectomized female mice and control male mice protected against LPS-induced bone deterioration findings. Collectively, sex-specific differences exist in LPS-induced airway inflammatory consequences with significant differences found in bone quantity and quality parameters. Male mice demonstrated susceptibility to bone loss and female animals were protected, which was modulated by estrogen. Therefore, sex differences influence the biologic response in the lung-bone inflammatory axis in response to inhalant LPS exposures.
Assuntos
Reabsorção Óssea/imunologia , Osso e Ossos/imunologia , Terapia de Reposição Hormonal , Inflamação/imunologia , Pulmão/imunologia , Animais , Reabsorção Óssea/tratamento farmacológico , Estradiol/uso terapêutico , Feminino , Inflamação/tratamento farmacológico , Exposição por Inalação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ovariectomia , Sexo , Tomografia Computadorizada por Raios XRESUMO
Experimental autoimmune myasthenia gravis (EAMG), an animal model of myasthenia gravis (MG), can be induced in C57BL/6 (B6, H-2 b) mice by 2-3 injections with Torpedo californica AChR (tAChR) in complete Freund's adjuvant. Some EAMG mice exhibit weight loss with muscle weakness. The loss in body weight, which is closely associated with bone structure, is particularly evident in EAMG mice with severe muscle weakness. However, the relationship between muscle weakness and bone loss in EAMG has not been studied before. Recent investigations on bone have shed light on association of bone health and immunological states. It is possible that muscle weakness in EAMG developed by anti-tAChR immune responses might accompany bone loss. We determined whether reduced muscle strength associates with decreased bone mineral density (BMD) in EAMG mice. EAMG was induced by two injections at 4-week interval of tAChR and adjuvants in two different age groups. The first tAChR injection was either at age 8 weeks or at 15 weeks. We measured BMD at three skeletal sites, including femur, tibia, and lumbar vertebrae, using dual energy X-ray absorptiometry. Among these bone areas, femur of EAMG mice in both age groups showed a significant decrease in BMD compared to control adjuvant-injected and to non-immunized mice. Reduction in BMD in induced EAMG at a later-age appears to parallel the severity of the disease. The results indicate that anti-tAChR autoimmune response alone can reduce bone density in EAMG mice. BMD reduction was also observed in adjuvant-injected mice in comparison to normal un-injected mice, suggesting that BMD decrease can occur even when muscle activity is normal. Decreased BMD observed in both tAChR-injected and adjuvant-injected mice groups were discussed in relation to innate immunity and bone-related immunology involving activated T cells and tumour necrosis factor-related cytokines that trigger osteoclastogenesis and bone loss.
Assuntos
Densidade Óssea/imunologia , Reabsorção Óssea/patologia , Debilidade Muscular/patologia , Miastenia Gravis Autoimune Experimental/patologia , Absorciometria de Fóton , Fatores Etários , Animais , Reabsorção Óssea/induzido quimicamente , Reabsorção Óssea/diagnóstico por imagem , Reabsorção Óssea/imunologia , Fêmur/diagnóstico por imagem , Fêmur/imunologia , Fêmur/patologia , Proteínas de Peixes/administração & dosagem , Adjuvante de Freund/administração & dosagem , Vértebras Lombares/diagnóstico por imagem , Vértebras Lombares/imunologia , Vértebras Lombares/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Debilidade Muscular/induzido quimicamente , Debilidade Muscular/diagnóstico por imagem , Debilidade Muscular/imunologia , Miastenia Gravis Autoimune Experimental/induzido quimicamente , Miastenia Gravis Autoimune Experimental/diagnóstico por imagem , Miastenia Gravis Autoimune Experimental/metabolismo , Receptores Colinérgicos/administração & dosagem , Índice de Gravidade de Doença , Tíbia/diagnóstico por imagem , Tíbia/imunologia , Tíbia/patologia , Fatores de Tempo , Torpedo/metabolismoRESUMO
OBJECTIVE: Proinflammatory molecules promote osteoclast-mediated bone erosion by up-regulating local RANKL production. However, recent evidence suggests that combinations of cytokines, such as tumor necrosis factor (TNF) plus interleukin-6 (IL-6), induce RANKL-independent osteoclastogenesis. The purpose of this study was to better understand TNF/IL-6-induced osteoclast formation and to determine whether RANK is absolutely required for osteoclastogenesis and bone erosion in murine inflammatory arthritis. METHODS: Myeloid precursors from wild-type (WT) mice or mice with either germline or conditional deletion of Rank, Nfatc1, Dap12, or Fcrg were treated with either RANKL or TNF plus IL-6. Osteoprotegerin, anti-IL-6 receptor (anti-IL-6R), and hydroxyurea were used to block RANKL, the IL-6R, and cell proliferation, respectively. Clinical scoring, histologic assessment, micro-computed tomography, and quantitative polymerase chain reaction (qPCR) were used to evaluate K/BxN serum-transfer arthritis in WT and RANK-deleted mice. Loss of Rank was verified by qPCR and by osteoclast cultures. RESULTS: TNF/IL-6 generated osteoclasts in vitro that resorbed mineralized tissue through a pathway dependent on IL-6R, NFATc1, DNAX-activation protein 12, and cell proliferation, but independent of RANKL or RANK. Bone erosion and osteoclast formation were reduced, but not absent, in arthritic mice with inducible deficiency of RANK. TNF/IL-6, but not RANKL, induced osteoclast formation in bone marrow and synovial cultures from animals deficient in Rank. Multiple IL-6 family members (IL-6, leukemia inhibitory factor, oncostatin M) were up-regulated in the synovium of arthritic mice. CONCLUSION: The persistence of bone erosion and synovial osteoclasts in Rank-deficient mice, and the ability of TNF/IL-6 to induce osteoclastogenesis, suggest that more than one cytokine pathway exists to generate these bone-resorbing cells in inflamed joints.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Artrite Experimental/genética , Reabsorção Óssea/genética , Fatores de Transcrição NFATC/genética , Osteogênese/genética , Receptor Ativador de Fator Nuclear kappa-B/genética , Animais , Artrite Experimental/imunologia , Reabsorção Óssea/imunologia , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Inibidores Enzimáticos/farmacologia , Hidroxiureia/farmacologia , Técnicas In Vitro , Interleucina-6/farmacologia , Camundongos , Camundongos Knockout , Osteogênese/efeitos dos fármacos , Osteogênese/imunologia , Osteoprotegerina/farmacologia , Ligante RANK/farmacologia , Reação em Cadeia da Polimerase em Tempo Real , Receptores de IgG/genética , Receptores de Interleucina-6/antagonistas & inibidores , Fator de Necrose Tumoral alfa/farmacologia , Microtomografia por Raio-XRESUMO
BACKGROUND: Interleukin (IL)-33 is a dual cytokine with both an alarmin role and a T helper 2 cell (Th2)-like inducing effect. It is involved in the pathogenesis of rheumatoid arthritis (RA) and its models; we recently demonstrated that exogenous IL-33 could inhibit collagen-induced arthritis (CIA) in C57BL/6 mice. However, its pathophysiological role in RA is unclear. Indeed, mice deficient in the IL-33 receptor ST2 show reduced susceptibility to arthritis, and the disease is not modified in IL-33-deficient mice. We examined the immune response in wild-type (WT) and IL-33-deficient mice with CIA. To further understand the role of endogenous IL-33 in inflammatory diseases, we studied its role in a skin psoriasis model. Mice on a C57BL/6 background were deficient in IL-33 but expressed lacZ under the IL-33 promoter. Therefore, IL-33 promotor activity could be analyzed by lacZ detection and IL-33 gene expression was analyzed by X-Gal staining in various mice compartments. Frequencies of CD4(+)FoxP3(+) regulatory T cells (Tregs) and Th1 and Th17 cells were evaluated by flow cytometry in WT and IL-33(-/-) mice. Bone resorption was studied by evaluating osteoclast activity on a synthetic mineral matrix. Psoriasis-like dermatitis was induced by application of imiquimod to the skin of mice. RESULTS: Severity of CIA was similar in IL-33(-/-) and WT littermates. Joints of IL-33(-/-) mice with CIA showed IL-33 promotor activity. In mice with CIA, frequencies of Tregs, Th1 and Th17 in the spleen or lymph nodes did not differ between the genotypes; osteoclast activity was higher but not significantly in IL-33(-/-) than WT mice. Psoriasis development did not differ between the genotypes. CONCLUSIONS: Despite its expression in the synovium of arthritic mice and normal keratinocytes, IL-33 is not required for CIA development in arthritis or psoriasis. Its absence does not induce a T cell shift toward Th1, Th17 or Treg subpopulations. Altogether, these data and our previous ones, showing that exogenous IL-33 can almost completely inhibit CIA development, suggest that this cytokine is not crucial for development of chronic inflammation. Studies of RA patients are needed to determine whether treatment targeting the IL-33/ST2 axis would be effective.
Assuntos
Artrite Experimental/imunologia , Artrite Reumatoide/imunologia , Interleucina-33/imunologia , Psoríase/imunologia , Adjuvantes Imunológicos/toxicidade , Aminoquinolinas/toxicidade , Animais , Reabsorção Óssea/imunologia , Modelos Animais de Doenças , Citometria de Fluxo , Imunofluorescência , Imiquimode , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Psoríase/induzido quimicamenteRESUMO
Nutrition is an important determinant of bone health and attainment of peak bone mass. Diets containing dried plum (DP) have been shown to increase bone volume and strength. These effects may be linked to the immune system and DP-specific polyphenols. To better understand these relationships, we studied DP in skeletally mature (6-month-old) and growing (1- and 2-month-old) C57Bl/6 male mice. In adult mice, DP rapidly (<2 weeks) increased bone volume (+32%) and trabecular thickness (+24%). These changes were associated with decreased osteoclast surface (Oc.S/BS) and decreased serum CTX, a marker of bone resorption. The reduction in Oc.S/BS was associated with a reduction in the osteoclast precursor pool. Osteoblast surface (Ob.S/BS) and bone formation rate were also decreased suggesting that the gain in bone in adult mice is a consequence of diminished bone resorption and formation, but resorption is reduced more than formation. The effects of DP on bone were accompanied by a decline in interleukins, TNF and MCP-1, suggesting that DP is acting in part through the immune system to suppress inflammatory activity and reduce the size of the osteoclast precursor pool. Feeding DP was accompanied by an increase in plasma phenolics, some of which have been shown to stimulate bone accrual. In growing and young adult mice DP at levels as low as 5% of diet (w/w) increased bone volume. At higher levels (DP 25%), bone volume was increased by as much as 94%. These data demonstrate that DP feeding dramatically increases peak bone mass during growth.
Assuntos
Desenvolvimento Ósseo , Reabsorção Óssea/prevenção & controle , Citocinas/antagonistas & inibidores , Alimentos em Conserva , Frutas , Alimento Funcional , Prunus domestica , Animais , Biomarcadores/sangue , Células da Medula Óssea/citologia , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Remodelação Óssea , Reabsorção Óssea/imunologia , Reabsorção Óssea/metabolismo , Reabsorção Óssea/patologia , Osso e Ossos/citologia , Osso e Ossos/imunologia , Osso e Ossos/metabolismo , Osso e Ossos/patologia , Células Cultivadas , Colágeno Tipo I/sangue , Citocinas/sangue , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos Endogâmicos C57BL , Osteoblastos/citologia , Osteoblastos/imunologia , Osteoblastos/metabolismo , Osteoblastos/patologia , Osteoclastos/citologia , Osteoclastos/imunologia , Osteoclastos/metabolismo , Osteoclastos/patologia , Fragmentos de Peptídeos/sangue , Peptídeos/sangueRESUMO
AIM: Aconiti Lateralis Radix Preparata is a traditional Chinese medicine used to treat chronic arthritis and is highly effective against rheumatoid arthritis. However, the effects of aconine, a derivative of aconitum alkaloids, on osteoclasts, which can absorb bone, remain unknown. Here, we investigated the effects of aconine on osteoclast differentiation and bone resorption in vitro. METHODS: The viability of mouse leukemic monocyte/macrophage cell line RAW264.7 was measured using CCK-8 assays. Osteoclast differentiation was induced by incubation of RAW264.7 cells in the presence of RANKL, and assessed with TRAP staining assay. Bone resorption was examined with bone resorption pits assay. The expression of relevant genes and proteins was analyzed using RT-PCR and Western blots. The activation of NF-κB and nuclear factor of activated T-cells (NFAT) was examined using stable NF-κB and NFATc1 luciferase reporter gene systems, RT-PCR and Western blot analysis. RESULTS: Aconine (0.125, 0.25 µmol/L) did not affect the viability of RAW264.7 cells, but dose-dependently inhibited RANKL-induced osteoclast formation and bone resorptive activity. Furthermore, aconine dose-dependently inhibited the RANKL-induced activation of NF-κB and NFATc1 in RAW264.7 cells, and subsequently reduced the expression of osteoclast-specific genes (c-Src, ß3-Integrin, cathepsin K and MMP-9) and the expression of dendritic cell-specific transmembrane protein (DC-STAMP), which played an important role in cell-cell fusion. CONCLUSION: These findings suggest that aconine inhibits RANKL-induced osteoclast differentiation in RAW264.7 cells by suppressing the activation of NF-κB and NFATc1 and the expression of the cell-cell fusion molecule DC-STAMP.
Assuntos
Aconitina/análogos & derivados , Adjuvantes Imunológicos/farmacologia , Proteínas de Membrana/genética , NF-kappa B/imunologia , Fatores de Transcrição NFATC/imunologia , Proteínas do Tecido Nervoso/genética , Osteoclastos/efeitos dos fármacos , Ligante RANK/imunologia , Aconitina/farmacologia , Animais , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/genética , Reabsorção Óssea/imunologia , Diferenciação Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Camundongos , Osteoclastos/citologia , Osteoclastos/imunologia , Osteoclastos/metabolismo , Células RAW 264.7RESUMO
Coenzyme Q10 (CoQ10) is a lipid-soluble antioxidant synthesized in human body. This enzyme promotes immune system function and can be used as a dietary supplement. Rheumatoid arthritis (RA) is an autoimmune disease leading to chronic joint inflammation. RA results in severe destruction of cartilage and disability. This study aimed to investigate the effect of CoQ10 on inflammation and Th17 cell proliferation on an experimental rheumatoid arthritis (RA) mice model. CoQ10 or cotton seed oil as control was orally administrated once a day for seven weeks to mice with zymosan-induced arthritis (ZIA). Histological analysis of the joints was conducted using immunohistochemistry. Germinal center (GC) B cells, Th17 cells and Treg cells of the spleen tissue were examined by confocal microscopy staining. mRNA expression was measured by real-time PCR and protein levels were estimated by enzyme-linked immunosorbent assay (ELISA). Flow cytometric analysis (FACS) was used to evaluate Th17 cells and Treg cells. CoQ10 mitigated the severity of ZIA and decreased serum immunoglobulin concentrations. CoQ10 also reduced RANKL-induced osteoclastogenesis, inflammatory mediators and oxidant factors. Th17/Treg axis was reciprocally controlled by CoQ10 treatment. Moreover, CoQ10 treatment on normal mouse and human cells cultured in Th17 conditions decreased the number of Th17 cells and enhanced the number of Treg cells. CoQ10 alleviates arthritis in mice with ZIA declining inflammation, Th17 cells and osteoclast differentiation. These findings suggest that CoQ10 can be a potential therapeutic substance for RA.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Osteoclastos/citologia , Osteoclastos/efeitos dos fármacos , Células Th17/citologia , Células Th17/efeitos dos fármacos , Ubiquinona/análogos & derivados , Animais , Artrite Experimental/induzido quimicamente , Artrite Experimental/diagnóstico , Artrite Experimental/tratamento farmacológico , Artrite Experimental/imunologia , Autoanticorpos/imunologia , Doenças Autoimunes/diagnóstico , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/imunologia , Modelos Animais de Doenças , Centro Germinativo/citologia , Centro Germinativo/imunologia , Humanos , Imunofenotipagem , Imunossupressores/administração & dosagem , Imunossupressores/farmacologia , Interleucina-17/metabolismo , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Camundongos , Baço/citologia , Baço/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Células Th17/imunologia , Células Th17/metabolismo , Ubiquinona/administração & dosagem , Ubiquinona/farmacologia , Zimosan/efeitos adversosRESUMO
Osteoclasts together with osteoblasts play pivotal roles in bone remodeling. The unique function and ability of osteoclasts to resorb bone makes them critical in both normal bone homeostasis and pathologic bone diseases such as osteoporosis and rheumatoid arthritis. Thus, new compounds that may inhibit osteoclastogenesis and osteoclast function may be of great value in the treatment of osteoclast-related diseases. In the present study, we examined the effect of jolkinolide B (JB), isolated from the root of Euphorbia fischeriana Steud on receptor activator of NF-κB ligand (RANKL)-induced osteoclast formation. We found that JB inhibited RANKL-induced osteoclast differentiation from bone marrow macrophages (BMMs) without cytotoxicity. Furthermore, the expression of osteoclastic marker genes, such as tartrate-resistant acid phosphatase (TRAP), cathepsin K (CtsK), and calcitonin receptor (CTR), was significantly inhibited. JB inhibited RANKL-induced activation of NF-κB by suppressing RANKL-mediated IκBα degradation. Moreover, JB inhibited RANKL-induced phosphorylation of mitogen-activated protein kinases (p38, JNK, and ERK). This study thus identifies JB as an inhibitor of osteoclast formation and provides evidence that JB might be an alternative medicine for preventing and treating osteolysis.
Assuntos
Diterpenos/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , NF-kappa B/imunologia , Osteoclastos/efeitos dos fármacos , Ligante RANK/imunologia , Animais , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/imunologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Diterpenos/isolamento & purificação , Medicamentos de Ervas Chinesas/isolamento & purificação , Euphorbia/química , Camundongos , Camundongos Endogâmicos C57BL , Osteoclastos/citologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Marine organisms have bioactive potential which has tremendous pharmaceutical promise. Emerging evidence highlights the importance of the interplay between bone and the immune system of which T lymphocytes and their product act as key regulators of bone resorption. In the present investigation we have analyzed the anti-osteoporotic effect of turbo methanol extract (TME) in the reversal of bone resoprtion. Forty-two female Swiss albino mice were used and randomly assigned into sham-operated group (sham) and six ovariectomized (OVX) subgroups, i.e. OVX with vehicle (OVX) that received daily oral administration of water ad libitum; OVX with estradiol (2mg/kg/day); and OVX with different doses of TME i.e. TME 100mg/kg, TME 50mg/kg, TME 25mg/kg and TME 12.5mg/kg. Oral administration of TME or estradiol started on the second week after ovariectomy for a period of 4weeks. We observed that the administration of TME increased the trabeculation in tibia and reduced the atrophy in the uterus. TME significantly decreased the serum alkaline phosphatase (ALP) and acid phosphatase (ACP) activity in OVX mice. Micro CT analysis revealed that the TME administration preserved the bone volume, connectivity density, trabecular number, trabecular thickness and trabecular separation in OVX mice. Bone mineralization was measured in different groups of mice by Raman spectroscopy. Reversal of bone resorption was observed in TME treated group of mice. To further investigate the mechanism of action of TME, we analyzed the T lymphocyte proliferation and profiles of cytokine TNFα and sRANKL in TME treated ovariectomized mice. Decrease in the elevation of T cell subsets was observed after the supplementation with TME. The extract significantly lowered the T cell proliferation responses to mitogens, phorbol 12-myristate 13-acetate (PMA) and ionomycin (Io) and phytohemagglutinin (PHA). A marked reduction in TNFα and sRANKL secretion in serum and TNFα in cell free supernatants of activated T lymphocytes was observed upon TME administration. TME could significantly inhibit the in vitro osteoclastogenesis and the bone resorption observed using artificial calcium coated slides. Collectively, these results indicate that TME has the potential to inhibit bone resorption and may prove to be a potential candidate for the development of an anti-osteoporosis drug.
Assuntos
Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/imunologia , Misturas Complexas/uso terapêutico , Metanol/química , Moluscos/química , Linfócitos T/imunologia , Fosfatase Ácida/sangue , Fosfatase Alcalina/sangue , Animais , Biomarcadores/sangue , Reabsorção Óssea/sangue , Reabsorção Óssea/diagnóstico por imagem , Calcificação Fisiológica/efeitos dos fármacos , Cálcio/sangue , Proliferação de Células/efeitos dos fármacos , Misturas Complexas/farmacologia , Feminino , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/imunologia , Camundongos , Mitógenos/farmacologia , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Osteoclastos/patologia , Osteogênese/efeitos dos fármacos , Ovariectomia , Ligante RANK/sangue , Análise Espectral Raman , Baço/efeitos dos fármacos , Baço/patologia , Linfócitos T/efeitos dos fármacos , Tíbia/diagnóstico por imagem , Tíbia/efeitos dos fármacos , Tíbia/patologia , Fator de Necrose Tumoral alfa/sangue , Útero/efeitos dos fármacos , Útero/patologia , Microtomografia por Raio-XRESUMO
Systemic bone loss is a hallmark of rheumatoid arthritis (RA). Inflammatory cytokines such as interleukin (IL)-6 promote bone resorption by osteoclasts. Sphingosine-1-phosphate (S1P) controls the migration of osteoclast precursor cells (OCPs) between the blood and bone marrow, in part via S1P receptors (S1PR1 and S1PR2) expressed on the surface of OCPs. OCPs (CD11b(+) Gr-1(low+med) ) isolated from bone marrow of DBA/1J mice were stimulated with IL-6. S1P-directed chemotaxis of OCPs was evaluated using a transwell plate. mRNA expression of S1PR1 and S1PR2 was measured. DBA/1J mice were immunized with bovine type II collagen (days 0 and 21) and anti-mouse IL-6 receptor antibody (MR16-1) was administered on days 0 and/or 21. Trabecular bone volume was analysed using micro-computed tomography. The percentage of OCPs in tibial bone marrow and S1PR1 and S1PR2 mRNA expression in OCPs were measured. IL-6 stimulation significantly decreased S1P-directed chemotaxis of OCPs. IL-6 induced S1PR2 mRNA expression, but not S1PR1 mRNA expression, in OCPs. Bone volume was significantly lower in arthritic mice than in non-arthritic control mice on day 35. Treatment of immunized mice with MR16-1 significantly inhibited bone loss. In MR16-1-treated mice, the percentage of OCPs and expression of S1PR2 mRNA was each decreased compared with arthritic mice on day 14, but not on day 35. IL-6 increased the number of OCPs in tibial bone marrow via up-regulating S1PR2, thus playing a crucial role in systemic bone loss induced by inflammation.
Assuntos
Artrite Experimental/imunologia , Reabsorção Óssea/metabolismo , Interleucina-6/fisiologia , Osteoclastos/metabolismo , Receptores de Lisoesfingolipídeo/fisiologia , Animais , Anticorpos Monoclonais Humanizados/imunologia , Densidade Óssea/imunologia , Células da Medula Óssea , Reabsorção Óssea/imunologia , Reabsorção Óssea/prevenção & controle , Movimento Celular , Colágeno , Expressão Gênica , Inflamação/imunologia , Lisofosfolipídeos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos DBA , Osteoclastos/citologia , RNA Mensageiro/biossíntese , Receptores de Interleucina-6/antagonistas & inibidores , Receptores de Lisoesfingolipídeo/genética , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Receptores de Esfingosina-1-Fosfato , Células-Tronco/citologiaRESUMO
Bone is continuously renewed through a dynamic balance between bone resorption and formation. This process is the fundamental basis for the maintenance of normal bone mass and architecture. Osteoclasts play a crucial role in both physiological and pathological bone resorption, and receptor activator of nuclear factor-κB ligand (RANKL) is the key cytokine that induces osteoclastogenesis. Here we summarize the recent advances in the understanding of osteoclastogenic signaling by focusing on the investigation of RANKL signaling and RANKL-expressing cells in the context of osteoimmunology. The context afforded by osteoimmunology will provide a scientific basis for future therapeutic approaches to diseases related to the skeletal and immune systems.
Assuntos
Remodelação Óssea , Reabsorção Óssea/metabolismo , Osteoclastos/metabolismo , Osteoprotegerina/metabolismo , Ligante RANK/metabolismo , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Transdução de Sinais , Animais , Artrite/imunologia , Artrite/metabolismo , Reabsorção Óssea/imunologia , Feminino , Humanos , Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Masculino , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/imunologiaRESUMO
OBJECTIVE: Rheumatoid arthritis, which is associated with elevated levels of S100A8 and S100A9, is characterized by severe bone erosions caused by enhanced osteoclast formation and activity. The aim of the present study was to investigate the role of S100A8 and S100A9 in osteoclastic bone destruction in murine antigen-induced arthritis (AIA). METHODS: Bone destruction was analyzed in the arthritic knee joints of S100A9-deficient mice in which S100A8 protein expression was also lacking, and in wild-type (WT) controls. Osteoclast precursors from S100A9-deficient and WT mice were differentiated into osteoclasts in vitro. Additionally, precursors were stimulated with S100A8, S100A9, or S100A8/A9 during osteoclastogenesis. Receptor involvement was investigated using an anti-receptor for advanced glycation end products (anti-RAGE)-blocking antibody, soluble RAGE, or Toll-like receptor 4 (TLR-4)-deficient osteoclast precursors. The formation of osteoclasts and actin rings, the regulation of osteoclast markers, and bone resorption were analyzed. RESULTS: Bone erosions and cathepsin K staining were significantly suppressed in S100A9-deficient mice after AIA induction. However, osteoclast precursors from S100A9-deficient mice developed normally into functional osteoclasts, which excludes a role for intrinsic S100A8/A9. In contrast to the results observed with S100A9 and S100A8/A9, the addition of S100A8 during osteoclastogenesis resulted in stimulation of osteoclast formation in conjunction with enhanced actin ring formation and increased bone resorption. Analysis of the putative receptor for S100A8 in osteoclastogenesis revealed that osteoclast differentiation and function could not be inhibited by blocking RAGE, whereas the increase in osteoclast numbers and enhanced bone resorption were completely abrogated using TLR-4-deficient osteoclast precursors. CONCLUSION: These results demonstrate that S100A8 stimulated osteoclast formation and activity and suggest that both S100A8 and TLR-4 are important factors in mediating osteoclastic bone destruction in experimental arthritis.
Assuntos
Artrite Experimental/metabolismo , Reabsorção Óssea/metabolismo , Calgranulina A/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Artrite Experimental/genética , Artrite Experimental/imunologia , Reabsorção Óssea/genética , Reabsorção Óssea/imunologia , Osso e Ossos/imunologia , Osso e Ossos/metabolismo , Calgranulina A/genética , Calgranulina A/imunologia , Catepsina K/imunologia , Catepsina K/metabolismo , Articulação do Joelho/imunologia , Articulação do Joelho/metabolismo , Camundongos , Camundongos Knockout , Osteoclastos/imunologia , Osteoclastos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/fisiologia , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologiaRESUMO
Clinical evidence indicates that fat is inversely proportional to bone mass in elderly obese women. However, it remains unclear whether obesity accelerates bone loss. In this report we present evidence that increased visceral fat leads to inflammation and subsequent bone loss in 12-month-old C57BL/6J mice that were fed 10% corn oil (CO)-based diet and a control lab chow (LC) for 6 months. As expected from our previous work, CO-fed mice demonstrated increased visceral fat and enhanced total body fat mass compared to LC. The adipocyte-specific PPARγ and bone marrow (BM) adiposity were increased in CO-fed mice. In correlation with those modifications, inflammatory cytokines (IL-1ß, IL-6, TNF-α) were significantly elevated in CO-fed mice compared to LC-fed mice. This inflammatory BM microenvironment resulted in increased superoxide production in osteoclasts and undifferentiated BM cells. In CO-fed mice, the increased number of osteoclasts per trabecular bone length and the increased osteoclastogenesis assessed ex-vivo suggest that CO diet induces bone resorption. Additionally, the up-regulation of osteoclast-specific cathepsin k and RANKL expression and down-regulation of osteoblast-specific RUNX2/Cbfa1 supports this bone resorption in CO-fed mice. Also, CO-fed mice exhibited lower trabecular bone volume in the distal femoral metaphysis and had reduced OPG expression. Collectively, our results suggest that increased bone resorption in mice fed a CO-enriched diet is possibly due to increased inflammation mediated by the accumulation of adipocytes in the BM microenvironment. This inflammation may consequently increase osteoclastogenesis, while reducing osteoblast development in CO-fed mice.
Assuntos
Reabsorção Óssea/imunologia , Reabsorção Óssea/patologia , Obesidade/imunologia , Obesidade/patologia , Adipócitos/imunologia , Adipócitos/patologia , Animais , Biomarcadores , Peso Corporal/imunologia , Medula Óssea/imunologia , Medula Óssea/patologia , Óleo de Milho/farmacologia , Gorduras na Dieta/farmacologia , Feminino , Fêmur/metabolismo , Fêmur/patologia , Expressão Gênica/imunologia , Gordura Intra-Abdominal/imunologia , Gordura Intra-Abdominal/patologia , Camundongos , Camundongos Endogâmicos C57BL , Osteoblastos/imunologia , Osteoblastos/patologia , Osteoclastos/imunologia , Osteoclastos/patologia , Osteoprotegerina/metabolismo , PPAR gama/genética , Ligante RANK/metabolismoRESUMO
Liposomes containing phosphatidylserine (PS) are engulfed by phagocytes including macrophages, microglia, and dendritic cells. PS liposomes (PSLs) mimic the effects of apoptotic cells on these phagocytes to induce the secretion of anti-inflammatory molecules and to inhibit the maturation of dendritic cells. However, the effects of PSLs on osteoclasts, which are also differentiated from the common myeloid precursors, remain to be determined. This study investigated the effects of PSLs on the osteoclastogenesis. In the rat bone marrow culture system, osteoclast precursors phagocytosed PSLs to secrete TGF-beta1 and PGE(2), which in turn inhibited osteoclastogenesis through the downregulation of receptor activator for NF-kappaB ligand, receptor activator of NF-kappaB, ICAM-1, and CD44. Consistent with these in vitro observations, i.m. injection of PSLs significantly increased the plasma level of TGF-beta1 and PGE(2) and decreased the expression of receptor activator for NF-kappaB ligand, receptor activator of NF-kappaB, and ICAM-1 in the skeletal tissues of ankle joints of rats with adjuvant arthritis (AA). A quantitative analysis using microcomputed tomography revealed that PSLs as well as TGF-beta1 together with PGE(2) significantly inhibited AA-induced trabecular bone loss. These observations strongly suggest that PSLs generate TGF-beta1 and PGE(2) release, leading to inhibit osteoclastogenesis and AA-induced trabecular bone loss. Because PS is a component of the cell membrane, PSLs therefore can be a potentially effective pharmacological intervention against abnormal bone loss, such as osteoporosis without deleterious side effects.
Assuntos
Reabsorção Óssea/prevenção & controle , Diferenciação Celular/imunologia , Regulação para Baixo/imunologia , Inibidores do Crescimento/administração & dosagem , Inibidores do Crescimento/fisiologia , Osteoclastos/imunologia , Fosfatidilserinas/administração & dosagem , Fosfatidilserinas/fisiologia , Animais , Artrite Experimental/imunologia , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Artrite Experimental/prevenção & controle , Reabsorção Óssea/imunologia , Reabsorção Óssea/metabolismo , Reabsorção Óssea/patologia , Células Cultivadas , Dinoprostona/metabolismo , Modelos Animais de Doenças , Feminino , Receptores de Hialuronatos/biossíntese , Molécula 1 de Adesão Intercelular/biossíntese , Lipossomos , Osteoclastos/metabolismo , Osteoclastos/patologia , Ligante RANK/antagonistas & inibidores , Ligante RANK/biossíntese , Ratos , Ratos Endogâmicos Lew , Receptor Ativador de Fator Nuclear kappa-B/antagonistas & inibidores , Receptor Ativador de Fator Nuclear kappa-B/biossíntese , Células-Tronco/citologia , Células-Tronco/imunologia , Células-Tronco/metabolismo , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Rheumatoid arthritis (RA) is a chronic and debilitating autoimmune disease characterized by chronic joint inflammation with subsequent cartilage and bone destruction. RA is emerging as a model of IL-17-driven autoimmune inflammatory disease. IL-17 is a marker for Th17 cells, with its master regulator being the retinoic acid receptor-related orphan receptor (RORgammat) regulated by STAT3 signaling. Glucuronoxylomannan (GXM), a polysaccharide representing the main component of the capsular material of the opportunistic yeast Cryptococcus neoformans, exhibits potent immunosuppressive properties both in vitro and in vivo. The present study investigates the effects of GXM treatment on the progression of collagen-induced arthritis. GXM suppressed clinical signs of collagen-induced arthritis and blocked joint erosion progression. This effect was mediated by down-regulation of key cytokines involved in the pathogenesis of RA such as TNF-alpha and IL-1beta, and up-regulation of the inhibitory cytokine IL-10. Moreover, a reduction of IL-6 and TGF-beta, which inhibit Th17 differentiation with consequent decreased IL-17 production at the local and systemic level, was observed. The effect of GXM on Th17 differentiation mirrored the reduction in STAT3 activation and inhibition of RORgammat synthesis. Consequently, this work highlights the beneficial properties of an efficacious compound that could eventually be destined to the clinic.
Assuntos
Artrite Reumatoide/imunologia , Cryptococcus neoformans/imunologia , Citocinas/biossíntese , Imunossupressores/administração & dosagem , Mediadores da Inflamação/metabolismo , Interleucina-17/fisiologia , Polissacarídeos/administração & dosagem , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Antígenos de Fungos/administração & dosagem , Antígenos de Fungos/imunologia , Artrite Experimental/tratamento farmacológico , Artrite Experimental/epidemiologia , Artrite Experimental/imunologia , Artrite Experimental/patologia , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/patologia , Reabsorção Óssea/imunologia , Reabsorção Óssea/patologia , Colágeno Tipo II/toxicidade , Citocinas/antagonistas & inibidores , Citocinas/fisiologia , Imunossupressores/imunologia , Incidência , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/fisiologia , Interleucina-17/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos DBA , Osteoclastos/imunologia , Osteoclastos/patologia , Polissacarídeos/imunologia , Polissacarídeos/uso terapêutico , Ligante RANK/biossíntese , Ligante RANK/genética , Índice de Gravidade de Doença , Linfócitos T Auxiliares-Indutores/metabolismo , Linfócitos T Auxiliares-Indutores/patologiaRESUMO
OBJECTIVE: In vitro spontaneous osteoclastogenesis from peripheral blood mononuclear cells (PBMCs) is increased in diseases with excessive bone loss. The purpose of this study was to reassess the role of T lymphocytes in this process. METHODS: Fresh or cryopreserved PBMCs obtained from healthy subjects and from patients with rheumatoid arthritis, psoriatic arthritis, and non-psoriatic spondylarthritis were cultured at high density and stained for tartrate-resistant acid phosphatase (TRAP). Resorption of mineralized matrix was assessed by a dentin disc assay. CD14+ monocytes and CD3+ T cells were selected using magnetically labeled antibodies. RESULTS: Numerous multinucleated, TRAP+, dentin-resorbing osteoclasts developed spontaneously from fresh PBMCs from healthy individuals. This process was abrogated by T cell depletion and was restored by exogenous macrophage colony-stimulating factor (M-CSF) and RANKL, indicating the important role of T cells in spontaneous osteoclastogenesis in vitro. Using physiologic freezing and thawing as a model for the activation of PBMCs, spontaneous osteoclastogenesis was significantly increased in cryopreserved versus fresh cells. Under these conditions, spontaneous osteoclastogenesis was not dependent on T lymphocytes, since it was not influenced by T cell depletion and persisted in purified CD14+ cell cultures supplemented with M-CSF and RANKL. In contrast to studies with fresh PBMCs, spontaneous osteoclastogenesis under these conditions did not appear to be clearly different between healthy subjects and patients with arthritis. CONCLUSION: Spontaneous osteoclastogenesis in vitro is dependent on T lymphocytes or on the direct activation of monocytic cells, depending on the test conditions. This variability warrants better validation of the relevance of this functional test for in vivo osteoclastogenesis.