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BACKGROUND: Hepatitis B (HB) is an infectious disease with unfavorable consequence for patients and involved in chronic inflammation of liver. The present study aimed to investigate whether High-mobility group protein B (HMGB)1/receptor for advanced glycation end products (RAGE) aggravates inflammation enhancing the expression of interleukin (IL)-17. METHODS: Mild and severe HB liver tissue and peripheral blood samples were obtained intra-operatively. Histological analysis of the livers was performed by immunohistochemistry. IL-1ß and IL-6 of liver tissue were detected by confocal microscopy staining. Relative mRNA expression was measured by real-time PCR and protein levels were measured by enzyme-linked immunosorbent assay. RESULTS: HMGB1, RAGE and IL-17 expression is increased in liver of HB patients with acute on chronic liver failure (ACLF) compared to healthy controls. HMGB1 treatment induced inflammatory cytokines including IL-17 in peripheral blood cells of HB patients. IL-17 also induced the expression of RAGE and IL-1ß in peripheral blood cells of HB patients with ACLF. On the other hands, the inhibitory factor of p38 and nuclear factor-kappa B reduced the expression of RAGE and IL-1ß in peripheral blood cells HB patients with ACLF. CONCLUSIONS: HMGB1, RAGE and IL-17 expression is increased in liver of severe HB patients. HMGB1 and RAGE interaction may contribute to the inflammation of liver enhancing the expression of IL-17, which can be possibly restored through the decline of the HMGB1/RAGE axis.
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Proteína HMGB1/metabolismo , Hepatitis B/sangre , Inflamación , Interleucina-17/metabolismo , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Adulto , Estudios de Casos y Controles , Ensayo de Inmunoadsorción Enzimática , Femenino , Regulación de la Expresión Génica , Humanos , Inmunohistoquímica , Interleucina-1beta/metabolismo , Hígado/metabolismo , Masculino , Microscopía Confocal , Microscopía Fluorescente , Persona de Mediana Edad , Estudios Prospectivos , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Inmunológicos/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
BACKGROUND: Surgical trauma elicits inflammatory responses, including the secretion of cytokines. Recent studies demonstrated that beta-blockers could reduce the expression of cytokines after injury. We therefore tested the effects of different doses of intraoperative esmolol on the inflammatory response after surgery. METHODS: Patients undergoing laparoscopic gastrectomy were randomly separated into 1 of 3 groups: saline, clinical dose, and subclinical dose groups. The levels of interleukin (IL)-6, IL-4, and IL-10 were quantified by sandwich enzyme-linked immunoassay after the induction of anesthesia (T0), at the end of peritoneal closure (T1), and 60 minutes after surgery (T2). Levels of C-reactive protein (CRP) were measured on postoperative day 1. RESULTS: At T2, the levels of IL-6 and IL-10 in the saline group were elevated significantly compared with at T0 or T1 (IL-6: 119.62 and 15.97 pg/mL at T2 and T0, respectively [P = .042]; IL-10: 27.27 and 7.03 pg/mL at T2 and T1, respectively [P = .037]). However, no changes were observed over time in the clinical dose group. In contrast, postoperative levels of IL-4 were decreased significantly in the clinical dose group compared with the saline group (2.14 vs 21.91 pg/mL, P = .022). In addition, the CRP levels on postoperative day 1 were lower in the esmolol-treated groups, in a dose-dependent manner. CONCLUSIONS: Serum IL-6 and IL-10 levels were increased over time, suggesting that laparoscopic surgery is a stressor, even though it causes minimal tissue injury. Treatment with esmolol decreased the inflammatory response and CRP production in a dose-dependent manner.
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Antagonistas de Receptores Adrenérgicos beta 1/administración & dosificación , Gastrectomía/efectos adversos , Laparoscopía/efectos adversos , Propanolaminas/administración & dosificación , Anciano , Proteína C-Reactiva/metabolismo , Relación Dosis-Respuesta a Droga , Femenino , Gastrectomía/métodos , Humanos , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Interleucinas/sangre , Cuidados Intraoperatorios , Laparoscopía/métodos , Masculino , Persona de Mediana EdadRESUMEN
Systemic lupus erythematosus (SLE) is an autoimmune disease in which abnormal immune responses are mediated by tissue-binding autoantibodies and immune complex deposition. Because most SLE patients are women of child-bearing age, oestrogen has been suggested to play an important role in SLE pathogenesis. One proposed role is to induce B-cell activation, culminating in increased autoantibody production. Interleukin-21 (IL-21) has been shown to be crucial in the differentiation of activated B cells into plasma cells. We therefore hypothesized that oestrogen up-regulates IL-21 production and induces subsequent B-cell activation in SLE patients. Peripheral blood was obtained from 22 SLE patients and 16 healthy controls. Expression levels of IL-21 and its receptor in serum, peripheral blood mononuclear cells, and CD4(+) T cells were higher in SLE patients than in healthy controls. Exposure of CD4(+) T cells from SLE patients to 17ß-oestradiol led to a dose- and time-dependent increase in IL-21 expression, which was abolished in the presence of mitogen-activated protein kinase (MAPK) (MAPK kinase, p38, Jun N-terminal kinase) inhibitors. B cells from healthy controls showed increased antibody production when they were co-cultured with oestrogen-treated CD4(+) T cells from SLE patients. Treatment with IL-21 antibody abrogated the increased antibody production of the co-culture systems. This study revealed the association between oestrogen and IL-21 in SLE patients. Oestrogen up-regulates IL-21 expression of CD4(+) T cells via MAPK-dependent pathways in SLE patients, which in turn induces increased antibody production by B cells.
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Linfocitos T CD4-Positivos/inmunología , Estradiol/farmacología , Estrógenos/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Interleucinas/inmunología , Lupus Eritematoso Sistémico/inmunología , Adulto , Formación de Anticuerpos/efectos de los fármacos , Linfocitos T CD4-Positivos/patología , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Regulación de la Expresión Génica/inmunología , Humanos , Interleucinas/biosíntesis , Lupus Eritematoso Sistémico/metabolismo , Lupus Eritematoso Sistémico/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/inmunología , Persona de Mediana Edad , Células Plasmáticas/inmunología , Células Plasmáticas/metabolismo , Células Plasmáticas/patologíaRESUMEN
OBJECTIVE: Bone destruction is a critical pathology involved in the functional disability caused by rheumatoid arthritis (RA). Osteoclasts, which are specialized bone-resorbing cells regulated by cytokines such as RANKL, are implicated in bone destruction in RA. The aim of this study was to determine whether interleukin-21 (IL-21), a potent immunomodulatory 4-α-helical bundle type 1 cytokine, has osteoclastogenic activity in patients with RA and in mice with collagen-induced arthritis (CIA). METHODS: The expression of IL-21 in synovial tissue was examined using immunohistochemistry. The concentrations of IL-21 in serum and synovial fluid were determined by enzyme-linked immunosorbent assay. The levels of RANKL and osteoclastogenic markers were measured using real-time polymerase chain reaction. CD14+ monocytes from patients with RA or mouse bone marrow cells were cocultured with fibroblast-like synoviocytes (FLS) from patients with RA or CD4+ T cells from mice with CIA in the presence of IL-21 and subsequently stained for tartrate-resistant acid phosphatase activity to determine osteoclast formation. RESULTS: IL-21 was up-regulated in the synovium, synovial fluid, and serum of patients with RA and in the synovium and serum of mice with CIA. IL-21 induced RANKL expression in mixed joint cells and CD4+ T cells from mice with CIA and in CD4+ T cells and FLS from patients with RA. Moreover, IL-21 enhanced in vitro osteoclastogenesis without the presence of RANKL-providing cells and by inducing RANKL expression in CD4+ T cells and FLS. CONCLUSION: Our data suggest that IL-21 promotes osteoclastogenesis in RA. We believe that therapeutic strategies targeting IL-21 might be effective for the treatment of patients with RA, especially in preventing bone destruction.
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Artritis Experimental/patología , Artritis Reumatoide/patología , Interleucinas/metabolismo , Osteoclastos/patología , Membrana Sinovial/patología , Fosfatasa Ácida/metabolismo , Animales , Artritis Experimental/metabolismo , Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Biomarcadores/metabolismo , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/patología , Células Cultivadas , Técnicas de Cocultivo , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Receptores de Lipopolisacáridos/metabolismo , Masculino , Ratones , Ratones Endogámicos DBA , Monocitos/metabolismo , Monocitos/patología , Osteoclastos/metabolismo , Ligando RANK/genética , Ligando RANK/metabolismo , Membrana Sinovial/metabolismoRESUMEN
OBJECTIVE: The interleukin (IL)-12 cytokine family is closely related to the development of T helper cells, which are responsible for autoimmune disease enhancement or suppression. IL-12 family members are generally heterodimers and share three α-subunits (p35, p19, and p28) and two ß-subunits (p40 and EBI3). However, a ß-sheet p40 homodimer has been shown to exist and antagonize IL-12 and IL-23 signaling 1. Therefore, we assumed the existence of a p40-EBI3 heterodimer in nature and sought to investigate its role in immune regulation. METHODS: The presence of the p40-EBI3 heterodimer was confirmed by ELISA, immunoprecipitation, and western blotting. A p40-EBI3 vector and p40-EBI3-Fc protein were synthesized to confirm the immunological role of this protein in mice with collagen-induced arthritis (CIA). The anti-inflammatory effects of p40-EBI3 were analyzed with regard to clinical, histological, and immune cell-regulating features in mice with CIA. RESULTS: Clinical arthritis scores and the expression levels of proinflammatory cytokines (e.g., IL-17, IL-1ß, IL-6, and TNF-α) were significantly attenuated in p40-EBI3-overexpressing and p40-EBI3-Fc-treated mice with CIA compared to vehicle-treated mice with CIA. Structural joint damage and vessel formation-related gene expression were also reduced by p40-EBI3 heterodimer treatment. In vitro, the p40-EBI3-Fc protein significantly suppressed the differentiation of Th17 cells and reciprocally induced CD4+CD25+Foxp3+ (regulatory T) cells. p40-EBI3 also inhibited osteoclast formation in a concentration-dependent manner. CONCLUSION: In this study, p40-EBI3 ameliorated proinflammatory conditions both in vivo and in vitro. We propose that p40-EBI3 is a novel anti-inflammatory cytokine involved in suppressing the immune response through the expansion of Treg cells and suppression of Th17 cells and osteoclastogenesis.
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Artritis Experimental , Enfermedades Autoinmunes , Interleucina-12 , Animales , Citocinas/uso terapéutico , Interleucina-12/química , Interleucina-12/metabolismo , Ratones , Antígenos de Histocompatibilidad Menor , Receptores de Citocinas/genética , Receptores de Citocinas/uso terapéutico , Linfocitos T Reguladores , Células Th17RESUMEN
In the present study, we investigated the effects of Galla Rhois (GR) on obesity and gene expression. We prepared a GR extract and various solvent fractions and evaluated the degree to which they inhibited adipocyte differentiation and adipogenesis in vitro. Among them, the GR ethyl acetate fraction (GE) had the lowest EC50 for adipocyte differentiation and adipogenesis and thus was selected for in vivo experiments. We induced obesity in C57BL/6 mice by providing them a high-fat diet (HFD). Then, GE (10-40 mg/kg) or orlistat (positive control, 4 mg/kg) was orally administered daily for six weeks. Mean body weights and weight gain were significantly lower in the GE40 group (40 mg/kg of GE) compared with the HFD group (p < 0.05). The most significant changes in serum glucose, total cholesterol, and triglyceride levels were confirmed in the GE40 group (p < 0.05). Epididymal fat was weighed and stained for body fat measurement, and significant differences were recorded from GE10 to GE40 (p < 0.05). Finally, 3T3-L1 pre-adipocytes were treated with GE, and cDNA from these cells was used for microarray analysis and qRT-PCR. Microarray analysis revealed 13 genes up-regulated and 21 genes down-regulated by GE. From the qRT-PCR analysis, we found that GE altered the mRNA expression of eosinophil peroxidase, glucose-dependent insulinotropic polypeptide receptor, and apolipoprotein B. Based on this study, we suggest that GR could be developed as an anti-obesity therapeutic agent.
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Adipogénesis/efectos de los fármacos , Fármacos Antiobesidad/farmacología , Productos Biológicos/farmacología , Obesidad/tratamiento farmacológico , Células 3T3-L1 , Adipocitos/citología , Adipocitos/efectos de los fármacos , Adipogénesis/genética , Animales , Peso Corporal/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Dieta Alta en Grasa , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/genética , Aumento de Peso/efectos de los fármacosRESUMEN
BACKGROUND/OBJECTIVES: Premenstrual syndrome (PMS) is a disorder characterized by repeated emotional, behavioral, and physical symptoms before menstruation, and the exact cause and mechanism are uncertain. Hyperprolactinemia interferes with the normal production of estrogen and progesterone, leading to PMS symptoms. Thus, we judged that the inhibition of prolactin hypersecretion could mitigate PMS symptoms. MATERIALS/METHODS: Hordeum vulgare L. extract (HVE), Chrysanthemum zawadskii var. latilobum extract (CZE), and Lomens-P0 the mixture of these extracts were tested in subsequent experiments. The effect of extracts on prolactin secretion at the in vitro level was measured in GH3 cells. Nitric oxide and pro-inflammatory mediator expression were measured in RAW 264.7 cells to confirm the anti-inflammatory effect. Also, the hyperprolactinemic Institute for Cancer Research (ICR) mice model was used to measure extract effects on prolactin and hormone secretion and uterine inflammation. RESULTS: Anti-inflammatory effects of and prolactin secretion suppress by HVE and CZE were confirmed through in vitro experiments (P < 0.05). Treatment with Lomens-P0 inhibited prolactin secretion (P < 0.05) and restored normal sex hormone secretion in the hyperprolactinemia mice model. In addition, extracts significantly inhibited the expression of pro-inflammatory biomarkers, including interleukin-1ß, and -6, tumor necrosis factor-α, inducible nitric oxide synthase, and cyclooxygenase-2 (P < 0.01). We used high-performance liquid chromatography analyses to identify tricin and chlorogenic acid as the respective components of HVE and CZE that inhibit prolactin secretion. The Lomens-P0, which includes tricin and chlorogenic acid, is expected to be effective in improving PMS symptoms in the human body. CONCLUSIONS: The Lomens-P0 suppressed the prolactin secretion in hyperprolactinemia mice, normalized the sex hormone imbalance, and significantly suppressed the expression of inflammatory markers in uterine tissue. This study suggests that Lomens-P0 may have the potential to prevent or remedy materials to PMS symptoms.
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Agrimonia pilosa (AP) and Rhus gall (RG) are traditional medicinal plants. The bioflavonoid composition standardized by HPLC analysis was named APRG64. Despite many studies reported to beneficial bioactivities of AP and RG, very limited range of toxicity tests have documented. So, we did experiment diversely on the toxicity tests of the substance APRG64. Genotoxicity (mammalian chromosomal aberration test, micronoucleus test) against APRG64, acute and sub-chronic toxicity test from rodent/non-rodent, and systemic safety pharmacology test were conducted. As a result of the test, genotoxicity against APRG64 was not observed. The NOAEL of rodents was confirmed as 2000 mg/kg/day and non-rodents was confirmed as 500 mg/kg/day. In addition, systemic safety pharmacological toxicity (effects on respiratory system, central nervous system, cardiovascular system) following administration of APRG64 was not observed. Finally, we accomplished ten potential toxicity tests and evaluated extensive safety of APRG64. Consequently, APRG64 may be a promising material for nutraceuticals and natural medicines.
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Dysfunction of T helper 17 (Th17) cells leads to chronic inflammatory disorders. Signal transducer and activator of transcription 3 (STAT3) orchestrates the expression of proinflammatory cytokines and pathogenic cell differentiation from interleukin (IL)-17-producing Th17 cells. However, the pathways mediated by STAT3 signaling are not fully understood. Here, we observed that Fos-related antigen 1 (FRA1) and JUNB are directly involved in STAT3 binding to sites in the promoters of Fosl1 and Junb. Promoter binding increased expression of IL-17 and the development of Th17 cells. Overexpression of Fra1 and Junb in mice resulted in susceptibility to collagen-induced arthritis and an increase in Th17 cell numbers and inflammatory cytokine production. In patients with rheumatoid arthritis, FRA1 and JUNB were colocalized with STAT3 in the inflamed synovium. These observations suggest that FRA1 and JUNB are associated closely with STAT3 activation, and that this activation leads to Th17 cell differentiation in autoimmune diseases and inflammation.
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OBJECTIVE: STAT-3 is a key transcriptional factor in the interleukin-6 (IL-6)-mediated differentiation of Th17 cells. Because Th17 is believed to be a central player in rheumatoid arthritis (RA), we sought to evaluate whether an endogenous inhibitor of the STAT3 gene, GRIM-19 (gene associated with retinoid-interferon-induced mortality 19), could attenuate the progression and severity of murine collagen-induced arthritis (CIA) through suppression of Th17 cells and, reciprocally, could increase expression of Treg cells. METHODS: Overexpression of GRIM-19 was produced either by intravenous/intramuscular administration of a GRIM-19 overexpression vector in DBA1/J mice or by development of GRIM-19-transgenic (Tg) mice on a C57BL/6 background. Clinical signs were scored for arthritis severity, and mouse splenocytes, serum, and joint tissue were obtained for immunostaining and histologic analyses. RESULTS: The numbers of CD4+IL-17+ cells and CD4+pSTAT3+ cells were decreased, while the numbers of CD4+CD25+Foxp3+ cells and CD4+pSTAT5+ cells were increased, in both GRIM-19 vector-transfected and GRIM-19-Tg mice. Administration of the GRIM-19 overexpression vector into mice with CIA markedly suppressed the clinical and histologic signs of arthritis in the affected joints. Similarly, when CIA was induced in GRIM-19-Tg mice, the arthritis phenotype was markedly attenuated and the expression of inflammatory cytokines (IL-1ß, IL-6, tumor necrosis factor α, and IL-17) in the arthritic joints was also significantly reduced. Moreover, bone marrow-derived monocyte/macrophages obtained from GRIM-19-Tg mice showed attenuated RANKL-induced osteoclastogenesis in vitro. CONCLUSION: GRIM-19 improved the clinical and histologic features of CIA and also inhibited osteoclast formation. These findings suggest that GRIM-19 may be a novel treatment agent for RA.
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Artritis Experimental/genética , Artritis Reumatoide/genética , NADH NADPH Oxidorreductasas/genética , Linfocitos T Reguladores/metabolismo , Células Th17/metabolismo , Animales , Artritis Experimental/metabolismo , Artritis Experimental/patología , Artritis Reumatoide/metabolismo , Artritis Reumatoide/patología , Diferenciación Celular/fisiología , Citocinas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , NADH NADPH Oxidorreductasas/metabolismo , Osteoclastos/metabolismo , Osteoclastos/patología , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Linfocitos T Reguladores/patología , Células Th17/patologíaRESUMEN
OBJECTIVES: The immune system releases cytokines during the stress response, and the balance between proinflammatory and anti-inflammatory cytokines is important. This prospective study was done to determine which cytokines are responsible for maintaining cytokine balance during live-donor liver transplant surgery. MATERIALS AND METHODS: Recipients undergoing live-donor liver transplant surgery due to cirrhotic hepatitis were allocated to a recipient group (n=44), and healthy donors were placed in the donor group (n=45). In donors, blood sampling for cytokine level analysis was performed after anesthetic induction (before the start of surgery, time point 1). In recipients, blood samples were collected before the start of surgery (time point 1), 60 minutes after the start of the anhepatic period (time point 2), and 60 minutes after reperfusion (time point 3). The proinflammatory cytokines measured were interleukin-1 ß, interleukin-6, and tumor necrosis factor-α; the anti-inflammatory cytokines were interleukin-10 and interleukin-4. Cytokines were quantified using sandwich enzyme-linked immunoassays. The time course of proinflammatory and anti-inflammatory cytokine concentrations during surgery in the recipient group was evaluated. RESULTS: Interleukin-6, interleukin-10 and tumor necrosis factor-α showed significant changes in concentration during surgery, with interleukin-6 reaching levels 40 times higher than the preoperative value at the anhepatic stage. Interleukin-10 reached a peak at the neohepatic phase, with values 60 times higher than the preoperative value. The preoperative concentrations of interleukin-6 and interleukin-10 in the recipient group were higher than those in the donor group with a median of 4.48 vs 1.98 pg/mL (P < .001) and 2.98 vs 1.22 (P = .026). CONCLUSIONS: Interleukin-6 and interleukin-10 play a major role in cytokine balance before and during live-donor liver transplant surgery.
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Citocinas/sangre , Hepatitis Viral Humana/cirugía , Homeostasis/fisiología , Inflamación/metabolismo , Cirrosis Hepática/cirugía , Trasplante de Hígado , Donadores Vivos , Adulto , Femenino , Hepatitis Viral Humana/complicaciones , Hepatitis Viral Humana/metabolismo , Humanos , Interleucina-10/sangre , Interleucina-1beta/sangre , Interleucina-4/sangre , Interleucina-6/sangre , Cirrosis Hepática/etiología , Cirrosis Hepática/metabolismo , Masculino , Persona de Mediana Edad , Periodo Perioperatorio , Estudios Prospectivos , Factor de Necrosis Tumoral alfa/sangreRESUMEN
OBJECTIVES: Magnesium has protective effects in ischaemia-reperfusion injury, and is involved in immunomodulation. We investigated the effects of magnesium pretreatment on the secretion of T helper (Th) cytokines and on the severity of post-reperfusion syndrome (PRS) in patients undergoing living donor liver transplantation (LDLT). METHODS: forty patients were allocated to two groups of 20 (magnesium and saline groups). Blood samples for cytokine analysis were collected before infusion of the study solution at the end of anhepatic phase (time point 1), as well as five min and 30 min after allograft reperfusion (time points 2 and 3, respectively). Levels of cytokines were quantified using a sandwich enzyme immunoassay test kit. RESULTS: The duration of PRS was shorter in the magnesium group (p = 0.038). The level of interferon (IFN)-γ released from Th1 was lower in the magnesium group at time point 3 (p = 0.009). Of the cytokines released from Th2 cells, interleukin (IL)-6 was present in higher concentrations in the magnesium group at time points 2 and 3 (p<0.05). The concentrations of IL-4 and IL-10, which were secreted from Th2 cells, were also higher in the magnesium group at time point 3 (p<0.05). The IFN-γ /IL-6, IFN-γ /IL-4 and IFN-γ /IL-10 ratios were lower in the magnesium group after allograft reperfusion. CONCLUSIONS: Magnesium pretreatment attenuated PRS and reinforced Th2 cell activity, shifting the Th1-to-Th2 cytokine balance towards Th2 in patients undergoing LDLT.
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Citocinas/sangre , Trasplante de Hígado , Donadores Vivos , Magnesio/uso terapéutico , Daño por Reperfusión/sangre , Daño por Reperfusión/tratamiento farmacológico , Linfocitos T Colaboradores-Inductores/metabolismo , Femenino , Humanos , Magnesio/administración & dosificación , Magnesio/farmacología , Masculino , Persona de Mediana Edad , Atención Perioperativa , Daño por Reperfusión/patología , Síndrome , Linfocitos T Colaboradores-Inductores/efectos de los fármacosRESUMEN
INTRODUCTION: Interleukin (IL)-32 and IL-17 play critical roles in pro-inflammatory responses and are highly expressed in the synovium of patients with rheumatoid arthritis (RA). We investigated the relations between these two cytokines (IL-17 and IL-32) for their ability to induce each other and to stimulate osteoclasts in RA fibroblast-like synoviocytes (FLSs) and T cells. METHODS: FLSs were isolated through surgical synovectomy obtained from patients with RA or osteoarthritis (OA). Real-time PCR were performed to evaluate the expression of IL-32, IL-17 and osteoclast-related genes. Immunohistochemical staining and tartrate-resistant acid phosphatase (TRAP) staining were performed to determine the distribution of inflammatory cytokines and the presence of osteoclastogenesis. RESULTS: IL-17 induced the expression of IL-32 in the FLSs from RA patients, as assessed by microarray. IL-32 production was increased by IL-17. IL-32 in the FLSs from RA patients induced the production of IL-17 in CD4+ T cells. IL-32 and IL-17 were colocalized near TRAP-positive areas in joint specimens. IL-17 and IL-32 synergistically induced the differentiation of osteoclasts, as demonstrated by the expression of osteoclast-related genes. IL-32 and IL-17 also could induce resorption by osteoclasts in a RANKL-dependent manner. CONCLUSIONS: IL-17 affected the expression of IL-32 in FLSs of RA patients and IL-32 induced the production of IL-17 in CD4+ T cells. Both IL-17 and IL-32 cytokines can reciprocally influence each other's production and amplify the function of osteoclastogenesis in the in RA synovium. Separately, IL-17 and IL-32 each stimulated osteoclastogenesis without RANKL. Together, the two cytokines synergistically amplified the differentiation of osteoclasts, independent of RANKL stimulation.
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Artritis Reumatoide/genética , Interleucina-17/genética , Interleucinas/genética , Osteoclastos/metabolismo , Osteogénesis/genética , Animales , Artritis Experimental/genética , Artritis Experimental/metabolismo , Artritis Experimental/patología , Artritis Reumatoide/metabolismo , Artritis Reumatoide/patología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Células Cultivadas , Técnicas de Cocultivo , Citocinas/genética , Citocinas/metabolismo , Sinergismo Farmacológico , Humanos , Inmunohistoquímica , Interleucina-17/metabolismo , Interleucina-17/farmacología , Interleucinas/metabolismo , Interleucinas/farmacología , Masculino , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , FN-kappa B/genética , FN-kappa B/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Osteoclastos/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Líquido Sinovial/citología , Líquido Sinovial/metabolismo , Células Th17/efectos de los fármacos , Células Th17/metabolismo , Transcriptoma/efectos de los fármacos , Transcriptoma/genéticaRESUMEN
INTRODUCTION: The study was undertaken to investigate the interrelation of toll-like receptor (TLR) and interleukin (IL)-17 in the salivary glands of patients with primary Sjogren's syndrome (pSS) and to determine the role of TLR and IL-17 in the pathophysiology of pSS. METHODS: The expressions of various TLRs, IL-17 and the cytokines involved in Th17 cell differentiation including IL-6, IL-23, tumor necrosis factor-alpha (TNF-α) and IL-1ß were examined by immunohistochemistry in salivary glands of pSS patients. The IL-17 producing CD4+ T cells (Th17 cells) were examined by flow cytometry and confocal staining in peripheral mononuclear blood cells (PMBCs) and salivary glands of pSS patients. After PBMCs were treated with TLR specific ligands, the induction of IL-17 and IL-23 was determined using real-time PCR and ELISA. The signaling pathway that mediates the TLR2 stimulated production of IL-17 and IL-23 was investigated by using treatment with specific signaling inhibitors. RESULTS: We showed that TLR2, TLR4, TLR6, IL-17 and the cytokines associated with Th17 cells were highly expressed in salivary glands of pSS patients but not in controls. The expressions of TLR2, TLR4 and TLR6 were observed in the infiltrating mononuclear cells and ductal epithelial cells, whereas IL-17 was mainly observed in infiltrating CD4+ T cells. The number of IL-17 producing CD4+ T cells was significantly higher in pSS patients both in PBMCs and minor salivary glands. The stimulation of TLR2, TLR4 and TLR6 additively induced the production of IL-17 and IL-23 from the PBMCs of pSS patients especially in the presence of TLR2 stimulation. IL-6, signal transducer and activator of transcription 3 (STAT3) and nuclear factor-kappaB (NF-kB) pathways were implicated in the TLR2 stimulated IL-17 and IL-23. CONCLUSIONS: Our data demonstrate that TLR2 ligation induces the production of IL-23/IL-17 via IL-6, STAT3 and NF-kB pathway in pSS. Therefore, therapeutic strategies that target TLR/IL-17 pathway might be strong candidates for treatment modalities of pSS.
Asunto(s)
Interleucina-17/biosíntesis , Interleucina-23/biosíntesis , Interleucina-6/fisiología , FN-kappa B/fisiología , Factor de Transcripción STAT3/fisiología , Síndrome de Sjögren/metabolismo , Receptor Toll-Like 2/metabolismo , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Células Cultivadas , Femenino , Humanos , Ligandos , Masculino , Glándulas Salivales/metabolismo , Transducción de Señal/fisiología , Síndrome de Sjögren/patología , Síndrome de Sjögren/fisiopatologíaRESUMEN
Osteoclastogenesis plays an important role in joint destruction in rheumatoid arthritis (RA). IL-15 is a pleiotropic proinflammatory cytokine that appears to help mediate the pathological bone loss. This study was undertaken to explore the signaling molecules essential for osteoclastogenesis mediated by IL-15 in rheumatoid synovial fibroblasts. Expression of phospholipase D1 (PLD1) and osteoclast-related gene expression in synovial tissues and their modulation by treatment with IL-15 and different inhibitors in synovial fibroblasts of RA patients were evaluated using immunohistochemistry and quantitative polymerase chain reaction. The levels of IL-15 in serum and synovial fluid were measured by ELISA. The effects of IL-15 and phosphatidic acid (PA) on osteoclast formation were evaluated in cocultures of rheumatoid synovial fibroblasts and peripheral blood monocytes or monocytes alone in the presence of M-CSF and RANKL. The levels of RANKL and PLD1 but not PLD2 were upregulated significantly by IL-15, and the RANKL level was significantly upregulated by PA in rheumatoid synovial fibroblasts. Blocking PA production with 1-butanol and siRNA against PLD1 significantly inhibited the IL-15-stimulated expression of RANKL and PLD1. IL-15 levels were significantly higher in serum and synovial fluid from patients with RA than in osteoarthritis patients and healthy controls. IL-15 and PA induced osteoclast formation through the mitogen-activated protein kinases (MAPKs) and NF-κB signaling pathways. Activation of PLD1 contributes to IL-15-mediated osteoclastogenesis via the MAPKs and NF-κB signaling pathways in rheumatoid synovial fibroblasts. Our data suggest that PLD1 might be an efficient therapeutic strategy for preventing bone destruction in rheumatoid arthritis.