RESUMO
Bone erosion is one of the primary features of inflammatory arthritis and is caused by excessive differentiation and activation of osteoclasts. Fc gamma receptors (FcγRs) have been implicated in osteoclastogenesis. Our recent studies demonstrate that joint-deposited lupus IgG inhibited RANKL-induced osteoclastogenesis. FcγRI is required for RANKL-induced osteoclastogenesis and lupus IgG-induced signaling transduction. We reviewed the results of studies that analyzed the association between FcγRs and bone erosion in inflammatory arthritis. The analysis revealed the dual roles of FcγRs in bone destruction in inflammatory arthritis. Thus, IgG/FcγR signaling molecules may serve as potential therapeutic targets against bone erosion.
Assuntos
Artrite/metabolismo , Remodelação Óssea , Osso e Ossos/metabolismo , Osteoclastos/metabolismo , Osteogênese , Receptores de IgG/metabolismo , Animais , Anti-Inflamatórios/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Artrite/tratamento farmacológico , Artrite/imunologia , Artrite/patologia , Remodelação Óssea/efeitos dos fármacos , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/imunologia , Osso e Ossos/patologia , Humanos , Imunoglobulina G/metabolismo , Imunoterapia , Osteoclastos/efeitos dos fármacos , Osteoclastos/imunologia , Osteoclastos/patologia , Osteogênese/efeitos dos fármacos , Ligante RANK/metabolismo , Receptores de IgG/antagonistas & inibidores , Receptores de IgG/imunologia , Transdução de SinaisRESUMO
Cannabis sativa is widely used for medical purposes and has anti-inflammatory activity. This study intended to examine the anti-inflammatory activity of cannabis on immune response markers associated with coronavirus disease 2019 (COVID-19) inflammation. An extract fraction from C. sativa Arbel strain (FCBD) substantially reduced (dose dependently) interleukin (IL)-6 and -8 levels in an alveolar epithelial (A549) cell line. FCBD contained cannabidiol (CBD), cannabigerol (CBG) and tetrahydrocannabivarin (THCV), and multiple terpenes. Treatments with FCBD and a FCBD formulation using phytocannabinoid standards (FCBD:std) reduced IL-6, IL-8, C-C Motif Chemokine Ligands (CCLs) 2 and 7, and angiotensin I converting enzyme 2 (ACE2) expression in the A549 cell line. Treatment with FCBD induced macrophage (differentiated KG1 cell line) polarization and phagocytosis in vitro, and increased CD36 and type II receptor for the Fc region of IgG (FcγRII) expression. FCBD treatment also substantially increased IL-6 and IL-8 expression in macrophages. FCBD:std, while maintaining anti-inflammatory activity in alveolar epithelial cells, led to reduced phagocytosis and pro-inflammatory IL secretion in macrophages in comparison to FCBD. The phytocannabinoid formulation may show superior activity versus the cannabis-derived fraction for reduction of lung inflammation, yet there is a need of caution proposing cannabis as treatment for COVID-19.
Assuntos
Anti-Inflamatórios/farmacologia , COVID-19/imunologia , Canabinoides/farmacologia , Cannabis/química , Células Epiteliais/imunologia , Macrófagos/imunologia , Extratos Vegetais/farmacologia , SARS-CoV-2/imunologia , Células A549 , Enzima de Conversão de Angiotensina 2/imunologia , Anti-Inflamatórios/química , COVID-19/patologia , Canabinoides/química , Citocinas/imunologia , Células Epiteliais/patologia , Células Epiteliais/virologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Humanos , Macrófagos/patologia , Macrófagos/virologia , Extratos Vegetais/química , Receptores de IgG/imunologia , Tratamento Farmacológico da COVID-19RESUMO
The pathogenesis of recurrent acute tonsillitis (Rtn) is to be further investigated. Polymorphonuclear neutrophils (PMN) often associate with the pathogenesis of acute and chronic inflammation. This study aims to identify the antigen-specific PMNs (sPMNs) isolated from the tonsillar tissues with recurrent acute inflammation. In this study, CD66b+ PMNs were isolated from surgically removed tonsils (40 tonsils were from 20 Rtn patients; 24 tonsils were from 12 tonsil tumour patients) by flow cytometry cell sorting. sPMNs were identified through immunological approaches. We found that compared with the control tonsil samples (from marginal non-tumour tissues of tonsil cancer), Rtn samples showed higher PMN frequency, higher levels of myeloperoxidase (MPO) and neutrophil elastase (NE), in which positive correlation was detected between the inflammatory scores in the Rtn tissues and PMN counts (r = .7352; p = .0002), or MPO (r = .6565, p = .0017), or NE (r = .6687, p = .0013). Upon exposure to tonsillar tissue protein extracts in the culture, a portion of Rtn PMNs was activated and released inflammatory mediators. A complex of tonsillar tissue-specific IgG and FcγRI was observed on the surface of Rtn PMNs; these PMNs could specifically recognize the Rtn tissue extracts and were designated the tonsillar antigen-specific PMNs (sPMNs). A signal transduction pathway of mitogen-activated protein kinase (MAPK)-nuclear factor of T cell activation (NFAT) was activated in sPMNs after exposure to Rtn tissue extracts. In summary, a fraction of sPMN in the Rtn tonsillar tissues was identified and characterized. The sPMNs can be activated upon exposure to tonsil-specific antigens. These sPMNs may contribute to the Rtn pathogenesis.
Assuntos
Antígenos/imunologia , Neutrófilos/imunologia , Tonsila Palatina/imunologia , Tonsilite/imunologia , Adolescente , Adulto , Idoso , Animais , Extratos Celulares/imunologia , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Neutrófilos/efeitos dos fármacos , Tonsila Palatina/efeitos dos fármacos , Peroxidase/metabolismo , Receptores de IgG/imunologia , Recidiva , Adulto JovemRESUMO
Myelodysplastic syndrome (MDS) is a clonal heterogeneous stem cell disorder driven by multiple genetic and epigenetic alterations resulting in ineffective hematopoiesis. MDS has a high frequency of immune suppressors, including myeloid-derived suppressor cells (MDSCs), that collectively result in a poor immune response. MDSCs in MDS patients express CD155 that ligates the T-cell immunoreceptor with immunoglobulin and ITIM domain (TIGIT) and delivers an inhibitory signal to natural killer (NK) cells. To mediate a productive immune response against MDS, negative regulatory checkpoints, like TIGIT, expressed on MDS NK cells must be overcome. NK cells can be directed to lyse MDS cells by bispecific killer engagers (BiKEs) that ligate CD16 on NK cells and CD33 on MDS cells. However, such CD16 × CD33 (1633) BiKEs do not induce the proliferative response in MDS NK cells needed to sustain their function. Here, we show that the addition of an NK stimulatory cytokine, interleukin-15 (IL-15), into the BiKE platform leads to productive IL-15 signaling without TIGIT upregulation on NK cells from MDS patients. Lower TIGIT expression allowed NK cells to resist MDSC inhibition. When compared with 1633 BiKE, 161533 trispecific killer engager (TriKE)-treated NK cells demonstrated superior killing kinetics associated with increased STAT5 phosphorylation. Furthermore, 161533 TriKE-treated MDS NK cells had higher proliferation and enhanced NK-cell function than 1633 BiKE-treated cells without the IL-15 linker. Collectively, our data demonstrate novel characteristics of the 161533 TriKE that support its application as an immunotherapeutic agent for MDS patients.
Assuntos
Células Matadoras Naturais/efeitos dos fármacos , Síndromes Mielodisplásicas/patologia , Células Supressoras Mieloides/patologia , Adulto , Anticorpos/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Proteínas Ligadas por GPI/imunologia , Células HL-60 , Humanos , Células Matadoras Naturais/imunologia , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/tratamento farmacológico , Síndromes Mielodisplásicas/imunologia , Células Supressoras Mieloides/imunologia , Receptores de IgG/imunologia , Receptores Imunológicos/metabolismo , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/imunologia , Células Tumorais Cultivadas , Adulto JovemRESUMO
BACKGROUND: Osteoclast-mediated bone erosion is a central feature of rheumatoid arthritis (RA). Immune complexes, present in a large percentage of patients, bind to Fcγ receptors (FcγRs), thereby modulating the activity of immune cells. In this study, we investigated the contribution of FcγRs, and FcγRIV in particular, during antigen-induced arthritis (AIA). METHODS: AIA was induced in knee joints of wild-type (WT), FcγRI,II,III-/-, and FcγRI,II,III,IV-/- mice. Bone destruction, numbers of tartrate-resistant acid phosphatase-positive (TRAP+) osteoclasts, and inflammation were evaluated using histology; expression of the macrophage marker F4/80, neutrophil marker NIMPR14, and alarmin S100A8 was evaluated using immunohistochemistry. The percentage of osteoclast precursors in the bone marrow was determined using flow cytometry. In vitro osteoclastogenesis was evaluated with TRAP staining, and gene expression was assessed using real-time PCR. RESULTS: FcγRI,II,III,IV-/- mice showed decreased bone erosion compared with WT mice during AIA, whereas both the humoral and cellular immune responses against methylated bovine serum albumin were not impaired in FcγRI,II,III,IV-/- mice. The percentage of osteoclast precursors in the bone marrow of arthritic mice and their ability to differentiate into osteoclasts in vitro were comparable between FcγRI,II,III,IV-/- and WT mice. In line with these observations, numbers of TRAP+ osteoclasts on the bone surface during AIA were comparable between the two groups. Inflammation, a process that strongly activates osteoclast activity, was reduced in FcγRI,II,III,IV-/- mice, and of note, mainly decreased numbers of neutrophils were present in the joint. In contrast to FcγRI,II,III,IV-/- mice, AIA induction in knee joints of FcγRI,II,III-/- mice resulted in increased bone erosion, inflammation, and numbers of neutrophils, suggesting a crucial role for FcγRIV in the joint pathology by the recruitment of neutrophils. Finally, significant correlations were found between bone erosion and the number of neutrophils present in the joint as well as between bone erosion and the number of S100A8-positive cells, with S100A8 being an alarmin strongly produced by neutrophils that stimulates osteoclast resorbing activity. CONCLUSIONS: FcγRs play a crucial role in the development of bone erosion during AIA by inducing inflammation. In particular, FcγRIV mediates bone erosion in AIA by inducing the influx of S100A8/A9-producing neutrophils into the arthritic joint.
Assuntos
Artrite Experimental/imunologia , Osso e Ossos/imunologia , Calgranulina A/imunologia , Calgranulina B/imunologia , Neutrófilos/imunologia , Receptores de IgG/imunologia , Animais , Artrite Experimental/genética , Artrite Experimental/metabolismo , Osso e Ossos/metabolismo , Osso e Ossos/patologia , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Articulação do Joelho/imunologia , Articulação do Joelho/metabolismo , Articulação do Joelho/patologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/metabolismo , Osteoclastos/imunologia , Osteoclastos/metabolismo , Receptores de IgG/genética , Receptores de IgG/metabolismo , Fosfatase Ácida Resistente a Tartarato/imunologia , Fosfatase Ácida Resistente a Tartarato/metabolismoRESUMO
OBJECTIVE: To study the correlation between the expression of Fcgamma receptor II b (FcgammaRII b) on B cells and rheumatoid arthritis (RA) patients of Shen deficiency syndrome (SDS). METHODS: There were 43 RA patients, including 26 of SDS and 17 of non-SDS. The expression levels of FcgammaRII b on naive B cells, memory B cells, and plasma blasts in the peripheral blood were detected by flow cytometry. The numbers of tender joints, numbers of swollen joints, erythrocyte sedimentation rate (ESR), rheumatoid factor (RF), and disease activity score (DAS28), the correlation between the distribution of B cells and the expression level of FcgammaRII b in RA patients were analyzed. Besides, another 21 healthy volunteers were recruited as the control group. RESULTS: The expression level of FcgammaRII b was 49.65% +/- 15.86% on memory B cells and 43.69% +/- 22.57% on plasma blasts in RA patients of SDS, significantly down-regulated when compared with those of the control group (64.03% +/- 6.01%, 66.59% +/- 10.18%, P < 0.01). The expression level of FcgammaRII b on memory B cells of RA patients of non-SDS was down-regulated more obviously when compared with that of the control group (52.70% +/- 9.52% versus 64.03% +/- 6.01%, P < 0.01). The expression level of FcgammaRII b on plasma blasts was obviously lower in RA patients of SDS than in RA patients of non-SDS (56.10% +/- 17.05%, P < 0.05). The expression level of FcgammaRII b on memory B cells was not correlated with numbers of tender joints, numbers of swollen joints, ESR, RF, or DAS28. CONCLUSIONS: The defective immunological tolerance of B cells in RA patients of SDS might be closely correlated with down-regulation of FcgammaRII b on memory B cells and plasma blasts. There might exist genetic abnormality of FcgammaRII b gene in RA patients of SDS, thus inducing loss of autoimmunity tolerance.
Assuntos
Artrite Reumatoide/sangue , Artrite Reumatoide/imunologia , Linfócitos B/metabolismo , Receptores de IgG/metabolismo , Adulto , Artrite Reumatoide/diagnóstico , Linfócitos B/imunologia , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Medicina Tradicional Chinesa , Pessoa de Meia-Idade , Receptores de IgG/imunologiaRESUMO
Allergen-IgE complexes are more efficiently internalized and presented by B cells than allergens alone. It has been suggested that IgG Abs induced by immunotherapy inhibit these processes. Food-allergic patients have high allergen-specific IgG levels. However, the role of these Abs in complex formation and binding to B cells is unknown. To investigate this, we incubated sera of peanut- or cow's milk-allergic patients with their major allergens to form complexes and added them to EBV-transformed or peripheral blood B cells (PBBCs). Samples of birch pollen-allergic patients were used as control. Complex binding to B cells in presence or absence of blocking Abs to CD23, CD32, complement receptor 1 (CR1, CD35), and/or CR2 (CD21) was determined by flow cytometry. Furthermore, intact and IgG-depleted sera were compared. These experiments showed that allergen-Ab complexes formed in birch pollen, as well as food allergy, contained IgE, IgG1, and IgG4 Abs and bound to B cells. Binding of these complexes to EBV-transformed B cells was completely mediated by CD23, whereas binding to PBBCs was dependent on both CD23 and CR2. This reflected differential receptor expression. Upon IgG depletion, allergen-Ab complexes bound to PBBCs exclusively via CD23. These data indicated that IgG Abs are involved in complex formation. The presence of IgG in allergen-IgE complexes results in binding to B cells via CR2 in addition to CD23. The binding to both CR2 and CD23 may affect Ag processing and presentation, and (may) thereby influence the allergic response.
Assuntos
Complexo Antígeno-Anticorpo/imunologia , Linfócitos B/imunologia , Hipersensibilidade Alimentar/imunologia , Imunoglobulina G/imunologia , Adolescente , Adulto , Idoso , Alérgenos/imunologia , Animais , Complexo Antígeno-Anticorpo/metabolismo , Linfócitos B/metabolismo , Betula/imunologia , Linhagem Celular , Ativação do Complemento/imunologia , Hipersensibilidade Alimentar/metabolismo , Humanos , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Imunoglobulina G/sangue , Camundongos , Pessoa de Meia-Idade , Pólen/imunologia , Ligação Proteica/imunologia , Receptores de Complemento/imunologia , Receptores de Complemento/metabolismo , Receptores de Complemento 3b/imunologia , Receptores de Complemento 3b/metabolismo , Receptores de Complemento 3d/imunologia , Receptores de Complemento 3d/metabolismo , Receptores de IgG/imunologia , Receptores de IgG/metabolismo , Rinite Alérgica Sazonal/imunologia , Rinite Alérgica Sazonal/metabolismo , Adulto JovemRESUMO
BACKGROUND: Conventional immunotherapy for peanut allergy using crude peanut extracts is not recommended because of the unacceptably high risk of anaphylaxis. Allergen-specific immunotherapy is not currently undertaken for peanut allergy. OBJECTIVES: The objective of this study was to develop a novel peanut-human fusion protein to block peanut-induced anaphylaxis. METHODS: We genetically designed and expressed a novel plant-human fusion protein composed of the major peanut allergen Ara h 2 and human IgG Fcγ1. We tested the Ara h 2-Fcγ fusion protein (AHG2)'s function in purified human basophils. Transgenic mice expressing human FcεRIα and a murine peanut allergy model were used. RESULTS: AHG2 inhibited histamine release induced by whole peanut extract (WPE) from basophils of patients with peanut allergy, whereas the fusion protein itself did not induce mediator release. AHG2 inhibited the WPE-induced, peanut-specific, IgE-mediated passive cutaneous anaphylaxis in hFcεRIα transgenic mice. AHG2 also significantly inhibited acute anaphylactic reactivity, including the prototypical decrease in body temperature in WPE-sensitized mice challenged with crude peanut extract. Histologic evaluation of the airways showed that AHG2 decreased peanut-induced inflammation, whereas the fusion protein itself did not induce airway inflammation in peanut-sensitized mice. AHG2 did not exert an inhibitory effect in mice lacking FcγRII. CONCLUSION: AHG2 inhibited peanut-specific IgE-mediated allergic reactions in vitro and in vivo. Linking specific peanut allergen to Fcγ can provide a new approach for the allergen immunotherapy of peanut allergy.
Assuntos
Albuminas 2S de Plantas/imunologia , Antígenos de Plantas/imunologia , Dessensibilização Imunológica , Glicoproteínas/imunologia , Hipersensibilidade a Amendoim/imunologia , Hipersensibilidade a Amendoim/prevenção & controle , Receptores de IgG/imunologia , Proteínas Recombinantes de Fusão/imunologia , Albuminas 2S de Plantas/química , Albuminas 2S de Plantas/genética , Alérgenos/imunologia , Anafilaxia/imunologia , Anafilaxia/prevenção & controle , Animais , Antígenos de Plantas/química , Antígenos de Plantas/genética , Arachis/imunologia , Basófilos/imunologia , Basófilos/metabolismo , Degranulação Celular/imunologia , Modelos Animais de Doenças , Feminino , Glicoproteínas/química , Glicoproteínas/genética , Liberação de Histamina/imunologia , Humanos , Imunoglobulina E/imunologia , Inflamação/imunologia , Inflamação/prevenção & controle , Camundongos , Camundongos Transgênicos , Hipersensibilidade a Amendoim/genética , Extratos Vegetais/imunologia , Receptores de IgG/química , Receptores de IgG/genética , Proteínas Recombinantes de Fusão/química , Sistema Respiratório/imunologiaRESUMO
INTRODUCTION: The effector functions of immunoglobulin G (IgG) are mediated by interaction of its Fc region with Fc receptors (FcγRs) and/or the complement system. The three main pathways of complement activation converge at C3. However, C3-independent pathways can activate C5 and other downstream complement components during IgG-initiated inflammatory responses. These C3-independent pathways of C5 activation are triggered by activating FcγRs in some systems or can be activated by factors of the coagulation cascade such as thrombin. Here we studied the interplay of C3, C5, and activating FcγRs in a model of spontaneous autoantibody-driven arthritis. METHODS: We utilized the K/BxN TCR transgenic mouse model of arthritis. We bred K/BxN mice bearing targeted or naturally-occurring mutations in one or more of the genes encoding complement components C3, C5, and FcRγ, the cytoplasmic signaling chain shared by the activating FcγRs. We measured arthritis development, the production of arthritogenic autoantibodies, T cell activation status and cytokine synthesis. In addition, we treated mice with anti-C5 monoclonal antibodies or with the thrombin inhibitor argatroban. RESULTS: We have previously shown that genetic deficiency of C5 protects K/BxN mice from the development of arthritis. We found here that C3-deficient K/BxN mice developed arthritis equivalent in severity to C3-sufficient animals. Arthritis also developed normally in K/BxN mice lacking both C3 and FcRγ, but could be ameliorated in these animals by treatment with anti-C5 monoclonal antibody or by treatment with argatroban. Production of arthritogenic autoantibodies, T cell activation, and T cell cytokine production were not affected by the absence of C3, C5, and/or FcRγ. CONCLUSIONS: In K/BxN mice, C5-dependent autoantibody-driven arthritis can occur in the genetic absence of both complement C3 and activating FcγRs. Our findings suggest that in this setting, thrombin activates C5 to provoke arthritis.
Assuntos
Artrite/imunologia , Autoanticorpos/imunologia , Complemento C3/imunologia , Complemento C5/imunologia , Receptores de IgG/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Antitrombinas/farmacologia , Arginina/análogos & derivados , Artrite/genética , Artrite/metabolismo , Autoanticorpos/sangue , Coagulação Sanguínea/efeitos dos fármacos , Coagulação Sanguínea/imunologia , Ativação do Complemento/efeitos dos fármacos , Ativação do Complemento/imunologia , Complemento C3/deficiência , Complemento C3/genética , Complemento C5/deficiência , Complemento C5/genética , Feminino , Citometria de Fluxo , Glucose-6-Fosfato Isomerase/imunologia , Imuno-Histoquímica , Masculino , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Transgênicos , Ácidos Pipecólicos/farmacologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de IgG/deficiência , Receptores de IgG/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , SulfonamidasRESUMO
AIM: The importance of the alteration of tumor infiltrative lymphocytes (CD4(+), CD8(+), CD16(+), and CD56(+)) in colorectal cancer prognosis is well known. In this study, we analyzed the effect of preoperative immunonutrition and different nutritional models on the clinical condition of colorectal cancer patients. METHODS: Twenty-eight colorectal cancer patients were grouped into 4 groups according to their nutrition regimens randomly and were given immunonutrition (IMN), standard enteral (SE), total parental nutrition (TPN), and normal nutrition (NN) regimens, all of which contained the same calorie-nitrogen content within a 7-day preoperative period. All patients had an endoscopic biopsy before and after the regimen, and the lymphocyte population infiltrating mucosal parts of the resected tumor tissue were evaluated. Immunohistochemical analysis of the tissue specimens was performed by staining with antihuman CD4(+), CD8(+), CD16(+), and CD56(+) antibodies. RESULTS: After nutrition, there were significant increases in each of the 4 groups of CD4(+) and CD8(+) cells within the tumor. Comparing the rates of augmentation, the increased rates of the CD8(+) cells infiltrating the tumor after nutrition in the patients who were fed with IMN were significantly more than the ones in other groups (P = .01). CD16(+) cell infiltration was significantly higher in all groups except the SE and IMN groups. The SE group had increased CD56(+) cell infiltration compared with the other groups. CONCLUSIONS: In the colorectal cancer patients who had nutrition in the 7-day preoperative period, except for the SE nutrition group, there were significant increases of infiltration of CD56(+) cells at the mucosal part of the tumor tissue within the CD4(+) and CD8(+) cell population. When postnutrition values were compared, there was a marked increase of CD8(+) cells in the IMN group.
Assuntos
Neoplasias Colorretais/imunologia , Neoplasias Colorretais/terapia , Nutrição Enteral , Alimentos Formulados , Linfócitos do Interstício Tumoral/imunologia , Nutrição Parenteral Total , Idoso , Arginina/administração & dosagem , Arginina/imunologia , Biópsia , Linfócitos T CD4-Positivos/imunologia , Antígeno CD56/imunologia , Linfócitos T CD8-Positivos/imunologia , Colonoscopia , Proteínas Alimentares/administração & dosagem , Proteínas Alimentares/imunologia , Ácidos Graxos Ômega-3/administração & dosagem , Ácidos Graxos Ômega-3/imunologia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Fenômenos Fisiológicos da Nutrição , Período Pré-Operatório , RNA/administração & dosagem , RNA/imunologia , Receptores de IgG/imunologia , TurquiaRESUMO
Fc receptors are a critical component of the innate immune system responsible for the recognition of cross-linked antibodies and the subsequent clearance of pathogens. However, in autoimmune diseases, these receptors play a role in the deleterious action of self-directed antibodies and as such are candidate targets for treatment. GMA161 is an aglycosyl, humanized version of the murine antibody 3G8 that targets the human low-affinity Fcγ receptor III (CD16). As CD16 expression and sequence have high species specificity, preclinical assessments were conducted in mice transgenic for both isoforms of human CD16, CD16A, and CD16B. This transgenic mouse model was useful in transitioning into phase I clinical trials, as it generated positive efficacy data in a relevant disease model and an acceptable single-dose safety profile. However, when GMA161 or its murine parent 3G8 were dosed repeatedly in transgenic mice having both human CD16 isoforms, severe reactions were observed that were not associated with significant cytokine release, nor were they alleviated by antihistamine administration. Prophylactic dosing with an inhibitor of platelet-activating factor (PAF), however, completely eliminated all signs of hypersensitivity. These findings suggest that (1) GMA161 elicits a reaction that is target dependent, (2) immunogenicity and similar adverse reactions were observed with a murine version of the antibody, and (3) the reaction is driven by the atypical hypersensitivity pathway mediated by PAF.
Assuntos
Anticorpos Monoclonais Humanizados , Doenças Autoimunes/tratamento farmacológico , Receptores de IgG/antagonistas & inibidores , Animais , Anticorpos Monoclonais Humanizados/efeitos adversos , Anticorpos Monoclonais Humanizados/imunologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Plaquetas/efeitos dos fármacos , Plaquetas/imunologia , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Feminino , Citometria de Fluxo , Proteínas Ligadas por GPI/antagonistas & inibidores , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Humanos , Imunoglobulina G/sangue , Contagem de Leucócitos , Masculino , Camundongos , Camundongos Transgênicos , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Fator de Ativação de Plaquetas/antagonistas & inibidores , Contagem de Plaquetas , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Receptores de IgG/genética , Receptores de IgG/imunologiaRESUMO
Mammalian cell culture systems are used predominantly for the production of therapeutic monoclonal antibody (mAb) products. A number of alternative platforms, such as Pichia engineered with a humanized N-linked glycosylation pathway, have recently been developed for the production of mAbs. The glycosylation profiles of mAbs produced in glycoengineered Pichia are similar to those of mAbs produced in mammalian systems. This report presents for the first time the comprehensive characterization of an anti-human epidermal growth factor receptor 2 (HER2) mAb produced in a glycoengineered Pichia, and a study comparing the anti-HER2 from Pichia, which had an amino acid sequence identical to trastuzumab, with trastuzumab. The comparative study covered a full spectrum of preclinical evaluation, including bioanalytical characterization, in vitro biological functions, in vivo anti-tumor efficacy and pharmacokinetics in both mice and non-human primates. Cell signaling and proliferation assays showed that anti-HER2 from Pichia had antagonist activities comparable to trastuzumab. However, Pichia-produced material showed a 5-fold increase in binding affinity to FcγIIIA and significantly enhanced antibody dependant cell-mediated cytotoxicity (ADCC) activity, presumably due to the lack of fucose on N-glycans. In a breast cancer xenograft mouse model, anti-HER2 was comparable to trastuzumab in tumor growth inhibition. Furthermore, comparable pharmacokinetic profiles were observed for anti-HER2 and trastuzumab in both mice and cynomolgus monkeys. We conclude that glycoengineered Pichia provides an alternative production platform for therapeutic mAbs and may be of particular interest for production of antibodies for which ADCC is part of the clinical mechanism of action.
Assuntos
Anticorpos Monoclonais Humanizados/imunologia , Anticorpos Monoclonais/imunologia , Pichia/genética , Receptor ErbB-2/imunologia , Proteínas Recombinantes/imunologia , Animais , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/metabolismo , Afinidade de Anticorpos/imunologia , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Área Sob a Curva , Ligação Competitiva/imunologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Fucose/metabolismo , Engenharia Genética , Humanos , Macaca fascicularis , Camundongos , Camundongos Endogâmicos C57BL , Pichia/metabolismo , Polissacarídeos/metabolismo , Ligação Proteica/imunologia , Receptores de IgG/imunologia , Receptores de IgG/metabolismo , Proteínas Recombinantes/farmacocinética , Proteínas Recombinantes/farmacologia , Trastuzumab , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
OBJECTIVE: The levels of both Fcγ receptor (FcγR) and the alarmins S100A8 and S100A9 are correlated with the development and progression of cartilage destruction during antigen-induced arthritis (AIA). This study was undertaken to study the active involvement of S100A8, S100A9, and S100A8/S100A9 in FcγR regulation in murine macrophages and synovium during AIA. METHODS: Recombinant murine S100A8 (rS100A8) was injected into normal mouse knee joints, and the synovium was isolated for analysis of FcγR messenger RNA (mRNA) expression by reverse transcription-polymerase chain reaction (RT-PCR). Macrophages, including bone marrow macrophages derived from Toll-like receptor 4-deficient (TLR-4(-/-)) mice, and polymorphonuclear cells (PMNs) were stimulated with S100 proteins, and levels of FcγR mRNA and protein were measured using RT-PCR and fluorescence-activated cell sorting analyses. AIA was induced in the knee joints of S100A9-deficient (S100A9(-/-)) mice, compared with wild-type (WT) controls, and the extent of cartilage destruction was determined using immunohistochemical analysis. RESULTS: Intraarticular injection of rS100A8 into the knee joints of normal mice caused a strong up-regulation of mRNA levels of activating FcγRI (64-fold increase) and FcγRIV (256-fold increase) in the synovium. Stimulation of macrophages with rS100A8 led to significant up-regulation of mRNA and protein levels of FcγRI and FcγRIV, but not FcγRIII, while the effects of S100A9 or S100A8/S100A9 complexes were less potent. Stimulation of PMNs (32Dcl3 cell line) with S100 proteins had no effect on FcγR expression. Up-regulation of FcγRI and FcγRIV was abrogated in rS100A8-stimulated macrophages from TLR-4(-/-) mice, indicating that the induction of FcγR expression by S100A8 is mediated by TLR-4. FcγR expression in the inflamed synovium of S100A9(-/-) mice was significantly lower on day 14 after arthritis induction when compared with WT controls, and these findings correlated with reduced severity of matrix metalloproteinase-mediated cartilage destruction. CONCLUSION: S100A8 is a strong promoter of activating FcγRI and FcγRIV in macrophages through the activation of TLR-4, and acts as a regulator of FcγR expression in inflamed synovium in chronic experimental arthritis.
Assuntos
Artrite Experimental/metabolismo , Calgranulina A/metabolismo , Macrófagos/metabolismo , Receptores de IgG/metabolismo , Membrana Sinovial/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Artrite Experimental/genética , Artrite Experimental/imunologia , Calgranulina A/administração & dosagem , Calgranulina A/imunologia , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/imunologia , Cartilagem Articular/metabolismo , Ensaio de Imunoadsorção Enzimática , Injeções Intra-Articulares , Articulação do Joelho/efeitos dos fármacos , Articulação do Joelho/imunologia , Articulação do Joelho/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de IgG/genética , Receptores de IgG/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estatísticas não Paramétricas , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/imunologia , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Regulação para CimaRESUMO
The dendritic cell (DC) lineage encompasses a diverse population of cells with unique subtype-specific functions. In peripheral blood, four DC subsets have been identified based on their distinct expression of CD1c, CD141, CD16, and CD123, and these subpopulations exhibit functional properties in immune responses. However, their respective roles in allergic diseases, such as rhinitis, are unclear. In this study, we have performed comparative assessments of DC subset frequencies and investigated their FcεRI expression levels in patients with allergic rhinitis. We demonstrate that the frequencies of CD1c+ and CD141+ DCs are elevated in grass pollen-allergic subjects compared with healthy controls, irrespectively of allergen stimulation. Among the DC subsets, CD1c+ DCs expressed the highest levels of FcεRI mRNA, and a large proportion expressed surface FcεRI. Furthermore, the FcεRI expression levels were augmented upon allergen challenge. Thus our data suggest that CD1c+ DCs influence allergen-specific immune responses. Research on their functional properties in allergy is warranted for development of future immunotherapies targeting specialized DC subsets.
Assuntos
Células Sanguíneas/metabolismo , Células Dendríticas/metabolismo , Receptores de IgG/metabolismo , Rinite Alérgica Sazonal/imunologia , Alérgenos/efeitos adversos , Antígenos CD/metabolismo , Antígenos de Diferenciação/metabolismo , Células Sanguíneas/imunologia , Células Sanguíneas/patologia , Contagem de Células , Linhagem da Célula , Células Dendríticas/imunologia , Células Dendríticas/patologia , Humanos , Imunização , Imunofenotipagem , Análise em Microsséries , Phleum , Pólen/efeitos adversos , Receptores de IgG/genética , Receptores de IgG/imunologia , Rinite Alérgica Sazonal/fisiopatologiaRESUMO
OBJECTIVE: To exploit the physiologic Fcgamma receptor IIb (CD32B) inhibitory coupling mechanism to control B cell activation by constructing a novel bispecific diabody scaffold, termed a dual-affinity retargeting (DART) molecule, for therapeutic applications. METHODS: DART molecules were constructed by pairing an Fv region from a monoclonal antibody (mAb) directed against CD32B with an Fv region from a mAb directed against CD79B, the beta-chain of the invariant signal-transducing dimer of the B cell receptor complex. DART molecules were characterized physicochemically and for their ability to simultaneously bind the target receptors in vitro and in intact cells. The ability of the DART molecules to negatively control B cell activation was determined by calcium mobilization, by tyrosine phosphorylation of signaling molecules, and by proliferation and Ig secretion assays. A DART molecule specific for the mouse ortholog of CD32B and CD79B was also constructed and tested for its ability to inhibit B cell proliferation in vitro and to control disease severity in a collagen-induced arthritis (CIA) model. RESULTS: DART molecules were able to specifically bind and coligate their target molecules on the surface of B cells and demonstrated a preferential simultaneous binding to both receptors on the same cell. DART molecules triggered the CD32B-mediated inhibitory signaling pathway in activated B cells, which translated into inhibition of B cell proliferation and Ig secretion. A DART molecule directed against the mouse orthologs was effective in inhibiting the development of CIA in DBA/1 mice. CONCLUSION: This innovative bispecific antibody scaffold that simultaneously engages activating and inhibitory receptors enables novel therapeutic approaches for the treatment of rheumatoid arthritis and potentially other autoimmune and inflammatory diseases in humans.
Assuntos
Anticorpos Biespecíficos/farmacologia , Anticorpos Monoclonais/farmacologia , Linfócitos B/efeitos dos fármacos , Imunossupressores/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Receptores de IgG , Animais , Anticorpos Biespecíficos/imunologia , Anticorpos Biespecíficos/farmacocinética , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacocinética , Complexo Antígeno-Anticorpo/imunologia , Complexo Antígeno-Anticorpo/metabolismo , Artrite Experimental/tratamento farmacológico , Artrite Experimental/imunologia , Linfócitos B/citologia , Linfócitos B/imunologia , Antígenos CD79/imunologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Dimerização , Feminino , Humanos , Imunoglobulinas/metabolismo , Imunossupressores/imunologia , Imunossupressores/farmacocinética , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Knockout , Receptores de IgG/imunologia , Transdução de Sinais , Baço/citologia , Alicerces TeciduaisRESUMO
Allograft rejection is a predominantly Th1 immune response. In this study, we showed that obaculactone, a natural compound derived from citrus fruit, prolonged skin graft survival in mice when treated after but not before transplantation. Furthermore, obaculactone inhibited alloantigen-specific production of Th1 cytokine IFN-gamma as well as proinflammatory cytokine IL-2, TNFalpha and IL-6. In parallel, IL-10 production was markedly up-regulated. Obaculactone significantly enhanced the percentage of CD4(+)CD25(+)Foxp3(+) Treg cells in the CD4(+) splenocytes without any effect on their inhibitory function. In vitro and in vivo tests showed obaculactone down-regulated T-bet expression in Th1 effector cells. Taken together, the unique immunomodulatory properties might qualify obaculactone as a putative, therapeutic compound for the treatment of Th1-driven diseases, including transplant rejection.
Assuntos
Citrus/química , Regulação para Baixo/efeitos dos fármacos , Sobrevivência de Enxerto/efeitos dos fármacos , Limoninas/farmacologia , Proteínas com Domínio T/efeitos dos fármacos , Células Th1/efeitos dos fármacos , Animais , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/imunologia , Sobrevivência de Enxerto/imunologia , Interferon gama/metabolismo , Limoninas/análise , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Extratos Vegetais/análise , Extratos Vegetais/farmacologia , Receptores de IgG/imunologia , Transplante de Pele/imunologia , Baço/efeitos dos fármacos , Baço/patologia , Proteínas com Domínio T/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Células Th1/imunologia , Transplante HomólogoRESUMO
Allergic asthma is characterized by airway eosinophilia, increased mucin production and allergen-specific IgE. Fc gamma receptor IIb (FcgammaRIIb), an inhibitory IgG receptor, has recently emerged as a negative regulator of allergic diseases like anaphylaxis and allergic rhinitis. However, no studies to date have evaluated its role in allergic asthma. Our main objective was to study the role of FcgammaRIIb in allergic lung inflammation. We used a murine model of allergic airway inflammation. Inflammation was quantified by BAL inflammatory cells and airway mucin production. FcgammaRIIb expression was measured by qPCR and flow cytometry and the cytokines were quantified by ELISA. Compared to wild type animals, FcgammaRIIb deficient mice mount a vigorous allergic lung inflammation characterized by increased bronchoalveolar lavage fluid cellularity, eosinophilia and mucin content upon ragweed extract (RWE) challenge. RWE challenge in sensitized mice upregulated FcgammaRIIb in the lungs. Disruption of IFN-gamma gene abrogated this upregulation. Treatment of naïve mice with the Th1-inducing agent CpG DNA increased FcgammaRIIb expression in the lungs. Furthermore, treatment of sensitized mice with CpG DNA prior to RWE challenge induced greater upregulation of FcgammaRIIb than RWE challenge alone. These observations indicated that RWE challenge upregulated FcgammaRIIb in the lungs by IFN-gamma- and Th1-dependent mechanisms. RWE challenge upregulated FcgammaRIIb on pulmonary CD14+/MHC II+ mononuclear cells and CD11c+ cells. FcgammaRIIb deficient mice also exhibited an exaggerated RWE-specific IgE response upon sensitization when compared to wild type mice. We propose that FcgammaRIIb physiologically regulates allergic airway inflammation by two mechanisms: 1) allergen challenge mediates upregulation of FcgammaRIIb on pulmonary CD14+/MHC II+ mononuclear cells and CD11c+ cells by an IFN-gamma dependent mechanism; and 2) by attenuating the allergen specific IgE response during sensitization. Thus, stimulating FcgammaRIIb may be a therapeutic strategy in allergic airway disorders.
Assuntos
Asma/imunologia , Inflamação/imunologia , Pulmão/imunologia , Receptores de IgG/imunologia , Ambrosia/química , Animais , Asma/genética , Asma/metabolismo , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Antígeno CD11c/imunologia , Antígeno CD11c/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Expressão Gênica , Humanos , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Inflamação/genética , Inflamação/metabolismo , Interferon gama/deficiência , Interferon gama/genética , Interferon gama/imunologia , Receptores de Lipopolissacarídeos/imunologia , Receptores de Lipopolissacarídeos/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Mucinas/metabolismo , Extratos Vegetais/imunologia , Receptores de IgG/deficiência , Receptores de IgG/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Th2/imunologia , Células Th2/metabolismoAssuntos
Anticorpos Monoclonais/farmacologia , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antígenos CD20/imunologia , Antineoplásicos/farmacologia , Ativação do Complemento/efeitos dos fármacos , Depleção Linfocítica/métodos , Animais , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais Murinos , Citotoxicidade Celular Dependente de Anticorpos/genética , Antígenos CD20/metabolismo , Antineoplásicos/imunologia , Antineoplásicos/metabolismo , Ativação do Complemento/genética , Ativação do Complemento/imunologia , Complemento C1q/imunologia , Complemento C1q/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Regiões Constantes de Imunoglobulina/genética , Regiões Constantes de Imunoglobulina/imunologia , Mutação de Sentido Incorreto , Ligação Proteica/genética , Ligação Proteica/imunologia , Receptores de IgG/genética , Receptores de IgG/imunologia , RituximabRESUMO
Numerous autoimmune and inflammatory disorders stem from the dysregulation of hematopoietic cell activation. The activity of inositol lipid and protein tyrosine phosphatases, and the receptors that recruit them, is critical for prevention of these disorders. Balanced signaling by inhibitory and activating receptors is now recognized to be an important factor in tuning cell function and inflammatory potential. In this review, we provide an overview of current knowledge of membrane proximal events in signaling by inhibitory/regulatory receptors focusing on structural and functional characteristics of receptors and their effectors Src homology 2 (SH2) domain-containing tyrosine phosphatase 1 and SH2 domain-containing inositol 5-phosphatase-1. We review use of new strategies to identify novel regulatory receptors and effectors. Finally, we discuss complementary actions of paired inhibitory and activating receptors, using Fc gammaRIIA and Fc gammaRIIB regulation human basophil activation as a prototype.