RESUMO
The immune system ensures optimum T-effector (Teff) immune responses against invading microbes and tumor antigens while preventing inappropriate autoimmune responses against self-antigens with the help of T-regulatory (Treg) cells. Thus, Treg and Teff cells help maintain immune homeostasis through mutual regulation. While Tregs can contribute to tumor immune evasion by suppressing anti-tumor Teff response, loss of Treg function can result in Teff responses against self-antigens leading to autoimmune disease. Thus, loss of homeostatic balance between Teff/Treg cells is often associated with both cancer and autoimmunity. Co-stimulatory and co-inhibitory receptors, collectively known as co-signaling receptors, play an indispensable role in the regulation of Teff and Treg cell expansion and function and thus play critical roles in modulating autoimmune and anti-tumor immune responses. Over the past three decades, considerable efforts have been made to understand the biology of co-signaling receptors and their role in immune homeostasis. Mutations in co-inhibitory receptors such as CTLA4 and PD1 are associated with Treg dysfunction, and autoimmune diseases in mice and humans. On the other hand, growing tumors evade immune surveillance by exploiting co-inhibitory signaling through expression of CTLA4, PD1 and PDL-1. Immune checkpoint blockade (ICB) using anti-CTLA4 and anti-PD1 has drawn considerable attention towards co-signaling receptors in tumor immunology and created renewed interest in studying other co-signaling receptors, which until recently have not been as well studied. In addition to co-inhibitory receptors, co-stimulatory receptors like OX40, GITR and 4-1BB have also been widely implicated in immune homeostasis and T-cell stimulation, and use of agonistic antibodies against OX40, GITR and 4-1BB has been effective in causing tumor regression. Although ICB has seen unprecedented success in cancer treatment, autoimmune adverse events arising from ICB due to loss of Treg homeostasis poses a major obstacle. Herein, we comprehensively review the role of various co-stimulatory and co-inhibitory receptors in Treg biology and immune homeostasis, autoimmunity, and anti-tumor immunity. Furthermore, we discuss the autoimmune adverse events arising upon targeting these co-signaling receptors to augment anti-tumor immune responses.
Assuntos
Autoimunidade , Homeostase/imunologia , Neoplasias/imunologia , Transdução de Sinais/imunologia , Linfócitos T Reguladores/imunologia , Animais , Antineoplásicos Imunológicos/uso terapêutico , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Antígeno CTLA-4/genética , Antígeno CTLA-4/imunologia , Regulação da Expressão Gênica , Proteína Relacionada a TNFR Induzida por Glucocorticoide/genética , Proteína Relacionada a TNFR Induzida por Glucocorticoide/imunologia , Humanos , Camundongos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/patologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Receptores OX40/genética , Receptores OX40/imunologia , Linfócitos T Citotóxicos/imunologia , Evasão Tumoral , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/genética , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologiaRESUMO
Virtually all efforts to generate an effective protection against the life-long, recurrent genital infections caused by HSV-2 have failed. Apart from sexual transmission, the virus can also be transmitted from mothers to neonates, and it is a key facilitator of HIV coacquisition. In this article, we uncover a nanoimmunotherapy using specially designed zinc oxide tetrapod nanoparticles (ZOTEN) with engineered oxygen vacancies. We demonstrate that ZOTEN, when used intravaginally as a microbicide, is an effective suppressor of HSV-2 genital infection in female BALB/c mice. The strong HSV-2 trapping ability of ZOTEN significantly reduced the clinical signs of vaginal infection and effectively decreased animal mortality. In parallel, ZOTEN promoted the presentation of bound HSV-2 virions to mucosal APCs, enhancing T cell-mediated and Ab-mediated responses to the infection, and thereby suppressing a reinfection. We also found that ZOTEN exhibits strong adjuvant-like properties, which is highly comparable with alum, a commonly used adjuvant. Overall, to our knowledge, our study provides the very first evidence for the protective efficacy of an intravaginal microbicide/vaccine or microbivac platform against primary and secondary female genital herpes infections.
Assuntos
Herpes Genital/tratamento farmacológico , Herpes Genital/imunologia , Herpesvirus Humano 2/efeitos dos fármacos , Herpesvirus Humano 2/imunologia , Nanopartículas/administração & dosagem , Nanopartículas/uso terapêutico , Óxido de Zinco/administração & dosagem , Óxido de Zinco/uso terapêutico , Animais , Células Cultivadas , Chlorocebus aethiops , Feminino , Células HeLa , Herpes Genital/patologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Nanopartículas/química , Tamanho da Partícula , Relação Estrutura-Atividade , Células Vero , Óxido de Zinco/farmacologiaRESUMO
GM-CSF was originally identified as a colony stimulating factor (CSF) because of its ability to induce granulocyte and macrophage populations from precursor cells. Multiple studies have demonstrated that GM-CSF is also an immune-modulatory cytokine, capable of affecting not only the phenotype of myeloid lineage cells, but also T-cell activation through various myeloid intermediaries. This property has been implicated in the sustenance of several autoimmune diseases like arthritis and multiple sclerosis. In contrast, several studies using animal models have shown that GM-CSF is also capable of suppressing many autoimmune diseases such as Crohn's disease, Type-1 diabetes, Myasthenia gravis and experimental autoimmune thyroiditis. Knockout mouse studies have suggested that the role of GM-CSF in maintaining granulocyte and macrophage populations in the physiological steady state is largely redundant. Instead, its immune-modulatory role plays a significant role in the development or resolution of autoimmune diseases. This is mediated either through the differentiation of precursor cells into specialized non-steady state granulocytes, macrophages and dendritic cells, or through the modulation of the phenotype of mature myeloid cells. Thus, outside of myelopoiesis, GM-CSF has a profound role in regulating the immune response and maintaining immunological tolerance.
Assuntos
Doenças Autoimunes/imunologia , Autoimunidade/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Tolerância Imunológica/imunologia , Células Mieloides/citologia , Animais , Diferenciação Celular/imunologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Granulócitos/citologia , Granulócitos/imunologia , Humanos , Ativação Linfocitária/imunologia , Macrófagos/citologia , Macrófagos/imunologia , Camundongos , Linfócitos T Reguladores/imunologiaRESUMO
Earlier, we had demonstrated that treatment with low dose of GM-CSF can prevent the development of experimental autoimmune thyroiditis (EAT), experimental autoimmune myasthenia gravis, and type 1 diabetes, and could also reverse ongoing EAT and experimental autoimmune myasthenia gravis. The protective effect was mediated through the induction of tolerogenic CD11C(+)CD8α(-) dendritic cells (DCs) and consequent expansion of Foxp3(+) regulatory T cells (Tregs). Subsequently, we showed that GM-CSF acted specifically on bone marrow precursors and facilitated their differentiation into tolerogenic dendritic cells (DCs; GM-CSF-induced bone marrow-derived DCs [GM-BMDCs]), which directed Treg expansion in a contact-dependent manner. This novel mechanism of Treg expansion was independent of TCR-mediated signaling but required exogenous IL-2 and cosignaling from DC-bound OX40L. In this study, we observed that OX40L-mediated signaling by GM-BMDCs, although necessary, was not sufficient for Treg expansion and required signaling by Jagged1. Concurrent signaling induced by OX40L and Jagged1 via OX40 and Notch3 receptors expressed on Tregs was essential for the Treg expansion with sustained FoxP3 expression. Adoptive transfer of only OX40L(+)Jagged1(+) BMDCs led to Treg expansion, increased production of IL-4 and IL-10, and suppression of EAT in the recipient mice. These results showed a critical role for OX40L- and Jagged1-induced cosignaling in GM-BMDC-induced Treg expansion.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Células Dendríticas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Membrana/metabolismo , Ligante OX40/metabolismo , Transdução de Sinais , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Animais , Antígenos B7/imunologia , Antígenos B7/metabolismo , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Membrana Celular/imunologia , Membrana Celular/metabolismo , Células Dendríticas/efeitos dos fármacos , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Proteína Jagged-1 , Ligantes , Ativação Linfocitária , Camundongos , Receptores Notch/metabolismo , Proteínas Serrate-Jagged , Tireoidite Autoimune/imunologia , Tireoidite Autoimune/metabolismoRESUMO
To prepare a novel Bispecific Antibody (BsAb) as a potential targeted therapy for T1D, we produced a "functionally inert" monoclonal antibody (mAb) against Glucose transporter-2 (GLUT-2) expressed on ß-cells to serve as an anchoring antibody. The therapeutic arm is an agonistic mAb against Cytotoxic T-Lymphocyte Antigen 4 (CTLA-4), a negative regulator of T-cell activation expressed on activated CD4+ T-cells. A BsAb was prepared by chemically coupling an anti-GLUT2 mAb to an agonistic anti-CTLA-4 mAb. This BsAb was able to bind to GLUT2 and CTLA-4 in vitro, and to pancreatic islets, both in vitro and in vivo. We tested the safety and efficacy of this BsAb by treating Non-Obese Diabetes (NOD) mice and found that it could delay the onset of diabetes with no apparent undesirable side effects. Thus, engagement of CTLA-4 on activated T cells from target tissue can be an effective way to treat type-1 diabetes.
Assuntos
Anticorpos Biespecíficos/administração & dosagem , Anticorpos Biespecíficos/imunologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Células Secretoras de Insulina/imunologia , Animais , Anticorpos Biespecíficos/efeitos adversos , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Antígeno CTLA-4/imunologia , Linhagem Celular , Cricetinae , Citocinas/biossíntese , Modelos Animais de Doenças , Feminino , Transportador de Glucose Tipo 2/biossíntese , Transportador de Glucose Tipo 2/imunologia , Camundongos , Camundongos Endogâmicos NOD , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Resultado do TratamentoRESUMO
The enforcement of sphingosine-1-phosphate (S1P) signaling network protects from radiation-induced pneumonitis. We now demonstrate that, in contrast to early postirradiation period, late postirradiation sphingosine kinase-1 (SphK1) and sphingoid base-1-phosphates are associated with radiation-induced pulmonary fibrosis (RIF). Using the mouse model, we demonstrate that RIF is characterized by a marked upregulation of S1P and dihydrosphingosine-1-phosphate (DHS1P) levels in the lung tissue and in circulation accompanied by increased lung SphK1 expression and activity. Inhibition of sphingolipid de novo biosynthesis by targeting serine palmitoyltransferase (SPT) with myriocin reduced radiation-induced pulmonary inflammation and delayed the onset of RIF as evidenced by increased animal lifespan and decreased expression of markers of fibrogenesis, such as collagen and α-smooth muscle actin (α-SMA), in the lung. Long-term inhibition of SPT also decreased radiation-induced SphK activity in the lung and the levels of S1P-DHS1P in the lung tissue and in circulation. In vitro, inhibition or silencing of serine palmitoyltransferase attenuated transforming growth factor-ß1 (TGF-ß)-induced upregulation of α-SMA through the negative regulation of SphK1 expression in normal human lung fibroblasts. These data demonstrate a novel role for SPT in regulating TGF-ß signaling and fibrogenesis that is linked to the regulation of SphK1 expression and S1P-DHS1P formation.
Assuntos
Inibidores Enzimáticos/farmacologia , Ácidos Graxos Monoinsaturados/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fibrose Pulmonar/prevenção & controle , Lesões Experimentais por Radiação/prevenção & controle , Serina C-Palmitoiltransferase/antagonistas & inibidores , Animais , Feminino , Regulação Enzimológica da Expressão Gênica/efeitos da radiação , Humanos , Camundongos , Fibrose Pulmonar/enzimologia , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Lesões Experimentais por Radiação/enzimologia , Lesões Experimentais por Radiação/genética , Lesões Experimentais por Radiação/patologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Tórax/efeitos da radiação , Fatores de Tempo , Fator de Crescimento Transformador beta/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/efeitos da radiaçãoRESUMO
Ig class switch recombination (CSR) is regulated through long-range intrachromosomal interactions between germline transcript promoters and enhancers to initiate transcription and create chromatin accessible to activation-induced deaminase attack. CSR occurs between switch (S) regions that flank Cmu and downstream C(H) regions and functions via an intrachromosomal deletional event between the donor Smicro region and a downstream S region. It is unclear to what extent S region primary sequence influences differential targeting of CSR to specific isotypes. We address this issue in this study by generating mutant mice in which the endogenous Sgamma3 region was replaced with size-matched Sgamma1 sequence. B cell activation conditions are established that support robust gamma3 and gamma1 germline transcript expression and stimulate IgG1 switching but suppress IgG3 CSR. We found that the Sgamma1 replacement allele engages in micro-->gamma3 CSR, whereas the intact allele is repressed. We conclude that S region identity makes a significant contribution to CSR. We propose that the Sgamma1 region is selectively targeted for CSR following the induction of an isotype-specific factor that targets the S region and recruits CSR machinery.
Assuntos
Linfócitos B/imunologia , Switching de Imunoglobulina/genética , Animais , Sequência de Bases , Southern Blotting , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Pesadas de Imunoglobulinas/imunologia , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Dados de Sequência Molecular , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Every year, influenza virus infection causes significant mortality and morbidity in human populations. Although egg-based inactivated viral vaccines are available, their effectiveness depends on the correct prediction of the circulating viral strains and is limited by the time constraint of the manufacturing process. Recombinant subunit vaccines are easier to manufacture with a relatively short lead time but are limited in their efficacy partly because the purified recombinant membrane proteins in the soluble form most likely do not retain their native membrane-bound structure. Nanodisc (ND) particles are soluble, stable, and reproducibly prepared discoid shaped nanoscale structures that contain a discrete lipid bilayer bound by two amphipathic scaffold proteins. Because ND particles permit the functional reconstitution of membrane/envelope proteins, we incorporated recombinant hemagglutinin (HA) from influenza virus strain A/New Caledonia/20/99 (H1N1) into NDs and investigated their potential to elicit an immune response to HA and confer immunity to influenza virus challenge relative to the commercial vaccines Fluzone and FluMist. HA-ND vaccination induced a robust anti-HA antibody response consisting of predominantly the immunoglobulin G1 (IgG1) subclass and a high hemagglutination inhibition titer. Intranasal immunization with HA-ND induced an anti-HA IgA response in nasal passages. HA-ND vaccination conferred protection that was comparable to that of Fluzone and FluMist against challenge with influenza virus strain A/Puerto Rico/8/1934 (H1N1).
Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/administração & dosagem , Imunidade Humoral , Vacinas contra Influenza/imunologia , Nanopartículas/uso terapêutico , Infecções por Orthomyxoviridae/prevenção & controle , Animais , Anticorpos Antivirais/biossíntese , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/uso terapêutico , Humanos , Vírus da Influenza A Subtipo H1N1/imunologia , Proteínas de Membrana/uso terapêutico , Camundongos , Nanopartículas/química , Infecções por Orthomyxoviridae/imunologia , Fosfolipídeos/uso terapêuticoRESUMO
Regulatory T cells (Tregs) play a pivotal role in immune-tolerance, and loss of Treg function can lead to the development of autoimmunity. Natural Tregs generated in the thymus substantially contribute to the Treg pool in the periphery, where they suppress self-reactive effector T cells (Teff) responses. Recently, we showed that OX40L (TNFSF4) is able to drive selective proliferation of peripheral Tregs independent of canonical antigen presentation (CAP-independent) in the presence of low-dose IL-2. Therefore, we hypothesized that OX40 signaling might be integral to the TCR-independent phase of murine and human thymic Treg (tTreg) development. Development of tTregs is a two-step process: Strong T-cell receptor (TCR) signals in combination with co-signals from the TNFRSF members facilitate tTreg precursor selection, followed by a TCR-independent phase of tTreg development in which their maturation is driven by IL-2. Therefore, we investigated whether OX40 signaling could also play a critical role in the TCR-independent phase of tTreg development. OX40-/- mice had significantly reduced numbers of CD25-Foxp3low tTreg precursors and CD25+Foxp3+ mature tTregs, while OX40L treatment of WT mice induced significant proliferation of these cell subsets. Relative to tTeff cells, OX40 was expressed at higher levels in both murine and human tTreg precursors and mature tTregs. In ex vivo cultures, OX40L increased tTreg maturation and induced CAP-independent proliferation of both murine and human tTregs, which was mediated through the activation of AKT-mTOR signaling. These novel findings show an evolutionarily conserved role for OX40 signaling in tTreg development and proliferation, and might enable the development of novel strategies to increase Tregs and suppress autoimmunity.
Assuntos
Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Receptores OX40/metabolismo , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Timócitos/imunologia , Animais , Proliferação de Células , Células Cultivadas , Citocinas/metabolismo , Feminino , Fatores de Transcrição Forkhead/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais , Timócitos/citologiaRESUMO
We have previously shown GM-CSF derived bone-marrow dendritic cells (G-BMDCs) can induce the selective expansion of Tregs through the surface-bound molecule OX40L; however, the physiological role of this ex vivo derived DC subset remained to be elucidated. We determined GM-CSF administration to mice induced the generation of in vivo derived OX40L+ DCs, phenotypically similar to ex vivo OX40L+G-BMDCs, in the spleen, brachial lymph nodes and liver. The generation of OX40L+ DCs correlated with increased percentages of functionally suppressive Tregs in the spleen, brachial lymph nodes, and liver of GM-CSF treated mice. DCs from GM-CSF treated mice expanded Tregs in CD4+ T-cell co-cultures in an OX40L dependent manner, suggesting OX40L+ DCs may play a role in peripheral Treg homeostasis. Furthermore, comparing the transcriptome data of OX40L+ DCs to that of all immune cell types revealed OX40L+ DCs to be distinct from steady-state immune cells and, microarray analysis of OX40L+G-BMDCs and OX40L-G-BMDCs revealed higher expression of molecules that are associated with tolerogenic phenotype and could play important roles in the function of OX40L+ DCs. These findings suggest that OX40L+ DCs may represent a unique DC subset induced under inflammatory conditions that may play an essential role in maintaining Treg homeostasis.
Assuntos
Células Dendríticas/imunologia , Glicoproteínas de Membrana/imunologia , Linfócitos T Reguladores/imunologia , Fatores de Necrose Tumoral/imunologia , Animais , Células Cultivadas , Células Dendríticas/metabolismo , Feminino , Citometria de Fluxo , Inflamação/genética , Inflamação/imunologia , Glicoproteínas de Membrana/análise , Glicoproteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Ligante OX40 , Linfócitos T Reguladores/metabolismo , Transcriptoma , Fatores de Necrose Tumoral/análise , Fatores de Necrose Tumoral/genéticaRESUMO
We have previously shown that OX40L/OX40 interaction is critical for TCR-independent selective proliferation of Foxp3+ Tregs, but not Foxp3- effector T-cells (Teff), when CD4+ T-cells are co-cultured with GM-CSF derived bone marrow dendritic cells (G-BMDCs). Events downstream of OX40L/OX40 interaction in Tregs responsible for this novel mechanism are not understood. Earlier, OX40L/OX40 interaction has been shown to stimulate CD4+ T-cells through the formation of a signalosome involving TRAF2/PKC-Ѳ leading to NF-kB activation. In this study, using CD4+ T-cells from WT and OX40-/- mice we first established that OX40 mediated activation of NF-kB was critical for this Treg proliferation. Although CD4+ T-cells from PKC-Ѳ-/- mice were also defective in G-BMDC induced Treg proliferation ex vivo, this defect could be readily corrected by adding exogenous IL-2 to the co-cultures. Furthermore, by treating WT, OX40-/-, and PKC-Ѳ-/- mice with soluble OX40L we established that OX40L/OX40 interaction was required and sufficient to induce Treg proliferation in vivo independent of PKC-Ѳ status. Although PKC-Ѳ is dispensable for TCR-independent Treg proliferation per se, it is essential for optimum IL-2 production by Teff cells. Finally, our findings suggest that OX40L binding to OX40 likely results in recruitment of TRAF1 for downstream signalling.
Assuntos
Proliferação de Células , Interleucina-2/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteína Quinase C-theta/metabolismo , Receptores OX40/metabolismo , Linfócitos T Reguladores/fisiologia , Fatores de Necrose Tumoral/metabolismo , Animais , Camundongos , Camundongos Knockout , NF-kappa B/metabolismo , Ligante OX40 , Proteína Quinase C-theta/deficiência , Receptores OX40/deficiênciaRESUMO
Regulatory T-cells (Tregs) play a pivotal role in maintaining peripheral tolerance. Increasing Treg numbers/functions has been shown to ameliorate autoimmune diseases. However, common Treg expansion approaches use T-Cell Receptor (TCR)-mediated stimulation which also causes proliferation of effector T-cells (Teff). To overcome this limitation, purified patient-specific Tregs are expanded ex vivo and transfused. Although promising, this approach is not suitable for routine clinical use. Therefore, an alternative approach to selectively expand functional Tregs in vivo is highly desired. We report a novel TCR-independent strategy for the selective proliferation of Foxp3+Tregs (without Teff proliferation), by co-culturing CD4+ T-cells with OX40 L+Jagged(JAG)-1+ bone marrow-derived DCs differentiated with GM-CSF or treating them with soluble OX40 L and JAG1 in the presence of exogenous IL-2. Tregs expanded using soluble OX40 L and JAG1 were of suppressive phenotype and delayed the onset of diabetes in NOD mice. Ligation of OX40 L and JAG1 with their cognate-receptors OX40 and Notch3, preferentially expressed on Tregs but not on Teff cells, was required for selective Treg proliferation. Soluble OX40L-JAG1-induced NF-κB activation as well as IL-2-induced STAT5 activation were essential for the proliferation of Tregs with sustained Foxp3 expression. Altogether, these findings demonstrate the utility of soluble OX40 L and JAG1 to induce TCR-independent Treg proliferation.
Assuntos
Células Dendríticas/imunologia , Ligante OX40/metabolismo , Linfócitos T Reguladores/fisiologia , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Fatores de Transcrição Forkhead/metabolismo , Humanos , Interleucina-2/metabolismo , Proteína Jagged-1/metabolismo , NF-kappa B/metabolismo , Receptor Notch3/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores OX40/metabolismo , Transdução de SinaisRESUMO
Earlier, we have shown that GM-CSF derived bone marrow (BM) dendritic cells (G-BMDCs) can expand Foxp3(+) regulatory T-cells (Tregs) through a TCR-independent, but IL-2 dependent mechanism that required OX40L/OX40 interaction. While some reports have shown suppression of autoimmunity upon treatment with an OX40 agonist, others have shown exacerbation of autoimmune disease instead. To better understand the basis for these differing outcomes, we compared the effects of OX40L treatment in 6-week-old pre-diabetic and 12-week-old near diabetic NOD mice. Upon treatment with OX40L, 6-week-old NOD mice remained normoglycemic and showed a significant increase in Tregs in their spleen and lymph nodes, while 12-week-old NOD mice very rapidly developed hyperglycemia and failed to show Treg increase in spleen or LN. Interestingly, OX40L treatment increased Tregs in the thymus of both age groups. However, it induced Foxp3(+)CD103(+)CD38(-) stable-phenotype Tregs in the thymus and reduced the frequency of autoreactive Teff cells in 6-week-old mice; while it induced Foxp3(+)CD103(-)CD38(+) labile-phenotype Tregs in the thymus and increased autoreactive CD4(+) T cells in the periphery of 12-week-old mice. This increase in autoreactive CD4(+) T cells was likely due to either a poor suppressive function or conversion of labile Tregs into Teff cells. Using ex vivo cultures, we found that the reduction in Treg numbers in 12-week-old mice was likely due to IL-2 deficit, and their numbers could be increased upon addition of exogenous IL-2. The observed divergent effects of OX40L treatment were likely due to differences in the ability of 6- and 12-week-old NOD mice to produce IL-2.
Assuntos
Ligante de CD40/metabolismo , Diabetes Mellitus/etiologia , Diabetes Mellitus/metabolismo , Transferência Adotiva , Fatores Etários , Animais , Glicemia , Antígenos CD40/metabolismo , Ligante de CD40/administração & dosagem , Citocinas/genética , Citocinas/metabolismo , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Mediadores da Inflamação/metabolismo , Linfonodos/imunologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos NOD , Ligação Proteica , Baço/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismoRESUMO
Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hematopoietic growth factor, that has been used as a therapeutic agent in facilitating bone marrow and stem cell transplantation and in other clinical cases like neutropenia. Although biologically active recombinant GM-CSF has been successfully produced in Escherichia coli, the reported levels are extremely poor. In this study we looked into the possible reasons for poor expression and found that protein toxicity coupled with protease-based degradation was the principal reason for low productivity. To overcome this problem we attached a signal sequence, as well as an amino-terminal His-tag fusion to the GM-CSF gene. This combination had a dramatic effect on expression levels, which increased from 0.8 microg/mL in the control to 40 microg/mL. When a larger fusion partner, such as the Maltose-binding protein (MBP-tag), was used the expression levels increased further to 69.5 microg/mL, which along with the MBP-tag represented approx 12% of the total cellular protein.
Assuntos
Escherichia coli/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/biossíntese , Sinais Direcionadores de Proteínas , Proteínas de Transporte/genética , Códon/genética , Escherichia coli/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Microbiologia Industrial , Proteínas Ligantes de Maltose , Periplasma/química , Periplasma/metabolismo , Plasmídeos/genética , Estabilidade de RNA , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Proteínas RecombinantesRESUMO
Granulocyte macrophage colony stimulating factor (GM-CSF) is generally recognized as an inflammatory cytokine. Its inflammatory activity is primarily due its role as a growth and differentiation factor for granulocyte and macrophage populations. In this capacity, among other clinical applications, it has been used to bolster anti-tumor immune responses. GM-CSF-mediated inflammation has also been implicated in certain types of autoimmune diseases, including rheumatoid arthritis and multiple sclerosis. Thus, agents that can block GM-CSF or its receptor have been used as anti-inflammatory therapies. However, a review of literature reveals that in many situations GM-CSF can act as an anti-inflammatory/regulatory cytokine. We and others have shown that GM-CSF can modulate dendritic cell differentiation to render them "tolerogenic," which, in turn, can increase regulatory T-cell numbers and function. Therefore, the pro-inflammatory and regulatory effects of GM-CSF appear to depend on the dose and the presence of other relevant cytokines in the context of an immune response. A thorough understanding of the various immunomodulatory effects of GM-CSF will facilitate more appropriate use and thus further enhance its clinical utility.
Assuntos
Citocinas/fisiologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/fisiologia , Mediadores da Inflamação/fisiologia , Animais , Anti-Inflamatórios/metabolismo , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Doenças Autoimunes/tratamento farmacológico , Autoimunidade , Citocinas/farmacologia , Citocinas/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/uso terapêutico , Humanos , Tolerância Imunológica , Imunossupressores/metabolismo , Imunossupressores/farmacologia , Imunossupressores/uso terapêutico , Imunoterapia , Inflamação/tratamento farmacológico , Inflamação/genética , Inflamação/metabolismo , Mediadores da Inflamação/farmacologia , Mediadores da Inflamação/uso terapêuticoRESUMO
Adenoid cystic carcinoma is a relatively rare epithelial tumor of the salivary glands accounting for about 5-10% of all salivary gland neoplasms. Approximately, 31% of salivary gland neoplasms affect minor salivary glands particularly the palate. It involves tongue in only 19.8% of cases and even rarely the dorsum of the tongue. We report such a rare case that affected dorsum of the tongue in a 45-year-old-female patient.
RESUMO
Multilocular cystic renal cell carcinoma (MCRCC), also known as multilocular clear cell renal cell carcinoma (RCC), is a rare cystic tumor of the kidney with an excellent outcome. It occurs in about 3.1-6% of the conventional RCC. It is usually included in the group of tumors of undetermined malignant potential with low nuclear grade. We present a case of MCRCC in a 30-year-old female patient presenting incidentally as an apparently benign-looking multicystic space occupying lesion in the upper pole of right kidney. Right-sided simple nephrectomy was performed, and on histopathologic examination it was found to be MCRCC, stage 1 with Fuhrman nuclear grade 1. Immunohistochemistry with epithelial membrane antigen and vimentin confirmed the diagnosis.
RESUMO
It is difficult to predict which urothelial neoplasm would subsequently recur or progress to muscle invasive tumours or produce metastasis.The aim and objective of the study were to evaluate the scope of immunohistochemical expression of p53 in human urothelial neoplasms with regard to grade, stage and outcome of the patients. Eighteen consecutive patients were taken and urothelial tumour samples were obtained from transurethral resection or surgical excision. Histopathological examinations were performed and the grading was done according to the WHO/ISUP consensus classification of urothelial neoplasms. Immunohistochemical staining for p53 was performed on formalin fixed paraffin embedded tissue sections with appropriate positive and negative control. It was found 3 patients with papillary urothelial neoplasm of low malignant potential (PUNLMP), 5 cases of papillary low grade urothelial carcinoma, 10 patients with papillary high grade urothelial carcinoma including 2 cases of invasive urothelial carcinoma. All three PUNLMP cases showed negative results. Four out of 5 low grade papillary urothelial carcinoma had nuclear p53 accumulation, while all of the 10 papillary high grade carcinoma had high p53 index. The finding of negative p53 staining in PUNLMPs and high p53 index in high grade papillary urothelial carcinomas and invasive carcinomas support the notion that mutation of p53 gene might be unrelated to the development of urothelial neoplasms but definitely play a crucial role in progression of the malignancy.
Assuntos
Carcinoma de Células de Transição/genética , DNA de Neoplasias/genética , Mutação , Proteína Supressora de Tumor p53/genética , Neoplasias da Bexiga Urinária/genética , Carcinoma de Células de Transição/metabolismo , Carcinoma de Células de Transição/patologia , Análise Mutacional de DNA , Humanos , Imuno-Histoquímica/métodos , Reprodutibilidade dos Testes , Estudos Retrospectivos , Proteína Supressora de Tumor p53/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologiaRESUMO
In our earlier work, we had shown that GM-CSF treatment of CBA/J mice can suppress ongoing thyroiditis by inducing tolerogenic CD8α(-) DCs, which helped expand and/or induce CD4(+)Foxp3(+) Tregs. To identify the primary cell type that was affected by the GM-CSF treatment and understand the mechanism by which Tregs were induced, we compared the effect of GM-CSF on matured spDCs and BMDC precursors in vitro. Matured spDCs exposed to GM-CSF ex vivo induced only a modest increase in the percentage of Foxp3-expressing T cells in cocultures. In contrast, BM cells, when cultured in the presence of GM-CSF, gave rise to a population of CD11c(+)CD11b(Hi)CD8α(-) DCs (BMDCs), which were able to expand Foxp3(+) Tregs upon coculture with CD4(+) T cells. This contact-dependent expansion occurred in the absence of TCR stimulation and was abrogated by OX40L blockage. Additionally, the BMDCs secreted high levels of TGF-ß, which was required and sufficient for adaptive differentiation of T cells to Foxp3(+) Tregs, only upon TCR stimulation. These results strongly suggest that the BMDCs differentiated by GM-CSF can expand nTregs and induce adaptive Tregs through different mechanisms.
Assuntos
Imunidade Adaptativa , Células da Medula Óssea/imunologia , Células Dendríticas/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/fisiologia , Células Progenitoras Linfoides/imunologia , Linfócitos T Reguladores/imunologia , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Diferenciação Celular/imunologia , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Tolerância Imunológica , Contagem de Linfócitos , Células Progenitoras Linfoides/citologia , Células Progenitoras Linfoides/metabolismo , Camundongos , Camundongos Endogâmicos CBA , Camundongos Knockout , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/metabolismoRESUMO
Autoimmune thyroid diseases (AITD) are one of the most common organ-specific autoimmune disorders, of which Hashimoto's thyroiditis (HT) and Graves' disease (GD) are 2 of the most common clinical expressions. HT is characterized by hypothyroidism that results from the destruction of the thyroid by thyroglobulin-specific T cell-mediated autoimmune response. In contrast, GD is characterized by hyperthyroidism due to excessive production of thyroid hormone induced by thyrotropin receptor-specific stimulatory autoantibodies. Cytokines play a crucial role in modulating immune responses that affect the balance between maintenance of self-tolerance and initiation of autoimmunity. However, the role of cytokines is often confusing and is neither independent nor exclusive of other immune mediators. A regulatory cytokine may either favor induction of tolerance against thyroid autoimmune disease or favor activation and/or exacerbation of autoimmune responses. These apparently contradictory functions of a given cytokine are primarily influenced by the nature of co-signaling delivered by other cytokines. Consequently, a thorough understanding of the role of a particular cytokine in the context of a specific immune response is essential for the development of appropriate strategies to modulate cytokine responses to maintain or restore health. This review provides a summary of recent research pertaining to the role of cytokines in the pathogenesis of AITD with a particular emphasis on the therapeutic applications of cytokine modulation.