RESUMO
Programmed cell death 1 (PD-1) is a premier cancer drug target for immune checkpoint blockade (ICB). Because PD-1 receptor inhibition activates tumor-specific T-cell immunity, research has predominantly focused on T-cell-PD-1 expression and its immunobiology. In contrast, cancer cell-intrinsic PD-1 functional regulation is not well understood. Here, we demonstrate induction of PD-1 in melanoma cells via type I interferon receptor (IFNAR) signaling and reversal of ICB efficacy through IFNAR pathway inhibition. Treatment of melanoma cells with IFN-α or IFN-ß triggers IFNAR-mediated Janus kinase-signal transducer and activator of transcription (JAK/STAT) signaling, increases chromatin accessibility and resultant STAT1/2 and IFN regulatory factor 9 (IRF9) binding within a PD-1 gene enhancer, and leads to PD-1 induction. IFNAR1 or JAK/STAT inhibition suppresses melanoma-PD-1 expression and disrupts ICB efficacy in preclinical models. Our results uncover type I IFN-dependent regulation of cancer cell-PD-1 and provide mechanistic insight into the potential unintended ICB-neutralizing effects of widely used IFNAR1 and JAK inhibitors.
Assuntos
Inibidores de Checkpoint Imunológico , Interferon Tipo I , Melanoma , Receptor de Morte Celular Programada 1 , Receptor de Interferon alfa e beta , Transdução de Sinais , Receptor de Morte Celular Programada 1/metabolismo , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Melanoma/tratamento farmacológico , Melanoma/imunologia , Melanoma/genética , Melanoma/metabolismo , Humanos , Receptor de Interferon alfa e beta/metabolismo , Receptor de Interferon alfa e beta/genética , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Animais , Linhagem Celular Tumoral , Camundongos , Interferon Tipo I/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/metabolismo , Fator Gênico 3 Estimulado por Interferon, Subunidade gama/genética , Interferon beta/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Janus Quinases/metabolismo , Camundongos Endogâmicos C57BL , Interferon-alfa/farmacologia , Interferon-alfa/metabolismo , FemininoRESUMO
Merkel cell carcinoma (MCC) is a rare and aggressive skin cancer. Inhibitors targeting the programmed cell death 1 (PD-1) immune checkpoint have improved MCC patient outcomes by boosting antitumor T cell immunity. Here, we identify PD-1 as a growth-promoting receptor intrinsic to MCC cells. In human MCC lines and clinical tumors, RT-PCR-based sequencing, immunoblotting, flow cytometry, and immunofluorescence analyses demonstrated PD-1 gene and protein expression by MCC cells. MCC-PD-1 ligation enhanced, and its inhibition or silencing suppressed, in vitro proliferation and in vivo tumor xenograft growth. Consistently, MCC-PD-1 binding to PD-L1 or PD-L2 induced, while antibody-mediated PD-1 blockade inhibited, protumorigenic mTOR signaling, mitochondrial (mt) respiration, and ROS generation. Last, pharmacologic inhibition of mTOR or mtROS reversed MCC-PD-1:PD-L1-dependent proliferation and synergized with PD-1 checkpoint blockade in suppressing tumorigenesis. Our results identify an MCC-PD-1-mTOR-mtROS axis as a tumor growth-accelerating mechanism, the blockade of which might contribute to clinical response in patients with MCC.
Assuntos
Carcinoma de Célula de Merkel , Neoplasias Cutâneas , Humanos , Antígeno B7-H1 , Carcinoma de Célula de Merkel/tratamento farmacológico , Carcinoma de Célula de Merkel/genética , Receptor de Morte Celular Programada 1 , Espécies Reativas de Oxigênio , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/genética , Serina-Treonina Quinases TORRESUMO
T-cell immunoglobulin mucin family member 3 (Tim-3) is an immune checkpoint receptor that dampens effector functions and causes terminal exhaustion of cytotoxic T cells. Tim-3 inhibitors are under investigation in immuno-oncology (IO) trials, because blockade of T-cell-Tim-3 enhances antitumor immunity. Here, we identify an additional role for Tim-3 as a growth-suppressive receptor intrinsic to melanoma cells. Inhibition of melanoma cell-Tim-3 promoted tumor growth in both immunocompetent and immunocompromised mice, while melanoma-specific Tim-3 overexpression attenuated tumorigenesis. Ab-mediated Tim-3 blockade inhibited growth of immunogenic murine melanomas in T-cell-competent hosts, consistent with established antitumor effects of T-cell-Tim-3 inhibition. In contrast, Tim-3 Ab administration stimulated tumorigenesis of both highly and lesser immunogenic murine and human melanomas in T-cell-deficient mice, confirming growth-promoting effects of melanoma-Tim-3 antagonism. Melanoma-Tim-3 activation suppressed, while its blockade enhanced, phosphorylation of pro-proliferative downstream MAPK signaling mediators. Finally, pharmacologic MAPK inhibition reversed unwanted Tim-3 Ab-mediated tumorigenesis in T-cell-deficient mice and enhanced desired antitumor activity of Tim-3 interference in T-cell-competent hosts. These results identify melanoma-Tim-3 blockade as a mechanism that antagonizes T-cell-Tim-3-directed IO therapeutic efficacy. They further reveal MAPK targeting as a combination strategy for circumventing adverse consequences of unintended melanoma-Tim-3 inhibition. SIGNIFICANCE: Tim-3 is a growth-suppressive receptor intrinsic to melanoma cells, the blockade of which promotes MAPK-dependent tumorigenesis and thus counteracts antitumor activity of T-cell-directed Tim-3 inhibition.
Assuntos
Receptor Celular 2 do Vírus da Hepatite A , Melanoma , Animais , Carcinogênese , Transformação Celular Neoplásica , Humanos , Imunoglobulinas , Melanoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , MucinasRESUMO
Monoclonal antibodies (abs) targeting the programmed cell death 1 (PD-1) immune checkpoint pathway have revolutionized tumor therapy. Because T-cell-directed PD-1 blockade boosts tumor immunity, anti-PD-1 abs have been developed for examining T-cell-PD-1 functions. More recently, PD-1 expression has also been reported directly on cancer cells of various etiology, including in melanoma. Nevertheless, there is a paucity of studies validating anti-PD-1 ab clone utility in specific assay types for characterizing tumor cell-intrinsic PD-1. Here, we demonstrate reactivity of several anti-murine PD-1 ab clones and recombinant PD-L1 with live B16-F10 melanoma cells and YUMM lines using multiple independent methodologies, positive and negative PD-1-specific controls, including PD-1-overexpressing and PD-1 knockout cells. Flow cytometric analyses with two separate anti-PD-1 ab clones, 29F.1A12 and RMP1-30, revealed PD-1 surface protein expression on live murine melanoma cells, which was corroborated by marked enrichment in PD-1 gene (Pdcd1) expression. Immunoblotting, immunoprecipitation, and mass spectrometric sequencing confirmed PD-1 protein expression by B16-F10 cells. Recombinant PD-L1 also recognized melanoma cell-expressed PD-1, the blockade of which by 29F.1A12 fully abrogated PD-1:PD-L1 binding. Together, our data provides multiple lines of evidence establishing PD-1 expression by live murine melanoma cells and validates ab clones and assay systems for tumor cell-directed PD-1 pathway investigations.
Assuntos
Antineoplásicos Imunológicos , Melanoma Experimental , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/farmacologia , Antígeno B7-H1 , Células Clonais , Humanos , CamundongosRESUMO
Vitiligo is a T cell-mediated inflammatory skin disorder characterized by the loss of epidermal melanocytes. However, the contribution of melanocytes to the physiopathology of the disease in response to the T-cell microenvironment remains unclear. Here, using NanoString technology and multiplex ELISA, we show that active vitiligo perilesional skin is characterized by prominent type 1 and 2 associated immune responses. The vitiligo skin T-cell secretome downregulated melanocyte function and adhesion while increasing melanocyte mitochondrial metabolism and expression of inflammatory cytokines and chemokines by epidermal cells. The Jak1/2 inhibitor ruxolitinib strongly inhibited such effects on epidermal cells. Our data highlight that vitiligo is more complex than previously thought, with prominent combined activities of both T helper type 1- and T helper type 2-related cytokines inducing inflammatory responses of epidermal cells. Melanocytes do not appear only to be a target of T cells in vitiligo but could actively contribute to perpetuate inflammation. Jak inhibitors could prevent the impact of T cells on epidermal cells and pigmentation, highlighting their potential clinical benefit in vitiligo.
Assuntos
Vitiligo , Citocinas/metabolismo , Epiderme/metabolismo , Humanos , Melanócitos/metabolismo , Linfócitos T/metabolismo , Vitiligo/patologiaRESUMO
Breast cancer (BC) is a public health problem worldwide, causing suffering and premature death among women. As a heterogeneous disease, BC-specific diagnosis and treatment are challenging. Ectonucleotidases are related to tumor development and their expression may vary among BC. miRNAs may participate in epigenetic events and may regulate ectonucleotidases in BC. This study aimed to evaluate the expression of ectonucleotidases according to BC subtypes and to predict if there is post-transcriptional regulation of them by miRNAs. MCF 10A (non-tumorigenic), MCF7 (luminal BC), and MDA-MB-231 (triple-negative BC - TNBC) breast cell lines were used and ENTPD1 (the gene encoding for NTPDase1) and NT5E (the gene encoding for ecto-5'-nucleotidase) gene expression was determined. Interestingly, the expression of ENTPD1 was only observed in MCF7 and NT5E was lower in MCF7 compared to MDA-MB-231 cell line. ATP, ADP, and AMP hydrolysis were observed on the surface of all cell lines, being higher in MDA-MB-231. Like qPCR, the activity of AMP hydrolysis was also lower in the MCF7 cells, which may represent a striking feature of this BC subtype. In silico analyses confirmed that the miRNAs miR-101-3p, miR-141-3p, and miR-340-5p were higher expressed in MCF7 cells and targeted NT5E mRNA. Altogether, data suggest that the regulation of NT5E by miRNAs in MCF7 lineage may direct the molecular profile of luminal BC. Thus, we suggest that the roles of ecto-5'-nucleotidase and the aforementioned miRNAs must be unraveled in TNBC to be possibly defined as diagnostic and therapeutic targets.
Assuntos
Neoplasias da Mama , MicroRNAs , Neoplasias de Mama Triplo Negativas , 5'-Nucleotidase/genética , 5'-Nucleotidase/metabolismo , Antígenos CD , Apirase , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , MicroRNAs/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologiaRESUMO
Loxoscelism is the most dangerous araneism form in Brazil and antivenom therapy is the recommended treatment. Antivenom is produced by horse immunization with Loxosceles spider venom, which is toxic for the producer animal. Moreover, due to the high amount of venom required for horse hyperimmunization, new strategies for antigens obtention have been proposed. In this sense, our research group has previously produced a non-toxic recombinant multiepitopic protein derived from Loxosceles toxins (rMEPLox). rMEPLox was a successful immunogen, being able to induce the production of neutralizing antibodies, which could be used in the Loxoscelism treatment. However, rMEPLox obtention procedure requires optimization, as its production needs to be scaled up to suit antivenom manufacture. Therefore, an effective protocol development for rMEPlox production would be advantageous. To achieve this objective, we evaluated the influence of different cultivation conditions for rMEPLox optimum expression. The optimum conditions to obtain large amounts of rMEPlox were defined as the use of C43(DE3)pLysS as a host strain, 2xTY medium, 0.6 mM IPTG, biomass pre induction of OD600nm = 0.4 and incubation at 30 °C for 16 h. Following the optimized protocol, 39.84 mg/L of soluble rMEPLox was obtained and tested as immunogen. The results show that the obtained rMEPLox preserved the previously described immunogenicity, and it was able to generate antibodies that recognize different epitopes of the main Loxosceles venom toxins, which makes it a promising candidate for the antivenom production for loxoscelism treatment.
Assuntos
Escherichia coli , Expressão Gênica , Aranhas/genética , Animais , Antivenenos/biossíntese , Antivenenos/genética , Antivenenos/imunologia , Antivenenos/isolamento & purificação , Escherichia coli/genética , Escherichia coli/metabolismo , Camundongos Endogâmicos BALB C , Diester Fosfórico Hidrolases/biossíntese , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/imunologia , Diester Fosfórico Hidrolases/isolamento & purificação , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/isolamento & purificação , Venenos de Aranha/biossíntese , Venenos de Aranha/genética , Venenos de Aranha/imunologia , Venenos de Aranha/isolamento & purificaçãoRESUMO
In Type 1 Diabetes (T1D), CD4+ T cells initiate autoimmune attack of pancreatic islet ß cells. Importantly, bioenergetic programs dictate T cell function, with specific pathways required for progression through the T cell lifecycle. During activation, CD4+ T cells undergo metabolic reprogramming to the less efficient aerobic glycolysis, similarly to highly proliferative cancer cells. In an effort to limit tumor growth in cancer, use of glycolytic inhibitors have been successfully employed in preclinical and clinical studies. This strategy has also been utilized to suppress T cell responses in autoimmune diseases like Systemic Lupus Erythematosus (SLE), Multiple Sclerosis (MS), and Rheumatoid Arthritis (RA). However, modulating T cell metabolism in the context of T1D has remained an understudied therapeutic opportunity. In this study, we utilized the small molecule PFK15, a competitive inhibitor of the rate limiting glycolysis enzyme 6-phosphofructo-2-kinase/fructose-2,6- biphosphatase 3 (PFKFB3). Our results confirmed PFK15 inhibited glycolysis utilization by diabetogenic CD4+ T cells and reduced T cell responses to ß cell antigen in vitro. In an adoptive transfer model of T1D, PFK15 treatment delayed diabetes onset, with 57% of animals remaining euglycemic at the end of the study period. Protection was due to induction of a hyporesponsive T cell phenotype, characterized by increased and sustained expression of the checkpoint molecules PD-1 and LAG-3 and downstream functional and metabolic exhaustion. Glycolysis inhibition terminally exhausted diabetogenic CD4+ T cells, which was irreversible through restimulation or checkpoint blockade in vitro and in vivo. In sum, our results demonstrate a novel therapeutic strategy to control aberrant T cell responses by exploiting the metabolic reprogramming of these cells during T1D. Moreover, the data presented here highlight a key role for nutrient availability in fueling T cell function and has implications in our understanding of T cell biology in chronic infection, cancer, and autoimmunity.
Assuntos
Linfócitos T CD4-Positivos/efeitos dos fármacos , Diabetes Mellitus Tipo 1/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Glicólise/efeitos dos fármacos , Fosfofrutoquinase-2/antagonistas & inibidores , Piridinas/farmacologia , Quinolinas/farmacologia , Transferência Adotiva , Animais , Antígenos CD/metabolismo , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/transplante , Células Cultivadas , Reprogramação Celular/efeitos dos fármacos , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/metabolismo , Modelos Animais de Doenças , Feminino , Masculino , Camundongos Endogâmicos NOD , Camundongos SCID , Fosfofrutoquinase-2/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Fatores de Tempo , Proteína do Gene 3 de Ativação de LinfócitosRESUMO
In contrast to the skin and the gut, where somatic stem cells and their niche are well characterized, a definitive pancreatic multipotent cell population in the adult pancreas has yet to be revealed. Of particular interest is whether such cells may be endogenous in patients with diabetes, and if so, can they be used for therapeutic purposes? In the current study, we used two separate reporter lines to target Cre-recombinase expression to the Lgr5- or glucagon-expressing cells in the pancreas. We provide evidence for the existence of a population of cells within and in the proximity of the ducts that transiently express the stem-cell marker Lgr5 during late gestational stages. Careful timing of tamoxifen treatment in Lgr5EGFP-IRES-CreERT2 ;R26 Tomato mice allowed us to show that these Lgr5-expressing progenitor cells can differentiate into α-cells during pregnancy. Furthermore, we report on a spontaneous lineage conversion of α- to ß-cells specifically after parturition. The contribution of Lgr5 progeny to the ß-cell compartment through an α-cell intermediate phase early after pregnancy appears to be part of a novel mechanism that would counterbalance against excessive ß-cell mass reduction during ß-cell involution.
Assuntos
Linhagem da Célula , Células Secretoras de Glucagon/citologia , Células Secretoras de Insulina/citologia , Pâncreas/citologia , Período Pós-Parto/metabolismo , Receptores Acoplados a Proteínas G/fisiologia , Células-Tronco/citologia , Animais , Apoptose , Diferenciação Celular , Feminino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Loxoscelism pose a health issue in the South America. The treatment for these accidents is based on the administration of antivenom produced in animals immunized with Loxosceles venom. In this work, a previously produced non-toxic multiepitopic chimeric protein (rMEPlox), composed of epitopes derived from the main toxins families (sphyngomielinase-D, metalloproteases, and hyaluronidases) of Loxosceles spider venoms, was used as antigen to produce monoclonal antibodies (mAbs). A selected anti-rMEPlox mAb (Lox-mAb3) reacted with metalloprotease from L. intermedia venom and showed cross-reactivity with metalloproteses from Brazilian and Peruvian Loxosceles laeta and Loxosceles gaucho venoms in immunoassays. The sequence recognized by Lox-mAb3 (184ENNTRTIGPFDYDSIMLYGAY205) corresponds to the C-terminal region of Astacin-like metalloprotease 1 and the amino acid sequence IGPFDYDSI, conserved among the homologs metalloproteases sequences, is important for antibody recognition. Lox-mAb3 neutralizes the fibrinogenolytic activity caused by metalloprotease from L. intermedia spider venom in vitro, which may lead to a decrease in hemorrhagic disturbances caused by Loxosceles envenomation. Our results show, for the first time, the use of a non-toxic multiepitopic protein for the production of a neutralizing monoclonal antibody against a metalloprotease of medically important Loxosceles venoms. These results contribute for the production improvement of therapeutic antivenom against loxoscelism.
Assuntos
Anticorpos Monoclonais Murinos/imunologia , Anticorpos Neutralizantes/imunologia , Proteínas de Artrópodes , Epitopos , Metaloendopeptidases , Diester Fosfórico Hidrolases , Venenos de Aranha , Aranhas , Animais , Proteínas de Artrópodes/química , Proteínas de Artrópodes/genética , Proteínas de Artrópodes/imunologia , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Feminino , Metaloendopeptidases/química , Metaloendopeptidases/genética , Metaloendopeptidases/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Diester Fosfórico Hidrolases/química , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/imunologia , Engenharia de Proteínas , Venenos de Aranha/química , Venenos de Aranha/genética , Venenos de Aranha/imunologiaRESUMO
Loss of melanocytes is the pathological hallmark of vitiligo, a chronic inflammatory skin depigmenting disorder induced by exaggerated immune response, including autoreactive CD8 T cells producing high levels of type 1 cytokines. However, the interplay between this inflammatory response and melanocyte disappearance remains to be fully characterized. Here, we demonstrate that vitiligo skin contains a significant proportion of suprabasal melanocytes, associated with disruption of E-cadherin expression, a major protein involved in melanocyte adhesion. This phenomenon is also observed in lesional psoriatic skin. Importantly, apoptotic melanocytes were mainly observed once cells were detached from the basal layer of the epidermis, suggesting that additional mechanism(s) could be involved in melanocyte loss. The type 1 cytokines IFN-γ and TNF-α induce melanocyte detachment through E-cadherin disruption and the release of its soluble form, partly due to MMP-9. The levels of MMP-9 are increased in the skin and sera of patients with vitiligo, and MMP-9 is produced by keratinocytes in response to IFN-γ and TNF-α. Inhibition of MMP-9 or the JAK/STAT signaling pathway prevents melanocyte detachment in vitro and in vivo. Therefore, stabilization of melanocytes in the basal layer of the epidermis by preventing E-cadherin disruption appears promising for the prevention of depigmentation occurring in vitiligo and during chronic skin inflammation.
Assuntos
Caderinas/metabolismo , Interferon gama/metabolismo , Sistema de Sinalização das MAP Quinases , Metaloproteinase 9 da Matriz/biossíntese , Melanócitos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Vitiligo/metabolismo , Animais , Humanos , Queratinócitos/metabolismo , Queratinócitos/patologia , Melanócitos/patologia , CamundongosRESUMO
Vitiligo is an autoimmune disease that results from the loss of melanocytes, associated with skin infiltration of CD8+ effector memory T cells with a Tc1 skewed immune response. NKG2D is an activating receptor found on immune cells, in particular natural killer and activated CD8+ T cells, that are able to produce a high amount of IFN-γ. Here we found that NKG2D expression was increased in vitiligo skin CD8+ effector memory T cells and was promoted by IL-15. Phenotypic and functional analyses showed that NKG2D+ CD8+ skin effector memory T cells displayed an activated phenotype and produced elevated levels of both IFN-γ and tumor necrosis factor-α. Additional experiments revealed that vitiligo skin dendritic cells expressed the NKG2D ligands MICA-MICB, and in vitro experiments showed that these ligands could be induced on dendritic cells by IFN-α. Cultures of IFN-α-stimulated dendritic cells with skin NKG2D+ CD8+ T cells potentiated the production of type 1 cytokines, which was next inhibited by blocking the NKG2D/MICA-MICB interaction. These data show that NKG2D is a potential marker of pathogenic skin CD8+ effector memory T cells during vitiligo. Therefore, targeting NKG2D could be an attractive strategy in vitiligo, a disease for which there is a strong need of innovative treatments.
Assuntos
Subpopulações de Linfócitos/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Linfócitos T Citotóxicos/imunologia , Vitiligo/imunologia , Adulto , Idoso , Biomarcadores/metabolismo , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Feminino , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Interleucina-15/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Subpopulações de Linfócitos/metabolismo , Masculino , Pessoa de Meia-Idade , Subfamília K de Receptores Semelhantes a Lectina de Células NK/antagonistas & inibidores , Cultura Primária de Células , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Pele/citologia , Pele/imunologia , Pele/patologia , Linfócitos T Citotóxicos/metabolismo , Vitiligo/tratamento farmacológico , Vitiligo/patologiaRESUMO
Lymphocyte activation gene-3 (LAG-3) is an inhibitory receptor expressed by CD4+ T cells and tempers their homeostatic expansion. Because CD4+ T cell proliferation is tightly coupled to bioenergetics, we investigate the role of LAG-3 in modulating naive CD4+ T cell metabolism. LAG-3 deficiency enhances the metabolic profile of naive CD4+ T cells by elevating levels of mitochondrial biogenesis. In vivo, LAG-3 blockade partially restores expansion and the metabolic phenotype of wild-type CD4+ T cells to levels of Lag3-/- CD4+ T cells, solidifying that LAG-3 controls these processes. Lag3-/- CD4+ T cells also demonstrate greater signal transducer and activator of transcription 5 (STAT5) activation, enabling resistance to interleukin-7 (IL-7) deprivation. These results implicate this pathway as a target of LAG-3-mediated inhibition. Additionally, enhancement of STAT5 activation, as a result of LAG-3 deficiency, contributes to greater activation potential in these cells. These results identify an additional mode of regulation elicited by LAG-3 in controlling CD4+ T cell responses.
Assuntos
Antígenos CD/fisiologia , Linfócitos T CD4-Positivos , Metabolismo Energético/genética , Mitocôndrias/fisiologia , Biogênese de Organelas , Animais , Antígenos CD/genética , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/ultraestrutura , Células Cultivadas , Feminino , Ativação Linfocitária/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/genética , Mitocôndrias/metabolismo , Fase de Repouso do Ciclo Celular/genética , Proteína do Gene 3 de Ativação de LinfócitosRESUMO
Psoriasis is one of the most common skin inflammatory diseases worldwide. The vitamin D3 analog calcipotriol has been used alone or in combination with corticosteroids in treating plaque psoriasis, but how it suppresses psoriatic inflammation has not been fully understood. Using an experimental mouse psoriasis model, we show that topical calcipotriol inhibited the pivotal IL-23/IL-17 axis and neutrophil infiltration in psoriatic skin, and interestingly, such effects were mediated through the vitamin D receptor (VDR) in keratinocytes (KCs). We further reveal that IL-36α and IL-36γ, which have recently emerged as key players in psoriasis pathogenesis, were effectively repressed by calcipotriol via direct VDR signaling in mouse KCs. Accordingly, calcipotriol treatment suppressed IL-36α/γ expression in lesional skin from patients with plaque psoriasis, which was accompanied by a reduced IL-23/IL-17 expression. In contrast, dexamethasone indirectly reduced IL-36α/γ expression in mouse psoriatic skin through immune cells. Furthermore, we demonstrate that calcipotriol and dexamethasone, in combination, synergistically suppressed the expression of IL-36α/γ, IL-23, and IL-17 in the established mouse psoriasis. Our findings indicate that the combination of calcipotriol and corticosteroid efficiently disrupts the IL-36 and IL-23/IL-17 positive feedback loop, thus revealing a mechanism underlying the superior efficacy of calcipotriol and corticosteroid combination therapy for psoriasis.
RESUMO
Vitiligo is a chronic autoimmune depigmenting skin disorder that results from a loss of melanocytes. Multiple combinatorial factors have been involved in disease development, with a prominent role of the immune system, in particular T cells. After repigmentation, vitiligo frequently recurs in the same area, suggesting that vitiligo could involve the presence of resident memory T cells (TRM). We sought to perform a thorough characterization of the phenotype and function of skin memory T cells in vitiligo. We show that stable and active vitiligo perilesional skin is enriched with a population of CD8 TRM expressing both CD69 and CD103 compared with psoriasis and control unaffected skin. CD8 TRM expressing CD103 are mainly localized in the epidermis. Expression of CXCR3 is observed on most CD8 TRM in vitiligo, including the population of melanocyte-specific CD8 T cells. CD8 TRM displayed increased production of IFN-γ and tumor necrosis factor-α with moderate cytotoxic activity. Our study highlights the presence of functional CD8 TRM in both stable and active vitiligo, reinforcing the concept of vitiligo as an immune memory skin disease. The CD8 TRM that remain in stable disease could play a role during disease flares, emphasizing the interest in targeting this cell subset in vitiligo.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/imunologia , Receptores CXCR3/metabolismo , Pele/imunologia , Vitiligo/imunologia , Adulto , Biópsia , Linfócitos T CD8-Positivos/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Receptores CXCR3/imunologia , Pele/citologia , Pele/patologia , Vitiligo/sangue , Vitiligo/patologiaRESUMO
The immune system is necessary for protecting against various pathogens. However, under certain circumstances, self-reactive immune cells can drive autoimmunity, like that exhibited in type 1 diabetes (T1D). CD4+ T cells are major contributors to the immunopathology in T1D, and in order to drive optimal T cell activation, third signal reactive oxygen species (ROS) must be present. However, the role ROS play in mediating this process remains to be further understood. Recently, cellular metabolic programs have been shown to dictate the function and fate of immune cells, including CD4+ T cells. During activation, CD4+ T cells must transition metabolically from oxidative phosphorylation to aerobic glycolysis to support proliferation and effector function. As ROS are capable of modulating cellular metabolism in other models, we sought to understand if blocking ROS also regulates CD4+ T cell activation and effector function by modulating T cell metabolism. To do so, we utilized an ROS scavenging and potent antioxidant manganese metalloporphyrin (MnP). Our results demonstrate that redox modulation during activation regulates the mTOR/AMPK axis by maintaining AMPK activation, resulting in diminished mTOR activation and reduced transition to aerobic glycolysis in diabetogenic splenocytes. These results correlated with decreased Myc and Glut1 upregulation, reduced glucose uptake, and diminished lactate production. In an adoptive transfer model of T1D, animals treated with MnP demonstrated delayed diabetes progression, concurrent with reduced CD4+ T cell activation. Our results demonstrate that ROS are required for driving and sustaining T cell activation-induced metabolic reprogramming, and further support ROS as a target to minimize aberrant immune responses in autoimmunity.