RESUMO
In 2007, it was shown that DNA sequence variants of the human NLRP1 gene are associated with autoimmune and autoinflammatory diseases affecting mainly the skin. However, at that time, the underlying cellular and molecular mechanisms were poorly characterized. Meanwhile, increasing evidence suggests that the NLRP1 inflammasome expressed by keratinocytes not only plays a part in the pathology of common inflammatory skin diseases and cancer development but also contributes to skin immunity. Understanding the mechanisms regulating NLRP1 activation in keratinocytes and the downstream events in human skin might pave the way for developing novel strategies for treating patients suffering from NLRP1-mediated skin diseases.
Assuntos
Dermatite , Dermatopatias , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Humanos , Inflamassomos/metabolismo , Queratinócitos/metabolismo , Proteínas NLRRESUMO
p62 is a highly conserved, multi-domain, and multi-functional adaptor protein critically involved in several important cellular processes. Via its pronounced domain architecture, p62 binds to numerous interaction partners, thereby influencing key pathways that regulate tissue homeostasis, inflammation, and several common diseases including cancer. Via binding of ubiquitin chains, p62 acts in an anti-inflammatory manner as an adaptor for the auto-, xeno-, and mitophagy-dependent degradation of proteins, pathogens, and mitochondria. Furthermore, p62 is a negative regulator of inflammasome complexes. The transcription factor Nrf2 regulates expression of a bundle of ROS detoxifying genes. p62 activates Nrf2 by interaction with and autophagosomal degradation of the Nrf2 inhibitor Keap1. Moreover, p62 activates mTOR, the central kinase of the mTORC1 sensor complex that controls cell proliferation and differentiation. Through different mechanisms, p62 acts as a positive regulator of the transcription factor NF-κB, a central player in inflammation and cancer development. Therefore, p62 represents not only a cargo receptor for autophagy, but also a central signaling hub, linking several important pro- and anti-inflammatory pathways. This review aims to summarize knowledge about the molecular mechanisms underlying the roles of p62 in health and disease. In particular, different types of tumors are characterized by deregulated levels of p62. The elucidation of how p62 contributes to inflammation and cancer progression at the molecular level might promote the development of novel therapeutic strategies.
RESUMO
Inflammasomes represent a group of protein complexes that contribute to host defense against pathogens and repair processes upon the induction of inflammation. However, aberrant and chronic inflammasome activation underlies the pathology of numerous common inflammatory diseases. Inflammasome assembly causes activation of the protease caspase-1 which in turn activates proinflammatory cytokines and induces a lytic type of cell death termed pyroptosis. Although NLRP1 (NACHT, leucine-rich repeat and pyrin domain containing 1) was the first inflammasome sensor, described almost 20 years ago, the molecular mechanisms underlying its activation and the resulting downstream events are incompletely understood. This is partially a consequence of the poor conservation of the NLRP1 pathway between human and mice. Moreover, recent evidence demonstrates a complex and multi-stage mechanism of NLRP1 inflammasome activation. In contrast to other inflammasome sensors, NLRP1 possesses protease activity required for proteolytic self-cleavage and activation mediated by the function-to-find domain (FIIND). CARD8 is a second FIIND protein and is expressed in humans but not in mice. In immune cells and AML (acute myeloid leukemia) cells, the anti-cancer drug talabostat induces CARD8 activation and causes caspase-1-dependent pyroptosis. In contrast, in human keratinocytes talabostat induces NLRP1 activation and massive proinflammatory cytokine activation. NLRP1 is regarded as the principal inflammasome sensor in human keratinocytes and UVB radiation induces its activation, which is believed to underlie the induction of sunburn. Moreover, gain-of-function mutations of NLRP1 cause inflammatory skin syndromes and a predisposition for the development of skin cancer. SNPs (single nucleotide polymorphisms) of NLRP1 are associated with several (auto)inflammatory diseases with a major skin phenotype, such as psoriasis or vitiligo. Here, we summarize knowledge about NLRP1 with emphasis on its role in human keratinocytes and skin. Due to its accessibility, pharmacological targeting of NLRP1 activation in epidermal keratinocytes represents a promising strategy for the treatment of the numerous patients suffering from NLRP1-dependent inflammatory skin conditions and cancer.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Inflamassomos/metabolismo , Inflamação/patologia , Queratinócitos/patologia , Neoplasias Cutâneas/patologia , Pele/patologia , Animais , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Queratinócitos/imunologia , Queratinócitos/metabolismo , Proteínas NLR , Proteínas de Neoplasias/metabolismo , Pele/imunologia , Pele/metabolismo , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/metabolismoRESUMO
The skin represents an indispensable barrier between the organism and the environment and is the first line of defense against exogenous insults. The transcription factor NRF2 is a central regulator of cytoprotection and stress resistance. NRF2 is activated in response to oxidative stress by reactive oxygen species (ROS) and electrophiles. These electrophiles oxidize specific cysteine residues of the NRF2 inhibitor KEAP1, leading to KEAP1 inactivation and, subsequently, NRF2 activation. As oxidative stress is associated with inflammation, the NRF2 pathway plays important roles in the pathogenesis of common inflammatory diseases and cancer in many tissues and organs, including the skin. The electrophile and NRF2 activator dimethyl fumarate (DMF) is an established and efficient drug for patients suffering from the common inflammatory skin disease psoriasis and the neuro-inflammatory disease multiple sclerosis (MS). In this review, we discuss possible molecular mechanisms underlying the therapeutic activity of DMF and other NRF2 activators. Recent evidence suggests that electrophiles not only activate NRF2, but also target other inflammation-associated pathways including the transcription factor NF-κB and the multi-protein complexes termed inflammasomes. Inflammasomes are central regulators of inflammation and are involved in many inflammatory conditions. Most importantly, the NRF2 and inflammasome pathways are connected at different levels, mainly antagonistically.
Assuntos
Fármacos Dermatológicos/farmacologia , Fumarato de Dimetilo/farmacologia , Fator 2 Relacionado a NF-E2/agonistas , Fator 2 Relacionado a NF-E2/metabolismo , Dermatopatias/tratamento farmacológico , Dermatopatias/metabolismo , Animais , Fármacos Dermatológicos/uso terapêutico , Fumarato de Dimetilo/uso terapêutico , Elétrons , Humanos , Inflamassomos/efeitos dos fármacos , Inflamassomos/metabolismo , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , NF-kappa B/efeitos dos fármacos , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
Inflammasome activation induces caspase-1-dependent secretion of the proinflammatory cytokine IL-1ß. In addition, caspase-1 activates the protein GSDMD in immune cells, causing pyroptosis, a lytic type of cell death. In contrast, UVB irradiation of human primary keratinocytes induces NLRP1 inflammasome activation, cytokine secretion, and caspase-1-dependent apoptosis, rather than pyroptosis. Here, we addressed the molecular mechanisms underlying the role of caspase-1 in UVB-induced cell death of human primary keratinocytes. We show that GSDMD is a poor substrate of caspase-1 in human primary keratinocytes and that its activation upon UVB irradiation supports secretion of IL-1ß. We screened for novel substrates of caspase-1 by a mass spectrometry-based approach and identified the specific cleavage of the major vault protein (MVP) at D441 by caspase-1 and -9. MVP is the main component of vaults, highly conserved ribonucleoprotein particles, whose functions are poorly understood. Cleavage of MVP is a common event occurring in human primary keratinocytes and fibroblasts undergoing apoptosis induced by different stimuli. In contrast, MVP cleavage could not be detected in pyroptotic cells. Cleavage of MVP by caspase-1 and -9 inactivates this cytoprotective protein. These results demonstrate a proapoptotic activity of caspase-1 and a crosstalk with caspase-9 upon inactivation of the cytoprotective MVP in apoptotic epithelial cells.
Assuntos
Apoptose , Caspase 1/metabolismo , Caspase 9/metabolismo , Células Epiteliais/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Ligação a Fosfato/metabolismo , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Biópsia , Fibroblastos/metabolismo , Humanos , Inflamassomos , Interleucina-1beta/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Espectrometria de Massas , Proteínas NLR , RNA Interferente Pequeno/metabolismo , Raios UltravioletaRESUMO
Epidermal growth factor receptor (EGFR) and MEK inhibitors (EGFRi/MEKi) are beneficial for the treatment of solid cancers but are frequently associated with severe therapy-limiting acneiform skin toxicities. The underlying molecular mechanisms are poorly understood. Using gene expression profiling we identified IL-36γ and IL-8 as candidate drivers of EGFRi/MEKi skin toxicity. We provide molecular and translational evidence that EGFRi/MEKi in concert with the skin commensal bacterium Cutibacterium acnes act synergistically to induce IL-36γ in keratinocytes and subsequently IL-8, leading to cutaneous neutrophilia. IL-36γ expression was the combined result of C. acnes-induced NF-κB activation and EGFRi/MEKi-mediated expression of the transcription factor Krüppel-like factor 4 (KLF4), due to the presence of both NF-κB and KLF4 binding sites in the human IL-36γ gene promoter. EGFRi/MEKi increased KLF4 expression by blockade of the EGFR/MEK/ERK pathway. These results provide an insight into understanding the pathological mechanism of the acneiform skin toxicities induced by EGFRi/MEKi and identify IL-36γ and the transcription factor KLF4 as potential therapeutic targets.
Assuntos
Receptores ErbB/imunologia , Interleucina-1/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Propionibacteriaceae/imunologia , Dermatopatias Bacterianas/imunologia , Animais , Receptores ErbB/genética , Humanos , Interleucina-1/genética , Queratinócitos/imunologia , Queratinócitos/microbiologia , Queratinócitos/patologia , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/imunologia , Sistema de Sinalização das MAP Quinases/genética , Camundongos , Camundongos Knockout , NF-kappa B/genética , NF-kappa B/imunologia , Dermatopatias Bacterianas/genética , Dermatopatias Bacterianas/patologiaRESUMO
The inflammasome protein NLRP1 is an important innate immune sensor in human keratinocytes, and, together with ASC and caspase-1, it mediates the activation and secretion of the proinflammatory cytokines IL-1ß and IL-18. These cytokines and inflammasomes can have partly opposing roles during tumorigenesis in mice. In contrast, ASC expression is impaired in different types of cancer in humans. In this study, we analyzed inflammasome activation and expression of inflammasome proteins, including their downstream cytokines, in squamous cell carcinomas, a type of nonmelanoma skin cancer derived from keratinocytes. We assessed mRNA and protein levels in human primary keratinocytes and skin carcinoma-derived SCC cell lines and detected a strong down-regulation of expression of NLRP1 inflammasome components, as well as reduced expression of the proinflammatory cytokines proIL-1ß and proIL-1α. Protein levels of NLRP1, ASC, caspase-1, and proIL-1ß were reduced in patient-derived SCC biopsy samples compared with healthy skin. Furthermore, the results suggest that expression of PYCARD (ASC), CASP1, IL1B, and NLRP1 is silenced by methylation in SCC cell lines. In conclusion, the down-regulation of the inflammasome pathway in SCCs might favor late tumor development in human skin.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Reguladoras de Apoptose/genética , Carcinoma de Células Escamosas/genética , Regulação Neoplásica da Expressão Gênica/imunologia , Inflamassomos/genética , Neoplasias Cutâneas/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/imunologia , Proteínas Reguladoras de Apoptose/metabolismo , Biópsia , Carcinogênese/genética , Carcinogênese/imunologia , Carcinoma de Células Escamosas/imunologia , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Citocinas , Regulação para Baixo , Humanos , Inflamassomos/imunologia , Inflamassomos/metabolismo , Queratinócitos/imunologia , Queratinócitos/patologia , Proteínas NLR , Cultura Primária de Células , RNA Mensageiro/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Pele/citologia , Pele/imunologia , Pele/patologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologiaRESUMO
By forming a protective barrier, epidermal keratinocytes represent the first line of defense against environmental insults. UVB radiation of the sun is a major challenge for the skin and can induce inflammation, aging, and eventually skin cancer. UVB induces an immune response in human keratinocytes resulting in activation and secretion of the proinflammatory cytokines proIL-1ß and -18. This is mediated by an assembly of protein complexes, termed inflammasomes. However, the mechanisms underlying sensing of UVB by keratinocytes, and particularly the types of inflammasomes required for cytokine secretion, are a matter of debate. To address these questions, we established a protocol that allows the generation of CRISPR/Cas9-targeted human primary keratinocytes. Our experiments showed an essential role of the NLRP1 rather than the NLRP3 inflammasome in UVB sensing and subsequent IL-1ß and -18 secretion by keratinocytes. Moreover, NLRP1 but not NLRP3 was required for inflammasome activation in response to nigericin, a potassium ionophore and well-established NLRP3 activator in immune cells. Because the CRISPR/Cas9-targeted cells retained their full differentiation capacity, genome editing of human primary keratinocytes might be useful for numerous research and medical applications.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Sistemas CRISPR-Cas , Edição de Genes/métodos , Inflamassomos/metabolismo , Queratinócitos/fisiologia , Radiodermite/genética , Raios Ultravioleta/efeitos adversos , Proteína 9 Associada à CRISPR/genética , Células Cultivadas , Humanos , Interleucina-18/metabolismo , Interleucina-1beta/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas NLR , Nigericina/farmacologia , Cultura Primária de Células , Radiodermite/metabolismoRESUMO
Allergen immunotherapy (AIT) is the only modality that can modify immune responses to allergen exposure, but therapeutic coverage is low. One strategy to improve AIT safety and efficacy is the use of new or improved adjuvants. This study investigates immune responses produced by microcrystalline tyrosine (MCT)-based vaccines as compared with conventional aluminum hydroxide (alum). Wild-type, immune-signaling-deficient, and TCR-transgenic mice were treated with different Ags (e.g., OVA and cat dander Fel d 1), plus MCT or alum as depot adjuvants. Specific Ab responses in serum were measured by ELISA, whereas cytokine secretion was measured both in culture supernatants by ELISA or by flow cytometry of spleen cells. Upon initiation of AIT in allergic mice, body temperature and further clinical signs were used as indicators for anaphylaxis. Overall, MCT and alum induced comparable B and T cell responses, which were independent of TLR signaling. Alum induced stronger IgE and IL-4 secretion than MCT. MCT and alum induced caspase-dependent IL-1ß secretion in human monocytes in vitro, but inflammasome activation had no functional effect on inflammatory and Ab responses measured in vivo. In sensitized mice, AIT with MCT-adjuvanted allergens caused fewer anaphylactic reactions compared with alum-adjuvanted allergens. As depot adjuvants, MCT and alum are comparably effective in strength and mechanism of Ag-specific IgG induction and induction of T cell responses. The biocompatible and biodegradable MCT seems therefore a suitable alternative adjuvant to alum-based vaccines and AIT.
Assuntos
Adjuvantes Imunológicos/farmacologia , Hidróxido de Alumínio/farmacologia , Dessensibilização Imunológica/métodos , Tirosina/farmacologia , Animais , Modelos Animais de Doenças , Hipersensibilidade/prevenção & controle , Imunoglobulina E/imunologia , Inflamassomos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Transdução de Sinais/imunologia , Receptores Toll-Like/imunologiaRESUMO
Hypoxia is a hallmark of cancer that is strongly associated with invasion, metastasis, resistance to therapy and poor clinical outcome. Tumour hypoxia affects immune responses and promotes the accumulation of macrophages in the tumour microenvironment. However, the signals linking tumour hypoxia to tumour-associated macrophage recruitment and tumour promotion are incompletely understood. Here we show that the damage-associated molecular pattern High-Mobility Group Box 1 protein (HMGB1) is released by melanoma tumour cells as a consequence of hypoxia and promotes M2-like tumour-associated macrophage accumulation and an IL-10 rich milieu within the tumour. Furthermore, we demonstrate that HMGB1 drives IL-10 production in M2-like macrophages by selectively signalling through the Receptor for Advanced Glycation End products (RAGE). Finally, we show that HMGB1 has an important role in murine B16 melanoma growth and metastasis, whereas in humans its serum concentration is significantly increased in metastatic melanoma. Collectively, our findings identify a mechanism by which hypoxia affects tumour growth and metastasis in melanoma and depict HMGB1 as a potential therapeutic target.
Assuntos
Macrófagos/metabolismo , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Hipóxia Tumoral , Animais , Proliferação de Células , Feminino , Proteína HMGB1/sangue , Proteína HMGB1/metabolismo , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Interleucina-10/metabolismo , Macrófagos/patologia , Masculino , Melanoma Experimental/sangue , Camundongos Endogâmicos C57BL , Fenótipo , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Neoplasias Cutâneas/sangue , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologiaRESUMO
Acne vulgaris is potentially a severe skin disease associated with colonization of the pilo-sebaceous unit by the commensal bacterium Propionibacterium acnes and inflammation. P. acnes is considered to contribute to inflammation in acne, but the pathways involved are unclear. Here we reveal a mechanism that regulates inflammatory responses to P. acnes. We show that IL-1ß mRNA and the active processed form of IL-1ß are abundant in inflammatory acne lesions. Moreover, we identify P. acnes as a trigger of monocyte-macrophage NLRP3-inflammasome activation, IL-1ß processing and secretion that is dependent on phagocytosis, lysosomal destabilization, reactive oxygen species, and cellular K+ efflux. In mice, inflammation induced by P. acnes is critically dependent on IL-1ß and the NLRP3 inflammasome of myeloid cells. These findings show that the commensal P. acnes-by activating the inflammasome-can trigger an innate immune response in the skin, thus establishing the NLRP3-inflammasome and IL-1ß as possible therapeutic targets in acne.