RESUMO
miRNAs are small noncoding RNAs that regulate mRNA targets in a cell-specific manner. miR-29 is expressed in murine and human skin, where it may regulate functions in skin repair. Cutaneous wound healing model in miR-29a/b1 gene knockout mice was used to identify miR-29 targets in the wound matrix, where angiogenesis and maturation of provisional granulation tissue was enhanced in response to genetic deletion of miR-29. Consistently, antisense-mediated inhibition of miR-29 promoted angiogenesis in vitro by autocrine and paracrine mechanisms. These processes are likely mediated by miR-29 target mRNAs released upon removal of miR-29 to improve cell-matrix adhesion. One of these, laminin (Lam)-c2 (also known as laminin γ2), was strongly up-regulated during skin repair in the wound matrix of knockout mice. Unexpectedly, Lamc2 was deposited in the basal membrane of endothelial cells in blood vessels forming in the granulation tissue of knockout mice. New blood vessels showed punctate interactions between Lamc2 and integrin α6 (Itga6) along the length of the proto-vessels, suggesting that greater levels of Lamc2 may contribute to the adhesion of endothelial cells, thus assisting angiogenesis within the wound. These findings may be of translational relevance, as LAMC2 was deposited at the leading edge in human wounds, where it formed a basal membrane for endothelial cells and assisted neovascularization. These results suggest a link between LAMC2, improved angiogenesis, and re-epithelialization.
Assuntos
Laminina , MicroRNAs , Humanos , Animais , Camundongos , Laminina/genética , Células Endoteliais , Transdução de Sinais/fisiologia , MicroRNAs/genética , Pele , Camundongos KnockoutRESUMO
The transcription factor NRF2 is well known as a master regulator of the cellular stress response. As such, activation of NRF2 has gained widespread attention for its potential to prevent tissue injury, but also as a possible therapeutic approach to promote repair processes. While NRF2 activation affects most or even all cell types, its effect on epithelial cells during repair processes has been particularly well studied. In response to tissue injury, these cells proliferate, migrate and/or spread to effectively repair the damage. In this review, we discuss how NRF2 governs repair of epithelial tissues, and we highlight the increasing number of NRF2 targets with diverse roles in regulating epithelial repair.
Assuntos
Fator 2 Relacionado a NF-E2 , Transdução de Sinais , Fator 2 Relacionado a NF-E2/metabolismo , Transdução de Sinais/fisiologia , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Regulação da Expressão Gênica , Células Epiteliais/metabolismo , Estresse OxidativoRESUMO
Collagens are fibrous proteins that are integral to the strength and stability of connective tissues. During collagen maturation, lysyl oxidases (LOX) initiate the cross-linking of fibers, but abnormal LOX activity is associated with impaired tissue function as seen in fibrotic and malignant diseases. Visualizing and targeting this dynamic process in healthy and diseased tissue is important, but so far not feasible. Here we present a probe for the simultaneous monitoring and targeting of LOX-mediated collagen cross-linking that combines a LOX-activity sensor with a collagen peptide to chemoselectively target endogenous aldehydes generated by LOX. This synergistic probe becomes covalently anchored and lights up in vivo and in situ in response to LOX at the sites where cross-linking occurs, as demonstrated by staining of normal skin and cancer sections. We anticipate that our reactive collagen-based sensor will improve understanding of collagen remodeling and provide opportunities for the diagnosis of fibrotic and malignant diseases.
Assuntos
Colágeno/metabolismo , Reagentes de Ligações Cruzadas/metabolismo , Peptídeos/metabolismo , Proteína-Lisina 6-Oxidase/metabolismo , Aldeídos/química , Aldeídos/metabolismo , Animais , Colágeno/química , Reagentes de Ligações Cruzadas/química , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Peptídeos/química , Proteína-Lisina 6-Oxidase/químicaRESUMO
Wound healing involves the concerted action of various lymphoid and in particular myeloid cell populations. To characterize and quantitate different types of myeloid cells and to obtain information on their kinetics during wound healing, we performed multiparametric flow cytometry analysis. In healthy mice, neutrophil numbers increased early after injury and returned to near basal levels after completion of healing. Macrophages, monocyte-derived dendritic cells (DCs), and eosinophils were abundant throughout the healing phase, in particular in early wounds, and Langerhans cells increased after wounding and remained elevated after epithelial closure. Major differences in healing-impaired diabetic mice were a much higher percentage of immune cells in late wounds, mainly as a result of neutrophil, macrophage, and monocyte persistence; reduced numbers and percentages of macrophages and monocyte-derived DCs in early wounds; and of Langerhans cells, conventional DCs, and eosinophils throughout the healing process. Finally, unbiased cluster analysis (PhenoGraph) identified a large number of different clusters of myeloid cells in skin wounds. These results provide insight into myeloid cell diversity and dynamics during wound repair and highlight the abnormal inflammatory response associated with impaired healing.
Assuntos
Células Mieloides/fisiologia , Cicatrização/fisiologia , Animais , Complicações do Diabetes/imunologia , Complicações do Diabetes/metabolismo , Diabetes Mellitus Experimental , Camundongos Endogâmicos C57BLRESUMO
Healing of skin wounds is orchestrated by various types of immune cells, but little is known about the role of FoxP3+ regulatory T cells (Tregs) in this process. Here, we determined if Tregs are important for wound healing in normal mice and if they contribute to the accelerated healing of mice overexpressing the growth and differentiation factor activin. Diphtheria toxin induced Treg depletion prior to injury caused impaired healing characterized by delayed reepithelialization, reduced wound contraction, and impaired vessel maturation. The accelerated wound repair of activin-transgenic mice was also abrogated. Mechanistically, we found a strong increase in IL-4 levels combined with overrepresentation of T-bet+ and GATA-3+ αß T cells in Treg-depleted 7-day wounds. In addition, numbers of IFN-γ- or IL-17A-producing CD4+ and CD4- T cells were elevated. These results demonstrate that Treg depletion in wounds facilitates the expansion of an αß T-cell population with features of Th1 and Th2 cells, and suggest that concomitant changes in the cytokine milieu disturb the healing process.
Assuntos
Fatores de Transcrição Forkhead/metabolismo , Linfócitos T Reguladores/fisiologia , Células Th1/imunologia , Células Th2/imunologia , Cicatrização/imunologia , Ativinas/imunologia , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Citocinas/metabolismo , Toxina Diftérica/imunologia , Fatores de Transcrição Forkhead/genética , Fator de Transcrição GATA3/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Regiões Promotoras Genéticas/genética , Proteínas com Domínio T/genética , Cicatrização/genéticaRESUMO
The nuclear factor-erythroid 2-related factor 2 (NRF2) transcription factor plays a central role in mediating the cellular stress response. Due to their antioxidant properties, compounds activating NRF2 have received much attention as potential medications for disease prevention, or even for therapy. Accumulating evidence suggests that activation of the NRF2 pathway also has a major impact on wound healing and may be beneficial in the treatment of chronic wounds, which remain a considerable health and economic burden. While NRF2 activation indeed shows promise, important considerations need to be made in light of corresponding evidence that also points towards pro-tumorigenic effects of NRF2. In this review, we discuss the evidence to date, highlighting recent advances using gain- and loss-of-function animal models and how these data fit with observations in humans.
Assuntos
Regulação da Expressão Gênica , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Cicatrização , Animais , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Citoproteção , Fibroblastos , Humanos , Queratinócitos/metabolismo , Mutação com Perda de Função , Estresse Oxidativo , FenótipoRESUMO
Exposure to particulate matter (PM), a major component of air pollution, contributes to increased morbidity and mortality worldwide. Inhaled PM induces innate immune responses by airway epithelial cells that may lead to the exacerbation or de novo development of airway disease. We have previously shown that 10-µm PM (PM10) activates the nucleotide-binding domain, leucine-rich repeat protein (NLRP) 3 inflammasome in human airway epithelial cells. Our objective was to determine the innate and adaptive immune responses mediated by the airway epithelium NLRP3 inflammasome in response to PM10 exposure. Using in vitro cultures of human airway epithelial cells and in vivo studies with wild-type and Nlrp3(-/-) mice, we investigated the downstream consequences of PM10-induced NLPR3 inflammasome activation on cytokine production, cellular inflammation, dendritic cell activation, and PM10-facilitated allergic sensitization. PM10 activates an NLRP3 inflammasome/IL-1 receptor I (IL-1RI) axis in airway epithelial cells, resulting in IL-1ß, CC chemokine ligand-20, and granulocyte/macrophage colony-stimulating factor production, which is associated with dendritic cell activation and lung neutrophilia. Despite these profound innate immune responses in the airway epithelium, the NLRP3 inflammasome/IL-1RI axis is dispensable for PM10-facilitated allergic sensitization. We demonstrate the importance of the lung NLRP3 inflammasome in mediating PM10 exposure-associated innate, but not adaptive, immune responses. Our study highlights a mechanism by which PM10 exposure can contribute to the exacerbation of airway disease, but not PM10-facilitated allergic sensitization.
Assuntos
Imunidade Adaptativa/efeitos dos fármacos , Proteínas de Transporte/imunologia , Imunidade Inata/efeitos dos fármacos , Material Particulado/efeitos adversos , Receptores Tipo I de Interleucina-1/imunologia , Mucosa Respiratória/imunologia , Transdução de Sinais/imunologia , Animais , Asma/induzido quimicamente , Asma/imunologia , Asma/patologia , Linhagem Celular Transformada , Citocinas/imunologia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Humanos , Inflamassomos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Proteína 3 que Contém Domínio de Pirina da Família NLR , Material Particulado/farmacologia , Mucosa Respiratória/patologiaRESUMO
Tissue repair and fibrosis involve the dynamic remodeling of collagen, and accurate detection of these sites is of utmost importance. Here, we use a collagen peptide sensor (1) to visualize collagen formation and remodeling during wound healing in mice and humans. We show that the probe binds selectively to sites of collagen formation and remodeling at different stages of healing. Compared to conventional methods, the peptide sensor localizes preferentially to areas of collagen synthesis and remodeling at the wound edge and not in matured fibrillar collagen. We also demonstrate its applicability for in vivo wound imaging and for discerning differential remodeling in wounds of transgenic mice with altered collagen dynamics. Our findings show the value of 1 as a diagnostic tool to rapidly identify the sites of matrix remodeling in tissue sections, which will aid in the conception of new therapeutic strategies for fibrotic disorders and defective tissue repair.
Assuntos
Proteína-Lisina 6-Oxidase , Cicatrização , Humanos , Camundongos , Animais , Proteína-Lisina 6-Oxidase/genética , Proteína-Lisina 6-Oxidase/metabolismo , Colágeno/metabolismo , Colágenos Fibrilares/genética , Fibrose , Peptídeos/farmacologiaRESUMO
Endothelial cell physiology is governed by its unique microenvironment at the interface between blood and tissue. A major contributor to the endothelial biophysical environment is blood hydrostatic pressure, which in mechanical terms applies isotropic compressive stress on the cells. While other mechanical factors, such as shear stress and circumferential stretch, have been extensively studied, little is known about the role of hydrostatic pressure in the regulation of endothelial cell behavior. Here we show that hydrostatic pressure triggers partial and transient endothelial-to-mesenchymal transition in endothelial monolayers of different vascular beds. Values mimicking microvascular pressure environments promote proliferative and migratory behavior and impair barrier properties that are characteristic of a mesenchymal transition, resulting in increased sprouting angiogenesis in 3D organotypic model systems ex vivo and in vitro. Mechanistically, this response is linked to differential cadherin expression at the adherens junctions, and to an increased YAP expression, nuclear localization, and transcriptional activity. Inhibition of YAP transcriptional activity prevents pressure-induced sprouting angiogenesis. Together, this work establishes hydrostatic pressure as a key modulator of endothelial homeostasis and as a crucial component of the endothelial mechanical niche.
Assuntos
Junções Aderentes , Pressão Hidrostática , Neovascularização Fisiológica , Transdução de Sinais , Proteínas de Sinalização YAP , Animais , Humanos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Junções Aderentes/metabolismo , Caderinas/metabolismo , Caderinas/genética , Movimento Celular , Células Endoteliais/metabolismo , Células Endoteliais/fisiologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Proteínas de Sinalização YAP/metabolismoRESUMO
Aging leads to biochemical and biomechanical changes in skin, with biological and functional consequences. Despite extensive literature on skin aging, there is a lack of studies which investigate the maturation of the tissue and connect the microscopic changes in the skin to its macroscopic biomechanical behavior as it evolves over time. The present work addresses this knowledge gap using multiscale characterization of skin in a murine model considering newborn, adult and aged mice. Monotonic uniaxial loading, tension relaxation with change of bath, and loading to failure tests were performed on murine skin samples from different age groups, complemented by inflation experiments and atomic force microscopy indentation measurements. In parallel, skin samples were characterized using histological and biochemical techniques to assess tissue morphology, collagen organization, as well as collagen content and cross-linking. We show that 1-week-old skin differs across nearly all measured parameters from adult skin, showing reduced strain stiffening and tensile strength, a thinner dermis, lower collagen content and altered crosslinking patterns. Surprisingly, adult and aged skin were similar across most biomechanical parameters in the physiologic loading range, while aged skin had lower tensile strength and lower stiffening behavior at large force values. This correlates with altered collagen content and cross-links. Based on a computational model, differences in mechanocoupled stimuli in the skin of the different age groups were calculated, pointing to a potential biological significance of the age-induced biomechanical changes in regulating the local biophysical environment of dermal cells. STATEMENT OF SIGNIFICANCE: Skin microstructure and the emerging mechanical properties change with age, leading to biological, functional and health-related consequences. Despite extensive literature on skin aging, only very limited quantitative data are available on microstructural changes and the corresponding macroscopic biomechanical behavior as they evolve over time. This work provides a wide-range multiscale mechanical characterization of skin of newborn, adult and aged mice, and quantifies microstructural correlations in tissue morphology, collagen content, organization and cross-linking. Remarkably, aged skin retained normal hydration and normal biomechanical function in the physiological loading range but showed significantly reduced properties at super-physiological loading. Our data show that age-related microstructural differences have a profound effect not only on tissue-level properties but also on the cell-level biophysical environment.
RESUMO
Granzyme B (GzmB) is a serine protease with intracellular and extracellular activities capable of regulating inflammation through cytokine processing and the apoptosis of effector cells. We tested the hypothesis that GzmB expression in T regulatory cells (Tregs) is required for the control of inflammatory responses and pathology during acute lung injury. To substantiate the clinical relevance of GzmB during lung injury, we performed GzmB immunohistochemistry on lung tissue from patients with acute respiratory distress syndrome (ARDS) and healthy control subjects. We also performed in vivo experiments with wild-type (WT) C57BL/6 and GzmB(-/-) mice exposed to a single intranasal instillation of bleomycin to model lung injury. Our results demonstrate that the expression of GzmB was elevated in ARDS lung sections, relative to healthy control samples. Bleomycin-exposed GzmB(-/-) mice exhibited greater morbidity and mortality, which was associated with increased numbers of lung lymphocytes. Bleomycin induced an equal increase in CD4(+)/CD25(+)/FoxP3(+) Treg populations in WT and GzmB(-/-) mice. GzmB expression was not significant in Tregs, with the majority of the expression localized to natural killer (NK)-1.1(+) cells. The expression of GzmB in NK cells of bleomycin-exposed WT mice was associated with greater lymphocyte apoptosis, reduced total lymphocyte numbers, and reduced pathology relative to GzmB(-/-) mice. Our data demonstrate that GzmB deficiency results in the exacerbation of lymphocytic inflammation during bleomycin-induced acute lung injury, which is associated with pathology, morbidity, and mortality.
Assuntos
Lesão Pulmonar Aguda/imunologia , Granzimas/imunologia , Células Matadoras Naturais/enzimologia , Pulmão/enzimologia , Pneumonia/imunologia , Síndrome do Desconforto Respiratório/imunologia , Linfócitos T Reguladores/enzimologia , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/enzimologia , Lesão Pulmonar Aguda/mortalidade , Administração Intranasal , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Bleomicina , Estudos de Casos e Controles , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Expressão Gênica , Granzimas/deficiência , Granzimas/genética , Humanos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/patologia , Pulmão/imunologia , Pulmão/patologia , Camundongos , Camundongos Knockout , Pneumonia/induzido quimicamente , Pneumonia/enzimologia , Pneumonia/mortalidade , Síndrome do Desconforto Respiratório/enzimologia , Síndrome do Desconforto Respiratório/genética , Síndrome do Desconforto Respiratório/patologia , Análise de Sobrevida , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologiaRESUMO
Atopic dermatitis is the most common inflammatory skin disease and is characterized by a deficient epidermal barrier and cutaneous inflammation. Genetic studies suggest a key role of keratinocytes in atopic dermatitis pathogenesis, but the alterations in the proteome that occur in the full epidermis have not been defined. Using a pressure-cycling technology and data-independent acquisition approach, we performed quantitative proteomics of epidermis from healthy volunteers and lesional and nonlesional patient skin. Results were validated by targeted proteomics using parallel reaction monitoring mass spectrometry and immunofluorescence staining. Proteins that were differentially abundant in the epidermis of patients with atopic dermatitis versus in healthy control reflect the strong inflammation in lesional skin and the defect in keratinocyte differentiation and epidermal stratification that already characterizes nonlesional skin. Most importantly, they reveal impaired activation of the NRF2-antioxidant pathway and reduced abundance of mitochondrial proteins involved in key metabolic pathways in the affected epidermis. Analysis of primary human keratinocytes with small interfering RNAâmediated NRF2 knockdown revealed that the impaired NRF2 activation and mitochondrial abnormalities are partially interlinked. These results provide insight into the molecular alterations in the epidermis of patients with atopic dermatitis and identify potential targets for pharmaceutical intervention.
Assuntos
Dermatite Atópica , Humanos , Dermatite Atópica/patologia , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Proteômica , Queratinócitos/metabolismo , Epiderme/patologia , Inflamação/patologia , Mitocôndrias/metabolismoRESUMO
Aging is associated with progressive skin fragility and a tendency to tear, which can lead to severe clinical complications. The transcription factor NRF2 is a key regulator of the cellular antioxidant response, and pharmacological NRF2 activation is a promising strategy for the prevention of age-related diseases. Using a combination of molecular and cellular biology, histology, imaging and biomechanical studies we show, however, that constitutive genetic activation of Nrf2 in fibroblasts of mice suppresses collagen and elastin expression, resulting in reduced skin strength as seen in aged mice. Mechanistically, the "aging matrisome" results in part from direct Nrf2-mediated overexpression of a network of microRNAs that target mRNAs of major skin collagens and other matrix components. Bioinformatics and functional studies revealed high NRF2 activity in aged human fibroblasts in 3D skin equivalents and human skin biopsies, highlighting the translational relevance of the functional mouse data. Together, these results identify activated NRF2 as a promoter of age-related molecular and biomechanical skin features.
Assuntos
MicroRNAs , Envelhecimento da Pele , Humanos , Camundongos , Animais , Idoso , Envelhecimento da Pele/genética , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Fibroblastos/metabolismo , Colágeno/genética , Colágeno/metabolismo , Pele/metabolismo , FenótipoRESUMO
Chronic, non-healing wounds constitute a major health problem, and the current therapeutic options are limited. Therefore, pharmaceuticals that can be locally applied to complicated wounds are urgently needed. Such treatments should directly target the underlying causes, which include diabetes mellitus, chronic local pressure and/or vascular insufficiency. A common consequence of these disorders is impaired wound angiogenesis. Here, we investigated the effect of topical application of a nitric oxide-releasing phosphodiesterase 5 inhibitor (TOP-N53)-containing liquid hydrogel on wound repair in mice. The drug-loaded hydrogel promoted re-epithelialization and angiogenesis in wounds of healthy and healing-impaired diabetic mice. Using a non-invasive label-free optoacoustic microscopy approach combined with automated vessel analysis, we show that the topical application of TOP-N53 formulation increases the microvascular network density and promotes the functionality of the newly formed blood vessels, resulting in enhanced blood perfusion of the wounds. These results demonstrate a remarkable healing-stimulating activity of topically applied TOP-N53 formulation, supporting its further development as a wound therapeutic.
RESUMO
The transcription factor nuclear factor-erythroid 2-related factor 2 (Nrf2) is widely recognized as a master regulator of the cellular stress response by facilitating the transcription of cytoprotective genes. As such, the Nrf2 pathway is critical in guarding the cell from the harmful effects of excessive reactive oxygen species/reactive nitrogen species (ROS/RNS) and in maintaining cellular redox balance. While excessive ROS/RNS are harmful to the cell, physiological levels of ROS/RNS play important roles in regulating numerous signaling pathways important for normal cellular function, including the synthesis of extracellular matrix (ECM). Recent advances have underscored the importance of ROS/RNS, and by extension, factors that influence redox-balance such as Nrf2, in regulating ECM production and deposition. In addition to reducing the oxidative burden in the cell, the discovery that Nrf2 can also directly target genes that regulate and form the ECM has cemented it as a multifaceted player in the regulation of ECM proteins, and provides new insight into its potential usefulness as a target for treating ECM-related pathologies.
RESUMO
Chronic wounds affect a large percentage of the population worldwide and cause significant morbidity. Unfortunately, efficient compounds for the treatment of chronic wounds are yet not available. Endothelial dysfunction, which is at least in part a result of compromised nitric oxide production and concomitant reduction in cGMP levels, is a major pathologic feature of chronic wounds. Therefore, we designed and synthesized a compound with a unique dual-acting activity (TOP-N53), acting as a nitric oxide donor and phosphodiesterase 5 inhibitor, and applied it locally to full-thickness skin wounds in healthy and healing-impaired mice with diabetes. TOP-N53 promoted keratinocyte proliferation, angiogenesis, and collagen maturation in healthy mice without accelerating the wound inflammatory response or scar formation. Most importantly, it partially rescued the healing impairment of mice with genetically determined type II diabetes (db/db) by stimulating re-epithelialization and granulation tissue formation, including angiogenesis. In vitro studies with human and murine primary cells showed a positive effect of TOP-N53 on keratinocyte and fibroblast migration, keratinocyte proliferation, and endothelial cell migration and tube formation. These results demonstrate a remarkable healing-promoting activity of TOP-N53 by targeting the major resident cells in the wound tissue.
Assuntos
Diabetes Mellitus Tipo 2/complicações , Pé Diabético/tratamento farmacológico , Doadores de Óxido Nítrico/farmacologia , Inibidores da Fosfodiesterase 5/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/genética , Pé Diabético/genética , Modelos Animais de Doenças , Feminino , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/fisiologia , Masculino , Camundongos , Camundongos Transgênicos , Neovascularização Fisiológica/efeitos dos fármacos , Doadores de Óxido Nítrico/uso terapêutico , Inibidores da Fosfodiesterase 5/uso terapêutico , Reepitelização/efeitos dos fármacosRESUMO
Psoriasis is a common inflammatory skin disorder that is characterized by keratinocyte hyperproliferation and abnormal differentiation, resulting in the thickening of the epidermis and stratum corneum. In this study, we investigated in vitro and in vivo pharmacological effects of tussilagonone (TGN), a sesquiterpenoid isolated from Tussilago farfara, on transcription factors relevant for the pathogenesis of psoriasis. TGN inhibited activation of NF-κB and STAT3, leading to the attenuated expression of psoriasis-related inflammatory genes and suppression of keratinocyte hyperproliferation. Mechanistically, we show that the inhibition of NF-κB and STAT3 by TGN is mediated through activation of the cytoprotective transcription factor NRF2. Evaluation of in vivo antipsoriatic effects of topical TGN in the imiquimod-induced psoriasis-like dermatitis mouse model demonstrated amelioration of imiquimod-induced phenotypical changes, lesion severity score, epidermal thickening, and reduction in dermal cellularity. The spleen index also diminished in TGN-treated mice, suggesting anti-inflammatory properties of TGN. Moreover, TGN significantly attenuated the imiquimod-induced mRNA levels of psoriasis-associated inflammatory cytokines and antimicrobial peptides and reduced epidermal hyperproliferation. Taken together, TGN, as a potent NRF2 activator, is a promising therapeutic candidate for the development of antipsoriatic agents derived from medicinal plants.
Assuntos
Anti-Inflamatórios/farmacologia , Fator 2 Relacionado a NF-E2/agonistas , Ácidos Pentanoicos/farmacologia , Psoríase/tratamento farmacológico , Sesquiterpenos/farmacologia , Administração Cutânea , Adulto , Animais , Anti-Inflamatórios/uso terapêutico , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Imiquimode/toxicidade , Queratinócitos/efeitos dos fármacos , Queratinócitos/patologia , Camundongos , Fator 2 Relacionado a NF-E2/metabolismo , Ácidos Pentanoicos/uso terapêutico , Psoríase/induzido quimicamente , Psoríase/imunologia , Psoríase/patologia , Sesquiterpenos/uso terapêutico , Tussilago/químicaRESUMO
The cytotoxic granzyme B (GrB)/perforin pathway has been traditionally viewed as a primary mechanism that is used by cytotoxic lymphocytes to eliminate allogeneic, virally infected and/or transformed cells. Although originally proposed to have intracellular and extracellular functions, upon the discovery that perforin, in combination with GrB, could induce apoptosis, other potential functions for this protease were, for the most part, disregarded. As there are 5 granzymes in humans and 11 granzymes in mice, many studies used perforin knockout mice as an initial screen to evaluate the role of granzymes in disease. However, in recent years, emerging clinical and biochemical evidence has shown that the latter approach may have overlooked a critical perforin-independent, pathogenic role for these proteases in disease. This review focuses on GrB, the most characterized of the granzyme family, in disease. Long known to be a pro-apoptotic protease expressed by cytotoxic lymphocytes and natural killer cells, it is now accepted that GrB can be expressed in other cell types of immune and nonimmune origin. To the latter, an emerging immune-independent role for GrB has been forwarded due to recent discoveries that GrB may be expressed in nonimmune cells such as smooth muscle cells, keratinocytes, and chondrocytes in certain disease states. Given that GrB retains its activity in the blood, can cleave extracellular matrix, and its levels are often elevated in chronic inflammatory diseases, this protease may be an important contributor to certain pathologies. The implications of sustained elevations of intracellular and extracellular GrB in chronic vascular, dermatological, and neurological diseases, among others, are developing. This review examines, for the first time, the multiple roles of GrB in disease pathogenesis.
Assuntos
Doença Crônica , Espaço Extracelular/enzimologia , Granzimas/fisiologia , Imunidade/fisiologia , Espaço Intracelular/enzimologia , Animais , Apoptose/fisiologia , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos Knockout , Perforina/fisiologiaRESUMO
Following tissue injury, cells produce reactive molecules that fight off invading pathogens, but these factors might also damage the host tissue. A new study has characterized a network of defense pathways that synergize to protect cells from collateral damage and drive repair.
RESUMO
Nrf2 is a key regulator of the antioxidant defense system, and pharmacological Nrf2 activation is a promising strategy for cancer prevention and promotion of tissue repair. Here we show, however, that activation of Nrf2 in fibroblasts induces cellular senescence. Using a combination of transcriptomics, matrix proteomics, chromatin immunoprecipitation and bioinformatics we demonstrate that fibroblasts with activated Nrf2 deposit a senescence-promoting matrix, with plasminogen activator inhibitor-1 being a key inducer of the senescence program. In vivo, activation of Nrf2 in fibroblasts promoted re-epithelialization of skin wounds, but also skin tumorigenesis. The pro-tumorigenic activity is of general relevance, since Nrf2 activation in skin fibroblasts induced the expression of genes characteristic for cancer-associated fibroblasts from different mouse and human tumors. Therefore, activated Nrf2 qualifies as a marker of the cancer-associated fibroblast phenotype. These data highlight the bright and the dark sides of Nrf2 and the need for time-controlled activation of this transcription factor.