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INTRODUCTION: TNFα-inducible matrix metalloproteinases play a critical role in the process of airway remodeling in respiratory inflammatory disease including asthma. The cationic host defense peptide LL-37 is elevated in the lungs during airway inflammation. However, the impact of LL-37 on TNFα-driven processes is not well understood. Here, we examined the effect of LL-37 on TNFα-mediated responses in human bronchial epithelial cells (HBECs). METHODS: We used a slow off-rate modified aptamer-based proteomics approach to define the HBEC proteome altered in response to TNFα. Abundance of selected protein candidates and signaling intermediates was examined using immunoassays, ELISA and Western blots, and mRNA abundance was examined by qRT-PCR. RESULTS: Proteomics analysis revealed that 124 proteins were significantly altered, 12 proteins were enhanced by ≥2-fold compared to unstimulated cells, in response to TNFα. MMP9 was the topmost increased protein in response to TNFα, enhanced by â¼10-fold, and MMP13 was increased by â¼3-fold, compared to unstimulated cells. Furthermore, we demonstrated that LL-37 significantly suppressed TNFα-mediated MMP9 and MMP13 in HBEC. Mechanistic data revealed that TNFα-mediated MMP9 and MMP13 production is controlled by SRC kinase and that LL-37 enhances related upstream negative regulators, namely, phospho-AKT (T308) and TNFα-mediated TNFAIP3 or A20. CONCLUSIONS: The findings of this study suggest that LL-37 may play a role in intervening in the process of airway remodeling in chronic inflammatory respiratory disease such as asthma.
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Remodelación de las Vías Aéreas (Respiratorias) , Péptidos Catiónicos Antimicrobianos , Asma , Bronquios , Catelicidinas , Células Epiteliales , Metaloproteinasa 13 de la Matriz , Metaloproteinasa 9 de la Matriz , Factor de Necrosis Tumoral alfa , Humanos , Péptidos Catiónicos Antimicrobianos/metabolismo , Asma/inmunología , Asma/metabolismo , Células Cultivadas , Células Epiteliales/metabolismo , Metaloproteinasa 13 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Proteómica , Mucosa Respiratoria/inmunología , Transducción de Señal , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Traffic-related air pollution (TRAP) exposure is associated with systemic health effects, which can be studied using blood-based markers. Although we have previously shown that high TRAP concentrations alter the plasma proteome, the concentration-response relationship between blood proteins and TRAP is unexplored in controlled human exposure studies. We aimed to identify concentration-dependent plasma markers of diesel exhaust (DE), a model of TRAP. Fifteen healthy non-smokers were enrolled into a double-blinded, crossover study where they were exposed to filtered air (FA) and DE at 20, 50 and 150 µg/m3 PM2.5 for 4h, separated by ≥ 4-week washouts. We collected blood at 24h post-exposure and used label-free mass spectrometry to quantify proteins in plasma. Proteins exhibiting a concentration-response, as determined by linear mixed effects models (LMEMs), were assessed for pathway enrichment using WebGestalt. Top candidates, identified by sparse partial least squares discriminant analysis and LMEMs, were confirmed using enzyme-linked immunoassays. Thereafter, we assessed correlations between proteins that showed a DE concentration-response and acute inflammatory endpoints, forced expiratory volume in 1 s (FEV1) and methacholine provocation concentration causing a 20% drop in FEV1 (PC20). DE exposure was associated with concentration-dependent alterations in 45 proteins, which were enriched in complement pathways. Of the 9 proteins selected for confirmatory immunoassays, based on complementary bioinformatic approaches to narrow targets and availability of high-quality assays, complement factor I (CFI) exhibited a significant concentration-dependent decrease (-0.02 µg/mL per µg/m3 of PM2.5, p = 0.04). Comparing to FA at discrete concentrations, CFI trended downward at 50 (-2.14 ± 1.18, p = 0.08) and significantly decreased at 150 µg/m3 PM2.5 (-2.93 ± 1.18, p = 0.02). CFI levels were correlated with FEV1, PC20 and nasal interleukin (IL)-6 and IL-1ß. This study details concentration-dependent alterations in the plasma proteome following DE exposure at concentrations relevant to occupational and community settings. CFI shows a robust concentration-response and association with established measures of airway function and inflammation.
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Contaminantes Atmosféricos , Emisiones de Vehículos , Humanos , Emisiones de Vehículos/toxicidad , Emisiones de Vehículos/análisis , Proteoma , Estudios Cruzados , Pruebas de Función Respiratoria , Interleucina-6 , Material Particulado/toxicidad , Material Particulado/análisis , Contaminantes Atmosféricos/toxicidad , Contaminantes Atmosféricos/análisisRESUMEN
IL-33 is an alarmin produced by stromal cells and is known to promote airway inflammation. IL-33 is a critical mediator of steroid-unresponsiveness in severe asthma. We have previously shown that IFNγ, a cytokine known to be elevated in airway inflammation and severe asthma, enhances the abundance of IL-33 in bronchial epithelial cells. Previous studies have shown that environmental insults such as particulate matter results in activation of the aryl hydrocarbon receptor (AhR) and IL-33 production. However, the role of AhR in cytokine-mediated IL-33 production is unknown. In this study, we demonstrate that the knockdown of AhR results in significant decrease in IFNγ-mediated IL-33 production and phosphorylation of STAT1 (Y701), in human bronchial epithelial cells. The findings of this report suggest that AhR may be an essential component in IFNγ-mediated IL-33 production in the lungs.
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Asma , Receptores de Hidrocarburo de Aril , Humanos , Receptores de Hidrocarburo de Aril/genética , Interleucina-33 , Citocinas , Células Epiteliales , Interferón gamma , InflamaciónRESUMEN
Curcumin, a component of the South-Asian spice turmeric, elicits anti-inflammatory functions. We have previously demonstrated that a highly bioavailable formulation of cucurmin, Cureit/Acumin™ (CUR), can suppress disease onset and severity, in a collagen-induced arthritis (CIA) mouse model. In a previous study, we have also shown that the abundance of antimicrobial host defence peptides, specifically cathelicidin (CRAMP) and calprotectin (S100A8 and S100A9), is significantly increased in the joint tissues of CIA mice. Elevated levels of cathelicidin and calprotectin have been associated with the pathogenesis of rheumatoid arthritis. Therefore, in this study, we examined the effect CUR administration on the abundance of cathelicidin and calprotectin in the joints, in a CIA mouse model. Here, we demonstrate that daily oral administration of CUR significantly reduces the elevated levels of CRAMP and calprotectin to baseline in the joints of CIA mice. We also show a linear correlation between the abundance of these peptides in the joints with serum inflammatory cytokines TNFα, IFNγ, and MCP-1. Overall, our results suggest that oral administration of a bioavailable CUR can suppress cathelicidin and calprotectin in the joints and regulate both local (joints) and systemic (serum) inflammation, in inflammatory arthritis.
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Artritis Experimental , Curcumina , Animales , Ratones , Péptidos Catiónicos Antimicrobianos , Catelicidinas , Complejo de Antígeno L1 de Leucocito , Artritis Experimental/tratamiento farmacológico , Modelos Animales de EnfermedadRESUMEN
Cell-free heme (CFH) is a product of hemoglobin, myoglobin and hemoprotein degradation, which is a hallmark of pathologies associated with extensive hemolysis and tissue damage. CHF and iron collectively induce cytokine storm, lung injury, respiratory distress and infection susceptibility in the lungs suggesting their key role in the progression of lung disease pathology. We have previously demonstrated that heme-mediated reactive oxygen species (ROS) induces platelet activation and ferroptosis. However, interaction of ferroptotic platelets and neutrophils, the mechanism of action and associated complications remain unclear. In this study, we demonstrate that heme-induced P-selectin expression and Phosphatidylserine (PS) externalization in platelets via ASK-1-inflammasome axis increases platelet-neutrophil aggregates in circulation, resulting in Neutrophil extracellular traps (NET) formation in vitro and in vivo. Further, heme-induced platelet activation in mice increased platelet-neutrophil aggregates and accumulation of NETs in the lungs causing pulmonary damage. Thus, connecting CFH-mediated platelet activation to NETosis and pulmonary thrombosis. As lung infections induce acute respiratory stress, thrombosis and NETosis, we propose that heme -mediated platelet activation and ferroptosis might be crucial in such clinical manifestations. Further, considering the ability of redox modulators and ferroptosis inhibitors like FS-1, Lpx-1 and DFO to inhibit heme-induced ferroptotic platelets-mediated NETosis and pulmonary thrombosis. They could be potential adjuvant therapy to regulate respiratory distress-associated clinical complications.
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Ferroptosis , Enfermedades Pulmonares , Síndrome de Dificultad Respiratoria , Trombosis , Ratones , Animales , Hemo , Activación Plaquetaria , Pulmón/patología , Trombosis/patologíaRESUMEN
Biological sex impacts disease prevalence, severity and response to therapy in asthma, however preclinical studies often use only one sex in murine models. Here, we detail sex-related differences in immune responses using a house dust mite (HDM)-challenge model of acute airway inflammation, in adult mice of two different strains (BALB/c and C57BL/6NJ). Female and male mice were challenged (intranasally) with HDM extract (~ 25 µg) for 2 weeks (N = 10 per group). Increase in serum HDM-specific IgE showed a female bias, which was statistically significant in BALB/c mice. We compared naïve and HDM-challenged mice to define immune responses in the lungs by assessing leukocyte accumulation in the bronchoalveolar lavage fluid (BALF), and profiling the abundance of 29 different cytokines in BALF and lung tissue lysates. Our results demonstrate specific sex-related and strain-dependent differences in airway inflammation. For example, HDM-driven accumulation of neutrophils, eosinophils and macrophages were significantly higher in females compared to males, in BALB/c mice. In contrast, HDM-mediated eosinophil accumulation was higher in males compared to females, in C57BL/6NJ mice. Differences in lung cytokine profiles indicated that HDM drives a T-helper (Th)17-biased response with higher IL-17 levels in female BALB/c mice compared to males, whereas female C57BL/6NJ mice elicit a mixed Th1/Th2-skewed response. Male mice of both strains showed higher levels of specific Th2-skewed cytokines, such as IL-21, IL-25 and IL-9, in response to HDM. Overall, this study details sex dimorphism in HDM-mediated airway inflammation in mice, which will be a valuable resource for preclinical studies in allergic airway inflammation and asthma.
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Asma , Pyroglyphidae , Femenino , Masculino , Ratones , Animales , Alérgenos , Caracteres Sexuales , Ratones Endogámicos C57BL , Dermatophagoides pteronyssinus , Inflamación , Ratones Endogámicos BALB C , CitocinasRESUMEN
BACKGROUND: The heterodimer interleukin (IL)-17A/F is elevated in the lungs in chronic respiratory disease such as severe asthma, along with the pro-inflammatory cytokine tumor necrosis factor-α (TNF-α). Although IL-17A/F and TNF-α are known to functionally cooperate to exacerbate airway inflammation, proteins altered by their interaction in the lungs are not fully elucidated. RESULTS: We used Slow Off-rate Modified Aptamer-based proteomic array to identify proteins that are uniquely and/or synergistically enhanced by concurrent stimulation with IL-17A/F and TNF-α in human bronchial epithelial cells (HBEC). The abundance of 38 proteins was significantly enhanced by the combination of IL-17A/F and TNF-α, compared to either cytokine alone. Four out of seven proteins that were increased > 2-fold were those that promote neutrophil migration; host defence peptides (HDP; Lipocalin-2 (LCN-2) and Elafin) and chemokines (IL-8, GROα). We independently confirmed the synergistic increase of these four proteins by western blots and ELISA. We also functionally confirmed that factors secreted by HBEC stimulated with the combination of IL-17A/F and TNF-α uniquely enhances neutrophil migration. We further showed that PI3K and PKC pathways selectively control IL-17A/F + TNF-α-mediated synergistic production of HDPs LCN-2 and Elafin, but not chemokines IL-8 and GROα. Using a murine model of airway inflammation, we demonstrated enhancement of IL-17A/F, TNF-α, LCN-2 and neutrophil chemokine KC in the lungs, thus corroborating our findings in-vivo. CONCLUSION: This study identifies proteins and signaling mediated by concurrent IL-17A/F and TNF-α exposure in the lungs, relevant to respiratory diseases characterized by chronic inflammation, especially neutrophilic airway inflammation such as severe asthma.
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Biological sex influences disease severity, prevalence and response to therapy in allergic asthma. However, allergen-mediated sex-specific changes in lung protein biomarkers remain undefined. Here, we report sex-related differences in specific proteins secreted in the lungs of both mice and humans, in response to inhaled allergens. Female and male BALB/c mice (7-8 weeks) were intranasally challenged with the allergen house dust mite (HDM) for 2 weeks. Bronchoalveolar lavage fluid (BALF) was collected 24 hour after the last HDM challenge from allergen-naïve and HDM-challenged mice (N=10 per group, each sex). In a human study, adult participants were exposed to nebulized (2 min) allergens (based on individual sensitivity), BALF was obtained after 24 hour (N=5 each female and male). The BALF samples were examined in immunoblots for the abundance of 10 proteins shown to increase in response to allergen in both murine and human BALF, selected from proteomics studies. We showed significant sex-bias in allergen-driven increase in five out of the 10 selected proteins. Of these, increase in eosinophil peroxidase (EPX) was significantly higher in females compared to males, in both mice and human BALF. We also showed specific sex-related differences between murine and human samples. For example, allergen-driven increase in S100A8 and S100A9 was significantly higher in BALF of females compared to males in mice, but significantly higher in males compared to females in humans. Overall, this study provides sex-specific protein biomarkers that are enhanced in response to allergen in murine and human lungs, informing and motivating translational research in allergic asthma.
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Alérgenos , Asma , Adulto , Alérgenos/efectos adversos , Animales , Asma/metabolismo , Biomarcadores/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Pulmón , Masculino , Ratones , Ratones Endogámicos BALB C , Pyroglyphidae , Caracteres SexualesRESUMEN
BACKGROUND: Traffic-related air pollution (TRAP) is a critical risk factor and major contributor to respiratory and cardiovascular disease (CVD). The effects of TRAP beyond the lungs can be related to changes in circulatory proteins. However, such TRAP-mediated changes have not been defined in an unbiased manner using a controlled human model. OBJECTIVE: To detail global protein changes (the proteome) in plasma following exposure to inhaled diesel exhaust (DE), a paradigm of TRAP, using controlled human exposures. METHODS: In one protocol, ex-smokers and never-smokers were exposed to filtered air (FA) and DE (300 µg PM2.5/m3), on order-randomized days, for 2 h. In a second protocol, independent never-smoking participants were exposed to lower concentrations of DE (20, 50 or 150 µg PM2.5/m3) and FA, for 4 h, on order-randomized days. Each exposure was separated by 4 weeks of washout. Plasma samples obtained 24 h post-exposure from ex-smokers (n = 6) were first probed using Slow off-rate modified aptamer proteomic array. Plasma from never-smokers (n = 11) was used for independent assessment of proteins selected from the proteomics study by immunoblotting. RESULTS: Proteomics analyses revealed that DE significantly altered 342 proteins in plasma of ex-smokers (n = 6). The top 20 proteins therein were primarily associated with inflammation and CVD. Plasma from never-smokers (n = 11) was used for independent assessment of 6 proteins, amongst the top 10 proteins increased by DE in the proteomics study, for immunoblotting. The abundance of all six proteins (fractalkine, apolipoproteins (APOB and APOM), IL18R1, MIP-3 and MMP-12) was significantly increased by DE in plasma of these never-smokers. DE-mediated increase was shown to be concentration-dependent for fractalkine, APOB and MMP-12, all biomarkers of atherosclerosis, which correlated with plasma levels of IL-6, a subclinical marker of CVD, in independent participants. CONCLUSION: This investigation details changes in the human plasma proteome due to TRAP. We identify specific atherosclerosis-related proteins that increase concentration-dependently across a range of TRAP levels applicable worldwide.
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Contaminantes Atmosféricos , Contaminación del Aire , Aterosclerosis , Contaminantes Atmosféricos/análisis , Contaminantes Atmosféricos/toxicidad , Contaminación del Aire/efectos adversos , Contaminación del Aire/análisis , Aterosclerosis/inducido químicamente , Aterosclerosis/etiología , Aterosclerosis/metabolismo , Humanos , Proteoma , Proteómica , Distribución Aleatoria , Emisiones de Vehículos/análisis , Emisiones de Vehículos/toxicidadRESUMEN
Innate defense regulator (IDR) peptides show promise as immunomodulatory therapeutics. However, there is limited understanding of the relationship of IDR peptide sequence and/or structure with its immunomodulatory activity. We previously reported that an IDR peptide, IDR-1002, reduces airway hyperresponsiveness (AHR) and inflammation in a house dust mite (HDM)-challenged murine model of airway inflammation. Here, we examined the sequence-to-function relationship of IDR-1002 in HDM-challenged mice and human bronchial epithelial cells (HBEC). We demonstrated that the tryptophan (W8) in the central hydrophobic region of IDR-1002 is required for the peptide to (i) suppress the pro-inflammatory cytokine IL-33, and induce anti-inflammatory mediators IL-1RA and stanniocalcin-1 in HBEC, and (ii) reduce IL-33 abundance, and eosinophil and neutrophil infiltration, in the lungs of HDM-challenged mice, without affecting the capacity to improve AHR, suggesting multimodal activity in vivo. Findings from this study can be used to design IDR peptides with targeted impact on immunomodulation and pathophysiology in respiratory diseases.
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Antiinflamatorios/farmacología , Péptidos Catiónicos Antimicrobianos/química , Inmunomodulación/efectos de los fármacos , Triptófano/química , Sustitución de Aminoácidos , Animales , Antiinflamatorios/química , Antiinflamatorios/uso terapéutico , Péptidos Catiónicos Antimicrobianos/farmacología , Péptidos Catiónicos Antimicrobianos/uso terapéutico , Asma/tratamiento farmacológico , Modelos Animales de Enfermedad , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Femenino , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Proteína Antagonista del Receptor de Interleucina 1/metabolismo , Interleucina-33/metabolismo , Ratones , Ratones Endogámicos BALB C , Infiltración Neutrófila/efectos de los fármacos , Análisis de Componente Principal , Estructura Secundaria de Proteína , Pyroglyphidae/patogenicidad , Triptófano/metabolismoRESUMEN
Bisphenol AF, an analogue of Bisphenol A, is an important raw material used in the production of plastic and rubber substances like plastic bottles and containers, toys, and medical supplies. Increased contamination of air, water, dust, and food with BPA/BPAF, poses an enormous threat to humans, globally. BPAF/BPA are endocrine-disrupting chemicals that mimic estrogen hormone, thus increasing the risks of various metabolic and chronic disorders. Exposure of human blood cells to BPA/BPAF induces oxidative stress and genotoxicity. However, its effects on platelets, which play central roles in hemostasis and thrombosis, are not well-documented. In this study, we demonstrate that BPAF induces RIPK1-inflammasome axis-mediated necroptosis in platelets, increasing procoagulant platelet levels in vivo and in vitro. We also show that BPAF-induced rise in procoagulant platelets worsens pulmonary thromboembolism in vivo. The elevated procoagulant platelets are shown to increase platelet-neutrophil/monocyte aggregates that mediate pathogenesis of CVD, thrombosis, and chronic inflammatory diseases. Our results demonstrate the toxic effects of BPAF on platelets and how it propagates the clinical complications by elevating procoagulant platelet numbers. Altogether, our study sends a cautionary message against extensive use of BPAF in the plastic and rubber industries, resulting in frequent human exposure to it, thus endangering platelet functions.
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Compuestos de Bencidrilo/toxicidad , Plaquetas/efectos de los fármacos , Disruptores Endocrinos/toxicidad , Necroptosis/efectos de los fármacos , Fenoles/toxicidad , Animales , Plaquetas/metabolismo , Femenino , Humanos , Inflamasomas/efectos de los fármacos , Inflamasomas/metabolismo , Masculino , Ratones , Embolia Pulmonar/fisiopatología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismoRESUMEN
OBJECTIVE: Curcumin (CUR), vitamin D3 (D3), and omega-3-fatty acids (O3FA) individually modulate inflammation and pain in arthritis. Although these supplements are widely used, their combinatorial effects have not been defined. In this study, we examined the effects of a D3 and O3FA (VO)-enriched diet in conjunction with a highly bioavailable form of CUR (Cureit/Acumin™) in a collagen-induced arthritis (CIA) murine model. METHODS: Male DBA/1J mice were acclimatized to VO-enriched diet and challenged with bovine collagen II (CII). Bioavailable CUR was administered daily by oral gavage from the onset of CII challenge. Disease severity was determined by monitoring joint thickness and standardized clinical score. Cellular infiltration and cartilage degradation in the joints were assessed by histology, serum cytokines profiled by Meso Scale Discovery multiplex assay, and joint matrix metalloproteinases examined by western blots. RESULTS: CUR by itself significantly decreased disease severity by ~ 60%. Administration of CUR in CIA mice taking a VO-enriched diet decreased disease severity by > 80% and maximally delayed disease onset and progression. Some of the disease-modifying effects was mediated by CUR alone, e.g., suppression of serum anti-collagen antibodies and decrease of cellular infiltration and MMP abundance in the joints of CIA mice. Although CUR alone suppressed inflammatory cytokines in serum of CIA mice, the combination of CUR and VO diet significantly enhanced the suppression (> 2-fold compared to CUR) of TNF, IFN-γ, and MCP-1, all known to be associated with RA pathogenesis. CONCLUSION: This study provides proof-of-concept that the combination of bioavailable CUR, vitamin D3, and O3FA substantially delays the development and severity of CIA. These findings provide a rationale for systematically evaluating these widely available supplements in individuals at risk for developing future RA.
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Artritis Experimental , Artritis Reumatoide , Curcumina , Animales , Artritis Experimental/tratamiento farmacológico , Bovinos , Curcumina/farmacología , Citocinas , Dieta , Modelos Animales de Enfermedad , Masculino , Ratones , Ratones Endogámicos DBA , Vitamina D , VitaminasRESUMEN
Cationic host defense peptides (CHDP) are immunomodulatory molecules that control infections and contribute to immune homeostasis. CHDP such as cathelicidin and calprotectin expression is altered in the arthritic synovium, and in the lungs of asthma and COPD patients. Recent studies suggest a link between airway inflammation and the immunopathology of arthritis. Therefore, in this study we compared the abundance of mouse cathelicidin (CRAMP), defensins, and calprotectin subunits (S100A8 and S100A9) in murine models of collagen-induced arthritis (CIA) and allergen house dust mite (HDM)-challenged airway inflammation. CRAMP, S100A8, and S100A9 abundance were significantly elevated in the joint tissues of CIA mice, whereas these were decreased in the lung tissues of HDM-challenged mice, compared to naïve. We further compared the effects of administration of two different synthetic immunomodulatory peptides, IG-19 and IDR-1002, on cathelicidin and calprotectin abundance in the two models. Administration of IG-19, which controls disease progression and inflammation in CIA mice, significantly decreased CRAMP, S100A8, and S100A9 levels to baseline in the joints of the CIA mice, which correlated with the decrease in cellular influx in the joints. However, administration of IDR-1002, which suppresses HDM-induced airway inflammation, did not prevent the decrease in the levels of cathelicidin and calprotectin in the lungs of HDM-challenged mice. Cathelicidin and calprotectin levels did not correlate with leukocyte accumulation in the lungs of the HDM-challenged mice. Results of this study suggest that endogenous cathelicidin and calprotectin abundance are disparately altered, and may be differentially regulated, within local tissues in airway inflammation compared to arthritis.
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Péptidos Catiónicos Antimicrobianos/metabolismo , Artritis Experimental/metabolismo , Asma/metabolismo , Articulaciones/metabolismo , Complejo de Antígeno L1 de Leucocito/metabolismo , Leucocitos/metabolismo , Pulmón/metabolismo , Alérgenos , Animales , Antígenos Dermatofagoides , Artritis Experimental/inducido químicamente , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/inmunología , Asma/inducido químicamente , Asma/tratamiento farmacológico , Asma/inmunología , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Colágeno Tipo II , Femenino , Factores Inmunológicos/farmacología , Articulaciones/efectos de los fármacos , Articulaciones/inmunología , Leucocitos/efectos de los fármacos , Leucocitos/inmunología , Lipopolisacáridos , Pulmón/efectos de los fármacos , Pulmón/inmunología , Masculino , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , CatelicidinasRESUMEN
IL-33 induces airway inflammation and hyper-responsiveness in respiratory diseases. Although defined as a therapeutic target, there are limited studies that have comprehensively investigated IL-33-mediated responses in the lungs in vivo. In this study, we characterized immunological and physiological responses induced by intranasal IL-33 challenge, in a mouse model. We identified specific cytokines, IL-4, IL-5, IL-6, IL-10, IP-10 and MIP1-α, that are increased in bronchoalveolar lavage and lung tissues by IL-33. Using transcriptomics (RNA-Seq) we demonstrated that 2279 transcripts were up-regulated and 1378 downregulated (≥ 2-fold, p < 0.01) in lung tissues, in response to IL-33. Bioinformatic interrogation of the RNA-Seq data was used to predict biological pathways and upstream regulators involved in IL-33-mediated responses. We showed that the mRNA and protein of STAT4, a predicted upstream regulator of IL-33-induced transcripts, was significantly enhanced in the lungs following IL-33 challenge. Overall, this study provides specific IL-33-induced molecular targets and endpoints that can be used as a resource for in vivo studies, e.g. in preclinical murine models examining novel interventions to target downstream effects of IL-33.
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Interleucina-33/inmunología , Pulmón/inmunología , Pulmón/metabolismo , Modelos Animales , Transcriptoma , Administración Intranasal , Animales , Femenino , Interleucina-33/administración & dosificación , Pulmón/patología , Ratones , Ratones Endogámicos BALB C , RNA-SeqRESUMEN
Melatonin is a chronobiotic hormone, which can regulate human diseases like cancer, atherosclerosis, respiratory disorders, and microbial infections by regulating redox system. Melatonin exhibits innate immunomodulation by communicating with immune system and influencing neutrophils to fight infections and inflammation. However, sustaining redox homeostasis and reactive oxygen species (ROS) generation in neutrophils are critical during chemotaxis, oxidative burst, phagocytosis, and neutrophil extracellular trap (NET) formation. Therefore, endogenous antioxidant glutathione (GSH) redox cycle is highly vital in regulating neutrophil functions. Reduced intracellular GSH levels and glutathione reductase (GR) activity in the neutrophils during clinical conditions like autoimmune disorders, neurological disorders, diabetes, and microbial infections lead to dysfunctional neutrophils. Therefore, we hypothesized that redox modulators like melatonin can protect neutrophil health and functions under GSH and GR activity-deficient conditions. We demonstrate the dual role of melatonin, wherein it protects neutrophils from oxidative stress-induced apoptosis by reducing ROS generation; in contrast, it restores neutrophil functions like phagocytosis, degranulation, and NETosis in GSH and GR activity-deficient neutrophils by regulating ROS levels both in vitro and in vivo. Melatonin mitigates LPS-induced neutrophil dysfunctions by rejuvenating GSH redox system, specifically GR activity by acting as a parallel redox system. Our results indicate that melatonin could be a potential auxiliary therapy to treat immune dysfunction and microbial infections, including virus, under chronic disease conditions by restoring neutrophil functions. Further, melatonin could be a promising immune system booster to fight unprecedented pandemics like the current COVID-19. However, further studies are indispensable to address the clinical usage of melatonin.
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Antioxidantes/uso terapéutico , Glutatión/metabolismo , Melatonina/uso terapéutico , Neutrófilos/efectos de los fármacos , Animales , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Infecciones por Coronavirus/tratamiento farmacológico , Evaluación Preclínica de Medicamentos , Femenino , Glutatión Reductasa/metabolismo , Humanos , Masculino , Melatonina/farmacología , Ratones , Mitocondrias/metabolismo , NADPH Oxidasas/metabolismo , Estrés Oxidativo/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Tratamiento Farmacológico de COVID-19RESUMEN
Curcumin, a major bioactive component of turmeric (Curcuma longa), is known for its multiple health benefits. Curcumin as such is a mixture of its analogs: bisdemethoxycurcumin (BDMC)-3%, and demethoxycurcumin (DMC)-17%. Although the effect of curcumin on platelets is documented, the effect of BDMC and DMC on platelets is less studied. Considering the indispensable role played by platelets in hemostasis, thrombosis, inflammation, and immunity, the present study evaluates the effect of curcumin, DMC and BDMC on platelet apoptosis. The components of curcumin were purified by silica-gel column chromatography. The purity and mass analysis of the purified curcuminoids was determined by RP-HPLC and LC-MS respectively. When analyzed for platelet apoptotic markers, only BDMC demonstrated increased incidence of platelet apoptotic markers including increase in intracellular Ca2+, decrease in ∆ψm, alteration in BCl-2 family proteins, the release of cytochrome c, caspase activation, and PS externalization via activation of ERK activation. ERK inhibitor PD98059 significantly alleviated BDMC induced decrease in ∆ψm, alteration in BCl-2, caspase-8 activation and PS externalization. Our results demonstrate that curcumin, DMC and BDMC differentially act on platelet in inducing apoptosis and the study highlights that the toxicity associated with curcumin therapy might be attributed to BDMC in the mammalian system.
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Plaquetas/efectos de los fármacos , Diarilheptanoides/toxicidad , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Apoptosis/efectos de los fármacos , Plaquetas/metabolismo , Curcumina/toxicidad , HumanosRESUMEN
OBJECTIVE: Epidemiological studies suggest vitamin D deficiency as a potential risk factor for rheumatoid arthritis (RA) development, a chronic autoimmune disorder highly prevalent in indigenous North American (INA) population. We therefore profiled the circulating levels of 25-hydroxyvitaminD [25(OH)D], an active metabolite of vitamin D, in a cohort of at-risk first-degree relatives (FDR) of INA RA patients, a subset of whom subsequently developed RA (progressors). METHODS: 2007 onward, serum samples from INA RA patients and FDR were collected at the time of a structured baseline visit and stored at -20°C. Anti-citrullinated protein antibodies (ACPA), 25(OH)D, hs-CRP, vitamin-D binding protein (VDBP) and parathyroid hormone (PTH) levels were determined using ELISA and rheumatoid factor (RF) seropositivity was determined by nephelometry. RESULTS: We demonstrate that 25 (OH) D concentrations were lower in winter than summer (P = 0.0538), and that serum 25(OH)D levels were higher in samples collected and stored after 2013 (P<0.0001). Analysis of samples obtained after 2013 demonstrated that 37.6% of study participants were 25(OH)D insufficient (<75nmol/L). Also, seropositive RA patients and FDR had lower 25(OH)D levels compared to ACPA-/FDR (P<0.05, P<0.01 respectively). Linear regression analysis showed 25(OH)D insufficiency was inversely associated with presence of RA autoantibodies. Longitudinal samples from 14 progressors demonstrated a consistent increase in 25(OH)D levels at the time they exhibited clinically detectable joint inflammation, without any significant change in VDBP or PTH levels. Spearman rank correlation analysis showed significant association between 25(OH)D and PTH levels, both in RA patients and progressors at RA onset time. CONCLUSION: We demonstrate that 25(OH)D levels in serum increased at RA onset in progressors. The potential role that vitamin D metabolites and their downstream effects play in RA transition requires further investigation.
Asunto(s)
Artritis Reumatoide/sangre , Biomarcadores , Vitamina D/análogos & derivados , Adulto , Anticuerpos Antiproteína Citrulinada/sangre , Anticuerpos Antiproteína Citrulinada/inmunología , Artritis Reumatoide/etiología , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana Edad , Factor Reumatoide/sangre , Factor Reumatoide/inmunología , Estaciones del Año , Vitamina D/sangreRESUMEN
The anti-endotoxin activity of the cationic peptide LL-37 and its derivative IG-19 is attributed to electrostatic interaction of the peptides' positive charge with negatively charged bacterial lipopolysaccharides (LPS), and in part to the alteration of intracellular mechanisms independent of peptide binding to LPS. We examined the immunomodulatory responses induced by IG-19 and four IG-19-derived scrambled peptides (IG-19a-d), in the presence and absence of LPS, in macrophages and peripheral blood-derived mononuclear cells. All peptides had identical net charge (+5) and amino acid composition, but different hydrophobicity and α-helical propensity. Peptide IG-19 suppressed LPS-induced cytokine/chemokine production by >90%, IG-19a and IG-19b suppressed it by 40-50%, and IG-19c and IG-19d did not suppress cytokine/chemokine production at all. In silico prediction algorithms and the peptide retention time (RT) on a C18 RP HPLC column indicated a linear association between α-helical propensity and hydrophobicity with the ability of the peptides to inhibit LPS-induced responses. Peptide RT exhibited a significant correlation (>70%) between the suppression of LPS-induced cytokine/chemokine production and peptide-induced production of the anti-inflammatory cytokine IL-1RA. These results indicate that RT on a C18 column can be used as a predictor for the immunomodulatory functions of cationic peptides. Overall, we demonstrated that the immunomodulatory functions of LL-37-derived peptides with identical positive charge and amino acid composition are directly associated with the predicted α-helical propensity and hydrophobicity of the peptides.
Asunto(s)
Antiinflamatorios/farmacología , Péptidos Catiónicos Antimicrobianos/química , Lipopolisacáridos/efectos adversos , Péptidos/farmacología , Antiinflamatorios/química , Simulación por Computador , Citocinas/metabolismo , Regulación hacia Abajo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Proteína Antagonista del Receptor de Interleucina 1/metabolismo , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/inmunología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Péptidos/química , Conformación Proteica en Hélice alfa , Células THP-1 , CatelicidinasRESUMEN
BACKGROUND: Arthritis is a common degenerative joint disease characterized by deterioration of articular cartilage, subchondral bone, and associated with immobility, pain and inflammation. The incessant action of reactive oxygen species (ROS) during progressive arthritis causes severe oxidative damage to vital organs and circulatory system. PURPOSE: In this study we investigated the ability of guggulipid (GL), a lipid rich extract from the gum resin of the plant Commiphora whighitii to suppress the progressive arthritis and associated liver oxidative stress both in vivo and in vitro. STUDY DESIGN/METHODS: The anti-arthritic ability of GL was demonstrated in vitro using IL-1ß stimulated bovine nasal cartilage model and in vivo Freund's complete adjuvant-induced arthritic rat model. Collagen/proteoglycan degradation and pro-inflammatory mediators were monitored in the harvested culture medium of nasal cartilage by estimating the levels of matrix metalloproteinases (MMPs), hydroxy proline, glycosaminoglycans and inflammatory mediators. Further, anti-arthritic ability of GL was evaluated in vivo by measuring enzymatic and non-enzymatic mediators of cartilage degradation, inflammation and oxidative stress markers. RESULTS: GL significantly inhibited the IL-1ß stimulated cartilage degradation in vitro by mitigating the MMPs activity, collagen degradation and secretion of pro-inflammatory mediators. Further, GL significantly reduced the adjuvant-induced paw swelling and body weight loss in vivo. GL remarkably reduced the MMPs and hyaluronidases activities in serum and bone homogenate along with altered hematological parameters. GL also mitigated the elevated bone resorbing enzymes cathepsins, exoglycosidases and phosphatases. Additionally, GL effectively mitigated ROS and oxidative stress-mediators recuperating the altered serum/liver oxidative stress and liver damage incurred during arthritic progression. CONCLUSION: In summary, the study clearly demonstrates the protective efficacy of GL against arthritis and its associated oxidative stress, particularly, liver oxidative damage. Hence, GL could be a potential alternative and complementary medicine to treat inflammatory joint diseases.