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Accurate screening of COVID-19 infection status for symptomatic patients is a critical public health task. Although molecular and antigen tests now exist for COVID-19, in resource-limited settings, screening tests are often not available. Furthermore, during the early stages of the pandemic tests were not available in any capacity. We utilized an automated machine learning (ML) approach to train and evaluate thousands of models on a clinical dataset consisting of commonly available clinical and laboratory data, along with cytokine profiles for patients (n = 150). These models were then further tested for generalizability on an out-of-sample secondary dataset (n = 120). We were able to develop a ML model for rapid and reliable screening of patients as COVID-19 positive or negative using three approaches: commonly available clinical and laboratory data, a cytokine profile, and a combination of the common data and cytokine profile. Of the tens of thousands of models automatically tested for the three approaches, all three approaches demonstrated > 92% sensitivity and > 88 specificity while our highest performing model achieved 95.6% sensitivity and 98.1% specificity. These models represent a potential effective deployable solution for COVID-19 status classification for symptomatic patients in resource-limited settings and provide proof-of-concept for rapid development of screening tools for novel emerging infectious diseases.
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COVID-19 , Citocinas , Aprendizado de Máquina , Humanos , COVID-19/diagnóstico , Citocinas/sangue , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/imunologia , Programas de Rastreamento/métodos , Masculino , Feminino , Sensibilidade e Especificidade , Pessoa de Meia-Idade , Adulto , IdosoRESUMO
PURPOSE: Experimental studies have shown that prolonged sitting for 2-8 h can cause changes to vascular and metabolic markers; the response of pro-inflammatory cytokines is relatively unexplored. The purpose of this study is to determine the response of interleukin-8 (IL-8) to prolonged and interrupted sitting. METHODS: Healthy participants (n = 24, 21.1 years ± 2.2, 50% female) completed a prolonged sitting session (4 h) and an interrupted sitting session (4 h of sitting with 3 min of walking at 60%HRmax, every 30 min) in random order. Saliva and capillary plasma were collected at the beginning (T1) and at the end of each session (T2). RESULTS: Salivary concentrations of IL-8 increased during the prolonged (T1 median: 22.09 pg/mL, T2 median: 86.18 pg/mL; p = < 0.01, ES - 0.55) and interrupted (T1 median: 22.09 pg/mL, T2 median: 51.99 pg/mL; p = 0.021, ES - 0.34) sessions; however, the increase during interrupted sitting was lower (PS median: 134.4%, range: - 43.96 to 1115.69 and IS median: 50.8%, range: - 75.5 to 356.35; p = 0.011, ES - 0.53). In the sub-sample of males, salivary IL-8 did not increase in the interrupted session (T1 median: 22.09, range: 3.496-699.12, and T2 median: 24.96, range: 5.11-533.5, p = > 0.05, ES - 0.16). No significant findings were observed for IL-8 in the plasma. CONCLUSION: Prolonged sitting appears to increase concentrations of the pro-inflammatory cytokine IL-8 while interrupting this sitting with short bouts of walking blunts this response. Sex appears to moderate this relationship; however, there appears to be a large amount of individual variability.
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Interleucina-8 , Feminino , Humanos , Masculino , Citocinas , Comportamento Sedentário , Caminhada/fisiologia , Adulto JovemRESUMO
High salt (NaCl) concentrations are found in a number of tissues under physiological and pathological conditions. Here, we analyzed the effects induced by high salt on the function of human neutrophils. The culture of neutrophils in medium supplemented with high salt (50 mM NaCl) for short periods (30-120 min) inhibited the ability of conventional agonists to induce the production of IL-8 and the activation of respiratory burst. By contrast, exposure to high salt for longer periods (6-18 h) resulted in the activation of neutrophils revealed by the production of high levels of IL-8, the activation of the respiratory burst, and a marked synergistic effect on the production of TNF-α induced by LPS. Increasing osmolarity of the culture medium by the addition of sorbitol or mannitol (100 mM) was shown to be completely unable to stimulate neutrophil responses, suggesting that high sodium but not an increased osmolarity mediates the activation on neutrophils responses. A similar biphasic effect was observed when the function of monocytes was analyzed. Short term exposure to high salt suppressed IL-8 and TNF-α production induced by LPS while culture for longer periods triggered the production of IL-8 but not TNF-α in the absence of LPS stimulation. Contradictory results have been published regarding how high salt modulates neutrophil function. Our results suggest that the modulation of neutrophil function by high salt is strongly dependent on the exposure time.
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Neutrófilos , Fator de Necrose Tumoral alfa , Humanos , Interleucina-8/farmacologia , Lipopolissacarídeos/farmacologia , Neutrófilos/patologia , Cloreto de Sódio/farmacologia , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Although extracellular carbonylated proteins (CPs) are found at higher levels in sun-exposed skin, their impact on the cellular functions of fibroblasts and their involvement in the progression of photoaging skin are not fully clarified. In our previous study, we reported that extracellular CPs increase levels of intracellular oxidative stress and result in the accumulation of newly synthesized CPs in normal human dermal fibroblasts (NHDF). Furthermore, fibroblasts exposed to CP-BSA, which is a model of extracellular CPs, had upregulated expression levels of mRNAs encoding matrix metalloproteinase-1 (MMP-1) and interleukin-8/CXCL8 (IL-8/CXCL8). These facts suggested the possibility that extracellular CPs induce a fragile structure in the dermis through the degradation of collagen and elastin. The purpose of this study was to characterize the efficacy of natural carotenoids, such as astaxanthin analogs, produced by Hematococus pluvialis (CHPs) to improve the impaired functions of fibroblasts exposed to CPs. CHPs suppressed the intracellular CP levels elevated by CP-BSA, restored mRNA expression levels of factors involved in the formation and assembly of collagen and elastin fibers and improved the formation of those fibers impaired by CP-BSA. We conclude that CHPs function as antiaging substances due to their restoration of the impaired formation of collagen and elastin fibers caused by extracellular soluble CPs.
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Fibroblastos/metabolismo , Carbonilação Proteica/efeitos dos fármacos , Carbonilação Proteica/fisiologia , Envelhecimento da Pele/efeitos dos fármacos , Xantofilas/farmacologia , Células Cultivadas , Colágeno/metabolismo , Derme/citologia , Elastina/metabolismo , Fibroblastos/efeitos dos fármacos , Expressão Gênica , Humanos , Interleucina-8/genética , Interleucina-8/metabolismo , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 1 da Matriz/metabolismo , Estresse Oxidativo/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Envelhecimento da Pele/genéticaRESUMO
Only scarce data pertaining to interleukin 8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) chemokines in human aneurysm can be found in the current literature. Therefore, the aim of this study was the evaluation of cerebrospinal fluid (CSF) and serum IL-8 and MCP-1 concentration in unruptured intracranial aneurysm (UIA) patients (n = 25) compared to the control group (n = 20). IL-8 and MCP-1 concentrations were measured with ELISA method. We demonstrated that CSF IL-8 concentration of UIA patients is significantly higher (p < 0.001) than that presented in the serum, which can indicate its local synthesis within central nervous system. CSF IL-8 concentration was also significantly related to aneurysm size, which may reflect the participation of IL-8 in the formation and development of brain aneurysms. IL-8 Quotient (CSF IL-8 divided by serum IL-8) in UIA patients was statistically higher compared to control individuals (p = 0.045). However, the diagnostic utility analysis did not equivocally indicate the diagnostic usefulness of the IL-8 Quotient evaluation in brain aneurysm patients. Nevertheless, this aspect requires further study.
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In up to 15% of acute myeloid leukemias (AMLs), a recurring chromosomal translocation, termed t(8;21), generates the AML1-eight-twenty-one (ETO) leukemia fusion protein, which contains the DNA-binding domain of Runt-related transcription factor 1 (RUNX1) and almost all of ETO. RUNX1 and the AML1-ETO fusion protein are coexpressed in t(8;21) AML cells and antagonize each other's gene-regulatory functions. AML1-ETO represses transcription of RUNX1 target genes by competitively displacing RUNX1 and recruiting corepressors such as histone deacetylase 3 (HDAC3). Recent studies have shown that AML1-ETO and RUNX1 co-occupy the binding sites of AML1-ETO-activated genes. How this joined binding allows RUNX1 to antagonize AML1-ETO-mediated transcriptional activation is unclear. Here we show that RUNX1 functions as a bona fide repressor of transcription activated by AML1-ETO. Mechanistically, we show that RUNX1 is a component of the HDAC3 corepressor complex and that HDAC3 preferentially binds to RUNX1 rather than to AML1-ETO in t(8;21) AML cells. Studying the regulation of interleukin-8 (IL8), a newly identified AML1-ETO-activated gene, we demonstrate that RUNX1 and HDAC3 collaboratively repress AML1-ETO-dependent transcription, a finding further supported by results of genome-wide analyses of AML1-ETO-activated genes. These and other results from the genome-wide studies also have important implications for the mechanistic understanding of gene-specific coactivator and corepressor functions across the AML1-ETO/RUNX1 cistrome.
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Subunidade alfa 2 de Fator de Ligação ao Core/genética , Histona Desacetilases/genética , Leucemia Mieloide Aguda/genética , Proteínas de Fusão Oncogênica/genética , Proteína 1 Parceira de Translocação de RUNX1/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Genoma Humano/genética , Humanos , Interleucina-8/genética , Leucemia Mieloide Aguda/patologia , Regiões Promotoras Genéticas , Ativação Transcricional/genética , Translocação Genética/genéticaRESUMO
IL-8 (CXCL-8) is a chemoattractant factor for myeloid leukocytes, that is produced in large quantities by many solid tumors. Levels of IL-8, which can act upon a variety of immune and nonimmune cells, can tell us a lot about tumors, including their size (positive association) and how likely they are to respond to immunotherapy (negative association). This is because the IL-8 produced by tumors can promote angiogenesis, recruit immunosuppressive cells like neutrophils and myeloid-derived suppressor cells (MDSCs), and stimulate epithelial-to-mesenchymal transition, which is a precursor to metastasis. In a pooled analysis of several clinical trials in kidney cancer, melanoma, and lung cancer, it was found that patients with higher baseline concentrations of IL-8 in the blood experienced worse outcomes and lower overall survival after being treated with immunotherapy. Currently, the field that relates IL-8 to immunotherapy is leading to numerous and promising clinical trials that combine the inhibition of IL-8 with existing immunotherapeutic therapies. For this reason, multiple constructs based on IL-8 agonists are being developed clinically by the pharmaceutical and biotech industries.
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Melanoma , Células Supressoras Mieloides , Humanos , Imunoterapia , Interleucina-8RESUMO
Thrombin plays a crucial role in lung inflammatory diseases such as asthma and chronic obstructive pulmonary disease (COPD). Thrombin induces the release of interleukin-8 (IL-8)/CXCL8 by lung epithelial cells, and this phenomenon plays a vital role in lung inflammation. Our previous studies have indicated that thrombin stimulates IL-8/CXCL8 expression through PI3K/Akt/IκB kinase (IKK)α/ß/nuclear factor-κB (NF-κB) and p300 pathways in human lung epithelial cells. In the present study, we explored the roles of mammalian target of rapamycin (mTOR) and p70S6 kinase (p70S6K) in thrombin-induced NF-κB activation and IL-8/CXCL8 release in human lung epithelial cells. In this study, we found that rapamycin (an mTOR inhibitor) and p70S6K siRNA diminished thrombin-induced IL-8/CXCL8 release. Thrombin induced mTOR Ser2448 phosphorylation and p70S6K Thr389 phosphorylation in a time-dependent manner. Moreover, rapamycin attenuated thrombin-stimulated p70S6K phosphorylation. We also found that transfection of cells with the dominant negative mutant of Akt (Akt DN) reduced the thrombin-induced increase in mTOR phosphorylation and p70S6K phosphorylation. Moreover, thrombin-stimulated p300 phosphorylation was attenuated by Akt DN, rapamycin, and p70S6K siRNA. Thrombin triggered p70S6K translocation from the cytosol to the nucleus in a time-dependent manner. Thrombin induced the complex formation of p70S6K, p300, and p65; acetylation of p65 Lys310, and recruitment of p70S6K, p300, and p65 to the κB-binding site of the IL-8/CXCL8 promoter region. In conclusion, these results indicate that thrombin initiates the Akt-dependent mTOR/p70S6K signaling pathway to promote p300 phosphorylation and NF-κB activation and finally induces IL-8/CXCL8 release in human lung epithelial cells.
Assuntos
Pulmão/imunologia , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais/imunologia , Serina-Treonina Quinases TOR/metabolismo , Trombina/metabolismo , Células A549 , Asma/imunologia , Asma/patologia , Proteína p300 Associada a E1A/metabolismo , Células Epiteliais/imunologia , Células Epiteliais/patologia , Humanos , Interleucina-8/metabolismo , Pulmão/patologia , Fosforilação/imunologia , Doença Pulmonar Obstrutiva Crônica/imunologia , Doença Pulmonar Obstrutiva Crônica/patologia , Fator de Transcrição RelA/metabolismoRESUMO
PURPOSE: To evaluate the expression of IL8/CXCL8 cytokine and its receptor CXCR1 in tear film and ocular surface of patients with ocular mucous membrane pemphigoid (oMMP). METHODS: Ten patients with oMMP in the quiescent phase, 20 patients with primary Sjogren syndrome (pSS) and 13 age- and sex-matched healthy controls (HCs) were included in this study. All patients undergone complete eye examination including lacrimal function tests and ocular surface staining assessed by ocular staining score. Ocular mucous membrane pemphigoid (oMMP) staging according to Mondino classification and dry eye severity grade according to Dry Eye Workshop 2007 classification were recorded. Tear samples and conjunctival epithelium were collected. IL-8 tear concentration was measured by enzyme-linked immunosorbent assay, and conjunctival IL8 was analysed by Western blot; conjunctival expression of CXCR1 was evaluated by immunohistochemistry. Il-8 and CXCR1 expression in oMMP patients were compared with HCand pSS patients and correlated with ocular clinical findings. RESULTS: Tear levels of IL-8 were significantly increased in patients with oMMP (260.1 ± 70 pg/ml) when compared to both HCs (98.5 ± 71.35 pg/ml; p = 0.001) and patients with pSS (96.3 ± 87.5 pg/ml; p = 0.001). Conjunctival expression of IL8 and CXCR1 was significantly increased in oMMP patients when compared to both healthy subjects and pSS patients. CONCLUSION: The significant increase of tear and conjunctival IL8 and CXCR1 levels in patients with oMMP when compared to healthy subjects and patients with Sjogren syndrome suggests that changes of IL8 pathway are specific of oMMP and may represent a potential biomarker of the disease and/or therapeutic target.
Assuntos
Túnica Conjuntiva/metabolismo , Interleucina-8/metabolismo , Penfigoide Mucomembranoso Benigno/metabolismo , Receptores de Interleucina-8A/metabolismo , Lágrimas/metabolismo , Adulto , Idoso , Biomarcadores/metabolismo , Western Blotting , Ensaio de Imunoadsorção Enzimática , Epitélio/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Penfigoide Mucomembranoso Benigno/diagnóstico , Síndrome de Sjogren/metabolismoRESUMO
BACKGROUND: The serum and tear levels of four inflammatory chemokines were evaluated in sulfur mustard (SM)-exposed with serious ocular problems. MATERIALS AND METHODS: In this study, 128 SM-exposed patients and 31 healthy control participants participated. Tear and serum levels of chemokines were assessed by ELISA method. RESULTS: There was no significant difference in the serum level of IL-8/CXCL8, CX3CL1/fractalkine, CCL2/MCP-1, and CCL5/RANTES between all SM-exposed subjects and control groups. The tear level of IL-8 in the SM-exposed group was lower than the control group, but the difference was not significant. In the SM-exposed group with the abnormalities in tear breakup time (TBUT) test, fundus and pannus formation were significantly higher than SM-exposed patients without these problems. CX3CL1 levels have significantly increased in SM-exposed group with blepharitis, pterygium, and conjunctival pigmentation as compared with the control group. Besides, significantly higher levels of CX3CL1 were observed in SM-exposed group with or without bulbar conjunctival hyperemia and abnormal vessels a well as with fundus abnormality compared to the control group. Only, SM-exposed group with subconjunctival fibrosis had significantly lower levels of CCL5 than SM-exposed group without this problem. CONCLUSION: The higher level of CX3CL1 and consistent levels of IL-8/CXCL8, MCP-1/CCL2, and RANTES/CCL5 in SM-exposed individuals may indicate an anti-inflammatory response against the destructive effects of SM gas. High tear level of IL-8/CXCL8 reflects the severity of ocular surface abnormalities, yet significantly low tear level found in mild SM-exposed subgroup compared with the control group. The lower levels of CX3CL1 and RANTES/CCL5 may represent the different pathophysiology which requires further studies.
Assuntos
Substâncias para a Guerra Química/toxicidade , Citocinas/metabolismo , Traumatismos Oculares/metabolismo , Gás de Mostarda/toxicidade , Lágrimas/metabolismo , Adulto , Citocinas/sangue , Traumatismos Oculares/sangue , Traumatismos Oculares/induzido quimicamente , Humanos , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de DoençaRESUMO
Cultured epithelial autograft (CEA) was the birth of skin tissue engineering and encompassed methodologies for the isolation and expansion of autologous basal keratinocytes for burn treatment that are still practiced at some specialised units around the world. One of the limitations of CEA, however, is the reliance on animal-derived material during the manufacturing process and despite all efforts to date, no xeno-free alternative with proven efficacy has been reported. Here, we investigate whether human-derived fibroblast feeder cells and human serum can sufficiently and effectively provide a suitable microenvironment for adult keratinocyte isolation and expansion. Human dermal fibroblasts and epidermal keratinocytes were isolated from discarded skin during abdominoplasty and breast reduction procedures and cultured in xeno-free conditions. We report that these xeno-free adult keratinocytes form similar numbers of colony-forming units as those cultured using the Green's methods; however, xeno-free keratinocytes express lower levels of α6 integrin (CD49f; a progenitor and stem cell marker). We identified IL-8 as a potential growth factor secreted by adult human fibroblasts that may enhance keratinocyte colony formation in human serum. Finally, we propose a step-by-step xeno-free isolation and cultivation methodology for adult keratinocytes that can be tested further in serial cultivation for clinical application.
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Células Alimentadoras , Queratinócitos/citologia , Engenharia Tecidual/métodos , Adulto , Autoenxertos , Proliferação de Células , Separação Celular , Técnicas de Cocultura , Feminino , Humanos , Integrina alfa6/metabolismo , Interleucina-8/metabolismo , SoroRESUMO
BACKGROUND: In low and middle income countries, human immunodeficiency virus (HIV) exposed, uninfected (HEU) infants demonstrate higher morbidity and mortality than their unexposed counterparts. To determine possible immune correlates of this effect, we investigated the impact of in utero HIV exposure on the uninfected neonatal immune milieu and maternal factors mediating these abnormalities in a cohort of vaginally delivered mother-infants. Samples of delivery and cord blood plasma were selected from 22 Kenyan HIV-infected women and their HIV exposed uninfected (HEU) infants drawn from the pre-ARV era, while 19 Kenyan HIV-uninfected (HU) women and their infants were selected from a control cohort. RESULTS: Compared to HU cord plasma, HEU cord plasma contained significantly higher levels of pro-inflammatory cytokines interleukins (IL)-6 and -8 (both p < 0.001) and significantly lower levels of CXC motif chemokine 11 (CXC11) (p < 0.001). Mediation analysis demonstrated that maternal HIV infection status was a significant determinant of infant IL-8 responses: HEU status was associated with a ninefold higher infant:mother (cord:delivery) plasma levels of IL-8 (p < 0.005), whereas maternal viral load was negatively associated with HEU IL-8 levels (p = 0.04) and not associated with HEU IL-6 levels. CONCLUSIONS: Exposure to maternal HIV infection drives an increase in prenatal IL-8 that is partially mediated by maternal cytokine levels. Differences between maternal and infant cytokine levels strongly suggest independent modulation in utero, consistent with prenatal immune activation. Elevated pro-inflammatory signals at birth may interfere with T cell responses at birth and subsequently influence immune maturation and the risk of morbidity and mortality in HEU infants.
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The rationale for developing histone deacetylase (HDAC) inhibitors (HDACi) as anticancer agents was based on their ability to induce apoptosis and cell cycle arrest in cancer cells. However, while HDACi have been remarkably effective in the treatment of hematological malignancies, clinical studies with HDACi as single agents in solid cancers have been disappointing. Recent studies have shown that, in addition to inducing apoptosis in cancer cells, class I HDACi induce IκB kinase (IKK)-dependent expression of proinflammatory chemokines, such as interleukin-8 (IL8; CXCL8), resulting in the increased proliferation of tumor cells, and limiting the effectiveness of HDACi in solid tumors. Here, we discuss the mechanisms responsible for HDACi-induced CXCL8 expression, and opportunities for combination therapies targeting HDACs and IKK in solid tumors.
Assuntos
Antineoplásicos/uso terapêutico , Inibidores de Histona Desacetilases/uso terapêutico , Quinase I-kappa B/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica , Apoptose/efeitos dos fármacos , Inibidores de Histona Desacetilases/administração & dosagem , Inibidores de Histona Desacetilases/farmacologia , Humanos , Quinase I-kappa B/metabolismo , Interleucina-8/metabolismo , Neoplasias/metabolismo , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologiaRESUMO
The role of the transcription factor NF-κB in shaping the cancer microenvironment is becoming increasingly clear. Inflammation alters the activity of enzymes that modulate NF-κB function, and causes extensive changes in genomic chromatin that ultimately drastically alter cell-specific gene expression. NF-κB regulates the expression of cytokines and adhesion factors that control interactions among adjacent cells. As such, NF-κB fine tunes tissue cellular composition, as well as tissues' interactions with the immune system. Therefore, NF-κB changes the cell response to hormones and to contact with neighboring cells. Activating NF-κB confers transcriptional and phenotypic plasticity to a cell and thereby enables profound local changes in tissue function and composition. Research suggests that the regulation of NF-κB target genes is specifically altered in cancer. Such alterations occur not only due to mutations of NF-κB regulatory proteins, but also because of changes in the activity of specific proteostatic modules and metabolic pathways. This article describes the molecular mode of NF-κB regulation with a few characteristic examples of target genes.
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Interleukin-8 (CXCL8) was originally described asa chemokine whose main function is the attraction of a polymorphonuclear inflammatory leukocyte infiltrate acting on CXCR1/2. Recently, it has been found that tumors very frequently coopt the production of this chemokine, which in this malignant context exerts different pro-tumoral functions. Reportedly, these include angiogenesis, survival signaling for cancer stem cells and attraction of myeloid cells endowed with the ability to immunosuppress and locally provide growth factors. Given the fact that in cancer patients IL-8 is mainly produced by tumor cells themselves, its serum concentration has been shown to correlate with tumor burden. Thus, IL-8 serum concentrations have been shown to be useful asa pharmacodynamic biomarker to early detect response to immunotherapy. Finally, because of the roles that IL-8 plays in favoring tumor progression, several therapeutic strategies are being developed to interfere with its functions. Such interventions hold promise, especially for therapeutic combinations in the field of cancer immunotherapy.
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Imunoterapia/métodos , Interleucina-8/metabolismo , Neoplasias , Biomarcadores Tumorais/metabolismo , Seguimentos , Humanos , Neoplasias/imunologia , Neoplasias/metabolismo , Neoplasias/terapiaRESUMO
Asthma and chronic obstructive pulmonary disease (COPD) are common chronic lung inflammatory diseases. Thrombin and interleukin (IL)-8/C-X-C chemokine ligand 8 (CXCL8) play critical roles in lung inflammation. Our previous study showed that c-Src-dependent IκB kinase (IKK)/IκBα/nuclear factor (NF)-κB and mitogen-activated protein kinase kinase kinase 1 (MEKK1)/extracellular signal-regulated kinase (ERK)/ribosomal S6 protein kinase (RSK)-dependent CAAT/enhancer-binding protein ß (C/EBPß) activation are involved in thrombin-induced IL-8/CXCL8 expression in human lung epithelial cells. In this study, we aimed to investigate the roles of p300 and C/EBPß-reliant IKKß expression in thrombin-induced IL-8/CXCL8 expression. Thrombin-induced increases in IL-8/CXCL8-luciferase activity and IL-8/CXCL8 release were inhibited by p300 small interfering (siRNA). Thrombin-caused histone H3 acetylation was attenuated by p300 siRNA. Stimulation of cells with thrombin for 12h resulted in increases in IKKß expression and phosphorylation in human lung epithelial cells. However, thrombin did not affect p65 expression. Moreover, 12h of thrombin stimulation produced increases in IKKß expression and phosphorylation, and IκBα phosphorylation, which were inhibited by C/EBPß siRNA. Finally, treatment of cells with thrombin caused increases in p300 and C/EBPß complex formation, p65 and C/EBPß complex formation, and recruitment of p300, p65, and C/EBPß to the IL-8/CXCL8 promoter. These results imply that p300-dependent histone H3 acetylation and C/EBPß-regulated IKKß expression contribute to thrombin-induced IL-8/CXCL8 expression in human lung epithelial cells. Results of this study will help clarify C/EBPß signaling pathways involved in thrombin-induced IL-8/CXCL8 expression in human lung epithelial cells.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/imunologia , Proteína p300 Associada a E1A/imunologia , Quinase I-kappa B/genética , Inflamação/imunologia , Interleucina-8/genética , Mucosa Respiratória/imunologia , Trombina/imunologia , Linhagem Celular , Células Epiteliais/citologia , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Humanos , Inflamação/genética , Pulmão/citologia , Pulmão/imunologia , Mucosa Respiratória/citologia , Mucosa Respiratória/metabolismoRESUMO
Airway inflammation plays a major role in the pathophysiology of lung inflammatory diseases such as asthma. Thrombin, a serine protease, is known to mediate central functions in thrombosis and hemostasis and also plays a critical role in lung inflammation via producing chemokine release including interleukin (IL)-8/CXCL8. Our previous studies showed that c-Src- and Rac-dependent nuclear factor (NF)-κB signaling pathways participate in thrombin-induced IL-8/CXCL8 release in human lung epithelial cells. In this study, we further investigated the role of casein kinase 2 (CK2)/mitogen stress-activated protein kinase 1 (MSK1)-dependent p65 phosphorylation in thrombin-induced NF-κB activation and IL-8/CXCL8 release. Thrombin-induced IL-8/CXCL8 release was inhibited by CK2 inhibitors (apigenin and tetrabromobenzotriazole, TBB), small interfering RNA of CK2ß (CK2ß siRNA), and MSK1 siRNA. Treatment of cells with thrombin caused increases in CK2ß phosphorylation at Ser209, which was inhibited by a protein kinase C α (PKCα) inhibitor (Ro-32-0432). Thrombin-induced MSK1 phosphorylation at Ser581 and Akt phosphorylation at Ser473 were inhibited by apigenin. Moreover, the thrombin-induced increase in IL-8/CXCL8 release was attenuated by p65 siRNA. Stimulation of cells with thrombin resulted in an increase in p65 phosphorylation at Ser276, which was inhibited by apigenin and MSK1 siRNA. Thrombin-induced κB-luciferase activity was also inhibited by apigenin and MSK1 siRNA. Taken together, these results show that thrombin activates the PKCα/CK2/MSK1 signaling pathways, which in turn initiates p65 phosphorylation and NF-κB activation, and ultimately induces IL-8/CXCL8 release in human lung epithelial cells.
Assuntos
Caseína Quinase II/metabolismo , Células Epiteliais/metabolismo , Interleucina-8/metabolismo , NF-kappa B/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Trombina/farmacologia , Animais , Apigenina/farmacologia , Caseína Quinase II/antagonistas & inibidores , Células Cultivadas , Células Epiteliais/efeitos dos fármacos , Humanos , Indóis/farmacologia , Pulmão/metabolismo , Fosforilação/efeitos dos fármacos , Pirróis/farmacologia , RNA Interferente Pequeno/farmacologia , Transdução de Sinais/efeitos dos fármacos , Trombina/antagonistas & inibidores , Fator de Transcrição RelA/metabolismo , Triazóis/farmacologiaRESUMO
The impact of complement activation and its possible relation to cytokine responses during malaria pathology was investigated in plasma samples from patients with confirmed Plasmodium falciparum malaria and in human whole-blood specimens stimulated with malaria-relevant agents ex vivo. Complement was significantly activated in the malaria cohort, compared with healthy controls, and was positively correlated with disease severity and with certain cytokines, in particular interleukin 8 (IL-8)/CXCL8. This was confirmed in ex vivo-stimulated blood specimens, in which complement inhibition significantly reduced IL-8/CXCL8 release. P. falciparum malaria is associated with systemic complement activation and complement-dependent release of inflammatory cytokines, of which IL-8/CXCL8 is particularly prominent.
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Ativação do Complemento/imunologia , Citocinas/metabolismo , Malária Falciparum/imunologia , Malária Falciparum/metabolismo , Adulto , Hemeproteínas/imunologia , Hemina/imunologia , Humanos , Malária Falciparum/epidemiologia , Malária Falciparum/fisiopatologia , Plasmodium falciparum/imunologiaRESUMO
Glioma cells release cytokines to stimulate inflammation that facilitates cell proliferation. Here, we show that Lipopolysaccharide (LPS) treatment could induce glioma cells to proliferate and this process was dependent on nucleotide receptor activation as well as interleukin-8 (IL-8/CXCL8) secretion. We observed that extracellular nucleotides controlled IL-8/CXCL8 and monocyte chemoattractant protein 1 (MCP-1/CCL2) release by U251MG and U87MG human glioma cell lines via P2X7 and P2Y6 receptor activation. The LPS-induced release of these cytokines was also modulated by purinergic receptor activation since IL-8 and MCP-1 release was decreased by the nucleotide scavenger apyrase as well as by the pharmacological P2Y6 receptor antagonists suramin and MRS2578. In agreement with these observations, the knockdown of P2Y6 expression decreased LPS-induced IL-8 release as well as the spontaneous release of IL-8 and MCP-1, suggesting an endogenous basal release of nucleotides. Moreover, high millimolar concentrations of ATP increased IL-8 and MCP-1 release by the glioma cells stimulated with suboptimal LPS concentration which were blocked by P2X7 and P2Y6 antagonists. Altogether, these data suggest that extracellular nucleotides control glioma growth via P2 receptor-dependent IL-8 and MCP-1 secretions.