RESUMEN
Monocytes/macrophages (MΦ), considered as plastic cells, can differentiate into either a pro-inflammatory (M1) subtype, also known as a classically activated subtype, or an anti-inflammatory alternatively activated subtype (M2) according to their microenvironment. Phenotypic markers of mouse polarized MΦ have been extensively studied, whereas their human counterparts remain less characterized. The main goal of this study was therefore to carefully characterize phenotypic and genomic markers of primary human MΦ generated from M-CSF-treated blood monocytes and polarized towards M1 or M2 subtype upon the action of lipopolysaccharide and interferon-γ (for M1) or interleukin (IL)-4 (for M2). Membrane expression of the markers CD80 and CD200R was found to be specific of human M1 and M2 polarized MΦ, respectively, whereas, by contrast, mannose receptor (CD206) expression did not discriminate between M1 and M2. mRNA expression analysis further identified six markers of M1 polarization (IL-12p35, CXCL10, CXCL11, CCL5, CCR7 and IDO1), five markers of M2 polarization (TGF-ß, CCL14, CCL22, SR-B1 and PPARγ) and transcription factors involved in MΦ polarization. Ability of human M-CSF-generated MΦ to polarize toward M1 or M2 subtype was also associated with enhanced secretion of TNFα, IL-1ß, IL-12p40, CXCL10 and IL-10 (for M1) or CCL22 (for M2). Moreover, the comparison of the expression of M1 markers in M-CSF- and GM-CSF-MΦ polarized towards M1 subtype has revealed similarities. In conclusion, we demonstrated that human M-CSF MΦ can polarize toward a M1 type after IFNγ/LPS stimulation. Moreover, the M1 and M2 markers of human polarized MΦ identified in the present study may be useful to better identify human MΦ subtypes, particularly at the tissue level, in order to better understand their respective roles in the development of pathologies.
Asunto(s)
Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Leucocitos Mononucleares/metabolismo , Factor Estimulante de Colonias de Macrófagos/farmacología , Macrófagos/metabolismo , Antígenos de Superficie/biosíntesis , Antígeno B7-1/biosíntesis , Biomarcadores/metabolismo , Diferenciación Celular , Células Cultivadas , Quimiocinas/biosíntesis , Citocinas/biosíntesis , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Humanos , Interferón gamma , Interleucina-4 , Lectinas Tipo C/biosíntesis , Leucocitos Mononucleares/efectos de los fármacos , Lipopolisacáridos , Factor Estimulante de Colonias de Macrófagos/metabolismo , Macrófagos/efectos de los fármacos , Receptor de Manosa , Lectinas de Unión a Manosa/biosíntesis , Receptores de Orexina , ARN Mensajero/biosíntesis , Receptores de Superficie Celular/biosíntesisRESUMEN
Co-exposure to environmental polycyclic aromatic hydrocarbons (PAHs) and interleukin (IL)-1ß induces expression of the tumor-promoting cytokine IL-6 in cancer cells. The present study was designed to determine whether such an IL-6 up-regulation also occurs in response to co-treatment by PAHs and tumor necrosis factor (TNF)-α, an inflammatory cytokine commonly found in tumor microenvironment. Co-exposure to the prototypical PAH benzanthracene (BZA) and TNF-α was found to markedly induce mRNA expression and secretion of IL-6 in human breast cancer cells MCF-7, whereas exposure to either BZA or TNF-α alone was without significant effect. Co-treatment by BZA and TNF-α-containing conditioned media from human inflammatory macrophages similarly up-regulated IL-6 expression in MCF-7 cells. BZA/TNF-α-mediated IL-6 induction in MCF-7 cells was counteracted by silencing aryl hydrocarbon receptor (AhR), known to mediates most of PAH effects. IL-6 up-regulation was moreover associated with NF-κB activation and was abolished by using chemical NF-κB inhibitors or knocking-down expression of the p65/RelA NF-κB subunit. Taken together, these data indicate that co-exposure to BZA/TNF-α induces IL-6 expression by AhR- and NF-κB-dependent pathways in MCF-7 cancer cells. This regulation of IL-6 by environmental PAHs, that is dependent of inflammatory cytokine microenvironment, may contribute to the well-known carcinogenic properties of these organic pollutants.
Asunto(s)
Benzo(a)Antracenos/toxicidad , Contaminantes Ambientales/toxicidad , Interleucina-6/metabolismo , FN-kappa B/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Activación Transcripcional/efectos de los fármacos , Factor de Necrosis Tumoral alfa/farmacología , Proteínas Adaptadoras Transductoras de Señales/genética , Humanos , Células MCF-7 , Macrófagos/citología , Fosfoproteínas/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Crecimiento Transformador beta/genética , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Aryl hydrocarbon receptor-dependent genomic effects of environmental polycyclic aromatic hydrocarbons (PAHs) have been shown to be modulated by non-genomic protein kinase C (PKC)-related pathways. The present study was designed to determine whether PKC activation may also impair estrogenic genomic response triggered by PAHs. Treatment by the PKC activator phorbol 12-myristate 13-acetate (PMA) was found to markedly and differentially impair the up-regulation of estrogenic markers triggered by the estrogenic PAH benzanthracene (BZA) in cultured human mammary cells; BZA-mediated mRNA up-regulation of pS2 and amphiregulin was thus increased, whereas that of progesterone receptor and CXCL12 was repressed. BZA/PMA cotreatment however failed to alter BZA-mediated increase of activity of a luciferase gene reporter construct driven by an estrogen response element, thus discarding any global effect of PMA toward BZA-triggered estrogen receptor activation. Various chemicals inhibiting PKCs or extracellular signal-regulated kinase (ERK) as well as the knock-down of PKCδ expression counteracted the PMA-mediated increase of pS2 mRNA up-regulation triggered by BZA, demonstrating that it was dependent on PKCs, including PKCδ isoform, and ERKs. This non-genomic modulation of estrogenic effects of PAHs by PKC activation may have to be considered when considering the deleterious effects of these environmental contaminants towards the endocrine system.
Asunto(s)
Contaminantes Ambientales/toxicidad , Estrógenos/toxicidad , Hidrocarburos Policíclicos Aromáticos/toxicidad , Proteína Quinasa C-delta/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Anfirregulina , Hidrocarburo de Aril Hidroxilasas/genética , Línea Celular , Quimiocina CXCL12/genética , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1B1 , Familia de Proteínas EGF , Glicoproteínas/genética , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Interleucina-6/genética , Proteína Quinasa C-delta/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genética , Receptores de Progesterona/genética , Factor Trefoil-1 , Proteínas Supresoras de Tumor/genéticaRESUMEN
Formaldehyde (FA) is a volatile organic compound (VOC), considered as a major indoor air pollutant and suspected to favor the development of inflammatory lung diseases. The present study was aimed at identifying cytokines/chemokines targeted by FA in human lung cells. This VOC was demonstrated to up-regulate interleukin-11 (IL-11) mRNA and secretion levels in a dose-dependent manner in cultured lung epithelial BEAS-2B cells. It concomitantly induced mRNA expression of transforming growth factor (TGF)-ß1, a fibrogenic marker regulated by IL-11. FA was also found to trigger an early phosphorylation of p38 and extracellular signal-regulated kinase (ERK)1/2 mitogen-activated protein kinases (MAPKs) in BEAS-2B cells, whose inhibition by ERK and p38 MAPK chemical inhibitors (U0126 and SB203580, respectively) counteracted FA-mediated induction of IL-11. In addition, FA increased phosphorylation of cAMP response element binding protein (CREB) and the use of small-interfering RNA targeting CREB demonstrated that this transcription factor was required for the up-regulation of IL-11 by FA. Implication of protein kinase C (PKC) in FA-induced IL-11 expression was moreover demonstrated by using RO-31-8220, a PKC inhibitor. We finally showed using SB203580 and RO-31-8220 that phosphorylation of CREB and CREB-promoter activity induced by FA are under the control of both p38 MAPK and PKC. Taken together, the results showed that FA uses different pathways to induce IL-11 expression in lung BEAS-2B cells. IL-11, well-known to contribute to lung inflammatory diseases, appears thus as a molecular target of FA, which could be involved in putative deleterious inflammatory and fibrogenic pulmonary effects of this VOC.