RESUMEN
Peritoneal macrophages (PMs), which resided in peritoneal cavity, are crucial to maintain tissue homeostasis and immunity. Macrophage self-renewal and polarization states are critical for PM population homeostasis and function. However, the underlying molecular mechanism that regulates self-renewal and polarization of PMs is still unclear and needs to be explored. Here, we demonstrated that PMs self-renewal was stimulated by granulocyte macrophage colony-stimulating factor (GM-CSF), but not by macrophage colony-stimulating factor (M-CSF). Pharmacological inhibition of Bromodomain & Extraterminal (BET) Proteins by either JQ1 or ARV-825 significantly reduced GM-CSF-dependent peritoneal macrophage self-renewal by abrogating cell proliferation and decreasing self-renewal-related gene expression, such as MYC and Klf4, at transcriptional and protein levels. In addition, transcriptomic analysis showed that JQ1 blocked alternative PMs polarization by downregulating key transcriptional factor IRF4 expression, but not the activation of AKT or STAT6 in PMs. These findings illustrated that the significance of BET family proteins in GM-CSF-induced PMs self-renewal and IL-4-induced alternative polarization.
RESUMEN
Pure plant extract luteolin has been demonstrated to possess numerous biological and immunological effects. However, how luteolin affects mice alveolar macrophages' self-renewal and polarization closely related to inflammatory and immunomodulatory is still unknown. In our study, the transcriptomic analysis showed that several self-renewal-related pathways in luteolin-pretreated alveolar macrophages were inhibited compared to the granulocyte-macrophage colony-stimulating factor (GM-CSF)-treated group. Ki-67 staining and EdU assay indicated that luteolin inhibited GM-CSF-induced alveolar macrophage proliferation. Moreover, GM-CSF-induced expressions of c-Myc and KLF4 were significantly suppressed by luteolin at transcriptional and protein levels. Besides, we found that luteolin promoted M1 macrophage polarization induced by LPS plus IFN-γ. At the same time, it inhibited M2 macrophage polarization induced by IL-4 in both alveolar and bone marrow-derived macrophages by detecting macrophage polarization-related gene expressions at mRNA and protein levels. We found that luteolin inhibited self-renewal and altered the polarization of primary alveolar macrophages. Taken together, our data will aid in a better understanding of the immunomodulatory effects of luteolin on the primary alveolar macrophages.
Asunto(s)
Factor Estimulante de Colonias de Granulocitos y Macrófagos , Macrófagos Alveolares , Animales , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Luteolina/metabolismo , Luteolina/farmacología , Macrófagos/metabolismo , RatonesRESUMEN
Cellular energy metabolism plays a crucial role in the regulation of macrophage polarization and in the execution of immune functions. A recent study showed that Slc6a8-mediated creatine uptake from exogenous supplementation modulates macrophage polarization, yet little is known about the role of the de novo creatine de novobiosynthesis pathway in macrophage polarization. Here, we observed that glycine amidinotransferase (GATM), the rate-limiting enzyme for creatine synthesis, was upregulated in alternative (M2) polarized macrophages, and was dependent on the transcriptional factor STAT6, whereas GATM expression was suppressed in the classical polarized (M1) macrophage. Next, we revealed that exogenous creatine supplementation enhanced IL-4-induced M2 polarization, confirming recent work. Furthermore, we revealed that genetic ablation of GATM did not affect expression of M1 marker genes (Nos2, IL1b, IL12b) or the production of nitric oxide in both peritoneal macrophages (PMs) and bone marrow-derived macrophages (BMDMs). By contrast, expression levels of M2 markers (Arg1, Mrc1, Ccl17 and Retnla) were lower following GATM deletion. Moreover, we found that deletion of GATM in resident alveolar macrophages (AMs) significantly blocked M2 polarization but with no obvious effect on the number of cells in knockout mice. Lastly, an upregulation of GATM was found in lung tissue and bronchoalveolar lavage fluid macrophages from HDM-induced asthmatic mice. Our study uncovers a previously uncharacterized role for the de novo creatine biosynthesis enzyme GATM in M2 macrophage polarization, which may be involved in the pathogenesis of related inflammatory diseases such as an T helper 2 (Th2)-associated allergic asthma.
Asunto(s)
Asma , Creatina , Amidinotransferasas , Animales , Creatina/metabolismo , Interleucina-4/metabolismo , Macrófagos , Ratones , Ratones Noqueados , Óxido Nítrico/metabolismoRESUMEN
Pyocyanin (PYO) is a major virulence factor secreted by Pseudomonas aeruginosa, and autophagy is a crucial homeostatic mechanism for the interaction between the pathogens and the host. It remains unknown whether PYO leads to autophagy in macrophages by regulating histone acetylation. The high mobility group nucleosomal binding domain 2 (HMGN2) has been reported to regulate the PYO-induced autophagy and oxidative stress in the epithelial cells; however, the underlying molecular mechanism has not been fully elucidated. In this study, PYO was found to induce autophagy in macrophages, and the mechanism might be correlated with the up-regulation of HMGN2 acetylation (HMGN2ac) and the down-regulation of H3K27 acetylation (H3K27ac) by modulation of the activities of acetyltransferases and deacetylases. Moreover, we further demonstrated that the up-regulated HMGN2ac enhances its recruitment to the Ulk1 promoter, while the down-regulation of H3K27ac reduces its recruitment to the Ulk1 promoter, thereby promoting or inhibiting the transcription of Ulk1. In conclusion, HMGN2ac and H3K27ac play regulatory roles in the PYO-induced autophagy in macrophages.
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Homólogo de la Proteína 1 Relacionada con la Autofagia/genética , Autofagia , Proteína HMGN2/metabolismo , Código de Histonas , Macrófagos Peritoneales/metabolismo , Acetilación , Animales , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Células Cultivadas , Humanos , Macrófagos Peritoneales/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , Regiones Promotoras Genéticas , Piocianina/farmacología , Células RAW 264.7 , Células THP-1 , Activación TranscripcionalRESUMEN
Quantum dots (QDs), are one kind of nanoscale semiconductor crystals with specific electronic and optical properties, offering near-infrared mission and chemically active surfaces. Increasing interest for QDs exists in developing theranostics platforms for bioapplications such as imaging, drug delivery and therapy. Here we summarized QDs' biomedical applications, toxicity, and immunological effects on the respiratory system. Bioapplications of QDs in lung include biomedical imaging, drug delivery, bio-sensing or diagnosis and therapy. Generically, toxic effects of nanoparticles are related to the generation of oxidative stresses with subsequent DNA damage and decreased lung cells viability in vitro and in vivo because of release of toxic metal ions or the features of QDs like its surface charge. Lastly, pulmonary immunological effects of QDs mainly include proinflammatory cytokines release and recruiting innate leukocytes or adaptive T cells.
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Puntos Cuánticos , Sistema Respiratorio , Animales , Técnicas Biosensibles , Portadores de Fármacos , Sistemas de Liberación de Medicamentos , Humanos , Inmunomodulación , Imagen Molecular , Puntos Cuánticos/química , Nanomedicina Teranóstica/métodosRESUMEN
Urinary tract infection is one of the most common bacterial infections which is mainly caused by Escherichia coli (UPEC). Autophagy plays a key role in immune response to eliminate invading pathogens. Exploring the effect of autophagy on UPEC infection and the molecular mechanisms will be benefit for the treatment of urinary tract infection. High-mobility group protein N2 (HMGN2), a highly conserved nuclear protein and an antibacterial peptide, has been associated with bacterial infection induced immune response; however, whether this function is due to the regulation of autophagy remains unclear. In this study, we demonstrate for the first time that HMGN2 is upregulated in UPEC infection of bladder epithelial cell line 5637 (BEC 5637). Furthermore, HMGN2 enhances autophagy in BEC 5637 via activation of AMPK and ULK1, whereas UPEC suppresses autophagy. In addition, the enhanced autophagy activity by HMGN2 overexpression or rapamycin boosts the proliferation of UPEC J96 in BEC 5637. In summary, our data indicate that HMGN2 activates autophagy via AMPK/ULK1 pathway which can be utilized by UPEC J96 for their proliferation within bladder epithelial cells.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Infecciones por Escherichia coli/metabolismo , Proteína HMGN2/metabolismo , Vejiga Urinaria/microbiología , Infecciones Urinarias/metabolismo , Animales , Autofagia , Línea Celular , Proliferación Celular , Células Epiteliales/citología , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Escherichia coli/crecimiento & desarrollo , Escherichia coli/metabolismo , Infecciones por Escherichia coli/microbiología , Femenino , Humanos , Ratones Endogámicos C57BL , Transducción de Señal , Vejiga Urinaria/citología , Vejiga Urinaria/metabolismo , Infecciones Urinarias/microbiologíaRESUMEN
Micro-RNAs (miRNAs) critically regulate several host defense mechanisms, but their roles in the bacteria-epithelium interplay remain unclear. Our results displayed that the expression of miR-155 and miR-23a were down-regulated in K. pneumoniae-infected pulmonary epithelial cells. The elevated bacterial adhesion on A549 cells followed the enhancement of the cellular levels of these two miRNAs. Meanwhile, a mechanistic study demonstrated that miR-155 promoted integrin α5ß1 function and resulted in the increased actin polymerization. Moreover, a non-histone nuclear protein, high mobility group nucleosomal-binding domain 2 (HMGN2) served as the potential target of miR-155 and miR-23a to regulate the integrin α5ß1 expression and K. pneumoniae adhesion. Furthermore, the expression of a known integrin transcription suppressor-Nuclear Factor-I (NFI) was also repressed by miR-155, which paralleled with its chromatin location in the promoter regions of integrin α5 and ß1. These results uncover novel links between miRNAs and integrin function to regulate bacterial adhesion, indicating a potential mechanism of host cell autonomous immune response to K. pneumoniae infection.
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Células Epiteliales Alveolares/metabolismo , Adhesión Bacteriana , Integrina alfa5beta1/metabolismo , Infecciones por Klebsiella/metabolismo , Klebsiella pneumoniae/metabolismo , MicroARNs/metabolismo , Línea Celular , Regulación hacia Abajo , Humanos , Transducción de SeñalRESUMEN
Uropathogenic Escherichia coli (UPEC), the primary uropathogen, adhere to and invade bladder epithelial cells (BECs) to establish a successful urinary tract infection (UTI). Emerging antibiotic resistance requires novel nonantibiotic strategies. Our previous study indicated that luteolin attenuated adhesive and invasive abilities as well as cytotoxicity of UPEC on T24 BECs through down-regulating UPEC virulence factors. The aims of this study were to investigate the possible function of the flavonoid luteolin and the mechanisms by which luteolin functions in UPEC-induced bladder infection. Firstly, obvious reduction of UPEC invasion but not adhesion were observed in luteolin-pretreated 5637 and T24 BECs sa well as mice bladder via colony counting. The luteolin-mediated suppression of UPEC invasion was linked to elevated levels of intracellular cAMP induced by inhibiting the activity of cAMP-phosphodiesterases (cAMP-PDEs), which resulting activation of protein kinase A, thereby negatively regulating Rac1-GTPase-mediated actin polymerization. Furthermore, p38 MAPK was primarily and ERK1/2 was partially involved in luteolin-mediated suppression of UPEC invasion and actin polymerization, as confirmed with chemical activators of p38 MAPK and ERK1/2. These data suggest that luteolin can protect bladder epithelial cells against UPEC invasion. Therefore, luteolin or luteolin-rich products as dietary supplement may be beneficial to control the UPEC-related bladder infections, and cAMP-PDEs may be a therapy target for UTIs treatment. © 2016 BioFactors, 42(6):674-685, 2016.
Asunto(s)
Antibacterianos/administración & dosificación , Luteolina/administración & dosificación , Infecciones Urinarias/prevención & control , Escherichia coli Uropatógena/efectos de los fármacos , Actinas/metabolismo , Administración Oral , Animales , Adhesión Bacteriana/efectos de los fármacos , Suplementos Dietéticos , Activación Enzimática , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Ratones Endogámicos C57BL , Pruebas de Sensibilidad Microbiana , Neuropéptidos/metabolismo , Multimerización de Proteína , Infecciones Urinarias/microbiología , Escherichia coli Uropatógena/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Proteína de Unión al GTP rac1/metabolismoRESUMEN
Colorectal cancer, a type of malignant neoplasm originating from the epithelial cells lining the colon and/or rectum, has been the third most frequent malignancy and one of the leading causes of cancer-related deaths in the US. As a bioflavonoid with high anticancer potential, quercetin (Qu) has been proved to have a prospective applicability in chemotherapy for a series of cancers. However, quercetin is a hydrophobic drug, the poor hydrophilicity of which hinders its clinical usage in cancer therapy. Therefore, a strategy to improve the solubility of quercetin in water and/or enhance the bioavailability is desired. Encapsulating the poorly water-soluble, hydrophobic agents into polymer micelles could facilitate the dissolution of drugs in water. In our study, nanotechnology was employed, and quercetin was encapsulated into the biodegradable nanosized amphiphilic block copolymers of monomethoxy poly(ethylene glycol)-poly(ε-caprolactone) (MPEG-PCL), attempting to present positive evidences that this drug delivery system of polymeric micelles is effective. The quercetin-loaded MPEG-PCL nanomicelles (Qu-M), with a high drug loading of 6.85% and a minor particle size of 34.8 nm, completely dispersed in the water and released quercetin in a prolonged period in vitro and in vivo. At the same time, compared with free quercetin, Qu-M exhibited improved apoptosis induction and cell growth inhibition effects in CT26 cells in vitro. Moreover, the mice subcutaneous CT26 colon cancer model was established to evaluate the therapy efficiency of Qu-M in detail, in which enhanced anti-colon cancer effect was proved in vivo: Qu-M were more efficacious in repressing the growth of colon tumor than free quercetin. In addition, better effects of Qu-M on inducing cell apoptosis, inhibiting tumor angiogenesis, and restraining cell proliferation were observed by immunofluorescence analysis. Our study indicated that Qu-M were a novel nanoagent of quercetin with an enhanced antitumor activity, which could serve as a promising potential candidate for colon cancer chemotherapy.
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Antineoplásicos Fitogénicos/farmacología , Neoplasias del Colon/tratamiento farmacológico , Portadores de Fármacos/química , Quercetina/química , Quercetina/farmacología , Animales , Antineoplásicos Fitogénicos/administración & dosificación , Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/farmacocinética , Apoptosis/efectos de los fármacos , Materiales Biocompatibles/química , Línea Celular Tumoral , Neoplasias del Colon/patología , Portadores de Fármacos/farmacología , Sistemas de Liberación de Medicamentos/métodos , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Micelas , Nanopartículas/química , Neovascularización Patológica/tratamiento farmacológico , Tamaño de la Partícula , Poliésteres/química , Polietilenglicoles/química , Quercetina/administración & dosificación , Quercetina/farmacocinética , Ratas Sprague-Dawley , Solubilidad , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Since bacterial invasion into host cells is a critical step in the infection process and the predominance of multiple-antibiotic-resistant Klebsiella (K.) pneumoniae strains, using molecular agents to interfere with K. pneumoniae invasion is an attractive approach for the prevention of infection and suppress the immune inflammatory response. In previous studies by our group, high-mobility group nucleosome-binding domain 2 (HMGN2) protein was shown to exhibit anti-bacterial activity in vitro. The objective of the present study was to investigate the effects of HMGN2 protein on the invasion of K. pneumoniae 03183 in vivo. The results showed that pre-treatment with 128 µg/ml HMGN2 significantly reduced K. pneumoniae 03183 invasion into mouse lungs and increased the mRNA expression of CXCL1 and LCN2 within 2 h. Immunohistochemical staining showed that F-actin expression was significantly decreased, and fluorescence microscopy and western blot analysis further demonstrated that HMGN2 significantly blocked K. pneumoniae 03183-induced actin polymerization. These changes implied that HMGN2 may provide protection against K. pneumoniae 03183 infection in vivo.
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Antibacterianos/farmacología , Proteína HMGN2/farmacología , Infecciones por Klebsiella/tratamiento farmacológico , Klebsiella pneumoniae/efectos de los fármacos , Pulmón/efectos de los fármacos , Neumonía Bacteriana/tratamiento farmacológico , Actinas/genética , Actinas/inmunología , Proteínas de Fase Aguda/agonistas , Proteínas de Fase Aguda/genética , Proteínas de Fase Aguda/inmunología , Animales , Antibacterianos/biosíntesis , Quimiocina CXCL1/agonistas , Quimiocina CXCL1/genética , Quimiocina CXCL1/inmunología , Femenino , Expresión Génica , Proteína HMGN2/biosíntesis , Interacciones Huésped-Patógeno/efectos de los fármacos , Humanos , Infecciones por Klebsiella/inmunología , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/patología , Klebsiella pneumoniae/fisiología , Lipocalina 2 , Lipocalinas/agonistas , Lipocalinas/genética , Lipocalinas/inmunología , Pulmón/microbiología , Ratones , Ratones Endogámicos C57BL , Proteínas Oncogénicas/agonistas , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/inmunología , Neumonía Bacteriana/inmunología , Neumonía Bacteriana/microbiología , Neumonía Bacteriana/patología , ARN Mensajero/genética , ARN Mensajero/inmunología , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/farmacologíaRESUMEN
Urinary tract infection (UTI), primarily caused by uropathogenic Escherichia coli (UPEC), is one of the most common infectious diseases worldwide. Emerging antibiotic resistance requires novel treatment strategies. Luteolin, a dietary polyphenolic flavonoid, has been confirmed as a potential antimicrobial agent. Here, we evaluated the sub-MICs of luteolin for potential properties to modulate the UPEC infection. We found that luteolin significantly decreased the attachment and invasion of UPEC J96 or CFT073 in human bladder epithelial cell lines T24. Meanwhile, obvious decreased expression of type 1 fimbriae adhesin fimH gene, lower bacterial surface hydrophobicity and swimming motility, were observed in luteolin-pretreated UPEC. Furthermore, luteolin could attenuate UPEC-induced cytotoxicity in T24 cells, which manifested as decreased activity of lactate dehydrogenase (LDH). Simultaneously, the inhibition of luteolin on UPEC-induced cytotoxicity was confirmed by ethidium bromide/acridine orange staining. Finally, the luteolin-pretreated UPEC showed a lower ability of biofilm formation. Collectively, these results indicated that luteolin decreased the attachment and invasion of UPEC in bladder epithelial cells, attenuated UPEC-induced cytotoxicity and biofilm formation via down-regulating the expression of adhesin fimH gene, reducing the bacterial surface hydrophobicity and motility.