Immunoglobulins and Serotonin modulate human macrophage polarization

1 p. Annual Scientific Meeting of the European Society for Clinical Investigation Cluj-Napoca, Romania 27– 30 May 2015Peer reviewe

Polysialylated neuropilin-2 enhances human dendritic cell migration through the basic C-terminal region of CCL21.

Free Access at: http://glycob.oxfordjournals.org/cgi/pmidlookup?view=long&pmid=20488940Dendritic cell (DC) migration to secondary lymphoid organs is a critical step to properly exert its role in immunity; and predominantly depends on the interaction of the chemokine receptor CCR7 with its ligands CCL21 and CCL19. Polysialic acid (PSA) has been recently reported to control CCL21-directed migration of mature DCs. Here; we first demonstrate that PSA present on human mature monocyte-derived dendritic cells did not enhance chemotactic responses to CCL19. We have also explored the molecular mechanisms underlying the selective enhancing effect of PSA on CCL21-driven chemotaxis of DCs. In this regard; we found out that prevention of DC polysialylation decreased CCL21 activation of JNK and Akt signaling pathways; both associated with CCR7-mediated chemotaxis. We also report that the enhanced PSA-mediated effect on DC migration towards CCL21 relied on the highly basic C-terminal region of this chemokine; and depended on the PSA acceptor molecule neuropilin-2 (NRP2) and on the polysialyltransferase ST8SiaIV. Altogether; our data indicate that the CCR7/CCL21/NRP2/ST8SiaIV functional axis constitutes an important guidance clue for DC targeting to lymphoid organs.This work was supported by research grant from Fondo de Investigaciones Sanitarias, Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo (FISPI0708879 to MAV).Peer reviewe

RUNX/AML and C/EBP factors regulate CD11a integrin expression in myeloid cells through overlapping regulatory elements

10 Figures. The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked ‘‘advertisement’’ in accordance with 18 U.S.C. section 1734.The CD11a/CD18 (leukocyte function-associated antigen 1 [LFA-1]) integrin mediates critical leukocyte adhesive interactions during immune and inflammatory responses. The CD11a promoter directs CD11a/CD18 integrin expression, and its activity in lymphoid cells depends on a functional RUNX1/AML-1–binding site (AML-110) within the MS7 sequence. We now report that MS7 contains a C/EBP-binding site (C/EBP-100), which overlaps with AML-110 and is bound by C/EBP factors in myeloid cells. C/EBP and RUNX/AML factors compete for binding to their respective cognate elements and bind to the CD11a promoter MS7 sequence in a cell lineage- and differentiation-dependent manner. In myeloid cells MS7 is primarily recognized by C/EBP factors in proliferating cells whereas RUNX/AML factors (especially RUNX3/AML-2) bind to MS7 in differentiated cells. RUNX3/AML-2 binding to the CD11a promoter correlates with increased RUNX3/AML-2 protein levels and enhanced CD11a/CD18 cell surface expression. The relevance of the AML-110 element is underscored by the ability of AML-1/ETO to inhibit CD11a promoter activity, thus explaining the low CD11a/CD18 expression in t(8;21)–containing myeloid leukemia cells. Therefore, the expression of the CD11a/CD18 integrin in myeloid cells is determined through the differential occupancy of the CD11a proximal promoter by transcription factors implicated in the pathogenesis of myeloid leukemia.From the Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain; Clínica Universitaria, Universidad de Navarra, Spain; Institute of Human Genetics, Aarhus, Denmark; Hospital Universitario Gregorio Maranón, Madrid, Spain; University of Colorado Health Sciences Center, Denver; and Department of Molecular Genetics, The Weizmann Institute of Science, Rehovot Israel. Supported by grants 08.3/0026/2000.1 from Comunidad Auto´noma de Madrid, 01/0063-01 from Fondo de Investigaciones Sanitarias, and SAF2002-04615- C02-01 from Ministerio de Ciencia y Tecnología (A.L.C.). We gratefully acknowledge Drs Ana Aranda and Aurora Sánchez-Pacheco for their very generous help with ChIP assays.Peer reviewe

Rosiglitazone-induced CD36 up-regulation resolves inflammation by PPARγ and 5-LO-dependent pathways.

PPARγ-achieved neuroprotection in experimental stroke has been explained by the inhibition of inflammatory genes, an action in which 5-LO, Alox5, is involved. In addition, PPARγ is known to promote the expression of CD36, a scavenger receptor that binds lipoproteins and mediates bacterial recognition and also phagocytosis. As phagocytic clearance of neutrophils is a requisite for resolution of the inflammatory response, PPARγ-induced CD36 expression might help to limit inflammatory tissue injury in stroke, an effect in which 5-LO might also be involved. Homogenates, sections, and cellular suspensions were prepared from brains of WT and Alox5(-/-) mice exposed to distal pMCAO. BMMs were obtained from Lys-M Cre(+) PPARγ(f/f) and Lys-M Cre(-) PPARγ(f/f) mice. Stereological counting of double-immunofluorescence-labeled brain sections and FACS analysis of cell suspensions was performed. In vivo and in vitro phagocytosis of neutrophils by microglia/macrophages was analyzed. PPARγ activation with RSG induced CD36 expression in resident microglia. This process was mediated by the 5-LO gene, which is induced in neurons by PPARγ activation and at least by one of its products--LXA4--which induced CD36 independently of PPARγ. Moreover, CD36 expression helped resolution of inflammation through phagocytosis, concomitantly to neuroprotection. Based on these findings, in addition to a direct modulation by PPARγ, we propose in brain a paracrine model by which products generated by neuronal 5-LO, such as LXA4, increase the microglial expression of CD36 and promote tissue repair in pathologies with an inflammatory component, such as stroke.This work was supported by grants from the Spanish Ministry of Economy and Competitiveness CSD2010-00045 (to M.A.M.) SAF2009-08145 and SAF2012-33216 (to M.A.M.), SAF2011-23354 (toI.L.), SAF2009-07466 and SAF2012-31483 (to M.R.), from Fondo Europeo de Desarrollo Regional (FEDER) “Instituto de Salud Carlos III” RETICS RD12/0014/0003 (to I.L. and from the local govern-ment of Madrid S2010/BMD-2336 (to M.A.M.) and S2010/BMD-2349 (to I.L.). I.B. and M.I.C. are fellows of the Spanish Ministry ofEconomy and Competitiveness. The authors thank Tamara Atanesand Roberto Cañadas for their technical assistance.S

TLR7 activation in M-CSF-dependent monocyte-derived human macrophages potentiates inflammatory responses and prompts neutrophil recruitment

1 p.-4 fig.Toll-like receptor 7 (TLR7) is an endosomal Pathogen-Associated Molecular Pattern (PAMP) receptor that senses single-stranded RNA (ssRNA) and whose engagement results in the production of type I IFN and pro-inflammatory cytokines upon viral exposure. Recent genetic studies have established that a dysfunctional TLR7-initiated signaling is directly linked to the development of SARS-CoV-2-induced severe COVID-19. We previously showed that TLR7 is preferentially expressed by macrophages generated in the presence of M-CSF (M-MØ), whose MAFB-dependent transcriptome resembles pathogenic pulmonary monocyte-derived macrophage subsets in severe COVID-19. We now report that TLR7 activation in M-MØ triggers a weak MAPK, NFkB and STAT1 activation and leads to defective production of type I IFN. Nonetheless, TLR7 engagement re-programs MAFB+ M-MØ towards a distinctive transcriptional profile. Specifically, TLR7-activated M-MØ acquired the expression of genes that characterize inflammatory macrophage subsets in COVID-19 and other inflammatory diseases, including genes encoding neutrophil-attracting chemokines (CXCL1-3, CXCL5, CXCL8) reported as biomarkers for severe COVID-19. Functionally, TLR7-activated M-MØ displayed enhanced proinflammatory responses towards secondary stimulation and a robust production of neutrophil-attracting chemokines (CXCL1, CXCL5, CXCL8), which was dependent on the transcription factors MAFB and AhR. Interestingly, CXCL1 and CXCL5 release from M-MØ was also promoted by SARS-CoV-2 but not by Virus-like particles. As defective TLR7 signaling and enhanced pulmonary neutrophil/lymphocyte ratio associate with severe COVID-19, these results suggest that targeting macrophage TLR7 might be a therapeutic strategy for viral infections where monocyte-derived macrophages exhibit a pathogenic role.This research work was also funded by the European Commission – NextGenerationEU (Regulation EU 2020/2094), through CSIC's Global Health Platform (PTI Salud Global)Peer reviewe

Impacto de la educación en la cultura del emprendimiento: estudio cualitativo

Entrepreneurship is a reality that is optimistic about the possibilities of development, progress and well-being. The economic policies of the most advanced countries encourage and promote actions for their citizens to implement their business ideas and generate their own jobs. In this research we will focus on the educational aspect that has traditionally been positively linked to formal entrepreneurship. The figure of the entrepreneur is key when generating social transformations and it is worth approaching not only his personal vision but also the perception that these promoters have different age groups.In this sense, the University of Burgos through the Interuniversity Program of Experience and in the framework of training for older people, launches concrete actions to meet the needs, experiences and expectations of university majors in relation to entrepreneurship. We consider it of great importance that the University, as a long-standing institution in the field of knowledge, supports the processes of entrepreneurship with all the resources at its disposal, among them, the great human capital that is not only among its faculty or staff, but also in their classrooms, mainly those of the elderly.We show in the present investigation the initial results of the qualitative study implemented through a discussion group in which the elders have discussed the concept of entrepreneurship, motives, aid, training and skills. The results will serve as a basis for future proposals and interventions.El emprendimiento es una realidad que se nos presenta optimista en cuanto a posibilidades de desarrollo, progreso y bienestar. Las políticas económicas de los países más avanzados animan y promueven acciones para que sus ciudadanos pongan en marcha sus ideas de negocio y generen sus propios puestos de trabajo. En la presente investigación nos centraremos en el aspecto educativo que tradicionalmente se ha vinculado positivamente con el emprendimiento formal. La figura del emprendedor es clave a la hora de generar transformaciones de tipo social y merece la pena acercarse no solo a su visión personal sino también a la percepción que de estos promotores tienen los diferentes grupos de edad.En este sentido, la Universidad de Burgos a través del Programa Interuniversitario de la Experiencia y en el marco de la formación para personas mayores, pone en marcha acciones concretas para conocer las necesidades, experiencias y expectativas de los mayores universitarios en relación con el emprendimiento. Consideramos de gran importancia que la Universidad, como institución de larga trayectoria en el campo del saber, apoye los procesos de emprendimiento con todos los recursos a su alcance, entre ellos, el gran capital humano que no solo se encuentra entre su profesorado o personal, sino también en sus aulas, principalmente en las de las personas mayores.Mostramos en la presente investigación los resultados iniciales del estudio cualitativo implementado a través de un grupo de discusión en el que los mayores han dialogado sobre el concepto de emprendimiento, motivos, ayudas, formación y competencias. Los resultados nos servirán de base para futuras propuestas e intervenciones

BRAIN & SPINAL CORD DAMAGE & REHABILITATION

Stroke and traumatic injury in brain or spinal cord are often life-threating conditions and major causes of death or permanent disability with high impact in the health care system. There are several stages of intervention to improve the neurological outcome. Acutely, fast interventions aiming to reestablish cerebral blood flow in ischemic stroke, to stop bleeding after brain hemorrhage, and to reduce edema after contusions are amongst mandatory actions. Current studies aim to develop accompanying strategies for brain cell protection based on enhancing endogenous protective mechanism, blocking cell death pathways, or through immunomodulation. After the acute phase, interventions are intended to promote recovery of function using rehabilitation with state-of-the-art technologies enabled by robotics. Other advanced strategies include cell, gene, and immune therapies, and brain function modulation with the aid of smart nanotechnologies. There is great expectation in the fast evolving novel approaches for improvement of neurological deficits in these unpredictable and devastating conditionPeer reviewe

AKT activation seems to be associated with apoptotic signals and not with pro-survival signals in a pristane-induced lupus model.

Several studies have shown that in addition to its role as a survival factor and tumor promoting agent, AKT is also able to exhibit pro-apoptotic effects under diverse conditions, including oxidative stress, cytokine stimulation and exposure to cytotoxic chemicals like staurosporine, methotrexate, docetaxel and etoposide. Moreover, phosphorylation of second mitochondria-derived activator of caspases (SMAC) by AKT promotes caspase-3 activation during etoposide-induced apoptosis in HeLa cells. Our data show that injection of pristane into the peritoneum induces apoptosis-mediated cell death of peritoneal exudate cells (PECs), as evidenced by the increased number of annexin V+ peritoneal cells and their increased levels of cleaved/active caspase-3. Indeed, the higher levels of activated caspase-3 protein in WT PECs, particularly at 2-weeks post pristane treatment, are indicative of a higher rate of apoptosis compared to Cd38¿/¿ cells. In contrast, no differences were observed in the levels of MCL-1, an anti-apoptotic protein and member of the BCL2 family. Furthermore, kinases ERK1/2 and AKT showed distinct activation kinetics in pristane-elicited PECs. Interestingly, caspase-3 activation followed similar kinetics to AKT activation in both WT and Cd38¿/¿ PECs, while ERK activation correlated with increased levels of MCL-1. In summary our data strongly suggest that in the pristane-induced lupus model AKT activation is associated with apoptotic signals and not with survival signals. Further studies, however, are required to identify specific pro- and anti-apoptotic target proteins that are phosphorylated by ERK or AKT following pristane treatment, and that regulate the apoptotic process

Macrophages from the synovium of active rheumatoid arthritis exhibit an activin A-dependent pro-inflammatory profile

12 p.-7 fig.Rheumatoid arthritis (RA) is a chronic inflammatory disease whose pathogenesis and severity correlates with the presence of macrophage-derived pro-inflammatory cytokines within the inflamed synovium. Macrophage-derived cytokines fuel the pathological processes in RA and are targets of clinically successful therapies. However, although macrophage polarization determines cytokine production, the polarization state of macrophages in RA joints remains poorly defined. To dissect the molecular basis for the tissue-damaging effects of macrophages in RA joints, we undertook the phenotypic and transcriptomic characterization of ex vivo isolated CD14(+) RA synovial fluid (RA-SF) macrophages. Flow cytometry and gene profiling indicated that RA-SF macrophages express pro-inflammatory polarization markers (MMP12, EGLN3, CCR2), lack expression of markers associated with homeostatic and anti-inflammatory polarization (IGF1, HTR2B) and exhibit a transcriptomic profile that resembles the activin A-dependent gene signature of pro-inflammatory in vitro-generated macrophages. In fact, high levels of Smad-activating activin A were found in RA-SF and, accordingly, the Smad signalling pathway was activated in ex vivo-isolated RA-SF macrophages. In vitro experiments on monocytes and macrophages indicated that RA-SF promoted the acquisition of pro-inflammatory markers (INHBA, MMP12, EGLN3, CCR2) but led to a significant reduction in the expression of genes associated with homeostasis and inflammation resolution (FOLR2, SERPINB2, IGF1, CD36), thus confirming the pro-inflammatory polarization ability of RA-SF. Importantly, the macrophage-polarizing ability of RA-SF was inhibited by an anti-activin A-neutralizing antibody, thus demonstrating that activin A mediates the pro-inflammatory macrophage-polarizing ability of RA-SF. Moreover, and in line with these findings, multicolour immunofluorescence evidenced that macrophages within RA synovial membranes (RA-SM) also express pro-inflammatory polarization markers whose expression is activin A-dependent. Altogether, our results demonstrate that macrophages from RA synovial fluids and membranes exhibit an MMP12(+) EGLN3(+) CCR2(+) pro-inflammatory polarization state whose acquisition is partly dependent on activin A from the synovial fluid.This study was supported by Instituto de Salud Carlos III (Grant Nos PI11/00165, to APK, andPI13/01454, to PSM); Comunidad de Madrid/FEDER(RAPHYME Programme; Grant No. S2010/BMD2350,to JLP, ALC and APK); Ministerio de Ciencia e Innovación (Grant Nos RIER RD12/009, to IGA, PSM, JLP,ALC and APK, and SAF2011-23801, to ALC); FIB-HGM (to APK); and Ministerio de Economia y Competitividad (Juan de la Cierva Contract No. JCI-2011-09836and a Miguel Servet contract, to EI and GC).Peer reviewe