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The expression and function of TRPV1 are influenced by its interaction with cellular proteins. Here, we identify Whirlin, a cytoskeletal PDZ-scaffold protein implicated in hearing, vision and mechanosensory transduction, as an interacting partner of TRPV1. Whirlin associates with TRPV1 in cell lines and in primary cultures of rat nociceptors. Whirlin is expressed in 55% of mouse sensory C-fibers, including peptidergic and non-peptidergic nociceptors, and co-localizes with TRPV1 in 70% of them. Heterologous expression of Whirlin increased TRPV1 protein expression and trafficking to the plasma membrane, and promoted receptor clustering. Silencing Whirlin expression with siRNA or blocking protein translation resulted in a concomitant degradation of TRPV1 that could be prevented by inhibiting the proteasome. The degradation kinetics of TRPV1 upon arresting protein translation mirrored that of Whirlin in cells co-expressing both proteins, suggesting a parallel degradation mechanism. Noteworthy, Whirlin expression significantly reduced TRPV1 degradation induced by prolonged exposure to capsaicin. Thus, our findings indicate that Whirlin and TRPV1 are associated in a subset of nociceptors and that TRPV1 protein stability is increased through the interaction with the cytoskeletal scaffold protein. Our results suggest that the Whirlin-TRPV1 complex may represent a novel molecular target and its pharmacological disruption might be a therapeutic strategy for the treatment of peripheral TRPV1-mediated disorders. (C) 2015 Elsevier B.V. All rights reserved.

Resumen: Introducción: La Organización Mundial de la Salud recomienda la educación interprofesional cuando estudiantes de 2 o más profesiones sanitarias aprenden juntos, para favorecer el desarrollo de las competencias interprofesionales que les van a permitir trabajar en equipo proporcionando la mejor asistencia sanitaria. En la Universidad Europea de Madrid existe un programa de educación interprofesional que trabaja las competencias de clarificación de roles, comunicación y autoridad, con estudiantes de diferentes titulaciones. Una de sus actividades es la de atención domiciliaria, en la que participan estudiantes de último curso de Medicina y de Enfermería. El objetivo de este estudio es analizar la percepción de los estudiantes que han participado en la actividad de atención domiciliaria durante el curso 2016-2017 en relación con las 3 competencias interprofesionales. Material y métodos: Estudio cualitativo descriptivo, de corte fenomenológico. Análisis de los fragmentos de texto de las reflexiones de los estudiantes acerca de la actividad de atención domiciliaria mediante el método de comparaciones constantes (codificación abierta y codificación axial). Resultados: Los estudiantes refieren que la actividad les ha ayudado a conocer mejor el rol y las tareas de los médicos y los enfermeros, de una forma diferente a las prácticas clínicas. En cuanto a la comunicación, la basan en el respeto y la confianza. Por último, en cuando a la autoridad, se decantan mayoritariamente por modelos democráticos y horizontales de toma de decisiones. Discusión: Los estudiantes perciben que la actividad resulta útil para el desarrollo de las 3 competencias interprofesionales, lo cual coincide con la literatura científica. Abstract: Introduction: The World Health Organization recommends implementing interprofessional education programs, where students from 2 or more health professions learn together, to improve interprofessional competency development that will help them to improve collaboration and the quality of care and services. At the Universidad Europea from Madrid there is a program on interprofessional education that develops the following competences: role, communication and authority, with students from several degrees. Home care is one of those activities, undertaken by students from the last years of medicine and nursing. The purpose of this research is to analyze the perception of the students about the interprofessional competences after taken the home care activity during the 2016-17 academic year. Material and methods: Phenomenological qualitative design. Student's written comments were analyzed using the constant comparative method (open and axial coding). Results: They got to know better the roles played by each other (doctors and nurses) thanks to the fact of working together. In relation to the way they communicate, they mentioned that adequate communication is based in respect and trust to each other. In relation to the different authority models, most of them tended to use and feel better in more horizontal and democratic ways of decision making. Discussion: Students reported that the activity is useful for improving their interprofessional competencies. It is coincident with other studies. Palabras clave: Educación interprofesional, Atención domiciliaria, Trabajo en equipo, Práctica colaborativa, Atención primaria, Competencias, Keywords: Interprofessional education, Home care, Team work, Collaborative practice, Primary health care, Competencies

In a previous research study, a protein from Bacillus subtilis was cloned in an overexpressing strain and produced in higher amounts than in natural conditions, with the aim of assessing whether this protein can be a candidate to be considered in the formulation of new biobased, bioinspired adhesives with industrial purposes. A bioadhesion assay, as well as the morphological analysis by scanning electron microscopy and confocal laser scanning microscopy, confirmed the involvement of the protein under study in the biofilm formation and adhesion processes of the bacterium. Furthermore, its suitability for being used as a polymeric base for adhesive formulations was assessed in terms of thermal properties, wettability of different substrates, surface tension, particle size distribution and bondability to polymeric adherends. The results obtained were promising. Nevertheless, it was found that the adhesive should be conveniently formulated in order to improve its performance. In this work, the purified protein has been modified using glutaraldehyde, which is commonly used in protein crosslinking. Structural changes produced in the protein, which could affect its suitable use as an adhesive, were assessed in terms of crystallinity, morphology, thermal properties and spectroscopic properties.

Understanding the regulation of T-cell responses during inflammation and auto-immunity is fundamental for designing efficient therapeutic strategies against immune diseases. In this regard, prostaglandin E2 (PGE2) is mostly considered a myeloid-derived immunosuppressive molecule. We describe for the first time that T cells secrete PGE2 during T-cell receptor stimulation. In addition, we show that autocrine PGE2 signaling through EP receptors is essential for optimal CD4(+) T-cell activation in vitro and in vivo, and for T helper 1 (Th1) and regulatory T cell differentiation. PGE2 was found to provide additive co-stimulatory signaling through AKT activation. Intravital multiphoton microscopy showed that triggering EP receptors in T cells is also essential for the stability of T cell-dendritic cell (DC) interactions and Th-cell accumulation in draining lymph nodes (LNs) during inflammation. We further demonstrated that blocking EP receptors in T cells during the initial phase of collagen-induced arthritis in mice resulted in a reduction of clinical arthritis. This could be attributable to defective T-cell activation, accompanied by a decline in activated and interferon-γ-producing CD4(+) Th1 cells in draining LNs. In conclusion, we prove that T lymphocytes secret picomolar concentrations of PGE2, which in turn provide additive co-stimulatory signaling, enabling T cells to attain a favorable activation threshold. PGE2 signaling in T cells is also required for maintaining long and stable interactions with DCs within LNs. Blockade of EP receptors in vivo impairs T-cell activation and development of T cell-mediated inflammatory responses. This may have implications in various pathophysiological settings. Sin financiación 4.557 JCR (2016) Q2, 57/190 Cell Biology, 39/151 Immunology UEM

Biofilms are aggregates of microorganisms in which cells adhere to each other and/or to a surface and are frequently embedded within a self-produced matrix of extracellular polymeric substance. This extracellular substance can be of interest in the search of new biobased adhesives. This research focuses on a protein from Bacillus subtilis. This protein has been highly produced in an overexpressing strain with the aim of assessing its involvement in biofilm formation and adhesion mechanisms. A bioadhesion assay showed that the overexpressing strain presented enhanced adherence to abiotic surfaces. Furthermore, scanning electron microscopy showed that cells in biofilms appeared to be completely encased in an extracellular matrix when the protein is overproduced. These results confirm the relationship between the expression of the selected protein and an increase in biofilm adherence to abiotic surfaces and allow considering the use of this protein as a base polymer in the formulation of bioinspired adhesives.

Functional materials with antimicrobial properties are being investigated in order to provide shoes with new properties and added value by incorporating natural antimicrobial agents to conventional materials (leather, fabrics, foams, etc). Microencapsulation is an effective method to protect these functional natural biocides from reactions with moisture, light, and oxygen. If a footwear material is treated with microencapsulated biocide agents, higher durability of functionality is expected. This article reports the evaluation of Melaleuca alternifolia (tea tree) oil which is a natural essential oil, as an antimicrobial agent for footwear application. Furthermore, microcapsules containing this essential oil were synthesized by in situ polymerization method using a melamine-formaldehyde resin as shell material in order to increase the durability of this natural biocide in footwear materials. Microcapsule characterization, such as particle size, morphology, and chemical properties was carried out for differ...

Prostanoids, including prostaglandins (PGs) and thromboxanes (TXs) are synthesized from arachidonic acid by the combined action of cyclooxygenases (COXs) and PG and TX synthases. Finally after their synthesis, prostanoids are quickly released to the extracellular medium exerting their effects upon interaction with prostanoid receptors present in the neighbouring cells. These agents exert important actions in the cardiovascular system, modulating vascular homeostasis and participating in the pathogenesis of vascular diseases as thrombosis and atherosclerosis. Among prostanoids, Tromboxane (TX)A(2), a potent platelet activator and vasoconstrictor and prostacyclin (PGI2), a platelet inhibitor and vasodilator, are the most important in controlling vascular homeostasis. Although multiple studies using pharmacological inhibitors and genetically deficient mice have demonstrated the importance of prostanoid-mediated actions on cardiovascular physiology, further analysis on the prostanoid mediated actions in the vascular system are required to better understand the benefits and risks for the use of COX inhibitors in cardiovascular diseases.

Francisella tularensis (Ft), a Gram-negative intracellular bacterium, is the etiologic agent of tularemia. Although attenuated for humans, i.p. infection of mice with 106 organisms. To examine the immunological consequences of Ft LVS infection on the innate immune response, the inflammatory responses of mice infected i.p. or intradermally were compared. Mice infected i.p. displayed greater bacterial burden and increased expression of proinflammatory genes, particularly in the liver. In contrast to most LPS, highly purified Ft LVS LPS (10 μg/ml) was found to be only minimally stimulatory in primary murine macrophages and in HEK293T cells transiently transfected with TLR4/MD-2/CD14, whereas live Ft LVS bacteria were highly stimulatory for macrophages and TLR2-expressing HEK293T cells. Despite the poor stimulatory activity of Ft LVS LPS in vitro, administration of 100 ng of Ft LVS LPS 2 days before Ft LVS challenge severely limited both bacterial burden and cytokine mRNA and protein expression in the absence of detectable Ab at the time of bacterial challenge, yet these mice developed a robust IgM Ab response within 2 days of infection and survived. These data suggest that prior administration of Ft LVS LPS protects the host by diminishing bacterial burden and blunting an otherwise overwhelming inflammatory response, while priming the adaptive immune response for development of a strong Ab response.

Adrenomedullin (AM) is a multifunctional evolutionarily highly conserved peptide. Although its genomic and amino acid (aa) sequences are known in several mammalian species and in fish, the structure of the AM gene remains unknown in intermediate phyla, including birds. Here, we report the structure and aa sequence of the chicken (c) AM ortholog. The cAM gene is located at the short arm of chromosome 5, which shows high synteny with the short arm of human (h) chromosome 11, where hAM is located. Key sequences in the third intron have been conserved which allow for an alternative splicing mechanism, similar to the one found in mammals. The preprohormone contains two peptides with high homology to human proadrenomedullin N-terminal 20 peptide (PAMP) and hAM. We found through real-time PCR and immunocytochemistry cAM mRNA and peptide expression in a variety of chicken tissues, which parallel patterns observed for mammals, with the exception that cAM levels are almost non-detectable in brain. Similarly to mammals, cAM expression is upregulated under hypoxic conditions and following dexamethasone treatment. These data demonstrate a high degree of homology between the cAM gene and its mammalian ortholog and evolutionary conservation of the regulatory mechanisms controlling its expression.

Toll-like receptor (TLR) proteins mediate cellular activation by microbes and microbial products. To delineate the role of TLR proteins in the development of host immune responses against mycobacteria, wild-type and TLR-deficient mice were infected with nonpathogenic Mycobacterium bovis bacillus Calmette-Guerin (BCG). Two weeks after intraperitoneal challenge with BCG, few bacilli were present in the lungs of wild-type and TLR4−/− mice, whereas bacterial loads were tenfold higher in the lungs of infected TLR2−/− mice. BCG challenge in vitro strongly induced proinflammatory cytokine secretion by macrophages from wild-type and TLR4−/− mice but not by TLR2−/− macrophages. In contrast, intracellular uptake, intracellular bacterial growth, and suppression of intracellular bacterial growth in vitro by interferon-γ (IFN-γ) were similar in macrophages from all three mouse strains, suggesting that BCG growth in the lungs of TLR2−/− mice was a consequence of defective adaptive immunity. Antigenic stimulation of splenocytes from infected wild-type and TLR4−/− mice induced T cell proliferation in vitro, whereas T cells from TLR2−/− mice failed to proliferate. Unexpectedly, activated CD4+ T cells from both TLR-deficient mouse strains secreted little IFN-γ in vitro compared with control T cells. A role for TLR4 in the control of bacterial growth and IFN-γ production in vivo was observed only when mice were infected with higher numbers of BCG. Thus, TLR2 and TLR4 appear to regulate distinct aspects of the host immune response against BCG.

Adrenomedullin (AM) is a multiregulatory peptide which is expressed in a wide range of tissues. In the pancreas, AM was first found in mammals, including man, and its colocalization with the pancreatic polypeptide (PP) was established in islet F cells. In addition, three different AM receptors have been characterized in B-cells. AM has been also located in the pancreatic cells of other vertebrate classes. The frequency and distribution of AM cells vary between different animals; they can be found scattered among the exocrine tissue, in the islets, or in ductal epithelia. The colocalization of AM with other hormones presents different patterns, although in birds, as in mammals, it seems to colocalize only with PP. The best-determined pancreatic AM function is the inhibition of insulin secretion in B-cells, which seems to be linked to a recently discovered binding protein, factor H. In relation to this physiological role, clinical data show that AM is raised in some groups of both types I and II diabetic patients and AM might have triggered the disease in a subset of them. On the other hand, AM pancreatic cells are also involved in the response to septic shock by increasing AM circulating levels. A third putative function is the inhibition of amylase secretion by the exocrine pancreatic cells. AM has been found in embryonic mammalian pancreas from the earliest stages of the development, colocalizing with all pancreatic hormones, although in adults only coexpression with PP is kept. AM may play a role in the growth and morphogenesis of the pancreas.

It is well known that there are many different natural organisms able to adhere strongly to surfaces even in highly adverse conditions where conventional synthetic adhesives would fail. In the case of bacteria, this adhesion may occur through the formation of biofilms by means of diverse mechanisms that involve biopolymers such as proteins.This has led several scientists to undertake research on the use of adhesive proteins as the base polymer for the formulation of adhesives. Due to the nature of these polymers, the impact of such adhesives on the environment is expected to be reduced, their waste disposal being possible without the appearance of substances that may be considered harmful to the environment. Moreover, their synthesis and formulation can be achieved in an aqueous medium.In this review, a brief analysis is made of the natural adhesion systems that have inspired the authors to propose obtaining bacterial adhesive proteins by industrial microbiology and microbial biotechnology techniques as a way to achieve the current aim of the industry to reduce the environmental impact of their business and to produce distinctive items with an added value.

Over the past years, a holistic approach has been applied to the study of the field of receptor signaling, permitting the analysis of how the interaction between receptors and their cellular environment determines receptor function and the study of the role of these receptors, under both normal and pathophysiological conditions, in whole organisms. This has been facilitated by the development of high-resolution microscopy techniques, which allow single-molecule or spatiotemporal resolution, or both, of signaling processes at the cellular and organismal levels. Concurrently, the role of these signaling pathways can be tested in increasingly sophisticated murine disease models. Finally, computational approaches aid in predicting and understanding receptor behavior. The program of the Madrid meeting reflected this integrated approach, highlighting signaling by and dynamics and regulation of immune cell receptors, the T cell receptor and B cell receptor, and signaling by and regulation of G protein-coupled receptors.

Obesity is a highly prevalent health problem in Western countries that leads to many important diseases such as type 2 diabetes and metabolic syndrome being now considered an inflammatory chronic disease. Adipocytes are no longer considered passive cells storing fat since they are major producers of inflammatory cytokines during obesity. Adipocytes and macrophages share many biological properties including the synthesis of similar molecules regulating inflammation. Fatty acid levels are elevated in obesity and induce inflammatory pathways by yet a mostly unknown mechanism, leading to the development of insulin and leptin resistance. Recent studies suggest that these effects could be mediated through the activation of toll-like receptors (TLR). TLR signalling pathways might contribute to the development of obesity-associated insulin resistance, thus representing a connection between innate immunity and metabolism. Here, we summarize the recent evidence for the important role that TLRs play in adipose tissue, obesity and insulin resistance.

Our understanding of the key players involved in the differential regulation of T-cell responses during inflammation, infection and auto-immunity is fundamental for designing efficient therapeutic strategies against immune diseases. With respect to this, the inhibitory role of the lipid mediator prostaglandin E2 (PGE2) in T-cell immunity has been documented since the 1970s. Studies that ensued investigating the underlying mechanisms substantiated the suppressive function of micromolar concentrations of PGE2 in T-cell activation, proliferation, differentiation and migration. However, the past decade has seen a revolution in this perspective, since nanomolar concentrations of PGE2 have been shown to potentiate Th1 and Th17 responses and aid in T-cell proliferation. The understanding of concentration-specific effects of PGE2 in other cell types, the development of mice deficient in each subtype of the PGE2 receptors (EP receptors) and the delineation of signalling pathways mediated by the EP receptors have enhanced our understanding of PGE2 as an immune-stimulator. PGE2 regulates a multitude of functions in T-cell activation and differentiation and these effects vary depending on the micro-environment of the cell, maturation and activation state of the cell, type of EP receptor involved, local concentration of PGE2 and whether it is a homeostatic or inflammatory scenario. In this review, we compartmentalize the various aspects of this complex relationship of PGE2 with T lymphocytes. Given the importance of this molecule in T-cell activation, we also address the possibility of using EP receptor antagonism as a potential therapeutic approach for some immune disorders.

The aryl hydrocarbon receptor repressor (AHRR) is a bHLH/Per-ARNT-Sim transcription factor located in a region of chromosome 5 (5p15.3) that has been proposed to contain one or more tumor suppressor genes. We report here consistent downregulation of AHRR mRNA in human malignant tissue from different anatomical origins, including colon, breast, lung, stomach, cervix, and ovary, and demonstrate DNA hypermethylation as the regulatory mechanism of AHRR gene silencing. Knockdown of AHRR gene expression in a human lung can- cer cell line using siRNA significantly enhanced in vitro anchorage-dependent and -independent cell growth as well as cell growth after transplantation into immunocompromised mice. In addition, knockdown of AHRR in non-clonable normal human mammary epithelial cells enabled them to grow in an anchorage-independent manner. Further, downregulation of AHRR expression in the human lung cancer cell line conferred resistance to apoptotic signals and enhanced motility and invasion in vitro and angiogenic potential in vivo. Ectopic expression of AHRR in tumor cells resulted in diminished anchorage-dependent and -independent cell growth and reduced angiogenic potential. These results therefore demonstrate that AHRR is a putative new tumor suppressor gene in multiple types of human cancers.

IFN regulatory factor (IRF)-2−/− mice are significantly more resistant to LPS challenge than wild-type littermates, and this was correlated with increased numbers of apoptotic Kupffer cells. To assess the generality of this observation, and to understand the role of IRF-2 in apoptosis, responses of peritoneal macrophages from IRF-2+/+ and IRF-2−/− mice to apoptotic stimuli, including the fungal metabolite, gliotoxin, were compared. IRF-2−/− macrophages exhibited a consistently higher incidence of apoptosis that failed to correlate with caspase-3/7 activity. Using microarray gene expression profiling of liver RNA samples derived from IRF-2+/+ and IRF-2−/− mice treated with saline or LPS, we identified >40 genes that were significantly down-regulated in IRF-2−/− mice, including Stat3, which has been reported to regulate apoptosis. Compared with IRF-2+/+ macrophages, STAT3α mRNA was up-regulated constitutively or after gliotoxin treatment of IRF-2−/− macrophages, whereas STAT3β mRNA was down-regulated. Phospho-Y705-STAT3, phospho-S727-STAT1, and phospho-p38 protein levels were also significantly higher in IRF-2−/− than control macrophages. Activation of the STAT signaling pathway has been shown to elicit expression of CASP1 and apoptosis. IRF-2−/− macrophages exhibited increased basal and gliotoxin-induced caspase-1 mRNA expression and enhanced caspase-1 activity. Pharmacologic inhibition of STAT3 and caspase-1 abolished gliotoxin-induced apoptosis in IRF-2−/− macrophages. A novel IFN-stimulated response element, identified within the murine promoter of Casp1, was determined to be functional by EMSA and supershift analysis. Collectively, these data support the hypothesis that IRF-2 acts as a transcriptional repressor of Casp1, and that the absence of IRF-2 renders macrophages more sensitive to apoptotic stimuli in a caspase-1-dependent process.

Activation of TLRs is most closely associated with induction of pro-inflammatory gene expression; however, expression of many other genes, including the TLR genes themselves, has also been shown to be modulated following TLR engagement. A large family of nuclear transcription factors, the interferon regulatory factors (IRFs), have been implicated in TLR signaling leading to pro-inflammatory gene expression. Given that IRF-1 and IRF-2 counter-regulate the transcriptional activity of many genes, we hypothesized that IRF-1 and IRF-2 might also regulate TLR gene expression following LPS stimulation of murine macrophages. mRNA derived from medium- or LPS-treated primary peritoneal macrophages was analyzed for TLR gene expression using quantitative real-time PCR. In wild-type macrophages, LPS up-regulated expression of TLRs 1—3 and 6—9 steady-state mRNA, while TLR4 mRNA was modestly downregulated. IRF-2—/ —macrophages responded to LPS with dysregulated expression of TLR3, TLR4, and TLR5 mRNA, whereas IRF-1 deficiency dampened LPS-induced mRNA expression for TLR3, TLR6, and TLR9. Functional studies revealed aberrant TLR3 signaling in IRF-2—/ —macrophages. Collectively, these findings reveal an additional level of complexity associated with TLR transcriptional regulation and suggest that the trans-acting factors, IRF-1 and IRF-2, contribute to the innate immune response to infections by regulating TLR gene expression.

The ubiquitously expressed canonical transient receptor potential (TRPC) ion channels are considered important in Ca2+ signal generation, but their mechanisms of activation and roles remain elusive. Whereas most studies have examined overexpressed TRPC channels, we used molecular, biochemical, and electrophysiological approaches to assess the expression and function of endogenous TRPC channels in A7r5 smooth muscle cells. Real time PCR and Western analyses reveal TRPC6 as the only member of the diacylglycerol-responsive TRPC3/6/7 subfamily of channels expressed at significant levels in A7r5 cells. TRPC1, TRPC4, and TRPC5 were also abundant. An outwardly rectifying, nonselective cation current was activated by phospholipase C-coupled vasopressin receptor activation or by the diacylglycerol analogue, oleoyl-2-acetyl-sn-glycerol (OAG). Introduction of TRPC6 small interfering RNA sequences into A7r5 cells by electroporation led to 90% reduction of TRPC6 transcript and 80% reduction of TRPC6 protein without any detectable compensatory changes in the expression of other TRPC channels. The OAG-activated nonselective cation current was similarly reduced by TRPC6 RNA interference. Intracellular Ca2+ measurements using fura-2 revealed that thapsigargin-induced store-operated Ca2+ entry was unaffected by TRPC6 knockdown, whereas vasopressin-induced Ca2+ entry was suppressed by more than 50%. In contrast, OAG-induced Ca2+ transients were unaffected by TRPC6 knockdown. Nevertheless, OAG-induced Ca2+ entry bore the hallmarks of TRPC6 function; it was inhibited by protein kinase C and blocked by the Src-kinase inhibitor, 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2). Importantly, OAG-induced Ca2+ entry was blocked by the potent L-type Ca2+ channel inhibitor, *nimodipine. Thus, TRPC6 activation probably results primarily in Na ion entry and depolarization, leading to activation of L-type channels as the mediators of Ca2+ entry. Calculations reveal that even 90% reduction of TRPC6 channels would allow depolarization sufficient to activate L-type channels. This tight coupling between TRPC6 and L-type channels is probably important in mediating smooth muscle cell membrane potential and muscle contraction.

Adrenomedullin (AM) is a potent vasodilator peptide present in the lung of mammals where it is expressed mainly in the columnar epithelium and alveolar macrophages. AM increases the secretion of phosphatidylcholine by type II pneumocytes, which suggests a role as an autocrine modulator of surfactant secretion. In this study we show the expression of an AM-like protein in the lung of the pigeon, Columba livia. Using an antibody against its human ortholog, AM-like immunoreactivity was found to be associated with membranous structures of the multivesicular bodies of type II pneumocytes. We also studied the differential expression of AM-like peptide in the lung of pigeons exposed to polluted city air vs cleaner countryside conditions and found that AM-like expression was higher in city animals. Similar results were obtained in an experimental study in which pigeons were exposed to increasing concentrations of a single pollutant, ozone. Taken together, our findings support the implication of AM in the response of type II pneumocytes to air pollutants.

Stimulation of the APC by Porphyromonas gingivalis LPS has been shown to result in the production of certain pro- and anti-inflammatory cytokines. However, the signaling pathways that regulate these processes are currently unknown. In the present study, the role of the phosphatidylinositol 3 kinase (PI3K)-Akt pathway in regulating P. gingivalis LPS-induced production of IL-10, IL-12 p40, and IL-12 p70 by human monocytes was investigated. P. gingivalis LPS selectively activates the PI3K-Akt pathway via Toll-like receptor 2, and inhibition of this pathway results in an abrogation of extracellular signal-regulated kinase 1/2 phosphorylation, whereas the activation of p38 and c-Jun N-terminal kinase 1/2 kinases were unaffected. Analysis of cytokine production following stimulation of monocytes with P. gingivalis LPS revealed that inhibition of the PI3K pathway differentially regulated IL-10 and IL-12 synthesis. IL-10 production was suppressed, whereas IL-12 levels were enhanced. Inhibition of P. gingivalis LPS-mediated activation of the PI3K-Akt pathway resulted in a pronounced augmentation of NF-κB p65 that was independent of IκB-α degradation. Furthermore, the ability of the PI3K-Akt pathway to modulate IL-10 and IL-12 production appears to be mediated by the selective suppression of extracellular signal-regulated kinase 1/2 activity, as the MEK1 inhibitor PD98059 closely mimicked the effects of wortmannin and LY294002 to differentially regulate IL-10 and IL-12 production by P. gingivalis LPS-stimulated monocytes. These studies provide new insight into how engagement of the PI3K-Akt pathway by P. gingivalis LPS affects the induction of key immunoregulatory cytokines that control both qualitative and quantitative aspects of innate and adaptive immunity.

In this study, tolerance induction by preexposure of murine macrophages to Toll-like receptor (TLR)2 and TLR4 agonists was revisited, focusing on the major signaling components associated with NF-κB activation. Pretreatment of macrophages with a pure TLR4 agonist (protein-free Escherichia coli (Ec) LPS) or with TLR2 agonists (Porphyromonas gingivalis LPS or synthetic lipoprotein Pam3Cys) led to suppression of TNF-α secretion, IL-1R-associated kinase-1, and IκB kinase (IKK) kinase activities, c-jun N-terminal kinase, and extracellular signal-regulated kinase phosphorylation, and to suppression of NF-κB DNA binding and transactivation upon challenge with the same agonist (TLR4 or TLR2 “homotolerance,” respectively). Despite inhibited NF-κB DNA binding, increased levels of nuclear NF-κB were detected in agonist-pretreated macrophages. For all the intermediate signaling elements, heterotolerance was weaker than TLR4 or TLR2 homotolerance with the exception of IKK kinase activity. IKK kinase activity was unperturbed in heterotolerance. TNF-α secretion was also suppressed in P. gingivalis LPS-pretreated, Ec LPS-challenged cells, but not vice versa, while Pam3Cys and Ec LPS did not induce a state of cross-tolerance at the level of TNF-α. Experiments designed to elucidate novel mechanisms of NF-κB inhibition in tolerized cells revealed the potential contribution of IκBε and IκBξ inhibitory proteins and the necessity of TLR4 engagement for induction of tolerance to Toll receptor-IL-1R domain-containing adapter protein/MyD88-adapter-like-dependent gene expression. Collectively, these data demonstrate that induction of homotolerance affects a broader spectrum of signaling components than in heterotolerance, with selective modulation of specific elements within the NF-κB signaling pathway.