Hepcidin is a peptide hormone that regulates iron homeostasis and acts as an antimicrobial peptide. It is expressed and secreted by a variety of cell types in response to iron loading and inflammation. Hepcidin mediates iron homeostasis by binding to the iron exporter ferroportin, inducing its internalization and degradation via activation of the protein kinase Jak2 and the subsequent phosphorylation of ferroportin. Here we have shown that hepcidin-activated Jak2 also phosphorylates the transcription factor Stat3, resulting in a transcriptional response. Hepcidin treatment of ferroportin-expressing mouse macrophages showed changes in mRNA expression levels of a wide variety of genes. The changes in transcript levels for half of these genes were a direct effect of hepcidin, as shown by cycloheximide insensitivity, and dependent on the presence of Stat3. Hepcidin-mediated transcriptional changes modulated LPS-induced transcription in both cultured macrophages and in vivo mouse models, as demonstrated by suppression of IL-6 and TNF-α transcript and secreted protein. Hepcidin-mediated transcription in mice also suppressed toxicity and morbidity due to single doses of LPS, poly(I:C), and turpentine, which is used to model chronic inflammatory disease. Most notably, we demonstrated that hepcidin pretreatment protected mice from a lethal dose of LPS and that hepcidin-knockout mice could be rescued from LPS toxicity by injection of hepcidin. The results of our study suggest a new function for hepcidin in modulating acute inflammatory responses.
Ivana De Domenico, Tian Y. Zhang, Curry L. Koening, Ryan W. Branch, Nyall London, Eric Lo, Raymond A. Daynes, James P. Kushner, Dean Li, Diane M. Ward, Jerry Kaplan
Necrosis stimulates inflammation, and this response is medically relevant because it contributes to the pathogenesis of a number of diseases. It is thought that necrosis stimulates inflammation because dying cells release proinflammatory molecules that are recognized by the immune system. However, relatively little is known about the molecular identity of these molecules and their contribution to responses in vivo. Here, we investigated the role of uric acid in the inflammatory response to necrotic cells in mice. We found that dead cells not only released intracellular stores of uric acid but also produced it in large amounts postmortem as nucleic acids were degraded. Using newly developed Tg mice that have reduced levels of uric acid either intracellularly and/or extracellularly, we found that uric acid depletion substantially reduces the cell death–induced inflammatory response. Similar results were obtained with pharmacological treatments that reduced uric acid levels either by blocking its synthesis or hydrolyzing it in the extracellular fluids. Importantly, uric acid depletion selectively inhibited the inflammatory response to dying cells but not to microbial molecules or sterile irritant particles. Collectively, our data identify uric acid as a proinflammatory molecule released from dying cells that contributes significantly to the cell death–induced inflammatory responses in vivo.
Hajime Kono, Chun-Jen Chen, Fernando Ontiveros, Kenneth L. Rock
Activated protein C (APC), the only FDA-approved biotherapeutic drug for sepsis, possesses anticoagulant, antiinflammatory, and barrier-protective activities. However, the mechanisms underlying its antiinflammatory functions are not well defined. Here, we report that the antiinflammatory activity of APC on macrophages is dependent on integrin CD11b/CD18, but not on endothelial protein C receptor (EPCR). We showed that CD11b/CD18 bound APC within specialized membrane microdomains/lipid rafts and facilitated APC cleavage and activation of protease-activated receptor–1 (PAR1), leading to enhanced production of sphingosine-1-phosphate (S1P) and suppression of the proinflammatory response of activated macrophages. Deletion of the γ-carboxyglutamic acid domain of APC, a region critical for its anticoagulant activity and EPCR-dependent barrier protection, had no effect on its antiinflammatory function. Genetic inactivation of CD11b, PAR1, or sphingosine kinase–1, but not EPCR, abolished the ability of APC to suppress the macrophage inflammatory response in vitro. Using an LPS-induced mouse model of lethal endotoxemia, we showed that APC administration reduced the mortality of wild-type mice, but not CD11b-deficient mice. These data establish what we believe to be a novel mechanism underlying the antiinflammatory activity of APC in the setting of endotoxemia and provide clear evidence that the antiinflammatory function of APC is distinct from its barrier-protective function and anticoagulant activities.
Chunzhang Cao, Yamei Gao, Yang Li, Toni M. Antalis, Francis J. Castellino, Li Zhang
HSCs are BM-derived, self-renewing multipotent cells that develop into circulating blood cells. They have been implicated in the repair of inflamed parenchymal tissue, but the signals that regulate their trafficking to sites of inflammation are unknown. As monocytes are recruited to sites of inflammation via chemoattractants that activate CCR2 on their surface, we investigated whether HSCs are also recruited to sites of inflammation through CCR2. Initial analysis indicated that in mice, CCR2 was expressed on subsets of HSCs and hematopoietic progenitor cells (HPCs) and that freshly isolated primitive hematopoietic cells (Lin–c-Kit+ cells) responded to CCR2 ligands in vitro. In vivo analysis indicated that after instillation of thioglycollate to cause aseptic inflammation and after administration of acetaminophen to induce liver damage, endogenous HSCs/HPCs were actively recruited to the peritoneum and liver, respectively, in WT but not Ccr2–/– mice. HSCs/HPCs recovered from the peritoneum successfully engrafted into the BM of irradiated primary and secondary recipients, confirming their self renewal and multipotency. Importantly, administration of exogenous WT, but not Ccr2–/–, HSCs/HPCs accelerated resolution of acetaminophen-induced liver damage and triggered the expression of genes characteristic of the macrophage M2 or repair phenotype. These findings reveal what we believe to be a novel role for CCR2 in the homing of HSCs/HPCs to sites of inflammation and suggest new functions for chemokines in promoting tissue repair and regeneration.
Yue Si, Chia-Lin Tsou, Kelsey Croft, Israel F. Charo
Down syndrome critical region gene 1 (DSCR-1) short variant (DSCR-1s) is an inhibitor of calcineurin/NFAT signaling encoded by exons 4–7 of DSCR1. We previously reported that VEGF induces DSCR-1s expression in endothelial cells, which in turn negatively feeds back to attenuate endothelial cell activation. Here, in order to characterize the role of the promoter that drives DSCR-1s expression in mediating inducible expression in vivo and to determine the functional relevance of DSCR-1s in inflammation, we targeted a DNA construct containing 1.7 kb of the human DSCR1s promoter coupled to the lacZ reporter to the hypoxanthine guanine phosphoribosyl transferase (Hprt) locus of mice. We determined that lacZ was uniformly expressed in the endothelium of transgenic embryos but was markedly downregulated postnatally. Systemic administration of VEGF or LPS in adult mice resulted in cyclosporine A–sensitive reactivation of the DSCR1s promoter and endogenous gene expression in a subset of organs, including the heart and brain. The DSCR1s promoter was similarly induced in the endothelium of tumor xenografts. In a mouse model of endotoxemia, DSCR-1s–deficient mice demonstrated increased sepsis mortality, whereas adenovirus-mediated DSCR-1s overexpression protected against LPS-induced lethality. Collectively, these data suggest that the DSCR1s promoter directs vascular bed–specific expression in activated endothelium and that DSCR-1s serves to dampen the host response to infection.
Takashi Minami, Kiichiro Yano, Mai Miura, Mika Kobayashi, Jun-ichi Suehiro, Patrick C. Reid, Takao Hamakubo, Sandra Ryeom, William C. Aird, Tatsuhiko Kodama
Mucosal diseases are often characterized by an inflammatory infiltrate that includes polymorphonuclear leukocytes (PMNs), monocytes, lymphocytes, and platelets. A number of studies have suggested that the interaction of platelets with leukocytes has an essential proinflammatory role. Here, we examined whether platelets migrate across mucosal epithelium, as PMNs are known to do, and whether platelets influence epithelial cell function. Initial studies revealed that human platelets did not efficiently transmigrate across human epithelial cell monolayers. However, in the presence of human PMNs, platelet movement across the epithelium was proportional to the extent of PMN transmigration, and strategies that blocked PMN transmigration diminished platelet movement. Furthermore, platelet-PMN comigration was observed in intestinal tissue derived from human patients with inflammatory bowel disease (IBD). The translocated platelets were found to release large quantities of ATP, which was metabolized to adenosine via a 2-step enzymatic reaction mediated by ecto-nucleotidases, including CD73 and ecto–nucleoside triphosphate diphosphohydrolases (ecto-NTPDases), expressed on the apical membrane of the intestinal epithelial cells. In vitro studies and a mouse model of intestinal inflammation were employed to define a mechanism involving adenosine-mediated induction of electrogenic chloride secretion, with concomitant water movement into the intestinal lumen. These studies demonstrate that ecto-NTPDases are expressed on the apical membrane of epithelial cells and are involved in what we believe to be a previously unappreciated function for platelets in the inflamed intestine, which might promote bacterial clearance under inflammatory conditions.
Thomas Weissmüller, Eric L. Campbell, Peter Rosenberger, Melanie Scully, Paul L. Beck, Glenn T. Furuta, Sean P. Colgan
Inflammation plays a critical role in the development of cardiovascular diseases. Infiltration of leukocytes to sites of injury requires their exit from the blood and migration across basement membrane; this process has been postulated to require remodeling of the ECM. Plasminogen (Plg) is a protease that binds to the ECM and, upon conversion to plasmin, degrades multiple ECM proteins. In addition, plasmin directly activates MMPs. Here, we used Plg–/– mice to investigate the role of Plg in inflammatory leukocyte migration. After induction of peritonitis by thioglycollate injection, we found that Plg–/– mice displayed diminished macrophage trans-ECM migration and decreased MMP-9 activation. Furthermore, injection of the active form of MMP-9 in Plg–/– mice rescued macrophage migration in this model. We used periaortic application of CaCl2 to induce abdominal aortic aneurysm (AAA) and found that Plg–/– mice displayed reduced macrophage infiltration and were protected from aneurysm formation. Administration of active MMP-9 to Plg–/– mice promoted macrophage infiltration and the development of AAA. These data suggest that Plg regulates macrophage migration in inflammation via activation of MMP-9, which, in turn, regulates the ability of the cells to migrate across ECM. Thus, targeting the Plg/MMP-9 pathway may be an attractive approach to regulate inflammatory responses and AAA development.
Yanqing Gong, Erika Hart, Aleksey Shchurin, Jane Hoover-Plow
Neutrophil granulocytes form the body’s first line of antibacterial defense, but they also contribute to tissue injury and noninfectious, chronic inflammation. Proteinase 3 (PR3) and neutrophil elastase (NE) are 2 abundant neutrophil serine proteases implicated in antimicrobial defense with overlapping and potentially redundant substrate specificity. Here, we unraveled a cooperative role for PR3 and NE in neutrophil activation and noninfectious inflammation in vivo, which we believe to be novel. Mice lacking both PR3 and NE demonstrated strongly diminished immune complex–mediated (IC-mediated) neutrophil infiltration in vivo as well as reduced activation of isolated neutrophils by ICs in vitro. In contrast, in mice lacking just NE, neutrophil recruitment to ICs was only marginally impaired. The defects in mice lacking both PR3 and NE were directly linked to the accumulation of antiinflammatory progranulin (PGRN). Both PR3 and NE cleaved PGRN in vitro and during neutrophil activation and inflammation in vivo. Local administration of recombinant PGRN potently inhibited neutrophilic inflammation in vivo, demonstrating that PGRN represents a crucial inflammation-suppressing mediator. We conclude that PR3 and NE enhance neutrophil-dependent inflammation by eliminating the local antiinflammatory activity of PGRN. Our results support the use of serine protease inhibitors as antiinflammatory agents.
Kai Kessenbrock, Leopold Fröhlich, Michael Sixt, Tim Lämmermann, Heiko Pfister, Andrew Bateman, Azzaq Belaaouaj, Johannes Ring, Markus Ollert, Reinhard Fässler, Dieter E. Jenne
Cigarette smoke (CS) inhalation causes an early inflammatory response in rodent airways by stimulating capsaicin-sensitive sensory neurons that express transient receptor potential cation channel, subfamily V, member 1 (TRPV1) through an unknown mechanism that does not involve TRPV1. We hypothesized that 2 α,β-unsaturated aldehydes present in CS, crotonaldehyde and acrolein, induce neurogenic inflammation by stimulating TRPA1, an excitatory ion channel coexpressed with TRPV1 on capsaicin-sensitive nociceptors. We found that CS aqueous extract (CSE), crotonaldehyde, and acrolein mobilized Ca2+ in cultured guinea pig jugular ganglia neurons and promoted contraction of isolated guinea pig bronchi. These responses were abolished by a TRPA1-selective antagonist and by the aldehyde scavenger glutathione but not by the TRPV1 antagonist capsazepine or by ROS scavengers. Treatment with CSE or aldehydes increased Ca2+ influx in TRPA1-transfected cells, but not in control HEK293 cells, and promoted neuropeptide release from isolated guinea pig airway tissue. Furthermore, the effect of CSE and aldehydes on Ca2+ influx in dorsal root ganglion neurons was abolished in TRPA1-deficient mice. These data identify α,β-unsaturated aldehydes as the main causative agents in CS that via TRPA1 stimulation mediate airway neurogenic inflammation and suggest a role for TRPA1 in the pathogenesis of CS-induced diseases.
Eunice Andrè, Barbara Campi, Serena Materazzi, Marcello Trevisani, Silvia Amadesi, Daniela Massi, Christophe Creminon, Natalya Vaksman, Romina Nassini, Maurizio Civelli, Pier Giovanni Baraldi, Daniel P. Poole, Nigel W. Bunnett, Pierangelo Geppetti, Riccardo Patacchini
In rodents and humans, alcohol exposure has been shown to predispose the pancreas to cholinergic or viral induction of pancreatitis. We previously developed a rodent model in which exposure to an ethanol (EtOH) diet, followed by carbachol (Cch) stimulation, redirects exocytosis from the apical to the basolateral plasma membrane of acinar cells, resulting in ectopic zymogen enzyme activation and pancreatitis. This redirection of exocytosis involves a soluble NSF attachment receptor (SNARE) complex consisting of syntaxin-4 and synapse-associated protein of 23 kDa (SNAP-23). Here, we investigated the role of the zymogen granule (ZG) SNARE vesicle-associated membrane protein 8 (VAMP8) in mediating basolateral exocytosis. In WT mice, in vitro EtOH exposure or EtOH diet reduced Cch-stimulated amylase release by redirecting apical exocytosis to the basolateral membrane, leading to alcoholic pancreatitis. Further reduction of zymogen secretion, caused by blockade of both apical and basolateral exocytosis and resulting in a more mild induction of alcoholic pancreatitis, was observed in Vamp8–/– mice in response to these treatments. In addition, although ZGs accumulated in Vamp8–/– acinar cells, ZG-ZG fusions were reduced compared with those in WT acinar cells, as visualized by electron microscopy. This reduction in ZG fusion may account for reduced efficiency of apical exocytosis in Vamp8–/– acini. These findings indicate that VAMP8 is the ZG-SNARE that mediates basolateral exocytosis in alcoholic pancreatitis and that VAMP8 is critical for ZG-ZG homotypic fusion.
Laura I. Cosen-Binker, Marcelo G. Binker, Cheng-Chun Wang, Wanjin Hong, Herbert Y. Gaisano
Intestinal macrophages play a central role in regulation of immune responses against commensal bacteria. In general, intestinal macrophages lack the expression of innate-immune receptor CD14 and do not produce proinflammatory cytokines against commensal bacteria. In this study, we identified what we believe to be a unique macrophage subset in human intestine. This subset expressed both macrophage (CD14, CD33, CD68) and DC markers (CD205, CD209) and produced larger amounts of proinflammatory cytokines, such as IL-23, TNF-α, and IL-6, than typical intestinal resident macrophages (CD14–CD33+ macrophages). In patients with Crohn disease (CD), the number of these CD14+ macrophages were significantly increased compared with normal control subjects. In addition to increased numbers of cells, these cells also produced larger amounts of IL-23 and TNF-α compared with those in normal controls or patients with ulcerative colitis. In addition, the CD14+ macrophages contributed to IFN-γ production rather than IL-17 production by lamina propria mononuclear cells (LPMCs) dependent on IL-23 and TNF-α. Furthermore, the IFN-γ produced by LPMCs triggered further abnormal macrophage differentiation with an IL-23–hyperproducing phenotype. Collectively, these data suggest that this IL-23/IFN-γ–positive feedback loop induced by abnormal intestinal macrophages contributes to the pathogenesis of chronic intestinal inflammation in patients with CD.
Nobuhiko Kamada, Tadakazu Hisamatsu, Susumu Okamoto, Hiroshi Chinen, Taku Kobayashi, Toshiro Sato, Atsushi Sakuraba, Mina T. Kitazume, Akira Sugita, Kazutaka Koganei, Kiyoko S. Akagawa, Toshifumi Hibi
In the liver, the JNK cascade is induced downstream of TNF receptors (TNFRs) in response to inflammatory, microbial, and toxic challenges. Sustained activation of JNK triggers programmed cell death (PCD), and hepatocyte survival during these challenges requires induction of the NF-κB pathway, which antagonizes this activation by upregulating target genes. Thus, modulation of JNK activity is crucial to the liver response to TNFR-mediated challenge. The basis for this modulation, however, is unknown. Here, we investigated the role of the NF-κB target Gadd45b in the regulation of hepatocyte fate during liver regeneration after partial hepatectomy. We generated Gadd45b–/– mice and found that they exhibited decreased hepatocyte proliferation and increased PCD during liver regeneration. Notably, JNK activity was markedly increased and sustained in livers of Gadd45b–/– mice compared with control animals after partial hepatectomy. Furthermore, imposition of a Jnk2-null mutation, attenuating JNK activity, completely rescued the regenerative response in Gadd45b–/– mice. Interestingly, Gadd45β ablation did not affect hepatotoxic JNK signaling after a TNFR-mediated immune challenge, suggesting specificity in the inducible hepatic program for JNK restraint activated during distinct TNFR-mediated challenges. These data provide a basis for JNK suppression during liver regeneration and identify Gadd45β as a potential therapeutic target in liver diseases.
Salvatore Papa, Francesca Zazzeroni, Yang-Xin Fu, Concetta Bubici, Kellean Alvarez, Kathryn Dean, Peter A. Christiansen, Robert A. Anders, Guido Franzoso
Septic shock results from an uncontrolled inflammatory response, mediated primarily by LPS. Cholesterol transport plays an important role in the host response to LPS, as LPS is neutralized by lipoproteins and adrenal cholesterol uptake is required for antiinflammatory glucocorticoid synthesis. In this study, we show that scavenger receptor B-I (SR-BI), an HDL receptor that mediates HDL cholesterol ester uptake into cells, is required for the normal antiinflammatory response to LPS-induced endotoxic shock. Despite elevated plasma HDL levels, SR-BI–null mice displayed an uncontrollable inflammatory cytokine response and a markedly higher lethality rate than control mice in response to LPS. In addition, SR-BI–null mice showed a lack of inducible glucocorticoid synthesis in response to LPS, bacterial infection, stress, or ACTH. Glucocorticoid insufficiency in SR-BI–null mice was due to primary adrenal malfunction resulting from deficient cholesterol delivery from HDL. Furthermore, corticosterone supplementation decreased the sensitivity of SR-BI–null mice to LPS. Plasma from control and SR-BI–null mice exhibited a similar ability to neutralize LPS, whereas SR-BI–null mice showed decreased plasma clearance of LPS into the liver and hepatocytes compared with normal mice. We conclude that SR-BI in mice is required for the antiinflammatory response to LPS-induced endotoxic shock, likely through its essential role in facilitating glucocorticoid production and LPS hepatic clearance.
Lei Cai, Ailing Ji, Frederick C. de Beer, Lisa R. Tannock, Deneys R. van der Westhuyzen
Sepsis is characterized by a systemic response to severe infection. Although the inflammatory phase of sepsis helps eradicate the infection, it can have detrimental consequences if left unchecked. Therapy directed against inflammatory mediators of sepsis has shown little success and has the potential to impair innate antimicrobial defenses. Heme oxygenase-1 (HO-1) and the product of its enzymatic reaction, CO, have beneficial antiinflammatory properties, but little is known about their effects on microbial sepsis. Here, we have demonstrated that during microbial sepsis, HO-1–derived CO plays an important role in the antimicrobial process without inhibiting the inflammatory response. HO-1–deficient mice suffered exaggerated lethality from polymicrobial sepsis. Targeting HO-1 to SMCs and myofibroblasts of blood vessels and bowel ameliorated sepsis-induced death associated with Enterococcus faecalis, but not Escherichia coli, infection. The increase in HO-1 expression did not suppress circulating inflammatory cells or their accumulation at the site of injury but did enhance bacterial clearance by increasing phagocytosis and the endogenous antimicrobial response. Furthermore, injection of a CO-releasing molecule into WT mice increased phagocytosis and rescued HO-1–deficient mice from sepsis-induced lethality. These data advocate HO-1–derived CO as an important mediator of the host defense response to sepsis and suggest CO administration as a possible treatment for the disease.
Su Wol Chung, Xiaoli Liu, Alvaro A. Macias, Rebecca M. Baron, Mark A. Perrella
Fibrin deposition within joints is a prominent feature of arthritis, but the precise contribution of fibrin(ogen) to inflammatory events that cause debilitating joint damage remains unknown. To determine the importance of fibrin(ogen) in arthritis, gene-targeted mice either deficient in fibrinogen (Fib–) or expressing mutant forms of fibrinogen, lacking the leukocyte receptor integrin αMβ2 binding motif (Fibγ390–396A) or the αIIbβ3 platelet integrin-binding motif (FibγΔ5), were challenged with collagen-induced arthritis (CIA). Fib– mice exhibited fewer affected joints and reduced disease severity relative to controls. Similarly, diminished arthritis was observed in Fibγ390–396A mice, which retain full clotting function. In contrast, arthritis in FibγΔ5 mice was indistinguishable from that of controls. Fibrin(ogen) was not essential for leukocyte trafficking to joints, but appeared to be involved in leukocyte activation events. Fib– and Fibγ390–396A mice with CIA displayed reduced local expression of TNF-α, IL-1β, and IL-6, which suggests that αMβ2-mediated leukocyte engagement of fibrin is mechanistically upstream of the production of proinflammatory mediators. Supporting this hypothesis, arthritic disease driven by exuberant TNF-α expression was not impeded by fibrinogen deficiency. Thus, fibrin(ogen) is an important, but context-dependent, determinant of arthritis, and one mechanism linking fibrin(ogen) to joint disease is coupled to αMβ2-mediated inflammatory processes.
Matthew J. Flick, Christine M. LaJeunesse, Kathryn E. Talmage, David P. Witte, Joseph S. Palumbo, Malinda D. Pinkerton, Sherry Thornton, Jay L. Degen
Triggering receptor expressed on myeloid cells–1 (TREM-1) potently amplifies acute inflammatory responses by enhancing degranulation and secretion of proinflammatory mediators. Here we demonstrate that TREM-1 is also crucially involved in chronic inflammatory bowel diseases (IBD). Myeloid cells of the normal intestine generally lack TREM-1 expression. In experimental mouse models of colitis and in patients with IBD, however, TREM-1 expression in the intestine was upregulated and correlated with disease activity. TREM-1 significantly enhanced the secretion of relevant proinflammatory mediators in intestinal macrophages from IBD patients. Blocking TREM-1 by the administration of an antagonistic peptide substantially attenuated clinical course and histopathological alterations in experimental mouse models of colitis. This effect was also seen when the antagonistic peptide was administered only after the first appearance of clinical signs of colitis. Hence, TREM-1–mediated amplification of inflammation contributes not only to the exacerbation of acute inflammatory disorders but also to the perpetuation of chronic inflammatory disorders. Furthermore, interfering with TREM-1 engagement leads to the simultaneous reduction of production and secretion of a variety of pro-inflammatory mediators such as TNF, IL-6, IL-8 (CXCL8), MCP-1 (CCL2), and IL-1β. Therefore, TREM-1 may also represent an attractive target for the treatment of chronic inflammatory disorders.
Mirjam Schenk, Axel Bouchon, Frank Seibold, Christoph Mueller
Monocyte recruitment to sites of inflammation is regulated by members of the chemokine family of chemotactic cytokines. However, the mechanisms that govern the migration of monocytes from bone marrow to blood and from blood to inflamed tissues are not well understood. Here we report that CC chemokine receptor 2 (CCR2) is highly expressed on a subpopulation of blood monocytes whose numbers are markedly decreased in CCR2–/– mice. In bone marrow, however, CCR2–/– mice had an increased number of monocytes, suggesting that CCR2 is critical for monocyte egress. Intravenous infusion of ex vivo–labeled WT or CCR2–/– bone marrow into WT recipient mice demonstrated that CCR2 is necessary for efficient monocyte recruitment from the blood to inflamed tissue. Analysis of mice lacking monocyte chemoattractant protein–1 (MCP-1), MCP-3, MCP-5, or MCP-2 plus MCP-5 revealed that MCP-3 and MCP-1 are the CCR2 agonists most critical for the maintenance of normal blood monocyte counts. These findings provide evidence that CCR2 and MCP-3/MCP-1 are critical for monocyte mobilization and suggest new roles for monocyte chemoattractants in leukocyte homeostasis.
Chia-Lin Tsou, Wendy Peters, Yue Si, Sarah Slaymaker, Ara M. Aslanian, Stuart P. Weisberg, Matthias Mack, Israel F. Charo
Duchenne muscular dystrophy (DMD) is a lethal X-linked disorder associated with dystrophin deficiency that results in chronic inflammation and severe skeletal muscle degeneration. In DMD mouse models and patients, we find that IκB kinase/NF-κB (IKK/NF-κB) signaling is persistently elevated in immune cells and regenerative muscle fibers. Ablation of 1 allele of the p65 subunit of NF-κB was sufficient to improve pathology in mdx mice, a model of DMD. In addition, conditional deletion of IKKβ in mdx mice elucidated that NF-κB functions in activated macrophages to promote inflammation and muscle necrosis and in skeletal muscle fibers to limit regeneration through the inhibition of muscle progenitor cells. Furthermore, specific pharmacological inhibition of IKK resulted in improved pathology and muscle function in mdx mice. Collectively, these results underscore the critical role of NF-κB in the progression of muscular dystrophy and suggest the IKK/NF-κB signaling pathway as a potential therapeutic target for DMD.
Swarnali Acharyya, S. Armando Villalta, Nadine Bakkar, Tepmanas Bupha-Intr, Paul M.L. Janssen, Micheal Carathers, Zhi-Wei Li, Amer A. Beg, Sankar Ghosh, Zarife Sahenk, Michael Weinstein, Katherine L. Gardner, Jill A. Rafael-Fortney, Michael Karin, James G. Tidball, Albert S. Baldwin, Denis C. Guttridge
Tight regulation of COX-2 expression is a key feature controlling eicosanoid production in atherosclerosis and other inflammatory syndromes. Adhesive interactions between platelets and monocytes occur in these conditions and deliver specific signals that trigger inflammatory gene expression. Using a cellular model of monocyte signaling induced by activated human platelets, we identified the central posttranscriptional mechanisms that regulate timing and magnitude of COX-2 expression. Tethering of monocytes to platelets and to purified P-selectin, a key adhesion molecule displayed by activated platelets, induces NF-κB activation and COX-2 promoter activity. Nevertheless, COX-2 mRNA is rapidly degraded, leading to aborted protein synthesis. Time-dependent signaling of monocytes induces a second phase of transcript accumulation accompanied by COX-2 enzyme synthesis and eicosanoid production. Here, generation of IL-1β, a proinflammatory cytokine, promoted stabilization of COX-2 mRNA by silencing of the AU-rich mRNA decay element (ARE) in the 3′-untranslated region (3'UTR) of the mRNA. Consistent with observed mRNA stabilization, activated platelets or IL-1β treatment induced cytoplasmic accumulation and enhanced ARE binding of the mRNA stability factor HuR in monocytes. These findings demonstrate that activated platelets induce COX-2 synthesis in monocytes by combinatorial signaling to transcriptional and posttranscriptional checkpoints. These checkpoints may be altered in disease and therefore useful as targets for antiinflammatory intervention.
Dan A. Dixon, Neal D. Tolley, Kristi Bemis-Standoli, Mark L. Martinez, Andrew S. Weyrich, Jason D. Morrow, Stephen M. Prescott, Guy A. Zimmerman
Multiple and paradoxical effects of airway smooth muscle (ASM) 7-transmembrane–spanning receptors activated during asthma, or by treatment with bronchodilators such as β2–adrenergic receptor (β2AR) agonists, indicate extensive receptor crosstalk. We examined the signaling of the prostanoid-EP1 receptor, since its endogenous agonist prostaglandin E2 is abundant in the airway, but its functional implications are poorly defined. Activation of EP1 failed to elicit ASM contraction in mouse trachea via this Gαq-coupled receptor. However, EP1 activation markedly reduced the bronchodilatory function of β2AR agonist, but not forskolin, indicating an early pathway interaction. Activation of EP1 reduced β2AR-stimulated cAMP in ASM but did not promote or augment β2AR phosphorylation or alter β2AR trafficking. Bioluminescence resonant energy transfer showed EP1 and β2AR formed heterodimers, which were further modified by EP1 agonist. In cell membrane [35S]GTPγS binding studies, the presence of the EP1 component of the dimer uncoupled β2AR from Gαs, an effect accentuated by EP1 agonist activation. Thus alone, EP1 does not appear to have a significant direct effect on airway tone but acts as a modulator of the β2AR, altering Gαs coupling via steric interactions imposed by the EP1:β2AR heterodimeric signaling complex and ultimately affecting β2AR-mediated bronchial relaxation. This mechanism may contribute to β-agonist resistance found in asthma.
Dennis W. McGraw, Kathryn A. Mihlbachler, Mary Rose Schwarb, Fahema F. Rahman, Kersten M. Small, Khalid F. Almoosa, Stephen B. Liggett
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