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Biomedical subjects

Kevin J Tracey

Publications and source records attributed to Kevin J Tracey.

At least 73 records · Page 4Linked to original sources
PubMed2003

Activation of gene expression in human neutrophils by high mobility group box 1 protein.

High mobility group box 1 (HMGB1) protein, a DNA binding protein that stabilizes nucleosomes and facilitates transcription, was recently identified as a late mediator of endotoxin lethality. High serum HMGB1 levels in patients with sepsis are associated with increased mortality, and administration of HMGB1 produces acute inflammation in animal models of lung injury and endotoxemia. Neutrophils occupy a critical role in mediating the development of endotoxemia-associated acute lung injury, but previously it was not known whether HMGB1 could influence neutrophil activation. In the present experiments, we demonstrate that HMGB1 increases the nuclear translocation of NF-kappaB and enhances the expression of proinflammatory cytokines in human neutrophils. These proinflammatory effects of HMGB1 in neutrophils appear to involve the p38 MAPK, phosphatidylinositol 3-kinase/Akt, and ERK1/2 pathways. The mechanisms of HMGB1-induced neutrophil activation are distinct from endotoxin-induced signals, because HMGB1 leads to a different profile of gene expression, pattern of cytokine expression, and kinetics of p38 activation compared with LPS. These findings indicate that HMGB1 is an effective stimulus of neutrophil activation that can contribute to development of a proinflammatory phenotype in diseases characterized by excessively high levels of HMGB1.

Biological Transport↗
PubMed2003

Neural inhibition of inflammation: the cholinergic anti-inflammatory pathway.

The innate immune system is activated by infection and injury to release pro-inflammatory cytokines, which activate macrophages and neutrophils and modulate specific cellular responses. The magnitude of the cytokine response is critical, because a deficient response may result in secondary infections, while an excessive response may be more injurious than the original insult. We recently described a neural pathway, termed the "cholinergic anti-inflammatory pathway", that reflexively monitors and adjusts the inflammatory response by inhibiting pro-inflammatory cytokine synthesis. Efferent signals in the vagus nerve provide a direct mechanism for neural regulation of the immune response that is rapid, localized, and integrated. Vagus nerve stimulation inhibits the release of TNF, HMGB1, and other cytokines, and protects against endotoxemia and ischemia-reperfusion injury. This newly identified physiological mechanism of maintaining immunological homeostasis suggests that novel therapeutics may effectively modulate inflammatory responses by activating the cholinergic anti-inflammatory pathway.

Acetylcholine↗
PubMed2002

Nicotinic acetylcholine receptor alpha7 subunit is an essential regulator of inflammation.

Excessive inflammation and tumour-necrosis factor (TNF) synthesis cause morbidity and mortality in diverse human diseases including endotoxaemia, sepsis, rheumatoid arthritis and inflammatory bowel disease. Highly conserved, endogenous mechanisms normally regulate the magnitude of innate immune responses and prevent excessive inflammation. The nervous system, through the vagus nerve, can inhibit significantly and rapidly the release of macrophage TNF, and attenuate systemic inflammatory responses. This physiological mechanism, termed the 'cholinergic anti-inflammatory pathway' has major implications in immunology and in therapeutics; however, the identity of the essential macrophage acetylcholine-mediated (cholinergic) receptor that responds to vagus nerve signals was previously unknown. Here we report that the nicotinic acetylcholine receptor alpha7 subunit is required for acetylcholine inhibition of macrophage TNF release. Electrical stimulation of the vagus nerve inhibits TNF synthesis in wild-type mice, but fails to inhibit TNF synthesis in alpha7-deficient mice. Thus, the nicotinic acetylcholine receptor alpha7 subunit is essential for inhibiting cytokine synthesis by the cholinergic anti-inflammatory pathway.

Acetylcholine↗
PubMed2002

Ethyl pyruvate prevents lethality in mice with established lethal sepsis and systemic inflammation.

Sepsis, a potentially fatal clinical syndrome, is mediated by an early (e.g., tumor necrosis factor and IL-1) and late [e.g., high mobility group B-1 (HMGB1)] proinflammatory cytokine response to infection. Specifically targeting early mediators has not been effective clinically, in part because peak mediator activity often has passed before therapy can be initiated. Late-acting downstream effectors, such as HMGB1, that mediate sepsis lethality may be more relevant therapeutic targets. Ethyl pyruvate (EP) recently was identified as an experimental therapeutic that significantly protects against lethal hemorrhagic shock. Here, we report that EP attenuates lethal systemic inflammation caused by either endotoxemia or sepsis even if treatment begins after the early tumor necrosis factor response. Treatment with EP initiated 24 h after cecal puncture significantly increased survival (vehicle survival = 30% vs. EP survival = 88%, P < 0.005). EP treatment significantly reduced circulating levels of HMGB1 in animals with established endotoxemia or sepsis. In macrophage cultures, EP specifically inhibited activation of p38 mitogen-activated protein kinase and NF-kappaB, two signaling pathways that are critical for cytokine release. This report describes a new strategy to pharmacologically inhibit HMGB1 release with a small molecule that is effective at clinically achievable concentrations. EP now warrants further evaluation as an experimental "rescue" therapeutic for sepsis and other potentially fatal systemic inflammatory disorders.

Animals↗
PubMed2002

HMGB-1, a DNA-binding protein with cytokine activity, induces brain TNF and IL-6 production, and mediates anorexia and taste aversion.

High-mobility group protein-1 (HMG-1 also termed HMGB-1), a DNA-binding protein, regulates gene transcription and stabilizes nucleosome formation. HMG-1 was recently implicated as a cytokine, because it is a late-acting mediator of endotoxin lethality that induces the release of pro-inflammatory cytokines from monocytes. Here it is shown that administration of HMG-1 into the cerebral ventricles decreases food intake (food intake=4.6g/mouse in controls vs 1.6g/mouse after 1 microg HMG-1 i.c.v.; P <0.05). Intracerebroventricular HMG-1 induced an increased in TNF and IL-6 expression in the brain, and mediated taste aversion with potencies equivalent to LPS. In a model of endotoxemia, passive immunization with anti-HMG-1 antibodies attenuated the development of hypophagia, indicating that HMG-1 is a mediator of sickness behaviour associated with endotoxemia.

Animals↗
PubMed2002

Lethal weight loss: the focus shifts to signal transduction.

A hallmark of life-threatening disease in vertebrates is cachexia, a syndrome of weight loss with progressive erosion of body protein. Tumor necrosis factor (TNF) and other endogenously derived factors are sufficient to mediate the pathophysiology of cachexia in vivo, but the downstream signaling pathways have remained a mystery until recently. Tracey describes the involvement of the stress-activated protein kinase p38 and the transcriptional regulators nuclear factor kappa B and peroxisome proliferator-activated receptor gamma coactivator-1 in causing alterations in myocytes and skeletal muscle physiology. Furthermore, soluble factors including TNF and proteolysis-inducing factor may enhance protein degradation through the ubiquitin-proteosome pathway.

Animals↗
PubMed2002

Neuroprotection in cerebral ischemia by neutralization of 3-aminopropanal.

Cerebral ischemia stimulates increased activity of polyamine oxidase, a ubiquitous enzyme that catabolizes polyamines to produce 3-aminopropanal. 3-Aminopropanal is a reactive aldehyde that mediates progressive neuronal necrosis and glial apoptosis. Here we report that increased levels of 3-aminopropanal-modified protein levels in humans after aneurysmal subarachnoid hemorrhage correlate with the degree of cerebral injury as measured by admission Hunt/Hess grade. In vitro screening of clinically approved drugs reveals that N-2-mercaptopropionyl glycine (N-2-MPG), an agent clinically approved for prevention of renal stones in patients with cysteinuria, significantly inhibits the cytotoxicity of 3-aminopropanal. N-2-MPG reacts with 3-aminopropanal to yield a nontoxic thioacetal adduct, as confirmed by electrospray ionization mass spectroscopy. Administration of N-2-MPG in clinically relevant doses to rats significantly reduces cerebral 3-aminopropanal-modified protein immunoreactivity and infarct volume in a standardized model of middle cerebral artery occlusion, even when the agent is administered after the onset of ischemia. These results implicate 3-aminopropanal as a therapeutic target for cerebral ischemia.

Aldehydes↗
PubMed2002

Pharmacological stimulation of the cholinergic antiinflammatory pathway.

Efferent activity in the vagus nerve can prevent endotoxin-induced shock by attenuating tumor necrosis factor (TNF) synthesis. Termed the "cholinergic antiinflammatory pathway," inhibition of TNF synthesis is dependent on nicotinic alpha-bungarotoxin-sensitive acetylcholine receptors on macrophages. Vagus nerve firing is also stimulated by CNI-1493, a tetravalent guanylhydrazone molecule that inhibits systemic inflammation. Here, we studied the effects of pharmacological and electrical stimulation of the intact vagus nerve in adult male Lewis rats subjected to endotoxin-induced shock to determine whether intact vagus nerve signaling is required for the antiinflammatory action of CNI-1493. CNI-1493 administered via the intracerebroventricular route was 100,000-fold more effective in suppressing endotoxin-induced TNF release and shock as compared with intravenous dosing. Surgical or chemical vagotomy rendered animals sensitive to TNF release and shock, despite treatment with CNI-1493, indicating that an intact cholinergic antiinflammatory pathway is required for antiinflammatory efficacy in vivo. Electrical stimulation of either the right or left intact vagus nerve conferred significant protection against endotoxin-induced shock, and specifically attenuated serum and myocardial TNF, but not pulmonary TNF synthesis, as compared with sham-operated animals. Together, these results indicate that stimulation of the cholinergic antiinflammatory pathway by either pharmacological or electrical methods can attenuate the systemic inflammatory response to endotoxin-induced shock.

Animals↗
PubMed2002

HMGB1 B box increases the permeability of Caco-2 enterocytic monolayers and impairs intestinal barrier function in mice.

BACKGROUND & AIMS: High mobility group (HMG) B1 is a nonhistone nuclear protein that was recently identified as a late-acting mediator of lipopolysaccharide-induced lethality in mice. The proinflammatory actions of HMGB1 have been localized to a region of the molecule called the B box. METHODS: To determine whether HMGB1 or B box are capable of causing derangements in intestinal barrier function, we incubated cultured Caco-2 human enterocytic monolayers with recombinant human HMGB1 or a 74-residue truncated form of the protein consisting of the B box domain. RESULTS: Both HMGB1 and B box increased the permeability of Caco-2 monolayers to fluorescein isothiocyanate-labeled dextran (FD4) in a time- and dose-dependent fashion. The increase in permeability was reversible following removal of the recombinant protein. Exposure of Caco-2 cells to B box resulted in increased expression of inducible nitric oxide synthase messenger RNA and increased production of NO. When we used various pharmacologic strategies to inhibit NO production or scavenge NO or peroxynitrite (ONOO(-)), we abrogated B box-induced hyperpermeability. Administration of B box to wild-type mice increased both ileal mucosal permeability to FD4 and bacterial translocation to mesenteric lymph nodes. These effects were not observed in inducible nitric oxide synthase knockout mice. CONCLUSIONS: These data support the view that HMGB1 and B box are capable of causing alterations in gut barrier function via a mechanism that depends on the formation of NO and ONOO(-).

Animals↗
PubMed2002

Cholinergic antiinflammatory pathway inhibition of tumor necrosis factor during ischemia reperfusion.

OBJECTIVE: Ischemia-reperfusion injury is a pathologic event characterized by tissue damage. It is mediated by tumor necrosis factor (TNF) and other cytokines that activate complement and proteases and stimulate fibrinolysis, degranulation of white blood cells, and free radical production. We recently reported that vagus nerve stimulation (VNS) suppresses endotoxin-induced cytokine synthesis through alpha bungarotoxin-sensitive cholinergic receptors. VNS protects against endotoxin-induced shock by inhibiting hepatic and cardiac synthesis of TNF. Here, the effects of VNS on suppression of ischemia-reperfusion injury and cytokine release were studied in a rat model of aortic occlusion. METHODS: Adult male Lewis rats were subjected to laparotomy and suprarenal aortic clamping for 15 minutes followed by reperfusion. Blood pressure, heart rate, and respiratory rate were recorded every 3 minutes for 90 minutes. Exposed cervical vagus nerves in the experimental group were stimulated for 5 minutes before and after aortic occlusion, with constant voltage (1 V, 2 ms, 5 Hz); sham-operated animals received no stimulation. TNF levels in serum and organs were measured with enzyme-linked immunosorbent assay ELISA (BioSource International, Camarillo, Calif). Data analysis was performed with the Student t test. RESULTS: Control animals had shock develop (mean, 59% decrease in blood pressure) whereas blood pressure in the stimulated animals did not decrease (control versus stimulated animals, P <.05). VNS significantly inhibited TNF levels in serum (7 +/- 1 ng/mL versus 45 +/- 6 ng/mL; P =.0008), heart (21 +/- 11 ng/g protein versus 85 +/- 15 ng/g protein; P =.01), and liver (16 +/- 2 ng/g protein versus 42 +/- 12 ng/g protein; P =.02). CONCLUSION: VNS significantly attenuates TNF synthesis and shock during reperfusion injury in a standard model of aortic occlusion. Clinical evaluation of VNS for this condition may be warranted.

Animals↗
PubMed2002

Globin attenuates the innate immune response to endotoxin.

Hemoglobin is an endotoxin (lipopolysaccharide; LPS)-binding protein that synergistically increases the release of proinflammatory cytokines from the innate immune system in response to LPS. It has been suggested that this activity of hemoglobin facilitates the recognition of Gram-negative bacteria in a wound, thereby maximizing immune efficiency. This synergy may be important to the pathogenesis of a broad spectrum of clinical conditions because elevated hemoglobin levels frequently are observed in patients after the transfusion of red cells, trauma, cardiopulmonary bypass surgery, hemolysis, in addition to other disorders. To determine the molecular basis of the specific hemoglobin-LPS synergy, in this article we tested the effects of globin itself on macrophage responses to LPS. Paradoxically, these studies revealed that globin suppressed tumor necrosis factor (TNF) synthesis in LPS-stimulated murine and human macrophage cultures. LPS comigrated with globin on non-denaturing electrophoretic gels, giving direct evidence for binding. Globin specifically inhibited LPS activity in the standard Limulus assay but did not inhibit interleukin-1beta-mediated TNF synthesis. Iron supplementation of macrophage cultures significantly increased interleukin-1beta-induced TNF release. Intraperitoneal administration of globin protected mice against both LPS-induced lethality and experimentally induced bacterial infection. Thus, the heme-iron moiety of hemoglobin, and not the binding of LPS to globin, enhanced macrophage responses to LPS.

Animals↗
PubMed2002

HMGB1 as a cytokine and therapeutic target.

HMGB1 is an abundant nuclear and cytoplasmic protein present in mammalian cells. It is traditionally known as a DNA binding protein involved in maintenance of nucleosome structure and regulation of gene transcription. Beyond these intracellular roles, we recently discovered that HMGB1 is released from activated macrophages and functions as a late mediator of lethal endotoxemia. Addition of HMGB1 to macrophage cultures activates cytokine release. When released into the extracellular milieu, HMGB1 causes systemic inflammatory responses including acute lung injury, epithelial barrier dysfunction, and death. Passive immunization with anti-HMGB1 antibodies confers significant protection against lethality induced by LPS administration and sepsis caused by cecal perforation in mice. Truncation of HMGB1 into individual structural domains revealed that the HMGB1 A box, a DNA-binding motif, specifically antagonizes the activity of HMGB1 and rescues mice from lethal sepsis caused by cecal perforation. Thus, strategies that target HMGB1 with specific antibodies or antagonists have potential for treating lethal systemic inflammatory diseases characterized by excessive HMGB1 release.

Antibodies↗
PubMed2002

HMGB1 as a DNA-binding cytokine.

HMGB1 (high mobility group box chromosomal protein 1), historically known as an abundant, nonhistone architectural chromosomal protein, is extremely conserved across species. As a nuclear protein, HMGB1 stabilizes nucleosomes and allows bending of DNA that facilitates gene transcription. Unexpectedly, recent studies identified extracellular HMGB1 as a potent macrophage-activating factor, signaling via the receptor for advanced glycation end-products to induce inflammatory responses. It is released as a late mediator during inflammation and participates in the pathogenesis of systemic inflammation after the early mediator response has resolved. HMGB1 occupies a critical role as a proinflammatory mediator passively released by necrotic but not apoptotic cells. Necrotic Hmgb1(-/-) cells mediate minimal inflammatory responses. Stimulated macrophages actively secrete HMGB1 to promote inflammation and in turn, stimulate production of multiple, proinflammatory cytokines. HMGB1 mediates endotoxin lethality, acute lung injury, arthritis induction, activation of macrophages, smooth muscle cell chemotaxis, and epithelial cell barrier dysfunction. HMGB1 is structurally composed of three different domains: two homologous DNA-binding sequences entitled box A and box B and a highly, negatively charged C terminus. The B box domain contains the proinflammatory cytokine functionality of the molecule, whereas the A box region has an antagonistic, anti-inflammatory effect with therapeutic potential. Administration of highly purified, recombinant A box protein or neutralizing antibodies against HMGB1 rescued mice from lethal sepsis, even when initial treatment was delayed for 24 h after the onset of infection, establishing a clinically relevant therapeutic window that is significantly wider than for other known cytokines.

Animals↗
PubMed2001

A new model of sciatic inflammatory neuritis (SIN): induction of unilateral and bilateral mechanical allodynia following acute unilateral peri-sciatic immune activation in rats.

Immune activation near healthy peripheral nerves may have a greater role in creating pathological pain than previously recognized. We have developed a new model of sciatic inflammatory neuritis to assess how such immune activation may influence somatosensory processing. The present series of experiments reveal that zymosan (yeast cell walls) acutely injected around the sciatic nerve of awake unrestrained rats rapidly (within 3h) produces low threshold mechanical allodynia in the absence of thermal hyperalgesia. Low (4 microg) doses of zymosan produce both territorial and extra-territorial allodynia restricted to the ipsilateral hindpaw. Higher (40-400 microg) doses of zymosan again produce both territorial and extra-territorial allodynia. However, allodynia is now expressed both in the ipsilateral as well as contralateral hindpaws. Several lines of evidence are provided that the appearance of this contralateral ('mirror') allodynia reflects local actions of zymosan on the sciatic nerve rather than spread of this immune activator to the general circulation. Since many clinical neuropathies result from inflammation/infection of peripheral nerves rather than frank physical trauma, understanding how immune activation alters pain processing may suggest novel approaches to pain control.

Acute Disease↗
PubMedDate not supplied

The inflammatory reflex.

Inflammation is a local, protective response to microbial invasion or injury. It must be fine-tuned and regulated precisely, because deficiencies or excesses of the inflammatory response cause morbidity and shorten lifespan. The discovery that cholinergic neurons inhibit acute inflammation has qualitatively expanded our understanding of how the nervous system modulates immune responses. The nervous system reflexively regulates the inflammatory response in real time, just as it controls heart rate and other vital functions. The opportunity now exists to apply this insight to the treatment of inflammation through selective and reversible 'hard-wired' neural systems.

Animals↗
PubMedDate not supplied

Anti-HMGB1 neutralizing antibody ameliorates gut barrier dysfunction and improves survival after hemorrhagic shock.

Intestinal barrier dysfunction occurs following hemorrhagic shock and resuscitation (HS/R). High-mobility group B1 (HMGB1) has been shown to increase the permeability of Caco-2 human enterocyte-like epithelial monolayers in vitro. In this study, we found that serum concentrations of HMGB1 were higher in blood samples obtained from 25 trauma victims with hemorrhagic shock than in 9 normal volunteers. We also studied whether treatment with anti-HMGB1 antibody can ameliorate HS/R-induced gut barrier dysfunction in mice. Animals were shocked by withdrawal of blood to maintain mean arterial pressure at 25 to 30 mmHg for 2 h. After resuscitation with shed blood plus Ringer's lactate solution, the mice were treated with either anti-HMGB1 antibody or nonimmune rabbit IgG. Serum HMGB1 concentrations were significantly higher in trauma victims than control mice. Treatment with anti-HMGB1 antibody improved survival at 24 h and ameliorated the development of ileal mucosal hyperpermeability to FITC-labeled dextran. At 24 h after HS/R, treatment with anti-HMGB1 antibody decreased bacterial translocation to mesenteric lymph nodes and was associated with lower circulating concentrations of IL-6 and IL-10. These data support the notion that HMGB1 is a mediator of HS/R-induced gut barrier dysfunction and suggest that anti-HMGB1 antibodies warrant further evaluation as a therapeutic to ameliorate the morbidity of HS/R in trauma patients.

Adolescent↗
PubMedDate not supplied

Structural basis for the proinflammatory cytokine activity of high mobility group box 1.

High mobility group box 1 (HMGB), a ubiquitous DNA-binding protein, has been implicated as a proinflammatory cytokine and late mediator of lethal endotoxemia. HMGB1 is released by activated macrophages. It amplifies and extends the inflammatory response by inducing cytokine release and mediating acute lung injury, anorexia, and the inflammatory response to tissue necrosis. The kinetics of HMGB1 release provide a wide therapeutic window for endotoxemia because extracellular levels of HMGB1 begin to increase 12 to 24 h after exposure to inflammatory stimuli. Here, we demonstrate that a DNA-binding domain of HMGB1, the B box, recapitulates the cytokine activity of full length HMGB1 and efficiently activates macrophages to release tumor necrosis factor (TNF) and other proinflammatory cytokines. Truncation of the B box revealed that the TNF-stimulating activity localizes to 20 amino acids (HMGB1 amino acids 89 to 108). Passive immunization of mice with antibodies raised against B box conferred significant protection against lethal endotoxemia or sepsis, induced by cecal perforation. These results indicate that a proinflammatory domain of HMGB1 maps to the highly conserved DNA-binding B box, making this primary sequence a suitable target in the design of therapeutics.

Animals↗