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Gregory P Downey

Publications and source records attributed to Gregory P Downey.

At least 19 recordsLinked to original sources
PubMed2006

Molecular pathogenesis of lymphangioleiomyomatosis: lessons learned from orphans.

Lymphangioleiomyomatosis (LAM) is a rare progressive cystic lung disease affecting young women. The pivotal observation that LAM occurs both spontaneously and as part of the tuberous sclerosis complex (TSC) led to the hypothesis that these disorders share common genetic and pathogenetic mechanisms. In this review we describe the evolution of our understanding of the molecular and cellular basis of LAM and TSC, beginning with the discovery of the TSC1 and TSC2 genes and the demonstration of their involvement in sporadic (non-TSC) LAM. This was followed by rapid delineation of the signaling pathways in Drosophila melanogaster with confirmation in mice and humans. This knowledge served as the foundation for novel therapeutic approaches that are currently being used in human clinical trials.

Animals↗
PubMed2006

Reactive oxygen and nitrogen species as signaling molecules regulating neutrophil function.

As a cornerstone of the innate immune response, neutrophils are the archetypical phagocytic cell; they actively seek out, ingest, and destroy pathogenic microorganisms. To achieve this essential role in host defense, neutrophils deploy a potent antimicrobial arsenal that includes oxidants, proteinases, and antimicrobial peptides. Importantly, oxidants produced by neutrophils, referred to in this article as reactive oxygen (ROS) and reactive nitrogen (RNS) species, have a dual function. On one hand they function as potent antimicrobial agents by virtue of their ability to kill microbial pathogens directly. On the other hand, they participate as signaling molecules that regulate diverse physiological signaling pathways in neutrophils. In the latter role, ROS and RNS serve as modulators of protein and lipid kinases and phosphatases, membrane receptors, ion channels, and transcription factors, including NF-kappaB. The latter regulates expression of key cytokines and chemokines that further modulate the inflammatory response. During the inflammatory response, ROS and RNS modulate phagocytosis, secretion, gene expression, and apoptosis. Under pathological circumstances such as acute lung injury and sepsis, excess production of ROS may influence vicinal cells such as endothelium or epithelium, contributing to inflammatory tissue injury. A better understanding of these pathways will help identify novel targets for amelioration of the untoward effects of inflammation.

Animals↗
PubMed2006

Air travel in women with lymphangioleiomyomatosis.

BACKGROUND AND OBJECTIVE: The safety of air travel in patients with pneumothorax-prone pulmonary diseases, such as lymphangioleiomyomatosis (LAM), has not been studied to any great extent. A questionnaire-based evaluation of air travel in patients with LAM was conducted to determine experiences aboard commercial aircraft. METHODS: A survey was sent to women listed in the US LAM Foundation registry (n = 389) and the UK LAM Action registry (n = 59) to assess air travel, including problems occurring during flight. Women reporting a pneumothorax in flight were followed up to ascertain further details about the incident. RESULTS: 327 (73%) women completed the survey. 308 women answered the travel section, of whom 276 (90%) had "ever" travelled by aeroplane for a total of 454 flights. 95 (35%) women had been advised by their doctor to avoid air travel. Adverse events reported included shortness of breath (14%), pneumothorax (2%, 8/10 confirmed by chest radiograph), nausea or dizziness (8%), chest pain (12%), unusual fatigue (11%), oxygen desaturation (8%), headache (9%), blue hands (2%), haemoptysis (0.4%) and anxiety (22%). 5 of 10 patients with pneumothorax had symptoms that began before the flight: 2 occurred during cruising altitude, 2 soon after landing and 1 not known. The main symptoms were severe chest pain and shortness of breath. DISCUSSION AND CONCLUSION: Adverse effects occurred during air travel in patients with LAM, particularly dyspnoea and chest pain. Hypoxaemia and pneumothorax were reported. The decision to travel should be individualised; patients with unexplained shortness of breath or chest pain before scheduled flights should not board. Patients with borderline oxygen saturations on the ground should be evaluated for supplemental oxygen therapy during flight. Although many women had been advised not to travel by air, most travelled without the occurrence of serious adverse effects.

Adult↗
PubMed2006

Phosphorylation of SHP-2 regulates interactions between the endoplasmic reticulum and focal adhesions to restrict interleukin-1-induced Ca2+ signaling.

Interleukin-1 (IL-1)-induced Ca2+ signaling in fibroblasts is constrained by focal adhesions. This process involves the proteintyrosine phosphatase SHP-2, which is critical for IL-1-induced phosphorylation of phospholipase Cgamma1, thereby enhancing IL-1-induced Ca2+ release and ERK activation. Currently, the mechanisms by which SHP-2 modulates Ca2+ release from the endoplasmic reticulum are not defined. We used immunoprecipitation and fluorescence protein-tagged SHP-2 or endoplasmic reticulum (ER)-protein expression vectors, and an ER-specific calcium indicator, to examine the functional relationships between SHP-2, focal adhesions, and IL-1-induced Ca2+ release from the ER. By total internal reflection fluorescence microscopy to image subplasma membrane compartments, SHP-2 co-localized with the ER-associated proteins calnexin and calreticulin at sites of focal adhesion formation in fibroblasts. IL-1beta promoted time-dependent recruitment of SHP-2 and ER proteins to focal adhesions; this process was blocked in cells treated with small interfering RNA for SHP-2 and in cells expressing a Y542F SHP-2 mutant. IL-1 stimulated inositol 1,4,5-trisphosphate receptor-mediated Ca2+ release from the ER subjacent to the plasma membrane that was tightly localized around fibronectin-coated beads and was reduced 4-fold in cells expressing Tyr-542 SHP-2 mutant. In subcellular fractions enriched for ER proteins, immunoprecipitation demonstrated that IL-1-enhanced association of SHP-2 with the type 1 inositol 1,4,5-trisphosphate receptor was dependent on Tyr-542 of SHP-2. We conclude that Tyr-542 of SHP-2 modulates IL-1-induced Ca2+ signals and association of the ER with focal adhesions.

Animals↗
PubMed2006

Cystic fibrosis mouse models.

Animal models of cystic fibrosis (CF) are powerful tools that enable the study of the mechanisms and complexities of human disease. Murine models have several intrinsic advantages compared with other animal models, including lower cost, maintenance, and rapid reproduction rate. Mice can be easily genetically manipulated by making transgenic or knockout mice, or by backcrossing to well-defined inbred strains in a reasonably short period of time. However, anatomic and immunologic differences between mice and humans mean that murine models have inherent limitations that must be considered when interpreting the results obtained from experimental models and applying these to the pathogenesis of CF disease in humans. This review will focus on the different CF mouse models available that represent diverse phenotypes observed in humans with CF and that can help researchers elucidate the diverse functions of the CFTR protein.

Animals↗
PubMed2006

Death of the septic monocyte: is more better?

Apoptosis is of pivotal importance in the pathogenesis of sepsis. Depending on the cell type involved and the time point of the disease process, apoptosis may be linked to either a good or a bad outcome. Work presented in this issue by Giamarellos-Bourboulis and coworkers suggests that an early increase in the apoptosis of blood monocytes is associated with improved survival in patients with varying degrees of sepsis. Although the mechanism by which monocyte apoptosis influences the outcome of sepsis cannot be determined by this study, these observations represent an important advance in our understanding of this complicated disease process.

Apoptosis↗
PubMed2006

CD44 is a phagocytic receptor.

CD44, a transmembrane adhesion molecule involved in binding and metabolism of hyaluronan, has additional functions in inflammatory and immune responses, contributing to the ingestion and clearance of particles and apoptotic cells. Our goal was to determine the specific role of CD44 in phagocytosis and whether it functions as a primary or accessory phagocytic receptor. Using hyaluronan-coated beads and erythrocytes coated with antiCD44 antibodies as the phagocytic prey, we determined that CD44 mediates efficient phagocytosis in primary murine peritoneal macrophages and in the murine macrophage cell line RAW 264.7. In RAW cells, the phagocytic index for anti-CD44-coated erythrocytes was 25 +/- 3 (mean +/- SEM) compared with less than 1 for erythrocytes coated with isotype-matched control antibodies. Uptake of anti-CD44-coated erythrocytes was abrogated by pretreatment with a blocking antibody to CD44 and was absent in primary cultures of CD44-deficient murine macrophages. Down-regulation of Fc receptors by aggregated IgG-induced internalization, which blocks uptake of IgG-coated particles, had no effect on CD44-mediated particle engulfment. Using a combination of immunoprecipitation, pharmacologic inhibition, and genetic deletion, we determined that CD44-mediated phagocytosis involves Syk, Rac1, and phosphatidylinositol 3-kinase and induced activation of the phagocyte oxidase. We conclude that CD44 is a competent phagocytic receptor that efficiently mediates internalization of large particles.

Animals↗
PubMed2006

Tyrosine phosphatase SHP-2 regulates IL-1 signaling in fibroblasts through focal adhesions.

Interleukin-1beta (IL-1beta) mediates destruction of matrix collagens in diverse inflammatory diseases including arthritis, periodontitis, and pulmonary fibrosis by activating fibroblasts, cells that interact with matrix proteins through integrin-based adhesions. In vitro, IL-1beta signaling is modulated by focal adhesions, supramolecular protein complexes that are enriched with tyrosine kinases and phosphatases. We assessed the importance of tyrosine phosphatases in regulating cell-matrix interactions and IL-1beta signaling. In human gingival fibroblasts plated on fibronectin, IL-1beta enhanced the maturation of focal adhesions as defined by morphology and enrichment with paxillin and alpha-actinin. IL-1beta also induced activation of ERK and recruitment of phospho-ERK to focal complexes/adhesions. Treatment with the potent tyrosine phosphatase inhibitor pervanadate, in the absence of IL-1beta, recapitulated many of these responses indicating the importance of tyrosine phosphatases. Immunoblotting of collagen bead-associated complexes revealed that the tyrosine phosphatase, SHP-2, was also enriched in focal complexes/adhesions. Depletion of SHP-2 by siRNA or by homologous recombination markedly altered IL-1beta-induced ERK activation and maturation of focal adhesions. IL-1beta-induced tyrosine phosphorylation of SHP-2 on residue Y542 promoted focal adhesion maturation. Association of Gab1 with SHP-2 in focal adhesions correlated temporally with activation of ERK and was abrogated in cells expressing mutant (Y542F) SHP-2. We conclude that IL-1beta mediated maturation of focal adhesions is dependent on tyrosine phosphorylation of SHP-2 at Y542, leading to recruitment of Gab1, a process that may influence the downstream activation of ERK.

Actinin↗
PubMed2006

Signalling platforms that modulate the inflammatory response: new targets for drug development.

Therapeutically controlling inflammation is essential for the clinical management of many high-prevalence human diseases. Drugs that block the pro-inflammatory cytokines tumour-necrosis factor-alpha and interleukin-1 (IL-1) can improve outcomes for rheumatoid arthritis and other inflammatory diseases but many patients remain refractory to treatment. Here we explore the need for developing new types of anti-inflammatory drugs and the emergence of novel drug targets based on the clustering of IL-1 receptors into multi-protein aggregates associated with cell adhesions. Interference with receptor aggregation into multi-protein complexes effectively abrogates IL-1 signalling. The exploration of the crucial molecules required for receptor clustering, and therefore signal transduction, offers new targets and scope for anti-inflammatory drug development.

Anti-Inflammatory Agents↗
PubMed2006

Up-regulation of functional CXCR4 expression on human lymphocytes in sepsis.

OBJECTIVE: Lymphocyte dysfunction has been documented in sepsis, and evidence suggests that lymphocyte infiltration contributes to tissue injury. The purpose of this study was to examine chemokine receptor expression and function in lymphocytes from septic patients and healthy donors. DESIGN: Observational study of septic patients and laboratory investigation of normal controls. SETTING: Tertiary care intensive care unit. PATIENTS AND SUBJECTS: Nine critically ill patients fulfilling criteria for the systemic inflammatory response syndrome and with a Sepsis Score of >/=3 were included in this study. Lymphocytes were also obtained from healthy volunteers. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The total number of circulating leukocytes in septic patients was markedly increased; however, lymphocyte counts were decreased. Chemokine receptor expression on lymphocytes was examined by flow cytometry. CXCR4 expression on lymphocytes from septic patients was increased whereas CCR5 was decreased and CCR7 was unchanged. Lipopolysaccharide stimulation of normal lymphocytes increased CXCR4 expression but decreased CCR5 and did not change CCR7 expression. This lipopolysaccharide-stimulated CXCR4 expression required 20 hrs of stimulation and was accompanied by increased messenger RNA. Lymphocytes from septic patients or after lipopolysaccharide treatment demonstrated enhanced actin polymerization and migration in response to CXCL12. Taken together, sepsis and lipopolysaccharide up-regulated CXCR4 expression and enhanced lymphocyte activation and migration in response to CXCL12. CONCLUSIONS: Blocking CXCR4 and CXCL12 function may provide a novel therapeutic method for controlling systemic inflammation and tissue injury in sepsis.

Actins↗
PubMed2006

Neutrophil granule contents in the pathogenesis of lung injury.

PURPOSE OF REVIEW: This review summarizes recent literature on the role of neutrophil granule contents in acute lung injury and the mechanisms by which these contribute to inflammatory tissue injury. RECENT FINDINGS: Neutrophil products such as elastase, reactive oxygen species, and antimicrobial peptides can alter pulmonary cell function in a nondegradative fashion through activation of cell surface receptors or modulation of signal transduction pathways. These effects can be either beneficial or detrimental to the host. SUMMARY: The primary function of neutrophils in the innate immune response--to contain and kill invading microbial pathogens--is achieved through a series of rapid and coordinated responses culminating in phagocytosis and intracellular killing of the pathogens. Neutrophils have a potent antimicrobial arsenal that includes oxidants, proteinases, and cationic peptides. Reactive oxygen species such as oxygen are produced by the phagocyte NADPH oxidase and are microbicidal. Granules within the neutrophil cytoplasm contain potent proteolytic enzymes and cationic proteins that can digest a variety of microbial substrates. These compounds are released directly into the phagosome, compartmentalizing both the pathogen and the cytotoxic products. Under pathological circumstances, however, unregulated release of microbicidal compounds into the extracellular space can paradoxically damage host tissues. Nonspecific inhibition of neutrophils is not clinically realistic, as it would leave the host vulnerable to infection. As the mechanisms of action of neutrophil granule contents are elucidated, therapeutic targets will be identified that will allow for suppression of neutrophils' detrimental effects while avoiding inhibition of their beneficial effects.

Animals↗
PubMed2006

Murine hepatitis virus strain 1 produces a clinically relevant model of severe acute respiratory syndrome in A/J mice.

Severe acute respiratory syndrome (SARS) is a life-threatening infectious disease which has been difficult to study and treat because of the lack of a readily available animal model. Intranasal infection of A/J mice with the coronavirus murine hepatitis virus strain 1 (MHV-1) produced pulmonary pathological features of SARS. All MHV-1-infected A/J mice developed progressive interstitial pneumonitis, including dense macrophage infiltrates, giant cells, and hyaline membranes, resulting in death of all animals. In contrast, other mouse strains developed only mild transitory disease. Infected A/J mice had significantly higher cytokine levels, particularly macrophage chemoattractant protein 1 (MCP-1/CCL-2), gamma interferon, and tumor necrosis factor alpha. Furthermore, FGL2/fibroleukin mRNA transcripts and protein and fibrin deposits were markedly increased in the lungs of infected A/J mice. These animals developed a less robust type I interferon response to MHV-1 infection than resistant C57BL/6J mice, and treatment with recombinant beta interferon improved survival. This study describes a potentially useful small animal model of human SARS, defines its pathogenesis, and suggests treatment strategies.

Animals↗
PubMed2006

Rare lung diseases I--Lymphangioleiomyomatosis.

The present article is the first in a series that will review selected rare lung diseases. The objective of this series is to promote a greater understanding and awareness of these unusual conditions among respirologists. Each article will begin with a case that serves as a focal point for a discussion of the pathophysiology and management of the particular condition. The first article is on lymphangioleiomyomatosis (LAM); subsequent articles will focus on pulmonary alveolar proteinosis, alpha-1-antitrypsin deficiency and primary ciliary dyskinesia. LAM is a rare, progressive and (without intervention) often fatal interstitial lung disease that predominantly affects women of childbearing age. LAM is characterized by progressive interstitial infiltration of the lung by smooth muscle cells, resulting in diffuse cystic changes of the lung parenchyma. The molecular basis of this disorder has been delineated over the past five years and LAM is now known to be a consequence of mutations in the tuberous sclerosis genes. This knowledge, combined with advances in our understanding of the signalling pathways regulated by these genes, has given rise to potential molecular therapies that hold great promise for treating this devastating disease.

Comorbidity↗
PubMed2006

Neutrophil apoptosis: a marker of disease severity in sepsis and sepsis-induced acute respiratory distress syndrome.

INTRODUCTION: Apoptosis of neutrophils (polymorphonuclear neutrophils [PMNs]) may limit inflammatory injury in sepsis and acute respiratory distress syndrome (ARDS), but the relationship between the severity of sepsis and extent of PMN apoptosis and the effect of superimposed ARDS is unknown. The objective of this study was to correlate neutrophil apoptosis with the severity of sepsis and sepsis-induced ARDS. METHODS: A prospective cohort study was conducted in intensive care units of three tertiary hospitals in Porto Alegre, southern Brazil. Fifty-seven patients with sepsis (uncomplicated sepsis, septic shock, and sepsis-induced ARDS) and 64 controls were enrolled. Venous peripheral blood was collected from patients with sepsis within 24 hours of diagnosis. All surgical groups, including controls, had their blood drawn 24 hours after surgery. Control patients on mechanical ventilation had blood collected within 24 hours of initiation of mechanical ventilation. Healthy controls were blood donors. Neutrophils were isolated, and incubated ex vivo, and apoptosis was determined by light microscopy on cytospun preparations. The differences among groups were assessed by analysis of variance with Tukeys. RESULTS: In medical patients, the mean percentage of neutrophil apoptosis (+/- standard error of the mean [SEM]) was lower in sepsis-induced ARDS (28% +/- 3.3%; n = 9) when compared with uncomplicated sepsis (57% +/- 3.2%; n = 8; p < 0.001), mechanical ventilation without infection, sepsis, or ARDS (53% +/- 3.0%; n = 11; p < 0.001) and healthy controls (69% +/- 1.1%; n = 33; p < 0.001) but did not differ from septic shock (38% +/- 3.7%; n = 12; p = 0.13). In surgical patients with sepsis, the percentage of neutrophil apoptosis was lower for all groups when compared with surgical controls (52% +/- 3.6%; n = 11; p < 0.001). CONCLUSION: In medical patients with sepsis, neutrophil apoptosis is inversely proportional to the severity of sepsis and thus may be a marker of the severity of sepsis in this population.

Adult↗
PubMed2005

Tyrosine phosphatase MEG2 modulates murine development and platelet and lymphocyte activation through secretory vesicle function.

MEG2, a protein tyrosine phosphatase with a unique NH2-terminal lipid-binding domain, binds to and is modulated by the polyphosphoinositides PI(4,5)P2 and PI(3,4,5)P3. Recent data implicate MEG2 in vesicle fusion events in leukocytes. Through the genesis of Meg2-deficient mice, we demonstrate that Meg2-/- embryos manifest hemorrhages, neural tube defects including exencephaly and meningomyeloceles, cerebral infarctions, abnormal bone development, and >90% late embryonic lethality. T lymphocytes and platelets isolated from recombination activating gene 2-/- mice transplanted with Meg2-/- embryonic liver-derived hematopoietic progenitor cells showed profound defects in activation that, in T lymphocytes, was attributable to impaired interleukin 2 secretion. Ultrastructural analysis of these lymphocytes revealed near complete absence of mature secretory vesicles. Taken together, these observations suggest that MEG2-mediated modulation of secretory vesicle genesis and function plays an essential role in neural tube, vascular, and bone development as well as activation of mature platelets and lymphocytes.

Animals↗
PubMed2005

Cyclic nucleotides modulate genioglossus and hypoglossal responses to excitatory inputs in rats.

RATIONALE: Previous studies modulating pharyngeal muscle activity with pharmacologic approaches have targeted membrane receptors on pharyngeal motoneurons. Whether modulation of intracellular pathways can increase pharyngeal muscle activity, however, has not been investigated but is relevant to pharmacologic treatments of obstructive sleep apnea. OBJECTIVES: To determine if modulating the second messenger cyclic adenosine-3'-5'-monophosphate (cAMP) at the hypoglossal motor nucleus (HMN) will increase genioglossus activity across sleep- wake states. METHODS: Forty-eight rats were implanted with electroencephalogram and neck electrodes to record sleep-wake states and genioglossus and diaphragm electrodes for respiratory muscle recordings. Microdialysis probes were inserted into the HMN to perfuse artificial cerebrospinal fluid and (1) forskolin (500 microM, adenylyl cyclase activator to increase cAMP), (2) a cAMP analog (500 microM), (3) iso-butyl-methylxanthine (IBMX; 300 microM, phosphodiesterase inhibitor), or (4) a cyclic guanosine-3'-5'-monophosphate (cGMP) analog (500 microM, 8-Br-cGMP). MEASUREMENTS AND MAIN RESULTS: Forskolin and the cAMP analog at the HMN increased respiratory-related and tonic genioglossus activities in wakefulness and non-REM sleep but not REM sleep. IBMX did not affect genioglossus activity in awake or sleeping rats. However, IBMX abolished the robust excitatory responses to serotonin and phenylephrine at the HMN, but responses to non-N-methyl-D-aspartate receptor activation remained. These effects of IBMX were mimicked by 8-Br-cGMP. CONCLUSIONS: Genioglossus responses to manipulation of cAMP at the HMN are differentially modulated by sleep-wake state. Selective abolition of serotonin and phenylephrine responses after IBMX suggests that under conditions of nonspecific phosphodiesterase inhibition the HMN is unresponsive to certain, otherwise potent, excitatory inputs. Similar responses with 8-Br-cGMP suggest this effect is likely mediated by cGMP pathways.

Animals↗
PubMed2005

Human neutrophil peptides induce interleukin-8 production through the P2Y6 signaling pathway.

Antimicrobial human neutrophil peptides (HNPs) play a pivotal role in innate host defense against a broad spectrum of prokaryotic pathogens. In addition, HNPs modulate cellular immune responses by producing the chemokine interleukin-8 (IL-8) in myeloid and epithelial cells and by exerting chemotaxis to T cells, immature dendritic cells, and monocytes. However, the mechanisms by which HNPs modulate the immune responses in the eukaryotic cells remain unclear. We demonstrated that, as with adenosine triphosphate (ATP) and uridine diphosphate (UDP), HNP stimulation of human lung epithelial cells selectively induced IL-8 production in 10 pro- and anti-inflammatory cytokines examined. HNP-induced IL-8 release was inhibited by treatment with the nucleotide receptor antagonists suramin and reactive blue. Transfection of lung epithelial cells with antisense oligonucleotides targeting specific purinergic P2Y receptors revealed that the P2Y6 (ligand of UDP) signaling pathway plays a predominant role in mediating HNP-induced IL-8 production.

Base Sequence↗