Short-chain fatty acids induce tissue plasminogen activator in airway epithelial cells via GPR41&43.

BACKGROUND: Chronic rhinosinusitis (CRS) is a heterogeneous chronic inflammatory disease generally divided based on the presence or absence of nasal polyps (NPs). One of the features of NPs is excessive fibrin deposition, which is associated with down-regulation of tissue plasminogen activator (t-PA) in NPs. As t-PA is expressed in epithelial cells, and epithelium is readily accessible to topical therapies, identifying compounds that can mediate the induction of t-PA would be a potential new strategy for the treatment of NPs. OBJECTIVE: The objective of this study was to determine whether short-chain fatty acids (SCFAs) can induce t-PA in airway epithelial cells via their known receptors GPR41 and GPR43. METHODS: We performed immunohistochemistry (IHC) to determine whether receptors for SCFAs, known as G protein-coupled receptor 41/free fatty acid receptor 3 (GPR41/FFAR3) and GPR43/FFAR2, are expressed in nasal tissue. Primary normal human bronchial epithelial (NHBE) cells were stimulated with different concentrations of SCFAs to test induction of t-PA, which was analysed by expression of mRNA and protein. Mediation of responses by SCFA receptors was evaluated by specific receptor gene silencing with siRNA. RESULTS: Immunohistochemistry study revealed that airway epithelial cells expressed GPR41 and GPR43. Acetic acid, propionic acid, butyric acid and valeric acid significantly induced t-PA expression from two- to tenfolds. The strongest inducer of t-PA from NHBE cells was propionic acid; cells stimulated with propionic acid released t-PA into the supernatant in its active form. Gene silencing of GPR41 and GPR43 revealed that induction of t-PA by SCFAs was dependent upon both GPR41 and GPR43. CONCLUSIONS AND CLINICAL RELEVANCE: Short-chain fatty acids were shown to induce airway epithelial cell expression of t-PA via GPR41 and GPR43. Topical delivery of potent compounds that activate these receptors may have value by reducing fibrin deposition and shrinking nasal polyp growth.

Chronic airway inflammation provides a unique environment for B cell activation and antibody production.

BACKGROUND: B cells play many roles in health and disease. However, little is known about the mechanisms that drive B cell responses in the airways, especially in humans. Chronic rhinosinusitis (CRS) is an inflammatory disease of the upper airways that affects 10% of Europeans and Americans. A subset of CRS patients develop nasal polyps (NPs), which are characterized by type 2 inflammation, eosinophils and group 2 innate lymphoid cells (ILC2s). We have reported that NP contain elevated levels of B cells and antibodies, making NP an ideal system for studying B cells in the airways. OBJECTIVE: We sought to determine the mechanisms that drive B cell activation and antibody production during chronic airway inflammation. METHODS: We analysed B cells from NP or tonsil, or after ILC2 coculture, by flow cytometry. Antibody production from tissue was measured using Luminex assays and the frequency of antibody-secreting cells by ELISpot. Formation of B cell clusters was assessed using immunohistochemistry. Expression of genes associated with B cell activation and class switch recombination was measured by qRT-PCR. RESULTS: NP contained significantly elevated frequencies of plasmablasts, especially those that expressed the extrafollicular marker Epstein-Barr virus-induced protein 2 (EBI2), but significantly fewer germinal centre (GC) B cells compared with tonsil. Antibody production and the frequency of antibody-secreting cells were significantly elevated in NP, and there was evidence for local class switch recombination in NP. Finally, ILC2s directly induced EBI2 expression on B cells in vitro. CONCLUSIONS AND CLINICAL RELEVANCE: Our data suggest there is a unique B cell activation environment within NP that is distinct from classic GC-mediated mechanisms. We show for the first time that ILC2s directly induce EBI2 expression on B cells, indicating that ILC2s may play an important role in B cell responses. B cell-targeted therapies may provide new treatment options for CRSwNP.

Elevated presence of myeloid dendritic cells in nasal polyps of patients with chronic rhinosinusitis.

BACKGROUND: Although chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by Th2 inflammation, the mechanism underlying the onset and amplification of this inflammation has not been fully elucidated. Dendritic cells (DCs) are major antigen-presenting cells, central inducers of adaptive immunity and critical regulators of many inflammatory diseases. However, the presence of DCs in CRS, especially in nasal polyps (NPs), has not been extensively studied. OBJECTIVE: The objective of this study was to characterize DC subsets in CRS. METHODS: We used real-time PCR to assess the expression of mRNA for markers of myeloid DCs (mDCs; CD1c), plasmacytoid DCs (pDCs; CD303) and Langerhans cells (LCs; CD1a, CD207) in uncinate tissue (UT) from controls and patients with CRS as well as in NP. We assayed the presence of DCs by immunohistochemistry and flow cytometry. RESULTS: Compared to UT from control subjects (n = 15) and patients with CRS without NP (CRSsNP) (n = 16) and CRSwNP (n = 17), mRNAs for CD1a and CD1c were significantly elevated in NPs (n = 29). In contrast, CD207 mRNA was not elevated in NPs. Immunohistochemistry showed that CD1c(+) cells but not CD303(+) cells were significantly elevated in NPs compared to control subjects or patients with CRSsNP. Flow cytometric analysis showed that CD1a(+) cells in NPs might be a subset of mDC1s and that CD45(+) CD19(-) CD1c(+) CD11c(+) CD141(-) CD303(-) HLA-DR(+) mDC1s and CD45(+) CD19(-) CD11c(+) CD1c(-) CD141(high) HLA-DR(+) mDC2s were significantly elevated in NPs compared to UT from controls and CRSsNP, but CD45(+) CD11c(-) CD303(+) HLA-DR(+) pDCs were only elevated in NPs compared to control UT. CONCLUSION AND CLINICAL RELEVANCE: Myeloid DCs are elevated in CRSwNP, especially in NPs. Myeloid DCs thus may indirectly contribute to the inflammation observed in CRSwNP.

Reduced expression of antimicrobial PLUNC proteins in nasal polyp tissues of patients with chronic rhinosinusitis.

BACKGROUND: Chronic rhinosinusitis (CRS) is a disease characterized by inflammation of the nasal mucosa and paranasal sinuses. This inflammation may result in part from decreased epithelial barrier and innate immune responses, leading to frequent bacterial and fungal colonization. The objectives of this study were to investigate the expression of innate immune proteins of the palate lung and nasal epithelium clone (PLUNC) family in patients with CRS. METHODS: Nasal tissue samples were collected from control subjects and CRS patients with and without nasal polyps. Expression of the members of the PLUNC family was analyzed by real-time PCR. Expression of SPLUNC1 and LPLUNC2 proteins was analyzed by ELISA, immunoblot, and immunohistochemical analysis. RESULTS: Levels of mRNA for most of the members of the PLUNC family were profoundly reduced in nasal polyps (NPs) compared to uncinate tissue from control subjects or patients with CRS. LPLUNC2 and SPLUNC1 proteins were decreased in NPs of patients with CRS compared to uncinate tissue from control subjects. Immunohistochemical data revealed that within submucosal glands of sinonasal tissues, SPLUNC1 and LPLUNC2 were differentially expressed, in serous and mucous cells, respectively. The decrease in the expression of these molecules is probably explained by a decrease in the number of glands in NPs as revealed by correlations with levels of the glandular marker lactoferrin. CONCLUSIONS: Decreased SPLUNC1 and LPLUNC2 in NPs reflect a profound decrease in the number of submucosal glands. Decreased glands may lead to a localized defect in the production and release of glandular innate defense molecules.

The in vivo transfer of antigen-induced airway reactions by bronchial lumen cells.

Rhesus monkeys with airway responses to aerosol challenge with Ascaris antigen constitute a primate model of inhalant asthma. Previous studies have shown that bronchial lavage cells from airway-reactive animals will release histamine or a slow-reactive substance of anaphylaxis after challenge with antigen. Because these bronchial lumen cells are the first cells in contact with inhaled antigen, they may play a role in induction of antigen-induced airway responses. To evaluate this possibility, bronchial lumen lavage cells from animals with airway reactivity were transferred to the bronchial lumens of animals with negative airway responses to antigen challenge. The transfer of the bronchial lumen cells resulted in transient airway reactivity of the recipients to aerosol antigen challenge. It is suggested that the mast cells which constitute a component of the bronchial lumen cells may be the active cell alone, or in combination with other cells, which results in this primate immunoglobulin E-mediated airway response and its transfer to nonreactivie recipients.

Ionophore and arachidonic acid stimulation of airway responses in rhesus monkeys.

Aerosolized doses of the ionophore, A23187, and arachidonic acid individually resulted in no airway response in rhesus monkeys. When these two agents were given simultaneously, by aerosol, an airway response occurred. The pulmonary function abnormalities that occurred qualitatively simulated those of an antigen-induced airway response. This is the first demonstration in our laboratory of two agents which singly will not produce a response but which are reactive when delivered in combination. Other fatty acids did not produce a similar response. The response to A23187 and arachidonic acid occurred only in rhesus monkeys from our colony which had been demonstrated to have airway responses to aerosolized antigen challenge, a response shown previously to be associated with hyperreactive airways to pharmacologic stimuli. The A23187 and arachidonic acid response was inhibited by aerosolized 5,8,11,14-eicosatetraynoic acid, an inhibitor of the cyclooxygenase and lipoxygenase pathways of arachidonic acid metabolism. Further, indomethacin, a prostaglandin synthetase inhibitor of the cyclooxygenase pathway, inhibited the response, although previous studies showed that this drug will potentiate an antigen-induced response in this animal model of asthma. The slow-reacting substance of anaphylaxis antagonist, FPL 55712, did not inhibit the A23187-arachidonic acid response under the conditions of these experiments. The mechanism of the A23187-arachidonic acid airway response in rhesus monkeys may or may not be the same as the antigen-induced response.

Reagin-mediated asthma in rhesus monkeys and relation to bronchial cell histamine release and airway reactivity to carbocholine.

Rhesus monkeys with persistent immediate-type cutaneous and respiratory responses (RR) to ascaris antigen (AA) were compared with rhesus monkeys with skin reactivity and no respiratory responses, and animals with no skin reactivity and no respiratory responses to inhaled antigen (NR). The RR group could not be distinguished from the nonresponding (NR) group by the cutaneous skin test titers, serum, or respiratory secretion IgE concentration. Leukocyte histamine (H) release due to anti-IgE was similar with peripheral blood leukocytes and bronchial lumen mast cells (MC) from RR and NR animals. The RR group of animals could be distinguished from the NR group by their degree of sensitivity to inhaled carbocholine and H release from respiratory MC exposed to AA. The RR group demonstrates consistent, persistent respiratory responses suitable for immunologic, pharmacologic, and physiologic studies. Finally, it was found that the IgE concentration in respiratory secretions of rhesus monkeys was comparatively higher than in serum, evidence for IgE as a secretory Ig in the respiratory tract of this species.

Six-month angiographic and clinical follow-up of patients prospectively randomized to receive either tirofiban or placebo during angioplasty in the RESTORE trial. Randomized Efficacy Study of Tirofiban for Outcomes and Restenosis.

OBJECTIVES: This study sought to investigate the effects of tirofiban versus placebo on the incidence of adverse cardiac outcomes and coronary artery restenosis at 6 months. BACKGROUND: Tirofiban is a highly selective, short-acting inhibitor of fibrinogen binding to platelet glycoprotein IIb/IIIa. In a recent clinical study, tirofiban reduced the incidence of adverse cardiovascular events at both 2 and 7 days after coronary angioplasty or directional coronary atherectomy. This reduction persisted but was no longer statistically significant at 30 days. METHODS: The Randomized Efficacy Study of Tirofiban for Outcomes and Restenosis (RESTORE) trial was a randomized, double-blind, placebo-controlled trial of tirofiban in patients undergoing balloon angioplasty or directional atherectomy within 72 h of presentation with either unstable angina pectoris or acute myocardial infarction. All patients received an initial bolus (10 microg/kg body weight over 3 min), followed by a 36-h infusion (0.15 microg/kg per min) of either tirofiban or placebo. RESULTS: At 6 months the composite end point (either death from any cause, new myocardial infarction, bypass surgery for angioplasty failure or recurrent ischemia, repeat target vessel angioplasty or stent insertion for actual or threatened abrupt closure) occurred in 1,070 placebo group patients (27.1%) and 1,071 tirofiban group patients (24.1%, p = 0.11). Analysis of 6-month coronary arteriograms by means of quantitative coronary arteriography showed no significant difference between placebo- and tirofiban-treated patients in either the incidence of a > or =50% diameter stenosis (57% vs. 51%, p = NS), a loss of > or =50% of lumen diameter gained (50% vs. 50%, p = NS) or a loss of > or =0.72 mm of lumen diameter (44% vs. 42%, p = NS). CONCLUSIONS: The 3% absolute reduction in the incidence of the composite end point at 6 months (27.1% placebo vs. 24.1% tirofiban) was similar to that previously reported at 2 days (8.7% vs. 5.4%, p < 0.005), and there does not appear to be any late effect of tirofiban on clinical end points between day 2 and 6 months. Tirofiban did not reduce the incidence of restenosis at 6 months when defined in a number of ways.

Idiopathic anaphylaxis. An attempt to estimate the incidence in the United States.

BACKGROUND: Idiopathic anaphylaxis has been described and classified, and increasing numbers of cases are being seen in the United States and abroad. Treatment regimens have been shown to be effective in prophylactic management. There is no available information about the number of cases in the United States. METHODS: We attempted to determine the number of cases of idiopathic anaphylaxis in the United States by mailing a questionnaire to all graduates (for the last 31 years) of the Northwestern University Allergy-Immunology Fellowship training program. RESULTS: Response to the questionnaire was 100%, and 633 cases were reported by this survey of 75 allergists. The current total number of identified cases of idiopathic anaphylaxis from all reports of cases in the United States is 1020. CONCLUSIONS: By extrapolation of the cases of idiopathic anaphylaxis reported by the allergists surveyed to the approximately 4000 allergists in the United States, the estimated number of cases in the United States is between 20,592 and 47,024. Idiopathic anaphylaxis is potentially fatal, represents a source of major medical health care costs, causes anxiety to patients and families, occurs in pediatric and adult populations, and is controlled by appropriate regimens. The estimated number of cases emphasizes the need for careful attention to idiopathic anaphylaxis by physicians.

Antihypertensive efficacy of once daily MK-521, a new nonsulfhydryl angiotensin-converting enzyme inhibitor.

The effects of the new nonsulfhydryl-containing oral converting-enzyme inhibitor MK-521 on blood pressure, heart rate, angiotensin-converting enzyme activity, plasma renin activity and plasma aldosterone concentration were assessed in 10 hypertensive patients. After a 2-week no-treatment period, patients received placebo and then 14 days each: MK-521 20 mg once daily, hydrochlorothiazide 50 mg once daily and the latter 2 in combination. During the last day of each treatment, the mean (+/- standard deviation) time-averaged (1- to 12-hour) standing diastolic blood pressure decreased from 106 +/- 8 (placebo) to 95 +/- 10 mm Hg with MK-521, 95 +/- 13 mm Hg with hydrochlorothiazide (p less than 0.05 vs placebo) and 88 +/- 11 mm Hg with the combination (p less than 0.05 vs all other treatments). The antihypertensive effect of MK-521 was maintained 24 hours after dosing. Heart rate did not change significantly after MK-521 treatment. MK-521 caused a marked suppression of converting enzyme activity for over 24 hours; plasma renin activity increased significantly after each active treatment and MK-521 significantly decreased the hydrochlorothiazide-induced elevation of plasma aldosterone concentration. In this short-term trial, MK-521 was well tolerated.

Hypersensitivity pneumonitis due to humidifier disease: seek and ye shall find.

STUDY OBJECTIVES: This study reports a classic case of hypersensitivity pneumonitis (HP) with classic histologic changes in lung tissue and the research used to identify the causative antigens. DESIGN: A patient with clinical, radiographic, pulmonary function abnormalities and a lung biopsy consistent with HP had no identifiable antigen exposure. SETTING: Evaluation of the patient's activities provided no suggestion of antigen exposure. Her home was evaluated. It was found that her humidifier ran continually without being cleaned but water was added periodically. MEASUREMENTS: Serologic analysis demonstrated precipitating antibodies against her humidifier water and ten antigens in the hypersensitivity lung disease serologic panel. CONCLUSION: Removal of the humidifier, cleaning of the house, and a course of prednisone resulted in the return of the patient to a normal state.

Aerosolized leukotriene D4 converts monkeys that are negative aerosolized ascaris responders to positive airway responders.

We designed studies to determine if Leukotriene D4 (LTD4) could alter airway reactivity such that rhesus monkeys with positive skin reactivity and consistently negative airway responses would respond to ascaris airway challenge. The experiments were complicated by the observation that aerosolized LTD4 would occasionally increase airway hyperreactivity in some monkeys used as controls such that an airway response occurred to saline, the diluent for ascaris antigen. In spite of this, we were able to demonstrate induction of airway responsiveness to ascaris antigen. These results demonstrate that LTD4 will induce airway hyperreactivity to a nonspecific stimulus such as aerosolized saline or to an allergen.

Combination therapy with tirofiban and enoxaparin in acute coronary syndromes.

BACKGROUND: Tirofiban, an intravenous glycoprotein IIb/IIIa antagonist, and enoxaparin, a low molecular weight heparin, have each been shown to be effective at reducing cardiac ischemic events compared to unfractionated heparin alone in separate trials of patients with unstable angina and non-Q-wave myocardial infarction. The combination of these agents may offer further therapeutic benefit. MATERIALS AND METHODS: Fifty-five patients with non-Q-wave myocardial infarction were randomized to receive double-blind treatment with tirofiban (0.1 microgram/kg/min i.v.) for 48-108 h coadministered with either enoxaparin (1 mg/kg sc q 12 h) (n=26) or unfractionated heparin (i.v. adjusted to activated partial-thromboplastin time) (n=27) to evaluate pharmacokinetics, pharmacodynamics, and safety. The primary objective of the study was to investigate the effect of unfractionated heparin versus enoxaparin on the plasma clearance of tirofiban. RESULTS: Coadministration of tirofiban and enoxaparin was generally well tolerated. Plasma clearance of tirofiban was 176.7+/-59.8 and 187.5+/-81.8 ml/min, respectively, for enoxaparin and unfractionated heparin-treated patients (P=NS). The mean difference was well within the prespecified criterion for comparability. Administration of tirofiban with enoxaparin vs. unfractionated heparin resulted in lesser variability and a trend towards greater inhibition of platelet aggregation using 5 microM adenosine phosphate agonist. More patients achieved target inhibition of platelet aggregation >70% in the tirofiban and enoxaparin group (84% vs. 65%, P=0.19). Median bleeding time was 21 min for tirofiban and enoxaparin vs. > or =30 min for tirofiban and unfractionated heparin (P=NS). For a given level of inhibition of platelet aggregation, bleeding time was less prolonged with tirofiban and enoxaparin than tirofiban and unfractionated heparin (adjusted mean bleeding time 19.6 vs. 24.9 min, P=0.02). Tirofiban plasma concentration and clearance were comparable whether coadministered with enoxaparin or unfractionated heparin. There were no major or minor bleeding events in either group by the TIMI criteria. INTERPRETATION: The more consistent inhibition of platelet aggregation and lower adjusted bleeding time of tirofiban and enoxaparin vs. tirofiban and unfractionated heparin support the therapeutic potential of combining these two agents. These data from the first clinical report of coadministration of a glycoprotein IIb/IIIa receptor antagonist and a low molecular weight heparin are consistent with prior data which show differential pharmacodynamic effects of enoxaparin and unfractionated heparin on platelet aggregation.

Immunoglobulin E antibody against environmental allergens in subjects with trimellitic anhydride-induced asthma.

The purpose of this study was to assess the relationship between atopy and the development of occupational asthma as a consequence of exposure to trimellitic anhydride (TMA). A case-control study was performed, which comprised 16 employees identified as having TMA-induced asthma and 44 similarly exposed controls. Specific immunoglobulin E measurements in response to cat, dust mite, ryegrass, and ragweed antigens were performed. Fifty-six percent of cases and 29% of controls were found to be atopic (P = 0.098). We demonstrated that there was a trend toward employees with TMA asthma being more atopic than those without TMA asthma. Atopy as an assessment of risk for the development of TMA asthma is unlikely to be useful, although further investigation may be warranted.

Total serum IgE in trimellitic anhydride-induced asthma.

The objective of this study was to determine whether total serum IgE levels are elevated in workers with trimellitic anhydride-induced asthma as compared with anhydride-exposed workers without an occupational immunologic syndrome. Sera from 12 highly exposed workers with trimellitic anhydride-induced asthma, and from 31 similarly exposed workers without occupational immunologic disease, were assayed for total immunoglobulin E (IgE) levels by Total IgE II FAST analysis. The mean total IgE levels were 176.74 ng/mL and 34.55 ng/mL respectively. The difference between the two groups was statistically significant, but considerable overlap of IgE levels between groups was seen. In conclusion, although mean total IgE levels are significantly different between TMA-exposed workers with or without occupational asthma, the significant amount of overlap and poor sensitivity of the test preclude the use of this assay in the individual evaluation of these workers.

Emergency medical recognition and management of idiopathic anaphylaxis.

Idiopathic anaphylaxis (IA) is a diagnosis of exclusion that is made when no identifiable causative factors can be found for an episode of anaphylaxis. IA is a potentially life-threatening disease that is the result of a nonimmunologic mast cell activation syndrome. Acute presentation and treatment of these patients is most often in the emergency department and is clinically the same as anaphylaxis from allergens. Since these episodes are unpredictable and often recurrent, these patients are at risk of death if not identified on acute presentation and managed appropriately. As an increasing number of patients are being diagnosed with IA, they will be presenting to emergency departments with initial and recurrent episodes of IA. Therefore, increased awareness of IA and coordinated care is needed so that the morbidity and mortality of this potentially fatal disease can be kept at a minimum.