Amygdalo-nigral inputs target dopaminergic and GABAergic neurons in the primate: a view from dendrites and soma.

The central nucleus (CeN) of the amygdala is an important afferent to the DA system that mediates motivated learning. We previously found that CeN terminals in nonhuman primates primarily overlap the elongated lateral VTA (parabrachial pigmented nucleus, PBP, A10), and retrorubral field(A8) subregion. Here, we examined CeN afferent contacts on cell somata and proximal dendrites of DA and GABA neurons, and distal dendrites of each, using confocal and electron microscopy (EM) methods, respectively. At the soma/proximal dendrites, the proportion of TH+ and GAD1+ cells receiving at least one CeN afferent contact was surprisingly similar (TH = 0.55: GAD1=0.55 in PBP; TH = 0.56; GAD1 =0.51 in A8), with the vast majority of contacted TH+ and GAD1+ soma/proximal dendrites received 1-2 contacts. Similar numbers of tracer-labeled terminals also contacted TH-positive and GAD1-positive small dendrites and/or spines (39% of all contacted dendrites were either TH- or GAD1-labeled). Overall, axon terminals had more symmetric (putative inhibitory) axonal contacts with no difference in the relative distribution in the PBP versus A8, or onto TH+ versus GAD1+ dendrites/spines in either region. The striking uniformity in the amygdalonigral projection across the PBP-A8 terminal field suggests that neither neurotransmitter phenotype nor midbrain location dictates likelihood of a terminal contact. We discuss how this afferent uniformity can play out in recently discovered differences in DA:GABA cell densities between the PBP and A8, and affect specific outputs. Significance statement: The amygdala's central nucleus (CeN) channels salient cues to influence both appetitive and aversive responses via DA outputs. In higher species, the broad CeN terminal field overlaps the parabrachial pigmented nucleus ('lateral A10') and the retrorubral field (A8). We quantified terminal contacts in each region on DA and GABAergic soma/proximal dendrites and small distal dendrites. There was striking uniformity in contacts on DA and GABAergic cells, regardless of soma and dendritic compartment, in both regions. Most contacts were symmetric (putative inhibitory) with little change in the ratio of inhibitory to excitatory contacts by region.We conclude that post-synaptic shifts in DA-GABA ratios are key to understanding how these relatively uniform inputs can produce diverse effects on outputs.

Corticotropin-releasing factor-dopamine interactions in male and female macaque: Beyond the classic VTA.

Dopamine (DA) is involved in stress and stress-related illnesses, including many psychiatric disorders. Corticotropin-releasing factor (CRF) plays a role in stress responses and targets the ventral midbrain DA system, which is composed of DA and non-DA cells, and divided into specific subregions. Although CRF inputs to the midline A10 nuclei ("classic VTA") are known, in monkeys, CRF-containing terminals are also highly enriched in the expanded A10 parabrachial pigmented nucleus (PBP) and in the A8 retrorubral field subregions. We characterized CRF-labeled synaptic terminals on DA (tyrosine hydroxylase, TH+) and non-DA (TH-) cell types in the PBP and A8 regions using immunoreactive electron microscopy (EM) in male and female macaques. CRF labeling was present mostly in axon terminals, which mainly contacted TH-negative dendrites in both subregions. Most CRF-positive terminals had symmetric profiles. In both PBP and A8, CRF symmetric (putative inhibitory) synapses onto TH-negative dendrites were significantly greater than asymmetric (putative excitatory) profiles. This overall pattern was similar in males and females, despite shifts in the size of these effects between regions depending on sex. Because stress and gonadal hormone shifts can influence CRF expression, we also did hormonal assays over a 6-month time period and found little variability in basal cortisol across similarly housed animals at the same age. Together our findings suggest that at baseline, CRF-positive synaptic terminals in the primate PBP and A8 are poised to regulate DA indirectly through synaptic contacts onto non-DA neurons.

Association of self-directed walking with toxicity moderation during chemotherapy for the treatment of early breast cancer.

BACKGROUND: In the field of exercise oncology, there is a need to quantify the potential benefits of moderate, self-directed physical activity during active treatment. In a pooled analysis of three identical single-arm intervention studies, we investigate the association of activity tracker steps with patient-reported toxicities during chemotherapy. METHODS: Women with early breast cancer who were enrolled in the intervention studies reported their symptom severity every 2-3 weeks throughout chemotherapy, and daily steps were documented through a Fitbit activity tracker. Relative risks (RR) and 95% confidence intervals (CI) were calculated using Poisson regression models with robust variance. For outcomes significant in unadjusted models, adjusted RRs were calculated controlling for race, age, and education level. Tracker step cut point (high step, low step) was determined by the means. Cumulative incidence functions of moderate, severe, and very severe (MSVS) symptoms were estimated using the Kaplan-Meier method and compared using a Cox proportional hazard model. RESULTS: In a sample of 283 women, mean age was 56 years and 76% were White. Mean tracker-documented steps/week were 29,625, with 55% walking below the mean (low step) and 45% above (high step). In multivariable analysis, high step patients had lower risk for fatigue [RR 0.83 (0.70, 0.99)] (p = 0.04), anxiety [RR 0.59 (0.42, 0.84)] (p = 0.003), nausea [RR 0.66 (0.46, 0.96)] (p = 0.03), depression [RR 0.59 (0.37, 0.03)] (p = 0.02), and ≥ 6 MSVS symptoms [RR 0.73 (0.54, 1.00)] (p = 0.05) and had 36% lower risk for dose reductions [RR 0.64 (95% CI 0.43, 0.97)] (p = 0.03). CONCLUSION: Self-directed walking at a rate of at least 30,000 steps/week may moderate the severity of treatment side effects during chemotherapy for early breast cancer. TRIAL NUMBERS: NCT02167932, NCT02328313, NCT03761706.

Translating Fear Circuitry: Amygdala Projections to Subgenual and Perigenual Anterior Cingulate in the Macaque.

Rodent fear-learning models posit that amygdala-infralimbic connections facilitate extinction while amygdala-prelimbic prefrontal connections mediate fear expression. Analogous amygdala-prefrontal circuitry between rodents and primates is not established. Using paired small volumes of neural tracers injected into the perigenual anterior cingulate cortex (pgACC; areas 24b and 32; a potential homologue to rodent prelimbic cortex) and subgenual anterior cingulate cortex (sgACC, areas 25 and 14c; a potential homologue to rodent infralimbic cortex) in a single hemisphere, we mapped amygdala projections to the pgACC and sgACC within single subjects. All injections resulted in dense retrograde labeling specifically within the intermediate division of the basal nucleus (Bi) and the magnocellular division of the accessory basal nucleus (ABmc). Areal analysis revealed a bias for connectivity with the sgACC, with the ABmc showing a greater bias than the Bi. Double fluorescence analysis revealed that sgACC and pgACC projections were intermingled within the Bi and ABmc, where a proportion were double labeled. We conclude that amygdala inputs to the ACC largely originate from the Bi and ABmc, preferentially connect to the sgACC, and that a subset collaterally project to both sgACC and pgACC. These findings advance our understanding of fear extinction and fear expression circuitry across species.

The neuroanatomic complexity of the CRF and DA systems and their interface: What we still don't know.

Corticotropin-releasing factor (CRF) is a neuropeptide that mediates the stress response. Long known to contribute to regulation of the adrenal stress response initiated in the hypothalamic-pituitary axis (HPA), a complex pattern of extrahypothalamic CRF expression is also described in rodents and primates. Cross-talk between the CRF and midbrain dopamine (DA) systems links the stress response to DA regulation. Classically CRF + cells in the extended amygdala and paraventricular nucleus (PVN) are considered the main source of this input, principally targeting the ventral tegmental area (VTA). However, the anatomic complexity of both the DA and CRF system has been increasingly elaborated in the last decade. The DA neurons are now recognized as having diverse molecular, connectional and physiologic properties, predicted by their anatomic location. At the same time, the broad distribution of CRF cells in the brain has been increasingly delineated using different species and techniques. Here, we review updated information on both CRF localization and newer conceptualizations of the DA system to reconsider the CRF-DA interface.

IL-3 up-regulates and activates human eosinophil CD32 and αMß2 integrin causing degranulation.

BACKGROUND: Eosinophils contribute to the pathogenesis of multiple diseases, including asthma. Treatment with antibodies targeting IL-5 or IL-5 receptor α reduces the frequency of asthma exacerbations. Eosinophil receptors for IL-5 share a common ß-chain with IL-3 and GM-CSF receptors. We recently reported that IL-3 is more potent than IL-5 or GM-CSF in maintaining the ERK/p90S6K/RPS6 ribosome-directed signaling pathway, leading to increased protein translation. OBJECTIVE: We aimed to determine disease-relevant consequences of prolonged eosinophil stimulation with IL-3. RESULTS: Human blood eosinophils were used to establish the impact of activation with IL-3 on IgG-driven eosinophil degranulation. When compared to IL-5, continuing exposure to IL-3 further induced degranulation of eosinophils on aggregated IgG via increased production and activation of both CD32 (low affinity IgG receptor) and αMß2 integrin. In addition, unlike IL-5 or GM-CSF, IL-3 induced expression of CD32B/C (FCGRIIB/C) subtype proteins, without changing CD32A (FCGRIIA) protein and CD32B/C mRNA expression levels. Importantly, these in vitro IL-3-induced modifications were recapitulated in vivo on airway eosinophils. CONCLUSIONS AND CLINICAL RELEVANCE: We observed for the first time upregulation of CD32B/C on eosinophils, and identified IL-3 as a potent inducer of CD32- and αMß2-mediated eosinophil degranulation.

Microglial P2Y12 is necessary for synaptic plasticity in mouse visual cortex.

Microglia are the resident immune cells of the brain. Increasingly, they are recognized as important mediators of normal neurophysiology, particularly during early development. Here we demonstrate that microglia are critical for ocular dominance plasticity. During the visual critical period, closure of one eye elicits changes in the structure and function of connections underlying binocular responses of neurons in the visual cortex. We find that microglia respond to monocular deprivation during the critical period, altering their morphology, motility and phagocytic behaviour as well as interactions with synapses. To explore the underlying mechanism, we focused on the P2Y12 purinergic receptor, which is selectively expressed in non-activated microglia and mediates process motility during early injury responses. We find that disrupting this receptor alters the microglial response to monocular deprivation and abrogates ocular dominance plasticity. These results suggest that microglia actively contribute to experience-dependent plasticity in the adolescent brain.

Nasal lavage VEGF and TNF-α levels during a natural cold predict asthma exacerbations.

BACKGROUND: Asthma exacerbations contribute to significant morbidity, mortality and healthcare utilization. Furthermore, viral infections are associated with asthma exacerbations by mechanisms that are not fully understood. OBJECTIVE: The aim of this analysis was to determine whether cytokine patterns in patients with colds could identify risks for subsequent asthma exacerbations. METHODS: We analysed cytokine levels in nasal lavage fluid (NLF) in 59 subjects (46 with asthma) with acute upper respiratory symptoms and after symptomatic resolution. Analyte choice was based on potential relevance to asthma exacerbations: antiviral (IFN-α, IFN-ß, IFN-γ, IFN-λ1, IP-10, TRAIL), cell recruiting (G-CSF, IL-1ß, IL-8, MCP-1, MCP-3, TNF-α), polarizing (CXCL13, IL-10, IL-13, IL-17, TSLP), and injury remodelling (fibronectin, IL-33, MMP-9, VEGF). RESULTS: The overall cytokine response induced during viral infections was not different between asthmatic and non-asthmatic individuals for a wide array of cytokines. However, mean levels of VEGF, TNF-α and IL-1ß were 1.7-, 5.1- and 4.7-fold higher in samples from asthma subjects who exacerbated in the first 3 weeks of the cold compared with those who did not exacerbate (P = 0.006, 0.01, 0.048, respectively). Using receiver operating characteristic curve-defined thresholds, high VEGF and TNF-α levels predicted a shorter time-to-exacerbation after NLF sampling (25% exacerbation rate: 3 vs. 45 days, and 3 vs. 26 days; P = 0.03, 0.04, respectively). CONCLUSION AND CLINICAL RELEVANCE: Although they produce similar cytokine responses to viral infection as non-asthmatics, asthmatics with higher levels of VEGF and TNF-α in NLF obtained during acute cold phases predicted subsequent asthma exacerbations in this cohort of patients with mild-to-moderate disease. In the future, stratifying the risk of an asthma exacerbation by cytokine profile may aid the targeting of personalized treatment and intervention strategies.

Anti-IL-5 attenuates activation and surface density of ß(2) -integrins on circulating eosinophils after segmental antigen challenge.

BACKGROUND: IL-5 activates α(M) ß(2) integrin on blood eosinophils in vitro. Eosinophils in bronchoalveolar lavage (BAL) following segmental antigen challenge have activated ß(2) -integrins. OBJECTIVE: To identify roles for IL-5 in regulating human eosinophil integrins in vivo. METHODS: Blood and BAL eosinophils were analysed by flow cytometry in ten subjects with allergic asthma who underwent a segmental antigen challenge protocol before and after anti-IL-5 administration. RESULTS: Blood eosinophil reactivity with monoclonal antibody (mAb) KIM-127, which recognizes partially activated ß(2) -integrins, was decreased after anti-IL-5. Before anti-IL-5, surface densities of blood eosinophil ß(2) , α(M) and α(L) integrin subunits increased modestly post challenge. After anti-IL-5, such increases did not occur. Before or after anti-IL-5, surface densities of ß(2) , α(M) , α(L) and α(D) and reactivity with KIM-127 and mAb CBRM1/5, which recognizes high-activity α(M) ß(2) , were similarly high on BAL eosinophils 48 h post-challenge. Density and activation state of ß(1) -integrins on blood and BAL eosinophils were not impacted by anti-IL-5, even though anti-IL-5 ablated a modest post-challenge increase on blood or BAL eosinophils of P-selectin glycoprotein ligand-1 (PSGL-1), a receptor for P-selectin that causes activation of ß(1) -integrins. Forward scatter of blood eosinophils post-challenge was less heterogeneous and on the average decreased after anti-IL-5; however, anti-IL-5 had no effect on the decreased forward scatter of eosinophils in post-challenge BAL compared with eosinophils in blood. Blood eosinophil KIM-127 reactivity at the time of challenge correlated with the percentage of eosinophils in BAL post-challenge. CONCLUSION AND CLINICAL RELEVANCE: IL-5 supports a heterogeneous population of circulating eosinophils with partially activated ß(2) -integrins and is responsible for up-regulation of ß(2) -integrins and PSGL-1 on circulating eosinophils following segmental antigen challenge but has minimal effects on properties of eosinophils in BAL. Dampening of ß(2) -integrin function of eosinophils in transit to inflamed airway may contribute to the decrease in lung inflammation caused by anti-IL-5.

Segmental allergen challenge enhances chitinase activity and levels of CCL18 in mild atopic asthma.

BACKGROUND: Allergic airway inflammation contributes to the airway remodelling that has been linked to increased obstruction and morbidity in asthma. However, the mechanisms by which allergens contribute to airway remodelling in humans are not fully established. CCL18, chitotriosidase (CHIT1) and YKL-40 are readily detectable in the lungs and contribute to remodelling in other fibrotic diseases, but their involvement in allergic asthma is unclear. OBJECTIVE: We hypothesized that CCL18, YKL-40 and CHIT1 bioactivity are enhanced in allergic asthma subjects after segmental allergen challenge and are related to increased pro-fibrotic and Th2-associated mediators in the lungs. METHODS: Levels of CCL18 and YKL-40 protein and chitotriosidase (CHIT1) bioactivity in bronchoalveolar lavage (BAL) fluid, as well as CCL18, YKL-40 and CHIT1 mRNA levels in BAL cells were evaluated in patients with asthma at baseline and 48 h after segmental allergen challenge. We also examined the correlation between CCL18 and YKL-40 levels and CHIT1 activity with the levels of other pro-fibrotic factors and chemokines previously shown to be up-regulated after allergen challenge. RESULTS: Chitotriosidase activity and YKL-40 and CCL18 levels were elevated after segmental allergen challenge and these levels correlated with those of other pro-fibrotic factors, T cell chemokines, and inflammatory cells after allergen challenge. CCL18 and YKL-40 mRNA levels also increased in BAL cells after allergen challenge. CONCLUSIONS AND CLINICAL RELEVANCE: Our results suggest that CCL18 and YKL-40 levels and CHIT1 activity are enhanced in allergic airway inflammation and thus may contribute to airway remodelling in asthma.

Human eosinophils release IL-1ß and increase expression of IL-17A in activated CD4+ T lymphocytes.

BACKGROUND: Differentiation and activation of CD4(+) T cells is controlled by various cytokines produced by innate immune cells. We have shown that eosinophils (EOS) have the potential to influence Th1 and Th2 cytokine generation by CD4(+) cells, but their influence on IL-17A (IL-17) has not been established. OBJECTIVE: The purpose of this study is to determine the effect of EOS on IL-17 production by lymphocytes. METHODS: Pre-activated CD4(+) T cells were cultured in the presence of either autologous EOS or EOS culture supernatants. Expression of IL-17 was determined by real-time quantitative PCR (qPCR) after 5 h and protein level was measured after 48 h. To determine the effect of allergen-induced airway EOS on IL-17, subjects with mild allergic asthma underwent bronchoscopic segmental bronchoprovocation with allergen (SBP-Ag) after a treatment with an anti-IL-5 neutralizing antibody (mepolizumab) to reduce airway eosinophilia. IL-17 mRNA was measured in bronchoalveolar lavage (BAL) cells by qPCR. RESULTS: In vitro, EOS significantly increased IL-17 production by CD4(+) T cells. Addition of exogenous IL-1ß increased expression of IL-17 mRNA by CD4(+) T cells. EOS expressed and released IL-1ß. Furthermore, levels of IL-1ß in EOS supernatants highly correlated with their ability to increase IL-17 expression by CD4(+) T cells, and neutralizing antibody to IL-1ß reduced expression of IL-17 mRNA. In vivo, reduction of EOS in the airway using mepolizumab was associated with diminished IL-17 expression after SBP-Ag. CONCLUSIONS AND CLINICAL RELEVANCE: Our data demonstrate that EOS can promote IL-17 production through the release of IL-1ß. Enhanced IL-17 cytokine production is another mechanism by which EOS may participate in pathogenesis of allergic airway inflammation in asthma.

Human airway and peripheral blood eosinophils enhance Th1 and Th2 cytokine secretion.

BACKGROUND: The effector function of eosinophils involves their release of toxic granule proteins, reactive oxygen species, cytokines, and lipid mediators. Murine studies have demonstrated that eosinophils can also enhance T cell function. Whether human eosinophils, in particular, airway eosinophils, have similar immunoregulatory activity has not been fully investigated. The aim of this study was to determine whether human blood and airway eosinophils can contribute to Th1 and Th2 cytokine generation from CD4+ T cells stimulated with superantigen. METHODS: Eosinophils were obtained from blood or bronchoalveolar lavage fluid 48 h after segmental allergen bronchoprovocation. Purified eosinophils were co-cultured with autologous CD4+ blood T cells in the presence of staphylococcal enterotoxin B (SEB). Cytokine levels in the supernatant fluid were determined by enzyme-linked immunosorbent assay (ELISA). Eosinophil expression of major histocompatibility complex (MHC) class II and co-stimulatory molecules was assessed by flow cytometry before culture, 24 h after granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulation, and 24 h after co-culture with CD4+ T cells and SEB. RESULTS: Interleukin (IL)-5, IL-13, and interferon (IFN)-gamma generation increased when CD4+ T cells were co-cultured with either blood or airway eosinophils in the presence of SEB. The ability of eosinophils to enhance cytokine generation was independent of their source (blood vs airway), activation by GM-CSF, or detectable expression of human leukocyte antigen (HLA)-DR, CD80, or CD86. CONCLUSION: Our data demonstrate that SEB-induced generation of Th1 and Th2 cytokines is increased in the presence of human blood and airway eosinophils. Thus, eosinophils can have an immunoregulatory function in pathogen-associated allergic diseases such as atopic dermatitis, chronic sinusitis, and asthma exacerbations.

Inflammatory changes associated with circadian variation in pulmonary function in subjects with mild asthma.

BACKGROUND: Nocturnal enhancement of airway inflammation has been demonstrated in patients with asthma who have a significant drop in pulmonary function at night. OBJECTIVE: To investigate the circadian changes in airway inflammation and their relationship with variations in pulmonary function in subjects with mild atopic asthma. METHODS: Twelve asthma subjects were admitted to the hospital for two separate 24-h visits. Bronchoalveolar lavage (BAL) was performed at 04:00 hours during one visit, and at 16:00 hours during another visit. BAL cells were analysed for lymphocyte phenotype and the capacity to secrete cytokines following ex vivo stimulation with phytohaemagglutinin (PHA). RESULTS: The numbers of BAL lymphocytes and the percentage of CD4+ T cells were higher at 04:00 hours compared with 16:00 hours. At 04:00 hours, the forced expiratory volume in 1 s (FEV1) was inversely correlated with BAL lymphocytes and CD4+ cells. PHA-induced generation of IL-5 by BAL cells correlated with BAL eosinophils and CD4+ cells. Moreover, there was a linear relationship between the relative change (16:00-04:00 hours) in IL-5 and circadian variation in FEV1. CONCLUSIONS: These data suggest that the circadian variation in lung function in asthma is associated with increased airway CD4+ lymphocyte numbers and their capacity to generate IL-5. Furthermore, in mild asthma, these circadian changes appear to fall into a continuous range, suggesting that day/night variations in airway inflammation and lung function occur on a continuum, rather than as an all-or-none phenomenon.

Auditory function at 14 years of age of very-low-birthweight.

The aim of the study was to determine audiological function at 14 years of age of very-low-birthweight (VLBW < or = 1500 g) children compared with a cohort of normal birthweight (NBW > 2499 g) children. Participants were consecutive surviving preterm children of birthweight < 1000 g born between 1977 and 1982 (n=86) and of birthweight 1000 to 1500 g born between 1980 and 1982 (n=124) and randomly selected NBW children born between 1981 and 1982 (n=60). Audiometric tests included pure tone audiometry, tympanometry, stapedius muscle reflexes, and measures of central auditory processing. Psychometric tests included measures of IQ, academic achievement, and behaviour. There were no significant differences in rates of hearing impairment, abnormal tympanograms, figure-ground problems, or digit recall between VLBW children and NBW control children. VLBW children had higher rates of some central auditory processing problems, which in turn were associated with poorer intellectual, academic, and behavioural progress.

Cognition, academic progress, behavior and self-concept at 14 years of very low birth weight children.

The aim of this study was to compare cognition, academic progress, behavior, and self-concept children of very low birth weight (VLBW, birth weight < 1501 g) born in the period 1980 to 1982 with randomly selected children of normal birth weight (NBW, birth weight > 2,499 g). At 14 years of age, 130 (84.4%) of 154 VLBW and 42 (70.0%) of 60 NBW children were assessed. Ten VLBW children and one NBW child who had cerebral palsy were excluded. VLBW children scored at a significantly lower level on all three composite scales of the Wechsler Intelligence Scale for Children, 3rd Edition. VLBW children were also significantly disadvantaged on more specific cognitive processes, including tests of visual processing and visual memory and on subtests reflecting learning and problem solving. Only in arithmetic was a difference between the groups discerned on tests of achievement. Significantly more VLBW children were rated by teachers as socially rejected and by their parents as having learning problems at school. VLBW children had significantly reduced self-esteem. VLBW children had more cognitive, academic, and behavioral problems and lower self-esteem at 14 years of age than NBW control subjects.

The relationship of sputum eosinophilia and sputum cell generation of IL-5.

BACKGROUND: Eosinophil recruitment to the airway after antigen challenge is regulated by many factors, including airway cell generation of cytokines. OBJECTIVES: The purpose of this study was to determine the relationship between sputum cell generation of IL-5 and the appearance of eosinophils in the sputum after antigen challenge. METHODS: Sputum samples from 11 allergic subjects were collected before and again 4 and 24 hours after antigen challenge. In 6 of these subjects, induced sputum samples were also obtained 48 hours and 7 days after challenge. Sputum leukocyte differential and cell counts and eosinophil-derived neurotoxin levels were determined. Sputum cells were then cultured with PHA (10 microg/mL) to stimulate IL-5 and IFN-gamma, which were measured in culture supernatants. RESULTS: An increase in sputum eosinophils and eosinophil-derived neurotoxin levels was detected at 4 hours after antigen challenge, with peak values at 24 hours. In contrast, significant increases in ex vivo generation of IL-5 by sputum cells was not seen until 24 hours after challenge. At 24 hours, PHA-induced IL-5 correlated with airspace eosinophil values (r (s) = 0.78, P <.01). In addition, the ratio of IFN-gamma/IL-5 decreased at 24 hours (P <.05) and had an inverse correlation with sputum eosinophils (r (s)= -0.68, P <.05). CONCLUSION: Although eosinophils are increased in the airway lumen as early as 4 hours, the ex vivo generation of IL-5 by sputum cells is first noted in samples obtained 24 hours after antigen challenge. This suggests that the early (4 hours) recruitment of eosinophils to the airway lumen may be regulated by factors other than IL-5 or that mucosal cells (rather than airspace cells) contribute to the IL-5 generation at this time point. Furthermore, IL-5 generation by airspace cells may be more responsible for either eosinophil recruitment or retention at later time points.

Inhaled budesonide decreases airway inflammatory response to allergen.

To define the mechanisms by which inhaled glucocorticosteroid regulates allergen-induced airway inflammation, a double-blind, placebo-controlled, cross-over study with inhaled budesonide was conducted in 14 subjects with allergic asthma. After baseline bronchoscopy and bronchoalveolar lavage (BAL), subjects were randomized to budesonide (400 microgram, twice daily) or placebo treatment for 4 wk. At the end of each treatment phase, whole-lung allergen inhalation challenge was performed, followed by BAL 48 h later. Budesonide treatment improved the FEV(1), attenuated both the immediate- and late-phase response to allergen, and prevented the increase in bronchial hyperresponsiveness after allergen challenge. Budesonide treatment also decreased allergen-induced airway eosinophilia. To determine the effects of budesonide on airway cell function, BAL cells were stimulated ex vivo with the T cell mitogen PHA, and cytokine generation was measured by ELISA. Budesonide decreased ex vivo generation of IL-5 and IFN-gamma by BAL cells. Ex vivo IL-5 production was significantly correlated with the number of airway eosinophils (r(s) = 0.61), and levels of eosinophil-derived neurotoxin (EDN) (r(s) = 0.57) and IL-5 (r(s) = 0.52) in BAL fluid. Moreover, PHA-induced IL-5 generation correlated with FEV(1) fall during the late-phase response to allergen (r(s) = 0.60). Budesonide decreased circulating eosinophils and serum levels of IL-5, but did not reduce IL-5 generation by peripheral blood mononuclear cells. The reduction in circulating eosinophils correlated with the decrease in levels of EDN (r(s) = 0.61) in BAL fluid and late response to inhaled allergen (r(s) = 0.51). These findings suggest that long-term treatment with inhaled budesonide reduces airway cell generation of cytokines, specifically IL-5, which then decreases circulating eosinophils and their availability for recruitment to the airway after allergen exposure.

Increased matrix metalloproteinase-9 in the airway after allergen challenge.

Persistent asthma is associated with airway inflammation, tissue damage, and deposition of extracellular matrix (ECM) proteins, which may be mediated, in part, through release of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinase-1 (TIMP-1). To investigate the role of allergen in the induction of MMP-9 and TIMP-1, bronchoscopy and segmental bronchoprovocation (SBP) with saline (SAL) and antigen (AG) were performed in 17 allergic subjects. Bronchoalveolar lavage (BAL) was done 5 min and 48 h after challenge and concentrations of MMP-9 and TIMP-1 in BAL fluid (BALF) were measured by ELISA. Forty-eight hours after AG challenge, concentrations of MMP-9 and TIMP-1 were increased in the airway, but not in serum. Zymography demonstrated that MMP-9 was the predominant metalloproteinase and was present in a latent proform. MMP-9 immunoreactivity was associated primarily with neutrophils, and concentrations of MMP-9 in BALF correlated with airway neutrophils and, to a lesser extent, with alveolar macrophages. These data suggest that AG challenge leads to generation of factors, including MMP-9, that may be associated with the initiation of bronchial injury, which may then lead to remodeling in asthma.

Enhanced cytokine generation by peripheral blood mononuclear cells in allergic and asthma subjects.

BACKGROUND: Although preferential expression of the Th2 cytokines, IL-4 and IL-5, has been described in atopic asthma, the role of IFN-gamma and IL-10 are less clear. OBJECTIVE: To determine the cytokine pattern of T cell mitogen-induced peripheral blood mononuclear cells obtained from atopic asthmatic (AA) subjects. METHODS: Peripheral blood mononuclear cells obtained from AA (n = 24), allergic rhinitis (AR) (n = 9), and normals (NL) (n = 9) were stimulated with phytohemagglutinin (PHA) and the generation of IL-4, IL-5, IFN-gamma, IL-10, and GM-CSF was quantified using ELISA. RESULTS: Compared with NL subjects, peripheral blood mononuclear cells from the atopic groups had increased generation of both IL-5 (AA, P = .001 and AR, P = .024) and IFN-gamma (AA, P = .037 and AR, P = .048) and decreased generation of IL-10 (AA, P = .038 and AR, P = .036). The absolute levels of cytokines did not differ between the two atopic groups; however, the ratio of IL-5/IL-10 was significantly higher in AA (P < .05), but not in AR when compared with NL subjects. CONCLUSION: The concomitant increase in the generation of IL-5 and IFN-gamma, with a decrease in IL-10 in the atopic groups suggests that in, at least a subset of these patients, there is potential expression of both Th2- and Th1-type cytokines. Furthermore, the increased IL-5 to IL-10 ratio could represent a key feature that distinguishes atopic asthmatic from non-asthmatic atopic subjects.

Antenatal corticosteroids and outcome at 14 years of age in children with birth weight less than 1501 grams.

OBJECTIVE: To determine whether exposure to antenatal corticosteroid therapy was associated with adverse effects on growth, sensorineural outcome, or lung function of children of birth weight <1501 g at 14 years of age. DESIGN: Cohort study. SETTING: The Royal Women's Hospital, Melbourne, Australia. SUBJECTS: One hundred fifty-four consecutive survivors born from October 1, 1980 to March 31, 1982. INTERVENTIONS: The mothers of 78 survivors (51%) had been given corticosteroids antenatally to accelerate fetal lung maturation. Treatment with antenatal corticosteroids was nonrandom. No mother received >1 course of corticosteroids. OUTCOME MEASURES: The children were assessed at 14 years of age, corrected for prematurity. All assessors were unaware of the exposure of the child to antenatal corticosteroids. The assessments included measurements of growth and neurological, cognitive, and lung function. Growth measurements were converted into z scores (standard deviation) for the appropriate age and gender. RESULTS: Of the 154 survivors, 130 (84%) were assessed at 14 years of age. Overall, the children exposed to antenatal corticosteroids were significantly taller (height z score; mean difference:.39; 95% confidence interval:.001-. 79) and had better cognitive functioning (Wechsler Intelligence Scale for Children-Third Edition Full Scale; IQ mean difference: 6. 2; 95% confidence interval:.8-11.6) than those not exposed to corticosteroids. There were no other differences in sensorineural outcomes between the groups. Lung function was not significantly different between the groups. No conclusions were altered by adjustment for confounding variables. CONCLUSIONS: Exposure to 1 course of antenatal corticosteroid therapy was associated with some clinically and statistically improved outcomes at 14 years of age in children of birth weight <1501 g, with no obvious adverse effects on growth or on sensorineural, cognitive, or lung function. corticosteroids, growth, cognitive, IQ, lung function, adolescence.