Neda Aberle

Interferon-γ release assay for the diagnosis of latent tuberculosis in children younger than 5 years of age.

Background: There are limited data available on interferon-&ggr; release assay (IGRA) performance in children up to 5 years of age, with documented exposure to active tuberculosis (TB). The aim of this study was to evaluate (1) the influence of infectivity of adult source cases on test results, (2) the impact of age, and (3) the level of agreement, between IGRA and tuberculin skin test (TST) results. Methods: A total of 142 Bacille Calmette-Guerin-vaccinated children up to 5 years of age were investigated because of a history of exposure to active TB. QuantiFERON-TB Gold In-Tube IGRA (QFT) and TST assays were performed. Results: Test results were significantly influenced by positive finding of cavitary lesions (QFT, odds ratio [OR] = 6.15; TST, OR = 7.48) and positive acid-fast bacilli (QFT, OR = 4.01; TST, OR = 4.47) in active TB contacts. QFT resulted in 1 indeterminate response (0.7%), attributable to low mitogen. There was no evidence for age having any effect on QFT performance. The 2 tests showed a moderate overall concordance (89%; &kgr; = 0.591) at a TST cutoff value of ≥10 mm. Conclusions: Association of positive QFT and TST results with risk factors for infection in child contacts (presence of cavitary lesions and acid-fast bacilli smear positivity in index cases) suggests that both the tests have good diagnostic accuracy. However, there was significant discord between results of the 2 tests that could not be definitively resolved. Thus, in a high-risk population of children up to 5 years of age, both tests (QFT and TST) should be performed and the child should be considered infected if either or both tests are positive.

Genetic Variation in Vascular Endothelial Growth Factor-A and Lung Function

RATIONALE Given the role of vascular endothelial growth factor (VEGF) in lung development, we hypothesized that polymorphisms in VEGF-A may be associated with lung function. OBJECTIVES The current study was designed to assess the role of genetic variants in VEGF-A as determinants of airway function from infancy through early adulthood. METHODS Association between five single-nucleotide polymorphisms (SNPs) in VEGF-A and lung function were assessed longitudinally in two unselected birth cohorts and cross-sectionally among infants. Replication with two SNPs was conducted in adults and children with asthma. We investigated the functionality of the SNP most consistently associated with lung function (rs3025028) using Western blotting to measure the ratio of plasma VEGF-A(165b)/panVEGF-A(165) among homozygotes. MEASUREMENTS AND MAIN RESULTS In two populations in infancy, C-allele homozygotes of rs3025028 had significantly higher VmaxFRC, forced expiratory flow(50), and forced expiratory flow(25-75) compared with other genotype groups. Among preschool children (age 3 yr), C allele of rs3025028 was associated with significantly higher specific airway conductance, with similar findings observed for lung function in school-age children. For FEV(1)/FVC ratio similar findings were observed among adolescents and young adults (birth cohort), and then replicated in adults and schoolchildren with asthma (cross-sectional studies). For rs3025038, plasma VEGF-A(165b)/panVEGF-A(165) was significantly higher among CC versus GG homozygotes (P ≤ 0.02) at birth, in school-age children, and in adults. CONCLUSIONS We report significant associations between VEGF-A SNP rs3025028 and parameters of airway function measured throughout childhood, with the effect persisting into adulthood. We propose that the mechanism may be mediated through the ratios of active and inhibitory isoforms of VEGF-A(165), which may be determined by alternative splicing.

17q12-21 and asthma: interactions with early-life environmental exposures.

BACKGROUND 17q12-21 polymorphisms are associated with asthma presence and severity across different populations. OBJECTIVE To extensively investigate the genes in this region among Croatian schoolchildren in a case-control study, taking account of early-life environmental exposures. METHODS We included 423 children with asthma and 414 controls aged 5 to 18 years. Fifty-one haplotype tagging single-nucleotide polymorphisms (SNPs) were genotyped (GSDMA, GSDMB, ORMDL3, IKZF3, ZPBP2, and TOP2). Data on exposure to smoking and furry pet ownership were collected using a validated questionnaire. Information on severe asthma exacerbations with hospital admission were retrieved from hospital notes. All patients underwent spirometry. RESULTS We found 2 SNPs (1 novel rs9635726 in IKZF3) to be associated with asthma. Among children with asthma, 4 SNPs (in ZPBP2, GSDMB, and GSDMA) were associated with hospital admissions and 8 SNPs with lung function. One SNP (rs9635726) remained significantly associated with a predicted forced expiratory volume in 1 second after false discovery rate correction. Nine markers across 5 genes showed interaction with early-life environmental tobacco smoke (ETS) exposure in relation to asthma and 2 with furry pet ownership. Among children with asthma, we observed significant interactions between early-life ETS exposure and 3 SNPs for lung function and among early-life ETS exposure, 3 SNPs (in ORMDL3 and GSDMA), and hospital admission with asthma exacerbation. Three SNPs (in ORMDL3) interacted with current furry pet ownership in relation to hospital admissions for asthma exacerbation. CONCLUSION Our results indicate that several genes in the 17q12-21 region may be associated with asthma. This study confirms that environmental exposures may need to be included into the genetic association studies.

Circulating levels of hormones, lipids, and immune mediators in post-traumatic stress disorder : a 3-month follow-up study

A number of peripheral blood analytes have been proposed as potential biomarkers of post-traumatic stress disorder (PTSD). Few studies have investigated whether observed changes in biomarkers persist over time. The aim of this study was to investigate the association of combat-related chronic PTSD with a wide array of putative PTSD biomarkers and to determine reliability of the measurements, i.e., correlations over time. Croatian combat veterans with chronic PTSD (n = 69) and age-matched healthy controls (n = 32), all men, were assessed at two time points separated by 3 months. Serum levels of lipids, cortisol, dehydroepiandrosterone-sulfate (DHEA-S), prolactin, and C-reactive protein were determined. Multiplex assay was used for the simultaneous assessment of 13 analytes in sera: cytokines [interferon-γ, interleukin (IL)-1β, IL-2, IL-4, IL-6, TNF-α], adhesion molecules (sPECAM-1, sICAM-1), chemokines (IL-8 and MIP-1α), sCD40L, nerve growth factor, and leptin. Group differences and changes over time were tested by parametric or non-parametric tests, including repeated measures analysis of covariance. Reliability estimates [intraclass correlation coefficient (ICC) and kappa] were also calculated. Robust associations of PTSD with higher levels of DHEA-S [F(1,75) = 8.14, p = 0.006)] and lower levels of prolactin [F(1,75) = 5.40, p = 0.023] were found. Measurements showed good to excellent reproducibility (DHEA-S, ICC = 0.50; prolactin, ICC = 0.79). Serum lipids did not differ between groups but significant increase of LDL-C after 3 months was observed in the PTSD group (t = 6.87, p < 0.001). IL-8 was lower in the PTSD group (t = 4.37, p < 0.001) but assessments showed poor reproducibility (ICC = −0.08). Stable DHEA-S and prolactin changes highlight their potential to be reliable markers of PTSD. Change in lipid profiles after 3 months suggests that PTSD patients may be more prone to hyperlipidemia. High intra-individual variability in some variables emphasizes the importance of longitudinal studies in investigations of PTSD biomarkers.

Asthma severity, polymorphisms in 20p13 and their interaction with tobacco smoke exposure.

We investigated the association between genetic variation in chromosomal region 20p13‐p12 (ADAM33 and flanking genes ATRN, GFRA4, SIGLEC1 and HSPA12B) and asthma. Amongst asthmatics, we then investigated the association between genetic variants and asthma severity. We evaluated the effect of environmental tobacco smoke (ETS) exposure in the context of genetic variants.

Decreased Toll-like receptor 8 expression and lower TNF-alpha synthesis in infants with acute RSV infection

BackgroundToll-like receptors (TLRs) are part of the innate immune system, able to recognize pathogen-associated molecular patterns and activate immune system upon pathogen challenge. Respiratory syncytial virus (RSV) is a RNA virus particularly detrimental in infancy. It could cause severe lower respiratory tract disease and recurrent infections related to inadequate development of anti-viral immunity. The reason could be inadequate multiple TLRs engagement, including TLR8 in recognition of single-stranded viral RNA and diminished synthesis of inflammatory mediators due to a lower expression.MethodsIntracellular TLR8 expression in peripheral blood monocytes from RSV-infected infants was profiled and compared to healthy adults and age matched controls. Whether the observed difference in TLR8 expression is a transitory effect, infants in convalescent phase (4-6 weeks later) were retested. Specific TLR8-mediated TNF-α production in monocytes during an acute and convalescent phase was analyzed.ResultsRSV-infected and healthy infants had lower percentage of TLR8-expressing monocytes than healthy adults whereas decreased of TLR8 protein levels were detected only for RSV-infected infant group. Lower protein levels of TLR8 in monocytes from RSV-infected infants, compared to healthy infants, negatively correlated with respiratory frequency and resulted in lower TNF-α synthesis upon a specific TLR8 stimulation. In the convalescent phase, levels of TLR8 increased, accompanied by increased TNF-α synthesis compared to acute infection.ConclusionsLower TLR8 expression observed in monocytes, during an acute RSV infection, might have a dampening impact on early anti-viral cytokine production necessary to control RSV replication, and subsequently initiate an adaptive Th1 type immune response leading to severe disease in infected infants.

Effect of house dust mite immunotherapy on transforming growth factor β1-producing T cells in asthmatic children

BACKGROUND Recent evidence suggests that regulatory T cells (Treg cells) and immunosuppressive cytokines, such as transforming growth factor BETA1 (TGF-BETA1) and interleukin 10 (IL-10), may have a role in clinically effective allergen specific immunotherapy (SIT). OBJECTIVE To evaluate the effect of SIT on the induction of Treg cells in house dust mite-allergic children and on the expression of specific Treg cell markers (cytotoxic T-lymphocyte-associated protein 4 [CTLA-4], IL-10, and TGF-BETA1). METHODS In this uncontrolled open-label study, the percentage of peripheral blood CD4+ Treg cells (CD69 CD45RO+CTLA-4+ and CD3+CD4+CD25+FOXP3+) and the expression of molecules associated with their functions (CTLA-4, TGF-BETA1, and IL-10) were analyzed using flow cytometry in 16 children allergic to house dust mites before and at 3 and 12 months of subcutaneous SIT. Clinical variables, such as symptom score, medication requirements, forced expiratory volume in 1 second, peak expiratory flow rate, and serum IgE levels, were also determined. Ten healthy children were included as controls. RESULTS All the clinical variables improved during immunotherapy. The percentage of CD4+CD25+CD69-CD45RO+ Treg cells remained unchanged. The percentage of CTLA-4+ -expressing Treg cells transiently increased after 3 months of immunotherapy, whereas the percentage of FOXP3+ Treg cells did not change after 1 year of immunotherapy. Levels of IL-10+ cells transiently decreased after 3 months of immunotherapy. Four children who required inhaled fluticasone propionate administration for significant symptom worsening had no statistically significant increase in TGF-BETA1-secreting T cells at 12 months of SIT, in contrast to 12 children without inhaled corticosteroid treatment. CONCLUSIONS The increase in TGF-BETA1-positive T cells only in children without significant symptom worsening requiring inhaled corticosteroid treatment limits the usefulness of TGF-BETA1 in monitoring response to allergen immunotherapy.

Genetic variants in endotoxin signalling pathway, domestic endotoxin exposure and asthma exacerbations

We investigated the interaction between genetic variants in endotoxin signalling pathway and domestic endotoxin exposure in relation to asthma presence, and amongst children with asthma, we explored the association of these genetic variants and endotoxin exposure with hospital admissions due to asthma exacerbations.

The increased type-1 and type-2 chemokine levels in children with acute RSV infection alter the development of adaptive immune responses.

Severe RSV infections and frequent recurrence could be related to the altered polarization of type-2/type-1 T cells. This increases the importance of determining distinctive chemokines and chemokine receptor profiles on memory T cells. We analyzed systemic adaptive T cell response in the acute (n = 17) and convalescent phase (n = 7) of RSV-infected children, in the acute (n = 11) and convalescent phase (n = 6) of children with other viral respiratory infections (adenovirus and influenza virus), and in healthy children (n = 18). Expression of CCR4 and CXCR3 on effector-memory (TEM) and central-memory (TCM) T cells was compared between tested groups. Serum concentrations of specific chemokines were determined. High CXCL10 levels were detected in acutely infected children regardless of virus pathogen, whereas increased CCL17 production was RSV-specific. Higher percentages of CCR4+ CD4 TEM cells in acute RSV infection were accompanied with higher percentages of CXCR3+ CD8 TEM cells, whereas the development of long-lived memory CXCR3+ CD4 and CD8 TCM cells seems to be compromised, as only children with other viral infections had higher percentages in the convalescent phase. Presence of type-2 and type-1 adaptive antiviral immune response, together with insufficient development of long-lived type-1 T cell memory, could play an important role in RSV pathogenesis and reinfection.