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GENE POLYMORPHISM OF IFNG +874 T/A AND TLR4 +896 A/G AND RECURRENT INFECTIONS AND WHEEZING IN TODDLERS WITH HISTORY OF BRONCHIOLITIS

Nuolivirta, Kirsi MD*; Hurme, Mikko MD; Halkosalo, Anne PhD; Koponen, Petri MD§; Korppi, Matti MD§; Vesikari, Timo MD; Helminen, Merja MD§

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The Pediatric Infectious Disease Journal: December 2009 - Volume 28 - Issue 12 - p 1121-1123
doi: 10.1097/INF.0b013e3181af37ee
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Abstract

The link from bronchiolitis to subsequent wheezing and asthma has frequently been reported, but the mechanisms are not known. Viruses, such as rhinovirus, and host immunity have been suggested as contributing factors.1–4 We recently found that IL10 polymorphism at −1082 A/G is associated with bronchiolitis requiring hospitalization in infants under 6 months of age but only when the causative agent is not respiratory syncytial virus (RSV).2 The aim of the present study was to evaluate the connection between polymorphisms of IL10 −1082 A/G, IFNG +874 T/A, IL18 −137 G/C, and TLR4 +896 A/G, and recurrent infections and wheezing in these children during the first 1½ years of life.

PATIENTS AND METHODS

Healthy, full-term infants hospitalized for bronchiolitis at less than 6 months of age between December 1st, 2001 and May 31st, 2002 and between October 28th, 2002 and May 31st, 2004 were recruited in the study. Bronchiolitis was defined as acute lower respiratory illness characterized by rhinorrhea, cough, and diffuse wheezes and rales. The viral etiology of bronchiolitis was determined as described recently.2 Children were invited to a follow-up visit between May 2003 and June 2005. The parents had recorded prospectively the illness history of the child during the follow-up period. At the follow-up visit, a questionnaire was used to obtain data on hospitalizations, breast-feeding, smoking and pets in the family, location of residence, form of day-care, number of siblings, history of allergy in the family, and the allergy status of the child (if diagnosed by the family doctor). Genotyping of IFNG +874 T/A (rs 2430561), IL10 −1082 A/G (rs1800896), IL18 −137 G/C (rs 187238), and TLR4 +896 A/G (rs4986790) gene polymorphisms has been described previously.2 Statistical analyses were performed using the Statistical Package of Social Science. The Ethics Committee of Tampere University Hospital District approved the study. Informed consent was obtained from parents before enrolling the children.

RESULTS

Study Population.

In all, 129 of the 203 children (63.5%) who were enrolled in the initial study participated in the follow-up study. The mean follow-up time of the children after the bronchiolitis episode was 15.6 ± 1.7 months (SD) (range: 11.3–19.5 months).The basic characteristics and the causative agent of the initial bronchiolitis episode of the children are shown in Table 1. The causative virus, place of residence, breast-feeding, parental smoking, keeping of pets in the family, form of day-care, allergy in the family, or number of siblings had no significant association with the number of infections, number of ear infections, use of antibiotics, placement of tympanostomy tubes, number of wheezing episodes, or use of corticosteroids (data not shown). Children with doctor-diagnosed allergy had more wheezing episodes per patient month than nonallergic children (0.10 ± 0.16 [SD] vs. 0.04 ± 0.07 [SD], P = 0.02) and were more likely to receive steroids than nonallergic children (11/32 [34.4%] vs. 13/97 [13.4%], P = 0.008).

T1-23
TABLE 1:
The Demographic and Clinical Characteristics of the Study Children
Gene Polymorphisms and Infection, Wheezing and Use of Corticosteroids.

The polymorphisms of IFNG, IL10, IL18, and TLR4 analyzed showed no association with doctor-diagnosed allergy, total serum IgE, or wheezing history of the child (data not shown). However, IFNG polymorphism at +874 T/A seemed to be associated with susceptibility to infections, so that allele A seemed to have a protective effect (Table 2). Children who were carriers of IFNG allele A were less likely to have tympanostomy tubes inserted than noncarriers (11/83 [13.3%] vs. 13/46 [28.3%], P = 0.037, χ2 test). The number of ear infections per patient month was also less, although not significantly, in children with the allele A (P = 0.07). The polymorphisms of IL10, IL18, and TLR4 studied were not associated with the infection histories (data not shown). However, carriers of TLR4+896 A/G allele G or IL10 −1082 A/G allele G were more likely to have tympanostomy tubes than noncarriers (8/21 [38.1%] vs. 16/108 [14.8%], P = 0.012, and 21/89 [23.6%] vs. 3/40 [7.5%], P = 0.030, respectively, χ2 test). The carriers of IFNG allele A were also less likely to have used corticosteroids for wheezing (10/83 [12.0%] vs. 14/46 [30.4%], P = 0.010, χ2 test), though no association was found with the numbers per patient month of wheezing episodes (Table 2). In multivariate logistic regression, the use of corticosteroids was associated with both allergy (aOR: 4.6 [95% CI: 1.2–18.1]) and IFNG allele A carriage (aOR: 0.23 [95% CI: 0.07–0.83]), with no interaction between the 3 variables (P = 0.88).

T2-23
TABLE 2:
Association of Genotype Frequencies and Allele Carrier Status of Gamma-Interferon (IFNG) +874 T/A With the Number and Number Per Patient Month of Infections, Ear Infections, Use of Antibiotics, and Wheezing Episodes in Children With History of Severe Bronchiolitis

DISCUSSION

The aim of this postbronchiolitis follow-up was to evaluate the association between cytokine gene polymorphisms and subsequent respiratory infections and wheezing during the first 1.5 years of life. We selected the gene polymorphisms of 4 cytokines that have been shown to influence the clinical presentation of bronchiolitis and/or susceptibility to asthma development. Recently, we have shown that infants who were hospitalized for bronchiolitis caused by other viruses than RSV, especially by rhinoviruses, were more likely than controls to be IL10 −1082 allele G noncarriers.2

In recent studies, recurrent wheezing has been common especially after rhinovirus-induced wheezing in infancy.1 In this study, we could not show any significant associations between the causative virus of bronchiolitis and subsequent infections or wheezing, allergic manifestations, or use of antibiotics or corticosteroids during the 1-year follow-up period. However, IFNG polymorphism at +874 seemed to influence the susceptibility to respiratory infections, and TLR4 polymorphism at +896 influenced the need of tympanostomy tube replacement. IFNG allele A at +874 also decreased the likelihood that corticosteroids were needed for postbronchiolitis wheezing.

IFNG is a pro-inflammatory cytokine which also seems to have direct antiviral activity. Production of IFNG is genetically controlled so that allele A at +874 is associated with low expression of IFNG.5 In earlier studies, IFNG allele A at +874 has been shown to decrease the risk of rejection and lung fibrosis and to increase the risk of infections, such as malaria.6,7 In addition, allele T at +874 has been shown to confer protection against chronic hepatitis B infection.8

In this study, allele A at +874 seemed to protect from respiratory infections and significantly decreased the need for corticosteroids used to prevent or treat postbronchiolitis wheezing. In logistic regression, this finding was robust to the adjustment for allergy, suggesting that, since steroids are used for recurrent wheezing, IFNG allele A carriage is an independent protective factor for recurrent wheezing after bronchiolitis. The finding that IFNG allele A at +874, which is associated with low IFNG production, decreased the need for corticosteroids, could be explained by a more controlled inflammatory reaction leading to less wheezing. Strong immune responses not only eliminate infecting organisms but also provoke inflammatory symptoms like wheezing. In addition, viral infections induce bronchial obstruction by neural reflexes,. In animal studies, IFNG has induced or at least enhanced bronchial obstruction by damaging the M2 receptors, which limit the release of acetylcholine from vagal nerve endings.9 This IFNG-mediated bronchial obstruction may be one of the mechanisms leading to recurrent wheezing during viral infections, especially in individuals who have a heightened IFNG response. Therefore, individuals with a more controlled, lower IFNG response may be protected, not from viral infections, but merely from repeated wheezing episodes.

Polymorphism of TLR4 at +896 was associated with the need of tympanostomy tubes so that carriers of allele G were more likely to require the tubes. TLR4 is a transmembrane protein which triggers the innate immune response to endotoxin.4,10TLR4 polymorphism was analyzed in this study, because TLR4 is also part of a receptor complex involved in the immune response against RSV, a major pathogen in bronchiolitis.4 The A/G polymorphism at +896 of TLR4 has been associated with the severity of RSV infection, as well as with lipopolysaccharide responsiveness in septic infections, so that allele G seems to increase susceptibility to severe RSV and gram-negative bacterial infections.4,10 Therefore, the allele G at +896 of TLR4 could predispose children to recurrent or chronic ear infections caused by gram-negative bacteria and also to more severe RSV infections and inflammatory reactions, further predisposing the child to recurrent otitis media and tympanostomy tube insertion. In this study, IFNG allele A at +874 was associated with less need for tympanostomy tubes.

Our results offer preliminary evidence that polymorphisms of IFNG +874 T/A, and of TLR4+896A/G had a significant association with the numbers and presentations of early-life respiratory infections in children with bronchiolitis at less than 6 months of age. Polymorphism at IFNG +874 T/A had no association with the number of postbronchiolitis wheezing episodes, but allele A carriage decreased the risk of repeated wheezing requiring treatment by corticosteroids. This finding was confirmed by multivariate analyses adjusted for allergy. The findings of this study have to be interpreted cautiously, since only 64% of the patients eligible for the study attended the follow-up visit, and only indirect outcome measures, the use of corticosteroids reflecting severe or repeated wheezing and the insertion of tympanostomy tubes reflecting repeated otitis media complicating respiratory infections, gave indisputably statistically significant results.

REFERENCES

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10.Lorenz E, Mira JP, Frees KL, et al. Relevance of mutations in the TLR4 receptor in patients with gram-negative septic shock. Arch Intern Med. 2002;162:1028–1032.
Keywords:

bronchiolitis; cytokine polymorphism; TLR4; recurrent infections

© 2009 Lippincott Williams & Wilkins, Inc.