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Keywords:

  • IgE-mediated allergy;
  • nitric oxide synthase;
  • NOS;
  • polymorphism;
  • steroid

Allergic diseases are considered to be multifactorial, involving complex interactions between genetic and environmental influences. There is increasing evidence that nitric oxide (NO) plays a key role in regulation of airway function and in the pathophysiology of inflammation in IgE-mediated allergic diseases (1). NO is produced in increased amounts in asthma and allergic rhinitis. NO in exhaled air correlates with asthma severity, sputum eosinophils, and with peak flow variability, probably related to airway inflammation (2). That is why the gene for endothelial NO synthase (ecNOS; endothelial constitutive nitric oxide synthase), one of three NO synthases, seems to be a candidate gene. The gene encoding ecNOS is located on chromosome 7p35–36 and a number of polymorphisms have been identified in this gene. A 27-bp repeat in intron 4 has been associated with atopic asthma in Koreans (3).

Our study was carried out (1) to determine whether the allele and genotype distributions in childhood patients with IgE-mediated allergic diseases in our Czech population differ from an earlier study; and (2) to see if this polymorphism is associated with the severity or intermediary phenotype markers of allergic diseases.

This was a cross-sectional study of 66 pediatric patients with IgE-mediated asthma and rhinitis and their combinations (45 boys, 21 girls; 14 ± 6 years). Children were investigated and treated at the Faculty Children Hospital Brno, according to standard protocols (4,5), using adequate controls. IgE-mediated allergy was recognized by specific IgE or positive skin prick test to any one or more allergens as described previously (6).

Project approval was granted by the local ethics committee at Medical Faculty MU Brno and written informed consent for genetic analysis was obtained from parents of all subjects.

We used peripheral blood leukocytes for DNA extraction. Then the 27-bp repeat polymorphism in intron 4 of the ecNOS gene was detected using our modified PCR method according to Lee et al. (3). An individual's genotype was evaluated as homozygote (bb), 602 bp (five repeats of the 27-bp); heterozygote (ab), 602 and 575 bp; or homozygote (aa), 575 bp (four repeats of the 27-bp).

We detected 6.1% (n = 4) subjects homozygous for the deletion (ecNOS 4a/a), 31.8% (n = 21) heterozygotes (ecNOS 4a/b) and 62.1% (n = 41) homozygotes for the common insertion allele (ecNOS 4b/b). There was a statistically significant difference in the frequencies of individual alleles (P = 0.0018) and the genotypes (P = 0.0108) of this polymorphism between patients whose clinical condition required compensation using topical corticoids (inhaled or nasal) (allele: a/b = 0.316/0.684; genotype: aa = 4 (10.5%), ab = 16 (42.1%), bb = 18 (47.4%)) and children whose compensation was achieved without corticoid therapy (allele: a/b = 0.089/0.911, genotype: aa = 0 (0%), ab = 5 (17.9%) and bb = 23 (82.1%)). We failed to find any association of genotype with intermediary phenotype.

Genetic, as well as environmental, factors may contribute not only to the manifestation of the disease, but also to its severity. The polymorphism studied here is responsible for part of the genetic variability in the control of the plasma NO levels (lower by about 20% in the aa homozygotes as compared to bb homozygotes) (7).

Our study ascertained the bb genotype in 62% of allergic patients, a much lower occurrence than in the Korean extrinsic asthmatics (86%), and even less than the Korean controls (75%) (3). More detailed analysis of our allergic children revealed statistically significant differences in the frequencies of alleles and the genotype combinations between the group of patients requiring inhaled or nasal steroid treatment (the frequency of the rarer a allele exceeding 30%, the frequency of the bb genotype being only 47%) and the group not requiring this therapy (allele a only 9%, genotype bb 82%). Thus we can speculate that although the manifestation of IgE-mediated allergic disease can be associated with the bb genotype (3), the a variant (or genotype aa) in the manifest allergic disease relates to a persisting airway inflammation where the clinical condition must be compensated by applying inhaled corticosteroids, i.e. a higher degree of therapy.

The discrepancy between the ‘risk’ allele associated by the Korean authors and our study is interesting; it could be related to genetic heterogeneity of allergic asthma and rhinitis which could, in addition, be population dependent. Reproduction of our findings in other populations could suggest whether association between the risk factors (genes in this case) and manifestation of IgE-mediated allergic disease (or its severity) can be generalized.

The study was funded from project CEZ: 307/98: 141100002 provided by the Ministry of Education, Youth and Physical Training of the Czech Republic.

References

  1. Top of page
  2. References
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    Barnes PJ. Nitric oxide and airway disease. Ann Med 1995;57:389393.
  • 2
    Lim S, Jatakanon A, John M, Gilbey T, O'Connor BJ, Chung KF, Barnes PJ. Effect of inhaled budesonide on lung function and airway inflammation. Assessment by various inflammatory markers in mild asthma. Am J Respir Crit Care Med 1999;57:2230.
  • 3
    Lee YC, Cheon KT, Lee HB, Kim W, Rhee YK, Kim DS. Gene-polymorphisms of endothelial nitric oxide synthase and angiotensin converting enzyme in patients with asthma. Allergy 2000;57:959963.
  • 4
    van Cauwenberge P, Bachert C, Passalacqua G, Bousquet J, Canonica GW, Durham SR, Fokkens WJ, Howarth PH, Lund V, Malling HJ, Mygind N, Passali D, Scadding GK, Wang DY. Consensus statement on the treatment of allergic rhinitis. European Academy of Allergology and Clinical Immunology. Allergy 2000;57:116134.
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    Von Mutius E. Presentation of new GINA guidelines for paediatrics. Clin Exp Allergy 2000;57:610.
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    Izakovièová Hollá L, Vaškù A, Izakoviè V, Znojil V. The interaction of the polymorphisms in transporter of antigen peptides (TAP) and lymphotoxin α (LT-α) genes and atopic diseases in the Czech population. Clin Exp Allergy 2001;57:14181423.
  • 7
    Tsukada T, Yokoyama K, Arai T, Takemoto F, Hara S, Yamada A, Kawaguchi Y, Hosoya T, IIgari J. Evidence of association of the ecNOS gene polymorphism with plasma NO metabolite levels in humans. Biochem Biophys Res Commun 1998;57:190193.

Accepted for publication 30 November 2001