Egypt J Pediatr Allergy Immunol 2012;10(2):81-86. Original article
Effects of β2-adrenergic receptor polymorphisms on asthma severity and response to salbutamol in Egyptian children Background: Several polymorphisms of the β2-adrenergic receptor (ADRB2) Khalid Salah, gene have been identified. There is mounting evidence that these polymorphisms Saed Morsy, are associated with significant variability in response to bronchodilator therapy and thus in severity and duration of asthmatic symptoms. Amal Atta * Objectives: to assess the frequency of ADRB2 polymorphisms at codon 16 in Egyptian asthmatic children and to study the association of these polymorphisms with asthma severity and response to inhaled salbutamol. Methods: This case-control study Departments of was conducted at pulmonology unit, Zagazig University children’s hospital Pediatrics and during the period from December 2010 to December 2011. One hundred Microbiology*, Faculty children (50 asthmatics and 50 controls) were enrolled into the study. For all of Medicine, Zagazig study population, detailed history taking, systematic physical examination, chest University, Egypt. x-ray, pulmonary function testing and ADRB2 genotyping were performed. Results: There was a significant increase in frequencies of Arg/Gly and Gly/Gly genotypes among asthmatic children in comparison with healthy controls (OR = 7.9; CI: 0.94-67.4, P<0.05 and OR= 4.5;CI: 0.91-22.7, P<0.05 respectively). On the other hand, there was a lower frequency of Arg/Arg genotype in asthmatic children than that in healthy controls (OR = 0.14; CI: 0.04-0.55, P<0.05). Moreover, statistical analysis revealed association of Arg/Arg genotype with mild asthma (OR = 5.77; 95% CI: 1.55-21.5, P<0.05) and association of Gly/Gly genotype with moderate/severe asthma (OR=0.057; 95% CI: 0.006-0.516, P<0.05). However, no difference in distribution of Arg/Gly genotypes among mild and moderate/severe asthmatics (OR = 0.79; 95% CI: 0.156-3.99, P>0.05). Regarding bronchodilator responsiveness, Gly/Gly and Arg/Gly genotypes were associated with reduced response, while Arg/Arg Correspondence: genotype was associated with favorable response to nebulized salbutamol. Conclusion: Polymorphisms of ADRB2 at codon 16 may be a determinant of asthma severity and response to salbutamol in Egyptian asthmatic children. Pediatrics, Faculty of Further studies are needed to demonstrate effects of other polymorphisms of ADRB2 gene on these outcomes. Keywords: Polymorphism; β2-adrenergic receptor (ADRB2) gene; asthma; Sharkiah, Egypt. Egyptian children; bronchodilator therapy.
Asthma aggregates within families and is a
Asthma is a disease characterized by hyper-
involvement of environmental and genetic
responsiveness of the airways to various stimuli,
components5. Understanding of the genetic basis of
which results in airway obstruction that is reversible
asthma may contribute toward identifying better
either spontaneously or as a result of treatment1.
Asthma prevalence has increased very considerably
Inhaled β2-adrenergic agonist medications are
in recent decades such that it is now one of the
the foundation of therapy for acute asthma
commonest chronic disorders in the world2. Asthma
exacerbation7. The β2-adrenergic receptor
is estimated to affect 300 million people world-
(ADRB2) protein is expressed on bronchial smooth
wide, with an expected increase to 400 million
muscle cells and mediates physiologic responses
worldwide by 20253. In a recent Egyptian study, the
including bronchodilation, vasodilatation and
overall prevalence of wheezing in the last year was
lipolysis8. ADRB2 gene located on chromosome
14.7% and of physician-diagnosed asthma was
5q31-32 is one of the candidate genes most
consistently identified as being associated with asthma-related phenotypes 9.
Several polymorphisms of ADRB2 have been
years. Subjects were included as controls only if
described in particular at codons 16 they reported no history of asthma or allergies, no Arginine/Glycine and 27 Glutamine /Glutamic acid
history or report of having experienced symptoms
which alter the receptor function in vitro10,11. In
of coughing, wheezing and shortness of breath in
adults, genetic variations in this receptor have been
the past 2 years and no other history of lung
linked to asthma severity12, bronchial hyper-
diseases, chronic illnesses, or medications.
responsiveness13 and lung function tests14.
For all study groups, detailed history taking,
The relationship between ADRB2 genotypes
systematic physical examination, chest x-ray,
and response to inhaled β2-adrenergic agonists is
pulmonary function testing and ADRB2 genotyping
controversial. Some studies15-21 have found that the
were performed. Informed consent was obtained
Arg/Arg genotype is associated with reduced from the children’s guardians. The study protocol response to these medications, whereas others22-25
was approved by the Pediatric Department Ethics
have found that the Gly/Gly genotype is associated
Knowledge of the genotype which is Three milliliters of venous blood were collected
associated with favorable response, could aseptically from every subject in a tube containing significantly affect treatment selection for asthma in
EDTA. This blood was incubated at -20°C
children. The current study will focus on the
temperature. After samples were enough, the blood
polymorphisms of ADRB2 gene at codon 16 as a
was prepared for DNA extraction and amplification.
previous study22 found no association between
DNA extraction was done using Axyprep Blood
polymorphisms at position 27 and response to
Genomic DNA Miniprep Kit (Axygen Biosciences,
inhaled β2-adrenergic agonists in asthmatic USA). DNA amplification was carried out by using children.
polymerase chain reaction (PCR) Master-Mix Gold
So, the objectives of this study were to assess
the frequencies of ADRB2 polymorphisms at codon
sequences of the forward and reverse prime used for
16 in Egyptian asthmatic children and the effects of
these polymorphisms on asthma severity and 5'-GCCTTCTTGCTGGCACCCCAT-3' and bronchodilator response to salbutamol. 5'-CAGACGCTCGAACTTGGCCATG-3'.
Then, for detection of the ADRB2 polymorphisms,
This case-control study was conducted at Pulmonary function testing: pulmonology unit, Zagazig University children's
Pulmonary function tests were performed using
hospital during the period from December 2010 to
Lilly Pneumatocho- graph (D-97204 Hochberg,
December 2011. One hundred children were Germany). Spirometry was performed according to enrolled in the study. They were divided into two
the American Thoracic Society standards27.
Spirometeric measurements included forced
Group I (asthmatic group) included 50 expiratory volume in one second (FEV1), forced
children, 26 males and 24 females. Their ages
vital capacity (FVC) and peak expiratory flow
ranged from 5-12 years with a mean value of 6.8
years. The diagnosis of asthma in these patients was
Pulmonary function test results were expressed
based on National Asthma Education and as a percentage of the predicted normal value. Prevention Program Guidelines26. Cases were Asthmatic children were instructed to stop any classified and subgrouped as follow: mild systemic bronchodilator or corticosteroid therapy intermittent and mild persistent asthmatics were
72 hours before tests and short acting β2-adrenergic
included into the group of mild asthma and
agonists were stopped 12 hours before the
moderate persistent and severe persistent asthmatics
were included as the group of moderate/severe
To assess response to a bronchodilator in
asthma. Recruited asthma patients were not in
asthmatic group, one dose of salbutamol (0.15
exacerbation state and did not have other mg/kg) was administered using ultrasonic nebulizer concomitant diseases that might affect lung (eMed A100, Italy) and measurement of lung function. Children with any chronic disease (other
function was performed before and 15 minutes after
salbutamol nebulization. Response to salbutamol
Group II (control group) included 50 healthy
was reported as a percent change in FEV1 between
children, 26 males and 24 females. Their ages
baseline and after salbutamol administration. We
ranged from 5-12 years with a mean value of 7.2
chose to explore changes in FEV1 after salbutamol
ADRB2 polymorphisms in asthmatic children
mobilizations as a measure of response to a
distribution between asthmatic and healthy children
bronchodilator, because this was the most objective,
immediate, and most common endpoint studied by
Distribution of ADRB2 gnenotypes at codon
16 among asthmatic and non-asthmatic children is
illustrated in table 2. There was a significant
increase of carriers of Arg/Gly and Gly/Gly
Data were analyzed using Statistical Package for
genotypes among asthmatic children in comparison
Social Sciences (SPSS), release 16. The to controls (OR = 7.9; CI: 0.94 - 67.4, P<0.05 and quantitative variables were expressed as means and
OR= 4.5; CI: 0.91-22.7, P<0.05 respectively). On
standard deviations. For comparison between two
the other hand, there was a lower frequency of
group means, t-test was used. For comparison
Arg/Arg genotype in asthmatic children than in
between three group means, one way ANOVA
controls (OR = 0.14; CI: 0.04- 0.55, P<0.05).
(analysis of variance) was used. Qualitative
Relation between polymorph-isms of ADRB2
variables were expressed by frequency and at codon 16 and asthma severity is shown in table 3. percentage and compared using chi-square test.
Statistical analysis revealed an association of
Also, odds ratio (OR) and 95% confidence interval
Arg/Arg genotype with mild asthma (OR= 5.77;
(CI) were calculated. For all tests a probability (P)
95% CI: 1.55-21.5, P<0.05). On other hand,
less than 0.05 was considered significant and less
Gly/Gly genotype was less frequent in mild asthma
than 0.001 was considered highly significant.
(OR= 0.057; 95% CI: 0.006-0.516, P<0.05).
However, no difference in distribution of Arg/Gly
The characteristics of the two study groups are
moderate/severe asthmatics (OR=0.79; 95% CI:
shown in table 1. The two groups were similar in
age and gender. A highly significant difference was
Regarding bronchodilator responsiveness,
observed in spirometric parameters between Gly/Gly and Arg/Gly genotypes were associated asthmatic and healthy children (P<0.001). Also,
with reduced response (FEV1change =11.4% and
significant difference was observed in genotype
9.1%, respectively), while Arg/Arg genotype was associated with favorable response (FEV1change = 17.3%) (Table 4).
Table 1. Demographic and clinical data of the study groups. Variable Asthmatic group Control group Age( year) , Mean ±SD Male gender, n (%) Genotype, n (%) Spirometric parameters, Mean±SD Asthma severity, n (%) Table 2. Distribution of ADRB2 genotypes at codon 16 in asthmatic and healthy children. Asthmatic group Control group Genotype Table 3. Relation between polymorphisms of β2AR at codon 16 and asthma severity. Moderate/ Mild asthma Genotype severe asthma Table 4. Relation between polymorphisms of β2AR at codon 16 and response to salbutamol
nebulization. Percentage of change of FEV1 (%), Mean±SD
FEV1: forced expiratory volume in one second , a Arg/Arg group vs. Arg/Gly group , b Arg/Arg group vs. Gly/Gly group, c Arg/Gly group vs. Gly/Gly group
moderate/severe asthma. On the other hand,
Genetic assessment revealed that asthmatic children
had frequencies of Arg/Arg of 70%, Arg/Gly of
moderate/severe asthma when compared with mild
14%, and Gly/Gly of 16%, while in the healthy
asthmatics. This indicates that the polymorphism at
children the frequency of Arg/Arg was 94%,
codon 16 of ADRB2 is a possible determinant of
Arg/Gly was 2%, and Gly/Gly was 4%. These
results revealed a significant difference of
A meta-analysis including a total of 28 studies
distribution of ADRB2 genotypes at codon 16
performed by Contopoulos-Ioannidis et al.32,
concluded that,Gly/Glyhad a much higher risk for
Globally, there are marked interethnic nocturnal asthma and asthma severity than the
differences in the frequency of ADRB2 Arg/Arg. polymorphisms. In Caucasian–American and
In contrast, Salama et al.30 found an association
African–American healthy individuals, the Arg/Gly
between Arg/Gly genotype with severe asthma
genotype was the most predominant (38.3% and
when compared to mild/moderate asthma. Also,
50.4% respectively), while Arg/Arg genotype was
present in lower frequencies (26.6% and 23.6%,
frequencies in severe asthma when compared with
respectively)28. In another study conducted on 128
Chinese asthmatics by Wang et al., the frequency of
In vitro functional studies33,34 indicated that
Arg/Arg was 40.6%, Arg/Gly was 42.2%, and
down-regulation of ADRB2 receptors occurs in
Gly/Gly was 17.2%, while in the healthy people the
individuals expressing Gly16 allele in response to
frequency of Arg/Arg was 27.9%, Arg/Gly was
circulating catecholamines or exogenously
administered β2-agonists. The Arg16 allele, which
In Egyptian population, Salama et al.30 found
demonstrates resistance to down-regulation, might
that the frequencies of ADRB2 genotypes at
therefore be expressed at greater levels than the
position 16 among healthy children, was 52.6% for
Gly16 allele within airways. Accordingly,
Arg/Arg, 5.3% for Arg/Gly and 42.1% for Gly/Gly.
individuals carrying the Gly/Gly genotype might be
In asthmatic children, these genotype frequencies
were different; 17.5% for Arg/Arg, 45% for
bronchoconstriction, and therefore have more
Arg/Gly and 37.5% for Gly/Gly. Although these
reactive airways than individuals carrying the
frequencies were different from our results, yet the
final conclusion was the same: higher frequencies
In our study, the Gly allele (Arg/Gly and
of Arg/Gly and Gly/Gly genotypes in asthmatic
Gly/Gly genotypes) was associated with resistance
children when compared with controls. In another
to the bronchodilator effect of inhaled short-acting
study, the frequencies of ADRB2 genotypes at
β 2-agonist (salbutamol) when compared with
position 16 among Egyptian healthy individuals,
Arg/Arg genotype. The association between
was 32.6% for Arg/Arg, 49% for Arg/Gly and
ADRB2 genotypes and response to inhaled β2
18.5% for Gly/Gly31. These differences reflect the
agonists is controversial, and discordant findings
need for wide-based, population studies.
have been reported. Finkelstein et al.22 conducted a
There was an association of Arg/Arg genotype
meta-analysis to examine the association between
with mild asthma when compared with ADRB2 polymorphisms and the response to inhaled
ADRB2 polymorphisms in asthmatic children
β2 -adrenergic agonists in children with asthma.
3. Masoli M, Fabian D, Holt S, Beasley R. The
They included 3 studies9,35,36 and found a significant
global burden of asthma: executive summary of the
association between favorable therapeutic response
GINA Dissemination Committee report. Allergy
to inhaled β2-adrenergic agonists and the Arg/Arg
genotype. Several studies came up with the same
4. Georgy V, Fahim HI, El Gaafary M, Walters S.
Prevalence and socioeconomic associations of asthma
However, when examining the influence of
and allergic rhinitis in northern Africa. Eur Respir J 2006; 28:756-62
ADRB2 polymorphisms on the response to long-
term and repeated dosages of inhaled β2 -adrenergic
5. Bijanzadeh M, Mahesh PA, Ramachandra NB.
An understanding of the genetic basis of asthma.
agonist therapy, the Arg/Arg genotype has been
more closely associated with reduced response15-20.
Moreover, in children with severe asthma
Hall IP. The future of asthma. BMJ 1997;
exacerbations, Carroll et al.21 found that children
whose genotypes were Gly/Gly had a more rapid
Baren JM, Zorc JJ. Contemporary approach to the
emergency department management of pediatric
response to inhaled β2 -adrenergic agonists.
asthma. Emerg Med Clin North Am 2002; 20:115–
discordant results reported by different authors.
8. Insel PA. Seminars in medicine of the Beth Israel
First, studies had included different patient
Hospital, Boston. Adrenergic receptors-evolving
populations, regarding age, disease severity, and
concepts and clinical implications. N Engl J Med
administered different β2-agonists to study patients.
9. Silverman EK, Kwiatkowski DJ, Sylvia JS,
Third, authors had used different outcome measures
Lazarus R, Drazen JM, Lange C, et al. Family-
and endpoints to assess drug-responsiveness.
based association analysis of beta2-adrenergic
Fourth, some authors had suggested that the
receptor polymorphisms in the childhood asthma
conflicting results among studies could be
management program. J Allergy Clin Immunol 2003;
polymorphisms that are commonly inherited 10. Giubergia V, Zelazko M, Roy A, Gravina LP, together (ADRB2 haplotypes), rather than by a
Gonzalez Pena H, Chertkoff L. Beta 2-adrenergic
single allele polymorphism37. Finally, race is both
polymorphisms and total serum IgE levels in children with asthma from Argentina. Ann Allergy Asthma
a biologic and a social construct and constitutes not
only genetic differences in individuals, but also the
behaviors, beliefs, and experiences that vary among
Qiu YY, Zhang XL, Yin KS . Association between
beta 2-adrenergic receptor genetic polymorphisms
and total serum IgE in asthmatic patients of Chinese
Han nationality. Respiration 2006;73(2):180-4.
ADRB2 gene at codon 16 may be a determinant of
12. Weir TD, Mallek N, Sandford AJ, Bai TR,
asthma severity and response to salbutamol in
Awadh N, Fitzgerald JM, et al. Beta2-Adrenergic
Egyptian asthmatic children. However, we cannot
receptor haplotypes in mild, moderate and fatal/near
exclude the possibility that other polymorphisms or
fatal asthma. Am J Respir Crit Care Med 1998;
complex haplotypes within the promoter and coding
regions of the ADRB2 gene or adjacent genes
13. Hall IP, Wheatley A, Wilding P, Liggett SB.
might contribute to the present results. Further
Association of Glu 27 beta 2-adrenoceptor
studies are needed to demonstrate effects of other
polymorphism with lower airway reactivity in
polymorphisms of ADRB2 gene on asthma-related
asthmatic subjects. Lancet 1995; 345: 1213– 4.
14. Summerhill E, Leavitt SA, Gidley H, Parry
R, Solway J, Ober C. Beta 2-Adrenergic receptor
arg16/ arg16 genotype is associated with reduced lung function, but not with asthma, in the Hutterites.
Am J Respir Crit Care Med 2000;162:599–602.
1. Weinberger M, Abu-Hasan M. Pseudo-asthma:
15. Taylor DR, Epton MJ, Kennedy MA, Smith AD,
when cough, wheezing, and dyspnea are not asthma.
Iles S, Miller AL, et al. Bronchodilator response
in relation to β2-adrenoceptor haplotype in patients
2. Anandan C, Nurmatov U, van Schayck O,
with asthma. Am J Respir Crit Care Med 2005;
Sheikh A. Is the prevalence of asthma declining?
Systematic review of epidemiological studies.
16. Lipworth BJ, Hall IP, Tan S, Aziz I, Coutie W.
28. Xie HG, Stein CM, Kim RB. Frequency of
Effects of genetic polymorphism on ex vivo and in
functionally important β2 adrenoreceptor
vivo function of the β2-adrenoceptors in asthmatic
polymorphisms varies markedly among African–
American, Caucasian and Chinese individuals.
17. Israel E, Drazen JM, Liggett SB, boushey HA,
Cherniac RM, Chinchilli VM, et al. The effect of
29. Wang Z, Chen C, Niu T, Wu D, Yang J, Wang
polymorphisms of the β2-adrenergic receptor on the
B, et al. Association of Asthma with β2-Adrenergic
response to regular use of albuterol in asthma. Am J
Receptor Gene Polymorphism and Cigarette
Respir Crit Care Med 2000; 162:75– 80.
Smoking. Am J Respir Crit Care Med 2001; 163:
18. Taylor DR, Drazen JM, Herbison GP, Yandava
CN, Hancox RJ, Town GI. Asthma exacerbations
30. Salama MS, Ashaat NA, Hamad AA. Genetic
during long term-agonist use: influence of β2-
association between common beta-2 adrenoreceptor
adrenoceptor polymorphism. Thorax 2000; 55:762–7.
polymorphism and asthma severity in school-age
19. Israel E, Chinchilli VM, Ford JG, Boushey
children. Egyptian Journal of Medical Human
HA, Cherniack R, Craig TJ. Use of regularly
Genetics; 12: 151-6.
scheduled albuterol treatment in asthma: genotype-
31. Hamdy S, Hiratsuka M, Narahar K, El-Enany
stratified, randomized, placebo-controlled cross-over
M, Moursi N, Ahmed MS, et al. Allele and
genotype frequencies of polymorphic DCP1, CETP,
20. Palmer CA, Lipworth BJ, Lee S, Ismail
ADRB2 and HTR2A in Egyptian population. Eur J
T, Macgregor DF, Mukhop-adhyay S. Arginine-
16 β-2 adrenoceptor genotype predisposes to
32. Contopoulos-Ioannidis DG, Manoli EN,
exacerbations in young asthmatics taking regular
Ioannidis JP. Meta-analysis of the association of
β2-adrenergic receptor polymorphisms with asthma
21. Carroll CL, Stoltz P, Schramm CM, Zucker AR.
phenotypes. J Allergy Clin Immunol 2005; 115:963–
Beta 2-adrenergic receptor polymorphisms affect
response to treatment in children with severe asthma
33. Green SA, Turki, J, Innis, M, Liggett SB. Amino-
exacerbations. Chest 2009;135(5):1186-92.
terminal polymorphisms of the human β2-adrenergic
22. Finkelstein Y, Bournissen FG, Hutson JR,
receptor impart distinct agonist-promoted regulatory
Shannon M. Polymorphism of the ADRB2 gene and
properties. Biochemistry 1994;33: 9414- 9.
response to inhaled beta-agonists in children with
34. Green SA, Cole G, Jacinto M, Innis M, Liggett
asthma: a meta-analysis. J Asthma 2009; 46:900–5.
SB. A polymorphism of the human β2-adrenergic
23. Kotani Y, Nishimura Y, Maeda H, Yokoyama M.
receptor within the fourth transmembrane domain
β2-adrenergic receptor polymorphisms affect airway
alters ligand binding and functional properties of the
responsiveness to salbutamol in asthmatics. J Asthma
receptor. J Biol Chem 1993; 268: 23116-21.
35. Martinez FD, Graves PE, Baldini M, Solomon
24. Lima JJ, Thomason DB, Mohamed MH, Eberle
S, Erickson R. Association between genetic
LV, Self TH, Johnson JA. Impact of genetic
polymorphisms of the β2-adrenoceptor and response
polymorphisms of the β2-adrenergic receptor on
to albuterol in children with and without a history of
albuterol bronchodilator pharmacodynamics. Clin
wheezing. J Clin Invest 1997; 100: 3184 –8.
36. Fu J, Chen H, Hu L, Zhang H, Ma Y, Chen Y.
25. Cho SH, Oh SY, Bahn JW, Choi JY, Chang
Association between the genetic polymorphisms of
beta2-adrenergic receptor gene and the asthma
bronchodilating response to short-acting beta-agonist
susceptibility and clinical phenotypes in a Chinese
and non-synonymous single-nucleotide population. Zhonghua Yi Xue Yi Chuan Xue Za Zhi polymorphisms of beta-adrenoceptor gene. Clin Exp
37. Hung CC, Tai JJ, Lin CJ, Lee MJ, Liou HH.
26. National Asthma Education and Prevention Program.
Complex haplotypic effects of the ABCB1 gene on
Expert Panel Report 3 (EPR-3): Guidelines for the
epilepsy treatment response. Pharmacogenomics
Diagnosis and Management of Asthma-Summary
Report 2007. J Allergy Clin Immunol 2007;120 : S94-138.
27. American Thoracic Society. Standardization of
spirometry, 1994 update. Am J Respir Crit Care Med 1995; 152:1107–36.
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