Interleukin-13 +2044 G/A and +1923C/T polymorphisms are associated with asthma susceptibility in Asians

Abstract A number of studies have reported that the interleukin 13 (IL-13) gene is associated with asthma susceptibility. However, the reported relationships between the +2044G/A and +1923C/T polymorphisms and asthma susceptibility are inconsistent, especially in Asian adults and children with atopic status. Meta-analysis was used to analyze combined data. The +2044G/A and +1923C/T polymorphisms were investigated using data from 18 and 11 studies, respectively. The results suggested that there was an association between asthma and the IL-13 +2044G/A polymorphisms: odds ratio (OR) 1.34, 95% confidence interval (CI) 1.03–1.75 for AA versus GG + GA and +1923C/T; OR 1.50, 95% CI 1.26–1.78 for TT versus CC; and OR 1.15, 95% CI 1.10–1.21 for TC versus CC. The subgroup meta-analysis demonstrated that IL-13 +2044G/A polymorphisms are associated with asthma: OR 1.47, 95% CI 1.06–2.04 for AA versus GG + GA and +1923C/T; OR 1.70, 95% CI 1.26–2.30 for TT versus CC; and OR 1.27, 95% CI 1.03–1.56 for TC versus CC. In particular, IL-13 +2044G/A polymorphisms are specifically associated with Asian ethnicity in both adults and children with atopic status. However, the 1923C/T polymorphisms were not significantly associated with age group or atopic status within the Asian subgroups. Further investigation using larger samples and meta-analysis is required. No publication bias was detected. This meta-analysis indicates that the IL13 +2044G/A and +1923C/T polymorphisms are risk factors for asthma, especially among Asians.


Introduction
Asthma is a chronic inflammatory disease in which many cells of the innate and adaptive immune systems act together with inflammatory cells to cause bronchial hyper-reactivity (BHR), airway smooth muscle contraction, mucus overproduction, and airway remodeling. Worldwide, more than 300 million people are affected by asthma. The total cost of the disease is estimated to exceed $18 billion annually in the USA, including direct medical costs and indirect costs of lost productivity. [1] The asymmetric functions of T helper cells 1 (Th1) and 2 (Th2) play an important role in causing asthma. [2,3] Mice lacking key Th2 cytokines interleukin (IL)-4, IL-5, or IL-13 have reduced inflammation reactions in asthma features in the ovalbumin model. [4] IL-13 is important for BHR and for goblet cell metaplasia, which can produce thick mucus containing the releasing mucins MUC5AC and MUC5B, which obstruct the airway lumen. [5,6] It is well-known that single-nucleotide polymorphisms (SNP) can be used to assess genetic disorders, on which basis multiple phenotypes have been associated with altered levels of IL-13. Two SNPs, positioned at regions +2044G/A and +1923C/T, have been linked to effective production of IL-13. Heinzmann et al [7] first reported an association between the IL13 G+2044A variant and high asthma risk in Japanese and British populations. Recent studies have suggested that IL-13 +1923C/T polymorphism plays a critical role in the development of asthma. Although +2044G/A polymorphisms have been correlated with asthma susceptibility in several studies, [8][9][10] neither the influences of age nor Asian populations have been described. One study [11] reported a relationship between +1923C/T polymorphisms and asthma susceptibility; however, the influences of atopic phenotypes and age were not analyzed in Asians. Thus, pooled analysis of all available studies is necessary.
In this study, we sought to determine the association between IL-13 +2044G/A and +1923C/T polymorphisms and asthma susceptibility in Asian specifically, by including recently published studies, to evaluate the pathogenesis of asthma from a novel perspective.

Search strategy and data extraction
This meta-analysis was not involved in an ethics committee or institutional review board. We considered all studies that examined the association between IL-13 and asthma, identified using PubMed, EMBASE, and China National Knowledge Infrastructure database (Wanfang database) from 2001 to 2014. Search terms were as follows: (gene or allele or polymorphism) and (IL-13), (+2044G/A), or (+1923C/T and asthma). Searches were performed in duplicate by 2 independent reviewers (QHM and JJQ). Only English and Chinese-language papers were included.
To explain the relationship between IL-13 and asthma susceptibility in Asians, different genotypes of +2044G/A and +1923C/T polymorphisms were extracted. The following information was extracted from each study: author, publication year, country, ethnicity, age group, positions, atopic status, and the number of cases. Disagreements were resolved by discussion between the authors.

Inclusion and exclusion criteria
All selected studies were evaluated for the following inclusion criteria: evaluation of the +2044G/A and +1923C/T polymorphisms in IL-13 gene and asthma risk; inclusion of odds ratio (OR) and 95% confidence interval (CI) estimates; use of a casecontrol design. Studies were excluded on the basis on the following criteria: not relevant to IL-13 +2044G/A and +1923C/ T polymorphism or asthma risk; reviews or abstracts; sample size not reported; and animal study. For overlapping studies, the most recent or largest-sample report was selected.

Assessment of quality scores
The quality of the included studies was evaluated by JJQ using a standardized quality assessment scoring system for studies of genetic association with asthma, as recommended by Thakkinstian et al. [12] The criteria covered the representativeness of cases and controls, ascertainment of cases and controls, genotyping examination, Hardy-Weinberg equilibrium (HWE), association assessment, and response rate. Total scores ranged from 0 (worst) to 15 (best). Any disagreement was adjudicated by a third investigator (JJQ). Low-quality studies with scores of 4 or lower were excluded from analyses.

Study characteristics
The process of selection is shown in Fig. 1. A total of 712 articles were chosen after searching and screening. Ultimately, 25 eligible case-control studies were included, all of which assessed the relationship between IL-13 +2044G/A and +1923C/T polymorphism and asthma risk; the studies represented 16,917 asthmatic cases and 32,522 controls.  Of these 25 studies, 18 reported IL-13 +2044G/A and 11 reported IL-13 +1923C/T. Eleven studies were conducted on adults, 7 on children, and 4 on both adults and children. Regarding ethnicity among the included studies, 15 studies examined Asians, 9 studies examined Caucasians, and 1 study examined Africans. The major characteristics of each group are in Table 1. Genotype numbers and results of +2044G/A and +1923C/T HWE examination are listed in Tables 2 and 3. 3.2. Meta-analysis of available data 3.2.1. IL-13 +2044G/A. Table 4 shows results of the metaanalysis for IL-13 +2044G/A and asthma. We analyzed the OR1 (genotype AA vs GG), OR2 (genotype GA vs GG), and OR3 (genotype AA vs GA); we choose the recessive model of analysis (genotypes AA vs GG+GA; OR 1.34, 95% CI 1.03-1.75, P = .031), which indicated that IL-13 +2044G/A increased risk of asthma (Fig. 2). To further evaluate the relationship between IL-13 +2044G/A and asthma, we conducted subgroup analyses. IL-13 +2044G/A was significantly associated with Asians (OR 1.47, 95% CI 1.06-2.04, P = .020) (Fig. 2). Next, we conducted further analyses on age and atopic status within the Asian subgroups. In short, the IL-13 +2044G/A polymorphism was significantly associated with asthma. In the subgroup based on Asian ethnicity, there was association between the IL-13 +2044G/A polymorphism and asthma, especially in children, indicating that geographic and age difference may lead to heterogeneity among studies. IL-13 +2044G/A maybe a risk factor in Asian children.
In sum, the IL-13 +1923C/T polymorphism was associated with asthma risk in our meta-analysis. IL-13 +1923C/T was associated with asthma among Asians; for TT versus CC, IL-13 Table 1 Main characteristics of 25 case-control studies in asthmatic patients showing methods for analyzing IL-13 gene promoter +2044 and +1923 regions.

Study heterogeneity and publication bias
Sensitivity analyses did not change the initial results. There was little modification of the estimates after exclusion of individual studies. Publication bias in the included literature did not show evidence of remarkable asymmetry (Fig. 4), as supported by Egger test (all P > .1).

Discussion
Asthma is 1 of the most common chronic airway inflammations in humans. Asthma is estimated to cause over 3000 deaths and cost over $56 billion annually in medical expenses. [41] The imbalance of type 1 and type 2 immune responses is the main cause of asthma. Th2 cells mainly secrete cytokines IL-4, IL-5, and IL-13, which stimulate type 2 immunity to induce an immune response. [42,43] IL-13 is an important mediator of asthma exacerbations. Clinical trial data for IL-13-directed therapies show a strong effect on decreasing asthma exacerbation, implicating IL-13 as a mediator of inflammation in asthma. [44] In addition, many experimental results have demonstrated that IL-13 may direct increased airway inflammation and is involved in airway remodeling in asthma. Introduction of exogenous IL-13 into murine airways results in eosinophilic and lymphocytic increases, airway muscle contraction, and airway hyper-responsiveness. [5,6] Furthermore, the inhibition of IL-13 alone in vivo prevents and reverses established mucus cell changes, indicating that IL-13 plays an important role in mucus hyperproduction. [45,46] These cumulative results suggest that IL-13 is the major effector of asthma exacerbation. Moreover, in the Human Genome Project, it was very popular to use SNPs to detect the localization of the genetic determinants of human disease. Numerous studies have revealed that the IL-13 +2044A/G SNP markedly increases the risk of developing asthma. [16][17][18][19][20][21][23][24][25][26][27]29,30,[32][33][34]37,40] Meanwhile, many studies have found that the IL-13 +1923C/T polymorphism is associated with asthma risk. [18,22,23,28,31,33,35,36,[38][39][40] Thus, it is interesting to study the +2044A/G and +1923C/T polymorphisms associated with asthma.

Conclusions
To the best of our knowledge, this is the first meta-analysis to explain the relationships between the +2044A/G and +1923C/T polymorphisms, and age and atopic status, specifically within Asian samples. Our study results suggest that the IL-13 +2044A/ G and +1923C/T polymorphisms are significantly associated with asthma risk; specifically, IL-13 +2044A/G is associated with Asian ethnicity among adults and children with atopic status. Further investigations should also consider gene-gene and geneenvironment interactions, and well-designed studies with larger samples should be considered.