Association between interleukin-4 C589T polymorphism and asthma in Iraq: Ameta-analysis

doi:10.21203/rs.3.rs-3792067/v1

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Association between interleukin-4 C589T polymorphism and asthma in Iraq: Ameta-analysis

https://doi.org/10.21203/rs.3.rs-3792067/v1

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Background

Asthma is a common disease with a complex risk architecture involving both genetic and environmental factors. Several studies have identified an association between interleukin (IL)-4 C-589T and asthma risk; however, the results remain inconclusive.

Aims

to identify the effect of IL-4 C589T polymorphisms on asthma susceptibility in Iraq.

Material and methods

PubMed, Web of Science, Scopus, and Google Scholar databases were searched for only case-control studies published on Iraqi asthmatic. The methodological quality assessment of the included studies was performed based on the Newcastle-Ottawa Quality Scale (NOS). Based on the heterogeneity, we conducted a meta-analysis using random-effect models. Pooled odds ratios (ORs) with a 95% confidence interval (CI) were calculated using the allele (C vs. T), homozygous (CC vs. TT), heterozygous (CT vs. TT), dominant (CC þ CT vs. TT), and recessive (CC vs. CT þ TT) genetic models to assess the strength of the relationship between IL-4 C589T polymorphism and asthma risk. In addition, the stability of our analysis was evaluated by heterogeneity, sensitivity, and study design, and publication bias analysis.

Result

We included 5 case-control studies with a total of 359 cases and 245 controls. All genetic models showed a higher risk of asthma, the overall analyses were a highly statistically significant. There was moderate to high heterogeneity among the genetic models. However, sensitivity analysis revealed highly significant differences in the pooled odds ratios of the rs2243250 polymorphism between asthmatic patients and control groups.

Conclusion

The meta-analysis identified a significant association between the IL-4 C-589T polymorphism and asthma. However, the substantial result variability indicates the need for well-designed case-control studies with a large population to more accurately estimate this association.

Asthma

IL-4

Iraq

Polymorphism

Asthma, a prevalent chronic non-communicable disease in both children and adults, is defined by fluctuating respiratory symptoms and variable airflow restriction. Both domestically and globally, the prevalence of asthma is increasing. It affects about 7.5% of the adult population. Additionally, the average annual asthma prevalence is higher in children (9.5%) than in adults (7.7%). It arises from complex gene-environment interactions, leading to diverse clinical presentations and variations in airway inflammation and remodeling ^[1–3].

Genome-wide association studies (GWAS) have shown that genetic variation is crucial in the onset and progression of asthma. Numerous candidate genes linked to asthma risk have been identified ^[4]. Several genetic epidemiological studies have identified an association between the gene polymorphisms of IL4 and asthma in different populations ^{[5, 6]}. Interleukin 4 (IL 4) is a cytokine associated with the cause of several allergic diseases such as asthma due to their function in the differentiation of T helper type 2 lymphocytes and induction of the IgE isotype switch ^[7]. IL-4 is a cytokine that plays a significant role in the immune system and has been associated with the development and exacerbation of asthma. IL-4 mediates important pro-inflammatory functions in asthma, including induction of the IgE isotype switch, expression of vascular cell adhesion molecule-1 (VCAM-1), promotion of eosinophil transmigration across endothelium, mucus secretion, and differentiation of T helper type 2 lymphocytes leading to cytokine release. IL-4 cytokine is a major cytokine in the etiology of thunderstorm asthma, atopic dermatitis, and allergic rhinitis. Genetic variations in the IL-4 gene may contribute to an individual's susceptibility to asthma and influence the severity of the condition ^{[8, 9]}.

Several case-control studies in Iraq have been conducted in recent years related to IL-4 rs2243250 polymorphism and asthma risk. However, consensus has not been reached regarding the association between rs2243250 polymorphism of the IL-4 gene and asthma risk. Therefore, we performed a meta-analysis of available case-control studies to systematically evaluate the potential association between thers2243250 polymorphism of the IL-4 gene and asthma risk.

Characteristics and Quality of Selected Studies:

We identified a total of articles from Scopus, PubMed, Web of Science, and Google Scholar databases. After duplicate entries were removed using EndNote, articles remained to be screened. The initial screening excluded studies that failed to meet the inclusion criteria. After filtering by availability and reading, consisting of reading the title, abstract, and keywords of each study, five articles were selected ^[10–14]. The study selection process was visually presented in Fig. 1, which displays the PRISMA flow diagram.

The included reports comprised 359 cases and 245 controls (Table 1). All studies were conducted on Iraqi people. With relation to the types of study all publications were the result of case–control studies and were of high quality according to the Newcastle-Ottawa scale (NOS). The samples used in the studies varied in terms of age group: two with adults, one with children, and two with mixed‑age groups (children and adults).

Quality assessments of the selected studies are shown in Table 2. According to the NOS, three studies ^[12–14] had a risk of bias due to differences between the number of cases and the number of case genotypes in hospital controls. However, they were still considered to be of moderate quality, scoring 5 or 6 points.

HWE was detected in the asthmatic patients and controls in all studies ^[10–14]. All studies suggested that the distribution of genotypes in the controls was consistent with HWE except one study ^[10], as shown in Table 3.

Meta-Analysis and Sensitivity Analysis Finding:

Due to the evident heterogeneity in study design among the included literature, we utilized the random-effect model to estimate the pooled effect sizes for all statistical analyses in this meta-analysis. Detailed results of the meta-analysis are presented in Table 4.

Five studies ^[10–14] with available data can be included to estimate the pool effect sizes under allele (C vs. T), homozygous (CC vs. TT), heterozygous (CT vs. TT) and recessive (CC vs. CT + TT) models. For the dominant (CC + CT vs. TT) model, there were five studies ^[10–14] with available data that can be included to estimate the pool effect sizes. Although all these five genetic models have a higher risk of asthma, the overall analyses were not statistically significant (Table 4). The table presents the odds ratios (OR) and corresponding 95% confidence intervals (CI) for different genetic models related to the IL-4 rs2243250 polymorphism and asthma risk. The "Total" row indicates the overall analysis, while the "Study design" rows show results for mixed-age and adult-specific studies. In the overall analysis, the heterozygous genetic model (CT vs TT) yielded a statistically significant result with an OR of 0.25 (95% CI: 0.08–0.83, p = 0.02), suggesting a lower risk of asthma associated with this genetic model. However, the other genetic models did not show statistically significant associations. When considering study design, the results for mixed-age and adult-specific studies varied. For mixed-age studies, none of the genetic models showed statistically significant associations. In adult-specific studies, the heterozygote genetic model (CT + TT) approached statistical significance with an OR of 0.38 (95% CI: 0.14, 1.06, p = 0.07).

For the overall meta-analyses, there was moderate to high heterogeneity (I2 between 71%-93%) among the genetic models. Sensitivity analysis conducted by omitting each study and cumulative statistics showed that the pooled ORs of the rs2243250 polymorphism were highly significant difference was noted between the asthmatic patient and control groups in the genetic IL‑4 rs2243250 polymorphism (CC genotype showed OR = 2.37, 95% CI = 1.67–3.37, P < 0.001; CT genotype showed OR = 0.23, 95% CI = 0.16–0.32, P < 0.001; and TT genotype showed OR = 2.74, 95% CI = 1.59–4.71, P = 0.008), as presented in Fig. 2.

Publication bias:

Funnel plot of IL-4 C‑589T CC, CT, and TT genotypes for all studies included in the meta-analysis were presented in Fig. 3. Standard error of the logarithm of the OR (SE (log [OR])) was plotted against the OR for each study. The symmetric, inverted funnel shape implies an absence of publication bias.

Literature search

The electronic databases PubMed, Web of Science, Scopus, and Google Scholar were searched using the following terms: “(IL4 or interleukin-4) and (polymorphism) and (asthma) and (Iraq)”. The search was performed in August 2023, with no language restriction. Search results were deduplicated using EndNote and then screened for eligibility based on title and abstract. Eligibility criteria included: (1) Investigated the relationship between IL-4 rs2243250 polymorphisms and risk of pediatric asthma; (2) Case-control study; (3) Study on human participants, not animal studies; (4) Genotype or allele frequency data were available in the original text; (5) The sample is composed of individuals of any age, of both sexes, with a diagnosis of asthma and independent of the time of diagnosis of the disease. Exclusion criteria: (1) non-original research, such as case reports, meeting abstracts, letters, etc; (2) IL-4 polymorphism in other countries; (3) The study evaluating the association between IL‑4 polymorphisms and other diseases; (4) Did not contain sufficient data. (5) IL‑4 polymorphisms in site other than IL-4 C589T.

Data extraction

All data were extracted independently by the two authors using the established search strategies and inclusion criteria. Data extracted from the literature included first author, year of publication, country, sample size, ethnicity, and genotype and allele distribution in the case and control groups.

Quality assessment

The quality of the case-control studies was assessed using the Newcastle-Ottawa scale, which evaluates various criteria including participant selection, group comparisons, and measurement of exposure factors. Scores on the Newcastle-Ottawa scale ranged from 0 to 9, with studies scoring 5 or higher classified as high-quality studies.

Statistical analysis:

Review Manager version 5.3 (Nordic Cochrane Centre, Cochrane Collaboration, Copenhagen, Sweden) was used to conduct the meta‑analysis, sequential analysis, and publication bias assessment. The odds ratio (OR) with a 95% confidence interval (CI) was employed to calculate the association between IL‑4 C‑589T polymorphisms and asthma risk. The pooled OR was determined using the Z‑test, which was considered as significant with P < 0.05.

A test for heterogeneity was performed separately for three ORs, that is the homozygous dominant model (CC), heterozygous model (CT), and homozygous recessive model (TT). A test for heterogeneity was performed separately for three ORs, the homozygous dominant model (CC), heterozygous model (CT), and homozygous recessive model (TT) for the IL‑4 C‑589T gene.

The study heterogeneity was assessed by the I2 test. The fixed‑effects model was used when the P value of the I2 test was < 25%. To estimate whether our results were substantially influenced by the presence of any individual study, a sensitivity analysis was performed by systematically omitting each included case–control study and recalculating the significance of the result.

Checking Hardy–Weinberg equilibrium (HWE), HWE was checked in the patient and control groups using the Chi‑square test. The funnel plot was used to examine the publication bias. In the absence of publication bias, it assumes that studies with high precision (large study effects) will be plotted near the average. Studies with low precision (small study effects) will be spread evenly on both sides of average, creating a roughly funnel‑shaped distribution. Deviation from this shape can indicate publication bias.

The present meta-analysis, based on 5 case-control studies with 359 cases and 245 controls, revealed that those subjects with the CT genotype vs. TT genotype of IL-4 rs2243250 polymorphism exhibited a lower risk of asthma associated with this genetic model. However, the other genetic models did not show statistically significant associations under the allele (C vs. T), homozygous (CC vs. TT), homozygous (CC vs TT), dominant (CC + CT vs TT), and recessive (CC vs. CT + TT) models.

Furthermore, the present meta-analysis reveals a highly significant association between the IL-4 C-589T polymorphism and asthma under the CC genotype, CT genotype, and TT genotype. However, a p-value of less than 0.001 in a test for the pool effect size indicates strong evidence of significant heterogeneity in the meta-analysis. The presence of significant heterogeneity carries important implications for the interpretation of the meta-analytic results, suggesting that the true effect size may vary across studies due to factors beyond random error or sampling variability. Consequently, it may be inappropriate to simply calculate a single overall effect size for the meta-analysis, as this would not adequately capture the variability in effect sizes across studies.

Certain limitations to this meta‑analysis need to be taken into consideration when interpreting the findings. First, the inclusion of a lower number of studies may limit the robustness and generalizability of the findings. A smaller pool of studies can impact the statistical power of the meta-analysis and may not fully capture the breadth of available evidence on the association between the IL-4 C-589T polymorphism and asthma. Second, high heterogeneity between study groups introduces a limitation, as it suggests substantial variability in the effect sizes reported by different studies. This heterogeneity can make it challenging to derive a single, precise estimate of the true effect size, potentially affecting the overall coherence and reliability of the meta-analytic results. Third, the variance of results between studies represents a limitation in that it may complicate the synthesis of findings and the drawing of definitive conclusions. The wide variability in reported results across studies may hinder the ability to arrive at a clear and consistent understanding of the relationship between the IL-4 C-589T polymorphism and asthma. Fourth, the insufficient original data on relevant risk factors or genes interacting with the IL-4 polymorphism limits the evaluation of environmental interactions. This limitation hampers the comprehensive assessment of how environmental factors and genetic interactions may influence the association between the IL-4 polymorphism and asthma, potentially constraining the depth of understanding in this area.

In conclusion, the meta-analysis revealed a strong association between the IL-4 C-589T polymorphism and asthma across different genotypes. However, significant heterogeneity in the results suggests caution is needed when interpreting the findings, as the true effect size may vary across studies. Large-scale studies with more polymorphisms are needed to investigate the functional effects of IL-4 polymorphisms on the development of asthma in Iraq.

Ethical Approval:

The procedures followed in this study were by the ethical standards of the responsible committee on human experimentation (institutional or regional) and with the Helsinki Declaration of 1975, as revised in 2000 (available at https://www.wma.net/policies-post/wma-declaration-of-helsinki-ethical-principles -for-medical-research-involving-human-subjects/). The study was approved by the Research Ethics Committee of Babylon University, a medical college in Babylon, Iraq (2022-05/12), and an IRB was obtained. The parents of participants who revealed readiness to participate in this study were supplied with written informed consent and verbal information regarding the aim of the study. Sample collection and processing. The identity of the participants has been kept anonymous and confidential. There has been no duplicate publication or submission of this manuscript elsewhere. The manuscript has been read and approved by all the authors, the requirements for authorship have been met, and each author believes that the manuscript represents honest work. Subject to acceptance, authors will transfer the copyright to the publisher. There is no ethical problem or conflict of interest to be declared for any of the co-authors.

Competing interests:

There is no conflict of interests

Funding:

There is no Funding

Data availability:

The data and materials are available from the corresponding authors upon request.

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Table 1

Characteristics of the studies
Study ID	year	Country	City	Study population	Sample size	Genotyping methods
Abood and Mohanad, ^[10]	2018	Iraq	Karbala	Children	100/100	ARMS-PCR
Gaidan ^[11]	2018	Iraq	Baghdad	Adult	45/40	Sequence
Hassan ^[12]	2022	Iraq	Thi-Qar	Mixed	60/30	RFLP-PCR
Hussein and Jaber ^[13]	2017	Iraq	Baghdad	Mixed	84/25	ARMS-PCR
Mahmood and Abdulla ^[14]	2021	Iraq	Babylon	Adult	70/50	ARMS-PCR

Table 2

Quality assessments of studies based on Newcastle Ottawa Scale.
Study ID	Selection				Comparability	Exposure			Total score
Study ID	A	B	C	D	E	F	G	H	Total score
Abood and Mohanad, ^[10]	*			*	**		*	*	6
Gaidan ^[11]	*			*	**		*	*	6
Hassan ^[12]	*			*	**		*	*	6
Hussein and Jaber ^[13]		*		*	*		*	*	5
Mahmood and Abdulla ^[14]			*	*	*	*	*	*	6
A- Is the case definition adequate B- Representativeness of the cases C- Selection of Controls D- Definition of Controls E- Comparability of cases and controls on the basis of the design or analysis F- Ascertainment of exposure G- Same method of ascertainment for cases and controls H- Non-Response rate

Table 3

Distribution of IL-‑4 genotype among asthmatic patients and controls
Study ID	Patients			HWE P-value	Control			HWE P-value
Study ID	CC (%)	CT (%)	TT (%)	HWE P-value	CC (%)	CT (%)	TT (%)	HWE P-value
Abood and Mohanad, ^[10]	66 (66%)	17 (17%)	17 (17%)	0.002**	7 (7%)	90 (90%)	3 (3%)	< 0.001**
Gaidan ^[11]	29 (64.44%)	13 (28.89%)	3 (6.67%)	0.576	35 (87.5%)	4 (10%)	1 (2.5%)	0.430
Hassan ^[12]	11 (18.3%)	17 (28.3%)	32 (53.3%)	0.057	10 (33.3%)	14 (46.7%)	6 (20.0%)	0.959
Hussein and Jaber ^[13]	42 (87.5%)	5 (10.42%)	1 (2.08%)	0.112	8 (32%)	13 (52%)	4 (16%)	0.736
Mahmood and Abdulla ^[14]	27 (36.0%)	32 (42.70%)	16 (21.3%)	0.686	13 (26.0%)	32 (64.0%)	5 (10.0%)	0.069

Table 4

Subgroup Analysis for IL-4 C589T Polymorphism Mixed
Study ID	No		Allele (C vs T)		Homozygous (CC vs TT)		Heterozygous (CT vs TT)		Dominant (CC + CT vs TT)		Recessive (CC vs CT + TT)
Study ID	No		OR (95%CI)	p-value	OR (95%CI)	p-value	OR (95%CI)	p-value	OR (95%CI)	p-value	OR (95%CI)	p-value
Total	5		1.22 [0.43, 3.40]	0.71	0.92 [0.24, 3.49]	0.90	0.25 [0.08, 0.83]	0.02*	0.41 [0.14, 1.16]	0.09	0.42 [0.07, 2.45]	0.34
Study design
Mixed		2	1.80 [0.08, 42.61]	0.71	1.87 [0.02, 181.63]	0.79	0.43 [0.07, 2.55]	0.35	1.22 [0.03, 47.44]	0.92	0.39 [0.01, 12.17]	0.59
Adult		2	0.59 [0.19, 1.82]	0.36	0.54 [0.19, 1.57]	0.26	0.38 [0.14, 1.06]	0.07	0.40 [0.15, 1.06]	0.07	1.49 [0.25, 8.85]	0.66

No competing interests reported.

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Association between interleukin-4 C589T polymorphism and asthma in Iraq: Ameta-analysis

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