Association between GABRG2 rs211037 polymorphism and febrile seizures: a meta-analysis

Emerging evidence has implied that the GABRG2 gene play a role in the mechanism of febrile seizure (FS), however, the relationship between GABRG2 rs211037 polymorphism and the risk of FS remains controversial. This meta-analysis was conducted to investigate the relationship of GABRG2 rs211037 polymorphism with the susceptibility to FS. MEDLINE, Embase, Cochrane Library and CNKI databases were searched (until April 6, 2019) for eligible studies on the relationship between GABRG2 rs211037 polymorphism and FS. We calculated the odds ratios (ORs) by a fixed or random model with the STATA 15.0 software. Subgroup analyses for the ethnicity, the source of the control, and age and sex matching of controls were conducted. A total of 8 studies consisting of 775 FS patients and 5162 controls were included in this study. Based on the overall data, he GABRG2 rs211037 polymorphism was not significantly associated with the risk of FS (TT + CT vs CC: OR = 0.95, 95%CI 0.64–1.41, P = 0.80). Notably, the GABRG2 rs211037 variant was significantly associated with decreased risk of FS in Asian populations (TT vs CT + CC: OR = 0.63, 95%CI 0.45–0.88, P = 0.006), but increased risk in Caucasian populations (CT vs CC: OR = 1.56, 95%CI 1.14–2.15, P = 0.006). Significant associations were also detected when healthy controls out of the whole controls were employed for comparison (TT vs CT + CC: OR = 0.59, 95% CI 0.45–0.77, P < 0.001) and when data from studies with age- and sex-matched controls were used (TT + CT vs CC: OR = 0.60, 95% CI 0.43–0.86, P = 0.001). The GABRG2 rs211037 polymorphism may decrease the risk of FS in Asian populations, while increasing the risk in Caucasian populations. Further well-designed studies with large sample sizes are essential to verify the conclusions in other ethnicities.


Background
Febrile seizure (FS) is a type of seizure related with fever, but the causes and mechanisms of FS remain to be determined. As the most common seizure subtype in children, FS affects 2-5% of children, especially those younger than 5 years [1]. Although the exact pathogenesis of FS remains obscure, genetic factors may act as an important factor [2].
The gamma-aminobutyric acid type A (GABAA) receptor (GABAAR) contains mainly α, β and γ subunits and mediates a great many inhibitory neurotransmission in the brain [3,4]. The γ2 subunit is vital for postsynaptic clustering and synaptic maintenance of GABAARs by affecting the kinetics of the GABAAR channels [5][6][7]. Mutations in the GABRG2 gene could produce a nonfunctioning or clipped γ protein, thereby disturbing subunit assembly, reducing the expression of surface receptor, decreasing the GABAergic inhibitory effect, and finally inducing epileptogenesis [8,9]. Previous studies have revealed several epilepsy risk variants of GABRG2, among which the rs211037 variant has attracted much attention. The GABRG2 rs211037 variant can disturb the expression levels of the GABAAR subunits by influencing transcription, mRNA stability, and translation efficiency, resulting in varied sensitivity to extrinsic environmental signals [10].
The relation between GABRG2 rs211037 polymorphism and the risk of FS has been investigated in previous studies. Haerian et al. conducted a meta-analysis to investigate the relationship between the GABRG2 gene polymorphism and epilepsy, and indicated that rs211037 was associated with FS in Asians. Nevertheless, the number of included articles was relatively small, so it may be underpowered to verify the association. Although several new studies have been published recently [11][12][13][14], some important factors, including the type of control, the

Literature search strategy
Studies related to the relation of GABRG2 rs211037 polymorphism with FS were searched in the MEDLINE, Embase, Cochrane Library and CNKI databases until April 6, 2019. The following search terms were used: ('febrile seizure'), ('GABRG2') and ('polymorphism' OR 'variant' OR 'mutation'). No language restriction was set on the literature search. Furthermore, we conducted a manual search from the references of reviews and eligible studies.

Inclusion and exclusion criteria
Xiaohui Yang and Jing Chi screened each eligible study independently, and Xiaosa Chi would rejudge the study if any disagreement. The inclusion criteria of publications were as follows: (1) assessing the association between GABRG2 rs211037 variants and FS; (2) genotype frequency data of both case and control groups were available; (3) having a case-control design; and (4) English or Chinese publications. Accordingly, the exclusion criteria were as follows: (1) providing insufficient genotype information; (2) animal studies or experiments in vitro; (3) family-based or linkage studies; (4) reviews and conference abstracts; and (5) case reports or lacking a control group. Besides, articles including subjects at the same hospital during overlapping times were regarded as duplications, and only the study with the largest sample size was included for analysis.

Data extraction
Two authors (Xiaohui Yang and Xiaomeng Wang) extracted the baseline data from the included studies seperately and repeatedly, and any discrepancies were figured out by discussion. We extracted the following information from each article: first author, year of publication, country, ethnicity of the subjects, source of controls (healthy control or patients), matching criteria of controls (age-, sex-matched or not), HWE of controls, study period, genotyping methods, quality of control, numbers of cases and controls, and frequencies of genotype.

Statistical analysis
We assessed the strength of the correlation between GABRG2 rs211037 polymorphism and the risk of FS by  , and other genetic models (TT vs CC, CT vs CC, and T vs C). Furthermore, subgroup-analyses were stratified by ethnicity, source of the control (non-FS or healthy control) and the matching criteria in controls (age-, sexmatched or not). We calculated the pooled OR using the Z test, and regarded P < 0.05 as statistical significance. We used Cochran's Q test and the I 2 statistic to estimate the inter-study heterogeneity among the eligible studies, and regarded I 2 ≥ 50% as statistical significance. In this condition, the random-effects model was applied to calculate the pooled OR. Otherwise, the fixed-effects model was applied. Moreover, publication bias was evaluated using visual inspection of Funnel plot which was obtained from Begg's test. All data were calculated and analyzed with the STATA software (version 15.0; Stata Corp, College Station, Texas).

Study selection
Altogether 472 potentially related articles were yielded at the initial database search, and 349 were left after duplicate removal (Fig. 1). After manual screening by titles and abstracts, 302 studies were excluded according to the exclusion criteria. Forty-seven full-text studies were used for further evaluation. Ultimately, 8 eligible studies consisting of 5937 subjects (775 FS patients and 5162 controls) were included in this study [11][12][13][14][15][16][17][18]. The detailed information of all included studies are present in Table 1. The genotype distributions of GABRG2 rs211037 polymorphism of included studies are shown in Table 2.
To eliminate the potential confounding factors in the control group, we conducted stratified analyses for healthy control subjects and age-, sex-matched controls. We found a significant relation between GABRG2 rs211037 polymorphism and the susceptibility to FS when healthy controls were employed for comparison (recessive model, OR = 0.59, 95% CI 0.45-0.77, P < 0.001, Fig. 4). When analysis was performed using data from studies with age and sex matched controls, significant association was detected between the GABRG2 rs211037 variant (CT + TT vs CC) and the susceptibility to FS (dominant model, OR = 0.60, 95% CI 0.43-0.86, P = 0.005, Table 3).

Publication bias
The symmetrical Begg's funnel plot indicated that there was no publication bias among included studies (Fig. 5).

Discussion
Evidence is emerging that the GABRG2 gene is implicated in the mechanisms of FS, however, the relationship between GABRG2 rs211037 polymorphism and the risk of FS is still controversial. Previous meta-analysis studies have demonstrated that the GABRG2 rs211037 polymorphisms is significantly relative to the risk of FS, but they are limited by small sample sizes. Thus, we conducted this meta-analysis to further explore the relationship.
Different from the previous meta-analyses, we found that GABRG2 rs211037 polymorphism was not significantly related to the risk of FS using data combining all ethnicities. However, the GABRG2 rs211037 polymorphism was significantly related to decreased risk of FS in Asian populations, but increased risk of FS in Caucasian populations. This suggested that ethnicity could modify  the impact of the GABRG2 gene on the risk of FS. In addition, the GABRG2 rs211037 polymorphism was associated with decreased risk of FS when healthy controls out of the whole controls were used for comparison. The γ2 subunit is the major component of GABAAR, and its decrease is reported to affect the phasic or synaptic transmission [19][20][21][22]. Studies about cultured hippocampal neurons indicated that the relation of γ2 subunit mutations with FS may be due to the decreased expression of mutant GABAAR on the synaptic surface [23]. In a family with febrile seizures and a GABRG2 variant R43Q, resting-state fMRI revealed increased functional connectivity within the somatosensory cortex, as compared to the age-matched controls [24]. Besides, GABRG2 variants may affect the function and expression of several epilepsy-related genes [25]. Thus, mutations in GABRG2 have been proposed as candidates of FS susceptibility genes. The GABRG2 rs211037 polymorphism may affect the expression of GABAAR subunits, modify the receptor composition, influence the reaction to extrinsic environmental signals, and eventually alter the neuroinflammatory pathway in FS [10,26,27]. In our study, the GABRG2 rs211037 polymorphism may be a protective factor for FS and play a role in the mechanisms of FS.
However, the results should be explained with caution due to the following limitations. First, relevant studies in other databases may be missed out. Second, stratified studies were not performed in Africans due to limited data. Therefore, our results need to be further verified in Africans. Third, the analysis of gene-gene and loci-loci interactions was not conducted on account of the insufficient data.

Conclusions
In conclusion, the current study indicated that the GABRG2 rs211037 polymorphism is significantly related to decreased risk of FS compared to healthy control. The GABRG2 rs211037 polymorphism might diversely contribute to the risk of FS in different ethnicities. Further studies are essential to verify the conclusions and reveal the underlying mechanisms.