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Introduction

Endometriosis manifests as ectopic endometrial cells outside the uterus. It is an intractable disease that causes infertility, dysmenorrhea and pelvic pain. Endometriosis occurs in 10% of women of childbearing age. Notably, the incidence of endometriosis has been rising in recent years (1). The pathogenesis of endometriosis remains to be elucidated.

Published reports indicate that endometriosis is a polygenic/multifactorial disease caused by interactions between multiple genes and the environment (2,3). In particular, a correlation has been identified between endometriosis and exposure to environmental toxins such as dioxin (4); dioxin and dioxin-like compounds have been implicated in the development of endometriosis (5,6).

The phase II conjugation enzymes usually function to inactivate environmental toxins. Among these, glutathione S-transferase (GST) may be critical for the detoxification of dioxins. Human GSTs are classified into two distinct categories: Soluble or cytosolic and membrane-bound microsomal. The soluble or cytosolic GSTs are subdivided into seven families named α, µ, ω, π, σ, θ and ζ (7). Genes in several of these families are polymorphic, including: GSTA2 in the α family, GSTM1 and GSTM3 in the µ family, GSTP1 in the π family, GSTO, GSTT1, and GSTT2 in the θ family, and GSTZ1 in the ζ family. Heritable allelic differences in GSTM1, GSTM3, GSTT1 and GSTP1 may have marked relevance for individual susceptibility to disease.

GSTM1 and GSTT1 are two candidate genes that may play an important role in the development of endometriosis. GSTM1 and GSTT1 are located on chromosomes 1p13.3 and 22q11.23, respectively. They are critical in the detoxification of the products of oxidative stress produced during the repair of the ovarian epithelium. GSTM1 and GSTT1 null alleles have reduced enzyme activity, a state that may contribute to inefficient detoxification of intermediates produced during stress. This may increase damage to various host genes and contribute to the pathogenesis of endometriosis (8,9).

A meta-analysis summarizing the literature up to the year 2005 suggested that the GSTT1 null genotype, but not the GSTM1 null genotype, was associated with an increased risk for endometriosis (7). In the years since 2005, additional reports investigating this topic have been published. The objective of the present study was to update the existing meta-analysis and reevaluate the possible associations between GSTM1, GSTT1 and combined GSTM1/GSTT1 (null genotype vs. wild-type) gene polymorphisms and susceptibility to endometriosis.

Materials and methods

Searches

For this systematic review and meta-analysis, PubMed (from January 1996 to January 2014), Embase (from January 1996 to January 2014), Chinese BioMedical Literature database (from January 1996 to January 2014) and Google Scholar (from January 1996 to January 2014) were searched. The following keywords were used: ‘endometriosis’, ‘polymorphisms’, ‘glutathione S-transferases’, ‘GSTM1’ and ‘GSTT1’ or their combinations.

Reference lists from articles identified by the electronic search were searched by hand. This process was performed iteratively until no additional articles could be identified.

Inclusion and exclusion criteria

Articles published in English or Chinese were included if they reported quantitative outcomes from case-control genetic association studies on GSTM1, GSTT1 or combined GSTM1/GSTT1 (null genotype vs. wild-type) gene polymorphisms and endometriosis versus non-endometriosis or healthy controls.

Studies were excluded if they were case reports, case-only studies, letters, reviews or meta-analyses; included subjects who were related; included cases of adenomyosis, which has unknown etiology (10); reported insufficient data; or were duplicate studies.

Selection of studies

Two reviewers (XYX and ZSJ) independently examined titles and abstracts to select eligible studies. Records were removed that were ongoing or unpublished studies, or were published as abstracts or conference proceedings. Where data sets were overlapping or duplicated, only the most recent information was included. The full text of potentially relevant studies was retrieved. Two reviewers (XYX and HJG) independently examined the full text records to determine which studies met the inclusion criteria. Disagreement about the selection of studies was resolved by discussion and consensus.

Data extraction and management

Two reviewers (XYX and ZSJ) independently extracted data from eligible studies including the first author's last name, publication year, study location, ethnicity, matching variability, diagnostic criteria, stages of disease, source of controls, numbers of cases and controls, and numbers and/or percentages of null genotypes. Disagreement about data extraction was resolved by discussion and consensus.

Assessment of quality of evidence in included studies

Two reviewers (YYL and HJG) independently assessed quality of evidence in the included studies using the 9-star Newcastle-Ottawa Scale, which considers selection, comparability and outcome evaluation criteria.

Assessment of heterogeneity

Heterogeneity was assessed using the χ2 test and I2 test. The I2 statistic was interpreted as follows: I2=0–40%, heterogeneity may not be important; I2= 30–60%, heterogeneity may be moderate; I2=50–90%, heterogeneity may be substantial; and I2=75–100%, considerable heterogeneity (11). If heterogeneity was present, meta-regression was used to find the source.

Assessment of reporting biases

A funnel plot of effect estimates against their standard errors (SEs) was created to assess possible reporting bias between studies. Funnel plot asymmetry was assessed using Egger's linear regression test and Begg's rank correlation test; P<0.05 suggested publication bias.

GSTM1/GSTT1 and risk for endometriosis

Two reviewers (XYX and HJG) independently combined data from trials using a fixed-effect model (DerSimonian and Laird method) when there was no significant heterogeneity in populations (I2<50%) and a random-effect model (Mantel-Haenszel method) when there was considerable heterogeneity. Variables were synthesized using odds ratios (ORs). A P-value of 0.05 was used as the cut-off value to determine statistical significance, and data are presented as the estimated OR with 95% confidence intervals (CIs). All statistical analyses were performed using STATA software, version 12.0 (StataCorp, College Station, TX, USA). Inconsistencies in data analysis were resolved through consensus and discussion with a third reviewer (ZSJ).

Sensitivity and subgroup analyses

Sensitivity analyses were performed to explore the impact of excluding outlying results. Subgroup analyses were performed by stratifying patients according to ethnicity (Caucasian, Asian or mixed), characteristics of controls (hospital patients or healthy individuals), and quality of evidence (high-quality or low-quality).

Results

Screening and selection

The searches identified 120 articles. Titles and abstracts were screened, and 36 studies were identified as potentially eligible for inclusion. The full text articles for these studies were retrieved. Following analysis of the full text articles, four studies were excluded and 32 studies were found to be eligible for inclusion according to the criteria used for considering studies in this review (Fig. 1).

Figure 1. PRISMA 2009 flow diagram showing the article screening and selection process. Using the search strategy, 120 articles were identified by the initial search, and 36 required further assessment. Finally, 32 articles were included in this review, one of which was considered as two studies. GSTM1, glutathione S-transferase µ1; GSTT1, glutathione S-transferase θ1.

Included studies

The characteristics of the included studies are shown in Table I. There were 32 case-control genetic association studies involving 3,990 cases of endometriosis and 4,625 controls. One publication addressed two groups of subjects with different ethnicities and was considered as two case-control genetic association studies (12); thus, the total number of studies was considered to be 33. Studies included data relevant to the GSTM1 genotype, GSTT1 genotype or the combined GSTM1/GSTT1 genotype. Of the 32 eligible studies, 20 were conducted in Asia (12–31), eight in Europe (32–39), two in North America (40,41), and two in South America (42,43). The evidence reported in 23 studies was identified as high-quality, and that in 10 studies was identified as low-quality.

Table I. Characteristics of included studies on the GSTM1, GSTT1 and combined GSTM1/GSTT1 gene polymorphisms.

Table I.

Characteristics of included studies on the GSTM1, GSTT1 and combined GSTM1/GSTT1 gene polymorphisms.

					GSTM1 (n)		GSTT1 (n)		GSTM1+GSTT1 (n)
First author, year	Ethnicity	Countries	Source of controls	Quality	Cases/null	Controls/null	Cases/null	Controls/null	Cases/null	Controls/null	Refs.
Baranov, 1996	Caucasian	Russia	Healthy individuals	Low	42/34	67/26					(33)
Baranova, 1999	Caucasian	Russia, France	Hospital patients	High	65/50	72/33	65/13	72/7			(32)
Baranov, 1999	Caucasian	Russia	Healthy individuals	Low	150/88	99/42					(34)
Hadfield, 2001	Caucasian	UK	Hospital patients	High	132/59	52/27	116/29	50/14			(35)
Baxter, 2001	Caucasian	England	Healthy individuals	High	84/40	219/107					(36)
Bischoff, 2002	Caucasian	USA	Hospital patients	Low	62/13	36/20					(40)
Ivaschenko, 2003	Caucasian	Russia	Hospital patients	High	74/42	40/17	74/27	40/6	74/16	40/2	(37)
Arvanitis, 2003	Caucasian	Greece	Healthy individuals	High	275/161	346/181	275/24	346/31	275/11	346/16	(38)
Peng, 2003	Asian	China	Hospital patients	High	76/50	80/37					(15)
Lin, 2003	Asian	China	Hospital patients	High	68/49	28/12	68/53	28/9			(16)
Morizane, 2004	Asian	Japan	Healthy individuals	Low	108/57	173/89	108/52	173/71	108/30	173/43	(17)
Hsieh, 2004	Asian	China	Hospital patients	High	150/95	159/8					(18)
De Carvalho, 2004	Mixed	Brazil	Hospital patients	Low	61/21	32/17					(42)
Ding, 2004	Asian	China	Healthy individuals	High	80/46	105/55	80/59	105/47	80/34	105/24	(12)
Ding, 2004	Asian	China	Healthy individuals	High	41/21	107/57	41/15	107/32	41/10	107/14	(12)
Babu, 2005	Caucasian	India	Hospital patients	High	310/121	215/64	310/42	215/34	310/14	215/11	(19)
Hur, 2005	Asian	Korea	Hospital patients	Low	194/112	259/145	194/104	259/125			(20)
Aban, 2007	Caucasian	Turkey	Hospital patients	High	150/88	150/65	150/59	150/44			(21)
Chang, 2007	Asian	China	Hospital patients	High	74/48	65/30	74/46	65/32	74/27	65/13	(30)
Kim, 2007	Asian	Korea	Hospital patients	High	316/183	256/146	316/178	256/124			(22)
Rozati, 2009	Caucasian	India	Hospital patients	High	97/26	102/15					(13)
Yang, 2009	Asian	China	Hospital patients	Low	216/134	216/100					(14)
Cao, 2009	Asian	China	Hospital patients	High	51/33	102/61	51/22	102/39			(23)
Wu, 2009	Asian	China	Hospital patients	High	96/63	85/40					(24)
Huang, 2010	Asian	China	Hospital patients	High	28/12	29/10					(25)
Trabert, 2011	Caucasian	USA	Healthy individuals	High	254/137	567/268					(41)
Hosseinzadeh, 2011	Caucasian	Iran	Healthy individuals	High	120/87	200/80					(26)
Wu, 2012	Asian	China	Healthy individuals	Low	121/57	171/52	121/40	171/33	121/23	171/15	(27)
Seifati, 2012	Caucasian	Iran	Hospital patients	High	101/51	142/74					(28)
Vichi, 2012	Caucasian	Italy	Hospital patients	High	181/104	162/85	181/20	162/32			(39)
Matsuzaka, 2012	Asian	Japan	Hospital patients	High	97/43	143/67	97/38	143/56			(29)
Frare, 2013	Mixed	Brazil	Healthy individuals	Low	50/25	46/34	50/16	46/27			(43)
Sachan, 2013	Caucasian	Iran	Healthy people	Low	66/27	100/16					(31)

[i] Null genotype vs. wild type gene polymorphisms and susceptibility to endometriosis. GSTM1, glutathione S-transferase µ1; GSTT1, glutathione S-transferase θ1.

Excluded studies

Of the 36 studies that were relevant to the GSTM1/GSTT1 genotype and endometriosis, four were excluded. Of these, three were duplicates (13,14,32), and one included subjects who were related (44).

GSTM1/GSTT1 and risk for endometriosis

Data reporting on the GSTM1 gene polymorphism are described in 33 case-control studies (3,990 cases of endometriosis and 4,625 controls). The meta-analysis demonstrated that there was a significant association between the GSTM1 null genotype and an increased risk for endometriosis (OR=1.56; 95% CI: 1.25–1.95; P<0.0001; Fig. 2A).

Figure 2. Association between GSTM1, GSTT1 and the combined GSTM1/GSTT1 null genotypes and susceptibility to endometriosis. (A) A total of 33 studies described the association between the GSTM1 null genotype and susceptibility to endometriosis [odds ratio (OR)=1.56; 95% confidence interval (CI): 1.25–1.95; P<0.0001]; and (B) 18 studies described the association between the GSTT1 null genotype and susceptibility to endometriosis (OR=1.31; 95% CI: 1.02–1.68; P=0.037). GSTM1, glutathione S-transferase μ1;. GSTT1, glutathione S-transferase θ1.

Subgroup analyses stratified by ethnicity (Caucasian: OR=1.599; 95% CI: 1.205–2.122; P=0.001; Asian: OR=1.772; 95% CI: 1.242–2.528, P=0.002), source of controls (hospital patients: OR=1.561; 95% CI: 1.151–2.117; P=0.004; healthy individuals: OR=1.569; 95% CI: 1.131–2.176; P=0.007), and quality of evidence (high-quality: OR=1.563; 95% CI: 1.253–1.949; P<0.0001) confirmed this finding.

Subgroup analysis stratified for mixed ethnicity (two case control studies involving 111 cases of endometriosis and 78 controls) demonstrated a significant association between the GSTM1 null genotype and a decreased risk for endometriosis (OR=0.404; 95% CI: 0.219–0.745; P=0.004; Table II). Compared with individual Caucasian and Asian populations, the difference was statistically significant (P<0.001; data shown in Table III).

Table II. Meta-analysis of the association between GSTM1, GSTT1 and combined GSTM1/GSTT1 (null genotype vs. wild-type) gene polymorphisms and susceptibility to endometriosis.

Table II.

Meta-analysis of the association between GSTM1, GSTT1 and combined GSTM1/GSTT1 (null genotype vs. wild-type) gene polymorphisms and susceptibility to endometriosis.

Group	No. of studies	No. of subjects (cases/controls)	OR [95%CI]	P-value
Total studies
GSTM1 genotype	33	3,990/4,625	1.563 [1.253–1.949]	<0.001
GSTT1 genotype	18	2,371/2,490	1.345 [1.044–1.733]	0.022
GSTM1+GSTT1 genotype	8	1,083/1,222	1.672 [1.291–2.166]	0.005
Caucasian
GSTM1 genotype	16	2,163/2,569	1.599 [1.205–2.122]	0.001
GSTT1 genotype	7	1,171/1,035	1.124 [0.745–1.697]	0.577
GSTM1+GSTT1 genotype	3	659/601	1.185 [0.717–1.961]	0.508
Asian
GSTM1 genotype	15	1,716/1,978	1.772 [1.242–2.528]	0.002
GSTT1 genotype	10	1,150/1,409	1.573 [1.186–2.085]	0.002
GSTM1+GSTT1 genotype	5	424/621	1.898 [1.404–2.565]	<0.001
Mixed
GSTM1 genotype	2	111/78	0.404 [0.219–0.745]	0.004
GSTT1 genotype	1	50/46
GSTM1+GSTT1 genotype	0
Controls from hospital patients
GSTM1 genotype	21	2,599/2,425	1.561 [1.151–2.117]	0.004
GSTT1 genotype	12	1,696/1,542	1.284 [0.963–1.712]	0.089
GSTM1+GSTT1 genotype	3	458/320	1.797 [1.081–2.989]	0.024
Controls from healthy individuals
GSTM1 genotype	12	1,391/2,200	1.569 [1.131–2.176]	0.007
GSTT1 genotype	6	675/948	1.315 [0.767–2.254]	0.320
GSTM1+GSTT1 genotype	5	625/902	1.657 [1.085–2.532]	0.001
High quality
GSTM1 genotype	23	2,920/3,426	1.563 [1.253–1.949]	<0.001
GSTT1 genotype	14	1,898/1,841	1.376 [1.020–1.858]	0.037
GSTM1+GSTT1 genotype	6	854/878	1.753 [1.265–2.430]	0.001
Low quality
GSTM1 genotype	10	1,070/1,199	1.259 [0.785–2.020]	0.340
GSTT1 genotype	4	473/649	1.121 [0.646–1.944]	0.684
GSTM1+GSTT1 genotype	2	229/344	1.542 [1.009–2.356]	0.045

[i] OR, odds ratio; CI, confidence interval; GSTM1, glutathione S-transferase µ1; GSTT1, glutathione S-transferase θ1.

Table III. Comparisons of subgroup analyses for GSTM1, GSTT1 and combined GSTM1/GSTT1 studies.

Table III.

Comparisons of subgroup analyses for GSTM1, GSTT1 and combined GSTM1/GSTT1 studies.

A, Analysis of the GSTM1 gene
		GSTM1 (n)
Subgroup	Subjects	Null	Normal	χ2	P-value
Ethnicity				3.245	0.72a
Caucasian	Cases	1,128	1,035
	Controls	1,120	1,449
Asian	Cases	1,003	713
	Controls	909	1,069
Mixed	Cases	46	65	18.737	<0.001b
	Controls	51	27	23.467	<0.001c
Source of controls				0.130	0.718
Hospital patients	Cases	1,397	1,202
	Controls	1,073	1,352
Healthy individuals	Cases	780	611
	Controls	1,007	1,193
Quality				0.825	0.364
High quality	Cases	1,609	1,311
	Controls	1,539	1,887
Low quality	Cases	568	502
	Controls	541	658
B, Analysis of the GSTT1 gene
		GSTT1 (n)
Subgroup	Subjects	Null	Normal	χ2	P-value
Ethnicity				6.766	0.009
Caucasian	Cases	214	957
	Controls	168	867
Asian	Cases	607	543
	Controls	568	841
Source of controls				0.638	0.425
Hospital patients	Cases	631	1,065
	Controls	522	1,020
Healthy individuals	Cases	206	469
	Controls	241	707
Quality				0.062	0.803
High quality	Cases	625	1,273
	Controls	507	1,334
Low quality	Cases	212	261
	Controls	256	393
C, Analysis of GSTM1+GSTT1 genes
		GSTM1+GSTT1 (n)
Subgroup	Subjects	Null	Normal	χ2	P-value
Ethnicity				7.642	0.006
Caucasian	Cases	41	618
	Controls	29	572
Asian	Cases	124	300
	Controls	109	152
Source of controls				0.091	0.763
Hospital patients	Cases	57	401
	Controls	26	294
Healthy individuals	Cases	108	517
	Controls	112	790
Quality				0.022	0.882
High quality	Cases	112	542
	Controls	80	598
Low quality	Cases	53	176
	Controls	58	286

a Caucasians vs..Asians

b Caucasians vs. mixed

c Asians vs. mixed.

Data reporting on the GSTT1 gene polymorphism are described in 18 case-control studies (2,371 cases of endometriosis and 2,490 controls). The meta-analysis demonstrated a significant association between the GSTT1 null genotype and an increased risk for endometriosis (OR=1.31; 95% CI: 1.02–1.68; P=0.037; Fig. 2B).

Subgroup analysis stratified by ethnicity demonstrated a significant association between the GSTT1 null genotype and an increased risk for endometriosis among Asians (OR=1.573; 95% CI: 1.186–2.085; P=0.002), but not among Caucasians (OR=1.124; 95% CI: 0.745–1.697; P=0.577).

Subgroup analyses stratified by the source of controls found no significant association between the GSTT1 null genotype and an increased risk for endometriosis among hospital-based studies (OR=1.284; 95% CI: 0.963–1.712; P=0.089) or among healthy individuals (OR=1.315; 95% CI: 0.767–2.254; P=0.320).

Subgroup analyses stratified by quality of evidence demonstrated a significant association between the GSTT1 null genotype and an increased risk for endometriosis among studies considered high-quality evidence (OR=1.376; 95% CI: 1.020–1.858; P=0.037), but not among studies considered low-quality evidence (OR=1.121, 95% CI: 0.646–1.944; P=0.684; Table II).

Data reporting on the combined GSTM1/GSTT1 gene polymorphism are described in eight case-control studies (1,083 cases of endometriosis and 1,222 controls). The meta-analysis demonstrated a significant association between the combined GSTM1/GSTT1 null genotype and an increased risk for endometriosis (OR=1.68, 95% CI: 1.29–2.17; P<0.0001; Fig. 2C).

This association was unchanged by subgroup analyses stratified by source of controls (hospital-based studies: OR=1.797; 95% CI: 1.081–2.989; P=0.024; healthy individuals: OR=1.657; 95% CI: 1.085–2.532; P=0.001) or quality of evidence (high-quality evidence: OR=1.753; 95% CI: 1.265–2.430; P=0.001; low-quality evidence: OR=1.542; 95% CI: 1.009–2.356, P=0.045; Table II).

Subgroup analysis stratified by ethnicity demonstrated a significant association between the combined GSTM1/GSTT1 null genotype and an increased risk for endometriosis among Asian populations (OR=1.898; 95% CI: 1.404–2.565; P<0.001), but not among Caucasian populations (OR=1.185; 95% CI: 0.717–1.961; P=0.508).

Visual inspection of a Funnel plot, Egger's test and Begg's rank correlation test revealed no significant publication bias for the GSTM1, GSTT1 and combined GSTM1/GSTT1 studies (Fig. 3; Table IV).

Figure 3. Assessment of publication bias for studies on (A) GSTM1, (B) GSTT1 and (C) combined GSTM1/GSTT1 genotypes. GSTM1, glutathione S-transferase µ1;. GSTT1, glutathione S-transferase θ1.

Table IV. Heterogeneity and publication bias of GSTM1, GSTT1 and combined GSTM1/GSTT1 studies.

Table IV.

Heterogeneity and publication bias of GSTM1, GSTT1 and combined GSTM1/GSTT1 studies.

	Heterogeneity		Publication bias (P-value)
Group	I2 value (%)	P-value	Egger's test	Begg's funnel plot
Total studies
GSTM1 genotype	81.8	<0.001	0.313	0.412
GSTT1 genotype	69.9	<0.001	0.557	0.705
GSTM1+GSTT1 genotype	44.7	0.081	0.170	1.000
Caucasian
GSTM1 genotype	79.2	<0.001	0.454	0.322
GSTT1 genotype	64.7	0.009	0.339	0.764
GSTM1+GSTT1 genotype	58.5	0.090	0.021	0.296
Asian
GSTM1 genotype	83.8	<0.001	0.098	0.083
GSTT1 genotype	62.4	0.004	0.160	0.210
GSTM1+GSTT1 genotype	13.6	0.081	0.340	0.806
Mixed
GSTM1 genotype	0.0	0.664	<0.001	0.317
Controls from hospital patients
GSTM1 genotype	83.6	<0.001	0.390	0.506
GSTT1 genotype	65.9	0.001	0.335	0.451
GSTM1+GSTT1 genotype	62.2	0.071	0.585	1.000
Controls from healthy individuals
GSTM1 genotype	79.4	<0.001	0.598	0.784
GSTT1 genotype	78.4	<0.001	0.431	0.707
GSTM1+GSTT1 genotype	45.9	0.116	0.531	1.000
High quality
GSTM1 genotype	80.9	<0.001	0.042	0.068
GSTT1 genotype	69.9	<0.001	0.530	0.189
GSTM1+GSTT1 genotype	49.0	0.081	0.641	1.000
Low quality
GSTM1 genotype	85.1	<0.001	0.788	0.516
GSTT1 genotype	77.2	0.004	0.347	1.000
GSTM1+GSTT1 genotype	62.9	0.101

[i] GSTM1, glutathione S-transferase µ1;. GSTT1, glutathione S-transferase θ1.

There was evidence of significant heterogeneity (I2>50%) between studies of GSTM1 and GSTT1, and those used in subgroup analyses, although not among studies of GSTM1/GSTT1 combined (Table IV). Therefore, the random-effect model was used in all analyses with the exception of the analysis of combined GSTM1/GSTT1 gene polymorphisms. For the GSTM1 and GSTT1 gene polymorphisms, a meta-regression was conducted in which publication year, ethnicity, source of controls, sample size, and quality of evidence were covariates. All the covariates were entered into the meta-regression model simultaneously, and the covariates that had the highest P-values were omitted one at a time in order to identify any sources of heterogeneity among them. However, the meta-regression analysis did not identify any of these covariates as a significant source of heterogeneity (Figs. 4 and 5).

Figure 4. Meta-regression for GSTM1 studies, with publication year, ethnicity, source of controls, sample size, and quality of evidence as covariates. All covariates were entered into the meta-regression model simultaneously, and the covariates with the highest P-values were omitted one at a time to identify sources of heterogeneity. The meta-regression did not identify any of these covariates as a significant source of heterogeneity. Variables were omitted in the following order: Size (A→B), source (B→C), publication year (C→D), ethnicity (D→E). GSTM1, glutathione S-transferase µ1.

Figure 5. Meta-regression for GSTT1 studies, with publication year, ethnicity, source of controls, sample size, and quality of evidence as covariates. All covariates were entered into the meta-regression model simultaneously, and covariates with the highest P-values were omitted one at a time to identify sources of heterogeneity. Meta-regression identified publication year as a significant source of heterogeneity (P=0.048), but after omitting this covariate heterogeneity remained substantial (I2=67.21%) Variables were omitted in the order: Source (A→B), quality (B→C), ethnicity (C→D), size (D→E). GSTT1, glutathione S-transferase θ1

To explore the effects of individual studies on the pooled OR estimates, a sensitivity analysis was performed, with the omission of one study at a time. The OR estimates for the GSTM1 polymorphism were not notably altered (Fig. 6A). The OR estimates for the GSTT1 and combined GSTM1/GSTT1 polymorphisms were altered when studies were excluded (Fig. 6B and C).

Figure 6. Sensitivity analyses investigating the association between the (A) GSTM1, (B) GSTT1 and (C) combined GSTM1/GSTT1 null genotypes and susceptibility to endometriosis; one study was omitted at a time. GSTM1, glutathione S-transferase µ1;. GSTT1, glutathione S-transferase θ1.

Discussion

In the present study, a meta-analysis of data from 33 studies was conducted to examine the associations between the GSTM1, GSTT1 and combined GSTM1/GSTT1 null genotypes and susceptibility to endometriosis. The risk for endometriosis was significantly increased in the presence of the GSTM1, GSTT1 and combined GSTM1/GSTT1 null genotypes compared with the wild-type. Subgroup analyses stratified by ethnicity, source of controls and quality of evidence confirmed this finding among several subgroups, but particularly among studies considered high-quality evidence. Notably, among patients of mixed ethnicity, the GSTM1 null genotype was significantly associated with a decreased risk for endometriosis compared with the wild-type.

A similar meta-analysis of 23 studies performed in 2005 demonstrated an increased risk for endometriosis in women with the GSTT1 null genotype (8). However, the authors requested that their findings be interpreted with caution as asymmetry in the funnel plot was evident, which was likely due to publication bias (8). This previous study did not include subgroup analyses or an evaluation of the combined GSTM1/GSTT1 null genotype-endometriosis association.

Previous meta-analyses have found that the GSTM1/GSTT1 gene polymorphism is associated with cervical cancer (45), breast cancer (46), bladder cancer (47), gastric cancer (48,49) and acute leukemia (50). In accordance with the observations of the present study, several studies have shown that the GSTM1 (OR=32.6, 95% CI: 15.07–70.32, P<0.0001) (18) and GSTT1 (OR>3; P<0.0001) null genotypes (12,16) are associated with an increased risk for endometriosis. However, other reports suggest the GSTM1 (OR=0.21, 95% CI: 0.09–0.52, P<0.0001; OR=0.35, 95% CI: 0.15–0.83, P<0.0001) (40,43), GSTT1 (OR≥5; P<0.0001) (16) and combined GSTM1/GSTT1 (OR=0.38; P<0.001) (17) null genotypes are associated with a decreased risk for endometriosis. These divergent results may be explained by differences in GSTM1/GSTT1 null genotype frequencies and study locations. The frequency of the GSTM1/GSTT1 null genotype may vary from 10 to 65% depending on the region and population studied (51). Different study locations may introduce confounding variables associated with variations in lifestyles and exposures to toxic substances of the study populations.

The results of the present study must be interpreted with caution due to the presence of substantial heterogeneity. Among analyses of the studies of GSTM1 and GSTT1, the cause of heterogeneity remains unclear, despite meta-regression analyses being conducted. Among the analyses of combined GSTM1/GSTT1 studies, subgroup and sensitivity analyses suggested that studies that included patients with advanced stage endometriosis caused most of the variability. Publication bias was unlikely to have influenced the findings.

In addition to the heterogeneity, there were several limitations to this study. Firstly, the composition of the endometriosis patient and control populations varied between studies. For instance, some studies included only patients with advanced endometriosis (17–20,22,27,35), while control populations consisted of a mixture of infertile (29), postmenopausal (43) and premenopausal (18,35) women, and newborn babies that had not been exposed to the environment (17). Furthermore, patients and controls were not always accurately matched by age or environmental exposures. Secondly, gene-gene or gene-environment interactions may jointly increase the risk for endometriosis; therefore, different lifestyle and environmental factors may contribute to differential genotypic frequencies in cases and controls. Attempts were made to mitigate inaccuracies associated with this limitation through a subgroup analysis stratified according to ethnicity. Thirdly, this study was based on published articles. As a positive result is more likely to be published, publication bias is an inherent limitation of all meta-analyses irrespective of the outcomes of the Egger's linear regression test and Begg's rank correlation test.

In conclusion, the present meta-analysis shows the GSTM1, GSTT1 and combined GSTM1/GSTT1 null genotypes are likely associated with increased susceptibility to endometriosis. These data are in contrast to those reported previously. Therefore, further studies reporting higher quality evidence are necessary to verify these conclusions.

Acknowledgements

The study was supported by Hubei Provincial Natural Science Foundation of China (Grant No.2013060602010236).

References