Abstract

Endometriosis is a common condition among women and can cause complications such as abdominal pain, dysmenorrhea, and infertility. One of the potential causes of this disease is autoimmunity. However, evidence regarding the role of autoimmunity is conflicting and inconclusive. The aim of this study was to investigate whether autoantibodies, a sign of autoimmunity, are present in people suffering from endometriosis. Relevant studies up to April 14, 2023 were identified by systematically searching Scopus, PubMed, Web of Science, Embase, and Google Scholar. This meta-analysis includes all qualified case-control studies of human populations that analyzed the association between serum autoantibodies and endometriosis. The odd ratios and 95% confidence intervals were calculated. In addition, heterogeneity and publication bias were examined, and subgroup analyses were performed based on region and target antigens. Forty-one studies were included, comparing 2,825 endometriosis patients with 4,158 healthy controls. The meta-analysis findings indicated a significant association between the presence of autoantibodies in the serum and an increased susceptibility to endometriosis (odds ratio: 4.242, confidence interval 95%: 3.824-4.706, p<0.001). In addition, there was a significant correlation between the presence of endometriosis and serum levels of anti-nuclear antibodies, B2 glycoprotein 1, CA125, carbonic anhydrase 1, cardiolipin, endometrial, laminin-1, smooth muscle, and syntaxin autoantibodies. Upon further analysis, it was found that the serum levels of these autoantibodies were higher in patients with endometriosis from North America than in those from other regions (p=0.001). The study revealed a significant correlation between serum autoantibodies and susceptibility to endometriosis, highlighting autoimmunity as a potential cause.

Keywords: Autoantibody, endometriosis, meta-analysis, serum

Introduction

Endometriosis is a prevalent medical condition characterized by endometrial tissue in extrauterine locations⁽¹⁾. This misplaced tissue responds to hormones and can cause internal bleeding, inflammation, and fibrosis⁽²⁾. A definitive diagnosis of endometriosis is performed after biopsy by surgery; therefore, its exact prevalence is uncertain⁽³⁾. However, it is estimated to affect 10-15% of women of reproductive age⁽⁴⁾. People affected by this disease experience symptoms such as pelvic pain, dysmenorrhea, and infertility⁽⁵⁾.

The exact etiology of this endometriosis remains unknown. However, previous research has shown that both genetic and environmental factors contribute to its development. Epigenetic abnormalities, anomalous estrogen production, retrograde menstruation, autoimmune responses, and allergic reactions are potential etiological factors^{(6, 7)}.

Several studies have been conducted on the role of autoimmune responses as a potential etiological factor in endometriosis. They have demonstrated that endometriosis is correlated with persistent regional inflammation and autoantibodies. Their results showed that women with endometriosis display immune system abnormalities similar to those seen in autoimmune diseases^{(8, 9)}. This includes disrupted immune surveillance, abnormal T and B-cell functions, heightened humoral immune response with increased autoantibodies in the serum, and inflammatory tissue damage^{(9, 10)}.

Although several studies have explored the connection between the presence of autoantibodies in the serum and the likelihood of developing endometriosis, the results have been inconsistent. The aim of this study was to analyze all relevant research and perform a meta-analysis to investigate this association.

Materials and Methods

Eligibility Criteria, Information Sources, and Search Strategy

This meta-analysis followed the Preferred Reporting Items for Systematic Reviews and Meta-analysis guidelines⁽¹¹⁾. Various databases, including Scopus, PubMed, Web of Science, EMBASE, and 30 pages of Google Scholar, were searched to identify relevant studies. The search was not restricted by language and covered the period from April 2023. The references of the included studies were also hand-searched. The search strategies consisted of the terms “endometriosis”, “autoantibodies”, “serum” and their related synonyms.

Women diagnosed with endometriosis based on standard criteria confirmed by laparoscopic sampling and/or histological examination will participate in this review. This study will compare endometriosis patients with healthy women of the same age group.

Study Selection

This meta-analysis included all available case-control studies that met the following criteria: Analysis of the association between autoantibodies and endometriosis patients provides necessary information for a meta-analysis.

The exclusion criteria were as follows: Letters, editorials, abstracts, conference abstracts, or publications lacking adequate information; studies that used women with diseases other than endometriosis, particularly autoimmune diseases; and studies that explored the presence of autoantibodies in bodily fluids or tissues other than serum.

Data Extraction

Two reviewers independently extracted the pertinent data from the selected studies. If necessary, the investigators resolved the differences through discussion and consultation with a third-party. The information extracted was entered into an Excel form, including the last name of the first author, date and location of the study, number of participants in both the case and control groups, mean age of participants, method of diagnosing endometriosis, subclass of autoantibodies, sample type, method of evaluating autoantibodies, and the number or mean value and standard deviation of autoantibody-positive cases and controls.

Assessment of the Risk of Bias

The articles’ quality was evaluated by two reviewers using the Newcastle-Ottawa Quality Assessment scale (NOS)⁽¹²⁾, and any discrepancies were resolved by a third reviewer. Articles that achieved a NOS score of 7 or above were classified as high quality, whereas those with scores between 5 and 7 were moderate quality.

Data Synthesis

All data were statistically analyzed using Comprehensive meta-analysis software version 3.0 (Biostat, USA). To determine the odds ratios (ORS) and their corresponding 95% confidence intervals (CIS), the statistical analysis used the groups’ sample size, mean serum autoantibody level, and standard deviation for both patients diagnosed with endometriosis and the healthy control group. Some studies presented continuous variables as median and quartile, which were converted to mean and standard deviation using the formula in the Cochrane Handbook^(13,14). The heterogeneity of the studies was assessed using Cochrane Q and I2 statistics. The random-effects model was used to estimate the outcome data in cases where the Cochrane Q p-value was less than 0.1 and the I2 value exceeded 50%, indicating the presence of statistical heterogeneity. Conversely, a fixed-effects model was employed in other instances. Subgroup analysis was performed to evaluate the impact of confounding variables on the outcomes of the meta-analysis. A sensitivity analysis was also conducted, in which each study was systematically excluded to assess the stability of the obtained results.

Results

Study Selection

After conducting a comprehensive search of the available resources, 1,053 studies were obtained. We then removed any duplicate sources and examined the titles and abstracts of 663 studies. Of these, 71 studies were selected for full-text review. Finally, after applying the inclusion and exclusion criteria, 41 studies were included in our meta-analysis. The process of literature screening and its outcomes are presented in Figure 1. The studies included in the analysis were assigned ratings ranging from zero to nine based on the Ottawa-Newcastle scale for case-control studies. As shown in Table 1, Table thirty-three studies were assessed as being of high quality, whereas eight studies were evaluated as being of moderate quality.

Study Characteristics

In line with the research methodology, we included a total of forty-one eligible case-control studies to investigate the correlation between the presence of autoantibodies and the risk of endometriosis. Eighteen studies were conducted in Europe^(15-32), eleven in Asia^(33-43), nine in North America^{(29, 44-51)}, and four in South America^(52-55). The basic characteristics of the included studies are listed in Table 1 and Table 2. Table 3 shows the number of studies investigating each autoantibody.

Risk of Bias of the Included Studies

Funnel plots, Begg’s rank correlation, and Egger’s regression tests were used to examine the existence of publication bias. Figure 2 and Table 3 present the outcomes of publication bias. Based on the results of statistical tests and asymmetry analysis of funnel plots, there is a possible publication bias in the studies that investigated pooled and anti-laminin-1 autoantibodies (as shown in Table 3). However, in other autoantibody studies, although a few exhibits slight visual asymmetry, statistical tests do not indicate any significant publication bias. The funnel plot diagrams, which show the likelihood of publication bias in studies on autoantibodies, were modified using the trim and fill test. The modifications did not result in any significant changes in studies that investigated the total and anti-laminin-1 autoantibodies (data not shown).

Synthesis of the Results

The findings of the meta-analysis indicate a substantial correlation between the existence of autoantibodies and susceptibility to endometriosis (OR: 4.242, CI 95%: 3.824-4.706, p<0.001) (Figure 3). Furthermore, individuals with endometriosis exhibited significantly higher levels of anti-nuclear antibodies (anti-ANA) (OR: 5.862, CI 95%: 3.454-9.950, p<0.001), B2 glycoprotein 1 (OR: 4.542, CI 95%: 1.360-15.175, p=0.014), CA125 (OR: 3.602, CI 95%: 1.485-8.733, p=0.005), carbonic anhydrase 1 (OR: 6.860, CI 95%: 3.043-15.468, p<0.001), cardiolipin (OR: 6.806, CI 95%: 3.369-13.749, p<0.001), endometrial (OR: 11.793, CI 95%: 2.382-58.383, p=0.002), laminin-1 (OR: 6.340, CI 95%: 3.151-12.757, p<0.001), smooth muscle (OR: 8.041, CI 95%: 4.442-14.557, p<0.001), and syntaxin (OR: 3.815, CI 95%: 1.249-11.649, p=0.019) autoantibodies than healthy controls. Statistical analysis found no significant relationship between autoantibodies against enolase (OR: 2.067, CI 95%: 0.739-5.782, p=0.167) and PDIK1L (PDLIM1 interacting kinase 1 like) (OR: 1.434, CI 95%: 0.241-8.528, p=0.692) and susceptibility to endometriosis.

The relevant forest plots are presented in Figure 4. It is worth mentioning that the conducted studies^{(52, 53, 55)} failed to detect anti-dsDNA autoantibodies in individuals suffering from endometriosis, indicating the absence of any correlation.

Heterogeneity Test and Subgroup Analysis

The I2 test and Cochrane Q statistic showed heterogeneity among studies analyzing total, anti-CA125, anti-endometrial, anti-enolase, anti-PDIK1L, and anti-syntaxin autoantibodies. However, studies related to other autoantibodies showed no significant between-study heterogeneity, as shown in Table 3. To investigate the cause of heterogeneity, subgroup analysis was conducted based on the region. The findings revealed that patients with endometriosis from North America had notably higher levels of autoantibody titers than patients from other regions (p=0.001), and the region of living was identified as one of the sources of heterogeneity. In addition, subgroup analysis based on differences in target antigens of autoantibodies demonstrated this factor as a cause of heterogeneity in the study of pooled autoantibodies (p<0.001).

Sensitivity Analysis

Sensitivity analyses were conducted to evaluate the potential impact of a single study on the overall effect of autoantibodies in endometriosis. The sensitivity analyses indicated that upon exclusion of each study, the general conclusions remained substantially unchanged. These analyses consistently exhibited the robustness of the meta-analysis outcomes.

Discussion

Endometriosis is a significant health issue that affects women, but its exact cause remains unknown⁽⁵⁶⁾. Several theories have been proposed to explain its origin, including autoimmunity⁽⁵⁷⁾. However, studies investigating the development of autoantibodies in patients with autoimmune conditions have produced conflicting results. Therefore, the research collected and analyze the current body of information on this topic. The findings of our study indicate that the serum of individuals diagnosed with endometriosis has significantly higher levels of autoantibodies than that of healthy controls.

Studies have shown an increase in both the activity and quantity of B-cells in the serum of individuals with endometriosis^{(58, 59)}. The increase in B-cell activity leads to an elevated production of antibodies, which is consistent with the findings of the current study. Research has shown that immune cells, particularly B-cells, demonstrate an increase within lesions associated with endometriosis⁽⁶⁰⁾. In addition, the secretion of cytokines responsible for activating B-cells, such as B lymphocyte-stimulators, also experiences an elevation within these specific regions⁽⁵⁹⁾. These findings validate the results of our study.

Endometriosis relies on estrogen for its development⁽⁶¹⁾ and estrogen has been identified as one of the potential mechanisms that increase the quantity and functionality of B-cells in individuals with endometriosis⁽⁶²⁾. Research suggests that estrogen initiates a genetic program that alters the survival and activation of B-cells, leading to a shift in the naive immune system toward autoreactivity⁽⁶²⁾. The exact cause of autoantibody formation in endometriosis is not well understood, but it is thought to be related to cellular damage and inflammation that occur in endometriosis-associated lesions. Abnormal exposure of self-antigens from damaged cells to the immune system triggers an autoimmune response, leading to the formation of autoantibodies that target cell-derived antigens⁽⁶³⁾.

This study suggests that differences in the target antigens of autoantibodies and continent of residence may contribute to the observed heterogeneity. The results of the subgroup analysis show that North American patients with endometriosis have higher levels of autoantibodies. Differences in genetic or environmental factors may explain this disparity. Additionally, according to the literature, autoimmune antibody manifestation varies among North American ethnic groups. Bruner et al.⁽⁶⁴⁾ discovered that African Americans with systemic lupus erythematosus exhibit significantly higher levels of anti-ANA autoantibody expression than other North American races. It is important to consider factors beyond ethnicity that can contribute to heterogeneity, such as sampling error, use of varied laboratory tests, body mass index, and age. It is crucial to study these factors in depth in future research efforts.

This meta-analysis is the most comprehensive study that systematically reviews and analyzes the relationship between  autoantibodies and endometriosis, synthesizing over three decades of research. In addition, this study included a large sample size of 2,731 patients diagnosed with endometriosis and 4,067 healthy controls, potentially providing a definitive outcome with high accuracy and minimal bias for the general population.

It is important to note that there are limitations to this study. The laboratory data were measured in different centers using various methods and detection kits, which may have resulted in inconsistencies. Furthermore, the use of varying cutoff points across investigations could lead to inconsistencies in the outcome. In addition, the limited and insufficient data regarding the participants in the selected studies hindered our ability to examine other factors that may have an impact on heterogeneity, such as age and BMI. Therefore, future research should focus on exploring this aspect in more detail.

Conclusion

In conclusion, our meta-analysis revealed that patients diagnosed with endometriosis exhibit a greater prevalence of autoantibodies than healthy individuals. Based on the findings of this study, it is likely that autoimmune reactions are associated with the progression of endometriosis. However, further studies are required to determine the mechanism underlying autoantibody production in endometriosis. Future research should also investigate autoantibody levels during different phases of endometriosis.

Ethics

Authorship Contributions

Design: S.F., M.H.M., Data Collection or Processing: R.H.M., N.A., Analysis or Interpretation: S.F., R.H.M., N.A., M.H.M., Literature Search: M.H.M., Writing: S.F., R.H.M., N.A., M.H.M.

Conflict of Interest: No conflict of interest was declared by the authors.

Financial Disclosure: The authors declared that this study received no financial support.