A causal association between gastroesophageal reflux disease and risk of heavy menstrual bleeding with regular cycle: A Mendelian randomization study

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A causal association between gastroesophageal reflux disease and risk of heavy menstrual bleeding with regular cycle: A Mendelian randomization study

https://doi.org/10.21203/rs.3.rs-3184160/v1

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Background: Clinical observations and retrospective studies have found that GERD is associated with hormone levels and digestive symptoms during menstruation. However, whether there is a genetic causal relationship between GERD and HMB with regular menstrual cycleshas not been reported.

Methods: In this study, we extracted instrumental variables from the dataset of GERD and HMB with regular menstrual cycles from the GWAS database, and then we used the R package "TwoSampleMR" and Mendelian randomization (MR) analysis to conduct causal inference analysis of GERD and HMB with regular menstrual cycles.

Results: We observed a possible causal relationship between GERD and HMB with regular menstrual cycles. GERD was positively correlated with menorrhagia.

Conclusion: Our results may provide evidence for a genetic causal effect of GERD on the etiological mechanism of HMB with regular menstrual cycles. For patients diagnosed with GERD, attention should be paid to monitoring the blood volume during menstruation.

mendelian randomization

causal relationship

gastroesophageal reflux disease

Heavy menstrual bleeding

Gastroesophageal reflux disease (GERD) is a common gastrointestinal disease with a low mortality rate. GERD has recently increased [1]. Generally report a burning feeling in the retrosternal area, raising into the chest and radiating toward the neck, throat, and occasionally the back[2]. The current diagnosis of GERD is based on the symptoms of esophagogastric mucosal lesions and gastroesophageal reflux. The diagnosis of GERD mainly includes a clinical history questionnaire, proton pump inhibition test, endoscopy and biopsy, dynamic reflux detection, and high-resolution esophageal manometry[3]. GRED is a chronic disease associated with a variety of risk factors, the main factors identified at present are genetics, body mass index, and smoking[4]. Previous studies have shown that GRED is related to female hormone levels, but the causal relationship between the two has not been clarified[5–9].

Heavy menstrual bleeding (HMB) or menorrhagia is defined as a loss of blood greater than or equal to 80mL per menstrual cycle[10, 11]. HMB affects women's health, reducing their physical, emotional, and social quality of life, and is a very common gynecological disease, affecting 2 to 5 in 10 women[12]. Some women experience symptoms during their periods, which can affect their quality of life, especially in younger women. A Spanish study found that dysmenorrhea was more likely to occur in women with heavy menstruation and was 6.6 times more likely to be accompanied by nausea during menstruation and 1.17 times more likely to be accompanied by vomiting during menstruation than women without menstruation[13]. The main causes of menorrhagia are first, uterine causes: organic, functional (functional uterine bleeding); Second, systemic causes: endocrine disorders, and blood diseases; Third, iatrogenic causes: IUD, use of anticoagulant drugs[14]. To our knowledge, the causal relationship between GERD and HMB with regular menstrual cycles has not been studied.

A few studies have found that estrogen may show sex differences in GERD symptoms[15, 16]. Only hormonal changes were observed in obesity-related hormone studies associated with GERD, which does not explain the high prevalence of GERD in obese patients[17]. GERD is closely related to hormones, but the causal relationship between GERD and HMB with regular menstrual cycles is not yet clear.

Mendelian randomization (MR) is a term that applies to the use of genetic variation to address causal questions about how modifiable exposures influence different outcomes. The principles of MR are based on Mendel's laws of inheritance and instrumental variable estimation methods, which enable the inference of causal effects in the presence of unobserved confounding[18]. On the basis of satisfying three assumptions (correlation hypothesis, exclusivity hypothesis, and independence hypothesis), MR has less confusion and bias than traditional epidemiological methods. To the best of our knowledge, no previous study has used the MR approach to test the causal effect of gastroesophageal reflux disease on the risk of Heavy menstrual bleeding with a regular cycle. Thus, this study aimed to examine whether GERD is causally associated with the occurrence of HMB with regular menstrual cycles., using a two-sample MR analysis.

2.1 | Data sources and selection of genetic Variants

We searched the GWAS database (https://gwas.mrcieu.ac.uk/), which houses a large collection of summary statistic data from hundreds of genome-wide association studies (GWASs). We used the publicly available summary statistics data sets of GWAS meta‐analyses for GERD in individuals of European descent (total n = 602,604; case = 129,080; control = 473,524; Table 1) as the exposure. To minimize the impact caused by linkage disequilibrium (LD), we set the threshold of statistical significance as “P < 5×10^− 8; LD r² < 0.001, distance = 10000kb” to identify the SNPs associated with GERD. We obtained summary statistics (R-squared and standard errors) for 75 SNPs (4 SNPs were lost in the later analysis, SNP = 79, Table S3, S6) associated with GERD as the IVs from the GWASs on GERD. We used the publicly available summary statistics data sets of GWAS meta‐analyses for HMB in individuals of European descent (total n = 463,010; case = 6,641, control = 456,369; GWAS ID: UKB-B-11572; SNP = 9,851,867; author: Ben Elsworth; year: 2018; consortium: MRC-IEU).

Table.1 Basic information on the GWAS applied in this study.

GWAS ID	Year	Trait	Consortium	Sample size	Number of SNPs
ebi-a-GCST90000514	2021	Excessive and frequent menstruation with regular cycle	NA	602,604	2,320,781
ukb-b-11572	2018	Gastroesophageal reflux disease	MRC-IEU	463,010	9,851,867

2.2 | Statistical analysis for Mendelian randomization

First, we evaluated the independent correlation between SNPS and GERD based on Mendelian randomization requirements. Second, we examined the association between each SNP and the risk of HMB with regular menstrual cycles. Third, we combined the above findings and used MR analysis to assess the uncompounded causal relationship between GERD and HMB with regular menstrual cycles. (Table 1). After collecting the GERD and HMB datasets from the GWAS database, we further performed MR Analysis using the "TwoSampleMR" package of the R program (version 3.4.2). The code for data processing refers to "The Causal Relationship Between Body Mass Index and the Risk of Osteoarthritis"[19] (PMID: 34103976; DOI: 10.2147/IJGM.S314180).

Three statistical methods including the inverse-variance weighted (IVW) method, weighted median estimator, and MR-Egger regression were used to investigate the causal relationship between GERD and regular menstrual cycles [20, 21]. The inverse-variance weighted (IVW) method uses a meta‐analysis approach to combine the Wald ratio estimates of the causal effect obtained from different SNPs and to provide a consistent estimate of the causal effect of the exposure on the outcome when each of the genetic variants satisfies the assumptions of an IV[22]. To explore its pleiotropy, we used weighted median and MR-Egger regression methods. MR-Egger regression provides a valid test of directional (unbalanced) pleiotropy and a valid test of the causal null hypothesis. Under this assumption, the slope estimate from MR-Egger regression is a consistent estimate of the true causal effect, and the intercept can be interpreted as an estimate of the average horizontal pleiotropic effect across the genetic variants[23, 24]. Since the traditional inverse variance weighting method can only produce a consistent estimate if all genetic variants in the analysis are valid instrumental variables, we use a weighted median estimator to combine data from multiple genetic variants into one causal estimate, which can better limit the error rate[20].

2.3 | Heterogeneity and sensitivity test

To assess whether the differences between SNP outcomes were significant, we performed a heterogeneity test. We also conducted an analysis to investigate the possibility of a single SNP drive.

3.1| Instrumental variables for Mendelian randomization

We selected 75 independent SNPs(There were 79SNPs in total, but four SNPs were lost during data analysis) from GWASs on GRED as the IVs (Table S3, Table 2, Fig. 1). 75 SNPS were positively correlated with HMB in regular menstrual cycles. A P-value of 5.00E‐ 08 corresponds to an F statistic > 30 for each single variant[25]. A threshold of F < 10 has been used to define a “weak IV.”

Table.2 MR estimates from each method of assessing the causal effect of GERD on the risk of HMB with normal menstrual cycles.

MR method	Number of SNPs	Beta	SE	pval	OR	95% CI
MR Egger	75	0.004	0.007	0.488	1.005	0.992 ~ 1.018
Weighted median	75	0.004	0.004	0.281	1.004	0.997 ~ 1.012
Inverse variance weighted	75	0.007	0.001	4.76E-10	1.007	1.005 ~ 1.009

Beta, beta coefficient; OR, odds ratio; MR, Mendelian randomization; SE, standard error; SNP, single nucleotide polymorphism; CI, confidence interval.

3.2 | Mendelian randomization results

The IVW method showed evidence to support a causal association between GERD and HMB in regular menstrual cycles. The IVW method showed evidence to support a causal association between GERD and HMB in regular menstrual cycles. (beta = 0.007, OR = 1.007, SE = 0.001, P = 4.6E-10, OR 95% confidence interval (95% CI) = 1.005 ~ 1.009); Table 2, Figs. 1 and 2). The MR-Egger method was used to assess potential directional pleiotropy[26]. MR‐Egger regression revealed that directional pleiotropy was unlikely to be biasing the result (Table S8, S9). The MR‐Egger analysis showed no causal association between GERD and HMB with regular menstrual cycles (beta = 0.005, OR = 1.005, SE = 0.007, P = 0.488; OR 95% CI: 0.992 ~ 1.018; Table 2, Figs. 1 and 2). However, the weighted median approach yielded evidence of a causal association between GERD and HMB with regular menstrual cycles.. The weighted median method suggests that there may not be a causal relationship between GERD and HMB with regular menstrual cycles (beta = 0.004, OR = 1,004, SE = 0.004, P = 0.281; OR 95% CI: 0.997 ~ 1.012; Table 2, Fig. 2). The IVW method suggests a causal effect of GERD on the risk of HMB with regular menstrual cycles, whereas the MR‐Egger and weighted median method suggests a null causal effect. Considering that compared with the MR-Egger analysis and weighted median method, the IVW method has the advantage of maintaining higher accuracy in estimation, and the direction of the above three models is consistent (OR > 1). the results of the MR analysis may support a potential causal association between GERD and HMB with regular menstrual cycles.

3.3 | Heterogeneity and sensitivity test

Horizontal pleiotropy tests showed no evidence of horizontal pleiotropy. However, The Cochran Q statistic may show evidence of heterogeneity (Table S7). Results from the “leave-one‐out” analysis demonstrated that no single SNP was driving the IVW point estimate. Asymmetry in the funnel plot indicates directional horizontal pleiotropy, which can bias MR methods; however, the funnel plot and MR Egger regression test showed no evidence of asymmetry (Fig. 3).

GERD is a multifaceted disorder that includes a range of syndromes that can be attributed to or exacerbated by gastroesophageal reflux. The epidemiology of GERD is associated with Western lifestyle, obesity, and the demise of Helicobacter pylori. Because of its prevalence and chronic nature, GERD is a significant financial burden when measured in physician visits, diagnoses, cancer surveillance protocols, and treatment[27]. In the Swedish study, a significant association was found between body weight and GERD symptoms, and estrogen may play an important role in GERD[7]. Many women experience vomiting or nausea during menstruation, and a few studies have found that elevated sex hormones during pregnancy may cause reflux, but other studies have found the opposite result[28]. However, due to confounding factors, it is difficult for classical epidemiological studies to explain the causal sequence of exposure factors and disease outcomes. The aim of this study was to explore the association between GERD and HMB with regular menstrual cycle risk through a two-sample Mendelian randomized study based on GWAS. The results suggest that there may be a causal relationship between gastroesophageal reflux and an increased risk of HMB with normal menstrual cycles.

In this study, we explored the relationship between GERD and the risk of HMB with normal menstrual cycles through a two-sample MR study based on GWAS. 75 SNPs that were significantly correlated with GERD were selected as instrumental variables. When IVW methods, weighted median estimators, and MR-Egger MR Methods were used with HMB with normal menstrual cycles study data from GWAS, a possible causal relationship between GERD and an increased risk of HMB with normal menstrual cycles were found.

The cause of the causal effect of GERD on HMB with normal menstrual cycles is not fully understood. Studies have evaluated the association between estrogen and GERD[16]. First, the distribution of GERD-related diseases varies widely. Women tend to have more symptoms of GERD than men, but men have more pathological complications of GERD than women[29]. These sex differences may be related to estrogen. Second, GERD is related to hormonal replacement therapy. Jacobson et al. studied postmenopausal hormone use and symptoms of GERD, which increase with estrogen dose and duration[15]. Third, GERD is associated with pregnancy. Van Thiel et al. studied that as pregnancy progresses, plasma progesterone and estrogen levels increase and LES pressure decreases throughout the course of pregnancy[30]. As we know, estrogen levels are usually supposed to drop during menstrual. When menstruation is excessive, it is usually caused by high levels of estrogen. In our causal effect study, GERD may increase estrogen levels and lead to heavy menstruation. GERD may promote the secretion of estrogen, which has not been found in previous studies. To our knowledge, this is the first study to look at a causal relationship between GERD and HMB with normal menstrual cycles.

The present study has several limitations. First, the effects of genetic variants on specific exposures (GERD) are unknown, and they may explain only a small percentage of the differences in specific exposures. Our analysis may be limited in its ability to detect an association. Second, the studies on GERD and HMB with normal menstrual cycles were based on participants of European descent. Since causation may depend on race and selection bias, further MR Studies in other populations are needed. Third, the GWAS meta-analysis we used for GERD did not include the pathological staging of GERD, which may have influenced the results. However, the available data do not allow us to perform MR Analysis according to the different pathological stages of GERD. Fourth, HMB with normal menstrual cycles data set is not segmented by age, and there may be bias in postmenopausal women and adolescent women. Finally, only the IVW method in our results showed evidence supporting causality, so the causal relationship between GERD and HMB with normal menstrual cycles may still need to be studied and analyzed.

In conclusion, the results of the MR Analysis suggest that GERD may be causally associated with an increased risk of HMB with normal menstrual cycles. However, the above causal relationship needs to be confirmed by repeated analysis. These results suggest that gastric reflux may play an important role in the occurrence and development of HMB with normal menstrual cycles. The current findings may provide an opportunity to identify the mechanisms by which GERD affects the risk of HMB with normal menstrual cycles. For patients diagnosed with GERD, attention should be paid to monitoring the blood volume during menstruation.

Author contributions

Xuetong Ren and Yuhua Wang designed the project; Xuetong Ren performed the analysis and wrote the manuscript. Haoyu Chen, Shaowei Liu and Haiyan Bai reviewed relevant literature; Yangang Wang designed the project and wrote the manuscript.

Data Availability

The data analysis code in this study comes from “He Y, Zheng C, He MH, Huang JR. The Causal Relationship Between Body Mass Index and the Risk of Osteoarthritis. Int J Gen Med. 2021 Jun 2;14:2227-2237. doi: 10.2147/IJGM.S314180. PMID: 34103976; PMCID: PMC8180293”.

Funding information

Natural Science Foundation of Hebei Province， Number: H2020423207

Acknowledgement

Not applicable.

Competing interests

The authors declare no competing interests.

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

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No competing interests reported.

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A causal association between gastroesophageal reflux disease and risk of heavy menstrual bleeding with regular cycle: A Mendelian randomization study

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Abstract

Figures

1 | INTRODUCTION

2 | MATERIALS AND METHODS

2.1 | Data sources and selection of genetic Variants

2.2 | Statistical analysis for Mendelian randomization

2.3 | Heterogeneity and sensitivity test

3 | RESULTS

3.1| Instrumental variables for Mendelian randomization

3.2 | Mendelian randomization results

3.3 | Heterogeneity and sensitivity test

4 | DISCUSSION

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