The Mechanistic Study Of Inflammatory Response Mediated Atrial Fibrillation Induced By Gastroesophageal Reflux Disease

Objective:Gastroesophageal reflux disease(GERD)refereed as an uncomfortable symptom(heartburn and acid reflux)caused by reflux of gastric contents in to the esophagus.It is the most common chronic disease of the digestive tract.Atrial fibrillation(AF)is the most common clinical arrhythmia disease,which can cause accelerations of patient mortality.The studies have suggested that gastroesophageal reflux disease may cause atrial fibrillation.In addition to damaging esophageal tissue,more and more clinical studies have reported that GERD may induce atrial fibrillation,which not only seriously affects the patients quality of life(reducing exercise capacity,increasing the risk of heart failure and stroke),but also significantly increases mortality.However,the clear mechanism of atrial fibrillation induced by gastroesophageal reflux disease remains deeper investigation.Inflammatory response is considered to be an important mechanism in the occurrence and development of both atrial fibrillation and gastroesophageal reflux disease.Based on the hypothesis that GERD induces esophageal inflammation and that inflammation can induce atrial fibrillation,we have proposed the following hypothesis: the gastric contents of GERD,bile acid and/or gastric acid reflux can leads to local inflammation of esophagus,that then local inflammation soaks into the whole esophagus layer,or excess amount of inflammatory cytokines which releases into the blood,resulting in increased levels of atrial inflammation,thereby subsequently inducing the occurrence and development of atrial fibrillation.In order to clarify the relationship between GERD-induced inflammation and atrial fibrillation,blood samples and esophageal mucosa tissues was taken under gastroscope examination to detect local inflammation and systemic inflammation in GERD with atrial fibrillation patients.Methods:A total of 50 patients who were diagnosed and treated in Xinjiang Uygur Autonomous Region People's Hospital between January 2017 to June 2018 was selected as research group basing on the inclusion,exclusion and diagnostic criteria.A total of fifteen persons selected as a healthy(control)group whom that finished a terms of physical test at the same time of period.General information(gender,age,BMI,etc.)of patients in each group was recorded.The GERD questionnaire,24-hour ambulatory electrocardiogram,24-hour p H monitoring,high-resolution esophageal manometry and gastroesophageal endoscopy were used for further investigation.The local mucosa at 3 cm of the dentate line of the esophagus were collected and stored as specimens.Hematoxylin-eosin(HE)staining,immunohistochemistry(IHC),real-time polymerase chain reaction(RT-PCR),enzyme-linked immunosorbent assay(ELISA)methods were subsequently used to analyze the degree of esophageal inflammation and detect expression levels of inflammatory cytokines [interleukin-6(IL-6),IL-8,IL-1?,and tumor necrosis factor-alpha,TNF-?],and high sensitive C-reactive protein(Hs-CRP)in blood.Results:(1)According to the clinical manifestations,GERD questionnaire scores and endoscopy,the patients were divided into gastroesophageal reflux disease with atrial fibrillation group(AF group),gastroesophageal reflux disease without atrial fibrillation group(non-AF group)and healthy control group.The control group consisted of 15 cases,10 males(66.7%)and 5 females(33.3%).The ratio of males to females was 2:1,with an average age of 32.29±4.61 years.In the non-AF group,there were 25 cases,16 males(64.0%)and 9 females(36.0%).The ratio of males to females was 1.77:1,with an average age of 38.86±11.33 years.There were 25 cases in AF group,18 males(72.0%)and 7 females(28.0%).The ratio of males to females was 2.57:1,with an average age of 43.57±10.81 years.The average age of AF group was higher than that of healthy control group,and there was significant statistical difference between the two groups(P<0.05).Smoking,drinking and body mass index(BMI)were not significantly different among the three groups(P>0.05).In the non-AF group,2 cases(8.0%)were complicated with hypertension,3 cases(12.0%)with hyperlipidemia and 2 case(8.0%%)with type 2 diabetes.In the AF group,3 cases(12.0%)were complicated with hypertension,4 cases(16.0%)with hyperlipidemia and 3 cases(12.0%)with type 2 diabetes.There was no significant statistical difference between the two groups(P>0.05).The GERDQ scores of non-AF group and AF group(3~7,8~10,11~14 and 15~18)were significantly higher than those of control group.There was a significant difference between the two groups(P<0.01).The scores of 3~7,8~10,11~14 and 15~18 in AF group were significantly higher than those in non-AF group(P<0.01).The 24-hour esophageal p H monitoring showed that the frequency of weak acid reflux(4<p H<7),acid reflux(p H?4),proximal esophageal acid reflux(%)and De Meester score in AF group were obviously significant as compare to non-AF group,with statistical significance(P<0.05).Howrver,the total number of reflux and proximal esophageal acid reflux(%)in AF group were slightly higher than those in non-AF group,but there was no significant difference between the two groups(P>0.05).High-resolution manometry(HRM)results showed that the length of lower esophageal sphincter(LES),the resting pressure of LES,the residual pressure of LES,the resting pressure of upper esophageal sphincter(UES),the residual pressure of UES and the contractile front velocity(contractile front velocity)in both groups.There was no significant difference in front velocity(CFV),intra bolus pressure(IBP)and distal contractile integral(DCI)between the two groups(P> 0.05).(2)The whole esophagus layer of the control group was smooth with intact mucosa and no erosive lesions were found under endoscopy.In the non-AF group,slight edema,congestion and erosive lesions were found in the esophageal mucosa at the gastroesophageal junction.In AF group,large-scale erosion,inflammatory exudation,edema and ulceration of esophageal mucosa at gastroesophageal junction were found under endoscopy.HE staining and light microscopy showed that the mucosa of the control group was basically intact,and there was no edema or neutrophil infiltration.Moderate neutrophil infiltration and severe inflammatory reaction were observed in non-AF group.In AF group,there were inflammatory changes in esophageal mucosa,such as infiltration of immune cells(neutrophils,eosinophils),prolongation of papilla,edema and congestion.Tissue histopathological inflammation(injury)score analysis showed that the scores of non-AF group and AF group were significantly higher than those of control group(P<0.001).The inflammation score of AF group reached the peak and was significantly higher than that of non-AF group(P<0.01).Immunohistochemical tests of inflammatory cytokines(IL-6,IL-8,IL-1?,and TNF-?)were performed after routine treatment of esophageal mucosa in each group.Microscopic observation showed that IL-6,IL-8,IL-1?,and TNF-? were mainly expressed in the esophageal epithelium,mucosa and submucosa.In the esophageal tissues of AF patient,the positive expressions of the early mentioned cytokines largely located in the whole esophageal mucosa,indicating severe tissue inflammation.RT-PCR results showed that the relative expression levels of inflammatory factors(IL-6,IL-8,IL-1?,and TNF-?)in non-AF group and AF group was significantly higher than that in control group(P<0.001).As compare to non-AF group,AF group markedly rised the transcription levels of the early mentioned cytokines(P<0.01).The results of ELISA showed that,compared with the control group,the concentration of inflammatory factors such as IL-6,IL-8,IL-1?,and TNF-? and Hs-CRP in the blood samples of non-AF group and AF group were increased significantly(P<0.001).AF group markedly up-regulated the expression levels of above mentioned inflammatory cytokines than non AF group,which statistically significant(P<0.01).Conclusion:This study in the present found that the acid reflux markers in AF group,such as weak acid reflux(4<p H<7),acid reflux(p H?4),proximal esophageal acid reflux(%)and De Meester score were markedly increased as compare to the non-AF group,suggesting that the severity of acid reflux was closely related to the occurrence and development of AF.In AF group,large-scale erosion,inflammatory exudation,edema and ulcer of esophageal mucosa at gastroesophageal junction were found under endoscopy.Under light microscopy,the esophageal mucosa of AF group showed severe inflammatory changes such as infiltration of immune cells(neutrophils,eosinophils),prolongation of papillae,edema and congestion.This study preliminarily confirmed the theory that GERD causes esophageal inflammation and inflammation further can induces atrial fibrillation.The result also shows that serious acid reflux exists objectively in the case of GERD combined with AF.GERD is closely related to AF occurrence.GERD may be an independent risk factor for atrial fibrillation,and atrial fibrillation is also considered as one of the extraesophageal symptoms of GERD.Inflammation may be involved in the pathogenesis of AF,and the degree of esophageal inflammation in GERD patients may affect to the atrium.GERD can also induce autoimmune response,leading to the occurrence and development of AF.