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Effect of Helicobacter pylori Eradication on the Development of Reflux Esophagitis and Gastroesophageal Reflux Symptoms: A Nationwide Multi-Center Prospective Study

Background/Aims A two-year, prospective, nationwide multicenter study was undertaken to evaluate the effect of Helicobacter pylori eradication on the development of reflux esophagitis (RE) and gastroesophageal reflux disease (GERD) symptoms in the Korean population. Methods In total, 1,489 subjects without RE were enrolled at the outpatient clinics of 12 tertiary hospitals nationwide, and 452 subjects underwent follow-up (F/U) for 2 years to evaluate the development of RE and GERD symptoms. Results RE was found in 33 subjects (7.3% of 452 subjects) and 14 subjects (7.3% of 192 subjects) during the first and second year of F/U, respectively. H. pylori status was not associated with the development of RE. RE was found in six (9.0%) of 67 H. pylori-negative patients, in 26 (11.2%) of 233 eradicated subjects and in eight (7.0%) of 114 noneradicated subjects (p=0.532). Multivariate analysis showed that age ≥60 years (odds ratio [OR], 7.11; 95% confidence interval [CI], 1.92 to 26.41), alcohol consumption (OR, 4.43; 95% CI, 1.03 to 19.19) and F/U cholesterol levels ≥200 mg/dL (OR, 5.03; 95% CI, 1.32 to 19.17) were significant risk factors for the development of RE. There was no significant difference in the development of GERD symptoms or weight according to H. pylori status during the 2-year F/U. Conclusions Eradication of H. pylori did not affect the development of reflux esophagitis or GERD symptoms among patients in outpatient gastroenterology clinics in South Korea.


INTRODUCTION
Helicobacter pylori (H. pylori) infection plays a major role in the pathogenesis of peptic ulcer disease, chronic gastritis, gastric mucosa-associated lymphoid tissue lymphoma (MALToma), and the development of gastric cancer. However, its role in reflux esophagitis (RE) or gastroesophageal reflux disease (GERD) is not fully understood. In addition, the effect of H. pylori eradication on RE or GERD is unclear. Several reports have shown an association of H. pylori eradication with the development of RE [1][2][3][4][5] or GERD symptoms. 6,7 However, contradictory results have been reported [8][9][10][11] and sometimes a beneficial effect of H. pylori eradication on GERD has been observed. [12][13][14][15][16] This issue regarding the effect of H. pylori eradication on the development of RE is important because RE often requires chronic treatment with a proton pump inhibitor (PPI) due to frequent relapse of symptoms. 17 The Maastricht III Consensus Report already extended the recommendations of H. pylori eradication to patients with dyspepsia, along with those who chronically use PPI or nonsteroid anti-inflammatory drugs (NSAID). 18 In South Korea, the eradication of H. pylori has been rapidly increasing 19,20 as well as the prevalence of RE 21 or Barrett's esophagus. 22 Therefore, if RE development increases after eradication of H. pylori, then the eradication of H. pylori should proceed with greater caution in South Korea.
Based on this background, a nationwide multicenter prospective study was undertaken to evaluate the effect of H. pylori eradication on the development of RE and GERD symptoms in South Koreans. In addition, factors affecting RE development were evaluated.

Subjects
This study was designed by the Scientific Committee of the Korean College of Helicobacter and Upper Gastrointestinal Research. A workshop was held on June 25, 2005 to approve the study design and to standardize the description of RE and gastroesophageal reflux symptoms in the questionnaire. Feedback from experts in this field and practical field tests were conducted to validate the reflux symptom questionnaire over 3 months. 23 Patients with gastritis, duodenal ulcers (DU) in the active or healing stages, and benign gastric ulcer (BGU) were enrolled in July 2006 for 1 year at the Gastroenterology Department of 12 tertiary hospitals nationwide. Most of the participants had gastroduodenal discomfort and underwent endoscopy to evaluate whether the symptoms were related to specific diseases such as gastric cancer, DU, BGU, or RE. Subjects with a previous history of gastric surgery, H. pylori eradication treatment, or systemic diseases requiring chronic medication were excluded. In addition, subjects current taking or with a history of taking H2 blocker or PPI within at least 1 month were excluded from this study. Subjects with upper gastrointestinal endoscopy results showing evidence of RE, gastric cancer, dysplasia, MALToma, or esophageal cancer were excluded from this study. Additionally subjects with endoscopic findings of BGU scars or DU scars were also excluded from enrollment to avoid confusion in the analysis. The Institutional Review Board of the 12 participating hospitals approved this study, and written informed consent was obtained from all participants.

Upper gastrointestinal endoscopic examinations
All study subjects underwent upper gastrointestinal endoscopic examinations. The endoscopic findings of RE in the lower esophagus were classified according to the Los Angeles (LA) classification based on the longest length of the mucosal break, and confluence of erosions. A hiatal hernia was endoscopically defined as a distance from the esophagogastric junction to the diaphragmatic impingement of more than 1 cm. The esophagogastric junction was defined as the proximal margin of the gastric mucosal fold. 24

Questionnaire
Patients who agreed to participate in the study were interviewed in person by an assistant using the gastroesophageal reflux questionnaire, before the endoscopy. The questionnaire included questions about seven reflux symptoms including heartburn, acid regurgitation, chest pain, hoarseness, globus sensation, cough, and epigastric soreness. The questions were as follows: 1) Have you experienced heartburn? 2) Have you experienced acid regurgitation? 3) Have you experienced chest pain? 4) Have you experienced hoarseness? 5) Have you experienced a globus sensation in the throat? 6) Have you experienced a frequent cough? 7) Have you experienced epigastric soreness? If the patients replied yes to at least one of the seven questions about reflux symptoms, then they were further questions about the reflux symptom periods. Body mass index (BMI) and the results of biochemical testing including glucose, cholesterol, triglyceride, and H. pylori tests were recorded by assistants.

H. pylori tests
Two biopsy specimens from the antrum and two from the body were fixed in formalin, and examed for the presence of H. pylori by modified Giemsa staining. One specimen from the lesser curvature of the antrum and one from the corpus were subjected to rapid urease testing (Campylobacter-like organism (CLO) test; Delta West, Bentley, Australia). H. pylori infection was diagnosed when at least one of these tests gave positive results.

Serum pepsinogen levels
Fasting serum was collected from all subjects at the beginning of the study. The samples were centrifuged immediately and stored at -70 o C until used. Serum concentrations of pepsinogen (PG) I and II were measured using a latex-enhanced turbidimetic immunoassay (L-TIA) (HBi Co., Seoul, Korea, imported from Shima Laboratories, Tokyo, Japan), and PG I to PG II ratios (PG I/II) were calculated. 25,26

Eradication treatments and follow-up study
When a subject with duodenal or gastric ulcers was found to be H. pylori-positive, the standard 7-day triple therapy (PPI b.i.d., 500 mg b.i.d. clarithromycin, and 1 g b.i.d. amoxicillin) was started for H. pylori eradication. Subjects with simple gastritis also received H. pylori eradication therapy when requested by the patient. To determine whether H. pylori had been eliminated, 13 C-urea breath testing was performed at least 4 weeks after completing the triple therapy. If H. pylori infection was found to present persistent after the first-line PPI-based triple therapy a 1-or 2-week course of bismuth-based quadruple therapy was started: 20 mg b.i.d. esomeprazole, and 300 mg q.i.d. tripotassium dicitrate bismuthate (DENOL®; Greencross, Seoul, Korea, three tabs taken 30 minutes before meals and one tab taken 2 hours after dinner), 500 mg t.i.d. metronidazole, and 500 mg q.i.d. tetracycline.
Initially enrolled subjects underwent endoscopy and filled out a questionnaire every year for 2 years. The endoscopists were unaware of the questionnaire results. Patients with recurrence or occurrence of BGU were excluded from this study. Every participant was asked whether new GERD symptoms (heartburn, acid regurgitation, chest pain, hoarseness, globus sensation, cough, and epigastric soreness) had developed during follow-up period. In addition, patients answered questions about duration and frequency of their symptom (a. 1-2 times per year, b. 1-2 times per month, c. 1-2 times per week, d. 3-4 times per week, e. daily). In addition, they were asked whether they took PPI or H 2 blocker to relieve these GERD symptoms, and were also asked about the duration of PPI or H 2 blocker treatment. If the participants refused the follow-up endoscopy at the second year in after numerous requests, an interview was conducted to evaluate changes in weight, smoking habits, and alcohol consumption. In addition, the questionnaire regarding gastroesophageal reflux symptoms was re-administered. The main reason for refusal of endoscopy at the second year was that the patient disliked undergoing the procedure.

Statistical analysis
The patients were categorized according to age in 10-year intervals. New development of reflux esophagitis and GERD symptoms (heartburn, acid regurgitation, chest pain, hoarseness, globus sensation, cough, and epigastric soreness) follow-up period was analyzed according to H. pylori eradication. Chi-square tests were performed to assess the association between each covariate and development of RE. Covariates that showed significant association by the chi-square tests were considered for the multiple logistic regression analyses. Model fit was assessed by the Hosmer-Lemeshow goodness-of-fit test. The variables that were significant in the model or improved model fit were included in the final model. The final models showed an appropriate goodness of fit (p>0.10). All analyses were performed using SAS statistical software (SAS Institute, Cary, NC, USA). A p-value of less than 0.05 was considered statistically significant.

Characteristics of participants
In total 1,489 subjects (855 males, 57.4%; 634 females, 42.6%) were included in the study. The mean age was 51.3 years and all participants were ethnically Korean. Follow-up endoscopy with the questionnaire was performed 1 year later and 452 subjects received an endoscopy 1 year after enrollment. The characteristics of 452 subjects are described in Table 1 including gender, age, BMI, endoscopic diagnosis, smoking habit, alcohol consumption, hiatal hernia, glucose, cholesterol, triglyceride, H. pylori infection, and pepsinogen levels. There were no statistical differences in these variables between the 1,489 initially enrolled and 452 follow-up subjects. Four hundred fifty-two subjects had DU in the active or healing stage (68, 15.1%), BGU in the active or healing stage (115, 25.4%), or gastritis (269, 59.5%). There were 385 individuals (85.2%) with H. pylori infections and the remaining 67 subjects (14.8%) were not infected. When reflux symptoms of 452 subjects were analyzed at enrollment, epigastric soreness was the most frequent symptom, reported by 127 subjects (28.1%), followed by heartburn (27 subjects, 6.0%), chest pain (25 subjects, 5.5%), acid regurgitation and globus sensation (21 subjects, 4.6%), cough (7 subjects, 1.5%), and hoarseness (2 subjects, 0.4%). Data are presented as mean±SD or number (%).

H. pylori treatment results and weight change during the 2-year follow-up
Among the 452 subjects who had received an endoscopy 1 year after enrollment, 192 (42.5%) another endoscopy in the second year of follow-up, and the remaining 260 subjects (57.5%) responded to a telephone interview. Initially, 67 subjects (14.8%) were diagnosed as H. pylori-negative. Among 385 H. pylori-positive cases, 233 cases (51.6%) were found to be eradicated, 114 subjects (25.2%) were not treated or the infec-tion was not eradicated, and 38 (8.4%) did not receive followup H. pylori tests ( Table 2). Participants were weighted prior to enrollment and 1 and 2 years after enrollment, there was no significant difference in weight based on the H. pylori status during follow-up.      lori infection (Table 3). New development of epigastric soreness was the most common symptom, especially in the H. pylorinegative group (10.5%) but there was no statistical significance associated with H. pylori infection. The percentage of participants with new development of other reflux symptoms such as heartburn, acid regurgitation, chest pain, hoarseness, globus sensation and cough were found to be 0% to 6.0%, there was no significant difference associated with H. pylori eradication (Table 3).

Development of RE during the 2-year follow-up
RE was found in 33 subjects (7.3% of 452 subjects) and 14 subjects (7.3% of 192 subjects) at the first and second year of follow-up, respectively. Among the 33 patients who developed RE at the 1st follow-up 30 (90.9%) showed LA-A and 3 (9.1%) showed LA-B (Fig. 1). Among the 30 subjects with LA-A at the first follow-up, 27 did not have RE at the second year and three subjects were found to have persistent LA-A (Fig. 1). Among the three subjects with LA-B, only one had persistent LA-B at the second year follow-up. Ten subjects who did not have RE at the first year follow-up turned out to have LA-A (nine subjects) or LA-B (one subject) at the second year follow-up. Finally, RE was found in 14 subjects at the second year follow-up (Fig. 1). In summary, there were 33 RE patients at the first year followup, and cases of RE were found in 14 patients RE patients at the second year follow-up. Three patients continuously had LA-A and one patient persistently showed LA-B at the first and second year follow-up, respectively. In total, RE developed in 43 patients during the 2-year follow-up period.

Analysis of risk factors for RE development
To evaluate the risk factors for RE during the 2-year followup period, univariate analysis was performed to examine gender, age, H. pylori eradication, smoking habit, alcohol consumption, initial finding of hiatal hernia, underlying disease, serum pepsinogen, weight changes, and cholesterol and triglyceride levels ( Table 4). Gender showed no correlation with RE development but there was an increase of RE incidence with age (14.4% ≥60 years vs 6.4% 40 to 59 years). H. pylori eradication was not associated with RE development of RE. That is, among 67 H. pylori-negative patients, RE development was found in 6 (9.0%), RE development was found in 26 (11.2%) out of 233 subjects in which H. pylori had been eradicated, and RE developed in 8 (7.0%) out of 133 patients with persistent H. pylori infections (p=0.532). Smoking showed a marginal difference in RE development, but alcohol consumption had a statistically difference impact (Table 5). That is, the nonalcohol-nonalcohol group had 4.3% RE, the alcohol-nonalcohol group 0%, which was lower than the nonalcohol-alcohol group 7.5%, and the alcoholalcohol group showed 15.2% (p=0.006). The presence of an underlying disease was found to influence the development of RE. RE development was frequent observed in patients with BGU (16.5%), this was more common than in patients with DU (11.8%) or gastritis (5.9%) (p=0.012). Serum pepsinogen level was also associated with development of RE. PG I levels were higher in the group that developed RE (75.9±17.9 ng/mL) than in the group that did not develop RE (60.6±12.1 ng/mL) (p=0.035); however, there was no significant difference in the PG II levels between these 2 groups. Finally, the PG I/II ratio was higher in the RE group (4.41±1.28) than in the group without RE (3.68±1.11) (p=0.043). Initial body weight was not significantly associated with the development of RE. However, body weight was increased in the RE group at follow-up, and this was significantly associated with RE development (p=0.028). Similar to body weight, cholesterol levels also showed a significant association with the development of RE in the univariate analysis. Initial cholesterol levels were similar between the groups with and without RE. However, the follow-up cholesterol levels were higher in the RE group (211.5±10.0) than in the group without RE (188.7±8.1). In contrast, triglyceride levels showed no sig-

DISCUSSION
Since H. pylori is a major cause of gastric cancer, eradication of H. pylori infections has become an important goal for the prevention of gastric cancer in high-risk groups. In South Korea, H. pylori eradication has rapidly increased from 13.9% in 2005 19 to 37.4% in 2007 among subjects presenting for a health checkup. 20 In addition, the prevalence of RE 21 or Barrett's esophagus is increasing 22 similar to other Asian countries. Furthermore, since the rates of on demand H. pylori treatment or relapse in RE patients are also high in South Korea, 15 it is very important to determine whether the risk of GERD or RE increases after H. pylori eradication. There have been several reports suggesting that GERD occurs after eradication of H. pylori in patients with or without peptic ulcers. 8,9,[27][28][29][30] This development of GERD following the eradication of H. pylori might be related to an increase in refluxate acidity due to the disappearance of the neutralizing effect of bacterial ammonia production and corpus gastritis induced by persistent H. pylori infection. 31,32 In contrast, there have been some reports of the beneficial effect of H. pylori eradication for ameliorating symptoms of GERD. [12][13][14][15][16] The reason for these conflicting reports could originate from the underlying gastric mucosal status. For instance, a meta-analysis showed that the frequency of GERD is not increased after eradication of H. pylori among dyspeptic patients, but there was the risk of GERD development increased 2-fold after H. pylori eradication among patients with peptic ulcers. 33 Similarly, the univariate analysis of the present multicenter prospective study showed that the development of RE was more frequent after treating active or healing cases of BGU (16.5%) or DU (11.8%) than gastritis (5.9%) (p=0.012) although this significance was not found in the multivariate analysis. In addition, PG I levels and PG I/ II ratios, which have been used as indicators of acid secretion, 34 were found to be significantly higher in the group that developed RE compared to the nonRE group. This could support the hypothesis that acid secretion determines the development of RE.
A lack of association between RE or GERD symptoms and H. pylori eradication in the present study might reflect the lower prevalence of RE or GERD in South Korea compared to other Eastern as well as Western countries. The prevalence of RE has increased in Japan from 3% in the late 1970s to 10% to 15% in the late 1990s based on the results of upper-gastrointestinal endoscopies, 35 and from 2.7% in the early 1990s to 13.4% in 2000 to 2001 in a Malaysian study. 36 In a South Korean study conducted in 1999, the prevalence of RE was 5.3% for patients with gastrointestinal symptoms, 37 and was 3.4% in a group of asymptomatic subjects presenting for a health check-up in 2001. 38 In 2006 RE prevalence increased to 7.9% among subjects presenting for a health check-up, 21 which is still lower than the levels reported in other Asian countries like Japan 35,39 and Malaysia. 36 Furthermore, the degree of RE found in the present study was relatively mild. That is, 39 out of 43 RE subjects (90.7%) had LA-A, while only four were found to have LA-B (9.3%). In addition, among 30 patients who participated in the follow-up during the second year 27 patients did not show definite mucosal breaks around z-line, suggesting that RE could be transient, which is consistent with the findings of a Japanese study. 5 Still, the precise reasons why the prevalence of RE or GERD is lower in South Korea than other Asian countries is not clear. This might be related to life style factors such as the consumption of certain kinds of food or a high prevalence of atrophic gastritis 21 due to the high infection rate of H. pylori. 19 Comparative studies on the prevalence of RE or GERD and its associated factors among South Korea and other Asian countries could help clarify this issue.
Another explanation for a lack of association between RE and H. pylori eradication in the present study could be related to the absence of definite weight gain after H. pylori eradication in Koreans. Previous studies showed that H. pylori infection is inversely related to obesity 40,41 and that increased body weight is often observed after H. pylori eradication. 42 Since ghrelin is mainly synthesized and secreted by the gastric mucosa, the inverse effect of H. pylori infection on body weight has been attributed to the difference in plasma ghrelin concentrations in patients with or without H. pylori infections. 43 However, in the present study there was no definite weight change observed according to H. pylori infection at the enrollment or H. pylori eradication after 2 years of follow-up. Since most of the previous studies were performed during short term follow-up (e.g., 12 weeks) after successful eradication, 40 it is possible that weight gain after H. pylori infection, which could be related to the increase of ghrelin, might be temporary. However, as the weight gain after successful H. pylori eradication was frequently observed in Japanese patients, 42 further study is necessary to confirm this finding in Koreans.
One of the characteristics of Korean patients with GERD is that they frequently complain of epigastric soreness or epigastric pain, which is usually classified as symptom of functional dyspepsia. This was similar to a Korean report showing that 18.8% patients that complained of functional dyspeptic symptoms were found to have RE and while 27.6% of patients with epigastric pain had RE. 44 This is the reason why epigastric soreness was included as one of the reflux symptoms in our study. When the occurrence of these symptoms were analyzed based on H. pylori eradication during the 2-year follow-up, there was no association of new development of any GERD symptoms (heartburn, acid regurgitation, chest pain, hoarseness, globus sensation, cough, or epigastric soreness) with H. pylori eradication. This fingding is rather different from the results of a Japanese study. 14 In the present study, the incidence of RE per year regardless of H. pylori eradication was 7.3%, which is similar to the RE prevalence rate, 7.9% reported in 2006. 21 Since RE was transient in the present study, new development and the disappearance of RE might be somehow balanced among Koreans. In addition, the rate of newly developed GERD symptoms (heartburn, acid regurgitation, chest pain, hoarseness, globus sensation, cough, and epigastric soreness) during the 2-year follow-up was only 0% to 10.45%. This is similar to the finding that the prevalence of GERD according to symptom occurrence was found to be 5.1% among health check-up subjects in 2006, 21 and 3.5% in the rural and urban areas of Asan-si in South Korea, in a 2000 to 2001 survey (n=1,471). 45 So far, there has been no prospective multicenter study examining the incidence of RE or GERD symptoms, nor the relationship of H. pylori eradication with RE or GERD symptoms in South Korea. From this point of view, the multicenter, prospective study presented here is very meaningful as it has provided clues regarding the incidence of RE and GERD symptoms among South Koreans.
The significant risk factors for reflux esophagitis are known to include male gender, alcohol consumption, smoking, BMI ≥25, abdominal obesity, and metabolic syndrome. 21,46,47 Similarly, the multivaraiate analysis of the present study showed that an age ≥60 (OR, 7.11; 95% CI, 1.92 to 26.41), alcohol consumption at follow-up (OR, 4.43; 95% CI, 1.03 to 19.2), and elevated cholesterol levels at follow-up (OR, 5.03; 95% CI, 1.32 to 19.2) were risk factors for the development of RE. Therefore, persons of advanced age (≥60 years-old) with significant alcohol intake and cholesterol levels over 200 mg/dL should be watchful for the development of RE.
The present study had several limitations. First, there was a significant 1-year endoscopic follow-up loss after initial enrollmen. Most of this follow-up loss may have occurred when the patients did not have any reflux or dyspeptic symptoms. Biennial endoscopy or barium gastrography has actually been recommended for gastric cancer screening starting at the age of 40 by the South Korean government since 2001. However, there was no statistical difference in gender, age, BMI, endoscopic diagnosis, smoking habits, alcohol use, occurrence of hiatal hernias, cholesterol levels, triglycerides, H. pylori infection, or pepsinogen levels between the 1,489 initially enrolled patients and 452 follow-up subjects. Second, the follow-up period was probably too short to detect the effect of H. pylori eradication on RE development. However, as pepsinogen I or PG I/II ratio reflecting the gastric acid secretion 34 has been already increased up to the level of H. pylori-negative group 1 year after H. py-lori eradication in Koreans, 21 there might be little possibility of further occurrence of RE due to H. pylori eradication after longtem follow-up more than 2 years.
In conclusion, our study demonstrated that the eradication of H. pylori did not cause the development of RE or GERD symptoms over 2 years of follow-up. These results support the hypothesis that H. pylori eradication could be performed in South Korea without concerns about an increased risk of GERD or RE development.

CONFLICTS OF INTEREST
No potential conflict of interest relevant to this article was reported.