Effect of Monotherapy Weifuchun Tablets on Chronic Atropic Gastritis: A Randomized Controlled Trial

Chronic atrophic gastritis is a common gastric disease which shows a positive correlation with the occurrence of gastric cancer. Traditional chinese medicines have been used in the treatment of chronic atrophic gastritis. There is no solid evidence for the curative effects of traditional chinese medicines. This study aimed to investigate the clinical ecacy of Weifuchun tablets on patients with chronic atrophic gastritis. 97 patients of chronic atrophic gastritis without current H. pylori infection were divided into two groups: an experiment group treated with Weifuchun and a control group treated with teprenone and folic acid for six months. Pathological examination of gastric biopsies and clinical symptoms assessment were performed for all patients before and after treatments. Primary outcome was the pathological response rate in each group evaluated by changes of histological scores.Secondary outcome was the clinical symptom scores.

Chronic atrophic gastritis (CAG) is a common troublesome gastric disease induced by various factors, such as H.pylori. infection, autoimmune attack and other uncertain environmental factors [1]. It is characterized by a chronic in ammatory condition of the gastric mucosa, with loss of normal glandular structures, and replacement by intestinal-type epitheluum, pyloric-type glands and brous tissue [1,2]. As most patients with chronic atrophic gastritis are asymptomatic, it is di cult to ascertain the exact prevalence rate.The Chinese Society of Digestive Endoscopology reported that 17.7% of the Chinese patients with gastrointestinal symptoms presented CAG at diagnostic upper endoscopy [3]. CAG shows a signi cant positive correlation with the occurrence of gastric cancer (GC), especially intestinal type GC [4].
Previous studies demonstrated that 0.1% and 1.44% of CAG developed into GC within 5 years of followup in Nertherland [5] and in China [6], respectively. It is of great importance to intervene and block the development of CAG to prevent the occurrence of gastric cancer.
At present, there are two main thoughts to treat CAG. One is to reduce the harmful stimulations to the gastric mucosa, and the other one is to enhance the defense of the gastric muccosa [7]. Therefore, investigators are keeping seeking effective treatments to block in ammation, promote tissue repair and reverse CAG, such as the regiments for Helicobacter pylori eradication [8] and using various gastric mucosal protectors to reduce tissue damage and protect gastric integrity [3]. Previous study reported that withdrawal of a drug which induced speci c loss of parietal cells could lead to complete restitution of the normal oxyntic mucosa [9]. This proposed that even the gastric mucosa with sever atrophy still had the capacity to self-renewal. That indicated that a small number of stem cells with the ability to proliferate and differentiate might still present in the gastric mucosa from patients with CAG.
In China, teprenone is frequently prescribed as an effective mucoprotective agent for gastric mucosal injuries and dyspepsia symptoms in chronic gastritis patients without H. pylori infection or history of gastroduodenal ulcer [10,11]. Folic acid, a one-carbon derivative of tetrahydrofolate, plays a critical role in the prevention of chromosome breakage and hypomethylation of DNA [12].It was reported that folic acid might treat atrophic gastritis by preventing or reversing precancerous lesions [13].We previously reported that combined application of teprenone and folic acid could signi cantly promote the reversal of gastric precancerous lesions, with an e cacy rate of pathological mucosa improvement reaching 49.8% [14].
The same as these western medicines, various Traditional Chinese Medicines (TCM) have been used in the treatment of CAG for decades [15][16][17]. However, there is no solid evidence for the curative effects of TCM because of the lack of high quality studies conducted with multi-center, large-sample size and longterm follow-up. Previous experimental study demonstrated that Weifuchun could reduce H. pylori-induced IL-8 and IL-4 production by blocking the NF-kappaB pathway [18]. This hinted that Weifuchun could be an effective drug to treat H. pylori-associated chronic gastritis. Although Weifuchun has been widely used in treatment of CAG in China,there is no high quality clinical study to report its e cacy on CAG among patients without H. pylori infection [19]. Thus, we carried out a randomized controlled clinical trial with long-term follow-up to determine the therapeutic effect of Weifuchun on CAG among H. pylori-negative or H. pylori eradicated successfully patients. Combined application of teprenone and folic acid was used as a positive control.

Subjects
This is a signle-center, randomized, controlled study. In total, 97 subjects diagnosed with CAG were recruited from the outpatient clinic from December 2017 to June 2018. All patients were required to provid a medical history and undergo gastroscopies with mucosa biopsies before and after interventions.

Inclusion criteria
Patients with CAG were diagnosed according to the criteria from Consensus on Chronic Gastritis in China (2017, Shanghai) [7] and the updated Sydney system [20]. Patients with H. pylori infeciton were required to eradicate H. pylori before the randomization. Only those whose H. pylori status were negative, determined by C 13 / C 14 urea breath test, were considered as eligible subjects to include.

Exclusion criteria
Patients were excluded if they were: 1) aged < 20 or > 70; 2) diagnosed or unable to exclude, malignant tumors, especially esophageal cancer and gastric cancer; diagnosed of peptic ulcer or Barrett's esophagus and other upper gastrointestinal diseases; 3) taking concomitant medication (such as proton pump inhibitors (PPI), prokinetic agents or other gastric mucosal protective agents) over 4 weeks during the trial; 4) pregnant or planning to pregnant, currently breastfeeding mothers; 5) serious primary diseases involving critical organs such as the heart, brain, lungs, liver, kidneys or the hematopoietic system; 6) unwilling to repeated endoscopy after treatment or to cooperate with researchers;.
Patients who met the inclusion and exclusion criteria described above were nally enrolled for randomization. Written informed consents and detailed records of concomitant mediciaion were obtained from all subjects.

Dosage and administration
Eligible patients were randomly assigned into two intervention groups. Patients in experiment group received four Weifuchun tablets (360mg) three times per day for six months. Weifuchun tablets were mainly prepared by medicinal herbs such as Hong Shen (Red Ginseng), Xiang Cha Cai (Isodon) and Zhi Ke (Citrus Aurantium), which was established by a high-performance liquid chromatography method (HuqingYuTang Pharmaceutical Co., Ltd., Hangzhou, Zhejiang, China; lot number: 20090120) [21].
Patients in control group received one teprenone tablet (50mg, Eisai China Pharmaceutical Co., Ltd. Suzhou, Jiangsu, China) and one folic acid tablet (10mg, Tianjing Lisheng Pharmaceutical Co., Ltd. Tianjing, China), three times per day for six months. Standard quadruple eradication regimen was given to the H. pylori-positive patients and two-week washout period was required to exclude the effects of H. pylori eradication before the randomization. During treatment, patients in both groups were required to discontinue any medications that could interfere with the results of the study.
Gastroscopy and mucosa biopsy Endoscopy procedures were performed in all participtants twice by senior endoscopists before and after interventions, respectively. Biopsy specimens were obtained from the lesser curvature of the antrum, 2-3 cm from the pylorus, gastric angle and middle of the corpus according to the updated Sydeny system [20].
The size and depth of the biopsy specimens were large enough to reach the muscularis mucosa [7]. H. pylori infection status was further determined by pathological examination.

Gastrointestinal symptom and adverse reaction assessment
Gastrointestinal symptoms of each participtant including epigastric pain, poor appetite, acid regurgitation, abdominal distention, belching, nausea and vomiting were evaluated by Gastrointestinal Symptom Rating Scale (GSRS) questionnaire before and after treatment as privously described [22]. Any adverse reactions occurred during the trials were recorded in details.

Outcome measurements
The primary outcome was the improvement of pathological ndings from the gastric mucosa after the treatments.The secondary outcome was the symptomatic bene ts from the treatments.
Pathological ndings including gastric mucosal in ammation, atrophy, intestinal metaplasis and allotypic hyperplasia were evaluated by a 4-point scale in severity to get a histological score (0: absent; 1: mild intestinal metaplasis and allotypic hyperplasis, and less than one third of the mucosa present in ammation and atrophy; 2: moderate intestinal metaplasis and allotypic hyperplasis, and one third to two third of the mucosa present in ammation and atrophy; 3: severe intestinal metaplasis and allotypic hyperplasis, and more than two third of the mucosa present in ammation and atrophy) [23].
The most advanced area in the stomach was selected as the object for diagnosis.
Individual gastrointestinal symptoms including (epigastric pain, poor appetite, acid regurgitation, abdominal distention, belching, nausea and vomiting) were assessed by a 4-point scale in severity as previously described: (0: absent, 1: mild symptoms easily to be tolerated; 2: moderate symptoms interfering with normal activities; 3: severe symptoms leading to inability to perform normal activities [22].
Response to treatment was determined by assessing changes of the scores from pathological evaluation and individual symptoms. Pathological effective was de ned by a > 1 point reduction in histological scores from baseline, and a ≥ 2 points reduction from baseline was further de ned as remarkably effective.

Statistical analysis
Continuous variables are expressed as mean ± standard deviation and categorical variables are expressed as frequencies (percentages). For comparisons of continuous variation from baseline to endpoint, Student's t -tests or Wilcoxon signed-ranks were performed for normal or abnormal distributed on variables, respectively. Response rates for the outcomes were analyzed by the Chi-squared tests or Fisher's exact tests. All statistical analyses were performed using GraphPad Prism 8.0 software (GraphPad, Inc., San Diego, CA, USA) and were considered statistically signi cant at a level of two side P < 0.05.

Subjects
By the time of project termination, a total of 52 patients had completed the study, as shown in Fig. 1. 24 patients (13 men, 11 women; mean age 55.29 ± 12.05 years) were from Weifuchun group and 28 patients (16 men, 12 women; mean age 54.21 ± 11.86 years) were from teprenone and folic acid group. Baseline demographic data including age and gender were similar between groups. Eleven patients in the Weifuchun group and nine patients in the control group occasionally took PPI or prokinetic drugs during intervention period less than four weeks.

Primary Outcome Assessment
The pathological response rates for in ammation, atrophy and intestinal metaplasia were 37.5%, 66.7% and 45.8% respectively in the Weifuchun treatment group and 21.4%, 66.7% and 42.9% in the control group, without statistically signi cant difference between groups. The remarkable response rates for atrophy and intestinal metaplasia were 33.3% and 20.8% respectively in the Weifuchun group and 33.3% and 21.4% in the folic acid group, with no signi cance between groups (Table 1 and Fig. 2).   Mucosal-protective agents and PPI are known to be the most commonly prescribed medications for chronic atrophic gastritis in China [3]. Teprenone (geranylgeranylacetone) is one kind of safe and effective mucosal-protective agent, which has been widely used for gastric mucosal injuries and dyspepsia symptoms in patients [10,24,25]. Folic acid plays a key role in DNA synthesis and cell replication [26]. It was reported that folic acid supplementation can improve gastric mucosal atrophy and intestinal metaplasia by involving DNA methylation and inhibition of cell proliferation [27,28]. Studies demonstrated that some patients with chronic atrophic gastritis had a low level of foliate and adequate folic acid supplementation could improve the histopathological status of the gastric mucosa and reduce the incidence of gastric cancer [29][30][31]. Therefore, we chose combined application of teprenone and folic acid as a positive control treatment for CAG in this study.
In China, TCMs have shown great advantages in the treatment of CAG, due to their multiple components and characteristics. There have been growing literatures demonstrating the notion that TCM herbal prescriptions can treat CAG effectively by relieving symptoms, inhibiting mucosal in ammation and reversing atrophy [32][33][34][35].Our previous study reported a kind of TCM, Weierkang tablets [36], could signi cantly improve atrophy and intestinal metaplasia, and promote the reversal of gastric precancerous lesions. In a multicenter randomized controlled trial from Beijing, Moluodan, could improve dysplasia scores in histopathology, and was superior to folic acid in improving epigastric pain, epigastric suffocation, belching and decreased appetite [15].
In our study, the pathological response rate for gastric atrophy and intestinal metaplasia in the Weifuchun group reached 66.6% and 45.80%, respectively, which represented the similar curative e cacy to the control group. Previous study reported that Weifuchun could dramatically improve clinical outcomes in patients with gastric precancerous lesions by targeting multiple pathways, such as involing NF-κB, RUNX3/TGF-beta/Smad, Hedgehog (Hh) and Wnt signaling pathways [37]. Red ginseng, a major component of Weifuchun, has been shown to induction of cell cycle arrest and apoptosis and inhibition of angiogenesis [38]. Amethystoides, another important ingredient of Weifuchun could promoting gastrointestinal motility [39]. Besides, the possible mechanism of Weifuchun in treating CAG and preventing malignant transformation might be regulating tumor suppressor genes and in ammatory factors by bio-network analysis [40]. Our results provided new evidences for regarding Weifuchun as an ideal treatment for reversal of chronic gastritis. Weifuchun has a good therapeutic effect on CAG and is worthy of further study.
S. Redéen et al. found that the severity of symptoms in patients with chronic gastritis did not correlate with endoscopic or histopathologic ndings [41]. Though Weifuchun showed similar improvement in pathological lesions as the western medicine regimen, both treatments failed to improve individual clinical symptoms in our study. Our clinical experience indicated that, patients often focus more on their physical symptoms, which could affect their quality of life and work. It was inevitable to take additional PPI or prokinetic drugs occasionally to relieve physical symptoms. This might account for the high rate of concomitant medications during this trial. We suppose that Weifuchun combined with prokinetic drugs or digestive enzyme agents may improve individual symptoms of CAG. Besides this, making individual adjustments to lifestyle and diet may also be a reasonable advice for relief of symptoms in CAG patients.
There are some limitations in our study. First of all, the accuracy and consistency of biopsies between pre-and post-treatment might affect the quality of study data and comparability of the results. Technique of the mucosa marking targeting biopsy (MTB) [42] by providing a safe and reliable long-term marker could be uesd in our future studies to improve the accuracy and consistency of biopsies. Secondly, the nal sample size was not large in this study. Multi-center and large-sample clinical studies are needed to further con rm the effect of Weifuchun in CAG.In addition, the drop-out rate in this study is high. Fifteen cases were lost to follow-up and 15 cases showed poor compliance. Long term (6 months) medication and required repeated invasive procedures (endoscopy) might be the main reasons for drop-out. In order to reduce the high drop-out rate, relavant measures should be taken in future studies, including expense deduction, periodic inspection, expert participation, facilitating consultation and prescription.

Conclusions
In summary, H. pylori-negative or H. pylori eradicated successfully chronic atrophic gastritis patients, Weifuchun treatment is effective for reducing pathological scores in chronic atrophic gastritis. Monotherapy of the Weifuchun treatments has the similar e cacy to the combination of teprenone and folic acid in the treatment of chronic atrophic gastritis.