CAG with IM is a benign, precancerous lesion with a tendency to develop into gastric cancer, in which IM is divided into complete IM and incomplete IM. Incomplete IM has more overexpressed oncogenes and molecular processes than intact IM, but the difference between histological subtypes of IM with or without GC is less than that of normal mucosa. Therefore, irrespective of the kind of IM, the possibility of canceration exists. A national multicenter cross-sectional study in China showed that the risk factors for CAG were related to not only common H. pylori infection, age, smoking, drinking, environmental, and drug factors but also insufficient acid production and pro-inflammatory genetic characteristics in the body3132. As no clear Western medicine exists for the prevention and treatment of precancerous lesions such as CAG with IM, the role of TCM in preventing precancerous lesions has gradually attracted the attention of researchers all over the world. 33,34
Professor Cai Gan, according to Li Dongyuan (ancient Chinese medicine scientists)stated “tonifying the spleen and stomach and dispelling evil fire” , “Evil fire is opposed to human vitality. If the evil fire is hyperactive, the human body will be unhealthy, and if the righteousness is sufficient, the human body will be healthy.”35. Gastric mucosal atrophy is the manifestation of weakness of spleen and stomach, and IM is caused by damp-heat, blood stasis, and heat for a long time, all belonging to the category of “evil fire.” Professor Cai Gan inherited and developed the theory of ancient Chinese medicine, produced E-Lian granule, took Liujunzi decoction (composed of C. pilosula, A. macrocephala, P. cocos, licorice, and Pi. ternata) as the base prescription for invigorating spleen and removing dampness; Radix Salviae Miltiorrhizae, A. sinensis, and Rhizoma Curcumae for promoting blood circulation, clearing heat, and removing blood stasis; and dandelion, Cop. chinensis, and H. diffusa for clearing heat, detoxification, and dispersing knots. E-Lian granule in the clinical treatment of CAG + IM has been proved so as to reverse gastric precancerous lesions, but the more comprehensive specific mechanism of the compound is not clear.
In this study, network pharmacology, GEO data, and reverse molecular docking were used to explore its microscopic mechanism based on integrated pharmacology platform TCMIP v2.0. TCMIP v2.0 originates from the data resources of the international authoritative database the Encyclopedia of Traditional Chinese Medicine, which integrates the data platform of TCM; integrates disease, syndrome, prescription, and medicine; simplifies complexity; and quickly mines the potential mechanism of famous doctors from different angles. The GEO database is a gene expression database created and maintained by the National Center for Biotechnology Information of the United States. Founded in 2000, it contains high-throughput gene expression data published by research institutions all over the world, all of which have been verified by clinical or experimental data, which increases the reliability of the TCM analysis.
In this study, 262 unique active components and 680 potential active targets of E-Lian granules were obtained using TCMIP V2.0 platform. All the drug ingredients obtained by TCMIP were adopted, but H. diffusa had no related data. Therefore, the active components of TCMSP and related literature were used to hang targets on the TCMIP V2.0 platform to obtain more reliable targets. At the same time, 2247 unique disease targets of CAG with IM were obtained by searching “Disease/Symptom Target Database” combined with GEO chip (GSE78523) and GeneCard database. “ELK–CAG+IM” compound targets and 38 core targets were obtained using Venn and Filter, respectively, such as ALB, TNF, PTGS2, RHOA, ESR1, HRAS, JUN, FOS, CASP3, and so forth. Through GO and KEGG enrichment analyses, E-Lian granule could reverse gastric precancerous lesions not only through the direct intervention of cancer pathway, gastric cancer pathway, and epithelial signal transduction in H. pylori infection but also through PI3K/AKT, VEGF, MAPK, cAMP, cGMP, Th1/Th2, and other pathways. Some of the conclusions of this study were also included in a similar study. The most important of the empirical prescription Qilian Shupi decoction is the cancer-related pathway (apoptosis, p53, and VEGF) in H. pylori infection and epithelial cell signal transduction.36
For example, in the PI3K/Akt pathway, the expression of PI3K and Akt phosphorylation was higher in the gastric precancerous lesion and gastric cancer groups than in the gastritis group, suggesting that the proliferative signal pathway of PI3K/Akt in gastric mucosa could be activated in the process of chronic gastritis–atrophic gastritis–IM–dysplasia–gastric cancer, which might be one of the pathogenic mechanisms of gastric cancer. 37
The infection rate of H. pylori was significantly higher in patients with early-stage gastric cancer than in those with advanced-stage gastric cancer (P < 0.05). Related experiments proved that H. pylori might promote the proliferation of tumor cells by activating the PI3K/Akt pathway in gastric cancer cells.38
With regard to the relationship between VEGF and CAG, related reports confirmed that the VEGF-634G > C polymorphism, especially the GG + GC genotype, was associated with the increased risk of gastric precancerous lesions transforming into gastric cancer and the increased level of VEGF39. The prescription of TCM under the principle of invigorating the spleen and regulating qi, activating blood circulation, and removing blood stasis might inhibit abnormal cell proliferation and differentiation of CAG by reducing the expression of ERK1, ERK2, and MAPK1 in the gastric tissue of rats. 40 After analyzing and comparing the expression of VEGF in the IM of gastric mucosa treated with H. pylori and eradication therapy. Rui et al. found that H. pylori could upregulate the expression of VEGF, but the expression of VEGF could not decrease to normal after the eradication therapy. A study comparing the positive rate of VEGF expression in the gastric mucosa of patients with acute simple gastritis and that of patients with CAG showed that the expression of VEGF in the gastric mucosa of patients with CAG significantly increased, but this was only one of the pathological changes of CAG and was not directly related to the degree of CAG. 41 Some studies also found that moxibustion at stomach meridian acupoints could significantly reduce the expression of proliferation factor VEGF in gastric mucosa of rats with precancerous lesions of CAG, inhibit the atypical proliferation of gastric mucosal cells, and promote the repair of the gastric mucosa.42
In terms of apoptosis, the scholars detected the proliferation index (PI), apoptosis index (AI), apoptosis–proliferation ratio, and apoptosis intensity in gastric antrum biopsies of patients with chronic superficial gastritis (CSG), CAG, and CAG + IM, respectively. The results showed a positive correlation between apoptosis and proliferation in CSG and CAG (r = 0.5475 and r = 0.5839), while the PI of CAG + IM PI was the highest. However, a negative correlation was found between apoptosis and proliferation (r = -0.6742). The intensity of apoptosis was extremely low, which was less than that of CSG. The atrophy of CAG might be caused by excessive apoptosis, while CAG + IM showed disturbance of apoptosis and excessive proliferation, which was similar to the cellular biological manifestations of dysplasia and carcinoma 43. Other related studies showed that precancerous lesions were nonspecific processes characterized by long-term cell degradation and proliferation. However, because it was a benign disease, even if some oncogenes were activated, cell metabolism might still be insufficient, especially nucleic acid metabolism and DNA repair, but these processes were reversible. 44 B-cell lymphoma-2(Bcl-2)is one of the most important oncogenes in the study of apoptosis. The low expression of the Bcl-2 gene suggests atrophic gastritis, IM, and intestinal-type gastric adenocarcinoma.45
In terms of the molecular mechanism of cAMP and cGMP, team 42 of Zhang Jingren (one of the masters of TCM in China) detected plasma cAMP and cGMP in 87 cases of atrophic gastritis with spleen qi deficiency syndrome. The two indexes of all patients were found to be lower than the normal low limit. Zhang's Weiwei'an granule (which is also a TCM prescription for treating atrophic gastritis) improved significantly after one or two courses of the treatment. To some extent, it reflected the essence of spleen and stomach qi deficiency of CAG.46 A report in 2005 showed that acupuncture could increase the content of cAMP in gastric mucosa and reduce the content of cGMP.47
Regarding the regulation of Th1/Th2 balance, for example, the Department of Gastroenterology of Hebei Hospital of TCM used the empirical prescription Huazhuo jiedu Hewei recipe (the empirical prescription of Hebei Hospital of TCM for treating atrophic gastritis) to treat CAG for 12 weeks; it upregulated Th2-type cytokine interleukin-4 (IL-4) and decreased the VEGF and EGF levels, but did not affect serum Th1 cytokine interferon-gamma (IFN-γ) level, thus regulating the Th1/Th2 balance 48. In a study of Shengyang Yiwei decoction (a classical prescription for the treatment of digestive diseases recorded in TCM) and the treatment of CAG rats, opposite results were obtained. Shengyang Yiwei decoction could increase the secretion of IFN-γ and decrease the secretion of IL-4 in gastric tissue and serum of rats, thus regulating the balance of Th1/Th2.49
In addition, E-Lian granule might reverse CAG + IM and improve the symptoms of dyspepsia through the brain or gastrointestinal hormone-related signals, such as cholinergic, 5-hydroxytryptamine (5-HT4), and dopaminergic receptors. Dopamine receptor antagonists, such as metoclopramide hydrochloride and domperidone, act directly on the gastrointestinal wall, increase the tension of the lower esophageal sphincter, prevent gastroesophageal reflux, and promote gastric emptying. E-Lian granule could stimulate selective 5-HT4 receptors, such as cisapride and mosapride, and induce the release of acetylcholine by stimulating cholinergic intermediate neurons in the gastrointestinal tract and 5-HT4 receptors in the myenteric plexus, thereby increasing gastrointestinal motility and improving the symptoms of dyspepsia. Previous clinical studies confirmed that the E-Lian granule was effective in improving epigastric pain, epigastric tiredness, and fatigue (P < 0.05 or P < 0.01).50