| Background:Gastric cancer is the fifth most common cancer worldwide.Helicobacter pylori infection,autoimmune status,genetic factors,and a variety of poor lifestyle habits are all involved in the development of gastric cancer.Chronic gastritis is initiated when pathological conditions influence the gastric mucosa over a long period of time.Studies have shown that chronic gastritis plays an important role in the development of gastric cancer,promoting the development of precancerous lesions such as intestinal epithelial metaplasia and heteroplasia,which eventually develop into intestinal-type gastric cancer.Characterized by concealed and atypical onset,gastric cancer has no obvious clinical symptoms at an early stage.Patients with gastric cancer are often in the middle or late stage when they are diagnosed the therapeutic effect is poor at this time.Therefore,it is of great clinical significance for the prediction and diagnosis of early gastric cancer.However,considering the invasive mode and misdiagnosis of endoscopic screening,requirements on skilled endoscopists and pathologists,and a high price,developing noninvasive methods to amend endoscopic screening would be highly needed.The development of transcriptome profiling revealed epigenetic changes and predictive signatures associated with the transformation of chronic gastritis into gastric carcinoma.Finding biomarkers in the transformation of gastritis to gastric cancer by transcriptome technology can facilitate for predicting early gastric cancer,and then take early intervention measures to prevent the occurrence of gastric cancer.Studies have found that the initiation of gastric diseases is associated with the dysbiosis of gastric flora and even intestinal flora,especially the decrease of probiotics such as butyrate-producing bacteria.Thus,restoring gastrointestinal microecology by supplementing probiotics has become an important intervention method for the prevention and treatment of gastric diseases.Clostridium butyricum,a butyric acid-producing bacteria,has been shown to be effective in the treatment of intestinal inflammatory diseases.However,the role of Clostridium butyricum in the prevention and treatment of gastritis is not completely clear,which needs to be further investigated.Recent studies have shown that probiotics combined with herbal medicines or prebiotics can improve the therapeutic effect when treating diseases caused by gastrointestinal mycobiome dysbiosis.However,due to the large differences in the results of different studies,the effect and mechanism of probiotics and microecology preparations on gastrointestinal microecology are still controversial,and further verification is required.Objective:The purpose of this study is to identify the key regulators in the complex molecular network of transformation of gastritis into cancer using transcriptome sequencing technology.Moreover,we further study the effect and mechanism of probiotics or microecology preparations containing probiotics and herbal medicine on gastritis using animal models of gastritis,which can provide a reasonable reference for the prediction,diagnosis,prevention,and treatment of early gastric cancer.Methods:(1)Both gastric cancer and paracancerous tissues were collected from 14 patients undergoing surgery for resection of gastric tumors in Zibo Municipal Hospital between 2019 and 2020.Then,RNA-seq was used to screen for the differentially expressed genes(DEGs)in gastric cancer and adjacent tissues.The potential key genes and pathways in gastric cancer were identified by GO and KEGG enrichment analysis,and thus the relationship between inflammation and gastric cancer would be assessed.In parallel,the human gastritis gene expression profile dataset GSE60427 obtained from the GEO database was used to screen for DEGs in gastritis in the R package.The common differentially expressed genes of gastritis and gastric cancer were selected,then PPI,GO,KEGG,and survival analyses were performed on these genes to find the key genes and pathways that may be involved in the transformation process from gastritis to carcinoma.(2)Experimental rat gastritis models were induced with synthetical method including MNNG,ranitidine hydrochloride,sodium deoxycholate,starvation and satiety disorder,and a high-salt diet.The expressions of inflammatory factors such as IL-1α,IL-6,IL-10,IFN-γ,and TGF-β1 in gastric mucosa of rats were detected by RT-qPCR.Then,the rats with gastritis were treated with Clostridium butyricum or Weichanghao(a compound microecology preparation composed of probiotics and Chinese medicinal herbs).There were four groups in the treatment phase:the gastritis model group,the prevention group with Clostridium butyricum(rats in this group were also given Clostridium butyricum before and during the modeling stage),the treatment group with Clostridium butyricum,and the treatment group with Weichanghao,and these interventions lasted 1.2,and 3 weeks.At different intervention stages,7-9 rats were randomly selected from each group to detect the expression of IL-1α,IL-6,IL-10,IFN-γ and TGF-β1 in gastric mucosa.Thus the inhibition effects and mechanism of Clostridium butyricum and Weichanghao in the progression of inflammation would be compared.After 3 weeks intervention,transcriptome sequencing was performed on the gastric mucosa of rats.GO and KEGG enrichment analyses were carried out to investigate the changes in gene expression and signaling pathways,which would help to further reveal the molecular mechanism of Clostridium butyricum and Weichanghao in blocking the transformation from inflammatory to cancer.To identify the beneficial metabolites of Clostridium butyricum,short-chain fatty acids and all other metabolites in the fermentation broth of Clostridium butyricum were assayed using LC-MS techniques.Results:(1)A total of 930 DEGs were screened in gastric cancer tissues by transcriptome sequencing.The results of GO and KEGG enrichment analyses suggested that immune and inflammatory processes play a key role in the progression of cancer.Biological process enrichment analysis revealed that DEGs in gastric cancer were mainly involved in the inflammatory response,myeloid leukocyte migration,cell proliferation and differentiation,cell adhesion,and vascular regeneration.KEGG enrichment analysis showed that 930 DEGs were enriched in various inflammation-related pathways such as PI3K/Akt,ECM-receptor interactions,AGE/RAGE,and PPAR pathway.Through the analysis of the GSE60427 dataset,a total of 940 DEGs of gastritis were identified,and there were 115 genes were common to DEGs in gastric cancer.The PPI,GO.KEGG.and surv ival analyses of these common DEGs suggested that APOE,ITGAX,C3,CLEC7A,HCK,TGFB1,TNFSF11,IGF1 and TREM1 may be key genes involved in the transformation of gastritis into cancer,and these key genes could be candidate biomarkers or potential therapeutic targets for early gastric cancer.(2)Rats were successfully induced into gastritis using the synthetical method.RT-qPCR results showed that the expressions IL-1α,IL-6,IL-10,IFN-γ,and TGF-β1 in stomach tissue of gastritis rats were significantly increased.After the intervention,the rats in the three intervention groups showed improvement in physical strength,mental status,and gastric mucosal morphology,and the expressions of IL-1α,IL-6,and TGF-β1 in gastric tissues were significantly reduced,indicating that both Clostridium butyricum and Wechanghao had therapeutic effects on gastritis.Several results demonstrated that Weichanghao had a better effect.The mechanism by which Clost ridium butyricum and Weichang inhibited gastritis and prevented the transformation of inflammation into cancer was interpreted by a comprehensive analysis of the transcriptome in gastric tissues,metabolites of Clostridium butyricum,and ingredients of Weichang.Clostridium butyricum inhibited the expression of cancer-promoting genes such as Atf4,Rps6kb1,Sgk3,Hspa5,and Fgd6,and blocked various inflammatory pathways including P13K/Akt,AGE/RAGE,and estrogen pathway.The anti-inflammatory effect of Clostridium butyricum depends on its beneficial metabolites such as short-chain fatty acids,terpenoids,linolenic acid,and steroidal saponins.Weichanghao decreased the expression of oncogenes like Hspa8,Hbegf,Cflar,and Dnaja1 in gastric tissue of rats,and suppressed pathways participating in inflammation and cancer such as TNF and AGE/RAGE pathway.The beneficial effects of Weichanghao might be attributed to the synergistic contribution of herbs,probiotics,and prebiotics.Conclusion:In this study,we found that APOE,ITGAX,C3,CLEC7A,HCK,TGFB1,TNFSF11,IGF1 and TREM1 may be key genes involved in the transformation of gastritis into cancer,which could be candidate biomarkers or potential therapeutic targets for early gastric cancer.In addition,animal studies showed that Clostridium butyricum and Weichanghao(a compound preparation composed of probiotics and Chinese medicinal herbs)were effective in the treatment of gastritis,and Weichanghao had a better effect.Both Clostridium butyricum and Weichanghao could prevent the malignant transformation of gastritis into gastric cancer by inhibiting the expression of oncogenes and blocking pathways related to inflammation and cancer such as PI3K/Akt and AGE/RAGE.Our research provides a reasonable reference for the prediction,diagnosis,and treatment of early gastric cancer during the process of transformation of inflammation to cancer. |
