Zuojin Pill attenuates Helicobacter pylori-induced chronic atrophic gastritis in rats and improves gastric epithelial cells function in GES-1 cells

https://doi.org/10.1016/j.jep.2021.114855Get rights and content

Highlights

  • Zuojin Pill alleviates chronic atrophic gastritis precancerous lesions of gastric carcinoma in GES-1 cells induced by Helicobacter pylori.

  • ZJP attenuates H. pylori-induced CAG by regulating the JMJD2B/COX-2/VEGF axis signaling pathway.

  • ZJP has therapeutic effects on H. pylori-induced CAG via inhibiting the HMGB1/NF-κB signaling pathway.

Abstract

Ethnopharmacological relevance

Zuojin pill (ZJP), a classical Chinese medicine formula, has been widely applied in Chinese clinical practice for the treatment of gastric injury such as acute gastric lesion, acute gastric mucosal injury, chronic unpredictable mild stress, gastroesophageal reflux disease, etc, thereby exerting anti-chronic atrophic gastritis (CAG) effects in traditional Chinese herbal medicine.

Aim of the study

This study was aimed to explore the therapeutic effects and molecular mechanisms of ZJP on Helicobacter pylori (H. pylori)-induced CAG based on the comprehensive approaches.

Materials and methods

Sprague-Dawley rats were infected with H. pylori for 8 weeks to establish CAG model. Then, rats in the ZJP groups received doses of 0.63, 1.26, and 2.52 g/kg ZJP for 4 weeks. Therapeutic effects of ZJP on serum indices and the histopathology of the gastric were analyzed in vivo. Moreover, GES-1 cells were infected with H. pylori to establish gastric epithelial cell injury model in vitro. Cell viability and gastric epithelial cell morphology were detected by a high-content screening (HCS) assay. Furthermore, the relative mRNA and protein expression of JMJD2B/COX-2/VEGF axis and HMGB1/NF-κB signaling pathway in vivo and in vitro were determined by RT-PCR and Western Blotting, respectively.

Results

The results showed that the therapeutic effects of ZJP on CAG rats were presented in down-regulation serum biochemical indices and alleviating histological damage of gastric tissue. ZJP could dose-dependently decrease the serum IL-6, MCP-1, PGE2, TNF-α, and VEGF level and significantly improved gastric tissue inflammatory lesions. Besides, ZJP has an effect on increasing cell proliferation of GES-1 cells, ameliorating H. pylori-induced gastric epithelial cell damage. It was found that ZJP has a down-regulating effect on inflammatory reaction and could inhibit the relative mRNA and protein expression of JMJD2B/COX-2/VEGF axis and HMGB1/NF-κB signaling pathway in vivo and in vitro, including JMJD2B, COX-2, VEGF, VEGFR1, and VEGFR2, which in turn reduced the damage of gastric mucosal cells.

Conclusions

The results suggested that ZJP exerts therapeutic effects on H. pylori-induced CAG by inhibiting the JMJD2B/COX-2/VEGF axis and HMGB1/NF-κB signaling pathway. These findings deeply explained why ZJP could be used to treat CAG clinically and clarified its pharmacological effect and potential mechanism in the treatment of CAG.

Introduction

Chronic atrophic gastritis (CAG) is a universal disease of the digestive system and one of the most common health concerns worldwide. Helicobacter pylori (H. pylori) infection is the strongest risk factor for the development of gastric carcinogenesis (Crew and Neugut, 2006). H. pylori infection can induce CAG, which develops during the premalignant periods of intestinal metaplasia and dysplasia, eventually leading to gastric cancer (Correa and Piazuelo, 2012). At present, traditional treatment options for H. pylori, include conventional triple therapy, bismuth-based quadruple therapy and proton pump inhibitors (Wang et al., 2017a). Regardless of the type, these conventional therapies can cause a series of serious adverse reactions, such as abdominal pain and constipation, as well as a decline in eradication rates. Therefore, the discovery of novel and safe drugs for clinical treatment is greatly needed. Copious evidence from China has indicated that traditional Chinese medicine (TCM) possess unlimited potential in treating H. pylori-induced CAG. As alternative therapies, TCMs are gaining increasing attention as clinical treatments worldwide (Wang, 2014). Zuojin pill (ZJP), which was recorded in the Danxi's experiential therapy, has been used for more than 700 years to treat gastrointestinal diseases. However, the underlying mechanism of ZJP in the treatment of H. pylori-induced CAG remains unclear.

ZJP consists of Coptidis Rhizoma (CR) and Euodiae Fructus (EF) in a 6:1 ratio (w/w) and was initially reported as a treatment for gastrointestinal disorders in an ancient medicine treatise during China's Yuan Dynasty. CR is the dried rhizome of Coptis chinensis Franch. (Ranunculaceae), Coptis deltoidea C.Y. Cheng et Hsiao, or Coptis teeta Wall. (Chinese Pharmacopoeia Commission, 2015). In the clinic, it is often utilized to treat diarrhea, abdominal fullness, vomiting, jaundice, toothache, diabetes and eczema (Wang et al., 2019). EF is the dried immature fruit of Euodia rutaecarpa (Juss.) Benth. (Rutaceae), Euodia rutaecarpa (Juss.) Benth. var. officinalis (Dode) Huang or Euodia rutaecarpa (Juss.) Benth. var. bodinieri (Dode) Huang (Chinese Pharmacopoeia Commission, 2015). EF is widely applied for the treatment of inflammation, headache, and hypertension (Xu et al., 2012). Alkaloids are the primary compounds of CR and EF, including berberine, coptisine, palmatine, evodiamine and rutaecarpine (Guo et al., 2019). ZJP is listed in the Chinese Pharmacopoeia (2015 edition) and is commonly prescribed for clinical patients suffering from esophagitis, gastritis, peptic ulcer and other disorders. To date, ZJP has been widely used in clinical applications. The mechanism by which ZJP treats CAG is still unclear. This study elucidates the effects of ZJP on H. pylori-induced CAG and the associated molecular mechanisms.

In this study, a H. pylori-induced CAG model was established to assess the therapeutic effectiveness and safety of ZJP in reducing inflammation and gastric mucosal cell injury. Gastric histopathology and serum inflammatory factors were assessed to evaluate the inflammatory levels and mucosal damage of CAG. The improvement of cell viability and morphology by ZJP was also detected by a cell model. In addition, the relative mRNA and protein expression levels of the JMJD2B/COX-2/VEGF axis and HMGB1/NF-κB signaling pathway were detected to better understand the therapeutic effects and potential mechanism of ZJP on H. pylori-induced CAG.

Section snippets

Materials

Coptidis Rhizoma (Lot: 18011901) and Euodiae Fructus (Lot: 17021602) were purchased from Beijing Lvye Pharmaceutical Co., Ltd. (Beijing, China). Product quality inspection document numbers were CP-18-01-22 (Coptidis Rhizoma) and CP-17-02-11 (Euodiae Fructus). All inspection results demonstrated that the quality of Coptidis Rhizoma and Euodiae Fructus met the quality requirements of the Chinese Pharmacopoeia (2020 Edition). Omeprazole (positive drugs) was purchased from Astrazeneca

Analysis of chemical constituents of ZJP

Firstly, the active chemical composition existed in the aqueous extract of ZJP was comprehensively identified. Positive ES mode of the total ion chromatograms is listed in Fig. 1. Six chemical compounds, including magnoflorine, berberine, coptisine, palmatine, evodiamine and rutaecarpine were totally identified. In addition, the corresponding chemical constituents were also quantitatively analyzed. The results are shown in Table 3.

Effects of ZJP on macroscopic pathological

After the rats were infected with H. pylori for 8 weeks,

Discussion

ZJP is a commonly used traditional Chinese prescription that was first written by Zhu Zhenheng. ZJP has a wide range of pharmacological actions in the digestive system, including bacteriostic, anti-inflammatory and analgesic effects. The protection offered by ZJP against ethanol-induced mucosal lesions is indicative of its outstanding gastroprotective effect, which may be related with the regulation of the NF-кB signaling pathway (Wang et al., 2015a). An increasing number of studies have shown

Conclusion

Taken together, our results confirmed that ZJP exerts a therapeutic effect in the H. pylori-induced CAG model. We also found that histone demethylase plays a vital role in the CAG model. Importantly, ZJP prevented gastric mucosal injury by inhibiting H. pylori-induced inflammation via the JMJD2B/COX-2/VEGF axis and HMGB1/NF-kB signaling pathway. The results of this study suggest a potential role for ZJP in the treatment of CAG, which needs to be further investigated.

Author contributions

JW ([email protected]) conceived the study and wrote the manuscript. SW ([email protected]) collected and prepared the samples. XM ([email protected]) checked the data. YZ ([email protected]) designed the study and amended the paper. All data were generated in-house, and no paper mill was used. All authors agree to be accountable for all aspects of work ensuring integrity and accuracy.

Data availability of statement

The datasets used and/or analyzed of this study are from corresponding author upon reasonable request.

Ethics approval and consent to participate

The animal study was reviewed and approved by the Chinese PLA General Hospital. No human studies are presented in this manuscript. No potentially identifiable human images or data is presented in this study.

Declaration of competing interest

We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome.

Acknowledgements and funding

This study was supported by the National Key R&D Program of China, China [No. 2018YFC1704500], Science Foundation of Sichuan Education Department, China [No.18ZA0186], and Xihua University Talent Introduction Project, China (Z211060).

References (53)

  • S. Xu et al.

    Pharmacokinetic comparisons of rutaecarpine and evodiamine after oral administration of Wu-Chu-Yu extracts with different purities to rats

    J. Ethnopharmacol.

    (2012)
  • T. Yang et al.

    Mechanism of berberine in treating Helicobacter pylori induced chronic atrophic gastritis through IRF8-IFN-γ signaling axis suppressing

    Life Sci.

    (2020)
  • Z. Zhao et al.

    Effect and mechanism of evodiamine against ethanol-induced gastric ulcer in mice by suppressing Rho/NF-кB pathway

    Int. Immunopharm.

    (2015)
  • U. Andersson et al.

    High mobility group 1 protein (HMG-1) stimulates proinflammatory cytokine synthesis in human monocytes

    J. Exp. Med.

    (2000)
  • S. Backert et al.

    Pathogenesis of Helicobacter pylori infection

    Helicobacter

    (2011)
  • M. Chen et al.

    A testing strategy to predict risk for drug-induced liver injury in humans using high-content screen assays and the 'rule-of-two' model

    Arch. Toxicol.

    (2014)
  • H.W. Chung et al.

    Serum high mobility group box-1 (HMGB1) is closely associated with the clinical and pathologic features of gastric cancer

    J. Transl. Med.

    (2009)
  • P. Correa et al.

    The gastric precancerous cascade

    J. Dig. Dis.

    (2012)
  • K.D. Crew et al.

    Epidemiology of gastric cancer. World

    J. Gastroenterol.

    (2006)
  • L. Guo et al.

    IRAK1 mediates TLR4-induced ABCA1 downregulation and lipid accumulation in VSMCs

    Cell Death Dis.

    (2015)
  • W. Guo et al.

    Integrating network pharmacology and pharmacological evaluation for deciphering the action mechanism of herbal formula Zuojin pill in suppressing hepatocellular carcinoma

    Front. Pharmacol.

    (2019)
  • F. Han et al.

    JMJD2B is required for Helicobacter pylori-induced gastric carcinogenesis via regulating COX-2 expression

    Oncotarget

    (2016)
  • D. Liu et al.

    Correlations among Helicobacter pylori infection and the expression of cyclooxygenase-2 and vascular endothelial growth factor in gastric mucosa with intestinal metaplasia or dysplasia

    J. Gastroenterol. Hepatol.

    (2010)
  • N. Liu et al.

    Helicobacter pylori promotes angiogenesis depending on Wnt/beta-catenin-mediated vascular endothelial growth factor via the cyclooxygenase-2 pathway in gastric cancer

    BMC Cancer

    (2016)
  • M.E. Morales et al.

    Lesiones gástricas en pacientes infectados con Helicobacter pylori: expresión de RAGE (receptor de productos de glicosilización avanzada) y otros inmunomarcadores [Expression of RAGE in Helicobacter pylori infested gastric biopsies]

    Rev. Med. Chile

    (2013)
  • S. Nomura et al.

    The correlation between platelet activation markers and HMGB1 in patients with disseminated intravascular coagulation and hematologic malignancy

    Platelets

    (2011)
  • Cited by (14)

    View all citing articles on Scopus
    View full text