Effect of Fuzheng Jiedu Prescription Combined with 5-Fu Against Postoperative Recurrence and Metastasis of Gastric Cancer in Mice： An Exploration Based on Endoplasmic Reticulum Stress / 中国实验方剂学杂志

ABSTRACT

Objective:To investigate the inhibitory effect of Fuzheng Jiedu prescription（FZJDP） combined with 5-fluorouracil （5-Fu） against postoperative recurrence and metastasis in gastric cancer-bearing mice and explore the possible mechanism based on changes in CD4⁺ T cells， CD8⁺ T cells， and regulatory T （Treg） cells of tumor microenvironment， endoplasmic reticulum stress （ERS） and phosphatidylinositol-3-kinase （PI3K）/protein kinase B（Akt）/mammalian target of rapamycin（mTOR） pathway. Method:Forty 615 mice were randomly divided into the model group，FZJDP （25 g·kg^-1） group，5-Fu（25 mg·kg^-1）group，and combined （25 g·kg^-1 FZJDP + 25 mg·kg^-1 5-Fu）group， with 10 mice in each group. Mouse forestomach carcinoma （MFC） cells were implanted into the inner side of footpad of the left hind paw and the transplanted tumor was then surgically excised to establish a postoperative recurrence model. Hematoxylin-eosin（HE） staining was conducted to observe the pathological changes in mice with gastric cancer recurrence and lung metastasis. The CD4⁺/CD8⁺ T cell ratio in recurrent tumor and the percentages of Treg cells ［CD4⁺，CD25⁺， and forkhead box protein P3 （FOXP3）⁺ cells］ in spleen were detected by flow cytometry. The contents of ERS-related proteins ［78-kDa glucose-regulated protein（GRP78），inositol-requiring enzyme 1 alpha （IRE1<italic>α</italic>），activating transcription factor 6（ATF6），and protein kinase R-like endoplasmic reticulum kinase（PERK）］ and the expression of related proteins in the PI3K/Akt/mTOR signaling pathway were determined by Western blot and immunohistochemistry （IHC）. Result:Compared with the model group， the combined group significantly increased the recurrent inhibition rate （<italic>P</italic><0.05）. The recurrent tumor weight was significantly decreased in each treatment group （<italic>P</italic><0.05）. The number of lung metastases and metastasis rate declined in each treatment group， and the lowest values were observed in the combined group， without any statistical significance. The CD4⁺/CD8⁺ T cell ratios in the FZJDP group and combined group were significantly elevated （<italic>P</italic><0.05）， while the percentages of Treg cells were reduced （<italic>P</italic><0.05）. However， 5-Fu resulted in a significant increase in Treg cell percentage （<italic>P</italic><0.05）. IHC results showed that the protein expression levels of ATF6 （<italic>P</italic><0.05，<italic>P</italic><0.01）， IRE1<italic>α</italic>（<italic>P</italic><0.05），and Akt （<italic>P</italic><0.01） in each treatment group were significantly down-regulated as compared with those in the model group. As revealed by Western blot， the GRP78 expression level in the 5-Fu group was lower than that in the model group （<italic>P</italic><0.05）， and the expression levels of PI3K， phosphorylated Akt （p-Akt）， and mTOR were significantly decreased in the 5-Fu group and the combined group （<italic>P</italic><0.05）. Conclusion:FZJDP combined with 5-Fu reduces postoperative recurrence and metastasis in tumor-bearing mice possibly by inhibiting PI3K/Akt/mTOR signaling pathway， diminishing ERS，and improving tumor immune microenvironment.