Exploring the Mechanism of Anti-Colorectal Cancer Action of Fushao Diqin Decoction Based on the Nrf2/SLC7A11/GPX4 Signaling Pathway

  OBJECTIVE  To explore the mechanism of action of Fushao Diqin Decoction in the treatment of colorectal cancer.

  METHODS  In vitro cell experiments were conducted using Fushao Diqin Decoction to treat colorectal cancer CT-26 cells, and the cell proliferation and migration abilities were detected. Flow cytometry was used to detect the levels of reactive oxygen species (ROS) in colorectal cancer CT-26 cells, as well as the levels of iron ions (Fe²⁺), malondialdehyde (MDA), and the activity of superoxide dismutase (SOD). PCR Array and Western blot methods were used to analyze and verify the differential gene expression of ferroptosis. Balb/c mice were randomly divided into a blank control group, a model group, an oxaliplatin group (1.5 mg·kg^-1·d^-1), a low-dose group of Fushao Diqin Decoction (4.49 g·kg^-1·d^-1), a medium dose group of Fushao Diqin Decoction (8.97 g·kg^-1·d^-1), and a high-dose group of Fushao Diqin Decoction (17.94 g·kg^-1·d^-1) for in vivo animal experiments. The effects of Fushao Diqin Decoction on Fe²⁺, ROS, MDA levels, SOD activity, and Nrf2, Keap1, SLC7A11 and GPX4 expression levels in mouse tumor tissues were tested.

  RESULTS  In vitro cell experiments showed that compared with the blank control group, Fushao Diqin Decoction significantly inhibited the proliferation and migration of colorectal cancer CT-26 cells in a dose-dependent manner. Fushao Diqin Decoction could increase the Fe²⁺ content (P < 0.05) and ROS level (P < 0.01) in colorectal cancer CT-26 cells, increase the MDA level in CT-26 cells of colorectal cancer (P < 0.01) and significantly reduce SOD activity (P < 0.01). Iron death PCR array analysis found that compared with the blank control group, after intervention with Fushao Diqin Decoction, the expression of genes GPX4 and SLC7A11 was significantly downregulated, while the expression of GSTA1, HMOX1, Ca9, Chac1, Keap1, Sqstm1, NOX1, FTH1, Tfr1, SAT2, Pparg, and Hamp was significantly upregulated. Western blot analysis revealed that after intervention with Fushao Diqin Decoction, the expression of Keap1 protein was upregulated (P < 0.01), while the expression of Nrf2, SLC7A11, and GPX4 proteins was downregulated (P < 0.01) in colorectal cancer CT-26 cells. The results of in vivo animal experiments showed that Fushao Diqin Decoction significantly inhibited the growth of subcutaneous transplanted tumors in mice (P < 0.05), increased the degree of tumor tissue necrosis, and levels of Fe²⁺, ROS, and MDA (P < 0.05, P < 0.01), decreased SOD activity (P < 0.01) and upregulated Keap1 protein expression (P < 0.01), while downregulated Nrf2, SLC7A11, and GPX4 protein expression (P < 0.01).

  CONCLUSION  Fushao Diqin Decoction has an anti-colorectal cancer effect and may promote ferroptosis in colorectal cancer cells by inhibiting the Nrf2/SLC7A11/GPX4 signaling pathway to exert its anti-colorectal cancer effect.