[Exocarpium Citri Grandis formula granules alleviate fatty liver disease in Zebrafish by maintaining iron homeostasis and suppressing lipid peroxidation and ferroptosis].

Abstract

Objectives: To investigate the therapeutic effect of Exocarpium Citri Grandis formula granules (ECGFG) on fatty liver disease (FLD) in zebrafish and explore the underlying mechanism.

Methods: Nonalcoholic fatty liver disease (NAFLD) and alcoholic fatty liver disease (ALD) models were established in zebrafish larvae at 3 days post fertilization (dpf), in which the treatment efficacy of 16, 32, or 64 μg/mL ECGFG was evaluated by examining zebrafish survival and liver pathologies and using whole-fish oil red O staining and RT-qPCR. The therapeutic mechanism of ECGFG for FLD was investigated using Prussian blue staining, DCFH-DA probe, MDA content detection, RT-qPCR assay and immunohistochemical staining for CAV1.

Results: In zebrafish models of NAFLD and ALD, treatment with ECGFG significantly reduced lipid accumulation and the expression levels of FASN, SREBP1, HMGCRA, TNF-α and IL-6, increased the expressions of Apoa1 and PPARα, and reduced iron deposition and the contents of MDA and ROS in the liver. In zebrafish models of NAFLD, treatment with ECGFG at the 3 doses significantly increased hepatic expressions of Tf, TfR, FPN and SLC7A11, and at the doses of 32 and 64 μg/mL, ECGFG obviously increased hepatic expression of GPX4. ALD fish models showed significantly increased hepatic expressions of Tf, TfR and FPN, which were effectively lowered by treatment with ECGFG at the 3 doses. ECGFG did not obviously affect the expression of SLC7A11, but its high dose (64 μg/mL) caused significant elevation of GPX4 expression. Both zebrafish models of NAFLD and ALD showed obviously increased CAV1 expression level in the liver, which was significantly reduced by treatment with 32 and 64 μg/mL ECGFG.

Conclusions: In zebrafish models of NAFLD and ALD, ECGFG can alleviate lipid accumulation and inflammatory response and lower the expression level of CAV1 to restore iron homeostasis and suppress lipid peroxidation and ferroptosis in the liver.