Background Bone marrow-derived macrophages (BMDMs) regulate hepatic progenitor cells (HPCs) differentiation, potentially via the Wnt signaling pathway. While M1-polarized BMDMs (M1-BMDMs) exert anti-fibrotic effects in the liver, Wnt5a is implicated in fibrosis progression. The specific influence of Wnt5a levels within M1-BMDMs on HPCs fate and cirrhosis development remains unclear. This study aimed to elucidate the relationship between M1-BMDM-derived Wnt5a and HPCs differentiation during cirrhosis progression. Methods First, Wnt5a protein expression was assessed in liver biopsy tissues from patients with hepatitis B-associated liver fibrosis. Second, cirrhosis was induced in rats using CCl4/2-AAF. In week 9, rats received intravenous injections of M1-BMDMs with Wnt5a knockdown (M1-BMDM Wnt5a−KD) or overexpression (M1-BMDM Wnt5a−OE); peripheral BMDMs recruitment was blocked using a CCR2 inhibitor. Fibrosis progression, ductular reaction (DR), and HPC differentiation were evaluated. In vitro, WB-F344 cells subjected to frizzled 2 (Fzd2) knockdown (WB-F344 Fzd2−KD) or overexpression (WB-F344 Fzd2−OE) were cultured with conditioned medium from M1-BMDM Wnt5a−KD (CM Wnt5a−KD) or M1-BMDM Wnt5a−OE (CM Wnt5a−OE). Results In patients with hepatitis B-related fibrosis, hepatic Wnt5a expression increased progressively with METAVIR fibrosis grade. In the rat cirrhosis model, M1-BMDMs Wnt5a−KD attenuated fibrosis, whereas M1-BMDMs Wnt5a−OE exacerbated it. Mechanistically, in vivo injection of M1-BMDMs Wnt5a−KD significantly inhibited HPCs differentiation into biliary epithelial cells (BECs), while M1-BMDM Wnt5a−OE promoted this differentiation. In vitro, CM Wnt5a−KD inhibited the differentiation of WB-F344 cells into BECs; this inhibition was potentiated by Fzd2 knockdown in WB-F344 cells but abrogated by Fzd2 overexpression. Conversely, under CM Wnt5a−OE conditions, WB-F344 Fzd2−OE cells exhibited increased cholangiocytic differentiation, an effect largely negated by Fzd2 knockdown. Conclusions M1-BMDMs Wnt5a−KD demonstrated superior therapeutic efficacy against cirrhosis compared to unmodified M1-BMDMs. Wnt5a/Fzd2 signaling mediated the crosstalk between M1-BMDMs Wnt5a−KD and HPCs, revealing a novel therapeutic target for cirrhosis treatment.

M1-BMDMs with Wnt5a deletion attenuate liver fibrosis by suppression of Wnt5a/Frizzled 2 axis in hepatic progenitors

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2025年8月30日

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Fei-fei Xing,
Dan-yang Wang,
Yan-nan Xu,
Xin-rui Zheng,
Shi-hao Zhang,
Meng-yao Zong,
Jun-yi Zhan,
De-xin Wang,
Wei Liu,
Jia-mei Chen,
Gao-feng Chen,
Ping Liu,
Cheng-hai Liu,
Yong-ping Mu

摘要

Background

Bone marrow-derived macrophages (BMDMs) regulate hepatic progenitor cells (HPCs) differentiation, potentially via the Wnt signaling pathway. While M1-polarized BMDMs (M1-BMDMs) exert anti-fibrotic effects in the liver, Wnt5a is implicated in fibrosis progression. The specific influence of Wnt5a levels within M1-BMDMs on HPCs fate and cirrhosis development remains unclear. This study aimed to elucidate the relationship between M1-BMDM-derived Wnt5a and HPCs differentiation during cirrhosis progression.

Methods

First, Wnt5a protein expression was assessed in liver biopsy tissues from patients with hepatitis B-associated liver fibrosis. Second, cirrhosis was induced in rats using CCl₄/2-AAF. In week 9, rats received intravenous injections of M1-BMDMs with Wnt5a knockdown (M1-BMDM ^Wnt5a−KD) or overexpression (M1-BMDM ^Wnt5a−OE); peripheral BMDMs recruitment was blocked using a CCR2 inhibitor. Fibrosis progression, ductular reaction (DR), and HPC differentiation were evaluated. In vitro, WB-F344 cells subjected to frizzled 2 (Fzd2) knockdown (WB-F344 ^Fzd2−KD) or overexpression (WB-F344 ^Fzd2−OE) were cultured with conditioned medium from M1-BMDM ^Wnt5a−KD (CM ^Wnt5a−KD) or M1-BMDM ^Wnt5a−OE (CM ^Wnt5a−OE).

Results

In patients with hepatitis B-related fibrosis, hepatic Wnt5a expression increased progressively with METAVIR fibrosis grade. In the rat cirrhosis model, M1-BMDMs ^Wnt5a−KD attenuated fibrosis, whereas M1-BMDMs ^Wnt5a−OE exacerbated it. Mechanistically, in vivo injection of M1-BMDMs ^Wnt5a−KD significantly inhibited HPCs differentiation into biliary epithelial cells (BECs), while M1-BMDM ^Wnt5a−OE promoted this differentiation. In vitro, CM ^Wnt5a−KD inhibited the differentiation of WB-F344 cells into BECs; this inhibition was potentiated by Fzd2 knockdown in WB-F344 cells but abrogated by Fzd2 overexpression. Conversely, under CM ^Wnt5a−OE conditions, WB-F344 ^Fzd2−OE cells exhibited increased cholangiocytic differentiation, an effect largely negated by Fzd2 knockdown.

Conclusions

M1-BMDMs ^Wnt5a−KD demonstrated superior therapeutic efficacy against cirrhosis compared to unmodified M1-BMDMs. Wnt5a/Fzd2 signaling mediated the crosstalk between M1-BMDMs ^Wnt5a−KD and HPCs, revealing a novel therapeutic target for cirrhosis treatment.