Novel regulators of hepatic macrophages in liver fibrosis

Abstract

Liver fibrosis is a common pathological process resulting from liver damage and subsequent inflammatory responses in various chronic liver diseases, leading to persistent structural and functional abnormalities in the liver. It can further progress to liver cirrhosis and hepatocellular carcinoma. Currently, no effective treatments are available for liver fibrosis, except for liver transplantation. Hepatic macrophages play essential roles in both the development and regression of liver fibrosis. Understanding the mechanisms by which hepatic macrophages regulate liver fibrosis could identify new therapeutic targets. In this review, we aim to summarize recent discoveries regarding the specific molecular mechanisms underlying the progression of liver fibrosis over the past 5 years, with a special focus on monocyte recruitment and macrophage polarization or differentiation, as well as their roles in disease progression.

Keywords: liver fibrosis; macrophage; macrophage polarization; molecular mechanisms; monocyte recruitment.

Figure 1

Hepatic macrophage subsets in mice and humans. KCs and MoMFs are the main components of hepatic macrophages. KCs mainly originate from the yolk sac or bone marrow. In humans, KCs exhibit CD45⁺CD11b⁺CD68⁺CD14⁺CD206⁺CD11c⁻CCR2⁻CD32⁺MARCO⁺TIM4⁺ phenotype. In mice, KCs can be marked as CD45⁺CD11b⁺F4/80⁺CD68⁺CD206⁺CD11c⁻MHC II⁺CCR2⁻CLEC4F⁺ TIM4⁺CLEC2⁺CRIg⁺. KCs are further distinguished into KC1 (CD206^loESAM⁻) and KC2 (CD206^hiESAM⁺) subgroups in mice. Infiltrating monocytes are mainly derived from bone marrow, spleen, and peritoneal macrophages. The monocytes further differentiate into MoMFs. In mice, MoMFs are categorized into two subsets: CX3CR1^intCCR2⁺CD62L⁺CD43^loLy6C^hi and CX3CR1^hiCCR2⁻CD62L⁻CD43^hiLy6C^lo monocytes. In humans, MoMFs are classified into three subtypes: CD14⁺⁺CD16⁻CCR2⁺CD11b⁺human leukocyte antigen DR(HLA-DR)⁺CX3CR1^lo (classical), CD14⁺⁺CD16⁺CCR2⁺CD11b⁺HLA-DR^hiTREM2^+/− (intermediate) and CD14⁺CD16⁺⁺ CCR2⁻CD11b⁺HLA-DR⁺CX3CR1^hiCD163⁺ (nonclassical) monocytes. Classically, macrophages polarize into M1 and M2 subtypes in response to different stimulations. In humans, M1 macrophages exhibit a CD80⁺CD86⁺HLA-DR^hiCD64⁺CD40⁺CCR7⁺ phenotype, while M2 macrophages represent a CD163⁺CD206⁺CD209⁺CD200R⁺CCR2⁻ phenotype. In mice, M1 subtypes can be identified as CD80⁺CD86⁺CD16/32⁺MHC II⁺iNOS⁺, while M2 can be recognized as CD206⁺CD163⁺CD209⁺FIZZ1⁺Ym1⁺.

Figure 2

Overview of the roles of hepatic macrophages in liver fibrosis. KC replenishment occurs predominantly by the self-renewal of resident stem cells to maintain liver homeostasis under the physiological conditions. Upon liver injury, KCs are activated by DAMPs, HMGB-1, mtDNA released from damaged hepatocytes, and highly secrete CCL2, TNF-α, IL-1β, and so on, which subsequently contribute to hepatocyte injury and Ly6C^hi monocyte infiltration to the damaged site, where they develop into Ly6C^hi macrophages in mice. The profibrogenic KCs and Ly6C^hi MoMFs promote HSC activation and exert profibrotic effects by expressing TGF-β, PDGF, and CTGF, leading to the excessive deposition of ECM and scar formation. During the regression stage, the KCs and Ly6C^hi monocytes undergo phenotype switch and degrade ECM by producing MMPs to exert antifibrotic effects upon phagocytosis or other resolution stimuli.

Figure 3

Key signaling pathways involved in hepatic macrophage polarization during liver fibrosis. Hepatic macrophage activation can be regulated by TLR4–MyD88–NF-κB, JAK–STAT, cGAS–STING pathways, etc. (1) TLR4–MyD88–NF-κB, RIP3 interferes with macrophage accumulation by regulating the ROCK1–TLR4–NF–κB signaling pathway. MANF disturbs S100A8/A9 heterodimer-mediated TLR4–NF-κB signaling pathway activation to inhibit M1 macrophage polarization. (2) JAK–STAT pathway, CXCL10 interacting with the receptor CXCR3 negatively regulates macrophages activation depending on SHP-1-mediated STAT6 activation. IL-4Rα regulates alternative macrophage activation in a STAT6-dependent manner. USP25 stabilizes STAT6 by reducing the K48-specific ubiquitination of STAT6. SOCS1 inhibits M1 macrophage polarization by blocking the JAK1/STAT1 pathway. FGF12 induces the Ly6C^lo phenotype switch to Ly6C^hi by inhibiting the JAK/STAT signaling pathway. IL-22 promotes M2 macrophage polarization by activating the STAT3 pathway. (3) cGAS–STING pathway, cGAS recognizes cytosolic mtDNA to promote STING phosphorylation and macrophage activation. XBP1 deficiency decreases mtDNA release and STING activation by promoting BNIP3-mediated mitophagy.

Figure 4

ncRNAs regulate the adhesion and activation of hepatic macrophages during liver fibrosis. The ncRNAs, including miRNAs, lncRNAs, and circRNAs, play important roles in the progression of liver fibrosis. (1) miRNAs, miR-155 promotes CCL2 expression to induce monocyte infiltration and influences M2 macrophage polarization by decreasing C/EBPβ expression. MiR-130a-3p, cooperating with miR-142-5p, controls macrophage polarization to the M2 phenotype and Ly6C^lo phenotype by targeting SOCS1 and PPAR-γ, respectively. MiR-21 negatively regulates M2 polarization by targeting PTEN. MiR-4524a-5p/TBP increased M2 macrophage polarization via the TIM3 and PI3K/mTOR pathways. MiR-690 maintains KC functions by downregulating Nadk transcription. Csi-let-7a-5p promotes M1 macrophage activation by blocking SOCS1- and Dectin1-mediated NF-κB signaling pathway. (2) lncRNAs, H19 enhances the M1 polarization of Kupffer cells and promotes the macrophage recruitment via CCL2 and IL-6. Lfar1 negatively regulates M1 macrophage activation through the NF-κB pathway and NLRP3 inflammasome-mediated proptosis. Helf, interacting with PTBP1, increases Pik3r5 mRNA stability and activates the AKT pathway to promote hepatic macrophage polarization to M1 phenotype. XIST/FTX also increases M1 macrophage polarization. SNHG20 induces M2 polarization through activating STAT6. (3) CircRNAs, Circ_1639 contributes to NF-κB pathway-mediated Kupffer cell activation by sponging miR-122 and regulating TNFRSF13C expression. CircDcbld2 promotes macrophage activation by binding miR-144-3p and regulating Et-1 expression.

Figure 5

Cellular crosstalk between hepatic macrophages and surrounding cells in hepatic fibrosis. The crosstalk between hepatic macrophages and other cells in the liver plays an important role during the hepatic fibrosis process. The ATP, IL-10, TGF-β, acetoacetate, HRG, cholesterol crystals within remnant lipid droplets, and DAMPs, including mtDNA, HMGB1, IL-33, etc., derived from damaged or dead hepatocytes, contribute to macrophage polarization. EGFR, CCL2, PSMP, CCL7, CYR61, glutamate, and EVs from hepatocytes are required for monocyte recruitment. Activated HSCs can release CSF1, TIMP-1, CCL2, CCL11, CXCL2, M-CSF, ICAM-1, VCAM-1, E-selectin, etc., which can promote macrophage recruitment. HSC-derived miR-99a-5p, CXCL10, ROS, EVs, and HSC-associated proteins such as FAP and BMP9 exert important roles in hepatic macrophage polarization. LSECs facilitate hepatic recruitment of monocytes by expressing CCL2, ICAM-1, VCAM-1, and VAP-1. DLL4 and TGF-β secreted by LSECs can induce monocytes to acquire and maintain Kupffer cell identity. MPO is associated with monocyte infiltration and hCLS formation. NETs stimulate macrophage pyroptosis. Neutrophils induce alternative activation of macrophages, potentially via miR-223. MSC-derived IL-4, IL-10, and exosomes containing miR-148a can regulate macrophage phenotype. Activated MAIT can switch macrophage phenotypes by regulating apoptotic, survival, and reprogramming-related genes. Splenic macrophage-secreted CCL2 induces the infiltration of circulating monocytes. Spleen-derived Lcn2 contributes to the infiltrated MoMF and KC activation. Hence, interfering with intercellular communication is a potential therapeutic strategy for liver fibrosis.

Figure 6

Metabolic reprogramming in hepatic macrophage activation during liver fibrogenesis. Metabolic reprogramming, such as carbohydrate metabolism, lipid metabolism, is responsible for macrophage activation. (1) Carbohydrate metabolism, M1 macrophage polarization mainly relies on aerobic glycolysis, while M2 macrophage polarization depends more on OXPHOS. GDF15 can preprogram the macrophages to commit to OXPHOS. PKM2 is a central enzyme for glycolysis. FSTL1 can bind and enhance the stability of PKM2. Both FSTL1 and HSPA12A can increase phosphorylation and nuclear translocation of PKM2. In contrast, ANXA5 inhibits phosphorylation and promotes PKM2 tetramer formation. PFKFB3 is another key activator of glycolysis. SGLT2 inhibitors and c-Rel can downregulate PFKFB3 in macrophages. (2) Lipid metabolism, CD36 is a key lipid transporter of hepatic macrophages. FGF21 can abolish HFCD-induced upregulation of CD36. ATF3 can increase RBP4 and Zdhhc4/5-mediated CD36 palmitoylation and block FOXO1 deacetylation to regulate gluconeogenesis. S1P is the most important lipid mediator, which can be catalytically synthesized by SPHK1. SPHK1 promotes macrophage polarization via the ASK1-JNK1/2-p38 signaling pathway and KLF4 SUMOylation. Activation of SPHK1 signaling contributes to macrophage recruitment by preventing miR-19b-3p-mediated inhibition of the CCL2–CCR2 axis. S1PR2/3 blockade downregulates the NLRP3 inflammasome and regulates BMDM activation. (3) Others, E2F2 downregulates the expression of SLC7A5 to mediate amino acid flux, resulting in enhanced glycolysis in a Leu‐mTORC1‐dependent manner. Bile acids activating FXR can repress KC activation and CCL2 release by upregulating IκBα. ATG5-mediated autophagy, Nup85-mediated CMA, and LAP are also associated with macrophage recruitment and polarization.