IDDF2024-ABS-0230 CCL2-CCR2 axis inhibition reduces liver fibrosis by restoring the immune cell landscape

<h3>Background</h3> C-C motif chemokine ligand 2 (CCL2), the most upregulated chemokine in injured LSECs, leads to hepatic C-C motif chemokine receptor 2 (CCR2)⁺ monocyte-derived macrophage accumulation. Therefore, we performed pharmacological CCL2-CCR2 inhibition and examined how CCL2-CCR2 inhibition restored hepatic immune landscapes related to fibrogenesis. <h3>Methods</h3> Liver injury and liver fibrosis were induced by carbon tetrachloride (CCl₄) in wild-type mice, LSEC-specific <i>Ccl2</i> deficiency mice, and <i>Ccr2</i> knockout mice. Murine intrahepatic leukocytes were isolated from olive oil/CCl₄-treated wild-type mice for mass cytometry by time-of-flight (CyTOF) analysis. Liver and primary mouse LSEC gene expression was analyzed using a PCR array and RNA sequencing. Cenicriviroc (CVC) was administrated to fibrotic mice in prevention and treatment models. Single-cell RNA sequencing was performed on the livers of olive oil/CCl₄/CVC-treated mice. <h3>Results</h3> CCL2 was the most upregulated chemokine in injured LSECs compared to control LSECs. The CCL2 secreted by injured LSECs leads to CCR2⁺ monocyte-derived macrophage accumulation in CCl₄-induced murine livers as determined by CyTOF. However, CCl₄-mediated liver fibrosis and macrophage accumulation were reduced in LSEC-specific <i>Ccl2</i> deficiency and CCR2 knockout mice. Thus, the CCL2-CCR2 angiocrine signaling axis is related to liver fibrogenesis, and the CVC was applied to block the CCL2-CCR2 angiocrine signaling axis. CVC was demonstrated to alleviate liver fibrosis by restoring the FSCN1⁺ macrophage and HERC6⁺ neutrophil landscape. Pathway analysis indicated that the STAT1, NFκΒ, and ERK signaling pathways might be involved in the anti-fibrotic effects of CVC. Consistently, both CVC treatment and <i>Ccr2</i> knockout decreased the levels of phosphorylated STAT1, NFkB, and ERK in the fibrotic liver.<i> In vitro</i>, CVC could transcriptionally suppress crucial profibrotic genes (<i>Xaf1</i>, <i>Slfn4</i>, <i>Slfn8, Ifi213</i>, and <i>Il1</i>b) in macrophages by inactivating the STAT1/NFkB/ERK signaling pathways. <h3>Conclusions</h3> In summary, activation of the angiocrine signal CCL2 in injured LSECs recruits CCR2⁺ monocyte-derived macrophages to the liver, leading to liver fibrosis. CVC restores the hepatic immune cell landscape by inhibiting profibrotic gene transcription <i>via</i> the CCR2-STAT1/NFkB/ERK signaling pathways.