# Caveolin-1 Deficiency in Macrophages Alleviates Carbon Tetra-Chloride-Induced Acute Liver Injury in Mice

## TL;DR

Deleting the Cav-1 protein in macrophages helps reduce liver damage in mice exposed to a toxic chemical.

## Contribution

This study shows that Cav-1 deficiency in macrophages improves liver recovery by enhancing their protective functions.

## Key findings

- Cav-1 KO BMMs reduced liver necrosis and apoptosis in CCl4-induced injury.
- Cav-1 KO BMMs increased M2 polarization and macrophage infiltration in injured livers.
- Adoptive transfer of Cav-1 KO BMMs showed therapeutic benefits in acute liver injury.

## Abstract

Bone marrow-derived macrophages (BMMs) exhibit dynamic behavior and functional capabilities in response to specific microenvironmental stimuli. Recent investigations have proved that BMMs play crucial roles in promoting necrotic lesion resolution. Despite substantial advancements in understanding their activation and interaction with injured livers, researchers face challenges to develop effective treatments based on manipulating BMMs function. Caveolin-1 (Cav-1) is the major structural protein on the plasma membrane. We previously reported that Cav-1 knockout (KO) mice exhibited less functional damage and necrosis in carbon tetrachloride (CCl4)-induced liver injury. We hypothesize that the activation and recruitment of BMMs are involved in the resolution of necrotic lesions in Cav-1 KO mice. Wild-type (WT) and Cav-1 KO mice were injected with CCl4 (10% v/v) to induce acute liver injury model. Blood samples and hepatic tissues were harvested for serum alanine transaminase (ALT) activity assessment, histopathological examination through hematoxylin–eosin (H&E) staining, and BMMs subpopulation analysis via flow cytometry. Then, primary BMMs were isolated and cultured to investigate the effect of Cav-1 on BMMs polarization, migration, and activation of STAT3 signal pathway. Validation of hepatic macrophage depletion was induced by administrating clodronate liposomes (CLs), and BMMs reconstitution was evaluated by EGFP labelled BMMs. Following this, hepatic macrophages were depleted by CLs, BMMs were isolated from Cav-1 KO, and WT mice were cultured and administrated to evaluate the protective role of Cav-1-deleted BMMs on the resolution of hepatocellular necrosis and apoptosis in acute liver injury. The BMMs ratio significantly increased from 2.12% (1D), 4.38% (1W), and 5.38% (2W) in oil control mice to 7.17%, 14.90%, and 19.30% in CCl4-treated mice (p < 0.01 or p < 0.001). Concurrently, Cav-1 positive BMMs exhibited a marked elevation from 6.41% at 1D to 24.90% by 2W (p = 0.0228). Cav-1 KO exerted protective effects by reducing serum ALT by 26% (p = 0.0265) and necrotic areas by 28% (p = 0.0220) and enhancing BMMs infiltration by 60% (p = 0.0059). In vitro, Cav-1 KO BMMs showed a decrease in CD206 fluorescence intensity (p < 0.001), a time-dependent upregulation of arginase-1 mRNA (p < 0.05 or p < 0.01), a 1.22-fold increase in phosphorylated STAT3 (p = 0.0036), and impaired wound healing from 12 to 24 h (p < 0.001). The macrophage-depleting action in livers by CL injection persists for a minimum of 48 h. Administrated EGFP+ BMMs emerged as the predominant population following CL injection for a duration of 48 h. Following clodronate liposome-mediated hepatic macrophage depletion, the adoptive transfer of Cav-1 KO BMMs demonstrated therapeutic efficacy in CCl4-induced acute liver injury. In CCl4-induced acute liver injury, the adoptive transfer of Cav-1 KO BMMs reduced necrosis by 12.8% (p = 0.0105), apoptosis by 25.2% (p = 0.0127), doubled macrophages infiltration (p = 0.0269), and suppressed CXCL9/10 mRNA expression (p = 0.0044 or p = 0.0385). BMMs play a key role in the resolution of liver necrotic lesions in CCl4-induced acute liver injury. Cav-1 depletion attenuates hepatocellular necrosis and apoptosis by accelerating BMMs recruitment and M2 polarization. Cav-1 in macrophages may represent a potential therapeutic target for acute liver injury.