Hepatic macrophages are central in the pathogenesis of chronic liver injury and have been proposed as potential targets in combatting fibrosis. Recent experimental studies in animal models revealed that hepatic macrophages are a remarkably heterogeneous population of immune cells that fulfill diverse functions in homeostasis, disease progression, and regression from injury. These range from clearance of pathogens or cellular debris and maintenance of immunological tolerance in steady state conditions; central roles in initiating and perpetuating inflammation in response to injury; promoting liver fibrosis via activating hepatic stellate cells in chronic liver damage; and, finally, resolution of inflammation and fibrosis by degradation of extracellular matrix and release of anti-inflammatory cytokines. Cellular heterogeneity in the liver is partly explained by the origin of macrophages. Hepatic macrophages can either arise from circulating monocytes, which are recruited to the injured liver via chemokine signals, or from self-renewing embryo-derived local macrophages, termed Kupffer cells. Kupffer cells appear essential for sensing tissue injury and initiating inflammatory responses, while infiltrating Ly-6C(+) monocyte-derived macrophages are linked to chronic inflammation and fibrogenesis. In addition, proliferation of local or recruited macrophages may possibly further contribute to their accumulation in injured liver. During fibrosis regression, monocyte-derived cells differentiate into Ly-6C (Ly6C, Gr1) low expressing 'restorative' macrophages and promote resolution from injury. Understanding the mechanisms that regulate hepatic macrophage heterogeneity, either by monocyte subset recruitment, by promoting restorative macrophage polarization or by impacting distinctive macrophage effector functions, may help to develop novel macrophage subset-targeted therapies for liver injury and fibrosis.

In addition to liver-resident Kupffer cells, infiltrating immune cells have recently been linked to the development of liver fibrosis. Blood monocytes are circulating precursors of tissue macrophages and can be divided into two functionally distinct subpopulations in mice: Gr1^hi (Ly6C^hi) and Gr1^lo (Ly6C^lo) monocytes. The role of these monocyte subsets in hepatic fibrosis and the mechanisms of their differential recruitment into the injured liver are unknown. We therefore characterized subpopulations of infiltrating monocytes in acute and chronic carbon tetrachloride (CCl₄)-induced liver injury in mice using flow cytometry and immunohistochemistry. Inflammatory Gr1^hi but not Gr1^lo monocytes are massively recruited into the liver upon toxic injury constituting an up to 10-fold increase in CD11b⁺F4/80⁺ intrahepatic macrophages. Comparing wild-type with C-C chemokine receptor (CCR2)-deficient and CCR2/CCR6–deficient mice revealed that CCR2 critically controls intrahepatic Gr1^hi monocyte accumulation by mediating their egress from bone marrow. During chronic liver damage, intrahepatic CD11b⁺F4/80⁺Gr1⁺ monocyte-derived cells differentiate preferentially into inducible nitric oxide synthase–producing macrophages exerting proinflammatory and profibrogenic actions, such as promoting hepatic stellate cell (HSC) activation, T helper 1–T cell differentiation and transforming growth factor β (TGF-β) release. Impaired monocyte subset recruitment in Ccr2 ^−/− and Ccr2 ^−/− Ccr6 ^−/− mice results in reduced HSC activation and diminished liver fibrosis. Moreover, adoptively transferred Gr1^hi monocytes traffic into the injured liver and promote fibrosis progression in wild-type and Ccr2 ^−/− Ccr6 ^−/− mice, which are otherwise protected from hepatic fibrosis. Intrahepatic CD11b⁺F4/80⁺Gr1⁺ monocyte-derived macrophages purified from CCl₄-treated animals, but not naïve bone marrow monocytes or control lymphocytes, directly activate HSCs in a TGF-β–dependent manner in vitro. Conclusion: Inflammatory Gr1⁺ monocytes, recruited into the injured liver via CCR2-dependent bone marrow egress, promote the progression of liver fibrosis. Thus, they may represent an interesting novel target for antifibrotic strategies. (Hepatology 2009;50:261–274.)