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 (CCl(4))-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 beta (TGF-beta) 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 CCl(4)-treated animals, but not naïve bone marrow monocytes or control lymphocytes, directly activate HSCs in a TGF-beta-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.