Abstract
IntroductionMurine Kupffer cells (KCs) comprise CD11bhi and F4/80hi subsets. Tissue-resident macrophages are known to express the tyrosine kinase receptors colony-stimulating factor 1 receptor (Csf1r) and Mer. However, the expression of Csf1r and Mer on KC subsets and the importance of these tyrosine kinases during liver regeneration (LR) are unknown.MethodsKCs from wild-type and Csf1r-GFP mice were characterized by flow cytometry. Partial hepatectomy (PH) was performed in mice treated with clodronate liposomes, a Csf1r small molecule inhibitor or depleting antibody, or a small molecule Mer inhibitor. Sera and livers were analyzed. The function of sorted KC subsets was tested in vitro.ResultsMer was specifically expressed on tissue-resident F4/80hi KCs, 55% of which also expressed Csf1r. Mer+Csf1r+ and Mer+Csf1r- KCs had distinct expression of macrophage markers. Csf1r inhibition in mice reduced F4/80hi KCs by approximately 50%, but did not affect CD11bhi KCs. Clodronate liposomes depleted F4/80hi KCs, but also altered levels of other intrahepatic leukocytes. Csf1r inhibition delayed LR, as demonstrated by a 20% reduction in liver-to-body weight ratios 7 days after PH. At 36h after PH, Csf1r inhibition increased serum ALT and histological liver injury, and decreased liver cell proliferation. A small molecule inhibitor of Mer did not alter the percentage of KCs or their proliferation and just modestly delayed LR. In vitro, Csf1r or Mer inhibition did not decrease KC viability, but did attenuate their cytokine response to stimulation.ConclusionsF4/80hi KCs are Mer+ and can be subdivided based on Csf1r expression. Csf1r or Mer inhibition each reduces KC cytokine production and delays LR.
Highlights
MethodsPartial hepatectomy (PH) was performed in mice treated with clodronate liposomes, a colony-stimulating factor 1 receptor (Csf1r) small molecule inhibitor or depleting antibody, or a small molecule Mer inhibitor
Murine Kupffer cells (KCs) comprise CD11bhi and F4/80hi subsets
Mer was expressed on tissue-resident F4/80hi KCs, 55% of which expressed colony-stimulating factor 1 receptor (Csf1r)
Summary
Partial hepatectomy (PH) was performed in mice treated with clodronate liposomes, a Csf1r small molecule inhibitor or depleting antibody, or a small molecule Mer inhibitor. Rat anti-mouse Csf1r (clone AFS98) monoclonal antibody or rat IgG2a isotype control (clone 2A3), both from BioXCell (West Lebanon, NH), was administered at a loading dose of 500 μg i.p. on day 1 followed by 250 μg on days 3 and 5. UNC2250, a selective Mer inhibitor,[20] was purchased from Selleckchem (Houston, TX) and administered by oral gavage at a dose of 3. Kupffer cell Csf1r and Mer in liver regeneration mg/kg twice daily. Mice were housed and bred in a pathogen-free facility at Memorial Sloan Kettering Cancer Center. The Institutional Animal Care and Use Committee of the Memorial Sloan Kettering Cancer Center approved all procedures. All animals received humane care according to the criteria outlined in the “Guide for the Care and Use of Laboratory Animals” prepared by the National Academy of Sciences and published by the National Institutes of Health
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