Retinoic acid and IFNγ induce parallel Arg1 and Nos2 gene expression in primary mouse Kupffer cells.

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Abstract Acute inflammation is associated with classically activated macrophages, while chronic inflammation is associated with fibrosis and alternatively activated (M2) macrophages. Liver fibrosis is a common complication of chronic liver inflammation, yet the molecular signals that promote M2 Kupffer cells, resident liver macrophages, are poorly defined. In the liver, activated hepatic stellate cells (HSC) are associated with fibrosis and release retinoic acid (RA). Here, we test the hypothesis that RA polarizes Kupffer cells to an M2 state. We contrast this to activation by IFNγ or lipopolysaccharide (LPS). Kupffer cells were isolated from C57BL/6J mice (N = 2–5) by sort-purification; plated at 30,000 per well for 24 hours; and then stimulated with 100 ng/mL LPS, 100 ng/mL IFNγ, 100 μM RA, or IFNγ + RA for 24 hours. Isolated RNA was analyzed by microfluidic qRT-PCR using assays for genes characteristic of macrophage activation. LPS induced Socs3, Ptgs2, Nos2, Tnf, Icam1, Il6, Il1a, Il10, and Marco. IFNγ induced Socs3, Ptgs2, Nos2, Tnf, Icam1, Ciita, Ccl12, and Klf4; and suppressed Mrc1. RA only induced expression of Arginase-1 (Arg1). Unexpectedly, co-treatment with RA and IFNγ induced additive expression. In summary, treatment of primary Kupffer cells with RA induces Arg1; co-treatment with IFNγ and RA simultaneously induces Arg1 and Nos2; and LPS, but not IFNγ, induces Il6 gene expression. These results support the hypothesis that HSC activation may induce aspects of alternative activation in Kupffer cells and demonstrate that classical and alternative activation fates of macrophages are not always mutually exclusive.