Fibroblast activation protein (FAP) is expressed on activated fibroblast. Its role in fibrosis and desmoplasia is controversial, and data on pharmacological FAP inhibition are lacking. We aimed to better define the role of FAP in liver fibrosis in vivo and in vitro.FAP expression was analyzed in mice and patients with fibrotic liver diseases of various etiologies. Fibrotic mice received a specific FAP inhibitor (FAPi) at 2 doses orally for 2 weeks during parenchymal fibrosis progression (6 weeks of CCl4) and regression (2 weeks off CCl4), and with biliary fibrosis (Mdr2-/-). Recombinant FAP was added to (co-)cultures of hepatic stellate cells (HSC), fibroblasts and macrophages. Fibrosis and inflammation related parameters were determined biochemically, by quantitative immunohistochemistry, PCR and transcriptomics.FAP+ fibroblasts/hepatic stellate cells (HSC) were α-SMA negative and located at interfaces of fibrotic septa next to macrophages in murine and human livers. In parenchymal fibrosis FAPi reduced collagen area, liver collagen content, α-SMA+ myofibroblasts, M2-type macrophages, serum ALT and AST, key fibrogenesis related transcripts, and increased hepatocyte proliferation 10-fold. During regression FAP was suppressed and FAPi ineffective. FAPi less potently inhibited biliary fibrosis. In vitro FAP siRNA reduced HSC α-SMA expression and collagen production, and FAPi suppressed their activation and proliferation. Compared to untreated macrophages, FAPi regulated macrophage profibrogenic activation and transcriptome, and their conditioned medium attenuated HSC activation, which was increased with addition of recombinant FAP.Pharmacological FAP inhibition attenuates inflammation-predominant liver fibrosis. FAP is expressed on subsets of activated fibroblasts/HSC and promotes both macrophage and HSC profibrogenic activity in liver fibrosis.Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.