Low molecular weight heparin inhibits circulating fibrocytes differentiation by modulating neuronal nitric oxide synthase and TGF-β1/Smad pathway

Abstract

Background/Aims: Circulating fbrocytes (CFs) have been placed at the center of a number of fbrosing conditions. Recently, attention has been drawn to the non-anticoagulant activities of low molecular weight heparin (LH), especially its anti-fbrotic effects. The purpose of this study was to investigate the effects of LH on CFs differentiation and possible underlying mechanisms. Methods/Results: CFs were cultured from human peripheral blood mononuclear cells and identifed by dual-immunofuorescence staining. Incubation with LH inhibited CFs trans-differentiation by upregulating CD34 and downregulating pro-Collagen I and a-SMA in a concentration- and time-dependent manner, all of which were detected by fow cytometry. Similar effects were observed after incubation with L-NAME, an inhibitor of NOS. NO production was measured by Griess methods and markedly decreased in CFs treated with LH. Three NOS isoforms were assessed by western blot and nNOS was the predominant isoform involved in this process. Additionally, LH and L-NAME had similar down-regulating effects on the expression of TGF-β1 and pSmad2/3, which indicated that TGF-β/Smad pathway might be a downstream signaling of nNOS/NO during LH treatment. Conclusion: These results suggested that LH could exhibit anti-fbrotic effects by inhibiting CFs transdifferentiation, in which the involvement of nNOS/NO and TGF-β/Smad pathway were identified. Copyright © 2012 S. Karger AG, Basel.