Abstract: Objecitve To investigate the role of macrophage migration inhibitory factor (MIF) in activation of primary hepatic stellate cells(HSC), and to explore the blocking effect of specific antisense oligonucleotides (ASONs) targeting MIF on the activation of primary HSCs. Methods Rat primary HSC were isolated from SD rats by in situ perfusion of collagenase and pronase and single-step Nycodenz density gradient centrifugation, and then cultured and activated. The expression of α-smooth muscle actin (α-SMA), the marker of HSC activation, was assessed by RT-PCR and immunocytochemistry (ICC). The expression of MIF was detected by RT-PCR, enzyme-linked immuno sorbent assay (ELISA), immunofluorescence (IF) or ICC. Completely activated primary HSC were transfected with MIF-specific antisense oligonucleotide (MIF-ASONs), mis-sense oligonucleotides of MIF, or blank liposome (negative-control). Results Quiescent HSC expressed very low level of MIF, while MIF gene and protein expression increased significantly during the process of HSC activation. Positive correlation was found between the expression of α-SMA and MIF (r=0.944, P<0.01). Compared with the control, activated primary HSC transfected with MIF-ASONs showed lower MIF and α-SMA expressions at gene and protein profiles (P<0.05). Conclusion The expression of MIF was up-regulated in the process of HSC activation. Activation in culture of primarily isolated rat HSC could be inhibited by MIF-ASONs, suggesting at least in part that, MIF might be a novel and important factor involved in HSC activation and hepatic fibrosis.