The Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway is widely involved in cell migration, apoptosis and inflammation. However, its exact mechanisms in severe acute pancreatitis (SAP) remain unclear. The aim of this study was to explore the activity of the JAK/STAT signaling pathway in pancreatic injury, investigate the functional mechanisms of SAP in vitro, and thus elucidate the underlying therapeutic effects for SAP in vivo. The activation of the JAK-1/STAT-1 signaling pathway and the expessions of TNF-α, IL-1β and IL-6 proteins were investigated in AR42J cells induced with cerulein and treated with either PBS, RPM, or AG490. One group of cells was left untreated as a control group. Subsequently the activity of NF-κB was evaluated. Rats were given RPM or AG490 just before the induction of SAP, the severity of which was assessed at 24 h. The findings revealed that the up-regulated expressions of JAK-1/STAT-1, STAT-3 protein were closely correlated with the transcription of TNF-α, IL-1β, and IL-6 in cerulein-stimulated cells. Administration of RPM or AG490 decreased the activity of NF-κB and inhibited the release of TNF-α, IL-1β, and IL-6. The reflective markers of severity of SAP were also decreased by RPM or AG490 treatment compared to SAP rats. This study indicates that the JAK-1/STAT-1 signaling pathway activity is an early event in pancreatic inflammatory injury. Therefore, early treatment with its inhibitors might be beneficial for attenuation of pancreatic injury in SAP.