Dynamic imaging of pancreatic NF-{kappa}B activation in live mice using AAV infusion and bioluminescence [Molecular Bases of Disease]

March 23rd, 2015 by Orabi, A. I., Sah, S., Javed, T. A., Lemon, K. L., Good, M. L., Guo, P., Xiao, X., Prasadan, K., Gittes, G. K., Jin, S., Husain, S. Z.

Nuclear factor κB (NF−κB) is an important signaling molecule that plays a critical role in the development of acute pancreatitis. Current methods for examining NF−κB activation involve infection of an adenoviral NF−κB−luciferase reporter into cell lines or electrophoretic mobility shift assay of lysate. The use of adeno−associated viruses (AAVs) has proven to be an effective method of transfecting whole organs in live animals. We examined whether intra−pancreatic duct infusion of AAV containing an NF−κB−luciferase reporter (AAV−NF−κB−luciferase) can reliably measure pancreatic NF−κB activation. We confirmed the infectivity of the AAV−NF−κB−luciferase reporter in HEK293 cells using a traditional luciferase readout. Mice were infused with AAV−NF−κB−luciferase 5 weeks before induction of pancreatitis (caerulein 50 μg/kg + LPS). Unlike transgenic mice which globally express NF−κB−luciferase, AAV−infused mice showed a 15−fold increase in pancreas−specific NF−κB bioluminescence following 12 hrs of caerulein compared to baseline luminescence (P<0.05). The specificity of the NF−κB−luciferase signal to the pancreas was confirmed by isolating the pancreas and adjacent organs and observing a predominant bioluminescent signal in the pancreas compared with liver, spleen, and stomach. A complementary mouse model of post−ERCP−pancreatitis also induced pancreatic NF−κB signals. Taken together these data provide the first demonstration that NF−κB activation can be examined in a live, dynamic fashion during pancreatic inflammation. We believe this technique offers a valuable tool to study real−time activation of NF−κB in vivo.