Sa1788 Tobacco Toxin NNK (4-[Methylnitrosamino]-1-[3-Pyridyl]-1-Butanone) Mediates Zymogen Activation in Murine and Human Pancreatic Acini

Abstract

Clinical observations indicate that cigarette smoking is an independent and dose-dependent risk factor for acute pancreatitis. Cigarette smoke has many potentially toxic components; one of the most active and injurious is the nicotine metabolite, NNK (4-[methylnitrosamino]1-[3-pyridyl]-1-butanone). Previously we have shown in rats that NNK mediates premature zymogen activation, a pivotal early pancreatitis event. Furthermore, pharmacologic evidence indicates this response occurs through non-neuronal α7 nicotinic acetylcholine receptors (α7nAChR) on the acini. In our current study we used a genetic mouse model to confirm whether NNK mediates zymogen activation through α7nAChR. In addition, we investigated whether NNK could similarly induce zymogen activation in isolated human acinar cells. Isolated acini from C57BL6 (wild type) mice were treated for 30 minutes with 100 nM NNK and zymogen (trypsinogen) activation was measured using a fluorogenic assay. NNK increased trypsinogen activation 3-fold. The presence of non-neuronal nicotinic acetylcholine receptors (α7 nAChR), a target for NNK, was detected in wild type mouse acinar cells using PCR. Treatment of acini isolated from α7 nAChR-/mice, with NNK, did not initiate zymogen activation. To further confirm the in vitro studies, NNK was given to mice (C57BL & α7 nAChR-/-) by IP injection (100 mg/kg/hour over 6 hours) and zymogen activation was assessed. In C57BL6 mice, the in vivo NNK treatment induced zymogen activation, whereas in α7 nAChR-/mice it did not. Finally, the presence of α7 nAChR was also detected by PCR in isolated human acinar cells. Treatment of these cells with 100 nM NNK caused a 3-fold increase in zymogen activation, versus an unstimulated control, similar to that seen in wild type mice. Levels of NNK-mediated zymogen activation in human acini were also comparable to that induced by hyperstimulatory concentrations of the muscarinic agonist carbachol, an agent used to induce pancreatitis responses in human acini. These studies suggest that exposure to the tobacco toxin NNK increases zymogen activation in both murine and humanmodels of acute pancreatitis. Further, activation of anα7 nAChR on the pancreatic acinar cell may have an important role in mediating pancreatitis effects of cigarette smoke.