KGF and Pancreatic Duct Figure 5. Activation of ERK1/2 pathway by KGF in the pancreas of neonatal diabetic rats. Two-day-old STZ diabetic rats were administrated with 3 mg/kg KGF, 20 minutes before the sacrifice. Pancreases were removed and pancreatic protein extracts were analyzed by western blot for the expression of P-ERK1/2 and total-ERK1/2 proteins. Four and five rats were analyzed in the STZ/saline and STZ/KGF group respectively. doi:10.1371/journal.pone.0004734.g005 proceeded to the inactivation of PI3K by wortmannin. Treatment of rats with the PI3K inhibitor abolished the effect of KGF on PDX1 expression as evidenced by a significant decrease in the number of PDX1 positive ductal cells in the pancreas of 2-day-old KGF treated diabetic pups pre-treated with wortmannin, compared to the KGF treated group, while this treatment did not alter the basal number of PDX1 expressing ductal cells (Figures 9A and 9B). Inactivation of PI3K/AKT pathway and its impact on KGF-induced ductal cell proliferation. We have also evaluated the rate of ductal cell proliferation in the STZ/ wortmannin/KGF group, as compared to the STZ/KGF group. This parameter was found to be similar in both groups (Figure 9C), indicating that the PI3K/AKT pathway is implicated in the differentiation but not in the proliferation process in the pancreatic ductal cells. Interestingly, the proliferation of acinar cells was dramatically reduced in the pancreas of wortmannin treated rats, either alone or associated with the treatment with KGF (not shown). Discussion Type 1 and type 2 diabetes are associated with absolute or relative loss of pancreatic beta cells. The shortage of islet tissue critically hampers the wide application of cell replacement therapy for type 1 diabetes and unfortunately patients with type 2 diabetes still suffer from the absence of curative treatments for the disease. Alternative cellular sources with potential to be induced to express beta cell phenotype can overcome beta cell deficiency and lead to major advances for the cure of this devastating disease. Pancreatic duct cells represent such an alternative source for generation of new beta cells [1] either ex-vivo prior to transplantation, or in vivo in the pancreas of diabetic patients. During pancreas organogenesis, all pancreatic cell types arise from common precursor cells residing in ductal structures which result from evagination and branching of the ventral and dorsal pancreatic buds [20–22]. Ducts are also important for the generation of beta cells in the post natal life. This has been illustrated during the compensatory attempt to replace the normal beta cell population, for instance in animal models of cell injury induced by subtotal pancreatectomy [23–25] or in neonatal pancreas following beta cell destruction by the toxic agent streptozotocin [9,26]. In these contexts apart from proliferation of existing beta cells, differentiation of precursor cells located in ductal epithelium participate to the recovery of the beta cell mass [9,23,27–33]. A substantial body of evidence in the literature thus designates pancreatic duct as an important source for generation of new beta cells both in vitro and in vivo [1,25,34]. Therefore, characterization of stimuli with potential to induce the expansion of ductal cells and/or to drive their differentiation into beta cells is of great importance. We and others have previously showed that in vivo adminis- tration of KGF induces ductal cell proliferation [5] and beta cell regeneration in diabetic rats [6]. Modulation of signaling pathways by external stimuli represents key strategy used by the cellular machinery to regulate cell proliferation and differentiation processes. The understanding of these mechanisms is essential for the outcome of regenerative therapies of diabetes based on the induction duct cell proliferation and differentiation. The present study was undertaken to dissect the intracellular signaling pathways which mediate the growth and differentiation promoting effects of KGF in pancreatic ductal cells. As a cell system to decipher the intracellular mechanisms underlying the effects of KGF on ductal cell proliferation in vitro, we used Panc-1 cells, a ductal cell line derived from human adenocarcinoma. This cell line has been used in several studies as a model for the induction of duct-to-beta cell differentiation [35,36]. As these cells have high proliferative capacity in the presence of serum, all the experiments were conducted in the context of serum starvation in order to induce quiescence. We showed that KGF PLoS ONE | www.plosone.org 7 March 2009 | Volume 4 | Issue 3 | e4734