Acute Pancreatitis and Pancreatic Cancer Figure 5. Nestin is expressed in human and mouse PanIN. (A–D) Human PanIN; mouse PanIN in NK (E–G) and PK (I–K). Overlays of CK19 (green), Nestin (red) and Hoescht (blue) show clear coexpression of Nestin with CK19 in the lesions (A, E, I). In contrast, in normal ducts, only CK19 is expressed (arrows)(H, L). (D) On human tissue, the secondary antibody alone generates high background in the stroma but no expression in PanIN (arrow). Scale bar: 20 mm. doi:10.1371/journal.pone.0027725.g005 similar again to the one observed in P/K mice following caerulein- induced AP [14]. These data demonstrate that N/K mice are a bona fide model of PDAC, and further support that the pancreatic Nestin lineage is the cell lineage that is mainly responsive to Kras mutation in the pancreas. The targeting of oncogenic Kras to the acinar compartment, using different acinar-specific promoters also leads to the development in adult mouse of PanINs [8,9] and infrequent invasive PDAC [7]. In N/K mice, KrasG12D is activated in acinar cells as well as in adult Nestin-expressing cells. These later cells represent less than 0.1% of the total pancreas, appear integrated in the acinar structure but do not display differentiation markers (manuscript in preparation, CC and MK). At this stage, their function remains unknown. It is important to note that the Nestin regulatory elements used for these experiments target all Nestin- expressing cells in the animal [17]. Cre expression can be traced occasionally in endothelial and mesenchymal cells but there are no data in the literature suggesting that pancreatic cancer could initiate in non parenchymal cells. Thus taken together, these data provide strong evidence for a non-ductal cell origin of PDAC at least in the mouse. Progression from PanIN to cancer is dramatically accelerated when the mice are subjected to environmental insults such as chronic pancreatitis [7,10]. These data correlate with human epidemiological studies showing that chronic pancreatitis increases the risk of developing PDAC [13]. Our data and others show that caerulein-induced AP acts as cancer promoter in PDAC mouse models [14,15,16], suggesting that AP, when occurring in an individual whose pancreas harbors a Kras mutation, could potentially increase PDAC risk. By which mechanisms does AP affect cancer progression? Recovery from AP in humans does not necessarily lead to irreversible damage and may be associated with pancreatic regeneration [28] as observed in the mouse. During the first 3 days following caerulein treatment, the mouse pancreas develop tubular structures in place of acini [23], and lineage tracing experiments using the elastase promoter suggest that part of these tubular structures are derived from acinar cells that appear to dedifferentiate, by going through a ductal-like metaplastic intermediate state before entering proliferation [23,29]. New acinar cells originate then essentially from preexisting acinar cells [22,23,29] and have to undergo a secondary ductal to acinar metaplasia (DAM), an essential step for pancreas regeneration [15]. Interestingly, following AP, acinar cells that expressed oncogenic Kras died or underwent ADM. Among the acinar structures undergoing ADM, some appeared to give rise to PanIN, at least during the short period in which the mice were analyzed [15]. Pancreas regeneration also appeared to be stalled suggesting that KrasG12D-expressing cells that underwent ADM were not able to undergo the reverse process DAM to generate new acinar cells [15]. We observed extensive ADM/low-grade PanIN formation in both P/K and N/K pancreata in the 2–6 months following AP arguing again that large number of pancreatic cells respond to oncogene activation. Still, only few higher grade lesions can be detected. It suggests that while most acinar cells do respond to a certain extent to oncogenic Kras following caerulein-induced AP by undergoing ADM, they are not competent to progress to higher grade lesions and PDAC. Nestin expression is upregulated in different models of pancreas regeneration and Nestin has been proposed as a transient marker of acinar dedifferentiation [26,29]. It is also known as a stem cell marker during both embryogenesis and adulthood in different organs. One could then speculate that pancreatic adult Nestin- expressing cells are in fact adult acinar progenitors and that pancreas regeneration following AP occurs in part via the expansion of these progenitors. As such, adult Nestin-expressing cells would potentially be more competent for cancer initiation and progression following AP-induced proliferation, as suggested by the expression of Nestin in all grade lesions both in human and in mice. More experiments will be required to evaluate these possibilities. Recent work showed that the inflammatory mediator Stat3 is activated at all stages of pancreatic cancer from low-grade PanIN to PDAC and is required for cancer progression [25,30]. AP causes transient Stat3 activation in normal pancreas. In P/K mice, caerulein-induced AP leads to a major upregulation of phospho- Stat3 both in neoplastic lesions and the surrounding microenvi- ronment in the 2–3 weeks following its induction. Indeed, PLoS ONE | www.plosone.org 5 November 2011 | Volume 6 | Issue 11 | e27725