Proteomics portrait of archival lesions of chronic pancreatitis

Abstract

Chronic pancreatitis is a chronic inflammatory disorder of the pancreas. The etiology is multi-fold, but all lead to progressive scarring and loss of pancreatic function. Early diagnosis is difficult; and the understanding of the molecular events that underlie this progressive disease is limited. In this study, we investigated differential proteins associated with mild and severe chronic pancreatitis in comparison with normal pancreas and pancreatic cancer. Paraffin-embedded formalin-fixed tissues from five well-characterized specimens each of normal pancreas (NL), mild chronic pancreatitis (MCP), severe chronic pancreatitis (SCP) and pancreatic ductal adenocarcinoma (PDAC) were subjected to proteomic analysis using a "label-free" comparative approach. Our results show that the numbers of differential proteins increase substantially with the disease severity, from mild to severe chronic pancreatitis, while the number of dysregulated proteins is highest in pancreatic adenocarcinoma. Important functional groups and biological processes associated with chronic pancreatitis and cancer include acinar cell secretory proteins, pancreatic fibrosis/stellate cell activation, glycoproteins, and inflammatory proteins. Three differential proteins were selected for verification by immunohistochemistry, including collagen 14A1, lumican and versican. Further canonical pathway analysis revealed that acute phase response signal, prothrombin activation pathway, and pancreatic fibrosis/pancreatic stellate cell activation pathway were the most significant pathways involved in chronic pancreatitis, while pathways relating to metabolism were the most significant pathways in pancreatic adenocarcinoma. Our study reveals a group of differentially expressed proteins and the related pathways that may shed light on the pathogenesis of chronic pancreatitis and the common molecular events associated with chronic pancreatitis and pancreatic adenocarcinoma.

Figure 1. Histology of chronic pancreatitis, pancreatic adenocarcinoma and normal pancreas.

a). non-disease pancreas; b). mild chronic pancreatitis; c). severe chronic pancreatitis; d). pancreatic ductal adenocarcinoma.

Figure 2. The workflow of the label-free quantitative proteomics pipeline for analyzing dissected formalin-fixed, paraffin-embedded tissue specimens.

Figure 3. Quantitative proteomics results.

a). evaluation of the reproducibility between the duplicates at protein level; b). histogram of protein ratio distribution in natural log scale.

Figure 4. The differentially expressed proteins in MCP, SCP and PDAC.

a). the number of differential proteins in MCP, SCP and PDAC using normal pancreas as a comparison. b). the overlap of the differential proteins with concurrent abundant changes in MCP, SCP and PDAC.

Figure 5. The expression of pancreatic acinar cell secretory enzymes and the related inhibitors.

Figure 6. The expression of some of the differential proteins related to ECM and stellate cell activation.

Figure 7. The expression of some of the differential proteins associated with wounding response and inflammation.

Figure 8. Immunohistochemical confirmation of proteomic findings in chronic pancreatitis and pancreatic adenocarcinoma.

Lumican (a–d) not seen in NL (a), but is overexpressed in acinar cells of MCP (b), SCP (c), and only in the stroma of PDAC (d). Versican (e–h) is mainly over-expressed in the stroma of MCP (f), SCP (g), and PDAC (h). Col14A1 (i–l) is over-expressed in acinar and ductal cells of MCP (j), and SCP (k), but not in PDAC (l).