Proteomic Analysis of Differentially Expressed Proteins in Peripheral Cholangiocarcinoma

Abstract

Cholangiocarcinoma is an adenocarcinoma of the liver which has increased in incidence over the last thirty years to reach similar levels to other liver cancers. Diagnosis of this disease is usually late and prognosis is poor, therefore it is of great importance to identify novel candidate markers and potential early indicators of this disease as well as molecules that may be potential therapeutic targets. We have used a proteomic approach to identify differentially expressed proteins in peripheral cholangiocarcinoma cases and compared expression with paired non-tumoral liver tissue from the same patients. Two-dimensional fluorescence difference gel electrophoresis after labeling of the proteins with cyanines 3 and 5 was used to identify differentially expressed proteins. Overall, of the approximately 2,400 protein spots visualised in each gel, 172 protein spots showed significant differences in expression level between tumoral and non-tumoral tissue with p<0.01. Of these, 100 spots corresponding to 138 different proteins were identified by mass spectrometry: 70 proteins were over-expressed whereas 68 proteins were under-expressed in tumoral samples compared to nontumoral samples. Among the over-expressed proteins, immunohistochemistry studies confirmed an increased expression of 14-3-3 protein in tumoral cells while α-smooth muscle actin and periostin were shown to be overexpressed in the stromal myofibroblasts surrounding tumoral cells. α-Smooth muscle actin is a marker of myofibroblast differentiation and has been found to be a Ian A. Darby, Karine Vuillier-Devillers, and Émilie Pinault have equally contributed to this work.

Periostin in intrahepatic cholangiocarcinoma: Pathobiological insights and clinical implications

“…As already noted in the previous section of this review, in the case of both ICC and PDAC, there are very convincing mRNA and protein data to support tumor stromal CAFs as being the primary if not exclusive source of Postn overexpressed in these respective desmoplastic GI cancers [41, 43, 45]. In agreement with the findings of Utispan et al .…”

“…In ICC, like in pancreatic ductal adenocarcinoma (PDAC), Postn has been largely, if not exclusively localized to the desmoplastic connective tissue stroma [30, 41–45]. Moreover, various methods including real-time-PCR, immunohistochemistry, and Western blotting have demonstrated myofibroblastic CAFs positive for α-SMA to be a major cellular source of Postn secreted into the desmoplastic stroma of human and rat cholangiocarcinoma in liver [43–46].…”

The role of the myofibroblast in tumor stroma remodeling

“…This expression of MMPs and TIMPs is stronger in cholangiocarcinomas with severe invasion 257 . Our lab has recently used a proteomic approach to identify differentially expressed proteins in peripheral cholangiocarcinoma cases and compared expression with paired non-tumor liver tissue from the same patients 258 . Overall, of the approximately 2,400 protein spots visualized in each gel, 172 protein spots showed significant differences in expression level between tumor and non-tumor tissue with p < 0.01.…”

Neural Stem Cell Properties of an Astrocyte Subpopulation Sorted by Sedimentation Field-Flow Fractionation

“…Protein zones of interest were excised and the proteins were digested using sequencing grade modified trypsin (Promega) as previously described. 17 The resulting peptides were analyzed by nanoLC-MS/MS using an LC Packings system (Dionex) coupled with a QTRAP mass spectrometer (Applied Biosystems). The QTRAP was operated in informationdependent acquisition mode with Analyst 1.4.1 software (Applied Biosystems).…”