Tumor-Stromal Interactions in a Co-Culture Model of Human Pancreatic Adenocarcinoma Cells and Fibroblasts and Their Connection with Tumor Spread.

Elena Prieto-García,Marina Alonso-Riaño,C Vanesa Díaz-García,Virginia Pardo-Marqués,Luis Paz-Ares,Inés García-Consuegra,M Teresa Agulló-Ortuño,Carlos Pérez,Alba Agudo-López,Sara Asensio-Peña,Carlos Gómez,Jorge Adeva,José A López-Martín

doi:10.3390/biomedicines9040364

Abstract

One key feature of pancreatic ductal adenocarcinoma (PDAC) is a dense desmoplastic reaction that has been recognized as playing important roles in metastasis and therapeutic resistance. We aim to study tumor–stromal interactions in an in vitro coculture model between human PDAC cells (Capan-1 or PL-45) and fibroblasts (LC5). Confocal immunofluorescence, Enzyme-Linked Immunosorbent Assay (ELISA), and Western blotting were used to evaluate the expressions of activation markers; cytokines arrays were performed to identify secretome profiles associated with migratory and invasive properties of tumor cells; extracellular vesicle production was examined by ELISA and transmission electron microscopy. Coculture conditions increased FGF-7 secretion and α-SMA expression, characterized by fibroblast activation and decreased epithelial marker E-cadherin in tumor cells. Interestingly, tumor cells and fibroblasts migrate together, with tumor cells in forming a center surrounded by fibroblasts, maximizing the contact between cells. We show a different mechanism for tumor spread through a cooperative migration between tumor cells and activated fibroblasts. Furthermore, IL-6 levels change significantly in coculture conditions, and this could affect the invasive and migratory capacities of cells. Targeting the interaction between tumor cells and the tumor microenvironment might represent a novel therapeutic approach to advanced PDAC.

Highlights

Pancreatic ductal adenocarcinomas (PDACs) are among the most lethal tumors worldwide, with a 5-year overall survival of around 9% [1]
We have shown an important decrease in the expression of the epithelial marker E-cadherin in tumor cells from our coculture model, suggesting that epithelial mesenchymal transition (EMT) occurs in this system and would facilitate the initiation of metastasis [4]
We have shown a coculture model of tumor cells and fibroblasts that recapitulates some features occurring in human PDAC and might be used for a better understanding of tumor–stromal interactions and to identify new targets for pharmacological intervention

Summary

Introduction

Pancreatic ductal adenocarcinomas (PDACs) are among the most lethal tumors worldwide, with a 5-year overall survival of around 9% [1]. The increasing knowledge of cellular and molecular mechanisms involved in pancreatic carcinogenesis has failed to translate into new therapeutic approaches This may be due in part to the fact that most studies mainly focus on targeting tumor cells and obviate the complex surrounding microenvironment, which plays an important role in the evolution of the disease [4,5]. One of the hallmarks of PDAC is the formation of abundant and dense stroma wrapping tumor cells, termed desmoplastic reaction, which leads to an increase in the interstitial pressure, and to poor tissue perfusion and hypoxia. This tumor-associated stroma may represent up to 80% of the tumor mass in the majority of patients [6]. A population of surrounding fibroblasts known as cancer-associated fibroblasts (CAFs) play a critical role in tumor progression by stimulating survival and proliferation signaling pathways through heterotypic signaling with cancer cells [13,14]

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