Design and characterization of a microfluidic device resembling exocrine pancreas

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal tumor affecting the exocrine pancreas. Its poor survival rate is attributed to late-stage symptom onset, invasive metastasis, and limited therapeutic options. To address these challenges, in vitro models that replicate the pancreatic tissue microenvironment are crucial for therapy development and to predict drug efficacy. Organ-on-chip technology offers a powerful approach to achieving these goals. In this thesis project, a microfluidic device is developed to model the cellular composition of the pancreatic acino-ductal unit. The device comprises a multilayer structure using Polydimethylsiloxane (PDMS), with three layers, two of them separated by a membrane. In particular, the bottom layer is loaded with human foreskin fibroblasts (HFF-1) embedded in a type I collagen hydrogel to mimic the stromal component, while the top layer reproduces the healthy or cancer epithelial component, using human pancreatic ductal epithelial cells (HPDE). An additional reservoir layer provides nutrients supply during the incubation time. The nanofibrous membrane, made of polycaprolactone and gelatin (PCL/Gel), enables the crosstalk between PDAC and stromal components, which is recognized as a key player in cancer progression. The fabrication process involved replica molding for the PDMS layers and SU-8 photolithography or 3d printing for the molds, whereas solution electrospinning was used to produce the membrane. Surface and morphological characterization of the molds and microfluidic device were performed using optical microscopy and fluidic characterization was conducted through insertion and diffusivity tests. In vitro studies allowed to evaluate cell proliferative capability and cellular response using various pancreatic tissue-specific cell lines seeded in the top layer, such as Human Pancreatic Duct Epithelial Cells in wild-type form that show non-mutated gene (HPDE-WT) to mimic healthy microenvironment, and Human Pancreatic Duct Epithelial Cells with oncogene KRAS mutation (HPDE-KRAS) or MIA-PaCa-2 cells for the pathological condition. Viability assays and fluorescence imaging confirmed the cellular functionality and the physiological morphological structure of cells in the different layers of the device. Overall, these studies demonstrated the successful optimization and characterization of this microfluidic device for modeling the pancreatic acino-ductal unit and studying the interaction between PDAC and stromal components. The system provides valuable insights into the tumor microenvironment and holds potential for drug development and understanding cellular crosstalk in PDAC.