Developing a Microfluidic Device for Adenoviral Transfection of Pancreatic Islets

Abstract

Diabetes is a global epidemic that is associated with the deterioration of pancreatic beta-cell mass and function. Beta-cell function within the islet has previously been studied ex vivo and the role of specific proteins is commonly investigated with overexpression models using viral transfection. However, poor penetration and reduced transfection efficiency limits the application of this technology in whole ex vivo islets. To improve transfection efficiency and maintain islet structure, we aim to develop a method to bring viral particles deeper into the tissue. Improving upon a previous microfluidic device design featuring hydrodynamic traps (nozzles) in series, a spiral-shaped microfluidic device with parallel nozzles was created to trap, transfect and deliver nutrients to islets. The device was engineered to provide sufficient resistance and pressure drop along the nozzles, allowing for homogeneous transfection throughout captured islets. Computer-generated models were used to optimize our design parameters to minimize pressure drop variations along with shear induced damage of islets. Through quantitative fluorescence microscopy, we have measured the media exchange rate through islets and have determined a range of operating flow-rates. We have also shown successful culturing of the loaded islets in our microfluidic device. Beta-cell-specific transfection in whole islets with desired genes will facilitate the investigation of islets in a more physiologically relevant environment. Therefore, we have developed a beta-cell specific shuttle vector in order to deliver fluorescent protein-tagged genes via adenoviral particles. Currently viral penetration efficiency is being tested in this device through fluorescence microscopy and western immunoblotting. We aim to create a multi-purpose platform that is not limited to the study of beta-cell biology in islets, but may be translated to the investigation of other tissue models and diseases.