A microfabricated array bioreactor for perfused 3D liver culture

TLDR

The authors designed and fabricated a perfused 3‑D liver bioreactor that supports tissue morphogenesis and allows repeated in‑situ light‑microscopy observation. The bioreactor uses silicon‑etched channel arrays with cell‑adhesive walls, a cell‑retaining filter, and a housing that delivers perfusate across the top and through each channel at flow rates matching oxygen demand while keeping shear stress below 2 dyne cm⁻², as verified by numerical fluid‑flow modeling. Primary rat hepatocytes cultured for up to two weeks reorganized into viable, tissue‑like structures, with pre‑aggregated spheroids enhancing morphogenesis and viability, and the system enabled repeated two‑photon imaging of structure and function. © 2002 Wiley Periodicals, Inc., Biotechnol Bioeng 78: 257–269.

Abstract

Abstract We describe the design, fabrication, and performance of a bioreactor that enables both morphogenesis of 3D tissue structures under continuous perfusion and repeated in situ observation by light microscopy. Three‐dimensional scaffolds were created by deep reactive ion etching of silicon wafers to create an array of channels (through‐holes) with cell‐adhesive walls. Scaffolds were combined with a cell‐retaining filter and support in a reactor housing designed to deliver a continuous perfusate across the top of the array and through the 3D tissue mass in each channel. Reactor dimensions were constructed so that perfusate flow rates meet estimated values of cellular oxygen demands while providing fluid shear stress at or below a physiological range (<2 dyne cm 2 ), as determined by comparison of numerical models of reactor fluid flow patterns to literature values of physiological shear stresses. We studied the behavior of primary rat hepatocytes seeded into the reactors and cultured for up to 2 weeks, and found that cells seeded into the channels rearranged extensively to form tissue like structures and remained viable throughout the culture period. We further observed that preaggregation of the cells into spheroidal structures prior to seeding improved the morphogenesis of tissue structure and maintenance of viability. We also demonstrate repeated in situ imaging of tissue structure and function using two‐photon microscopy. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 78: 257–269, 2002.