Joerg C Gerlach
University Of Pittsburgh At Pittsburgh
Project start date: 2011-09-01
Project end date: 2015-05-31
Sponsored Links Excellgen http://Excellgen.com
Grants awarded to Joerg C Gerlach
3D Culture Of MES Cells In Four-Compartment Bioreactors
Joerg C Gerlach, Professor Of Surgery And Bioengineering
University Of Pittsburgh At Pittsburgh 350 Thackeray Hall Pittsburgh, Pa 15260
Grant 5R21EB005739-02 from National Institute Of Biomedical Imaging And Bioengineering IRG: ISD
Abstract: Embryonic stem (ES) cell research and scale-up for development of possible clinical therapies is limited by the existing 2D dish culture methods. Our proposed studies present a new approach, in which ES cells are expanded under 3D medium perfusion conditions within four-compartment hollow fiber-based bioreactors. The design of the bioreactors allows integral oxygenation and efficient transfer of nutrients and waste products to and from the cells, cultured at high density involving minimal solute gradients within the cell compartment. Additionally, the interwoven fibers provide a scaffold allowing the cells to form 3D structures where the size of cellular aggregates is limited by the spacing between the hollow fibers. We propose that the well-controlled and versatile culture environment provided by our bioreactor is ideal for both large-scale expansion of undifferentiated ES cells and directed differentiation of ES cells using numerous strategies, including controlled exposure of the cells to molecular reagents and compartmentalized co-culture with mature cells. The objective of this 2-year project is to take the first step toward applying our bioreactor technology to ES cell research, by expanding and maintaining undifferentiated mouse embryonic stem (mES) cells in laboratory-scale versions of our bioreactor. We hypothesize that undifferentiated mES cells can be expanded and maintained in the perfused 3D environment provided by our bioreactor, and that within this 3D culture model mES cell pluripotency can be maintained by culturing mES cells and fibroblast feeder cells in two separate bioreactors perfused within one circuit (compartmentalized co-culture). The specific aim of the project is to 1. Develop a 3D culture model for bioreactor expansion and maintenance of undifferentiated mES cells, incorporating compartmentalized co-culture of mES cells with fibroblast feeder cells and allowing for enzymatic mES cell harvesting. The research plan consists of the following tasks 1.1 Develop the 3D culture model incorporating direct co-culture of mES cells and feeder cells; 1.2 Develop the culture model incorporating compartmentalized co-culture of mES cells and feeder cells; and 1.3 Develop a protocol for enzymatic mES cell harvesting from intact bioreactors. Completion of the project will provide a solid foundation for future studies on 1) large-scale, potentially automated bioreactor expansion of ES cells; and 2) bioreactor-based directed differentiation of ES cells under perfused 3D tissue-density conditions.
Keywords: bioreactor, embryonic stem cell, mixed tissue /cell culture, pluripotent stem cell, technology /technique development, tissue /cell culture, cell differentiation, cell proliferation, animal tissue, biotechnology
Project start date: 2005-09-06
Project end date: 2007-07-31
5R21EB005739-02 (2006): $181263
1R21EB005739-01 (2005): $203350