Optimization of Fibrin Scaffolds for Differentiation of Murine Embryonic Stem Cells into Neural Lineage Cells
Date
2006-12
Authors
Willerth, Stephanie
Arendas, Kelly J
Gottlieb, David I
Sakiyama-Elbert, Shelly E
Journal Title
Journal ISSN
Volume Title
Publisher
Elsevier
Abstract
The objective of this research was to determine the appropriate cell culture conditions for embryonic stem (ES) cell proliferation and differentiation in fibrin scaffolds by examining cell seeding density, location, and the optimal concentrations of fibrinogen, thrombin, and aprotinin (protease inhibitor). Mouse ES cells were induced to become neural progenitors by adding retinoic acid for 4 days to embryoid body (EB) cultures. For dissociated EBs, the optimal cell seeding density and location was determined to be 250,000 cells/cm² seeded on top of fibrin scaffolds. For intact EBs, three dimensional (3D) cultures with one EB per 400 μL fibrin scaffold resulted in greater cell proliferation and differentiation than two dimensional (2D) cultures. Optimal concentrations for scaffold polymerization were 10 mg/mL of fibrinogen and 2 NIH units/mL of thrombin. The optimal aprotinin concentration was determined to be 50 μg/mL for dissociated EBs (2D) and 5 μg/mL for intact EBs in 3D fibrin scaffolds. Additionally, after 14 days in 3D culture EBs differentiated into neurons andastrocytes as indicated by immunohistochemisty. These conditions provide an optimal fibrin scaffold for evaluating ES cell differentiation and proliferation in culture, and for use as a platform for neural tissue engineering applications, such as the treatment for spinal cord injury.
Description
Author manuscript, post peer review (post-print)
Keywords
nerve tissue engineering, cell spreading, cell culture, progenitor cells
Citation
Willerth, S.M. et al (2006) Optimization of Fibrin Scaffolds for Differentiation of Murine Embryonic Stem Cells into Neural Lineage Cells. Biomaterials 27(36): 5990–6003.