title: Microfluidic engineered high cell density three-dimensional neural cultures.
abstract: Three-dimensional (3D) neural cultures with cells distributed throughout a thick, bioactive protein scaffold may better represent neurobiological phenomena than planar correlates lacking matrix support. Neural cells in vivo interact within a complex, multicellular environment with tightly coupled 3D cell-cell/cell-matrix interactions; however, thick 3D neural cultures at cell densities approaching that of brain rapidly decay, presumably due to diffusion limited interstitial mass transport. To address this issue, we have developed a novel perfusion platform that utilizes forced intercellular convection to enhance mass transport. First, we demonstrated that in thick (>500 microm) 3D neural cultures supported by passive diffusion, cell densities <or=5.0 x 10(3) cells mm(-3) were required for survival. In 3D neuronal and neuronal-astrocytic co-cultures with increased cell density (10(4) cells mm(-3)), continuous medium perfusion at 2.0-11.0 microL min(-1) improved viability compared to non-perfused cultures (p < 0.01), which exhibited widespread cell death and matrix degradation. In perfused cultures, survival was dependent on proximity to the perfusion source at 2.00-6.25 microL min(-1) (p < 0.05); however, at perfusion rates of 10.0-11.0 microL min(-1) survival did not depend on the distance from the perfusion source, and resulted in a preservation of cell density with >90% viability in both neuronal cultures and neuronal-astrocytic co-cultures. This work demonstrates the utility of forced interstitial convection in improving the survival of high cell density 3D engineered neural constructs and may aid in the development of novel tissue-engineered systems reconstituting 3D cell-cell/cell-matrix interactions.
authors.affiliation: Wallace H Coulter Department of Biomedical Engineering, Parket H Petit Institute for Bioengineering and Bioscience, Laboratory for Neuroengineering, Georgia Institute of Technology, Atlanta, GA, USA. dkacy@neuro.gatech.edu
authors.forename: D Kacy
authors.initials: DK
authors.lastname: Cullen
authors.forename: Jelena
authors.initials: J
authors.lastname: Vukasinovic
authors.forename: Ari
authors.initials: A
authors.lastname: Glezer
authors.forename: Michelle C
authors.initials: MC
authors.lastname: Laplaca
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D019149:Bioreactors; D002452:Cell Count; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods; D002478:Cells, Cultured; D004867:Equipment Design; D019544:Equipment Failure Analysis; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D009415:Nerve Net / Q000166:cytology* / Q000502:physiology*; D009474:Neurons / Q000166:cytology* / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D023822:Tissue Engineering / Q000295:instrumentation* / Q000379:methods