Ensemble analysis of angiogenic growth in three-dimensional microfluidic cell cultures.
We demonstrate ensemble three-dimensional cell cultures and quantitative analysis of angiogenic growth from uniform endothelial monolayers. Our approach combines two key elements: a micro-fluidic assay that enables parallelized angiogenic growth instances subject to common extracellular conditions, and an automated image acquisition and processing scheme enabling high-throughput, unbiased quantification of angiogenic growth. Because of the increased throughput of the assay in comparison to existing three-dimensional morphogenic assays, statistical properties of angiogenic growth can be reliably estimated. We used the assay to evaluate the combined effects of vascular endothelial growth factor (VEGF) and the signaling lipid sphingoshine-1-phosphate (S1P). Our results show the importance of S1P in amplifying the angiogenic response in the presence of VEGF gradients. Furthermore, the application of S1P with VEGF gradients resulted in angiogenic sprouts with higher aspect ratio than S1P with background levels of VEGF, despite reduced total migratory activity. This implies a synergistic effect between the growth factors in promoting angiogenic activity. Finally, the variance in the computed angiogenic metrics (as measured by ensemble standard deviation) was found to increase linearly with the ensemble mean. This finding is consistent with stochastic agent-based mathematical models of angiogenesis that represent angiogenic growth as a series of independent stochastic cell-level decisions.
Department of Mechanical Engineering, Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts, United States of America. wfarahat@mit.edu
Waleed A
WA
Farahat
Levi B
LB
Wood
Ioannis K
IK
Zervantonakis
Alisha
A
Schor
Sharon
S
Ong
Devin
D
Neal
Roger D
RD
Kamm
H Harry
HH
Asada
D043925:Angiogenesis Inducing Agents / Q000494:pharmacology*; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology; D006801:Humans; D008246:Lysophospholipids / Q000494:pharmacology; D044085:Microfluidics / Q000295:instrumentation / Q000379:methods*; D018919:Neovascularization, Physiologic / Q000187:drug effects* / Q000502:physiology*; D013110:Sphingosine / Q000031:analogs & derivatives / Q000494:pharmacology; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D043925:Angiogenesis Inducing Agents; D008246:Lysophospholipids; D042461:Vascular Endothelial Growth Factor A; C060506:sphingosine 1-phosphate; D013110:Sphingosine