Schlett K$, Czirok A, Tárnok K, Vicsek T, Madarász E*:
Dynamics of cell aggregation during in vitro neurogenesis by immortalized neuroectodermal progenitors
Published in J Neurosci Res 2000 Apr 15;60(2):184-94 (pdf)

$ Department of Physiology, Eötvös University, Budapest

Early events of in vitro neuronal development were studied by inducing neuron formation in a neuroectodermal cell line, NE-4C/A3, derived from the embryonic forebrain vesicles of p53-deficient mice. Neuronal differentiation was initiated by treating the cells with all-trans retinoic acid (RA). By the second day of RA treatment compact cell aggregates were formed. The first signs of neuronal
cell fate decision were revealed inside the aggregates. To elucidate the process of aggregate formation, the dynamics of cell clustering and the migration of individual cells were investigated by a novel computer-controlled videomicroscopic system. Besides real-time observation of cell motility, the system allowed statistical analysis of large sets of data providing quantitative evaluation of cell locomotion during an early, critical phase of RA induced
neuron formation. The results showed that chemoattractants did not play a principal role in cell aggregation. Retinoic acid, on the other hand, was found to cause a rapid decrease in the average migratory velocity without changing the randomness of migratory routes. The data indicated that aggregation was facilitated by increased cohesion upon incident collision of randomly encountering
cells. The resulting compact cell clusters provided the structural conditions for contact communication apparently needed for the neuronal differentiation of NE-4C/A3 cells.


Untreated culture of neuroectodermal progenitor cells:1.7Mb mpeg stream
Objective: 10X (Field of view: 500um X 370um). Duration of recording: 2 days. Number of frames: 525 (5 min/frame).
Cells grow until confluency and occasionally form small clusters.

Culture treated with Retinoic acid: 1.7Mbyte mpeg stream.
Objective: 10X (Field of view: 500um X 370um). Duration of recording: 2 days. Number of frames: 574 (5 min/frame).
After some time cells form tight, multicellular aggregates in which the differentiation starts. Note that the aggregates are motile, but individual cells can never break free.

Extra-long history of a Retinoic acid-treated culture:9Mbyte mpeg stream.
Objective: 3X (Field of view: 1240um X 920um). Duration of recording: 4 days. Number of frames: 1356 (4 min/frame).
After 3 days the aggregation process is reversed, and a monolayer develops with already differentiated cells, some of them extending long processes.