Assembling neurospheres: dynamics of neural progenitor/stem cell aggregation probed using an optical trap.
Optical trapping (tweezing) has been used in conjunction with fluid flow technology to dissect the mechanics and spatio-temporal dynamics of how neural progenitor/stem cells (NSCs) adhere and aggregate. Hitherto unavailable information has been obtained on the most probable minimum time (∼5 s) and most probable minimum distance of approach (4-6 µm) required for irreversible adhesion of proximate cells to occur. Our experiments also allow us to study and quantify the spatial characteristics of filopodial- and membrane-mediated adhesion, and to probe the functional dynamics of NSCs to quantify a lower limit of the adhesive force by which NSCs aggregate (∼18 pN). Our findings, which we also validate by computational modeling, have important implications for the neurosphere assay: once aggregated, neurospheres cannot disassemble merely by being subjected to shaking or by thermal effects. Our findings provide quantitative affirmation to the notion that the neurosphere assay may not be a valid measure of clonality and "stemness". Post-adhesion dynamics were also studied and oscillatory motion in filopodia-mediated adhesion was observed. Furthermore, we have also explored the effect of the removal of calcium ions: both filopodia-mediated as well as membrane-membrane adhesion were inhibited. On the other hand, F-actin disrupted the dynamics of such adhesion events such that filopodia-mediated adhesion was inhibited but not membrane-membrane adhesion.
UM-DAE Centre for Excellence in Basic Science, University of Mumbai, Kalina Campus, Mumbai, India. brainwave@cbs.ac.in
Uma
U
Ladiwala
Himanish
H
Basu
Deepak
D
Mathur
D000818:Animals; D002448:Cell Adhesion / Q000502:physiology; D002449:Cell Aggregation / Q000502:physiology; D002478:Cells, Cultured; D005260:Female; D007150:Immunohistochemistry; D008297:Male; D058953:Neural Stem Cells / Q000166:cytology*; D009474:Neurons / Q000166:cytology*; D052898:Optical Tweezers; D051381:Rats; D017207:Rats, Sprague-Dawley