Quantitating intracellular transport of polyplexes by spatio-temporal image correlation spectroscopy.
Quantitatively understanding how nonviral gene delivery vectors (polyplexes) are transported inside cells is essential before they can be optimized for gene therapy and medical applications. In this study, we used spatio-temporal image correlation spectroscopy (ICS) to follow polymer-nucleic acid particles (polyplexes) of various sizes and analyze their diffusive-like and flow behaviors intracellularly to elucidate the mechanisms responsible for their transport. ICS is a quantitative imaging technique that allows the assessment of particle motion in complex systems, although it has not been widely used to date. We find that the internalized polyplexes are able to use microtubule motors for intracellular trafficking and exhibit different transport behaviors for short (<10 s) versus long (approximately 60 s) correlation times. This motion can be explained by a memory effect of the microtubule motors. These results reveal that, although microtubule motor biases may be present for short periods of time, resulting in a net directional velocity, the overall long-term motion of the polyplexes is best described as a random walk-like process. These studies suggest that spatio-temporal ICS is a powerful technique for assessing the nature of intracellular motion and provides a quantitative tool to compare the transport of different objects within a living cell.
Kulkarni
Rajan P
RP
Option in Biochemistry and Molecular Biophysics, California Institute of Technology, Pasadena, CA 91125, USA.
Wu
David D
DD
Davis
Mark E
ME
Fraser
Scott E
SE
D001692:Biological Transport / Q000502:physiology; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D008863:Microspheres; D008870:Microtubules / Q000378:metabolism*; D020409:Molecular Motor Proteins / Q000378:metabolism*; D011094:Polyethyleneimine / Q000737:chemistry / Q000378:metabolism; D013057:Spectrum Analysis / Q000379:methods*; D047392:beta-Cyclodextrins / Q000737:chemistry / Q000378:metabolism
D020409:Molecular Motor Proteins; D047392:beta-Cyclodextrins; D011094:Polyethyleneimine; C031215:betadex