Effects of cell culture scaffold stiffness on cell membrane damage induced by sonoporation.
OBJECTIVE
As basic studies to realize in vivo sonoporation, rates of cell membrane damage during sonoporation were evaluated using monolayer cells cultured on scaffolds with different degrees of stiffness.


METHODS
Four types of scaffolds, constructed using collagen gel, 10 and 30 % acrylamide gels, and a coverslip, were used for cultivation of monolayer cells. Young's moduli measured using an atomic force microscope were in the range 0.09-8.6 kPa for the gel scaffolds, whereas Young's modulus for living cells was 4.5 kPa. Cells with attached microbubbles were exposed to one-shot pulsed ultrasound of 8.0/-1.3 MPa in peak positive/negative pressures with durations of 3, 100, and 10,000 cycles.


RESULTS
Cell membrane damage was visualized by fluorescence microscopy using propidium iodide. The 3-cycle ultrasound pulse had no significant effect; however, the rates of damage caused by 100-cycle and 10,000-cycle pulses showed a strong tendency for higher rates of damage with a higher Young's modulus.


CONCLUSIONS
The experimental results indicate that the stiffness of the underlying layer of adherent cells should be considered as an essential parameter of the sonoporation condition and that the optimum exposure conditions for in vivo sonoporation should be determined with consideration of the physical properties of underlying tissues.
Graduate School of Information Science and Technology, Hokkaido University, N14W9, Kita-ku, Sapporo, 060-0814, Japan. kudo@bme.ist.hokudai.ac.jp.
Nobuki
N
Kudo
Graduate School of Information Science and Technology, Hokkaido University, N14W9, Kita-ku, Sapporo, 060-0814, Japan.
Yuto
Y
Kinoshita
D020106:Acrylamide; D018929:Cell Culture Techniques / Q000295:instrumentation*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000473:pathology / Q000502:physiology*; D002478:Cells, Cultured; D003094:Collagen; D004396:Coloring Agents; D004548:Elasticity; D005782:Gels; D006801:Humans; D045423:Microbubbles; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D011419:Propidium; D054457:Tissue Scaffolds; D000069453:Ultrasonic Waves
D004396:Coloring Agents; D005782:Gels; D020106:Acrylamide; D011419:Propidium; D003094:Collagen
Cell damage; Gel scaffold; Microbubbles; Scaffold stiffness; Sonoporation