TY - JOUR
T1 - The role of RhoA kinase (ROCK) in cell alignment on nanofibers Mohammad Nahid Andalib1
AU - Lee, Jeong Soon
AU - Ha, Ligyeom
AU - Dzenis, Yuris
AU - Lim, Jung Yul
N1 - Funding Information:
The authors thank the funding supports from NSF NIRT-0709333, NSF CMMI-0600675, NSF CBET-1140065, AFOSR FA9550-11-1-0204 (all to Dzenis); AO Foundation Research Grant S-10-7L, NE DHHS Stem Cell Research Grant 2011-05, AHA Scientist Development Grant 12SDG12030109, Osteology Foundation Grant 12-006, UNL Interdisciplinary Research Grant (all to Lim).
PY - 2013/8
Y1 - 2013/8
N2 - While the potential of nanofibers as tissue engineering scaffolds has been demonstrated, very little has been revealed as regards the molecular mechanism by which cells sense and respond to nanofibers. It was hypothesized that RhoA kinase (ROCK), one of the vital cell tension signaling cascades, plays a role in regulating cell alignment on nanofibers. To test this, unidirectionally aligned and randomly distributed nanofibers, both with an average diameter of -130 nm, were fabricated with poly(L-lactic acid) (PLLA). A flat PLLA film was used as the control. Mesenchymal stem cells (MSCs, C3H10T1/2) displayed high fidelity in cell orientation along aligned nanofibers, and showed an increased cell spreading area on random nanofibers. Interestingly, cells cultured on aligned nanofibers displayed significantly greater ROCK expression relative to cells on a flat surface, as assessed by immunoblotting. To further test the role of ROCK, MSCs with ROCK small hairpin RNA (shRNA) were established. It is notable that, even when ROCK was stably knocked down via shRNA, cells could still display preferred orientation along aligned nanofibers. However, MSCs with shRNA-ROCK displayed a significantly decreased cell major axis length following aligned nanofibers compared with shRNA vector control, suggesting that ROCK may be involved in cell elongation on aligned nanofibers. Along with the reduction in cell length, cell area was decreased with ROCK silencing. These cell morphological changes induced by shRNA-ROCK were generally maintained on a flat surface and random nanofibers. A pharmacological ROCK inhibitor, Y-27632, produced results similar to those of shRNA-ROCK. The data on the role of ROCK in regulating cell alignment on nanofibers may provide a new mechanistic insight into nanofiber control of cells.
AB - While the potential of nanofibers as tissue engineering scaffolds has been demonstrated, very little has been revealed as regards the molecular mechanism by which cells sense and respond to nanofibers. It was hypothesized that RhoA kinase (ROCK), one of the vital cell tension signaling cascades, plays a role in regulating cell alignment on nanofibers. To test this, unidirectionally aligned and randomly distributed nanofibers, both with an average diameter of -130 nm, were fabricated with poly(L-lactic acid) (PLLA). A flat PLLA film was used as the control. Mesenchymal stem cells (MSCs, C3H10T1/2) displayed high fidelity in cell orientation along aligned nanofibers, and showed an increased cell spreading area on random nanofibers. Interestingly, cells cultured on aligned nanofibers displayed significantly greater ROCK expression relative to cells on a flat surface, as assessed by immunoblotting. To further test the role of ROCK, MSCs with ROCK small hairpin RNA (shRNA) were established. It is notable that, even when ROCK was stably knocked down via shRNA, cells could still display preferred orientation along aligned nanofibers. However, MSCs with shRNA-ROCK displayed a significantly decreased cell major axis length following aligned nanofibers compared with shRNA vector control, suggesting that ROCK may be involved in cell elongation on aligned nanofibers. Along with the reduction in cell length, cell area was decreased with ROCK silencing. These cell morphological changes induced by shRNA-ROCK were generally maintained on a flat surface and random nanofibers. A pharmacological ROCK inhibitor, Y-27632, produced results similar to those of shRNA-ROCK. The data on the role of ROCK in regulating cell alignment on nanofibers may provide a new mechanistic insight into nanofiber control of cells.
KW - Cell alignment
KW - Mesenchymal stem cells
KW - Nanofibers
KW - RhoA kinase
KW - Small hairpin RNA
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U2 - 10.1016/j.actbio.2013.04.013
DO - 10.1016/j.actbio.2013.04.013
M3 - Article
C2 - 23587628
AN - SCOPUS:84891715859
SN - 1742-7061
VL - 9
SP - 7737
EP - 7745
JO - Acta Biomaterialia
JF - Acta Biomaterialia
IS - 8
ER -