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Effects of ROCK inhibitor Y‐27632 on cell fusion through a microslit
Author(s) -
Wada KenIchi,
Hosokawa Kazuo,
Ito Yoshihiro,
Maeda Mizuo
Publication year - 2015
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.25641
Subject(s) - chemistry
ABSTRACT We previously reported a direct cytoplasmic transfer method using a microfluidic device, in which cell fusion was induced through a microslit (slit‐through‐fusion) by the Sendai virus envelope (HVJ‐E) to prevent nuclear mixing. However, the method was impractical due to low efficiency of slit‐through‐fusion formation and insufficient prevention of nuclear mixing. The purpose of this study was to establish an efficient method for inducing slit‐through‐fusion without nuclear mixing. We hypothesized that modulation of cytoskeletal component can decrease nuclear migration through the microslit considering its functions. Here we report that supplementation with Y‐27632, a specific ROCK inhibitor, significantly enhances cell fusion induction and prevention of nuclear mixing. Supplementation with Y‐27632 increased the formation of slit‐through‐fusion efficiency by more than twofold. Disruption of F‐actin by Y‐27632 prevented nuclear migration between fused cells through the microslit. These two effects of Y‐27632 led to promotion of the slit‐through‐fusion without nuclear mixing with a 16.5‐fold higher frequency compared to our previous method (i.e., cell fusion induction by HVJ‐E without supplementation with Y‐27632). We also confirmed that mitochondria were successfully transferred to the fusion partner under conditions of Y‐27632 supplementation. These findings demonstrate the practicality of our cell fusion system in producing direct cytoplasmic transfer between live cells. Biotechnol. Bioeng. 2015;112: 2334–2342. © 2015 Wiley Periodicals, Inc.