TY - JOUR
T1 - Application of infrared laser to the zebrafish vascular system:Gene induction, tracing, and ablation of single endothelial cells
AU - Kimura, Eiji
AU - Deguchi, Tomonori
AU - Kamei, Yasuhiro
AU - Shoji, Wataru
AU - Yuba, Shunsuke
AU - Hitomi, Jiro
PY - 2013/6
Y1 - 2013/6
N2 - Objective-Infrared laser-evoked gene operator is a new microscopic method optimized to heat cells in living organisms without causing photochemical damage. By combining the promoter system for the heat shock response, infrared laser- evoked gene operator enables laser-mediated gene induction in targeted cells. We applied this method to the vascular system in zebrafish embryos and demonstrated its usability to investigate mechanisms of vascular morphogenesis in vivo. Approach and Results-We used double-transgenic zebrafish with fli1:nEGFP to identify the endothelial cells, and with hsp:mCherry to carry out single-cell labeling. Optimizing the irradiation conditions, we finally succeeded in inducing the expression of the mCherry gene in single targeted endothelial cells, at a maximum efficiency rate of 60%. In addition, we indicated that this system could be used for laser ablation under certain conditions. To evaluate infrared laser-evoked gene operator, we applied this system to the endothelial cells of the first intersegmental arteries, and captured images of the connection between the vascular systems of the brain and spinal cord. Conclusions-Our results suggest that the infrared laser-evoked gene operator system will contribute to the elucidation of the mechanisms underlying vascular morphogenesis by controlling spatiotemporal gene activation in single endothelial cells, by labeling or deleting individual vessels in living embryos.
AB - Objective-Infrared laser-evoked gene operator is a new microscopic method optimized to heat cells in living organisms without causing photochemical damage. By combining the promoter system for the heat shock response, infrared laser- evoked gene operator enables laser-mediated gene induction in targeted cells. We applied this method to the vascular system in zebrafish embryos and demonstrated its usability to investigate mechanisms of vascular morphogenesis in vivo. Approach and Results-We used double-transgenic zebrafish with fli1:nEGFP to identify the endothelial cells, and with hsp:mCherry to carry out single-cell labeling. Optimizing the irradiation conditions, we finally succeeded in inducing the expression of the mCherry gene in single targeted endothelial cells, at a maximum efficiency rate of 60%. In addition, we indicated that this system could be used for laser ablation under certain conditions. To evaluate infrared laser-evoked gene operator, we applied this system to the endothelial cells of the first intersegmental arteries, and captured images of the connection between the vascular systems of the brain and spinal cord. Conclusions-Our results suggest that the infrared laser-evoked gene operator system will contribute to the elucidation of the mechanisms underlying vascular morphogenesis by controlling spatiotemporal gene activation in single endothelial cells, by labeling or deleting individual vessels in living embryos.
KW - Endothelial cells
KW - Gene expression
KW - Heat-shock response
KW - Infrared rays
KW - Morphogenesis
KW - Zebrafish
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UR - http://www.scopus.com/inward/citedby.url?scp=84879105395&partnerID=8YFLogxK
U2 - 10.1161/ATVBAHA.112.300602
DO - 10.1161/ATVBAHA.112.300602
M3 - Article
C2 - 23539214
AN - SCOPUS:84879105395
VL - 33
SP - 1264
EP - 1270
JO - Arteriosclerosis, Thrombosis, and Vascular Biology
JF - Arteriosclerosis, Thrombosis, and Vascular Biology
SN - 1079-5642
IS - 6
ER -