Novel protein transduction method by using 11R: an effective new drug delivery system for the treatment of cerebrovascular diseases.
OBJECTIVE
A motif of 11 consecutive arginines (11R) is reported to be one of the most effective protein transduction domains for introducing proteins into the cell membrane. We therefore examined the transduction efficiency of 11R in cerebral arteries.


METHODS
Basilar arteries (BAs) obtained from rats were incubated with either 11R-enhanced green fluorescent protein (11R-EGFP) or EGFP without 11R. After incubation, expression of 11R-EGFP or EGFP in BA serial sections was observed by fluorescence microscope. In an additional in vivo experiment, 11R-EGFP or EGFP was injected into the cisterna magna with or without subarachnoid hemorrhage. The 11R-EGFP or EGFP was injected just after the autologous blood injection, and then the expression of 11R-EGFP or EGFP in BA sections was also observed by fluorescence microscope.


RESULTS
The 11R-EGFP signal was much stronger than that of EGFP in all layers of the rat BA, in both in vivo and ex vivo experiments. Moreover, the 11R-EGFP was transduced into the BA immediately (2 hours after the injection). Interestingly, 11R-fused fluorescent protein was transduced especially into the tunica media of the BA.


CONCLUSIONS
The 11R-fused fluorescent protein effectively penetrates into all layers of the rat BA, especially into the tunica media. This is the first study to our knowledge to demonstrate the successful transduction of a protein transduction domain fused protein into the cerebral arteries.
Ogawa
Tomoyuki
T
Department of Neurological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Shikata-cho, Okayama, Japan. tom@md.okayama-u.ac.jp
Ono
Shigeki
S
Ichikawa
Tomotsugu
T
Arimitsu
Seiji
S
Onoda
Keisuke
K
Tokunaga
Koji
K
Sugiu
Kenji
K
Tomizawa
Kazuhito
K
Matsui
Hideki
H
Date
Isao
I
D000818:Animals; D001120:Arginine / Q000493:pharmacokinetics* / Q000627:therapeutic use; D001488:Basilar Artery / Q000187:drug effects / Q000378:metabolism; D016503:Drug Delivery Systems / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000493:pharmacokinetics; D008297:Male; D008856:Microscopy, Fluorescence; D010446:Peptide Fragments / Q000737:chemistry / Q000493:pharmacokinetics* / Q000627:therapeutic use; D017434:Protein Structure, Tertiary / Q000502:physiology; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013346:Subarachnoid Space / Q000187:drug effects / Q000378:metabolism
D010446:Peptide Fragments; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D001120:Arginine