Positive correlation between the generation of reactive oxygen species and activation/reactivation of transgene expression after hydrodynamic injections into mice.
OBJECTIVE
Hydrodynamic injection has been shown to reactivate silenced transgene expression in mouse liver. In this study, the roles of inflammatory cytokines and reactive oxygen species (ROS) in the reactivation were examined.


METHODS
Production of inflammatory cytokines and ROS by hydrodynamic injection of saline was examined in mice that had received a hydrodynamic injection of a plasmid expressing Gaussia luciferase. The level of reporter gene expression was used as an indicator of the reactivation. The involvement of cytokines and ROS was examined by depleting Kupffer cells or by pre-administration of antioxidants, respectively.


RESULTS
A hydrodynamic injection of saline induced a significant production of interleukin (IL)-6. Depleting Kupffer cells using clodronate liposomes markedly reduced the IL-6 production but had no significant effect on the transgene expression. On the other hand, an injection of catalase or N-acetylcysteine significantly inhibited the hydrodynamic injection-induced reactivation of silenced transgene expression. The silenced expression was also reactivated by carbon tetrachloride, an inducer of oxidative stress in the liver, in a dose-dependent manner, and this reactivation was significantly inhibited by catalase.


CONCLUSIONS
These findings show a positive correlation between the generation of ROS and the reactivation of silenced transgene expression after hydrodynamic injections.
Department of Biopharmaceutics and Drug Metabolism, Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto, 606-8501, Japan.
Naomi
N
Takiguchi
Yuki
Y
Takahashi
Makiya
M
Nishikawa
Yuriko
Y
Matsui
Yasushi
Y
Fukuhara
Daihi
D
Oushiki
Kazuki
K
Kiyose
Kenjiro
K
Hanaoka
Tetsuo
T
Nagano
Yoshinobu
Y
Takakura
D000818:Animals; D002251:Carbon Tetrachloride / Q000494:pharmacology; D004002:Clodronic Acid / Q000494:pharmacology; D004247:DNA / Q000008:administration & dosage / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression / Q000187:drug effects; D020868:Gene Silencing / Q000187:drug effects*; D018014:Gene Transfer Techniques; D057446:Hydrodynamics; D007275:Injections, Intravenous; D015850:Interleukin-6 / Q000097:blood / Q000276:immunology; D007552:Isotonic Solutions / Q000008:administration & dosage / Q000494:pharmacology; D007728:Kupffer Cells / Q000187:drug effects / Q000473:pathology; D008081:Liposomes; D008099:Liver / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D008156:Luciferases / Q000235:genetics; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism*; D019076:Transgenes; D014409:Tumor Necrosis Factor-alpha / Q000097:blood / Q000276:immunology
D015850:Interleukin-6; D007552:Isotonic Solutions; D008081:Liposomes; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D004002:Clodronic Acid; D004247:DNA; D002251:Carbon Tetrachloride; D008156:Luciferases