Inflammatory cell-specific transgene expression system responding to Ikappa-B kinase beta activation.
BACKGROUND
Control of inflammation is essential for the clinical management of many common human diseases. However, there are few generally applicable strategies to convert an abnormal intracellular signal into a gene expression that leads to normalization of the intracellular environment. Recently, we proposed a novel strategy termed D-RECS (i.e. drug or gene delivery system responding to cellular signals) to convert an intracellular signal to transgene expression. In the present study, we applied this concept to inflammatory cells using Ikappa-B kinase as a signal molecule that triggers the gene expression.


METHODS
Candidate cationic substrates of Ikappa-B kinase (IKK)beta were synthesized and their reactivity was investigated. Then, polymers grafted with these peptides were prepared by radical polymerization. Polymer/DNA complexes (polyplexes) were prepared by mixing plasmid DNAs with the polymers. The behaviour of these polyplexes by adding IKKbeta was examined. Furthermore, changes of gene expression were evaluated after the microinjection of polyplex into living cells under conditions of nuclear factor (NF)-kappaB activation.


RESULTS
Synthetic peptides with additional lysine residues were well phosphorylated by IKKbeta. Both the polymer and the polyplex were also phosphorylated by IKKbeta. The results of gel shift assay showed that the polyplex was disintegrated and free DNA was released in the presence of IKKbeta. The polyplex comprising-green fluorescent protein plasmid DNA and the polymer expressed the transgene in living cells exposed to a pro-inflammatory stimulus.


CONCLUSIONS
Our concept of cell-specific gene expression was demonstrated to work in inflammatory cells. This method may provide a unique strategy for gene therapy exclusively in inflammatory cells.
Department of Microbiology, St Marianna University School of Medicine, Kawasaki, Japan. asai@marianna-u.ac.jp
Daisuke
D
Asai
Akira
A
Tsuchiya
Jeong-Hun
JH
Kang
Kenji
K
Kawamura
Jun
J
Oishi
Takeshi
T
Mori
Takuro
T
Niidome
Yoko
Y
Shoji
Hideki
H
Nakashima
Yoshiki
Y
Katayama
D000818:Animals; D004789:Enzyme Activation; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D006801:Humans; D051550:I-kappa B Kinase / Q000235:genetics / Q000378:metabolism*; D007249:Inflammation / Q000378:metabolism*; D051379:Mice; D015394:Molecular Structure; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D019076:Transgenes
D010455:Peptides; D051550:I-kappa B Kinase