TLR9-dependent systemic interferon-beta production by intravenous injection of plasmid DNA/cationic liposome complex in mice.
BACKGROUND
The type I interferon (IFN) response to DNA/cationic liposome complex, or lipoplex, has been reported in cultured cells, but little is known about the response in vivo. Studies of the pro-inflammatory cytokine response to lipoplex have shown the importance of the unmethylated CpG dinucleotide (CpG motif) and its receptor, Toll-like receptor (TLR)-9.


METHODS
CpG- and non-CpG lipoplex consisting of CpG- or non-CpG plasmid DNA, respectively, and N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride/cholesterol liposomes were intravenously injected into mice. IFN-beta and interleukin (IL)-6 in the serum and organs were measured by the enzyme-linked immunosorbent assay. The involvement of TLR9, phagocytic cells and the spleen in the responses was evaluated using TLR9(-/-), clodronate liposome-treated-, and splenectomized mice, respectively. Accumulation of blood cells in the lung was evaluated histologically.


RESULTS
CpG lipoplex induced a large increase in the levels of IFN-beta and IL-6 in the serum, liver, spleen, lung and kidney, whereas non-CpG lipoplex scarcely had any effect. Neither formulation led to significant cytokine production in TLR9(-/-) mice. Clodronate liposome-treated mice showed a large reduction in both IFN-beta and IL-6 levels. Splenectomized mice receiving CpG lipoplex also showed a significantly low production of IL-6 but a similar level of IFN-beta production to that of unsplenectomized mice. A large number of monocytes were found in the capillary vessels around the pulmonary alveoli of mice receiving lipoplex.


CONCLUSIONS
These findings indicate that, in contrast to the production of IL-6 from splenic macrophages, IFN-beta is produced from phagocytic cells other than splenic macrophages after the injection of CpG lipoplex through the TLR9-dependent pathway.
Yoshida
Hiroyuki
H
Department of Biopharmaceutics and Drug Metabolism, Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto, Japan.
Nishikawa
Makiya
M
Yasuda
Sachiyo
S
Mizuno
Yumiko
Y
Toyota
Hiroyasu
H
Kiyota
Tsuyoshi
T
Takahashi
Rei
R
Takakura
Yoshinobu
Y
D000818:Animals; D002412:Cations; D002469:Cell Separation; D004002:Clodronic Acid / Q000494:pharmacology; D018899:CpG Islands / Q000235:genetics; D004247:DNA / Q000008:administration & dosage*; D007275:Injections, Intravenous; D016899:Interferon-beta / Q000096:biosynthesis* / Q000097:blood; D015850:Interleukin-6 / Q000096:biosynthesis / Q000097:blood; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects; D008081:Liposomes; D008168:Lung / Q000098:blood supply / Q000166:cytology / Q000187:drug effects; D051379:Mice; D008813:Mice, Inbred ICR; D010957:Plasmids / Q000008:administration & dosage*; D013156:Splenectomy; D051217:Toll-Like Receptor 9 / Q000378:metabolism*
D002412:Cations; D015850:Interleukin-6; D008081:Liposomes; D051217:Toll-Like Receptor 9; D004002:Clodronic Acid; D016899:Interferon-beta; D004247:DNA