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Chemical modifications on siRNAs avoid Toll-like-receptor-mediated activation of the hepatic immune system in vivo and in vitro.
- Source :
-
International immunology [Int Immunol] 2014 Jan; Vol. 26 (1), pp. 35-46. Date of Electronic Publication: 2013 Sep 24. - Publication Year :
- 2014
-
Abstract
- Objectives: The therapeutic application of small interfering RNAs (siRNAs) is limited by the induction of severe off-target effects, especially in the liver. Therefore, we assessed the potential of differently modified siRNAs to induce the hepatic innate immune system in vitro and in vivo.<br />Methods: Primary isolated liver cells were transfected with siRNAs against apolipoprotein B1 (APOB1), luciferase (LUC) or galactosidase (GAL). For in vivo use, siRNAs were formulated in lipid nanoparticles (LNPs) and administered intravenously to C57BL/6 mice. Liver tissue was collected 6-48 h after injection and knock-down efficiency or immune responses were determined by quantitative reverse-transcription-linked PCR.<br />Results: Unmodified GAL siRNA transiently induced the expression of TNF-α, IL-6, IL-10, IFN-β and IFN-sensitive gene 15 in vivo, whereas a formulation of 2'-O-methylated-LUC siRNA had no such effects. Formulation of unmodified APOB1-specific siRNA suppressed APOB1 mRNA levels by ~80% in the liver 48h after application. The results were paralleled in vitro, where transfection of liver cells with unmodified siRNAs, but not with chemically modified siRNAs, led to cell-type-specific induction of immune genes. These immune responses were not observed in MYD88-deficient mice or in chloroquine-treated cells in vitro.<br />Conclusions: Our data indicate that siRNAs activate endosomal Toll-like receptors in different liver-derived cell types to various degrees, in vitro. LNP-formulated siRNA selectively leads to hepatic knock-down of target genes in vivo. Here, off-target immune responses are restricted to non-parenchymal liver cells. However, 2'-O-methyl modifications of siRNA largely avoid immune-stimulatory effects, which is a crucial prerequisite for the development of safe and efficient RNA-interference-based therapeutics.
- Subjects :
- Animals
Immunity, Innate genetics
Interleukin-10 genetics
Interleukin-10 immunology
Interleukin-6 genetics
Interleukin-6 immunology
Liver
Mice
Mice, Inbred C57BL
RNA, Messenger genetics
RNA, Messenger immunology
RNA, Small Interfering genetics
Toll-Like Receptors genetics
Transfection methods
Tumor Necrosis Factor-alpha genetics
Tumor Necrosis Factor-alpha immunology
Hepatocytes immunology
Immune System immunology
Immunity, Innate immunology
RNA, Small Interfering chemistry
RNA, Small Interfering immunology
Toll-Like Receptors immunology
Subjects
Details
- Language :
- English
- ISSN :
- 1460-2377
- Volume :
- 26
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- International immunology
- Publication Type :
- Academic Journal
- Accession number :
- 24065781
- Full Text :
- https://doi.org/10.1093/intimm/dxt040