Abstract
Adeno-associated viral (AAV) vectors are characterised by low immunogenicity, although humoral and cellular responses may be triggered upon infection. Following systemic administration high levels of vector particles accumulate within the liver. Kupffer cells (KCs) are liver resident macrophages and an important part of the liver innate immune system. Decreased functional activity of KCs can contribute to exaggerated inflammatory response upon antigen exposure. Heme oxygenase-1 (HO-1) deficiency is associated with considerably reduced numbers of KCs. In this study we aimed to investigate the inflammatory responses in liver and to characterise two populations of hepatic macrophages in adult wild type (WT) and HO-1 knockout (KO) mice following systemic administration of one or two doses (separated by 3 months) of self-complementary (sc)AAV9 vectors. At steady state, the livers of HO-1 KO mice contained significantly higher numbers of monocyte-derived macrophages (MDMs), but significantly less KCs than their WT littermates. Three days after re-administration of scAAV9 we observed increased mRNA level of monocyte chemoattractant protein-1 (Mcp-1) in the livers of both WT and HO-1 KO mice, but the protein level and the macrophage infiltration were not affected. Three days after the 1st and 3 days after the 2nd vector dose the numbers of AAV genomes in the liver were comparable between both genotypes indicating similar transduction efficiency, but the percentage of transgene-expressing MDMs and KCs was higher in WT than in HO-1 KO mice. In the primary culture, KCs were able to internalize AAV9 particles without induction of TLR9-mediated immune responses, but no transgene expression was observed. In conclusion, in vivo and in vitro cultured KCs have different susceptibility to scAAV9 vectors. Regardless of the presence or absence of HO-1 and initial numbers of KCs in the liver, scAAV9 exhibits a low potential to stimulate inflammatory response at the analysed time points.
Highlights
Recombinant adeno-associated viral (AAV) vectors, derived from a replication-defective, nonpathogenic parvovirus with a small single-stranded DNA genome, have emerged as an attractive gene therapy tool due to safe, stable and long-term transgene expression
Higher expression of monocyte chemoattractant protein-1 (Mcp-1) was detected in Heme oxygenase-1 (HO-1) KO mice injected with one dose of scAAV9-green fluorescent protein (GFP) when compared to untreated control or scAAV9-empty vector (Fig 1A)
MCP-1 protein level was higher in the livers of heme oxygenase-1 knock-out (HO-1 KO) than of wild type (WT) mice (Fig 1C) and there was a slight, but statistically not significant increase in MCP-1 protein level in HO-1 KO mice 3 days after re-administration of either scAAV9-empty or scAAV9-GFP as compared to untreated HO-1 KO control (Fig 1C)
Summary
Recombinant adeno-associated viral (AAV) vectors, derived from a replication-defective, nonpathogenic parvovirus with a small single-stranded (ss) DNA genome, have emerged as an attractive gene therapy tool due to safe, stable and long-term transgene expression. Following cellular entry and translocation to the nucleus, ssAAV genome must be converted to a double-stranded (ds) form to enable transgene expression. This rate-limiting step in AAV transduction is omitted when the selfcomplementary (sc) AAV vectors, developed by the elimination of a nicking site in one of the inverted terminal repeats (ITRs) [2], are used. AAV vectors generated from several naturally occurring AAV serotypes have different cell tropism and kinetics of transgene expression (reviewed in: [1]) Their ability to efficiently transduce terminally differentiated or non-proliferating cells provides considerable, long-term in vivo gene transfer in various cell types e.g. muscle cells, hepatocytes, neurons, retinal cells, and many others (reviewed in: [1]). Systemic delivery of AAV9 results in liver transduction [5]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
