1. ¹Center for Molecular and Vascular Biology, University of Leuven, Leuven, Belgium
2. ²EM unit Pathology, University of Maastricht, Maastricht, The Netherlands

Correspondence: Dr B De Geest, Center for Molecular and Vascular Biology, University of Leuven, Campus Gasthuisberg, Herestraat 49, 3000 Leuven, Belgium. E-mail: Bart.Degeest@med.kuleuven.be

³These authors contributed equally to this work.

Received 11 August 2006; Revised 5 November 2006; Accepted 6 November 2006; Published online 18 January 2007.

Abstract

Sinusoidal fenestrae may restrict the transport of gene transfer vectors according to their size. Using Vitrobot technology and cryo-electron microscopy, we show that the diameter of human adenoviral serotype 5 vectors is 93 nm with protruding fibers of 30 nm. Thus, a diameter of fenestrae of 150 nm or more is likely to be sufficient for passage of vectors from the sinusoidal lumen to the space of Disse and subsequent uptake of vectors in hepatocytes. The average diameter of fenestrae in New Zealand White rabbits (1031.3 nm) was 1.4-fold (P<0.0001) lower than in C57BL/6 mice (1415.4 nm). The percentage of sinusoidal fenestrae with a diameter larger than 150 nm was 10-fold (P<0.01) lower in rabbits (3.20.24%) than in C57BL/6 mice (325%), and this resulted in 8.8-fold (P=0.01) lower transgene DNA levels in hepatocytes in rabbits after adenoviral transfer. Injection of N-acetylcysteine combined with transient liver ischemia preceding intraportal transfer in rabbits increased the percentage of sinusoidal fenestrae above 150 nm 2.0-fold (P<0.001) and increased transgene DNA levels in hepatocytes 6.6-fold (P<0.05). In conclusion, species differences in transgene DNA uptake in hepatocytes after adenoviral transfer correlate with the diameter of fenestrae.