Genetic Manipulation of Liver Sinusoidal Endothelial Cells

摘要

The liver sinusoidal endothelial cells (SECs) possess unique hyaluronan receptors that recognize and internalize hyaluronic acid (HA). This characteristic was used in the development of a system for targeting foreign DNA to SECs. A gene carrier system was prepared by coupling HA (number-average molecular weight: 1.5 x 104) to poly-L-lysine (PLL, number-average molecular weight: 4.6 x 104) in a 1:1 weight ratio by reductive ami-nation reaction. The resulting copolymer (PLL-g-HA) was mixed with various amounts of DNA in 154mM NaCl at 4 deg C. Inter-polyelectrolyte complex formation between PLL-g-HA and DNA exhibited minimal self-aggregation, explaining the highly soluble nature of the complex. The agarose gel retardation assay revealed that the titration point representing the minimum proportion of PLL-g-HA required to retard the DNA completely occurred at a 1:1 copolymer to DNA charge ratio. Intravenous injection of the [32P]pSV p-Gal plasmid complexed to PLL-g-HA in Wistar rats demonstrated specific hepatic targeting with >93% of the injected counts taken up by the liver in 1 h. Further, using a fluorescein isothiocyanate-labeled DNA, it was shown that the PLL-g-HA/DNA complex was distributed exclusively in the SECs. Seventy-two hours after injection of 90 l g of pSV p-Gal in a PLL-g-HA-complexed form, a large number of SECs expressing ?-galactosidase were detected. Thus, the PLL-g-HA/DNA system permits targeted delivery of exogenous genes selectively to the liver SECs, providing a novel strategy for manipulation of SEC functions.