Studies Of Preparation And Targeting Effects On Liver Of Lactosaminated Nanoparticles Loaded With Exogenous Gene

Objective To establish and identify ASGP receptor- mediated and positive liver targeting nanoparticles-gene model with alginate hydroxyapatite-poly-L-lysine nanoparticles as the carrier; To compare the effects lactosaminated ones loaded with exogenous gene on liver targeting via peripheral intravenous route.Methods Preparation of alginate-poly-L-lysine based on control of "gelification phenomenon " of alginate by calcium ions and preparation of hydroxyopatite with collosol-gel method, both nanoparticles further modified with lactosaminated-poly-L-lysine which were synthesized by reductive lactosamination .With the enhanced green fluorescent protein(pEGFP-C1) as the reporter gene, we establish a receptor-mediated and active liver targeting nanoparticles-gene model. The potential of adsorbing DNA on nanoparticles was analyzed by electrophoresis and spectrophotometer. Then different complexes were transferred into the rats body by peripheral intravenous route and the liver targeting and other organs distribution characteristics were investigated using radioisotope tracing assay.Results1 HAP nanoparticle presented as needle -like particle with a diameter of 20nm by TEM, and can be effectively combined with PLL through modifying potential resulting in no apparent increase in diameter. Alginate-PLL nanoparticle ,spherical is 250nm in diameter. The optional preparation condition: 18mmol/l CaCl2 0.06% alginate and first adding CaCl2 were chosen .2 Agarose gel electrophoresis showed both nanoparticles could effectively combine with DNA and the optimal proportion of PLL-PCHNP and DNA is 30: 1 (w/w); DNA mixed ratio of AlgPLL was 68.3?.1% by spectrophotometer.3 According to the liver uptake curve, the radioactivities in liver for galactosylated HAPNP was higher than relativeone(t3=3.033,P<0.05)with apparent stastical difference. The radioactivities for HAP-PLLNP reached to the peaks at 30 min after injection, declined thereafter .While galactosylated HAP-PLLNP gradually increase and persist in high level, decline after 45 min and continue to persist in low level. The radioactivities in liver for galactosylated AlgLacPLLNP was slightly higher than relative one(t3=1.054,P>0.05) with no apparent stastical difference. From biodistribution at 30min post injection, AlgPLLNP , the first targeting organ of AlgPLLNP was lung, while for AlgLacPLLNP was liver. Leastsignificant difference (LSD) show no difference between lung and liver. Conclusions1. It was conformed that the properties of AlgNP depended on the concentration of Alg and CaCl2 and the sequence of adding CaCl2 or PLL to solution alginate. 18mmol/l CaCl2 0.06% alginate and first adding CaCl2 were chosen to prepare AlgNP in this study. Hydroxyapatitate PLL modification relates toits potential.2. Alg-LacPLL nanosponge are sponge-like nanoparticle which encapsulates DNA within its aqueous core. HAP-LacPLLNP , complexing nanoparticle , are coated with cationic polymers which attracts DNA by electrostatic charges.3. Lactosaminated nanoparticles can be effectively targeted to the liver by peripheral intravenous route ,and better than nonlactosaminated ones. Lactosaminated hydroxyapatite-poly-L-lysine nanoparticles has the the potential to be gene carrier targeting to liver.