188. Transfer of DNA in Tissues Using Local Fast Injection

Abstract

Top of pageAbstract The simple injection of naked DNA into skeletal muscle is known to result in the expression of the injected gene even though at very low and variable levels. We report here that, with respect to the local (intratissular) slow injection of the DNA (in about 25 seconds), the local fast injection in 1 to 2 seconds results in an increase in gene expression from 10 times in tumors and 100 times in liver, up to 500 times in the skeletal muscle. We also show that the fast liquid injection affects cell membrane and we demonstrate that the main mechanism of the increase in DNA uptake and gene expression is the enhancement of the receptor-mediated endocytosis of the injected DNA. However, at the highest speed of injection, there is not a complete inhibition of this receptor-mediated endocytosis process by means of a huge excess of a competitive inhibitor (heparin), suggesting the existence of another minor process for DNA uptake. Using a classical marker of membrane permeability (propidium iodide) we show that this small amount of non heparin-inhibitable DNA uptake could result from the direct permeabilisation of the cells by the high speed/high pressure DNA injection. This hydroporation could be similar to the permeabilisation found in the hydrodynamics method based upon the fast intravascular DNA injection. Neither the hydroporation nor the increase in DNA receptor-mediated endocytosis induce histologically detectable lesions, and the level of cell stress in the muscle, as measured by Hsp70 gene transcription induction, is similar whatever the injection speed. Thus the fast intratissular injection of DNA is a safe, easy and quick way to achieve local gene expression using naked DNA.