DNA transfection of macaque and murine respiratory tissue is greatly enhanced by use of a nuclease inhibitor

Abstract

This work provides evidence that using direct nuclease inhibitors will enhance lung transfection and that nuclease activity is present in all lung fluids tested, which can be inhibited by the use of direct DNase inhibitors.

Improved chitosan-mediated gene delivery based on easily dissociated chitosan polyplexes of highly defined chitosan oligomers

“…The impact of protecting DNA against degradation by extracellular DNase present in the lung fluid, on the efficiency of gene expression after intratracheal administration, was recently found to increase the efficiency of naked DNA 50-to 80-fold. 36,37 We therefore studied the stability of DPn18 and DP15-21 polyplexes against DNase at a DNase concentration (1.0 U) corresponding to those found in the mouse lung (o0.6 U). 36 Indeed, our finding that the luciferase gene expression in mouse lung increased with increasing stability of polyplexes against degradation by DNase (Figures 2a and 8a), confirmed that DNA degradation by DNase is a major barrier to efficient in vivo gene expression in the lung, and indicated that chitosan oligomers provided complete protection against this degradation at the optimal charge ratio 60:1 (+/À).…”

Characterization of complexation of PVP copolymer with DNA

“…It has been reported that intracellular degradation of the DNA significantly limits the gene transfer efficiency. [33] Several studies on non-viral gene transfer were performed with PVP which can encapsulate DNA and protect it from intracellular degradation, [34] and with naked plasmid [35,36] which are known to be less susceptible to intracellular degradation. Thus, the PVP copolymer P(NVP-co-TrpAMT) as a gene carrier has a potential application for gene delivery.…”

Uptake of Foreign Nucleic Acids in Kidney Tubular Epithelial Cells Deficient in Proapoptotic Endonucleases

“…E xtracellular DNA uptake occurs during normal and cancer tissue growth (Bergsmedh et al, 2006;Yan et al, 2006) and during viral and bacterial infections (Chu et al, 2006;Metifiot et al, 2007), and is routinely used in genetic manipulations and experimental animals (Tanswell et al, 1998;Glasspool-Malone et al, 2002;Freitas et al, 2007). The entry of foreign DNA (fDNA) is harmful to the host cell (Li et al, 1999), causing DNA-dependent cell death, whereas DNase treatment before transfection prevented cell death (Stacey et al, 1993).…”

“…The uptake of DNA is restricted by a number of cell defense enzymes, the core of which consists of DNA endonucleases (Tanswell et al, 1998;Glasspool-Malone et al, 2002). Despite attempts to protect the fDNA by modifications, lipid or viral packaging, increased rate of DNA delivery, or by precise targeting to a tissue, our knowledge about the protection against the cellular defense system remained insufficient (Tanswell et al, 1998;Glasspool-Malone et al, 2002;Freitas et al, 2007).…”

“…Despite attempts to protect the fDNA by modifications, lipid or viral packaging, increased rate of DNA delivery, or by precise targeting to a tissue, our knowledge about the protection against the cellular defense system remained insufficient (Tanswell et al, 1998;Glasspool-Malone et al, 2002;Freitas et al, 2007). Endocytosis of DNA normally leads to its lysosomal delivery, with DNases playing major role in degrading fDNA (Torchilin, 2006).…”