Receptor-mediated Transfection of Murine and Ovine Mammary Glandsin Vivo

Abstract

Transfection of HC-11 murine epithelial mammary cells as well as murine and sheep mammary glands were carried out using insulin-containing constructs that deliver DNA by receptor-mediated endocytosis to receptor-expressing cells. In vivo transfection of mammary gland tissue with the luciferase gene was carried out by introducing the DNA constructs into the mammary ducts of both mice and sheep. The successful transfection of ewe mammary glands was demonstrated by the detection of luciferase activity in mammary gland biopsy material up to a month after a single administration of the construct. To test whether products of expression of transfected genes could be secreted into the milk in this system, the N-terminal secretory signal sequences of bovine ␤-lactoglobulin or the entire coding sequence of human ␣-lactalbumin were fused to the N terminus of the luciferase gene. After transfection with the modified luciferases, both murine and sheep milk could be shown to contain luciferase activity, whereas mice, which had been transfected with the nonmodified luciferase gene, did not secrete any activity in the milk. This approach demonstrates for the first time the possibility of gene transfer in vivo into mammary gland epithelial cells using constructs delivering DNA via receptor-mediated endocytosis.

Targeted intracellular delivery of photosensitizers to enhance photodynamic efficiency

“…122 Adenoviruses have been used to increase the delivery of a variety of proteins into cells, 123,124 while the incorporation of adenovirus particles into gene transfer complexes has been shown to enhance gene delivery, through effecting the release of endocytosed DNA into the cytoplasm. 125 We have used attenuated Ad5 adenovirus strain dL312 (deleted for the tumourigenic E1A region) in combination with the most potent photosensitizing NLS-containing construct, P10-chlorin e6-insulin (see Fig. 2a), to demonstrate that the nuclear accumulation of the construct and its photodynamic efficiency in situ can be significantly increased (see Fig.…”

New Cationized LDL-DNA Complexes: Their Targeted Delivery to Fibroblasts in Culture

“…In many of the applications reported, peptides or proteins are conjugated to poly-L-lysine, which serves to bind and condense the exogenous DNA. Ligands used in this manner include asialoorosomucoid (Wu and Wu 1988), transferrin (Wagner et al 1990(Wagner et al , 1991Strydom et al 1993), antibodies (Trubetskoy et al 1992), insulin (Rosenkranz et al 1992;Sobolev et al 1998), and α 2 -macroglobulin (Schneider et al 1996). Low density lipoproteins (LDL), described as spheroidal microemulsions (Segrest et al 2001), have also been explored as gene delivery vehicles.…”

Gene transfer facilitated by a cellular targeting molecule, reovirus protein σ1

“…This DNA-ligand complex is internalized by targeted cells when the ligand binds to its respective cell-surface receptor. Such receptor-mediated gene transfer has been accomplished using a variety of receptors by conjugating DNA to ligands such as asialo-orosomucoid, 1,2 transferrin, [3][4][5] lectins, 6 folate, 7 lung surfactant protein, 8 insulin, 9 and monoclonal antibodies. 10,11 Receptor-mediated gene transfer has some advantages over the other methods of in vivo gene transfer.…”