Intracytoplasmic gene delivery for in vitro transfection with cytoskeleton-specific immunoliposomes.
A novel and highly efficient method of in vitro gene transfection has been developed. This method employs direct intracytoplasmic gene delivery into embryonic cardiocytes using neutral cytoskeletal-antigen specific immunoliposomes (CSIL). These immunoliposomes target cardiocytes specifically under reversible hypoxic conditions. Two independent reporter genes, pGL2 and pSV-beta-galactosidase, were used to verify CSIL-transfection (CSIL-fection). The efficiency of CSIL-fection with firefly luciferase pGL2 vector was 30+ times greater than controls consisting of hypoxic cardiocytes treated with plain liposomes (PL) or normoxic cardiocytes treated with CSIL, PL or naked DNA. CSIL-fection was also compared to cationic liposome transfection. Net cationic liposome transfection appeared to be more efficient than CSIL-fection for pGL2 vectors. However, a smaller number of viable cells was observed in the cationic liposome treated cultures than in the CSIL treated cultures. Therefore, to determine whether more cells were transfected with cationic liposomes or CSIL, pSV-beta-galactosidase vector was used. CSIL-fection with pSV-beta-galactosidase vector produced at least 40 times more transfected cells than those transfected with cationic liposomes. No transfection with pSV-beta-galactosidase vectors was obtained with IgG-liposome, PL or naked DNA treatments. Targeted enhanced efficiency of transfection by this novel method could have practical therapeutic applications in the genetic modification of cells ex vivo that could then be reimplanted into patients for gene therapy.
Khaw
B A
BA
Center for Cardiovascular Targeting, Department of Pharmaceutical Sciences, Bouve College of Health Sciences, Northeastern University, Boston, MA 02115, USA. b.khaw@nunet.neu.edu
daSilva
J
J
Vural
I
I
Narula
J
J
Torchilin
V P
VP
D000818:Animals; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism*; D004247:DNA / Q000008:administration & dosage / Q000032:analysis / Q000493:pharmacokinetics; D004337:Drug Carriers; D005079:Excipients; D018390:Gene Targeting / Q000379:methods*; D017930:Genes, Reporter; D008081:Liposomes; D008156:Luciferases / Q000235:genetics; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D009218:Myosins / Q000276:immunology; D010316:Particle Size; D051381:Rats; D014162:Transfection / Q000379:methods*; D001616:beta-Galactosidase / Q000235:genetics
D004337:Drug Carriers; D005079:Excipients; D008081:Liposomes; D004247:DNA; D008156:Luciferases; D001616:beta-Galactosidase; D009218:Myosins