Enhanced transfection of primary cortical cultures using arginine-grafted PAMAM dendrimer, PAMAM-Arg.
PAMAM-Arg is a cationic arginine-grafted polyamidoamine (PAMAM) dendrimer. In the previous study, we reported that PAMAM-Arg facilitates transfection in a range of mammalian cell types. In the present study, we investigated the transfection efficiency of PAMAM-Arg in primary cortical cultures, which are known to be extremely vulnerable to exogenous gene transfection. PAMAM-Arg/DNA complexes showed particularly high transfection efficiencies and low cytotoxicity in primary cortical cells, as compared to other gene carriers such as, native PAMAM, polyethylenimine (BPEI), and Lipofectamine. Efficient transfection was not limited to neurons but extended to all three glial cells, astrocytes, microglia, and oligodendrocytes, present in these primary cortical cultures. The potential use of PAMAM-Arg was demonstrated by efficient gene knock-down by transfecting HMGB1 shRNA-expressing plasmid. The numbers of green fluorescent protein (GFP)-positive and HMGB1-negative cells indicated that PAMAM-Arg/shRNA-expressing plasmid complex suppressed target gene expression in over 40% of cells, which is the highest level achieved to date in primary cortical culture by any gene carrier. Here, we present evidence of the successful delivery and expression of both a reporter gene and of a shRNA-expressing plasmid in primary cortical cells, which demonstrates the potential of PAMAM-Arg for mediating gene delivery to primary neuronal cells.
Kim
Jung-Bin
JB
Department of Anatomy and Center for Advanced Medical Education (BK21), Inha University School of Medicine, 7-241 Shinheung-dong, Jung-Gu Inchon, 400-712, South Korea.
Choi
Joon Sig
JS
Nam
Kihoon
K
Lee
Minhyung
M
Park
Jong-Sang
JS
Lee
Ja-Kyeong
JK
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000378:metabolism*; D050091:Dendrimers; D015536:Down-Regulation / Q000235:genetics; D015870:Gene Expression / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D024243:HMGB1 Protein / Q000235:genetics; D008156:Luciferases / Q000235:genetics; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry* / Q000633:toxicity; D010957:Plasmids / Q000235:genetics; D011073:Polyamines / Q000737:chemistry* / Q000633:toxicity; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000235:genetics; D014162:Transfection / Q000379:methods*
D050091:Dendrimers; D024243:HMGB1 Protein; C104700:PAMAM Starburst; D010455:Peptides; D011073:Polyamines; D012330:RNA, Double-Stranded; D049452:Green Fluorescent Proteins; C015462:polyarginine; D008156:Luciferases