A two-stage poly(ethylenimine)-mediated cytotoxicity: implications for gene transfer/therapy.
Poly(ethylenimine) (PEI) is a cationic macromolecule commonly used in gene transfer/therapy protocols with high transfection efficiency both in vitro and in vivo. PEI is also cytotoxic, but the molecular basis of its cytotoxicity is poorly understood. Here, we have demonstrated that branched (25 kDa) and linear (750 kDa) PEI can both induce membrane damage and initiate apoptosis in three clinically relevant human cell lines (Jurkat T cells, umbilical vein endothelial cells, and THLE3 hepatocyte-like cells). We have defined Phase I toxicity as early necrotic-like changes (30 min) resulting from compromised membrane integrity, assessed by considerable lactate dehydrogenase release and phosphatidylserine translocation from the inner plasma membrane to the outer cell surface. Phase II cytotoxicity (24 h) was due to activation of a "mitochondrially mediated apoptotic program," resulting from PEI-induced channel formation in the outer mitochondrial membrane. This led to the release of proapoptotic cytochrome c, subsequent activation of caspase 3, and alteration in mitochondrial membrane potential as a result of caspase translocation into the mitochondria. The reported observations have important implications for the design and execution of gene therapy protocols as well for controlling intracellular distribution of drugs with cationic-based polymer-delivery systems.
Moghimi
S Moein
SM
Molecular Targeting and Polymer Toxicology Group, School of Pharmacy, University of Brighton, Brighton BN2 4GJ, UK. s.m.moghimi@brighton.ac.uk
Symonds
Peter
P
Murray
J Clifford
JC
Hunter
A Christy
AC
Debska
Grazyna
G
Szewczyk
Adam
A
D017209:Apoptosis; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects; D011094:Polyethyleneimine / Q000633:toxicity*; D021381:Protein Transport
D011094:Polyethyleneimine; D045304:Cytochromes c; D007770:L-Lactate Dehydrogenase; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases