title: Polyplex exposure inhibits cell cycle, increases inflammatory response, and can cause protein expression without cell division.
abstract: We sought to evaluate the relationship between cell division and protein expression when using commercial poly(ethylenimine) (PEI)-based polyplexes. The membrane dye PKH26 was used to assess cell division, and cyan fluorescent protein (CFP) was used to monitor protein expression. When analyzed at the whole population level, a greater number of cells divided than expressed protein, regardless of the level of protein expression observed, giving apparent consistency with the hypothesis that protein expression requires cells to pass through mitosis in order for the transgene to overcome the nuclear membrane. However, when the polyplex-exposed population was evaluated for the amount of division in the protein-expressing subpopulation, it was observed that substantial amounts of expression had occurred in the absence of division. Indeed, in HeLa S3 cells, this represented the majority of expressing cells. Of interest, the doubling time for both cell lines was slowed by ~2-fold upon exposure to polyplexes. This change was not altered by the origin of the plasmid DNA (pDNA) transgene promoter (cytomegalovirus (CMV) or elongation factor-1 alpha (EF1α)). Gene expression arrays in polyplex-exposed HeLa S3 cells showed upregulation of cell cycle arrest genes and downregulation of genes related to mitosis. Chemokine, interleukin, and toll-like receptor genes were also upregulated, suggesting activation of proinflammatory pathways. In summary, we find evidence that a cell division-independent expression pathway exists, and that polyplex exposure slows cell division and increases inflammatory response.
authors: Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors: Rebecca L
authors: RL
authors: Matz
authors: Blake
authors: B
authors: Erickson
authors: Sriram
authors: S
authors: Vaidyanathan
authors: Jolanta F
authors: JF
authors: Kukowska-Latallo
authors: James R
authors: JR
authors: Baker
authors: Bradford G
authors: BG
authors: Orr
authors: Mark M
authors: MM
authors: Banaszak Holl
mesh_terms: D000818:Animals; D019556:COS Cells; D002453:Cell Cycle / Q000187:drug effects*; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D015870:Gene Expression; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007249:Inflammation; D008938:Mitosis; D009685:Nuclear Envelope / Q000378:metabolism; D020648:Peptide Elongation Factor 1 / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011094:Polyethyleneimine / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D013997:Time Factors; D019076:Transgenes
chemical_list: C485182:Cyan Fluorescent Protein; D020648:Peptide Elongation Factor 1; D049452:Green Fluorescent Proteins; D011094:Polyethyleneimine