Cationic lipid-mediated transfection of cells in culture requires mitotic activity

Abstract

Cationic lipid-based delivery systems such as lipoplexes or aphidicolin exhibited 20-fold lower reporter gene activity stabilized plasmid-lipid particles (SPLP) represent a safer than asynchronous control cells upon incubation with lipoalternative to viral systems for gene therapy applications.plexes. When cells arrested in the G1 phase were allowed We studied the impact of cell cycle status on the efficiency to proceed though the cell cycle in the presence of the lipoof transfection of human ovarian carcinoma tumor cells plex or SPLP, transgene expression was found to coincide using two cationic-lipid based delivery systems. Cells with the transition of cells from the G2/M phase into the arrested in the G1 phase of the cell cycle by treatment with G1 phase of the subsequent cell cycle. In addition, higher aphidicolin were compared with an asynchronous dividing levels of reporter gene expression were observed when the population of cells. Treatment of the cells with aphidicolin cells were incubated with lipoplexes or SPLP during, or just had no effect on the rate of internalization of the lipid forbefore, mitosis. These results suggest that it may be possmulated DNA or on the level of gene expression observible to augment cationic lipid-mediated transfection by able in stably transfected cells. However, cells treated with manipulating the cell cycle status of the target cells.

Transfection of large plasmids in primary human myoblasts

“…Since it is believed that the majority of the DNA enters the cell nucleus during mitosis, we synchronized the myoblasts with aphidicolin 27,28 to verify the effect of cell cycle synchronization on PEI adenofection. No significant enhancement of transfection efficiency was observed (Table 1).…”

“…51 This explains why rapidly dividing cells are more readily transfected. To try to enhance nuclear entry of plasmid DNA during mitosis, we synchronized the myoblasts with aphidicolin 27,28 but no enhancement of transfection was observed. Synchronizing the cells at mitosis when the nucleus is disrupted could perhaps be more efficient in enhancing transfections than synchronizing the cells in G1/S as we did.…”

Hypoxia-inducible transgene expression in differentiated human NT2N neurons – a cell culture model for gene therapy of postischemic neuronal loss

“…This finding is in line with the fact that postmitotic cells have a lower expression rate after lipoplex transfection than highly mitotic (tumor) cells. 14 The cationic liposomal preparations LipofectAmine and DOTAP were further tested for efficiency of cell transduction in the presence or absence of fetal calf serum (FCS) in the culture medium. NT2 cells and NT2N neurons were efficiently transfected with the pCMV-luc vector in medium without FCS (Optimem; Gibco BRL), while addition of 10% FCS (Sigma, Deisenhofen, Germany) drastically reduced the efficiency of transduction in both cell types.…”

“…Mortimer et al 14 showed that mitotic cells have increased transfectability compared with the low transfection rates in postmitotic cells. In agreement with these findings, our studies showed that the transgene expression in postmitotic NT2N neurons was four-fold lower than in undifferentiated NT2 cells.…”

Synthesis of a novel series of cationic lipids that can act as efficient gene delivery vehicles through systematic heterocyclic substitution of cholesterol derivatives

“…The positively charged head-group was generally believed to allow interactions between the cationic lipids, the negatively charged DNA, as well as cell membrane through charge/charge interactions. 39,40 The presence of nitrogen and oxygen atoms in the heterocyclic ring might further contribute to this charge/charge interaction of liposome and plasmid DNA and stabilize the binding between the cationic liposomes and DNA.…”