Cellular uptake and transfection activity of DNA complexes based on poly(ethylene glycol)-poly(l-glutamine) copolymer with PAMAM G2

Abstract

Poly(ethylene glycol)-poly(L-glutamine) (PEG-PLGA) copolymer EA-G2 (or EA-G1) was prepared by aminolysis of poly(ethylene glycol)-poly(L-benzyl glutamate) (PEG-PBLG) using PAMAM G2 (or G1). The chemical structure of PEG-PLGA was confirmed by FT-IR, 1 H-NMR, DSC and GPC. The performances of the EA-G2 (or EA-G1) were assayed by enzyme degradation, MTT method and agarose gel electrophoresis. The particle size, zeta potential and morphology of EA-G2 (or EA-G1)/pDNA complexes were inspected by DLS and AFM. The cellular uptake mechanism was evaluated by endocytic inhibiting test, cell uptake test and observation of CLSM. The transfection activity was measured by flow cytometry. The EA-G2 (or EA-G1) exhibited good biodegradability, low cytotoxicity and great ability to combine with pDNA. The EA-G2 (or EA-G1) complexes exhibited particle sizes in the range 120-180 nm and zeta potentials in the range 20-40 mV, which were suitable for cell uptake. The cellular uptake of the EA-G2 complexes occurred mainly through clathrin-dependent and caveolin-mediated endocytosis, and at 6 h in 10% FBS and in serum-free media, the percentages of complex uptake reached 89.0% or 72.7%, respectively. EA-G2 complexes could efficiently mediate pEGFP-Cl into the cell nuclei. EA-G2 complexes displayed enhancing transfection efficiency and better serum tolerance. The results suggest that the EA-G2 has potential to be used as a biodegradable, efficient and serum-resistant gene vector.

Magnetic ternary nanohybrids for nonviral gene delivery of stem cells and applications on cancer therapy

“…Interestingly, the transfection efficiency of PAE MTN was unexpectedly enhanced by adding chlorpromazine, a clathrin-mediated endocytosis inhibitor. Blocking the clathrin-mediated endocytosis pathway might increase the probability of transporting PAE MTN through caveolea-mediated endocytosis and macropinocytosis pathways 45, 46 and leads to enhanced transfection efficiency. Enhancement of cell internalization of PAE MTN due to the magnetic field was also visualized using TOTO3-labeled DNA.…”

Synthetic <I>In Vitro</I> Delivery Systems for Plasmid DNA in Eukaryotes

“…120 Recently, Sarkar et al, 121 conjugated N -maleyl chitosan (NMCTS) by Michael type addition reaction to improve solubility, transfection efficiency and low the toxicity (Table III). Pan et al 122 reported as well high transfection activity with moderate toxicity of a copolymer made of poly(ethylene glycol)-poly-(L-glutamine) mixed with high molecular weight PAMAMs (Table III).…”

Star‐Shaped Polypeptides: Synthesis and Opportunities for Delivery of Therapeutics

“…Feng and co‐workers have examined the transfection efficacy and cellular uptake of a PEG‐PGA copolypeptide grafted to two generations of PAMAM dendrimer . Using conventional ROP techniques, the polypeptide was prepared via PEG‐NH 2 initiation of the BLG NCA monomer.…”