Cationic glycopolymers for the delivery of pDNA to human dermal fibroblasts and rat mesenchymal stem cells

Abstract

Progenitor and pluripotent cell types offer promise as regenerative therapies but transfecting these sensitive cells has proven difficult. Herein, a series of linear trehalose-oligoethyleneamine “click” copolymers were synthesized and examined for their ability to deliver plasmid DNA (pDNA) to two progenitor cell types, human dermal fibroblasts (HDFn) and rat mesenchymal stem cells (RMSC). Seven polymer vehicle analogs were synthesized in which three parameters were systematically varied: the number of secondary amines (4–6) within the polymer repeat unit (Tr433, Tr530, and Tr632), the end group functionalities [PEG (Tr4128PEG-a, Tr4118PEG-b), triphenyl (Tr4107-c), or azido (Tr499-d)], and the molecular weight (degree of polymerization of about 30 or about 100) and the biological efficacy of these vehicles was compared to three controls: Lipofectamine 2000, JetPEI, and Glycofect. The trehalose polymers were all able to bind and compact pDNA polyplexs, and promote pDNA uptake and gene expression [luciferase and enhanced green fluorescent protein (EGFP)] with these primary cell types and the results varied significantly depending on the polymer structure. Interestingly, in both cell types, Tr433 and Tr530 yielded the highest luciferase gene expression. However, when comparing the number of cells transfected with a reporter plasmid encoding enhanced green fluorescent protein, Tr433 and Tr4107-c yielded the highest number of HDFn cells positive for EGFP. Interestingly, with RMSC, all of the higher molecular weight analogs (Tr4128PEG-a, Tr4118PEG-b, Tr4107-c, Tr499-d) yielded high percentages of cells positive for EGFP (30–40%).

Cationic, helical polypeptide-based gene delivery for IMR-90 fibroblasts and human embryonic stem cells

“…31, 32, 34 The ring-opening polymerization of VB-Glu-NCA with PEG-amine as the macroinitiator yielded PEG- block -poly(γ-(4-vinylbenzyl)-L-glutamate) (PEG- b -PVBLG) with controlled molecular weights (MWs) and narrow molecular-weight distributions (Scheme 2). At the VB-Glu-NCA/PEG-amine ratio of 100, the obtained M n of 20.7 × 10 3 g·mol -1 agreed well with the theoretical M n of 24.6 × 10 3 g·mol -1 and had a narrow molecular weight distribution of 1.21 (entry 1, Table 1). Two PEG- b -PVBLG copolymers were prepared with degrees of polymerization (DP) of 76 (PEG- b -PVBLG 76 ) and 287 (PEG- b -PVBLG 287 ) of the PVBLG block (Table 1).…”

Exploring the Mechanism of Plasmid DNA Nuclear Internalization with Polymer-Based Vehicles

“…45 Additionally, it would seem that the carbohydrate residues in G4 allow it to transfect primary and mesenchymal stem cells to a greater extent than JetPEI™. 46 Taken together, the data from these prior experiments as well as the current data illustrate how slight changes in polymer structure (in this case, the number of hydroxyl groups per repeat unit) can impact their pDNA delivery mechanisms.…”

Spinal Cannabinoid Receptor Type 2 Agonist Reduces Mechanical Allodynia and Induces Mitogen-Activated Protein Kinase Phosphatases in a Rat Model of Neuropathic Pain

“…it may induce antinociception in vivo. 23 However, we have demonstrated that triptolide alone does not modify MKP-1, MKP-3, or MAPKs in lipopolysaccharide-stimulated microglial cells. 31 Moreover, our laboratory and others have shown that triptolide effectively blocks MKP-1 and MKP-3 when the expression of these MKPs is enhanced, and consequently blocks pharmacological anti-inflammatory effects in vitro.…”