Research Repository

See what's under the surface

In-vitro evaluation of the effect of polymer structure on uptake of novel polymer-insulin polyelectrolyte complexes by human epithelial cells.

Ibie, Chidinma O.; Knott, Rachel M.; Thompson, Colin J.

Authors

Chidinma O. Ibie

Rachel M. Knott

Colin J. Thompson

Abstract

The biocompatibility and cellular uptake of polymer, insulin polyelectrolyte complexes (PECs) prepared using polyallylamine-based polymers was evaluated in-vitro using Caco-2 cell monolayers as a predictive model for human small intestinal epithelial cells. Poly(allyl amine) (PAA) and Quaternised PAA (QPAA) were thiolated using either carbodiimide mediated conjugation to N-acetylcysteine (NAC) or reaction with 2-iminothiolane hydrochloride yielding their NAC and 4-thiobutylamidine (TBA) conjugates, respectively. The effect of polymer quaternisation and/or thiolation on the IC50 of PAA was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay carried out on Caco-2 cells (with and without a 24 h recovery period after samples were removed). Uptake of PECs by Caco-2 cells was monitored by microscopy using fluorescein isothiocyanate (FITC) labelled insulin and rhodamine-labelled polymers at polymer:insulin ratios (4:5) after 0.5, 1, 2 and 4 h incubation in growth media ({plusmn}calcium) and following pre-incubation with insulin. MTT results indicated that quaternisation of PAA was associated with an improvement in IC50 values; cells treated with QPAA (0.001-4 mg mL?1) showed no signs of toxicity following a 24 h cell recovery period, while thiolation of QPAA resulted in a decrease in the IC50. Cellular uptake studies showed that within 2-4 h, QPAA and QPAA-TBA insulin PECs were taken up intracellularly, with PECs being localised within the perinuclear area of cells. Further investigation showed that uptake of PECs was unaffected when calcium-free media was used, while presaturating insulin receptors affected the uptake of QPAA, insulin PECs, but not QPAA-TBA PECs. The biocompatibility of PAA and uptake of insulin was improved by both thiol and quaternary substitution.

Journal Article Type Article
Publication Date Feb 1, 2015
Journal International journal of pharmaceutics
Print ISSN 0378-5173
Electronic ISSN 1873-3476
Publisher Elsevier
Peer Reviewed Peer Reviewed
Volume 479
Issue 1
Pages 103-117
Institution Citation IBIE, C., KNOTT, R. and THOMPSON, C.J. 2015. In-vitro evaluation of the effect of polymer structure on uptake of novel polymer-insulin polyelectrolyte complexes by human epithelial cells. International journal of pharmaceutics [online], 479(1), pages 103-117. Available from: https://doi.org/10.1016/j.ijpharm.2014.12.058
DOI https://doi.org/10.1016/j.ijpharm.2014.12.058
Keywords Fluorescence microscopy; Insulin; MTT assay; Polyelectrolyte complex; Thiolation; Quaternization

Files




Downloadable Citations