Proceedings of The Physiological Society

Physiology 2016 (Dublin, Ireland) (2016) Proc Physiol Soc 37, PCB191

Poster Communications

Use of quantum dots as a potential method for delivery of growth factors to the developing placenta

M. Karolczak-Bayatti1, J. Horn1, L. Harris1, M. Westwood1, J. Aplin1

1. Institute of Human Development, Manchester University, Manchester, United Kingdom.

The defective development of the placenta plays a major role in miscarriage, intrauterine growth restriction and pre-eclampsia. In vivo animal studies have demonstrated the importance of insulin like growth factor I (IGF-I) for the development of the placenta and embryo. In humans, IGF-I levels in umbilical cord blood correlate with fetal weight at birth while in vitro, IGF-I regulates turnover of cytotrophoblasts, which continuously proliferate ensuring appropriate growth and functioning of the placenta. Therefore manipulation of the IGF-I axis might offer a therapeutic route to improve the performance of the placenta during pregnancy. Quantum Dots (QD), have been used to deliver a range of molecules such as growth factors, antibodies and drugs to various tissues. Using QD conjugated to IGF-I and a placental targeting peptide (iRGD), as a means of selectively delivering IGF-I to the placenta, we investigated the transcellular IGF-I route from the syncytial microvillus membrane (MVM) to the underlying cytotrophoblasts and analysed IGF-I turnover in placenta. First trimester placental explants were pre-incubated for 1h with the clathrin-dependent endocytosis inhibitor CPMZ (1h, 100uM) and subsequently treated with IGF-I (20nM), QD-biotinylated-IGF-I (QD-IGF; IGF 100nM) or iRGD-QD-IGF for 30min-24h. Immunohistochemistry (IHC) and Western blotting (WB) with antibodies to phosphorylated (p)IGFR, pAKT, p70S6K, pGSK, and the endosomal markers clathrin and EEA1, were used to investigate IGF-I-mediated signalling and turnover. All IGF conjugates induced phosphorylation of the IGFR after 30min (WB), with iRGD-QD-IGF triggering a higher response as compared to QD-IGF (WB, t-test p<0.05). IHC revealed two stages of IGF-I signalling: incubation with IGF-I for 5 min triggered phosphorylation of IGFR at MVM, followed by translocation of the p70S6K to the apical and basal membranes of the syncytium. Longer incubation (15-30min) generated exclusive activation of IGFR in the cytotrophoblasts, and was followed by phosphorylation of AKT and GSK. CPMZ inhibited IGF-I stimulated pAKT and p70S6K, but not pIGFR. Following internalisation, QD-IGF co-localised with clathrin but not the early endosomal marker EEA-1, and accumulated near the basal syncytial membrane, whereas iRGD-QD-IGF crossed this membrane and was internalised by cytotrophoblasts. QD-IGF-I is a useful tool to investigate the delivery, turnover and effects of IGF-I on placental tissues and might be considered as an effective delivery vehicle for growth factors to the placenta. Conjugates of IGF-I and QD are efficient in triggering an IGFR response in the syncytium and underlying cytotrophoblast, followed by the activation of signalling related to mitosis (pAKT and pGSK) and protein synthesis (p70S6K). However, the IGF-I transcytosis route from MVM to cytotrophoblast layer requires further investigation.

Where applicable, experiments conform with Society ethical requirements