Oral Presentation 6th Annual Meeting for Australasian Society for Stem Cell Research 2013

Using embryonic stem cells to provide human neurons for drug screening (#7)

Ana Antonic 1 , Mirella Dottori 2 , Jessie Leung 2 , Geoffrey Donnan 1 , David Howells 1
  1. Florey Neuroscience Institutes, Heidelberg, Vic, Australia
  2. University of Melboure, Melbourne, Vic, Australia

Despite most neuroprotectants working in animal models of stroke, none have been shown to be successful in the clinic. While stem cell based therapies could be of benefit, an alternative use of stem cells is to create an in vitro screening system in which human embryonic stem cells (hESCs) differentiated into neurons are used to test candidate drugs. Aims: to differentiate hESC lines into neurons, develop a model of ischaemic injury and test potential therapeutic agents. Induction of neuronal differentiation of hESCs was achieved by using the bone morphogenic inhibitor protein, Noggin. The resulting progenitors were then grown in the presence of EGF and bFGF until they formed neurospheres. Removal of the growth factors allowed the neurospheres to differentiate into neurons which were then cultured for 11 days prior to injury. Two methods of injury were used: (i) oxygen-glucose deprivation (OGD) and (ii) H2O2 induced oxidative stress. Four therapies (hypothermia to 33°C, melatonin at the concentration range of 10nM to 1mM, VAS2870 at 10µM to 50µM and NXY-059 at 1µM to1mM) were tested and cell death was quantified using LDH assay.Hypothermia reduced H2O2 and OGD induced cell death by 53% and 45% respectively at 24 hours. The neuroprotective effect of hypothermia diminished with time however, it was neuroprotective even when administered six hours after H2O2 induction. 100µM Melatonin and 50µVAS2870 showed the highest neuroprotective effect and both drugs reduce H2O2 and OGD injury by >60% at 24 hours. The neuroprotective effect of both melatonin and VAS2870 diminished with time and they had no effect on cell death when administered six hours after injury. NXY-059 provided no effect on neuronal cell survival in any of the injury models.This study provides the first investigation of protection of hESC derived neurons by four different potential therapeutic agents. These results suggest that hESCs have a potential to be differentiated into different populations of neurons, and could provide a useful drug screening tool.