Inducing neuroprotective responses in astrocytes using human stem cell models

Amyotrophic lateral sclerosis (ALS) is a debilitating neurodegenerative disorder characterised by progressive loss of motor neurons in the spinal cord, brain stem and motor cortex. While research has predominantly focused on neuronal involvement in ALS, recent studies have explored non-cell autonomous pathomechanisms and highlighted the complex interplay between motor neurons and non-neuronal cells in the CNS, such as astrocytes. Under normal conditions, astrocytes play myriad supportive roles in maintaining brain homeostasis. In neurological diseases, such as ALS and ischaemic stroke, astrocytes undergo a phenotypic switch to become reactive, thus acquiring potential to exert both pro- and anti-inflammatory properties. These states have been coined A1 (neurotoxic) and A2 (neuroprotective) respectively, however astrocyte reactivity is more likely a heterogenous spectrum rather than two distinct states. While the A1 state has been characterised in vitro, an A2 state is yet to be functionally validated. Although hypoxia is thought to facilitate neurodegeneration, known markers of protective reactivity are expressed by astrocytes under moderate hypoxic conditions. Against this background, my PhD will first establish a human induced pluripotent stem cell model of (functionally validated) neuroprotective astrocytes using a range of established assays and some that I will develop myself. Where assays require developing, all preliminary work will be performed in HEK293s for experimental tractability. I will next address the molecular mechanisms by which human astrocytes mount neuroprotective responses by comparing the transcriptome and proteome of protective astrocytes with already established models of basal and toxic/deleterious astrocytes. I will finally systematically examine how neuroprotective mechanisms are disrupted in ALS. Crucially, the directed differentiation paradigm together with functional assays, co-culture assays and other essential 'know-how' is already established and routinely performed in my host lab, which mitigates risk and allows me to focus on my specific biological questions.