NKX2-5 is expressed in the heart throughout life. We targeted eGFP sequences to the NKX2-5 locus of human embryonic stem cells (hESCs); NKX2-5(eGFP/w) hESCs facilitate quantification of cardiac differentiation, purification of hESC-derived committed cardiac progenitor cells (hESC-CPCs) and cardiomyocytes (hESC-CMs) and the standardization of differentiation protocols. We used NKX2-5 eGFP(+) cells to identify VCAM1 and SIRPA as cell-surface markers expressed in cardiac lineages.
The role of microglia cells in Alzheimer’s disease (AD) is well recognized, however their molecular and functional diversity remain unclear. Here, we isolated amyloid plaque-containing (using labelling with methoxy-XO4, XO4+) and non-containing (XO4−) microglia from an AD mouse model. Transcriptomics analysis identified different transcriptional trajectories in ageing and AD mice. XO4+ microglial transcriptomes demonstrated dysregulated expression of genes associated with late onset AD. We further showed that the transcriptional program associated with XO4+ microglia from mice is present in a subset of human microglia isolated from brains of individuals with AD. XO4− microglia displayed transcriptional signatures associated with accelerated ageing and contained more intracellular post-synaptic material than XO4+ microglia, despite reduced active synaptosome phagocytosis. We identified HIF1α as potentially regulating synaptosome phagocytosis in vitro using primary human microglia, and BV2 mouse microglial cells. Together, these findings provide insight into molecular mechanisms underpinning the functional diversity of microglia in AD.
Embryonic stem cells (ESCs) will become a source of models for a wide range of adult differentiated cells, providing that reliable protocols for directed differentiation can be established. Stem-cell technology has the potential to revolutionize drug discovery, making models available for primary screens, secondary pharmacology, safety pharmacology, metabolic profiling and toxicity evaluation. Models of differentiated cells that are derived from mouse ESCs are already in use in drug discovery, and are beginning to find uses in high-throughput screens. Before analogous human models can be obtained in adequate numbers, reliable methods for the expansion of human ESC cultures will be needed. For applications in drug discovery, involving either species, protocols for directed differentiation will need to be robust and affordable. Here, we explore current challenges and future opportunities in relation to the use of stem-cell technology in drug discovery, and address the use of both mouse and human models.
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