The challenges of accurately modelling diseases in vitro are great. A prime example is found in the central nervous system (CNS), where the complexities of the multicellular systems increase the difficulty in obtaining primary cells, especially those from patients. The use of stem-cell derived cultures is changing this, enabling neurons to be cultured from progenitor cells. More recently it has been possible to obtain human induced pluripotent stem cells (iPSC) derived from patients, effectively opening up a new source of patient cells. A number of publications have come out using these cells, and a recent double-publication from a group in Cambridge looking at Alzheimer’s disease illustrate not only the technical process, but how well the disease state can be modelled using this technology.
The first, published in Nature Neuroscience (2), describes the multi-step differentiation of human cerebral cortical neurons from pluripotent stem cells. The authors describe how they developed a multistep process for the differentiation of human cortical cells from embryonic stem (ES) and induced pluripotent stem cells (iPSC). The publication goes into some depth regarding the different phases these cells undergo during this differentiation, but critically after two months, these cells form multicellular cultures including both cortical neurons as well as astrocytes. These neurons form glutamatergic synapses that contain NMDA and AMPA receptors and whole cells patch clamp demonstrating? they have mature electrical properties and form functional excitatory synapses.
As though it wasn’t enough to publish the technical achievement differentiating? a cortical neuronal culture from iPSC and ES, the group went on to publish a follow-up in Science Translational Medicine (1), immediately demonstrating the application of this technology. They used the iPSC derived from Down syndrome patients from early Alzheimer’s pathology to model the neuronal cortex of a patient with Alzheimer’s disease.
The classic pathological hallmarks of Alzheimer’s disease are amyloid plaques composed of the amyloid Aβ (Aβ) peptide, (formed from the amyloid precursor protein) and neurofibrillary tangles comprising hyperphosphorylated forms of Tau. The amyloid precursor protein (APP) gene is encoded on chromosome 21 and therefore patients with Down syndrome (caused by trisomy of chromosome 21) develop early onset Alzheimer’s, commonly by 35 years of age.
The group took iPSC from a patient with Down syndrome (DS) and using the method set out in the neuroscience paper differentiated them into cortical neurons in the same way as they did in the first paper with control iPSCs and ES. The group proceeded to measure whether there was Alzheimer’s pathology in these cultures.
They measured levels of Aβ40 and Aβ42 peptide production from 2-4 weeks after the onset of neuronal differentiation. In DS cultures, the levels of both Aβ40 and Aβ42 were significantly higher than in control iPSC. In the DS cultures, these Aβ peptides also form amyloid aggregates that are not visible in the control cultures. They also measured the abundance of Tau phosphorylated at Ser202 and Thr205. This demonstrated that although phosphorylated Tau was expressed in both DS and control neurons, in control cultures phospho Tau was diffusely localised in the primary axons whereas in DS neurons it was aberrantly localised into linear foci in the cell bodies and dendrites in the DS cells. This matched the distribution found within the patient CNS.
The recent failure of so many late-stage Alzheimer’s drugs highlights the need to bring new compounds through the pipeline. Being able to elegantly model such a complex disease is perfect for a drug screening program, or phenotypic screen, to either hit Aβ or phosphorylated Tau. However, the screen can also be used for basic science to monitor disease progression potentially unravelling new targets, e.g. to elucidate the link between Aβ aggregation and hyperphosphorylated Tau. The development of such an elegant in vitro culture system such as this makes all of these more possible.
1. Shi Y, Kirwan P, Smith J, MacLean G, Orkin SH, Livesey FJ. A human stem cell model of early Alzheimer’s disease pathology in Down syndrome. Science translational medicine 4: 124ra29, 2012.
2. Shi Y, Kirwan P, Smith J, Robinson HPC, Livesey FJ. Human cerebral cortex development from pluripotent stem cells to functional excitatory synapses. Nature neuroscience 15: 477–86, S1, 2012.