In early phases of Alzheimer?s disease (AD) only specific types of neurons are affected by pathological lesions. As the disease progresses, additional neurons types gradually start expressing signs of pathology and degeneration throughout the brain. A major obstacle to treating Alzheimer?s disease (AD) is our lack of understanding of the molecular mechanisms underlying this selective neuronal vulnerability. Neurons from the layer II of the entorhinal cortex (ECII) are the most vulnerable neurons of the brain, but there are currently no in vitro culture system for these neurons, preventing both an in-depth analysis of genes involved in neuronal vulnerability, and the search for drugs inhibiting early neurodegeneration affecting these neurons. In the parent RF1 grant, we proposed to identify transcription factors (TFs) involved in specifying ECII neuron identity, using both detailed molecular profiles for ECII and control neurons, which we generated at different points of their development, and cutting-edge functional genomics analyses. These TFs will then be used to drive the differentiation of induced pluripotent stem cells (iPSC) into ECII neurons. While iPSCs provide a versatile platform that can be differentiated into many different neuron types, concerns have been raised that neurons derived from iPSCs are biologically very young, as the iPSC generation protocol ?resets? the biological clock of a cell. AD-associated lesions are age-dependent, and we think that an in vitro paradigm adapted to this study of aging is crucial. In this administrative supplement, we thus want to adapt this iPSC protocol for the direct reprogramming of human fibroblasts, using the TFs identified in the parent grant. Directly reprogrammed fibroblasts are not reset biologically and present a biological age that correlates with the age of the fibroblast donor. After having optimized a protocol for direct reprogramming of fibroblasts into ECII neurons, we will provide a proof of concept that these ECII neurons are a good model for probing selective neuronal vulnerability by studying Tau pathology in neurons reprogrammed from fibroblasts of aging donors and of donors with AD mutations. Overall these ECII neurons directly reprogrammed from fibroblasts would be the first in vitro model for the study of the interplay between selective neuronal vulnerability in AD and aging.
In Alzheimer?s diseases (AD), only certain regions of the brain are initially affected, followed later by more wide-ranging degeneration. In our parent grant, we proposed to establish the first cell culture system to model the neurons that are most vulnerable in AD, using stem cells. It was recently shown that culture systems involving stem cells generate cells that are young biologically, and not perfectly suited for the study of age- associated degeneration. We now propose to modify our initial protocol using ?fibroblast direct reprogramming?, a protocol that does not involve stem cells, and that yields cells than can be biologically much older. This cell culture system would allow AD researchers to investigate, for the first time, the interaction between age and neuronal vulnerability to AD.
