Sandoe, J. | Search bioRxiv

bioRxiv2022

Rapid iPSC inclusionopathy models shed light on formation, consequence and molecular subtype of alpha-synuclein inclusions

Intracellular inclusions accompanying neurodegeneration are histopathologically and ultrastructurally heterogeneous but the significance of this heterogeneity is unclear. iPSC models, while promising for disease modeling, do not form inclusions in a reasonable timeframe and suffer from limited tractability. Here, we developed an iPSC toolbox utilizing piggyBac-based or targeted transgenes to rapidly induce CNS cells with concomitant expression of aggregation-prone proteins. This system is amenable to screening and longitudinal tracking at single-cell and single-inclusion resolution. For proof-of-principle, cortical neuron -synuclein "inclusionopathy" models were engineered to form inclusions through exogenous seeding or -synuclein mutation. These models recapitulated known fibril- and lipid-rich inclusion subtypes, uncovering dynamic interactions between them, and refined the classification of inclusions in postmortem brain. Genetic-modifier and protein-interaction screens pinpointed proteins like RhoA whose sequestration into specific inclusion subtypes is likely to be toxic. This iPSC platform should enhance our understanding of proteinaceous pathologies in neurodegeneration and facilitate therapeutics development.

neuroscience↗

bioRxiv2021

A Multiparametric Activity Profiling Platform for Neuron Disease Phenotyping and Drug Screening

Patient stem cell-derived models enable imaging of complex disease phenotypes and the development of scalable drug discovery platforms. Current preclinical methods for assessing cellular activity do not, however, capture the full intricacies of disease-induced disturbances, and instead typically focus on a single parameter, which impairs both the understanding of disease and the discovery of effective therapeutics. Here, we describe a cloud-based image processing and analysis platform that captures the intricate activity profile revealed by GCaMP fluorescent recordings of intracellular calcium changes and enables discovery of molecules that correct 153 parameters that define the amyotrophic lateral sclerosis motor neuron disease phenotype. In a high-throughput screen we identified compounds that revert the multiparametric disease profile to that found in healthy cells, a novel and robust measure of therapeutic potential quite distinct from unidimensional screening. This platform can guide the development of therapeutics that counteract the multifaceted pathological features of diseased cellular activity.

neuroscience↗