Human‐specific transcriptome of ventral and dorsal midbrain dopamine neurons

Objective Neuronal loss in the substantia nigra pars compacta (SNpc) in Parkinson's disease (PD) is not uniform as dopamine neurons from the ventral tier are lost more rapidly than those of the dorsal tier. Identifying the intrinsic differences that account for this differential vulnerability may provide a key for developing new treatments for PD. Method Here we compared the RNA‐sequenced transcriptomes of ~100 laser captured micro‐dissected SNpc neurons from each tier from seven healthy controls. Results Expression levels of dopaminergic markers were similar across the tiers while markers specific to the neighbouring ventral tegmental area were virtually undetected. After accounting for unwanted sources of variation, we identified one hundred and six differentially expressed genes (DEGs) between the SNpc tiers. The genes higher in the dorsal/resistant SNpc tier neurons displayed coordinated patterns of expression across the human brain, their protein products had more interactions than expected by chance and they demonstrated evidence of functional convergence. No significant shared functionality was found for genes higher in the ventral/vulnerable SNpc tier. Surprisingly but importantly, none of the identified DEGs were among the familial PD genes or genome‐wide associated loci. Finally, we found some DEGs in opposite tier orientation between human and analogous mouse populations. Interpretation Our results highlight functional enrichments of vesicular trafficking, ion transport/homeostasis and oxidative stress genes showing higher expression in the resistant neurons of the SNpc dorsal tier. Furthermore, the comparison of gene expression variation in human and mouse SNpc populations strongly argues for the need of human‐focused Omics studies

Deep phenotyping for precision medicine in Parkinson's disease

A major challenge in medical genomics is to understand why individuals with the same disorder have different clinical symptoms and why those who carry the same mutation may be affected by different disorders. In every complex disorder, identifying the contribution of different genetic and non-genetic risk factors is a key obstacle to understanding disease mechanisms. Genetic studies rely on precise phenotypes and are unable to uncover the genetic contributions to a disorder when phenotypes are imprecise. To address this challenge, deeply phenotyped cohorts have been developed for which detailed, fine-grained data have been collected. These cohorts help us to investigate the underlying biological pathways and risk factors to identify treatment targets, and thus to advance precision medicine. The neurodegenerative disorder Parkinson's disease has a diverse phenotypical presentation and modest heritability, and its underlying disease mechanisms are still being debated. As such, considerable efforts have been made to develop deeply phenotyped cohorts for this disorder. Here, we focus on Parkinson's disease and explore how deep phenotyping can help address the challenges raised by genetic and phenotypic heterogeneity. We also discuss recent methods for data collection and computation, as well as methodological challenges that have to be overcome

Pro-maturational effects of human iPSC-derived cortical astrocytes upon iPSC-derived cortical neurons

Astrocytes influence neuronal maturation and function by providing trophic support, regulating the extracellular environment, andmodulating signaling at synapses. The emergence of induced pluripotent stem cell (iPSC) technology offers a human system with whichto validate and re-evaluate insights from animal studies. Here, we set out to examine interactions between human astrocytes and neuronsderived from a common cortical progenitor pool, thereby recapitulating aspects ofin vivocortical development. We show that the corticaliPSC-derived astrocytesexhibit many of the molecular and functional hallmarks of astrocytes. Furthermore, optogenetic and electrophys-iological co-culture experiments reveal that the iPSC-astrocytes can actively modulate ongoing synaptic transmission and exertpro-maturational effects upon developing networks of iPSC-derived cortical neurons. Finally, transcriptomic analyses implicate syn-apse-associated extracellular signaling in the astrocytes’ pro-maturational effects upon the iPSC-derived neurons. This work helps laythe foundation for future investigations into astrocyte-to-neuron interactions in human health and disease

Postmitotic cell longevity–associated genes: a transcriptional signature of postmitotic maintenance in neural tissues

Different cell types have different post-mitotic maintenance requirements. Nerve cells, however, are unique in this respect as they need to survive and preserve their functional complexity for the entire lifetime of the organism and failure at any level of their supporting mechanisms, leads to a wide range of neurodegenerative conditions. Whether these differences across tissues arise from the activation of distinct cell type-specific maintenance mechanisms or the differential activation of a common molecular repertoire is not known. In order to identify the transcriptional signature of post-mitotic cellular longevity (PMCL), we compared whole genome transcriptome data from human tissues ranging in longevity from 120 days to over 70 years and found a set of 81 genes whose expression levels are closely associated with increased cell longevity. Using expression data from ten independent sources, we found that these genes are more highly co-expressed in longer living tissues and are enriched in specific biological processes and transcription factors targets compared to randomly selected gene samples. Crucially, we found that PMCL-associated genes are down-regulated in the cerebral cortex and substantia nigra of Alzheimer’s and Parkinson’s disease patients as well as Hutchinson-Gilford progeria-derived fibroblasts, and that this down regulation is specifically linked to their underlying association with cellular longevity. Moreover, we found that sexually dimorphic brain expression of PMCL-associated genes reflects sexual differences in lifespan in humans and macaques. Taken together our results suggest that PMCL-associated genes are part of a generalized machinery of post-mitotic maintenance and functional stability in both neural and non-neural cells and support the notion of a common molecular repertoire differentially engaged in different cell types with different survival requirements

Single-cell transcriptomics and In vitro lineage tracing reveals differential susceptibility of human iPSC-derived midbrain dopaminergic neurons in a cellular model of Parkinson's Disease

Advances in stem cell technologies open up new avenues for modelling development and diseases. The success of these pursuits, however, relies on the use of cells most relevant to those targeted by the disease of interest, for example, midbrain dopaminergic neurons for Parkinson’s disease. In the present study, we report the generation of a human induced pluripotent stem cell (iPSC) line capable of purifying and tracing nascent midbrain dopaminergic progenitors and their differentiated progeny via the expression of a Blue Fluorescent Protein (BFP). This was achieved by CRISPR/Cas9-assisted knock-in of BFP and Cre into the safe harbour locus AAVS1 and an early midbrain dopaminergic lineage marker gene LMX1A, respectively. Immunocytochemical analysis and single-cell RNA sequencing of iPSC-derived neural cultures confirm developmental recapitulation of the human fetal midbrain and high-quality midbrain cells. By modelling Parkinson’s disease-related drug toxicity using 1-Methyl-4-phenylpyridinium (MPP+), we showed a preferential reduction of BFP+ cells, a finding demonstrated independently by cell death assays and single-cell transcriptomic analysis of MPP+ treated neural cultures. Together, these results highlight the importance of disease-relevant cell types in stem cell modelling

Combining multi-omics and drug perturbation profiles to identify novel treatments that improve disease phenotypes in spinal muscular atrophy [preprint]

Spinal muscular atrophy (SMA) is a neuromuscular disorder caused by loss of survival motor neuron (SMN) protein. While SMN restoration therapies are beneficial, they are not a cure. We aimed to identify novel treatments to alleviate muscle pathology combining transcriptomics, proteomics and perturbational datasets. This revealed potential drug candidates for repurposing in SMA. One of the lead candidates, harmine, was further investigated in cell and animal models, improving multiple disease phenotypes, including SMN expression and lifespan. Our work highlights the potential of multiple, parallel data driven approaches for development of novel treatments for use in combination with SMN restoration therapies

Lipopolysaccharide distinctively alters human microglia transcriptomes to resemble microglia from Alzheimer's disease mouse models

Alzheimer's disease (AD) is the most common form of dementia, and risk-influencing genetics implicates microglia and neuroimmunity in the pathogenesis of AD. Induced pluripotent stem cell (iPSC)-derived microglia (iPSC-microglia) are increasingly used as a model of AD, but the relevance of historical immune stimuli to model AD is unclear. We performed a detailed cross-comparison over time on the effects of combinatory stimulation of iPSC-microglia, and in particular their relevance to AD. We used single-cell RNA sequencing to measure the transcriptional response of iPSC-microglia after 24 h and 48 h of stimulation with prostaglandin E2 (PGE2) or lipopolysaccharide (LPS)+interferon gamma (IFN-γ), either alone or in combination with ATPγS. We observed a shared core transcriptional response of iPSC-microglia to ATPγS and to LPS+IFN-γ, suggestive of a convergent mechanism of action. Across all conditions, we observed a significant overlap, although directional inconsistency to genes that change their expression levels in human microglia from AD patients. Using a data-led approach, we identify a common axis of transcriptomic change across AD genetic mouse models of microglia and show that only LPS provokes a transcriptional response along this axis in mouse microglia and LPS+IFN-γ in human iPSC-microglia. This article has an associated First Person interview with the first author of the paper

Combining multiomics and drug perturbation profiles to identify muscle-specific treatments for spinal muscular atrophy

Spinal muscular atrophy (SMA) is a neuromuscular disorder caused by loss of survival motor neuron (SMN) protein. While SMN restoration therapies are beneficial, they are not a cure. We aimed to identify potentially novel treatments to alleviate muscle pathology combining transcriptomics, proteomics, and perturbational data sets. This revealed potential drug candidates for repurposing in SMA. One of the candidates, harmine, was further investigated in cell and animal models, improving multiple disease phenotypes, including lifespan, weight, and key molecular networks in skeletal muscle. Our work highlights the potential of multiple and parallel data-driven approaches for the development of potentially novel treatments for use in combination with SMN restoration therapies