Cell-type-resolved mosaicism reveals clonal dynamics of the human forebrain.

Debate remains around the anatomical origins of specific brain cell subtypes and lineage relationships within the human forebrain1-7. Thus, direct observation in the mature human brain is critical for a complete understanding of its structural organization and cellular origins. Here we utilize brain mosaic variation within specific cell types as distinct indicators for clonal dynamics, denoted as cell-type-specific mosaic variant barcode analysis. From four hemispheres and two different human neurotypical donors, we identified 287 and 780 mosaic variants, respectively, that were used to deconvolve clonal dynamics. Clonal spread and allele fractions within the brain reveal that local hippocampal excitatory neurons are more lineage-restricted than resident neocortical excitatory neurons or resident basal ganglia GABAergic inhibitory neurons. Furthermore, simultaneous genome transcriptome analysis at both a cell-type-specific and a single-cell level suggests a dorsal neocortical origin for a subgroup of DLX1+ inhibitory neurons that disperse radially from an origin shared with excitatory neurons. Finally, the distribution of mosaic variants across 17 locations within one parietal lobe reveals that restriction of clonal spread in the anterior-posterior axis precedes restriction in the dorsal-ventral axis for both excitatory and inhibitory neurons. Thus, cell-type-resolved somatic mosaicism can uncover lineage relationships governing the development of the human forebrain.

APOE4/4 is linked to damaging lipid droplets in Alzheimer's disease microglia.

Several genetic risk factors for Alzheimer's disease implicate genes involved in lipid metabolism and many of these lipid genes are highly expressed in glial cells1. However, the relationship between lipid metabolism in glia and Alzheimer's disease pathology remains poorly understood. Through single-nucleus RNA sequencing of brain tissue in Alzheimer's disease, we have identified a microglial state defined by the expression of the lipid droplet-associated enzyme ACSL1 with ACSL1-positive microglia being most abundant in patients with Alzheimer's disease having the APOE4/4 genotype. In human induced pluripotent stem cell-derived microglia, fibrillar Aß induces ACSL1 expression, triglyceride synthesis and lipid droplet accumulation in an APOE-dependent manner. Additionally, conditioned media from lipid droplet-containing microglia lead to Tau phosphorylation and neurotoxicity in an APOE-dependent manner. Our findings suggest a link between genetic risk factors for Alzheimer's disease with microglial lipid droplet accumulation and neurotoxic microglia-derived factors, potentially providing therapeutic strategies for Alzheimer's disease.

Epigenomic landscape of the human dorsal root ganglion: sex differences and transcriptional regulation of nociceptive genes.

Gene expression is influenced by chromatin architecture via controlled access of regulatory factors to DNA. To better understand gene regulation in the human dorsal root ganglion (hDRG) we used bulk and spatial transposase-accessible chromatin technology followed by sequencing (ATAC-seq). Using bulk ATAC-seq, we detected that in females diverse differentially accessible chromatin regions (DARs) mapped to the X chromosome and in males to autosomal genes. EGR1/3 and SP1/4 transcription factor binding motifs were abundant within DARs in females, and JUN, FOS and other AP-1 factors in males. To dissect the open chromatin profile in hDRG neurons, we used spatial ATAC-seq. The neuron cluster showed higher chromatin accessibility in GABAergic, glutamatergic, and interferon-related genes in females, and in Ca2+- signaling-related genes in males. Sex differences in transcription factor binding sites in neuron-proximal barcodes were consistent with the trends observed in bulk ATAC-seq data. We validated that EGR1 expression is biased to female hDRG compared to male. Strikingly, XIST, the long-noncoding RNA responsible for X inactivation, hybridization signal was found to be highly dispersed in the female neuronal but not non-neuronal nuclei suggesting weak X inactivation in female hDRG neurons. Our findings point to baseline epigenomic sex differences in the hDRG that likely underlie divergent transcriptional responses that determine mechanistic sex differences in pain.

Dopamine-driven Increase in IL-1ß in Myeloid Cells is Mediated by Differential Dopamine Receptor Expression and Exacerbated by HIV.

The catecholamine neurotransmitter dopamine is classically known for regulation of central nervous system (CNS) functions such as reward, movement, and cognition. Increasing evidence also indicates that dopamine regulates critical functions in peripheral organs and is an important immunoregulatory factor. We have previously shown that dopamine increases NF-κB activity, inflammasome activation, and the production of inflammatory cytokines such as IL-1ß in human macrophages. As myeloid lineage cells are central to the initiation and resolution of acute inflammatory responses, dopamine-mediated dysregulation of these functions could both impair the innate immune response and exacerbate chronic inflammation. However, the exact pathways by which dopamine drives myeloid inflammation are not well defined, and studies in both rodent and human systems indicate that dopamine can impact the production of inflammatory mediators through both D1-like dopamine receptors (DRD1, DRD5) and D2-like dopamine receptors (DRD2, DRD3, and DRD4). Therefore, we hypothesized that dopamine-mediated production of IL-1ß in myeloid cells is regulated by the ratio of different dopamine receptors that are activated. Our data in primary human monocyte-derived macrophages (hMDM) indicate that DRD1 expression is necessary for dopamine-mediated increases in IL-1ß, and that changes in the expression of DRD2 and other dopamine receptors can alter the magnitude of the dopamine-mediated increase in IL-1ß. Mature hMDM have a high D1-like to D2-like receptor ratio, which is different relative to monocytes and peripheral blood mononuclear cells (PBMCs). We further confirm in human microglia cell lines that a high ratio of D1-like to D2-like receptors promotes dopamine-induced increases in IL-1ß gene and protein expression using pharmacological inhibition or overexpression of dopamine receptors. RNA-sequencing of dopamine-treated microglia shows that genes encoding functions in IL-1ß signaling pathways, microglia activation, and neurotransmission increased with dopamine treatment. Finally, using HIV as an example of a chronic inflammatory disease that is substantively worsened by comorbid substance use disorders (SUDs) that impact dopaminergic signaling, we show increased effects of dopamine on inflammasome activation and IL-1ß in the presence of HIV in both human macrophages and microglia. These data suggest that use of addictive substances and dopamine-modulating therapeutics could dysregulate the innate inflammatory response and exacerbate chronic neuroimmunological conditions like HIV. Thus, a detailed understanding of dopamine-mediated changes in inflammation, in particular pathways regulating IL-1ß, will be critical to effectively tailor medication regimens.

Studying neurodegenerative diseases in culture models

Neurodegenerative diseases are pathological conditions that have an insidious onset and chronic progression. Different models have been established to study these diseases in order to understand their underlying mechanisms and to investigate new therapeutic strategies. Although various in vivo models are currently in use, in vitro models might provide important insights about the pathogenesis of these disorders and represent an interesting approach for the screening of potential pharmacological agents. In the present review, we discuss various in vitro and ex vivo models of neurodegenerative disorders in mammalian cells and tissues.