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The loss of skeletal muscle mass and size, or muscle atrophy, is a common human experience, linked to disability, for which there are no widely accepted pharmacological therapies. Piezo1 is a mechanosensitive cation channel that opens upon alteration of the plasma membrane lipid bilayer, such as through increased membrane tension. In this issue of the JCI, Hirata et al. identified Piezo1 and its downstream effectors, Krüppel-like factor 15 (KLF15) and interleukin-6 (IL-6), as an important signaling pathway in a murine model of disuse atrophy. Through genetic and pharmacological modulation of the pathway, the authors demonstrated that immobilization resulted in downregulation of Piezo1 and basal intracellular calcium concentration ([Ca2+]i), increasing expression of Klf15 and its downstream target Il6 and thereby inducing muscle atrophy. Piezo1 has been considered a therapeutic target for diverse disorders, including atherosclerosis and kidney fibrosis, and with this publication should now also be considered a viable target for disuse atrophy.

The use of human pluripotent stem cells for in vitro disease modelling and clinical applications requires protocols that convert these cells into relevant adult cell types. Here, we report the rapid and efficient differentiation of human pluripotent stem cells into vascular endothelial and smooth muscle cells. We found that GSK3 inhibition and BMP4 treatment rapidly committed pluripotent cells to a mesodermal fate and subsequent exposure to VEGF-A or PDGF-BB resulted in the differentiation of either endothelial or vascular smooth muscle cells, respectively. Both protocols produced mature cells with efficiencies exceeding 80% within six days. On purification to 99% via surface markers, endothelial cells maintained their identity, as assessed by marker gene expression, and showed relevant in vitro and in vivo functionality. Global transcriptional and metabolomic analyses confirmed that the cells closely resembled their in vivo counterparts. Our results suggest that these cells could be used to faithfully model human disease. Cowan and colleagues report a method to generate mature endothelial or vascular smooth muscle cells from human pluripotent stem cells with high efficiency and purity.

Mitochondria are key organelles in regulating the metabolic state of a cell. In the brain, mitochondrial oxidative metabolism is the prevailing mechanism for neurons to generate ATP. While it is firmly established that neuronal function is highly dependent on mitochondrial metabolism, it is less well-understood how astrocytes function rely on mitochondria. In this study, we investigate if astrocytes require a functional mitochondrial electron transport chain (ETC) and oxidative phosphorylation (oxPhos) under physiological and injury conditions. By immunohistochemistry we show that astrocytes expressed components of the ETC and oxPhos complexes . Genetic inhibition of mitochondrial transcription by conditional deletion of ( ) led to dysfunctional ETC and oxPhos activity, as indicated by aberrant mitochondrial swelling in astrocytes. Mitochondrial dysfunction did not impair survival of astrocytes, but caused a reactive gliosis in the cortex under physiological conditions. Photochemically initiated thrombosis induced ischemic stroke led to formation of hyperfused mitochondrial networks in reactive astrocytes of the perilesional area. Importantly, mitochondrial dysfunction significantly reduced the generation of new astrocytes and increased neuronal cell death in the perilesional area. These results indicate that astrocytes require a functional ETC and oxPhos machinery for proliferation and neuroprotection under injury conditions.

Mutations in glucocerebrosidase cause the lysosomal storage disorder Gaucher’s disease and are the most common risk factor for Parkinson’s disease. Therapies to restore the enzyme’s function in the brain hold great promise for treating the neurological implications. Thus, we developed blood-brain barrier penetrant therapeutic molecules by fusing transferrin receptor-binding moieties to β-glucocerebrosidase (referred to as GCase-BS). We demonstrate that these fusion proteins show significantly increased uptake and lysosomal efficiency compared to the enzyme alone. In a cellular disease model, GCase-BS rapidly rescues the lysosomal proteome and lipid accumulations beyond known substrates. In a mouse disease model, intravenous injection of GCase-BS leads to a sustained reduction of glucosylsphingosine and can lower neurofilament-light chain plasma levels. Collectively, these findings demonstrate the potential of GCase-BS for treating GBA1 -associated lysosomal dysfunction, provide insight into candidate biomarkers, and may ultimately open a promising treatment paradigm for lysosomal storage diseases extending beyond the central nervous system. Mutations in glucocerebrosidase (GCase) cause the lysosomal storage disorder Gaucher’s disease and are the most common risk factor for Parkinson’s disease. Using a fusion protein comprising GCase and a transferrin receptor antibody fragment, the authors show that the transferrin receptor pathway can be therapeutically exploited to both pass the blood-brain barrier and efficiently target lysosomal GCase deficiency.

Neural stem cells (NSCs) generate new granule cells throughout life in the mammalian hippocampus. Canonical Wnt signaling regulates the differentiation of NSCs towards the neuronal lineage. Here we identified the prospero-related homeodomain transcription factor Prox1 as a target of β-catenin-TCF/LEF signaling in vitro and in vivo. Prox1 overexpression enhanced neuronal differentiation whereas shRNA-mediated knockdown of Prox1 impaired the generation of neurons in vitro and within the hippocampal niche. In contrast, Prox1 was not required for survival of adult-generated granule cells after they had matured, suggesting a role for Prox1 in initial granule cell differentiation but not in the maintenance of mature granule cells. The data presented here characterize a molecular pathway from Wnt signaling to a transcriptional target leading to granule cell differentiation within the adult brain and identify a stage-specific function for Prox1 in the process of adult neurogenesis.

Human induced pluripotent stem cells (hiPSCs) are invaluable to study developmental processes and disease mechanisms particularly in the brain. hiPSCs can be differentiated into mature and functional dopaminergic (DA) neurons. Having robust protocols for the generation of differentiated DA neurons from pluripotent cells is a prerequisite for the use of hiPSCs to study disease mechanisms, for drug discovery, and eventually for cell replacement therapy. Here, we describe a protocol for generating and expanding large numbers of homogeneous midbrain floor plate progenitors (mFPPs) that retain efficient DA neurogenic potential over multiple passages and can be cryobanked. We demonstrate that expanded mFPPs have increased DA neuron potential and differentiate more efficiently and rapidly than progenitors generated by standard protocols. In addition, this novel method results in increased numbers of DA neurons that in vitro show characteristic electrophysiological properties of nigrostriatal DA neurons, produce high levels of dopamine, and integrate into host mice when grafted in vivo . Thus, we describe a robust method for producing human mesencephalic DA neurons from hiPSCs.

Economic efficiency has been a popular explanation for how networks self-organize within the developing nervous system. However, the precise nature of the economic negotiations governing this putative organizational principle remains unclear. Here, we address this question further by combining large-scale electrophysiological recordings to characterize the functional connectivity of developing neuronal networks in vitro, with a generative modeling approach capable of simulating network formation. We find that the best fitting model uses a homophilic generative wiring principle in which neurons form connections to other neurons which are spatially proximal and have similar connectivity patterns to themselves. Homophilic generative models outperform more canonical models in which neurons wire depending upon their spatial proximity either alone or in combination with the extent of their local connectivity. This homophily-based mechanism for neuronal network emergence accounts for a wide range of observations that are described, but not sufficiently explained, by traditional analyses of network topology. Using rodent and human neuronal cultures, we show that homophilic generative mechanisms can accurately recapitulate the topology of emerging cellular functional connectivity, representing an important wiring principle and determining factor of neuronal network formation in vitro.

Here, we have developed a highly sensitive immunoassay for Dcx to characterize expression in brain and cerebrospinal fluid (CSF) of rodents. We demonstrate that Dcx is widely expressed during development in various brain regions and as well can be detected in cerebrospinal fluid of rats (up to 30 days postnatal). While Dcx protein level decline in adulthood and were detectable in neurogenic regions of the adult rodent brain, similar levels were also detectable in brain regions expected to bear no neurogenesis including the cerebral cortex and CA1/CA3 enriched hippocampus. We monitored DCX protein levels after paradigms to increase or severely decrease adult hippocampal neurogenesis, namely physical activity and cranial radiation, respectively. In both paradigms, Dcx protein- and mRNA-levels clearly reflected changes in neurogenesis in the hippocampus. However, basal Dcx-levels are unaffected in non-neurogenic regions (e.g. CA1/CA3 enriched hippocampus, cortex). These data suggest that there is a substantial \"non-neurogenic\" pool of Dcx- protein, whose regulation can be uncoupled from adult neurogenesis suggesting caution for the interpretation of such studies.

Mutations in GBA1, the gene encoding the lysosomal hydrolase glucocerebrosidase (GCase), are the strongest common genetic risk factor for Parkinson’s Disease (PD). However, these mutations are incompletely penetrant, which suggests that there are likely genetic modifiers of GCase function. To identify such genes, we implemented a live cell GCase activity-based CRISPR-platform to enable genome-wide screening for novel regulators of lysosomal GCase activity. Among the screening hits, we find significant enrichment of genes linked to development and progression of PD through genome-wide association studies (GWAS). Moreover, we identify two lysosomal lipid transporter genes, including those encoding the lysosphospholipid transporter SPNS1 and the cholesterol transporter NPC1, and find an allele of SPNS1 that is associated with increased risk of PD. We show that disruption of SPNS1 does not affect GCase protein levels but impairs its lysosomal function. Collectively, these data suggest that dysfunction of many PD-associated genes converge to impact lysosomal GCase activity and thereby contribute to disease pathogenesis. A better understanding of the impacts of these and the other GCase modulators identified here should help unravel the important, yet complex, relationship between GBA1 and PD.

The loss of skeletal muscle mass and size, or muscle atrophy, is a common human experience, linked to disability, for which there are no widely accepted pharmacological therapies. Piezo1 is a mechanosensitive cation channel that opens upon alteration of the plasma membrane lipid bilayer, such as through increased membrane tension. In this issue of theJCI, Hirata et al. identified Piezol and its downstream effectors, Krüppel-like factor 15 (KLF15) and interleukin-6 (IL-6), as an important signaling pathway in a murine model of disuse atrophy. Through genetic and pharmacological modulation of the pathway, the authors demonstrated that immobilization resulted in downregulation of Piezol and basal intracellular calcium concentration ([Ca2·].), increasing expression of Klf15 and its downstream target Il6 and thereby inducing muscle atrophy. Piezol has been considered a therapeutic target for diverse disorders, including atherosclerosis and kidney fibrosis, and with this publication should now also be considered a viable target for disuse atrophy.