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Summary: Neurons in the neocortex are generated during embryonic development. While the adult ventricular-subventricular zone (V-SVZ) contains cells with neural stem/progenitors’ characteristics, it remains unclear whether it has the capacity of producing neocortical neurons. Here, we show that generating neurons with transcriptomic resemblance to upper layer neocortical neurons continues in the V-SVZ of mouse models of a human condition known as periventricular heterotopia by abrogating Flna and Flnb. We found such surplus neurogenesis was associated with V-SVZ’s upregulation of oxidative phosphorylation, mitochondrial biogenesis, and vascular abundance. Additionally, spatial transcriptomics analyses showed V-SVZ’s neurogenic activation was coupled with transcriptional enrichment of genes in diverse pathways for energy metabolism, angiogenesis, cell signaling, synaptic transmission, and turnovers of nucleic acids and proteins in upper cortical layers. These findings support the potential of generating neocortical neurons in adulthood through boosting brain-wide vascular circulation, aerobic adenosine triphosphate synthesis, metabolic turnover, and neuronal activity.

Abstract Background Children of mothers with gestational diabetes mellitus (GDM) are more prone to acquire type 2 diabetes and obesity as adults. Due to this link, early intervention strategies that alter the gut microbiome may benefit the mother and kid long-term. This work uses metagenomic and transcriptome sequencing to investigate how probiotics affect gut microbiota dysbiosis and inflammation in GDM. Methods GDM and control metagenomic sequencing data were obtained from the SRA database. This metagenomic data helped us understand gut microbiota abundance and function. KEGG detected and extracted functional pathway genes. Transcriptome sequencing data evaluated GDM-related gene expression. Finally, GDM animal models were given probiotics orally to evaluate inflammatory response, regulatory immune cell fractions, and leptin protein levels. Results GDM patients had more Fusobacteria and Firmicutes, while healthy people had more Bacteroidetes. Gut microbiota composition may affect GDM by altering the L-aspartate and L-asparagine super pathways. Mannan degradation and the super pathway of L-aspartate and L-asparagine synthesis enhanced in GDM mice with leptin protein overexpression. Oral probiotics prevent GDM by lowering leptin. Oral probiotics increased Treg, Tfr, and Breg cells, which decreased TNF-α and IL-6 and increased TGF-β and IL-10, preventing inflammation and preserving mouse pregnancy. Conclusion Dysbiosis of the gut microbiota may increase leptin expression and cause GDM. Oral probiotics enhance Treg, Tfr, and Breg cells, which limit the inflammatory response and assist mice in sustaining normal pregnancy. Thus, oral probiotics may prevent GDM, enabling targeted gut microbiota modulation and maternal and fetal health.

Hemorrhage is a leading cause of death in trauma. Tourniquets are effective at controlling extremity hemorrhage and have saved lives. However, tourniquets can cause ischemia reperfusion injury of limbs, leading to systemic inflammation and other adverse effects, which results in secondary damage to the kidney, lung, and liver. A clinically relevant animal model is critical to understanding the pathophysiology of this process and developing therapeutic interventions. Despite the importance of animal models, tourniquet-induced lower limb ischemia/reperfusion (TILLIR) models to date lack a hemorrhage component. We sought to develop a new TILLIR model that included hemorrhage and analyze the subsequent impact on kidney, lung and liver injuries. Four groups of mice were examined: group 1) control, group 2) hemorrhage, group 3) tourniquet application, and group 4) hemorrhage and tourniquet application. The hemorrhagic injury consisted of the removal of 15% of blood volume through the submandibular vein. The tourniquet injury consisted of orthodontic rubber bands applied to the inguinal area bilaterally for 80 min. Mice were then placed in metabolic cages individually for 22 h to collect urine. Hemorrhage alone did not significantly affect transcutaneous glomerular filtration rate (tGFR), blood urea nitrogen (BUN) or urinary kidney injury molecule-1 (KIM-1) levels. Without hemorrhage, TILLIR decreased tGFR by 46%, increased BUN by 162%, and increased KIM-1 by 27% (p < 0.05 for all). With hemorrhage, TILLIR decreased the tGFR by 72%, increased BUN by 395%, and increased urinary KIM-1 by 37% (p < 0.05 for all). These differences were statistically significant (p < 0.05). While hemorrhage had no significant effect on TILLIR-induced renal tubular degeneration and necrosis, it significantly increased TILLIR-induced lung total injury scores and congestion, and fatty liver. In conclusion, hemorrhage exacerbates TILLIR-induced acute kidney injury and structural damage in the lung and liver.

Summary: Aging is an intricate process characterized by multiple hallmarks including stem cell exhaustion, genome instability, epigenome alteration, impaired proteostasis, and cellular senescence. Whereas each of these traits is detrimental at the cellular level, it remains unclear how they are interconnected to cause systemic organ deterioration. Here we show that abrogating Brap, a BRCA1-associated protein essential for neurogenesis, results in persistent DNA double-strand breaks and elevation of histone H2A mono- and poly-ubiquitination (H2Aub). These defects extend to cellular senescence and proteasome-mediated histone H2A proteolysis with alterations in cells’ proteomic and epigenetic states. Brap deletion in the mouse brain causes neuroinflammation, impaired proteostasis, accelerated neurodegeneration, and substantially shortened the lifespan. We further show the elevation of H2Aub also occurs in human brain tissues with Alzheimer’s disease. These data together suggest that chromatin aberrations mediated by H2Aub may act as a nexus of multiple aging hallmarks and promote tissue-wide degeneration.

Summary: The NDE1 gene encodes a scaffold protein essential for brain development. Although biallelic NDE1 loss of function (LOF) causes microcephaly with profound mental retardation, NDE1 missense mutations and copy number variations are associated with multiple neuropsychiatric disorders. However, the etiology of the diverse phenotypes resulting from NDE1 aberrations remains elusive. Here we demonstrate Nde1 controls neurogenesis through facilitating H4K20 trimethylation-mediated heterochromatin compaction. This mechanism patterns diverse chromatin landscapes and stabilizes constitutive heterochromatin of neocortical neurons. We demonstrate that NDE1 can undergo dynamic liquid-liquid phase separation, partitioning to the nucleus and interacting with pericentromeric and centromeric satellite repeats. Nde1 LOF results in nuclear architecture aberrations and DNA double-strand breaks, as well as instability and derepression of pericentromeric satellite repeats in neocortical neurons. These findings uncover a pivotal role of NDE1/Nde1 in establishing and protecting neuronal heterochromatin. They suggest that heterochromatin instability predisposes a wide range of brain dysfunction.

Abstract The mechanisms by which lower limb ischemia/reperfusion induces acute kidney injury (AKI) remain largely uncharacterized. We hypothesized that tourniquet‐induced lower limb ischemia/reperfusion (TILLIR) would inhibit mitochondrial function in the renal cortex. We used a murine model to show that TILLIR of the high thigh regions inflicted time‐dependent AKI as determined by renal function and histology. This effect was associated with decreased activities of mitochondrial complexes I, II, V and citrate synthase in the kidney cortex. Moreover, TILLIR reduced mRNA levels of a master regulator of mitochondrial biogenesis PGC‐1α, and its downstream genes NDUFS1 and ATP5o in the renal cortex. TILLIR also increased serum corticosterone concentrations. TILLIR did not significantly affect protein levels of the critical regulators of mitophagy PINK1 and PARK2, mitochondrial transport proteins Tom20 and Tom70, or heat‐shock protein 27. TILLIR had no significant effect on mitochondrial oxidative stress as determined by mitochondrial ability to generate reactive oxygen species, protein carbonylation, or protein levels of MnSOD and peroxiredoxin1. However, TILLIR inhibited classic autophagic flux by increasing p62 protein abundance and preventing the conversion of LC3‐I to LC3‐II. TILLIR increased phosphorylation of cytosolic and mitochondrial ERK1/2 and mitochondrial AKT1, as well as mitochondrial SGK1 activity. In conclusion, lower limb ischemia/reperfusion induces distal AKI by inhibiting mitochondrial function through reducing mitochondrial biogenesis. This AKI occurs without significantly affecting PINK1‐PARK2‐mediated mitophagy or mitochondrial oxidative stress in the kidney cortex.

Summary: Cells initiate fate decisions during G1 phase by converting extracellular signals into distinctive cell cycle kinetics. The DNA replication timing is determined in G1 phase; lengthened G1 and hastened S phases correlate with increased neurogenic propensity of neural progenitor cells (NPCs), although the underlying molecular control remains elusive. Here, we report that proper G1 phase completion in NPCs requires Brap, a Ras-Erk signaling modulator with ubiquitin E3 ligase activity. We identified Skp2 and Skp2-associated SCF ubiquitin ligase as a key target of Brap-mediated polyubiquitination. Loss of Brap resulted in elevated Skp2, which increased p27Kip1 destruction, leading to G1 phase truncation and premature S phase entry. The aberrantly executed G1 in Brap-mutant NPCs, followed by hindered S phase progression and increased G2 phase arrest, which together prolonged the cell cycle, impeded neuronal differentiation and culminated in microcephaly. These findings demonstrate that neuronal differentiation is potentiated during G1 phase by Brap-directed cascade of events in cell signaling and protein turnover. : Lanctot et al. show that the capacity of neural progenitors to generate cerebral cortical neurons relies on the integration of cell signaling and ubiquitin-mediated protein turnover to complete G1 phase of the cell cycle. Keywords: cell cycle, G1 phase, G1/S transition, stem/progenitor, differentiation, ubiquitination, cerebral cortical neurogenesis

Mutations in the filamin A (FlnA) gene are frequently associated with severe arterial abnormalities, although the physiological role for this cytoskeletal element remains poorly understood in vascular cells. We used a conditional mouse model to selectively delete FlnA in smooth muscle (sm) cells at the adult stage, thus avoiding the developmental effects of the knockout. Basal blood pressure was significantly reduced in conscious smFlnA knockout mice. Remarkably, pressure-dependent tone of the resistance caudal artery was lost, whereas reactivity to vasoconstrictors was preserved. Impairment of the myogenic behavior was correlated with a lack of calcium influx in arterial myocytes upon an increase in intraluminal pressure. Notably, the stretch activation of CaV1.2 was blunted in the absence of smFlnA. In conclusion, FlnA is a critical upstream element of the signaling cascade underlying the myogenic tone. These findings allow a better understanding of the molecular basis of arterial autoregulation and associated disease states.

The mechanically activated non-selective cation channel Piezo1 is a determinant of vascular architecture during early development. Piezo1-deficient embryos die at midgestation with disorganized blood vessels. However, the role of stretch-activated ion channels (SACs) in arterial smooth muscle cells in the adult remains unknown. Here, we show that Piezo1 is highly expressed in myocytes of small-diameter arteries and that smooth-muscle-specific Piezo1 deletion fully impairs SAC activity. While Piezo1 is dispensable for the arterial myogenic tone, it is involved in the structural remodeling of small arteries. Increased Piezo1 opening has a trophic effect on resistance arteries, influencing both diameter and wall thickness in hypertension. Piezo1 mediates a rise in cytosolic calcium and stimulates activity of transglutaminases, cross-linking enzymes required for the remodeling of small arteries. In conclusion, we have established the connection between an early mechanosensitive process, involving Piezo1 in smooth muscle cells, and a clinically relevant arterial remodeling.

Neuronal fate-restricted intermediate progenitors (IPs) are derived from the multipotent radial glia (RGs) and serve as the direct precursors for cerebral cortical neurons, but factors that control their neurogenic plasticity remain elusive. Here we report that IPs’ neuron production is enhanced by abrogating filamin function, leading to the generation of periventricular neurons independent of normal neocortical neurogenesis and neuronal migration. Loss of Flna in neural progenitor cells (NPCs) led RGs to undergo changes resembling epithelial-mesenchymal transition (EMT) along with exuberant angiogenesis that together changed the microenvironment and increased neurogenesis of IPs. We show that by collaborating with β-arrestin, Flna maintains the homeostatic signaling between the vasculature and NPCs, and loss of this function results in escalated Vegfa and Igf2 signaling, which exacerbates both EMT and angiogenesis to further potentiate IPs’ neurogenesis. These results suggest that the neurogenic potential of IPs may be boosted in vivo by manipulating Flna-mediated neurovascular communication.

Successfully completing the S phase of each cell cycle ensures genome integrity. Impediment of DNA replication can lead to DNA damage and genomic disorders. In this study, we show a novel function for NDE1, whose mutations cause brain developmental disorders, in safeguarding the genome through S phase during early steps of neural progenitor fate restrictive differentiation. Nde1 mutant neural progenitors showed catastrophic DNA double strand breaks concurrent with the DNA replication. This evoked DNA damage responses, led to the activation of p53-dependent apoptosis, and resulted in the reduction of neurons in cortical layer II/III. We discovered a nuclear pool of Nde1, identified the interaction of Nde1 with cohesin and its associated chromatin remodeler, and showed that stalled DNA replication in Nde1 mutants specifically occurred in mid-late S phase at heterochromatin domains. These findings suggest that NDE1-mediated heterochromatin replication is indispensible for neuronal differentiation, and that the loss of NDE1 function may lead to genomic neurological disorders.

Radial glial cells (RGCs) are distinctive neural stem cells with an extraordinary slender bipolar morphology and dual functions as precursors and migration scaffolds for cortical neurons. Here we show a novel mechanism by which the Lis1-Nde1 complex maintains RGC functions through stabilizing the dystrophin/dystroglycan glycoprotein complex (DGC). A direct interaction between Nde1 and utrophin/dystrophin allows for the assembly of a multi-protein complex that links the cytoskeleton to the extracellular matrix of RGCs to stabilize their lateral membrane, cell-cell adhesion, and radial morphology. Lis1-Nde1 mutations destabilized the DGC and resulted in deformed, disjointed RGCs and disrupted basal lamina. Besides impaired RGC self-renewal and neuronal migration arrests, Lis1-Nde1 deficiencies also led to neuronal over-migration. Additional to phenotypic resemblances of Lis1-Nde1 with DGC, strong synergistic interactions were found between Nde1 and dystroglycan in RGCs. As functional insufficiencies of LIS1, NDE1, and dystroglycan all cause lissencephaly syndromes, our data demonstrated that a three-dimensional regulation of RGC's cytoarchitecture by the Lis1-Nde1-DGC complex determines the number and spatial organization of cortical neurons as well as the size and shape of the cerebral cortex.

Hemorrhage is a leading cause of death in trauma. Tourniquets are effective at controlling extremity hemorrhage and have saved lives. However, tourniquets can cause ischemia reperfusion injury of limbs, leading to systemic inflammation and other adverse effects, which results in secondary damage to the kidney, lung, and liver. A clinically relevant animal model is critical to understanding the pathophysiology of this process and developing therapeutic interventions. Despite the importance of animal models, tourniquet-induced lower limb ischemia/reperfusion (TILLIR) models to date lack a hemorrhage component. We sought to develop a new TILLIR model that included hemorrhage and analyze the subsequent impact on kidney, lung and liver injuries. Four groups of mice were examined: group 1) control, group 2) hemorrhage, group 3) tourniquet application, and group 4) hemorrhage and tourniquet application. The hemorrhagic injury consisted of the removal of 15% of blood volume through the submandibular vein. The tourniquet injury consisted of orthodontic rubber bands applied to the inguinal area bilaterally for 80 min. Mice were then placed in metabolic cages individually for 22 h to collect urine. Hemorrhage alone did not significantly affect transcutaneous glomerular filtration rate (tGFR), blood urea nitrogen (BUN) or urinary kidney injury molecule-1 (KIM-1) levels. Without hemorrhage, TILLIR decreased tGFR by 46%, increased BUN by 162%, and increased KIM-1 by 27% (p < 0.05 for all). With hemorrhage, TILLIR decreased the tGFR by 72%, increased BUN by 395%, and increased urinary KIM-1 by 37% (p < 0.05 for all). These differences were statistically significant (p < 0.05). While hemorrhage had no significant effect on TILLIR-induced renal tubular degeneration and necrosis, it significantly increased TILLIR-induced lung total injury scores and congestion, and fatty liver. In conclusion, hemorrhage exacerbates TILLIR-induced acute kidney injury and structural damage in the lung and liver. [ABSTRACT FROM AUTHOR]

Mutations in the human Filamin A (FLNA) gene disrupt neuronal migration to the cerebral cortex and cause cardiovascular defects. Complete loss of FIna in mice results in embryonic lethality with severe cardiac structural defects involving ventricles, atria, and outflow tracts, as well as widespread aberrant vascular patterning. Despite these widespread developmental defects, migration and motility of many cell types does not appear to be affected. Instead, FIna-null embryos display abnormal epithelial and endothelial organization and aberrant adherens junctions in developing blood vessels, heart, brain, and other tissues. Essential roles for FLNA in intercellular junctions provide a mechanism for the diverse developmental defects seen in patients with FLNA mutations. [ABSTRACT FROM AUTHOR]

LIS1 and nuclear distribution gene E (NudE) are partner proteins in a conserved pathway regulating the function of dynein and microtubules. Here, we present data that cytoplasmic LEKI (cytLEKI), a large protein containing a spectrin repeat and multiple leucine zippers, is a component of this pathway through its direct inter- action with NudE, as determined by a yeast two-hybrid screen. We identified the binding domains in each molecule, and coimmuno-precipitation and colocalization studies confirmed the specificity of the interaction between cytLEKI and NudE. Confocal deconvolution analysis revealed that cytLEKI exhibits colocalization with endogenous NudE and with the known NudE binding partners, LISI and dynein. By localizing the NudE-binding domain of cytLEK1 to a small domain within the molecule, we were able to disrupt cytLEK1 function by using a dominant negative approach in addition to LEK1 knockdown and, thus, examine the role of the cytLEK1-NudE interaction in cells. Consistent with a defect in the LISI pathway, disruption of cytLEK1 function resulted in alteration of microtubule organization and cellular shape. The microtubule network of cells became tightly focused around the nucleus and resulted in a rounded cell shape. Additionally, cells exhibited a severe inability to repolymerize their microtubule networks after nocodazole challenge. Taken together, our studies revealed that cytLEK1 is essential for cellular functions regulated by the LISI pathway. [ABSTRACT FROM AUTHOR]

Filamins were discovered as the first family of non-muscle actin-binding protein. They are lage cytoplasmic proteins that cross-link cortical actin into a dynamic three-dimensional structure. Filamins have also been reported to interact with a large number of cellular proteins of great functional diversity, suggesting that they are unusually versatile signalling scaffolds. More recently, genetic mutations in filamin A and B have been reported to cause a wide range of human diseases, suggesting that different diseases highlight distinct filamin interactions. [ABSTRACT FROM AUTHOR]