Molecular and cellular dynamics of the developing human neocortex at single-cell resolution.

The development of the human neocortex is a highly dynamic process and involves complex cellular trajectories controlled by cell-type-specific gene regulation1. Here, we collected paired single-nucleus chromatin accessibility and transcriptome data from 38 human neocortical samples encompassing both the prefrontal cortex and primary visual cortex. These samples span five main developmental stages, ranging from the first trimester to adolescence. In parallel, we performed spatial transcriptomic analysis on a subset of the samples to illustrate spatial organization and intercellular communication. This atlas enables us to catalog cell type-, age-, and area-specific gene regulatory networks underlying neural differentiation. Moreover, combining single-cell profiling, progenitor purification, and lineage-tracing experiments, we have untangled the complex lineage relationships among progenitor subtypes during the transition from neurogenesis to gliogenesis in the human neocortex. We identified a tripotential intermediate progenitor subtype, termed Tri-IPC, responsible for the local production of GABAergic neurons, oligodendrocyte precursor cells, and astrocytes. Remarkably, most glioblastoma cells resemble Tri-IPCs at the transcriptomic level, suggesting that cancer cells hijack developmental processes to enhance growth and heterogeneity. Furthermore, by integrating our atlas data with large-scale GWAS data, we created a disease-risk map highlighting enriched ASD risk in second-trimester intratelencephalic projection neurons. Our study sheds light on the gene regulatory landscape and cellular dynamics of the developing human neocortex.

Vitamin D Deficiency Accelerates Coronary Artery Disease Progression in Swine.

OBJECTIVE: The role of vitamin D deficiency in coronary artery disease (CAD) progression is uncertain. Chronic inflammation in epicardial adipose tissue (EAT) has been implicated in the pathogenesis of CAD. However, the molecular mechanism underlying vitamin D deficiency-enhanced inflammation in the EAT of diseased coronary arteries remains unknown. We examined a mechanistic link between 1,25-dihydroxyvitamin D-mediated suppression of nuclear factor-κB (NF-κB) transporter, karyopherin α4 (KPNA4) expression and NF-κB activation in preadipocytes. Furthermore, we determined whether vitamin D deficiency accelerates CAD progression by increasing KPNA4 and nuclear NF-κB levels in EAT. APPROACH AND RESULTS: Nuclear protein levels were detected by immunofluorescence and Western blot. Exogenous KPNA4 was transported into cells by a transfection approach and constituted lentiviral vector. Swine were administered vitamin D-deficient or vitamin D-sufficient hypercholesterolemic diet. After 1 year, the histopathology of coronary arteries and nuclear protein expression of EAT were assessed. 1,25-dihydroxyvitamin D inhibited NF-κB activation and reduced KPNA4 levels through increased vitamin D receptor expression. Exogenous KPNA4 rescued 1,25-dihydroxyvitamin D-dependent suppression of NF-κB nuclear translocation and activation. Vitamin D deficiency caused extensive CAD progression and advanced atherosclerotic plaques, which are linked to increased KPNA4 and nuclear NF-κB levels in the EAT. CONCLUSIONS: 1,25-dihydroxyvitamin D attenuates NF-κB activation by targeting KPNA4. Vitamin D deficiency accelerates CAD progression at least, in part, through enhanced chronic inflammation of EAT by upregulation of KPNA4, which enhances NF-κB activation. These novel findings provide mechanistic evidence that vitamin D supplementation could be beneficial for the prevention and treatment of CAD.

FOXO1 Mediates Vitamin D Deficiency-Induced Insulin Resistance in Skeletal Muscle.

Prospective epidemiological studies have consistently shown a relationship between vitamin D deficiency, insulin resistance, and type 2 diabetes mellitus (DM2). This is supported by recent trials showing that vitamin D supplementation in prediabetic or insulin-resistant patients with inadequate vitamin D levels improves insulin sensitivity. However, the molecular mechanisms underlying vitamin D deficiency-induced insulin resistance and DM2 remain unknown. Skeletal muscle insulin resistance is a primary defect in the majority of patients with DM2. Although sustained activation of forkhead box O1 (FOXO1) in skeletal muscle causes insulin resistance, a relationship between vitamin D deficiency and FOXO1 activation in muscle is unknown. We generated skeletal muscle-specific vitamin D receptor (VDR)-null mice and discovered that these mice developed insulin resistance and glucose intolerance accompanied by increased expression and activity of FOXO1. We also found sustained FOXO1 activation in the skeletal muscle of global VDR-null mice. Treatment of C2C12 muscle cells with 1,25-dihydroxyvitamin D (VD3) reduced FOXO1 expression, nuclear translocation, and activity. The VD3-dependent suppression of FOXO1 activation disappeared by knockdown of VDR, indicating that it is VDR-dependent. Taken together, these results suggest that FOXO1 is a critical target mediating VDR-null signaling in skeletal muscle. The novel findings provide the conceptual support that persistent FOXO1 activation may be responsible for insulin resistance and impaired glucose metabolism in vitamin D signaling-deficient mice, as well as evidence for the utility of vitamin D supplementation for intervention in DM2.

A role for p38 mitogen-activated protein kinase and c-myc in endothelin-dependent rat aortic smooth muscle cell proliferation.

We have demonstrated recently that endothelin (ET) stimulates rat aortic smooth muscle cell proliferation through an extracellular signal-regulated kinase (ERK)-dependent mechanism. Approximately 70% of ET-dependent [3H]-thymidine incorporation in these cells signals through this system. In the present study, we show that the residual mitogenic activity requires an intact p38 mitogen-activated protein kinase (p38 MAPK) system and increased c-myc gene expression. ET increased [3H]-thymidine incorporation in rat aortic smooth muscle cells approximately 5-fold. p38 MAPK inhibition with SB203580 or ERK/ERK kinase inhibition with PD98059 each effected approximately 70% inhibition in ET-dependent DNA synthesis, whereas the combination led to nearly complete blockade of the ET effect. ET also increased c-myc RNA levels and c-Myc protein levels in these cells. The increment in c-Myc expression was blocked by SB203580 but not by PD98059. Use of antisense oligonucleotides directed against the translation start site of the c-myc transcript, but not scrambled oligonucleotide sequence, resulted in approximately 60% decrease in ET-dependent [3H]-thymidine incorporation. The combination of antisense c-myc and PD98059 resulted in near complete inhibition of ET-dependent DNA synthesis. Both ET and c-Myc increased expression and promoter activity of E2F, a transcription factor that has been linked to enhanced cell cycle activity. The ET-dependent increment in E2F promoter activity was suppressed after treatment with SB203580 or antisense c-myc but not by PD98059 or a scrambled oligonucleotide sequence. Collectively, these findings demonstrate that ET uses 2 complementary signal transduction cascades (ERK and p38 MAPK) to control proliferative activity of vascular smooth muscle cells.