The anterior pituitary gland plays a central role in regulating various physiological processes, including body growth, reproduction, metabolism and stress response. Here, we perform single-cell RNA-sequencing (scRNA-seq) of 4113 individual cells from human fetal pituitaries. We characterize divergent developmental trajectories with distinct transitional intermediate states in five hormone-producing cell lineages. Corticotropes exhibit an early intermediate state prior to full differentiation. Three cell types of the PIT-1 lineage (somatotropes, lactotropes and thyrotropes) segregate from a common progenitor coexpressing lineage-specific transcription factors of different sublineages. Gonadotropes experience two multistep developmental trajectories. Furthermore, we identify a fetal gonadotrope cell subtype expressing the primate-specific hormone chorionic gonadotropin. We also characterize the cellular heterogeneity of pituitary stem cells and identify a hybrid epithelial/mesenchymal state and an early-to-late state transition. Here, our results provide insights into the transcriptional landscape of human pituitary development, defining distinct cell substates and subtypes and illustrating transcription factor dynamics during cell fate commitment.

Brain drug delivery is one of the most important bottlenecks in the development of drugs for the central nervous system. Cumulative evidence has emerged that extracellular vesicles (EVs) play a key role in intercellular communication. Exosomes, a subgroup of EVs, have received the most attention due to their capability in mediating the horizontal transfer of their bioactive inclusions to neighboring and distant cells, and thus specifically regulating the physiological and pathological functions of the recipient cells. This native and unique signaling mechanism confers exosomes with great potential to be developed into an effective, precise, and safe drug delivery system. Here, we provide an overview into the challenges of brain drug delivery and the function of exosomes in the brain under physiological and pathological conditions, and discuss how these natural vesicles could be harnessed for brain drug delivery and for the therapy of brain diseases.

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FSHD is characterized by the misexpression of DUX4 in skeletal muscle. Although DUX4 upregulation is thought to be the pathogenic cause of FSHD, DUX4 is lowly expressed in patient samples, and analysis of the consequences of DUX4 expression has largely relied on artificial overexpression. To better understand the native expression profile of DUX4 and its targets, we performed bulk RNA-seq on a 6-day differentiation time-course in primary FSHD2 patient myoblasts. We identify a set of 54 genes upregulated in FSHD2 cells, termed FSHD-induced genes. Using single-cell and single-nucleus RNA-seq on myoblasts and differentiated myotubes, respectively, we captured, for the first time, DUX4 expressed at the single-nucleus level in a native state. We identified two populations of FSHD myotube nuclei based on low or high enrichment of DUX4 and FSHD-induced genes ("FSHD-Lo" and "FSHD Hi", respectively). FSHD-Hi myotube nuclei coexpress multiple DUX4 target genes including DUXA, LEUTX and ZSCAN4, and also upregulate cell cycle-related genes with significant enrichment of E2F target genes and p53 signaling activation. We found more FSHD-Hi nuclei than DUX4-positive nuclei, and confirmed with in situ RNA/protein detection that DUX4 transcribed in only one or two nuclei is sufficient for DUX4 protein to activate target genes across multiple nuclei within the same myotube. DUXA (the DUX4 paralog) is more widely expressed than DUX4, and depletion of DUXA suppressed the expression of LEUTX and ZSCAN4 in late, but not early, differentiation. The results suggest that the DUXA can take over the role of DUX4 to maintain target gene expression. These results provide a possible explanation as to why it is easier to detect DUX4 target genes than DUX4 itself in patient cells and raise the possibility of a self-sustaining network of gene dysregulation triggered by the limited DUX4 expression.

Background The births of more than 8 million infants have been enabled globally through assisted reproductive technologies (ARTs), including conventional in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) with either fresh embryo transfer (ET) or frozen embryo transfer (FET). However, the safety issue regarding ARTs has drawn growing attention with accumulating observations of rising health risks, and underlying epigenetic mechanisms are largely uncharacterized. Methods In order to clarify epigenetic risks attributable to ARTs, we profiled DNA methylome on 137 umbilical cord blood (UCB) and 158 parental peripheral blood (PPB) samples, histone modifications (H3K4me3, H3K4me1, H3K27me3 and H3K27ac) on 33 UCB samples and transcriptome on 32 UCB samples by reduced representation bisulfite sequencing (RRBS), chromatin immunoprecipitation sequencing (ChIP-seq), and RNA sequencing (RNA-seq), respectively. Findings We revealed that H3K4me3 was the most profoundly impacted by ICSI and freeze-thawing operation compared with the other three types of histone modifications. IVF-ET seemed to introduce less disturbance into infant epigenomes than IVF-FET or ICSI-ET did. ARTs also decreased the similarity of DNA methylome within twin pairs, and we confirmed that ART per se would introduce conservative changes locally through removal of parental effect. Importantly, those unique and common alterations induced by different ART procedures were highly enriched in the processes related to nervous system, cardiovascular system and glycolipid metabolism etc., which was in accordance with those findings in previous epidemiology studies and suggested some unexplored health issues, including in the immune system and skeletal system. Interpretation Different ART procedures can induce local and functional epigenetic abnormalities, especially for DNA methylation and H3K4me3, providing an epigenetic basis for the potential long-term health risks in ART-conceived offspring. Funding sources This study was funded by National Natural Science Foundation of China (81,730,038; 81,521,002), National Key Research and Development Program (2018YFC1004000; 2017YFA0103801; 2017YFA0105001) and Strategic Priority Research Program of the Chinese Academy of Sciences (XDA16020703). Yang Wang was supported by Postdoctoral Fellowship of Peking-Tsinghua Center for Life Science.

A third of the population sustains a bone fracture, and the pace of fracture healing slows with age. The slower pace of repair is responsible for the increased morbidity in older individuals who sustain a fracture. Bone healing progresses through overlapping phases, initiated by cells of the monocyte/macrophage lineage. The repair process ends with remodeling. This last phase is controlled by osteoclasts, which are bone-specific multinucleated cells also of the monocyte/macrophage lineage. The slower rate of healing in aging can be rejuvenated by macrophages from young animals, and secreted proteins from macrophage regulate undifferentiated mesenchymal cells to become bone-forming osteoblasts. Macrophages can derive from fetal erythromyeloid progenitors or from adult hematopoietic progenitors. Recent studies show that fetal erythromyeloid progenitors are responsible for the osteoclasts that form the space in bone for hematopoiesis and the fetal osteoclast precursors reside in the spleen postnatally, traveling through the blood to participate in fracture repair. Differences in secreted proteins between macrophages from old and young animals regulate the efficiency of osteoblast differentiation from undifferentiated mesenchymal precursor cells. Interestingly, during the remodeling phase osteoclasts can form from the fusion between monocyte/macrophage lineage cells from the fetal and postnatal precursor populations. Data from single cell RNA sequencing identifies specific markers for populations derived from the different precursor populations, a finding that can be used in future studies. Here, we review the diversity of macrophages and osteoclasts, and discuss recent finding about their developmental origin and functions, which provides novel insights into their roles in bone homeostasis and repair.

The majority of Alzheimer’s disease (AD) cases are late-onset and occur sporadically, however most mouse models of the disease harbor pathogenic mutations, rendering them better representations of familial autosomal-dominant forms of the disease. Here, we generated knock-in mice that express wildtype human Aβ under control of the mouse App locus. Remarkably, changing 3 amino acids in the mouse Aβ sequence to its wild-type human counterpart leads to age-dependent impairments in cognition and synaptic plasticity, brain volumetric changes, inflammatory alterations, the appearance of Periodic Acid-Schiff (PAS) granules and changes in gene expression. In addition, when exon 14 encoding the Aβ sequence was flanked by loxP sites we show that Cre-mediated excision of exon 14 ablates hAβ expression, rescues cognition and reduces the formation of PAS granules.

Proper development of fetal germ cells (FGCs) is vital for the precise transmission of genetic and epigenetic information through generations. The transcriptional landscapes of human FGC development have been revealed; however, the epigenetic reprogramming process of FGCs remains elusive. Here, we profiled the genome-wide DNA methylation and chromatin accessibility of human FGCs at different phases as well as gonadal niche cells at single-cell resolution. First, we found that DNA methylation levels of FGCs changed in a temporal manner, whereas FGCs at different phases in the same embryo exhibited comparable DNA methylation levels and patterns. Second, we revealed the phase-specific chromatin accessibility signatures at the promoter regions of a large set of critical transcription factors and signaling pathway genes. We also identified potential distal regulatory elements including enhancers in FGCs. Third, compared with other hominid-specific retrotransposons, SVA_D might have a broad spectrum of binding capacity for transcription factors, including SOX15 and SOX17. Finally, using an in vitro culture system of human FGCs, we showed that the BMP signaling pathway promoted the cell proliferation of FGCs, and regulated the WNT signaling pathway by orchestrating the chromatin accessibility of its ligand genes. Our single-cell epigenomic atlas and functional assays provide valuable insights for understanding the strongly heterogeneous, unsynchronized, yet highly robust nature of human germ cell development.