A defining feature of embryonic stem cells (ESCs) is the ability to differentiate into all three germ layers. Pluripotency is maintained in part by a unique transcription network that maintains ...expression of pluripotency-specific transcription factors and represses developmental genes. While the mechanisms that establish this transcription network are well studied, little is known of the posttranscriptional surveillance pathways that degrade differentiation-related RNAs. We report that the surveillance pathway mediated by the RNA exosome nuclease complex represses ESC differentiation. Depletion of the exosome expedites differentiation of human ESCs into all three germ layers. LINE-1 retrotransposons and specific miRNAs, lncRNAs, and mRNAs that encode developmental regulators or affect their expression are all bound by the exosome and increase in level upon exosome depletion. The exosome restrains differentiation in part by degrading transcripts encoding FOXH1, a transcription factor crucial for mesendoderm formation. Our studies establish the exosome as a regulator of human ESC differentiation and reveal the importance of RNA decay in maintaining pluripotency.
Direct conversion or reprogramming of human postnatal cells into endothelial cells (ECs), bypassing stem or progenitor cell status, is crucial for regenerative medicine, cell therapy, and ...pathophysiological investigation but has remained largely unexplored.
We sought to directly reprogram human postnatal dermal fibroblasts to ECs with vasculogenic and endothelial transcription factors and determine their vascularizing and therapeutic potential.
We utilized various combinations of 7 EC transcription factors to transduce human postnatal dermal fibroblasts and found that ER71/ETV2 (ETS variant 2) alone best induced endothelial features. KDR
(kinase insert domain receptor) cells sorted at day 7 from ER71/ETV2-transduced human postnatal dermal fibroblasts showed less mature but enriched endothelial characteristics and thus were referred to as early reprogrammed ECs (rECs), and did not undergo maturation by further culture. After a period of several weeks' transgene-free culture followed by transient reinduction of ER71/ETV2, early rECs matured during 3 months of culture and showed reduced ETV2 expression, reaching a mature phenotype similar to postnatal human ECs. These were termed late rECs. While early rECs exhibited an immature phenotype, their implantation into ischemic hindlimbs induced enhanced recovery from ischemia. These 2 rECs showed clear capacity for contributing to new vessel formation through direct vascular incorporation in vivo. Paracrine or proangiogenic effects of implanted early rECs played a significant role in repairing hindlimb ischemia.
This study for the first time demonstrates that ER71/ETV2 alone can directly reprogram human postnatal cells to functional, mature ECs after an intervening transgene-free period. These rECs could be valuable for cell therapy, personalized disease investigation, and exploration of the reprogramming process.
The diencephalon, an integral component of the forebrain, governs a spectrum of crucial functions, ranging from sensory processing to emotional regulation. Yet, unraveling its unique development, ...intricate connectivity, and its role in neurodevelopmental disorders has long been hampered by the scarcity of human brain tissue and ethical constraints. Recent advancements in stem cell technology, particularly the emergence of brain organoids, have heralded a new era in neuroscience research. Although most brain organoid methodologies have hitherto concentrated on directing stem cells toward telencephalic fates, novel techniques now permit the generation of region-specific brain organoids that faithfully replicate precise diencephalic identities. These models mirror the complexity of the human diencephalon, providing unprecedented opportunities for investigating diencephalic development, functionality, connectivity, and pathophysiology
. This review summarizes the development, function, and connectivity of diencephalic structures and touches upon developmental brain disorders linked to diencephalic abnormalities. Furthermore, it presents current diencephalic organoid models and their applications in unraveling the intricacies of diencephalic development, function, and pathology in humans. Lastly, it highlights thalamocortical assembloid models, adept at capturing human-specific aspects of thalamocortical connections, along with their relevance in neurodevelopmental disorders.
Embryonic stem cells (ESCs) maintain pluripotency through unique epigenetic states. When ESCs commit to a specific lineage, epigenetic changes in histones and DNA accompany the transition to ...specialized cell types. Investigating how epigenetic regulation controls lineage specification is critical in order to generate the required cell types for clinical applications. Uhrf1 is a widely known hemi-methylated DNA-binding protein, playing a role in DNA methylation through the recruitment of Dnmt1 and in heterochromatin formation alongside G9a, Trim28, and HDACs. Although Uhrf1 is not essential in ESC self-renewal, it remains elusive how Uhrf1 regulates cell specification. Here we report that Uhrf1 forms a complex with the active trithorax group, the Setd1a/COMPASS complex, to maintain bivalent histone marks, particularly those associated with neuroectoderm and mesoderm specification. Overall, our data demonstrate that Uhrf1 safeguards proper differentiation via bivalent histone modifications.
AbstractA plant (jack bean) extract can function as a urease catalyst, similar to those of certain microorganisms such as Sporosarcina pasteurii, which can decompose urea into carbonate and ammonium ...ions. Carbonate ions decomposed from urea can combine with calcium ions to form calcium carbonate (usually calcite mineral). In this study, calcium chloride, calcium hydroxide, or calcium nitrate was added to urea solution with plant extract, and then mixed with Nakdong River sand to precipitate calcite within the sand-grain matrix. The mixed sand was compacted into a cylindrical specimen and cured for 3 days at room temperature. A specimen without plant extract was prepared for comparison. A series of unconfined compression tests and analytical methods was carried out on sand treated with various amounts of urea and different calcium sources. With increasing amounts of urea, the unconfined compressive strength (UCS) of the sand treated with plant extract increased 10-fold up to 317 kPa, compared to that of the sand without plant extract. Its strength was similar to that of 4% cemented sand with high-early strength portland cement. The strength of the specimen containing calcium chloride was higher than that with calcium hydroxide or calcium nitrate.
There is wide variability in the propensity of somatic cells to reprogram into pluripotency in response to the Yamanaka factors. How to segregate these variabilities to enrich for cells of specific ...traits that reprogram efficiently remains challenging. Here we report that the variability in reprogramming propensity is associated with the activity of the MKL1/SRF transcription factor and concurs with small cell size as well as rapid cell cycle. Reprogramming progressive cells can be prospectively identified by their low activity of a widely used synthetic promoter, CAG. CAGlow cells arise and expand during cell cycle acceleration in the early reprogramming culture of both mouse and human fibroblasts. Our work illustrates a molecular scenario underlying the distinct reprogramming propensities and demonstrates a convenient practical approach for their enrichment.
Extensive cellular heterogeneity exists in cultures undergoing Yamanaka reprogramming. The ability to identify reprogramming progressive cells has practical and mechanistic implications. We report that reprogramming progressive cells can be conveniently identified by the low activity of a widely used synthetic promoter, the CAG promoter. CAGlow cells are small in size, cycle fast and display low endogenous MKL1/SRF activity.
Induced pluripotent stem cells (iPSCs) generated from somatic cells of patients can be used to model different human diseases. They may also serve as sources of transplantable cells that can be used ...in novel cell therapies. Here, we analyzed neuronal properties of an iPSC line derived from a patient with a juvenile form of Huntington's disease (HD) carrying 72 CAG repeats (HD‐iPSC). Although its initial neural inducing activity was lower than that of human embryonic stem cells, we found that HD‐iPSC can give rise to GABAergic striatal neurons, the neuronal cell type that is most susceptible to degeneration in HD. We then transplanted HD‐iPSC‐derived neural precursors into a rat model of HD with a unilateral excitotoxic striatal lesion and observed a significant behavioral recovery in the grafted rats. Interestingly, during our in vitro culture and when the grafts were examined at 12 weeks after transplantation, no aggregate formation was detected. However, when the culture was treated with a proteasome inhibitor (MG132) or when the cells engrafted into neonatal brains were analyzed at 33 weeks, there were clear signs of HD pathology. Taken together, these results indicate that, although HD‐iPSC carrying 72 CAG repeats can form GABAergic neurons and give rise to functional effects in vivo, without showing an overt HD phenotype, it is highly susceptible to proteasome inhibition and develops HD pathology at later stages of transplantation. These unique features of HD‐iPSC will serve as useful tools to study HD pathology and develop novel therapeutics. Stem Cells2012;30:2054–2062
Display omitted
•CO2/microalgae were converted into both syngas and biochar.•CO2 expedited a reaction with VOCs, resulting in enhanced syngas formation.•Biochar produced from pyrolysis showed a ...catalytic capability for pyrolysis.•Ni impregnated biochar significantly promoted CO and H2 production.
This study proposes a sustainable waste-to-energy/biochar platform using a toxic microalgal biomass waste. In particular, CO2-feeding pyrolysis of Microcystis aeruginosa (M. aeruginosa) waste was investigated, focusing on the analysis of gaseous pyrolysates and properties of biochar with a construction of mass balance. Also, the catalytic capability of biochar produced from M. aeruginosa was explored to reinforce the mechanistic impact of CO2 on the pyrolysis process within the overall process level. Ni impregnated biochar composite was further synthesized and used as a catalyst to promote syngas formation in the CO2-feeding pyrolysis process of M. aeruginosa.
Transcriptomic changes in specific brain regions can influence the risk of alcohol use disorder (AUD), but the underlying mechanism is not fully understood. We investigated AUD-associated miRNA-mRNA ...regulatory networks in multiple brain regions by analyzing transcriptomic changes in two sets of postmortem brain tissue samples and ethanol-exposed human embryonic stem cell (hESC)-derived cortical interneurons. miRNA and mRNA transcriptomes were profiled in 192 tissue samples (Set 1) from eight brain regions (amygdala, caudate nucleus, cerebellum, hippocampus, nucleus accumbens, prefrontal cortex, putamen, and ventral tegmental area) of 12 AUD and 12 control European Australians. Nineteen differentially expressed miRNAs (fold-change>2.0 & P < 0.05) and 97 differentially expressed mRNAs (fold-change>2.0 & P < 0.001) were identified in one or multiple brain regions of AUD subjects. AUD-associated miRNA-mRNA regulatory networks in each brain region were constructed using differentially expressed and negatively correlated miRNA-mRNA pairs. AUD-relevant pathways (including CREB Signaling, IL-8 Signaling, and Axonal Guidance Signaling) were potentially regulated by AUD-associated brain miRNA-mRNA pairs. Moreover, miRNA and mRNA transcriptomes were mapped in additional 96 tissue samples (Set 2) from six of the above eight brain regions of eight AUD and eight control European Australians. Some of the AUD-associated miRNA-mRNA regulatory networks were confirmed. In addition, miRNA and mRNA transcriptomes were analyzed in hESC-derived cortical interneurons with or without ethanol exposure, and ethanol-influenced miRNA-mRNA regulatory networks were constructed. This study provided evidence that alcohol could induce concerted miRNA and mRNA expression changes in reward-related or alcohol-responsive brain regions. We concluded that altered brain miRNA-mRNA regulatory networks might contribute to AUD development.
BACKGROUND:Human pluripotent stem cell (hPSC)–derived endothelial cells (ECs) have limited clinical utility because of undefined components in the differentiation system and poor cell survival in ...vivo. Here, we aimed to develop a fully defined and clinically compatible system to differentiate hPSCs into ECs. Furthermore, we aimed to enhance cell survival, vessel formation, and therapeutic potential by encapsulating hPSC-ECs with a peptide amphiphile (PA) nanomatrix gel.
METHODS:We induced differentiation of hPSCs into the mesodermal lineage by culturing on collagen-coated plates with a glycogen synthase kinase 3β inhibitor. Next, vascular endothelial growth factor, endothelial growth factor, and basic fibroblast growth factor were added for endothelial lineage differentiation, followed by sorting for CDH5 (VE-cadherin). We constructed an extracellular matrix–mimicking PA nanomatrix gel (PA-RGDS) by incorporating the cell adhesive ligand Arg-Gly-Asp-Ser (RGDS) and a matrix metalloproteinase-2–degradable sequence. We then evaluated whether the encapsulation of hPSC-CDH5 cells in PA-RGDS could enhance long-term cell survival and vascular regenerative effects in a hind-limb ischemia model with laser Doppler perfusion imaging, bioluminescence imaging, real-time reverse transcription–polymerase chain reaction, and histological analysis.
RESULTS:The resultant hPSC-derived CDH5 cells (hPSC-ECs) showed highly enriched and genuine EC characteristics and proangiogenic activities. When injected into ischemic hind limbs, hPSC-ECs showed better perfusion recovery and higher vessel-forming capacity compared with media-, PA-RGDS–, or human umbilical vein EC–injected groups. However, the group receiving the PA-RGDS–encapsulated hPSC-ECs showed better perfusion recovery, more robust and longer cell survival (> 10 months), and higher and prolonged angiogenic and vascular incorporation capabilities than the bare hPSC-EC–injected group. Surprisingly, the engrafted hPSC-ECs demonstrated previously unknown sustained and dynamic vessel-forming behaviorinitial perivascular concentration, a guiding role for new vessel formation, and progressive incorporation into the vessels over 10 months.
CONCLUSIONS:We generated highly enriched hPSC-ECs via a clinically compatible system. Furthermore, this study demonstrated that a biocompatible PA-RGDS nanomatrix gel substantially improved long-term survival of hPSC-ECs in an ischemic environment and improved neovascularization effects of hPSC-ECs via prolonged and unique angiogenic and vessel-forming properties. This PA-RGDS–mediated transplantation of hPSC-ECs can serve as a novel platform for cell-based therapy and investigation of long-term behavior of hPSC-ECs.
PDF
