Development Growth & Differentiation最新文献

From September 16 to 19, 2024, an international symposium to celebrate the centennial of the discovery of the gastrula organizer by Hans Spemann and Hilde Mangold, was held at the University of Freiburg, Germany, where they studied embryology. There were 41 plenary lectures, 11 short talks, and 182 poster presentations, with more than 300 participants from 23 countries. The symposium covered research topics broadly related to developmental, cell, genome, and evolutionary biology, mainly focused on early animal development. In addition to in vivo studies on topics such as gastrulation, embryonic patterning, cell polarity, and morphogenesis, recent studies using gastruloids and organoids, which recapitulate embryogenesis and organogenesis in in vitro cell culture, were also presented at this symposium, entitled Self-Organization in Biology. Most of the reported studies used vertebrate models such as mice, frogs, and zebrafish; however, evolutionary studies involving invertebrate and plant models were also presented. Presentations employing traditional methods such as cell transplantation and phenotype screening, and state-of-the-art technologies such as single-cell omics, high-resolution imaging, and computational analysis showed that experimental embryology has a long history, to which studies of the organizer have contributed significantly. Here we discuss memorable aspects of the symposium in the hope that this report will encourage young scientists to actively participate in face-to-face international conferences.

5′Hox genes regulate pattern formation along the axes of the limb. Previously, we showed that Hoxa13/Hoxd13 double-mutant newts lacked all digits of the forelimbs during development and regeneration, showing that newt Hox13 is necessary for digit formation in development and regeneration. In addition, we found another unique phenotype. Some of the Hox13 crispant newts showed hindlimb defects, in which whole or almost whole hindlimbs were lost, suggesting a novel function of Hox13 in limb development. Using germline mutants, we showed that mutation in Hox13 led to hindlimb defects. The limb buds of Hox13 crispants formed, however, did not show outgrowth. Expression of Fgf10 and Tbx4, which are involved in limb outgrowth, decreased in the hindlimb buds of Hox13 crispants. In addition, hindlimb defects were observed in both Fgf10 and Tbx4 crispant newts. Finally, Fgf10 and Tbx4 interacted with Hox13 genetically. Our results revealed a novel function of Hox13 in regulating the outgrowth of the newt hindlimb bud through interaction with Fgf10 and Tbx4.

Transcription factors collaborate with epigenetic regulatory factors to orchestrate cardiac differentiation for heart development, but the underlying mechanism is not fully understood. Here, we report that GATA-6 induces cardiac differentiation but peroxisome proliferator-activated receptor α (PPARα) reverses GATA-6-induced cardiac differentiation, possibly because GATA-6/PPARα recruits the polycomb protein complex containing EZH2/Ring1b/BMI1 to the promoter of the cardiac-specific α-myosin heavy chain (α-MHC) gene and suppresses α-MHC expression, which ultimately inhibits cardiac differentiation. Furthermore, Ring1b ubiquitylates PPARα and GATA-6. By overexpression and knockout of EZH2/BMI1, it was demonstrated that the polycomb protein complex inhibits cardiac differentiation induced by GATA-6 and PPARα. Together, our results demonstrate that the polycomb protein complex interacts with GATA-6/PPARα to inhibit cardiac differentiation, a finding that could facilitate the development of new therapies for congenital heart disease.

In the embryonic neuroepithelium (NE), neural progenitor cells undergo cell cycle-dependent interkinetic nuclear migration (IKNM) along the apicobasal axis. Extensive IKNM supports increasing cell production rates per unit apical surface, as typically observed in the mammalian telencephalic NE. Apical nucleokinesis during the G2 phase is an essential premitotic event, but its occurrence has not yet been quantitatively analyzed at a large 3D-scale with sufficient spatiotemporal resolution. Here, we comprehensively analyzed apically migrating nuclei/somata in reference to their surroundings from embryonic day (E)11 to E13 in the mouse telencephalon. The velocity of apical nucleokinesis decreased, with more frequent nuclear pausing occurring at E12 and E13, whereas the nuclear density in the middle NE zone (20–40-μm deep) increased. This result, together with the results of Shh-mediated overproliferation experiments in which the nuclear density was increased in vivo at E11, suggests that apical nucleokinesis is physically influenced by the surrounding nuclei. Mean square displacement analysis for nuclei being passed by the apically migrating nuclei via horizontal sectioning in toto-recorded movies revealed that the “tissue fluidity” or physical permissiveness of the NE to apical nucleokinesis gradually decreased (E11 > E12 > E13). To further investigate the spatial relationship between preexisting mitoses and subsequent premitotic apical nucleokinesis, the horizontal distribution of mitoses was cumulatively (~3 hr) analyzed under in toto monitoring. The four-dimensional cumulative apical mitoses presented a “random”, not “clustered” or “regular”, distribution pattern throughout the period examined. These methodologies provide a basis for future comparative studies of interspecies differences.

Avian species are essential resources for human society, with their preservation and utilization heavily dependent on primordial germ cells (PGCs). However, efficient methods for isolating live PGCs from embryos remain elusive in avian species beyond chickens, and even in chickens, existing techniques have shown limited efficiency. In this study, we present a rapid, simple, and cost-effective method for labeling and sorting circulating-stage PGCs across various avian species, including Carinatae and Ratitae, using Lycopersicon Esculentum (Tomato) lectin (LEL). Notably, this method demonstrates high sorting efficiency by identifying a wide range of PGC subtypes while preserving the proliferative and migratory potential of chicken PGCs. This approach is anticipated to significantly contribute to the conservation, research, and agricultural industries related to avian species globally.

Cardiovascular disease is the leading cause of mortality worldwide. Myocardial injury resulting from ischemia can be fatal because of the limited regenerative capacity of adult myocardium. Mammalian cardiomyocytes rapidly lose their proliferative capacities, with only a small fraction of adult myocardium remaining proliferative, which is insufficient to support post-injury recovery. Recent investigations have revealed that this decline in myocardial proliferative capacity is closely linked to perinatal metabolic shifts. Predominantly glycolytic fetal myocardial metabolism transitions towards mitochondrial fatty acid oxidation postnatally, which not only enables efficient production of ATP but also causes a dramatic reduction in cardiomyocyte proliferative capacity. Extensive research has elucidated the mechanisms behind this metabolic shift, as well as methods to modulate these metabolic pathways. Some of these methods have been successfully applied to enhance metabolic reprogramming and myocardial regeneration. This review discusses recently acquired insights into the interplay between metabolism and myocardial proliferation, emphasizing postnatal metabolic transitions.

Mitochondria are unique organelles that have their own genome (mtDNA) and perform various pivotal functions within a cell. Recently, evidence has highlighted the role of mitochondria in the process of stem cell differentiation, including differentiation of neural stem cells (NSCs). Here we studied the importance of mtDNA function in the early differentiation process of NSCs in two cell culture models: the CGR8-NS cell line that was derived from embryonic stem cells by a lineage selection technique, and primary NSCs that were isolated from embryonic day 14 mouse fetal forebrain. We detected a dramatic increase in mtDNA content upon NSC differentiation to adapt their mtDNA levels to their differentiated state, which was not accompanied by changes in mitochondrial transcription factor A expression. As chemical mtDNA depletion by ethidium bromide failed to generate living ρ° cell lines from both NSC types, we used inhibition of mtDNA polymerase-γ by 2′-3′-dideoxycytidine to reduce mtDNA replication and subsequently cellular mtDNA content. Inhibition of mtDNA replication upon NSC differentiation reduced neurogenesis but not gliogenesis. The mtDNA depletion did not change energy production/consumption or cellular reactive oxygen species (ROS) content in the NSC model used. In conclusion, mtDNA replication is essential for neurogenesis but not gliogenesis in fetal NSCs through as yet unknown mechanisms, which, however, are largely independent of energy/ROS metabolism.

Both development and regeneration depend on signaling centers, which are sources of locally secreted tissue-patterning molecules. As many signaling centers are decommissioned before the end of embryogenesis, a fundamental question is how signaling centers can be re-induced later in life to promote regeneration after injury. Here, we use the axolotl salamander model (Ambystoma mexicanum) to address how the floor plate is assembled for spinal cord regeneration. The floor plate is an archetypal vertebrate signaling center that secretes Shh ligand and patterns neural progenitor cells during embryogenesis. Unlike mammals, axolotls continue to express floor plate genes (including Shh) and downstream dorsal–ventral patterning genes in their spinal cord throughout life, including at steady state. The parsimonious hypothesis that Shh+ cells give rise to functional floor plate cells for regeneration had not been tested. Using HCR in situ hybridization and mathematical modeling, we first quantified the behaviors of dorsal–ventral spinal cord domains, identifying significant increases in gene expression level and floor plate size during regeneration. Next, we established a transgenic axolotl to specifically label and fate map Shh+ cells in vivo. We found that labeled Shh+ cells gave rise to regeneration floor plate, and not to other neural progenitor domains, after tail amputation. Thus, despite changes in domain size and downstream patterning gene expression, Shh+ cells retain their floor plate identity during regeneration, acting as a stable cellular source for this regeneration signaling center in the axolotl spinal cord.

The development of new technologies opens new avenues in the research field. Gene knockout is a key method for analyzing gene function in mice. Currently, conditional gene knockout strategies are employed to examine temporal and spatial gene function. However, phenotypes are sometimes not observed because of the time required for depletion due to the long half-life of the target proteins. Protein knockdown using an improved auxin-inducible degron system, AID2, overcomes such difficulties owing to rapid and efficient target depletion. We observed depletion of AID-tagged proteins within a few to several hours by a simple intraperitoneal injection of the auxin analog, 5-Ph-IAA, which is much shorter than the time required for target depletion using conditional gene knockout. Importantly, the loss of protein is reversible, making protein knockdown useful to measure the effects of transient loss of protein function. Here, we also established several mouse lines useful for AID2-medicated protein knockdown, which include knock-in mouse lines in the ROSA26 locus; one expresses TIR1(F74G), and the other is the reporter expressing AID-mCherry. We also established a germ-cell-specific TIR1 line and confirmed the protein knockdown specificity. In addition, we introduced an AID tag to an endogenous protein, DCP2 via the CAS9-mediated gene editing method. We confirmed that the protein was effectively eliminated by TIR1(F74G), which resulted in the similar phenotype observed in knockout mouse within 20 h.