As a possible target for pro- or anti-angiogenic therapy, the process of angiogenesis has become more widely recognized in disease and is essential in normal growth and development. The current study examined the immunological activities of macrophages during angiogenesis in quail embryos. CD68 was used to identify perivascular macrophages, which also retained expression of some stem cell markers, including CD34 and CD117, and also CD21. The macrophages also expressed vascular endothelial growth factor (VEGF), a key promoter of angiogenesis, and matrix metalloproteinase-9 (MMP-9), which degrades matrix components to facilitate the formation of new blood vessels. The growth factor bone morphogenetic protein (BMP) and platelet-derived growth factor receptor-alpha (PDGF-α), which are often associated with mesenchymal cell recruitment and vascularization, were also expressed in the perivascular macrophages. More recently, studies investigating macrophages have become more prominent due to their proposed effects on endothelial cells. Macrophages are also increasingly considered potential targets for pro- or anti-angiogenic therapies, and this research provides insights into protein expression and roles in angiogenesis.

Archaeopteryx, the iconic 150-million-year-old basal bird, lies at the heart of the debate on the origin of avian flight. While its asymmetric wing feathers indicate some flight capability, its limited shoulder mobility and reduced feather asymmetry suggest it lacked the capacity for the single, powerful leap used by modern birds for rapid take-off. Observations of extant birds reveal that many species, including corvids, employ multiple bipedal leaps with minimal wing assistance to gradually accelerate during take-off, particularly in low-stress contexts. Given Archaeopteryx's robust hindlimbs, we hypothesized that it could have used a similar multi-leap strategy to become airborne. We therefore tested the hypothesis that Archaeopteryx could generate sufficient take-off velocity via multiple hops or leaps. To test this, we developed a biomechanical model based on experimental take-off data from living birds, adapted to Archaeopteryx's anatomy. By analysing joint moments at the hip, knee, and ankle, alongside muscle capacity, we estimated its take-off velocity. Our results demonstrate that Archaeopteryx could have achieved its minimum sustainable flight speed in as few as two to three leaps, without requiring the energetically demanding single leap of modern birds. This study provides the first quantitative support for a ground-up, multi-leap take-off mechanism in early birds, distinct from the single-leap strategy usually observed today. We propose that the modern avian take-off may have evolved from this primitive multi-leap behaviour, offering new insights into the biomechanical transition from terrestrial locomotion to powered flight.

Translating the bioelectric code remains one of the core challenges to widespread biomedical translation of bioelectric interventions, as well as a better evolutionary understanding of how developmental ionic signaling evolved to become the basis of neural intelligence. To advance our understanding, it is crucial to develop model systems that allow simultaneous, quantitative study of diverse bioelectric parameters in living states, which must also be linked to cell- and tissue-level outcomes. Here, we apply quantitative Fluorescent Lifetime Imaging (FLIM) optical estimation of membrane potential (Vmem[oe]) to map the bioelectric dynamics of spreading Xenopus laevis neural crest explants (NCEs) over > 17 hours. We identify a slow hours-scale bioelectric component and distinct faster minutes-scale and seconds-scale components. These dynamics often span multiple cells, consistent with roles in the collective behavior of NCEs. We then use information theory to show that minutes-scale NCE Vmem[oe] dynamics are largely distinct from calcium dynamics. Finally, we provide a survey of diverse bioelectric events revealing a deep complexity in collective bioelectric dynamics, likely involving tunneling nanotubes in their transmission, which suggests numerous avenues for further investigation. These combined data demonstrate FLIM as a powerful tool for observing subtle bioelectric signals at the large temporal and spatial scale needed for developmental biology studies.

Vascularization of the central nervous system and the postnatal retina proceeds through angiogenic growth that relies in part on the basement membrane (BM) to support endothelial cell-cell interactions and maintain barrier integrity during vessel formation. In addition to the core structural components of the BM, matricellular proteins are present as minor constituents, however, their contributions to endothelial cohesion and barrier stability during angiogenesis remain poorly defined. Here, we identify the matricellular protein Fibulin-1 (Fbln1), a regulator of cell-matrix interactions, as an essential factor during mid-gestation that prevents the formation of dilated, tortuous capillaries that give rise to glomeruloid vascular lesions and intracerebral hemorrhage. Fbln1-deficient brains exhibit vascular malformations characterized by reduced type IV collagen deposition and diminished CD31 localization at endothelial junctions. Although overall pericyte coverage across the vasculature is maintained, the glomeruloid lesions lack pericyte investment. In the developing retina, loss of Fbln1 similarly results in hemorrhage and tortuous capillaries with glomeruloid vascular abnormalities that disrupt the formation of both the superficial and deep vascular plexuses. Together, these findings demonstrate that Fbln1 is a critical determinant of brain and retinal vascular development and is required to preserve endothelial barrier integrity during angiogenesis.

Following standardized vertebrate embryonic staging criteria, this study establishes a normal developmental staging table for the critically endangered Chinhai Spiny Newt, Echinotriton chinhaiensis, a narrowly distributed salamandrid species endemic to Ningbo, Zhejiang Province, eastern China. At a constant temperature of 20 °C, the entire embryonic period from oviposition to hatching lasted approximately 31.4 days, and the full developmental process was subdivided into 36 sequential stages. Stages 1-17 encompassed cleavage and neurulation, whereas Stages 18-36 corresponded to organogenesis-including tail bud outgrowth, external gill development, eye formation, and limb differentiation-until hatching. We systematically documented the diagnostic morphological traits of key embryonic structures and generated a concise chronological schematic illustrating the complete 36-stage developmental sequence. Comparative analyses across caudate species revealed evident heterochronic developmental shifts. The unique ontogenetic characteristics of E. chinhaiensis likely represent adaptive modifications to its ephemeral breeding microhabitats. This refined staging system improves our understanding of amphibian embryonic developmental patterns, clarifies how phylogenetic constraints and ecological selection shape evolutionary developmental trajectories, and provides fundamental baseline data for future comparative ontogenetic studies and conservation efforts for this endangered newt.

In binocular animals, retinal ganglion cell (RGC) axons selectively decussate at the optic chiasm. Selective decussation is directed by the radial glia and midline neurons in the optic chiasm. Radial glia attach to the underlying pial basement membrane (PBM) that is rich in β2 laminins. Here, we asked whether, and if so, how, β2 laminins in the PBM impact the selective decussation of RGC axons. β2 laminins are expressed in the PBM during the peak and late phases of RGC axon growth, and their deletion results in the fragmentation of the PBM and dysmorphic radial glia that lack properly elaborated endfeet. Genetic deletion of the β2 laminin subunit increased the proportion of non-decussating RGC axons in the ipsilateral tract. Consistent with the increase in the proportion of ipsilateral axons, we found persistent expression of the ipsilateral guidance cue EphrinB2 in radial glia during the late phase of axonal decussation and an increased number of ipsilateral RGCs in the ventrotemporal retina. These results suggest that in the optic chiasm β2 laminins are required for the integrity of the PBM and radial glia organization, and that this in turn may influence the production of EphrinB2. Our results also suggest that β2 laminins in the retina are necessary for neurogenesis and the laterality fate of ipsilateral RGCs, a function possibly mediated by their role in maintaining the attachment of retinal progenitor cells (RPCs) to the inner limiting membrane (ILM).

Immune cells in invertebrates and vertebrates are constantly exposed to environmental stressors, such as oxidative stress, UV radiation, toxins or infection, causing cellular damage of lipids, proteins but also DNA. Unresolved DNA damage in immune cells compromises genome integrity and stability, and can result in cellular dysfunction, apoptosis, or senescence and thereby contributing to immunodeficiencies, cancer or chronic inflammatory diseases. To counteract the effects and maintain genome integrity, cells employ a highly conserved DNA damage response (DDR) machinery. Beyond its canonical role of DNA repair, DDR has emerged as an active regulator of diverse physiological functions. This non-canonical (nc)DDR has been connected in the regulation of immune cells in vertebrates and invertebrates. Especially in Drosophila melanogaster it has been shown that ncDDR guides both lineage differentiation and functional adaptation during immune responses. These findings link ncDDR and genome plasticity to regulation of cell fate decisions and functional specialization of hemocytes and make Drosophila an attractive tool to dissect the role of ncDDR in immune cells during development, health but also disease settings. In this review, we discuss and summarize the available literature on the emerging concept of ncDDR as a critical layer of immune cell development and function in Drosophila and potentially similar mechanisms in mammals.

Regeneration abilities vary among species, but they also vary within and across individuals of a single species. Many annelids (segmented worms) have variable regenerative ability along their anterior-posterior (AP) body axis. However, satisfactory explanations for the diverse regenerative outcomes at different amputation positions remain elusive. The annelid Capitella teleta demonstrates robust posterior regenerative ability but lacks anterior regenerative ability, although anterior regenerative ability has not been rigorously assessed at multiple amputation sites. In this study, we characterize regenerative ability along the AP axis of C. teleta by comparing the post-amputation response of tail fragments cut between segments 1 and 2, 4 and 5, 10 and 11, and 15 and 16. Cell proliferation, gene expression, and protein expression assays demonstrate that the anterior-most amputation site progresses to later stages of regeneration than more posterior amputation sites, but regeneration does not reach completion. The distribution of neoblast-like cells after amputation suggests that these cells do not substantially contribute to formation of the anterior-facing blastema. Additionally, regeneration outcome is not improved following activation of Wnt/β-catenin signaling in tail fragments amputated between segments 1 and 2. Conversely, inhibition of Wnt/β-catenin signaling at the same amputation position leads to increased regenerative ability with distinct phenotypes consistent with anterior regeneration. Although not all differentiated anterior structures form, this result hints at partial anteriorization. Taken together, our data show the value of comparing regenerative abilities of tissues within an individual organism and demonstrate potential of increasing regenerative ability in regeneration-deficient tissues.

The respiratory system of Drosophila melanogaster consists of a network of interconnected gas-filled epithelial tubes. Stellate-shaped terminal cells at the termini of each tracheal branch serve as the primary source of oxygen and grow continuously throughout larval stages in response to hypoxic signals. Each cytoplasmic branch of a terminal cell hollows out to form subcellular seamless tubes that lack epithelial junctions. It is known that over the course of larval development, tracheal terminal cells grow by extending new branches and lengthening existing branches. By the end of larval development, each branch of these highly ramified cells contains a subcellular lumen; the relationship between branch and tube extension over this period of immense growth has yet to be explored. Here we demonstrate that branch extension and intracellular lumen formation are spatially and temporally coupled throughout larval stages, however, nascent tubes formed during a larval instar are fluid-filled and do not inflate with air until the following molt. This has interesting implications for the delivery of oxygen to target tissues during the first 48 h of larval growth.

The vertebrate neural retina, the image forming component of the eye, offers a useful, clinically-relevant system for understanding nervous system development and regeneration. Within vertebrates, the teleost fish retina specifically has been the subject of intense study for its regenerative response to neuronal damage - a response that does not take place in mammals. While the field has recently focused upon molecular and cellular processes that initiate this response and generate new neurons, it is also important to evaluate the ultimate "success" of this process. True neural regeneration requires that the cells and tissues that replace those lost to damage, also restore their original structure and function. From this outcomes-based perspective, does retinal regeneration recapitulate the results of development? This paper reviews ∼50 years of effort and knowledge regarding the regeneration of retinal neuronal types and patterns, wiring of regenerated neurons within the retina and beyond, and physiological and behavioral studies of function of regenerated retina. Emphasis is placed upon goldfish and zebrafish model systems, as these have been popular, productive models for revealing both the process and the outcome of retinal regeneration. The outcome, as accumulated data suggest, is considered remarkable in the sense that several features of structure and function are restored. Regenerated retinas are structurally similar in comparison with undamaged retinas that developed and grew normally, with notable local patterning errors. A body of evidence suggests that these structural errors are not of great consequence for the re-establishment of retinal function that is centrally integrated to support simple, visually-mediated behaviors. However, functional tests have been limited in scope and scale, lacking rigorous applications of more sophisticated approaches to evaluate visual system circuitry. Such approaches are becoming possible in the zebrafish model. Fish models are upheld as outstanding resources for providing current and future knowledge of regeneration that can be translated to treat human retinal disorders.

The oculocutaneous albinism 2 protein is required for melanin synthesis in vertebrates and its loss of function leads to one of the most common forms of skin and eye albinism, Oculocutaneous albinism (OCA). Using a zebrafish disease model for OCA2, we explore developmental explanations for pleiotropic traits observed in humans with OCA2. At multiple stages, we find that oca2 mutant zebrafish exhibit abnormal number and location of iridophores, an accessory pigment cell type that arises from neural crest cells in fish and amphibians. In addition, oca2 mutants display defects during eye development including delayed choroid fissure closure and significant changes in retinal cell differentiation and organization. Our transcriptomic analysis of oca2 mutants revealed alterations in gene networks involved in differentiation of neural crest cells, retinal neurons, and the retinal pigment epithelium (RPE) including Notch and Wnt/β-catenin pathway genes. Together, our data provide information about developmental consequences of oca2 mutation and support the possibility that the Oca2 protein may function in neural crest differentiation and eye development.

Epigenetic regulation via DNA demethylation and histone modification is essential for tissue development. We investigated the role of the Drosophila ten-eleven translocation protein (dTet) in eye development and its interaction with the oncohistone H3.3K27M. dTet overexpression in eye imaginal discs caused strong downregulation of retinal determination genes (toy, ey, so, eya, dac) and developmental arrest, leading to partial or complete eye loss. Enhanced dTet knockdown using sensitized conditions also resulted in eye developmental defects, supporting a dosage-dependent role for dTet during retinal morphogenesis. DNA-binding analysis using published ChIP-seq data revealed that dTet binds the promoters of these genes as well as wingless (wg), a negative regulator of eye specification, explaining its dual role in repressing retinal programs and activating Wnt/Wg signaling. Co-expression with H3.3K27M suppressed the mutation-induced overgrowth phenotype, whereas dTet loss enhanced it, indicating context-dependent modulation of tissue growth. Many dTet targets have conserved human orthologs implicated in congenital eye disorders, highlighting evolutionary conservation and potential translational relevance. These findings establish dTet as an important regulator of eye development.