Coelomocytes is the generic name for a collection of cellular morphotypes, present in many coelomate animals, and highly variable among echinoderm classes. The roles attributed to the major types of these free circulating cells present in the coelomic fluid of echinoderms include immune response, phagocytic digestion and clotting. Our main aim in this study was to characterize coelomocytes found in the coelomic fluid of Marthasterias glacialis (class Asteroidea) by using a combination of flow cytometry (FC), imaging flow cytometry (IFC) and fluorescence plus transmission electron microscopy (TEM). Two coelomocyte populations (P1 and P2) identified through flow cytometry were subsequently studied in terms of abundance, morphology, ultrastructure, cell viability and cell cycle profiles. Ultrastructurally, P2 diploid cells were present as two main morphotypes, similar to phagocytes and vertebrate thrombocytes, whereas the smaller P1 cellular population was characterized by low mitotic activity, a relatively undifferentiated cytotype and a high nucleus/cytoplasm ratio. In the present study we could not rule out possible similarities between haploid P1 cells and stem-cell types in other animals. Additionally, we report the presence of two other morphotypes in P2 that could only be detected by fluorescence microscopy, as well as a morphotype revealed via combined microscopy/FC. This integrative experimental workflow combined cells physical separation with different microscopic image capture technologies, enabling us to better tackle the characterization of the heterogeneous composition of coelomocytes populations.

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The direct method was applied to characterize the fracture behavior of a structural adhesive obtaining the cohesive law for Mode I. Additionally, the effect of the substrate material in the results was analyzed. The Double Cantilever Beam (DCB) tests were applied to achieve the fracture toughness in Mode I and the J-integral approach was used to measure the critical energy release rate, JIc . The results were then compared and validated using the Compliance Based Beam Method (CBBM). By differentiation of the J-integral values with respect to the crack tip opening displacement (CTOD), the cohesive law was obtained. Moreover, a numerical validation was performed, by comparing the load-displacement experimental results with the numerical data obtained with the Abaqus software. Furthermore, Digital Image Correlation (DIC) analysis was applied resorting to a Python script aiming to optimize the process. The obtained results indicate that the material substrate does not influence the value of the fracture toughness but could positively influence the precision of the obtained cohesive law. Moreover, it was observed that the direct method could predict the fracture behavior of the epoxy adhesive.

The potential to regenerate a damaged body part is expressed to a different extent in animals. Echinoderms, in particular starfish, are known for their outstanding potential to regenerate cell, tissue, organ, and body parts. For instance, humans have restricted abilities to restore organ systems being dependent on limited sources of stem cells. In particular, the potential to regenerate the central nervous system is extremely limited, explaining the lack of natural mechanisms that could overcome the development of neurodegenerative diseases and the presence of traumatisms. Therefore, understanding the molecular and cellular mechanisms of regeneration in starfish could lead to the development of new therapeutic approaches in humans. In this study, we tackle the problem of starfish central nervous system regeneration by examining anatomical and behavioral traits, including external anatomic anomalies, the dynamics of coelomocytes populations and neuronal tissue architecture. We noticed that several anatomic anomalies were evident and detected that the injured arm could not be used anymore to lead the starfish movement. Those seem to be related to defense mechanisms and protection of the wound. In particular, histology showed that tissue patterns during regeneration resemble those described in holothurians and in starfish arm tip regeneration. Flow cytometry coupled with imaging flow cytometry unveiled a new coelomocyte population during the late phase of the regeneration process. Morphotypes of previously characterized coelomocytes populations were described based on IFC data. Further studies of this new coelomocyte population might provide insights on their involvement in radial nerve cord regeneration.

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