Ti–48Al–2Cr–2Nb (at.%) (γ-TiAl), a gamma titanium aluminide alloy originally designed for aerospace applications, appears to have excellent potential as implant material. Thermal treatment of γ-TiAl renders this alloy extremely corrosion resistant in vitro, which could improve its biocompatibility. In this study, the surface oxides produced by thermal oxidation (at 500°C, and at 800°C for 1 h in air) on γ-TiAl were characterized by X-ray photoelectron spectroscopy (XPS). hFOB 1.19 cell adhesion on thermally oxidized γ-TiAl was examined in vitro by a hexosaminidase assay, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) after 1, 7 and 14 days. Ti–6Al–4V surfaces were used for comparison. Hexosaminidase assay data and CLSM analysis of focal contacts and cytoskeleton organization showed no differences in cell attachment on autoclaved and both heat-treated γ-TiAl surfaces at the different time points. SEM images showed well organized multi-layers of differentiated cells adhered on thermally oxidized γ-TiAl surfaces at day 14. Unexpectedly, thermally oxidized Ti–6Al–4V surfaces oxidized at 800°C exhibited cytotoxic effects on hFOB 1.19 cells. Our results indicate that thermal oxidation of γ-TiAl seems to be a promising method to generate highly corrosion resistant and biocompatible surfaces for implant applications.

The mesentery, a newly minted organ, plays various anatomical and physiological roles during animal development. In echinoderms, and particularly in members of the class Holothuroidea (sea cucumbers) the mesentery plays an additional unique role: it is crucial for the process of intestinal regeneration. In these organisms, a complete intestine can form from cells that originate in the mesentery. In this review, we focus on what is known about the changes that take place in the mesentery and what has been documented on the cellular and molecular mechanisms involved. We describe how the events that unfold in the mesentery result in the formation of a new intestine.

No abstract

The ability to culture different cell types is essential for answering many questions in developmental and regenerative biology. Studies in marine organisms, in particular echinoderms, have been limited by the lack of well-described cellular culture systems. Here we describe a cell culture system, for normal or regenerating holothurian cells, that allows cell characterization by immunohistochemistry and scanning electron microscopy. These cell cultures can now be used to perform multiple types of experiments in order to explore the cellular, biochemical, and genomic aspects of echinoderm regenerative properties.

Tissue engineering leads to the development of biomaterial scaffolds where its biocompatibility and bioactivity are often improved after performing physical or chemical surface modification treatments. Micropatterning, soft lithography, and biofabrication are also approaches that provide a biomimetic microenvironment but have proven very costly and time consuming. In this concern, an appropriate substrate with suitable sites for cell attachment represents a major factor in cell behavior and biological functions. For this reason, our strategy was to fabricate a standard fibrous biomaterial with reproducible surface topography, incorporating microbeads and nanofeatures, and show the positive outcomes of the new substrate reflected on cell functions of bone cells. The electrospun polycaprolactone (PCL) beads-on-string membranes were obtained by adjusting the spinning solution at different concentrations until continuous beads were formed. Cell adhesion and proliferation, on the PCL scaffold, were analyzed the subsequent 2 days after initial culture. Complementary studies of cytoskeleton spreading and differentiation were analyzed after 7 and 14 days of the initial incubation. The scanning electron microscopy (SEM) images showed evidence of the formation of beads-on-string nanofibers and suggested that as-formed microstructures worked as attachment sites for osteoblasts. We investigated cell proliferation using anti-BrdU fluorescence assay, and results show a similar proliferation rate of cells cultured between PCL scaffolds and control. Finally, Phalloidin TRITC and antisialoprotein antibody were used to analyze cell spreading and differentiation after 7 and 14 days, respectively. This work shows a low-cost fabrication method to produce a biodegradable scaffold with micro/nanostructured characteristics that favor cell adhesion, proliferation, maturation, and subsequent differentiation of osteoblasts. According to the results, the biocompatibility of PCL beads-on-string could be comparable to other complex biomaterials, and we conclude that our scaffold is optimal for applications in bone tissue regeneration.

Interfacial surface properties, both physical and chemical, are known to play a critical role in achieving longterm stability of cell−biomaterial interactions. Novel bone tissue engineering technologies, which provide a suitable interface between cells and biomaterials and mitigate aseptic osteolysis, are sought and can be developed via the incorporation of nanostructured materials. In this sense, engineered nanobased constructs provide an effective interface and suitable topography for direct interaction with cells, promoting faster osseointegration and anchoring. Therefore, herein we have investigated the surface functionalization, biocompatibility, and effect of cellulose-nanodiamond conjugates on osteoblast proliferation and differentiation. Cellulose nanocrystals (CNC) were aminated through a 3aminopropyltriethyoxysilane (APTES) silylation, while nanodiamonds (ND) were treated with a strong acid oxidation reflux, as to produce carboxyl groups on the surface. Thereafter, the two products were covalently joined through an amide linkage, using a common bioconjugation reaction. Human fetal osteoblastic cells (hFOB) were seeded for 7 days to investigate the in vitro performance of the cellulose-nanodiamond conjugates. By employing immunocytochemistry, the bone matrix expression of osteocalcin (OC) and bone sialoprotein (BSP) was analyzed, demonstrating the viability and capacity of osteoblasts to proliferate and differentiate on the developed composite. These results suggest that cellulose-nanodiamond composites, which we call oxidized biocompatible interfacial nanocomposites (oBINC), have the potential to serve as a biointerface material for cell adhesion, proliferationand differentiation because of their osteoconductive properties and biocompatibility; furthermore, they show promising applications for bone tissue regeneration.

The cellular events underlying intestine regrowth in the sea cucumber Holothuria glaberrima have been described by our group. Currently, the molecular and signaling mechanisms involved in this process are being explored. One of the limitations to our investigations has been the absence of suitable cell culture methodologies, required to advance the regeneration studies. An in vitro system, where regenerating intestine explants can be studied in organ culture, was established previously by our group. However, a detailed description of the histological properties of the cultured gut explants was lacking. Here, we used immunocytochemical techniques to study the potential effects of the culture conditions on the histological characteristics of explants, comparing them to the features observed during gut regeneration in our model in vivo. Additionally, the explant outgrowths were morphologically described by phase-contrast microscopy and SEM. Remarkably, intestine explants retain most of their original histoarchitecture for up to 10 days, with few changes as culture time increases. The most evident effects of the culture conditions on explants over culture time were the reduction in the proliferative rate, the loss of the polarity in the localization of proliferating cells, and the appearance of a subpopulation of putative spherulocytes. Finally, cells that migrated from the gut explants could form net-like monolayers, firmly attached to the culture substrate. Overall, regenerating explants in organ culture represent a powerful tool to perform short-term studies of processes associated with gut regeneration in H. glaberrima under controlled conditions.

Organ regeneration is a rare phenomenon in higher vertebrates. However, members of closely related groups such as echinoderms have striking regeneration capacities. In studies of intestinal regeneration using the sea cucumber Holothuria glaberrima, recent results have shown the involvement of the Wnt signaling pathway. One important component in this intracellular cascade is β‐catenin, that, depending on its phosphorylation state, activates transcription of Wnt target genes. In one case, phosphorylation of the tyrosine residue 489 (Y489) increases transcriptional activity of the β‐catenin/Tcf complex. To determine if Y489‐β‐catenin phosphorylation is involved in intestinal regeneration, normal and regenerating intestines of H. glaberrima specimens were dissected at different post‐evisceration stages. Tissues were processed for immunohistochemistry using anti‐PY489‐β‐catenin antibody and assessed by counting the temporal and spatial appearance of labeled cells in the distal, medial and proximal ends of the mesentery and gut rudiment. In normal non‐eviscerated animals, the antibody labeled about the 14% of cells in the coelomic lining and 31.2% of cells in the luminal epithelium. In regenerating specimens, PY489‐β‐catenin was expressed at all stages of regeneration from 3‐dpe up to 14‐dpe, with a peak increment of 40.6% of cells labeled in the mesothelium of the 7‐dpe intestinal rudiment. To determine the role of activating or inhibiting the WNT/β‐catenin pathway on the expression of the PY489 labeling, we treated tissue explants with the Wnt pathway activator LiCl and inhibitor iCRT14. When compared to controls there was no significant difference in the number of PY489‐β‐catenin immunoreactive cells observed in the LiCl‐treated explants, while there was a significant increase in immunoreactive cells in the iCRT14‐treated explants. Our results suggest that the Wnt/β‐catenin signaling is activated during intestinal regeneration in H. glaberrima and that phosphorylation in the Y489 residue of β‐catenin is important for the Wnt role in intestinal regeneration. In particular, the results suggest a possible role for Wnt pathway in the dedifferentiation status of the mesothelium once the initial intestinal blastema is formed. Our study might provide the basis for the role of the Wnt signaling pathway during organogenesis in holothurians and thus provide a better understanding of general regenerative processes.Support or Funding InformationFunded by NIH R15NS081686, NSF IOS‐1252679 and the University of Puerto Rico.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.