The lack of EphB3 receptor prevents bone loss in mouse models of osteoporosis.

Bone homeostasis is a complex process in which some Eph kinase receptors and their ephrin ligands appear to be involved. In the present study we address this issue by examining, both in vitro and in vivo, the role of EphB2 and EphB3 in MSC differentiation into bone tissue. This was firstly evaluated by RT-qPCR and histological staining in MSCs cultured in specific mediums revealing that, whereas EphB2-/- MSCs mainly expressed pro-adipogenic transcription factors, EphB3-/- MSCs showed abundant osteogenic transcripts, such as Runx2, Msx2 and Sp7. To clarify the underlying molecular mechanisms, we found that the lack of EphB3 signaling alters the genetic profile of differentiating MSCs, reducing the expression of many inhibitory molecules and antagonists of the BMP signaling pathway, and increasing Bmp7 expression, a robust bone inductor. Then, to confirm the osteogenic role of EphB3 in vivo, we studied the condition of two mouse models of induced bone loss (ovariectomy or long-term glucocorticoid treatment). Interestingly, in both models, both WT and EphB2-/- mice equally developed the disease but EphB3-/- mice did not exhibit the typical bone loss, nor an increase in urine Ca2+ or blood serum CTX-1. This phenotype in EphB3-KO mice could be due to their significantly higher proportions of osteoprogenitor cells and preosteoblasts, and their lower number of osteoclasts, as compared with WT and EphB2-KO mice. Thus, we conclude that EphB3 acts as a negative regulator of the osteogenic differentiation, and its absence prevents bone loss in mice subjected to ovariectomy or dexamethasone treatment.

Osteoporosis affects more than 200 million people, mostly women. Our work shows that the EphB3 receptor restricts bone formation, and its absence prevents bone loss in osteoporotic mice. The bone protection observed in EphB3-deficient mice is due to the presence of more bone-forming cells and fewer bone-degrading cells. Molecularly, we found that when there's no EphB3 in mesenchymal stem cells, some bone-promoting genes are increased while many inhibitors are reduced. Therefore, this receptor could become a key target for new therapies that would help to improve the quality of life for those suffering from bone diseases. We're really excited to share our findings with a broad audience, including patients, healthcare professionals, researchers, and the life sciences industry.

Complications in Olecranon Fracture Surgery: A Comparison of Tension Band Vs. Plate Osteosynthesis.

Background: The purpose of this study is to compare the incidence of complications associated with tension band wiring (TBW) versus plate osteosynthesis (POS) in the treatment of olecranon fractures. Methods: We performed a retrospective cohort study of operatively treated adult olecranon fractures from an integrated healthcare system by multiple surgeons from January 2008 to December 2011. Patients were divided into two cohorts: fractures fixed using the tension band technique and fractures fixed using plate osteosynthesis. The study was limited to the Orthopedic Trauma Association classification of olecranon fracture type 21-B1, with subtypes 1-3. Outcome measures were loss of fracture fixation requiring revision, postoperative infection, stiffness requiring surgery, and symptomatic hardware removal (HWR). Univariate and multivariable logistic regressions were performed to test the associations between the type of internal fixation and outcomes. Results: A total of 321 olecranon fractures were included (median age: 61 years old, 57 % female); 153 participants were treated with TBW, and 168 patients with POS. There was one failure in the TBW group and two in the POS group (P=0.62). There were no significant differences in the infection rates (TBW 5%, POS 9%, P=0.20) and no reoperations for stiffness. The HWR occurred significantly more often in TBW (29%) than in POS (14%) (OR=0.39, P=0.001). The association between POS and decreased HWR remained highly significant (OR=0.40, P=0.003) after adjusting for clinical variables. Conclusion: In this large study comparing POS and TBW for 21-B1 olecranon fractures, no difference in fixation failure, infection, or postoperative stiffness was noted. A significantly greater risk of symptomatic hardware occurred in TBW. These findings may assist surgeons and patients in considering the risks and benefits of TBW and POS as treatment options for displaced olecranon fractures.

Oral Trypanosoma cruzi Acute Infection in Mice Targets Primary Lymphoid Organs and Triggers Extramedullary Hematopoiesis.

During the acute phase of Chagas disease, Trypanosoma cruzi circulation through the bloodstream leads to high tissue parasitism in the host. In primary lymphoid organs, progenitor cell reduction paralleled transient immunosuppression. Herein we showed that acute oral infection in mice promotes diffuse parasitism in bone marrow cells at 14 and 21 days post-infection (dpi), with perivascular regions, intravascular regions, and regions near the bone being target sites of parasite replication. Phenotypic analysis of hematopoietic differentiation in the bone marrow of infected mice showed that the cell number in the tissue is decreased (lineage-negative and lineage-positive cells). Interestingly, analysis of hematopoietic branching points showed that hematopoietic stem and progenitor cells (HSPCs) were significantly increased at 14 dpi. In addition, the pool of progenitors with stem plasticity (HSC-MPP3), as well as multipotent progenitors (MPPs) such as MPP4, also showed this pattern of increase. In contrast, subsequent progenitors that arise from MPPs, such as common lymphoid progenitors (CLPs), lymphoid-primed MPPs (LMPPs), and myeloid progenitors, were not enhanced; conversely, all presented numeric decline. Annexin V staining revealed that cell death increase in the initial hematopoietic branching point probably is not linked to CLPs and that myeloid progenitors decreased at 14 and 21 dpi. In parallel, our investigation provided clues that myeloid progenitor decrease could be associated with an atypical expression of Sca-1 in this population leading to a remarkable increase on LSK-like cells at 14 dpi within the HSPC compartment. Finally, these results led us to investigate HSPC presence in the spleen as a phenomenon triggered during emergency hematopoiesis due to mobilization or expansion of these cells in extramedullary sites. Splenocyte analysis showed a progressive increase in HSPCs between 14 and 21 dpi. Altogether, our study shows that the bone marrow is a target tissue in T. cruzi orally infected mice, leading to a hematopoietic disturbance with LSK-like cell bias accounting on HSPCs possibly affecting myeloid progenitor numbers. The LMPP and CLP reduction converges with defective thymocyte development. Lastly, it is tempting to speculate that the extramedullary hematopoiesis seen in the spleen is a mechanism involved in the hematological maintenance reported during the acute phase of oral T. cruzi infection.

Polycyclic aromatic hydrocarbons in fine road dust from a coal-utilization city: Spatial distribution, source diagnosis and risk assessment.

Coal combustion can release large amounts of polycyclic aromatic hydrocarbons (PAHs), which persist in various environment matrices (e.g., road dust) and hence cause the carcinogenic risk to human health. In this study, an exhaustive characterization of road dust samples coupling their physicochemical characteristics and stable isotope compositions (δ13C and δ15N) was conducted to evaluate the source, level, spatial distribution, and carcinogenic risk of PAHs in a typical coal-utilization city. Concentrations of Σ16 PAHs ranged from 605.5 to 25,374.3 ng/g with a mean concentration of 4083.0 ng/g. Pollution levels of sites around the coal-fired power plant (Zone 1) were significantly higher than those in other zones (p < 0.05). PAH concentration showed significant correlations with both C and N fractions (p < 0.01). Compositions of δ13C and δ15N in road dust coupled to principal component analysis and the partitions and diagnostic ratios of PAHs contributed to associating road dust to several local sources of contamination. Coal combustion and vehicular exhaust were major sources of PAHs around the power plant and urban area. Results of incremental lifetime cancer risk showed 81.5% of all sampling sites suffered potential risk (>10-6) for both adults and children, while children around the power plant suffered the highest risk. Despite the estimation of only potential risk being posed by PAHs in road dust, human exposure to the various environmental matrices, scientific and systematic assessment of carcinogenic risks by PAHs in the total environment warrants further investigations.

Natural estrogens enhance the engraftment of human hematopoietic stem and progenitor cells in immunodeficient mice.

Hematopoietic Stem and Progenitor Cells are crucial in the maintenance of lifelong production of all blood cells. These Stem Cells are highly regulated to maintain homeostasis through a delicate balance between quiescence, self-renewal and differentiation. However, this balance is altered during the hematopoietic recovery after Hematopoietic Stem and Progenitor Cell Transplantation. Transplantation efficacy can be limited by inadequate Hematopoietic Stem Cells number, poor homing, low level of engraftment, or limited self-renewal. As recent evidences indicate that estrogens are involved in regulating the hematopoiesis, we sought to examine whether natural estrogens (estrone or E1, estradiol or E2, estriol or E3 and estetrol or E4) modulate human Hematopoietic Stem and Progenitor Cells. Our results show that human Hematopoietic Stem and Progenitor Cell subsets express estrogen receptors, and whose signaling is activated by E2 and E4 on these cells. Additionally, these natural estrogens cause different effects on human Progenitors in vitro. We found that both E2 and E4 expand human Hematopoietic Stem and Progenitor Cells. However, E4 was the best tolerated estrogen and promoted cell cycle of human Hematopoietic Progenitors. Furthermore, we identified that E2 and, more significantly, E4 doubled human hematopoietic engraftment in immunodeficient mice without altering other Hematopoietic Stem and Progenitor Cells properties. Finally, the impact of E4 on promoting human hematopoietic engraftment in immunodeficient mice might be mediated through the regulation of mesenchymal stromal cells in the bone marrow niche. Together, our data demonstrate that E4 is well tolerated and enhances human reconstitution in immunodeficient mice, directly by modulating human Hematopoietic Progenitor properties and indirectly by interacting with the bone marrow niche. This application might have particular relevance to ameliorate the hematopoietic recovery after myeloablative conditioning, especially when limiting numbers of Hematopoietic Stem and Progenitor Cells are available.

Coordinated effects of air pollution control devices on PAH emissions in coal-fired power plants and industrial boilers.

Coal-fired power plants are important sources of polycyclic aromatic hydrocarbon (PAH) emissions in the world. The effects of various air pollution control devices (APCDs) on PAH emissions were investigated by analyzing samples from inlets and outlets of APCDs in six coal-fired power plants (A-F) and two coal-fired industrial boilers (G and H). The APCDs were electrostatic precipitators (ESPs), wet flue gas desulfurization systems (WFGDs), and wet ESPs (WESPs). The PAH congener patterns for the coal-fired plants were similar. Gas-phase PAHs were dominant in flue gases, and the most abundant PAH was naphthalene. Three- and four-ring PAHs were dominant in fly ash. Positive correlations were found between the PAH and total organic carbon contents of fly ash (R2 0.87) and slag (R2 0.92). Plants D-F, equipped with low-low-temperature ESPs (LLT-ESPs) and WESPs discharged the lowest PAHs. Circulating water was an important source of PAHs in the desulfurization except in plant A, which used desalinated seawater rather than circulating water in the desulfurization process. WESPs decreased PAH concentrations by an average of 20.67%, which can be spread to other plants to reduce PAHs.

Contrasting effects of microplastics on sorption of diazepam and phenanthrene in soil.

Microplastics have attracted extensive attention regarding their role in the cycling of organic pollutants in aquatic environments. However, the influence of microplastics on the sorption of organic pollutants in soil is unclear. Herein, we investigated the sorption of polar diazepam and nonpolar phenanthrene to two soils (Inceptisol and Oxisol). Batch sorption experiments were used to evaluate the effect of polyethylene (PE), polypropylene (PP), and polystyrene (PS) microplastics at addition rates of 0.1%, 1%, and 10% (w/w). The addition of microplastics significantly decreased the overall sorption of diazepam at 10%, and increased the sorption of phenanthrene at 1%, while the effects were negligible at other addition rates. Decreased sorption of diazepam was attributed to its lower sorption affinity to microplastics than to soil. Microplastics, even at 0.1%, substantially decreased the relative distribution of phenanthrene in soil, particularly for PE in the Oxisol with lower foc. The sorption affinity of phenanthrene followed the order PE > soil organic carbon (SOC) > PP > PS, suggesting that PE can be a significant sink of phenanthrene in contaminated soils. Overall, microplastics can change the sorption of diazepam and phenanthrene to soil and therefore affect their mobility and environmental risk in soil ecosystems.

Eph/ephrin Signaling and Biology of Mesenchymal Stromal/Stem Cells.

Mesenchymal stromal/stem cells (MSCs) have emerged as important therapeutic agents, owing to their easy isolation and culture, and their remarkable immunomodulatory and anti-inflammatory properties. However, MSCs constitute a heterogeneous cell population which does not express specific cell markers and has important problems for in vivo homing, and factors regulating their survival, proliferation, and differentiation are largely unknown. Accordingly, in the present article, we review the current evidence on the relationships between Eph kinase receptors, their ephrin ligands, and MSCs. These molecules are involved in the adult homeostasis of numerous tissues, and we and other authors have demonstrated their expression in human and murine MSCs derived from both bone marrow and adipose tissue, as well as their involvement in the MSC biology. We extend these studies providing new results on the effects of Eph/ephrins in the differentiation and immunomodulatory properties of MSCs.

Eph/Ephrin-mediated stimulation of human bone marrow mesenchymal stromal cells correlates with changes in cell adherence and increased cell death.

BACKGROUND: Mesenchymal stromal cells (MSC) are components of connective tissues and, in vitro, cell entities characterized by cell adhesion and immunophenotyping, although specific markers for their identification are lacking. Currently, MSC derived from either human bone marrow (BM-MSC) or adipose tissue (Ad-MSC) are considered the main sources of MSC for cell therapy. Eph receptors and their ligands, Ephrins, are molecules involved in cell adhesion and migration in several tissues and organs. In the current study, we analyze the pattern of Eph/Ephrin expression in MSC and evaluate the effects of blockade and stimulation of these receptor/ligand pairs on their biology. METHODS: Eph/Ephrin expression was analyzed in both BM-MSC and Ad-MSC by qRT-PCR. Then, we supplied BM-MSC cultures with either blocking or activating compounds to evaluate their effects on MSC proliferation, survival, and cell cycle by FACS. Changes in cytoskeleton and integrin α5ß1 expression were studied in stimulated BM-MSC by immunofluorescence microscopy and FACS, respectively. RESULTS: Higher numbers of Eph/Ephrin transcripts occurred in BM-MSC than in Ad-MSC. In addition, the blocking of Eph/Ephrin signaling correlated with decreased numbers of BM-MSC due to increased proportions of apoptotic cells in the cultures but without variations in the cycling cells. Unexpectedly, activation of Eph/Ephrin signaling by clustered Eph/Ephrin fusion proteins also resulted in increased proportions of apoptotic MSC. In this case, MSC underwent important morphological changes, associated with altered cytoskeleton and integrin α5ß1 expression, which did not occur under the blocking conditions. CONCLUSIONS: Taken together, these results suggest that Eph/Ephrin activation affects cell survival through alterations in cell attachment to culture plates, affecting the biology of BM-MSC.

Can a Proper T-Cell Development Occur in an Altered Thymic Epithelium? Lessons From EphB-Deficient Thymi.

For a long time, the effects of distinct Eph tyrosine kinase receptors and their ligands, ephrins on the structure, immunophenotype, and development of thymus and their main cell components, thymocytes (T) and thymic epithelial cells (TECs), have been studied. In recent years, the thymic phenotype of mutant mice deficient in several Ephs and ephrins B has been determined. Remarkably, thymic stroma in these animals exhibits important defects that appear early in ontogeny but little alterations in the proportions of distinct lymphoid cell populations. In the present manuscript, we summarize and extend these results discussing possible mechanisms governing phenotypical and functional thymocyte maturation in an absence of the critical T-TEC interactions, concluding that some signaling mediated by key molecules, such as MHCII, CD80, ß5t, Aire, etc. could be sufficient to enable a proper maturation of thymocytes, independently of morphological alterations affecting thymic epithelium.

Increased epithelial-free areas in thymuses with altered EphB-mediated thymocyte-thymic epithelial cell interactions.

Epithelial-free areas, present in both thymic cortex and medulla, have been studied in WT and EphB-deficient mice that have important alterations in the development of thymic epithelium due to the lack of proper thymocyte-thymic epithelial cell interactions. In both WT and mutant thymuses, the number and size of epithelial-free areas are significantly larger in the medulla than in the cortex. The two parameters show a reverse correlation: low numbers of these areas course with large epithelial-free areas and vice versa. However, their structure and cell content are similar in mutant and WT thymuses. Cortical epithelial-free areas just contain DP thymocytes, while the medullary ones consist of SP cells, blood vessels, mesenchyme-derived ER-TR7+ cells and components of the extracellular matrix (i.e., collagen IV, fibronectin, laminin). Other components, such as desmin, αSMA, PDGFRß and Ng2, frequently associated with blood vessel walls, also appear. Vimentin, although present in medullary epithelial-free areas, does not co-express with epithelial cells. Other markers related to epithelial-mesenchymal transitions, such as Snail, Slug or FSP1, are not expressed. These results suggest that alterations in the cell interactions between distinct thymic cell components that induce both increased proportions of apoptotic thymic epithelial cells and altered behavior of the mesenchyme associated with the medullary vasculature could explain the appearance of these areas and their differences in the cortex and medulla.

Community-Reviewed Biological Network Models for Toxicology and Drug Discovery Applications.

Biological network models offer a framework for understanding disease by describing the relationships between the mechanisms involved in the regulation of biological processes. Crowdsourcing can efficiently gather feedback from a wide audience with varying expertise. In the Network Verification Challenge, scientists verified and enhanced a set of 46 biological networks relevant to lung and chronic obstructive pulmonary disease. The networks were built using Biological Expression Language and contain detailed information for each node and edge, including supporting evidence from the literature. Network scoring of public transcriptomics data inferred perturbation of a subset of mechanisms and networks that matched the measured outcomes. These results, based on a computable network approach, can be used to identify novel mechanisms activated in disease, quantitatively compare different treatments and time points, and allow for assessment of data with low signal. These networks are periodically verified by the crowd to maintain an up-to-date suite of networks for toxicology and drug discovery applications.

Eph/ephrin-B-mediated cell-to-cell interactions govern MTS20(+) thymic epithelial cell development.

Thymus development is a complex process in which cell-to-cell interactions between thymocytes and thymic epithelial cells (TECs) are essential to allow a proper maturation of both thymic cell components. Although signals that control thymocyte development are well known, mechanisms governing TEC maturation are poorly understood, especially those that regulate the maturation of immature TEC populations during early fetal thymus development. In this study, we show that EphB2-deficient, EphB2LacZ and EphB3-deficient fetal thymuses present a lower number of cells and delayed maturation of DN cell subsets compared to WT values. Moreover, deficits in the production of chemokines, known to be involved in the lymphoid seeding into the thymus, contribute in decreased proportions of intrathymic T cell progenitors (PIRA/B(+)) in the mutant thymuses from early stages of development. These features correlate with increased proportions of MTS20(+) cells but fewer MTS20(-) cells from E13.5 onward in the deficient thymuses, suggesting a delayed development of the first epithelial cells. In addition, in vitro the lack of thymocytes or the blockade of Eph/ephrin-B-mediated cell-to-cell interactions between either thymocytes-TECs or TECs-TECs in E13.5 fetal thymic lobes coursed with increased proportions of MTS20(+) TECs. This confirms, for the first time, that the presence of CD45(+) cells, corresponding at these stages to DN1 and DN2 cells, and Eph/ephrin-B-mediated heterotypic or homotypic cell interactions between thymocytes and TECs, or between TECs and themselves, contribute to the early maturation of MTS20(+) TECs.

MR spectroscopy of intracranial tuberculomas: A singlet peak at 3.8 ppm as potential marker to differentiate them from malignant tumors.

PURPOSE: The diagnosis of intracranial tuberculomas is often challenging. Our purpose is to describe the most common metabolic patterns of tuberculomas by MR spectroscopy (MRS) with emphasis on potential specific markers. METHODS: Single-voxel MRS short echo time was performed in 13 cases of tuberculomas proven by histology and/or response to anti-mycobacterial therapy. For comparison MRS was also performed in 19 biopsy-proven malignant tumors (13 high-grade gliomas and six metastasis). Presence of metabolic peaks was assessed visually and categorical variables between groups were compared using chi-square. Metabolite ratios were compared using Mann-Whitney test and diagnostic accuracy of the metabolite ratios was compared using receiver-operating characteristic (ROC) curves analysis. RESULTS: Spectroscopic peaks representing lipids and glutamate/glutamine (Glx) as well as a peak at ∼3.8 ppm were well defined in 77% (10/13), 77% (10/13) and 69% (nine of 13) of tuberculomas, respectively. Lipid and Glx peaks were also present in most of the malignant lesions, 79% (15/19) and 74% (14/19) respectively. However, a peak at ∼3.8 ppm was present in only 10% (two of 19) of the tumor cases (p < 0.001). Higher Cho/Cr and mI/Cr ratios helped discriminate malignant lesions with an area under the ROC curve of 0.86 (SE: 0.078, p < 0.002, CI: 0.7-1) and 0.8 (SE: 0.1, p < 0.009, CI: 0.6-1), respectively. Threshold values between 1.7-1.9 for Cho/Cr and 0.8-0.9 for mI/Cr provided high specificity (91% for both metabolites) and adequate sensitivity (75% and 80%, respectively) for discrimination of malignant lesions. CONCLUSION: A singlet peak at ∼3.8 ppm is present in the majority of tuberculomas and absent in most malignant tumors, potentially a marker to differentiate these lesions. The assignment of the peak is difficult from our analysis; however, guanidinoacetate (Gua) is a possibility. Higher Cho/Cr and mI/Cr ratios should favor malignant lesions over tuberculomas. The presence of lipids and Glx is non-specific.

Eph/Ephrins-Mediated Thymocyte-Thymic Epithelial Cell Interactions Control Numerous Processes of Thymus Biology.

Numerous studies emphasize the relevance of thymocyte-thymic epithelial cell (TECs) interactions for the functional maturation of intrathymic T lymphocytes. The tyrosine kinase receptors, Ephs (erythropoietin-producing hepatocyte kinases) and their ligands, ephrins (Eph receptor interaction proteins), are molecules known to be involved in the regulation of numerous biological systems in which cell-to-cell interactions are particularly relevant. In the last years, we and other authors have demonstrated the importance of these molecules in the thymic functions and the T-cell development. In the present report, we review data on the effects of Ephs and ephrins in the functional maturation of both thymic epithelial microenvironment and thymocyte maturation as well as on their role in the lymphoid progenitor recruitment into the thymus.

EphB2 and EphB3 play an important role in the lymphoid seeding of murine adult thymus.

Adult thymuses lacking either ephrin type B receptor 2 (EphB2) or EphB3, or expressing a truncated form of EphB2, the forward signal-deficient EphB2LacZ, have low numbers of early thymic progenitors (ETPs) and are colonized in vivo by reduced numbers of injected bone marrow (BM) lineage-negative (Lin(-)) cells. Hematopoietic progenitors from these EphB mutants showed decreased capacities to colonize wild type (WT) thymuses compared with WT precursors, with EphB2(-/-) cells exhibiting the greatest reduction. WT BM Lin(-) cells also showed decreased colonizing capacity into mutant thymuses. The reduction was also more severe in EphB2(-/-) host thymuses, with a less severe phenotype in the EphB2LacZ thymus. These results suggest a major function for forward signaling through EphB2 and, to a lesser extent, EphB3, in either colonizing progenitor cells or thymic stromal cells, for in vivo adult thymus recruitment. Furthermore, the altered expression of the molecules involved in thymic colonization that occurs in the mutant thymus correlates with the observed colonizing capacities of different mutant mice. Reduced production of CCL21 and CCL25 occurred in the thymus of the 3 EphB-deficient mice, but their expression, similar to that of P-selectin, on blood vessels, the method of entry of progenitor cells into the vascular thymus, only showed a significant reduction in EphB2(-/-) and EphB3(-/-) thymuses. Decreased migration into the EphB2(-/-) thymuses correlated also with reduced expression of both ephrinB1 and ephrinB2, without changes in the EphB2LacZ thymuses. In the EphB3(-/-) thymuses, only ephrinB1 expression appeared significantly diminished, confirming the relevance of forward signals mediated by the EphB2-ephrinB1 pair in cell recruitment into the adult thymus.

Assessment of the Internal Fit of Lithium Disilicate Crowns Using Micro-CT.

PURPOSE: The aim of this study was to compare the internal fit of lithium disilicate crowns fabricated using digital technology with those fabricated by conventional means. MATERIALS AND METHODS: Forty-five lithium disilicate crowns were fabricated: 15 using digital impression and computer-aided design/computer-aided machining technique (group 1), 15 from the same digital impressions, but using a conventional die and laboratory fabrication process (group 2), and 15 using a conventional poly (vinyl siloxane) (PVS) impression and laboratory fabrication process (group 3). Tooth #15 was prepared for all-ceramic restoration on an ivorine typodont, which was digitized and a replica milled in zirconia to serve as master model. The master zirconia model was used for the impression procedures. Duplicate dies of the master zirconia die were made in polyurethane, enabling the internal fit of each crown to be evaluated using X-ray microcomputed tomography. The total volume of the internal space between the crown and die, the mean and maximum thickness of this space, and the percentage of the space that was at or below 120 µm thickness was calculated for each group and statistically tested for significant difference using one-way ANOVA, with post hoc Scheffé analysis. RESULTS: Group 1 crowns resulted in a smaller volume of internal space (12.49 ± 1.50 mm(3)) compared to group 2 (15.40 ± 2.59 mm(3) ) and to those of group 3 (18.01 ± 2.44 mm(3)). The mean thickness of the internal space for group 1 (0.16 ± 0.01 mm) and for group 2 (0.17 ± 0.03 mm) was significantly lower than that of group 3 (0.21 ± 0.03 mm). The average percentage of the internal space of a thickness of 120 µm and below was different between the three groups: 46.73 ± 5.66% for group 1, 37.08 ± 17.69% for group 2, and 22.89 ± 9.72% for group 3. Three-dimensional renderings of the internal space were also created. CONCLUSIONS: The results of this study suggested that pressed and milled IPS e. max crowns from LAVA COS digital impressions had a better internal fit to the prepared tooth than pressed IPS e.max crowns from PVS impressions in terms of total volume of internal space, average thickness of internal space, and percentage of internal space at or below 120 µm.

Conditioned deletion of ephrinB1 and/or ephrinB2 in either thymocytes or thymic epithelial cells alters the organization of thymic medulla and favors the appearance of thymic epithelial cysts.

Our understanding about medullary compartment, its niches composition and formation is still limited. Previous studies using EphB2 and/or EphB3 knockout mice showed an abnormal thymic development that affects mainly to the epithelial component, including the cortex/medulla distribution, thymic epithelial cell (TEC) morphology and different epithelial-specific marker expression. We have already demonstrated that the lack of ephrinB1 and/or ephrinB2, either on thymocytes or on TECs, alters the cell intermingling processes necessary for thymus organization and affect cortical TEC subpopulations. In the present work, we have used the Cre-LoxP model to selectively delete ephrinB1 and/or ephrinB2 in thymocytes (EfnB1(thy/thy), EfnB2(thy/thy), EfnB1(thy/thy)EfnB2(thy/thy) mice) or TECs (EfnB1(tec/tec), EfnB2(tec/tec), EfnB1(tec/tec)EfnB2(tec/tec) mice) and have analyzed their role on the medullary compartment. In all the studied mutants, medullary areas are smaller and more compact than in the wt thymuses. In most of them, we observe abundant big cysts and a higher proportion of UEA(hi)MTS10(-) cells than in wt mice, which are often forming small cysts. On EfnB1(tec/tec)EfnB2(tec/tec), changes affecting organ size and medullary compartment start at perinatal stage. Our data shed some light on knowledge about wt medulla histological structure and cysts meaning and formation process and on the role played by ephrinB in them.

Ephrin-B-dependent thymic epithelial cell-thymocyte interactions are necessary for correct T cell differentiation and thymus histology organization: relevance for thymic cortex development.

Previous analysis on the thymus of erythropoietin-producing hepatocyte kinases (Eph) B knockout mice and chimeras revealed that Eph-Eph receptor-interacting proteins (ephrins) are expressed both on T cells and thymic epithelial cells (TECs) and play a role in defining the thymus microenvironments. In the current study, we have used the Cre-LoxP system to selectively delete ephrin-B1 and/or ephrin-B2 in either thymocytes (EfnB1(thy/thy), EfnB2(thy/thy), and EfnB1(thy/thy)EfnB2(thy/thy) mice) or TECs (EfnB1(tec/tec), EfnB2(tec/tec), and EfnB1(tec/tec)EfnB2(tec/tec) mice) and determine the relevance of these Eph ligands in T cell differentiation and thymus histology. Our results indicate that ephrin-B1 and ephrin-B2 expressed on thymocytes play an autonomous role in T cell development and, expressed on TECs, their nonautonomous roles are partially overlapping. The effects of the lack of ephrin-B1 and/or ephrin-B2 on either thymocytes or TECs are more severe and specific on thymic epithelium, contribute to the cell intermingling necessary for thymus organization, and affect cortical TEC subpopulation phenotype and location. Moreover, ephrin-B1 and ephrin-B2 seem to be involved in the temporal appearance of distinct cortical TECs subsets defined by different Ly51 levels of expression on the ontogeny.

Eph/ephrinB signalling is involved in the survival of thymic epithelial cells.

The signals that determine the survival/death of the thymic epithelial cells (TECs) component during embryonic development of the thymus are largely unknown. In this study, we combine different in vivo and in vitro experimental approaches to define the role played by the tyrosine kinase receptors EphB2 and EphB3 and their ligands, ephrinsB, in the survival of embryonic and newborn (NB) TECs. Our results conclude that EphB2 and EphB3 are involved in the control of TEC survival and that the absence of these molecules causes increased apoptotic TEC proportions that result in decreased numbers of thymic cells and a smaller-sized gland. Furthermore, in vitro studies using either EphB2-Fc or ephrinB1-Fc fusion proteins demonstrate that the blockade of Eph/ephrinB signalling increases TEC apoptosis, whereas its activation rescues TECs from cell death. In these assays, both heterotypic thymocyte-TEC and homotypic TEC-TEC interactions are important for Eph/ephrinB-mediated TEC survival.