Endothelial-Mesenchymal Transition of Brain Endothelial Cells: Possible Role during Metastatic Extravasation

Cancer progression towards metastasis follows a defined sequence of events described as the metastatic cascade. For extravasation and transendothelial migration metastatic cells interact first with endothelial cells. Yet the role of endothelial cells during the process of metastasis formation and extravasation is still unclear, and the interaction between metastatic and endothelial cells during transendothelial migration is poorly understood. Since tumor cells are well known to express TGF-β, and the compact endothelial layer undergoes a series of changes during metastatic extravasation (cell contact disruption, cytoskeletal reorganization, enhanced contractility), we hypothesized that an EndMT may be necessary for metastatic extravasation. We demonstrate that primary cultured rat brain endothelial cells (BEC) undergo EndMT upon TGF-β1 treatment, characterized by the loss of tight and adherens junction proteins, expression of fibronectin, β1-integrin, calponin and α-smooth muscle actin (SMA). B16/F10 cell line conditioned and activated medium (ACM) had similar effects: claudin-5 down-regulation, fibronectin and SMA expression. Inhibition of TGF-β signaling during B16/F10 ACM stimulation using SB-431542 maintained claudin-5 levels and mitigated fibronectin and SMA expression. B16/F10 ACM stimulation of BECs led to phosphorylation of Smad2 and Smad3. SB-431542 prevented SMA up-regulation upon stimulation of BECs with A2058, MCF-7 and MDA-MB231 ACM as well. Moreover, B16/F10 ACM caused a reduction in transendothelial electrical resistance, enhanced the number of melanoma cells adhering to and transmigrating through the endothelial layer, in a TGF-β-dependent manner. These effects were not confined to BECs: HUVECs showed TGF-β-dependent SMA expression when stimulated with breast cancer cell line ACM. Our results indicate that an EndMT may be necessary for metastatic transendothelial migration, and this transition may be one of the potential mechanisms occurring during the complex phenomenon known as metastatic extravasation.     

Derepressed Hyphal Growth and Reduced Virulence in a VH1 Family-related Protein Phosphatase Mutant of the Human Pathogen Candida albicans

Mitogen-activated protein (MAP) kinases are pivotal components of eukaryotic signaling cascades. Phosphorylation of tyrosine and threonine residues activates MAP kinases, but either dual-specificity or monospecificity phosphatases can inactivate them. The Candida albicans CPP1 gene, a structural member of the VH1 family of dual- specificity phosphatases, was previously cloned by its ability to block the pheromone response MAP kinase cascade in Saccharomyces cerevisiae. Cpp1p inactivated mammalian MAP kinases in vitro and acted as a tyrosine-specific enzyme. In C. albicans a MAP kinase cascade can trigger the transition from the budding yeast form to a more invasive filamentous form. Disruption of the CPP1 gene in C. albicans derepressed the yeast to hyphal transition at ambient temperatures, on solid surfaces. A hyphal growth rate defect under physiological conditions in vitro was also observed and could explain a reduction in virulence associated with reduced fungal burden in the kidneys seen in a systemic mouse model. A hyper-hyphal pathway may thus have some detrimental effects on C. albicans cells. Disruption of the MAP kinase homologue CEK1 suppressed the morphological effects of the CPP1 disruption in C. albicans. The results presented here demonstrate the biological importance of a tyrosine phosphatase in cell-fate decisions and virulence in C. albicans.     

Versatility of microglial bioenergetic machinery under starving conditions

Microglia are highly dynamic cells in the brain. Their functional diversity and phenotypic versatility brought microglial energy metabolism into the focus of research. Although it is known that microenvironmental cues shape microglial phenotype, their bioenergetic response to local nutrient availability remains unclear.In the present study effects of energy substrates on the oxidative and glycolytic metabolism of primary – and BV-2 microglial cells were investigated. Cellular oxygen consumption, glycolytic activity, the levels of intracellular ATP/ADP, autophagy, mTOR phosphorylation, apoptosis and cell viability were measured in the absence of nutrients or in the presence of physiological energy substrates: glutamine, glucose, lactate, pyruvate or ketone bodies.All of the oxidative energy metabolites increased the rate of basal and maximal respiration. However, the addition of glucose decreased microglial oxidative metabolism and glycolytic activity was enhanced. Increased ATP/ADP ratio and cell viability, activation of the mTOR and reduction of autophagic activity were observed in glutamine-supplemented media. Moreover, moderate and transient oxidation of ketone bodies was highly enhanced by glutamine, suggesting that anaplerosis of the TCA-cycle could stimulate ketone body oxidation.It is concluded that microglia show high metabolic plasticity and utilize a wide range of substrates. Among them glutamine is the most efficient metabolite. To our knowledge these data provide the first account of microglial direct metabolic response to nutrients under short-term starvation and demonstrate that microglia exhibit versatile metabolic machinery. Our finding that microglia have a distinct bioenergetic profile provides a critical foundation for specifying microglial contributions to brain energy metabolism.     

Contingency checking and self-directed behaviors in giant manta rays: Do elasmobranchs have self-awareness?

Elaborate cognitive skills arose independently in different taxonomic groups. Self-recognition is conventionally identified by the understanding that one’s own mirror reflection does not represent another individual but oneself, which has never been proven in any elasmobranch species to date. Manta rays have a high encephalization quotient, similar to those species that have passed the mirror self-recognition test, and possess the largest brain of all fish species. In this study, mirror exposure experiments were conducted on two captive giant manta rays to document their response to their mirror image. The manta rays did not show signs of social interaction with their mirror image. However, frequent unusual and repetitive movements in front of the mirror suggested contingency checking; in addition, unusual self-directed behaviors could be identified when the manta rays were exposed to the mirror. The present study shows evidence for behavioral responses to a mirror that are prerequisite of self-awareness and which has been used to confirm self-recognition in apes.     

Glial architecture of the ghost shark (Callorhinchus milii, Holocephali, Chondrichthyes) as revealed by different immunohistochemical markers

This article presents the first study on the glial architecture of a representative species of Holocephali, Callorhinchus milii (ghost shark). Holocephali are a small subclass of Chondrichthyes, with only a few extant genera, and those are considered to have a brain organization more similar to squalomorph sharks than to galeomorph sharks, skates, and rays. Three different astroglial markers--glial fibrillary acidic protein, S-100 protein, and glutamine synthetase (GS)--were investigated by immunohistochemical methods, applying both diaminobenzidine (DAB) and fluorescent techniques. They revealed similar glial structures, although most of them were detected by immunohistochemical reaction against GS and visualized by DAB. The predominant elements were radial ependymoglia spanning the area between the ventricular and meningeal surfaces, as in squalomorph sharks. Other similar features were the light appearance of myelinated neural tracts devoid of immunoreactivity, and the glial architecture of the reticular formation of the brain stem, cerebellum, and tectum, the latter with recognizable layers. The immunoreactivity of the vascular walls was similar; however, it is believed that different cell types form the blood-brain barrier in chimeras and in elasmobranchs. Some glial structures, however, resembled those of skates, rays, and galeomorph sharks. In C. milii astrocyte-like elements were observed in the telencephalon, using GS and S-100, although typical astrocyte-rich regions were not found. In some areas, especially the telencephalon, not only endfeet but also cell bodies were observed to be attached to the meningeal surface, with processes extending into the brain substance.     

Quantitative trends of zebra mussels in Lake Balaton (Hungary) in 2003–2005 at different water levels

During the extremely dry period between 2000 and 2003, the water level of Lake Balaton decreased by 82 cm and 80% of the stony littoral, an important habitat for the zebra mussel (Dreissena polymorpha), became dry. A recovery period started in 2004 due to intense precipitation, which increased water levels in the lake. Seasonal and spatial variations of the relative abundance, population density, population structure and biomass of the zebra mussel and the relative abundance of the amphipod Chelicorophium curvispinum were monitored in the period of 2003–2005 at four different shoreline sections and in two different portions (on the bottom and near the surface portion of the rip-rap) of Lake Balaton. Along with these studies, a quantitative survey of mussel larvae found in the plankton and of the abundance of mussel feeding diving ducks were made. As a consequence of the water level fall, on the dried part of the stony littoral, numerous zebra mussel druses perished. Following the dry period in early 2004, the relative abundance of the mussel on the bottom stones was smaller than in 2003 and the bottom community was dominated by C. curvispinum. By the end of 2004 and during 2005, the water level returned to normal and the surfaces of the reinundated stones were conducive to the successful colonization of zebra mussels. Hence, they returned as the dominant fauna in 2005. The stones near the surface might provide a new substrate for the recruitment of zebra mussels, probably offering more suitable substrata for the settlement in 2005 than in 2003. Therefore, the new substrata available in 2005 may have encouraged better and more rapid zebra mussel colonization than before. Zebra mussels may be better competitors for new space than C. curvispinum. A minor change of water-level fluctuation in 2005 and the reduction in population size of the mussel feeding waterfowl could have contributed to the intensive spread of zebra mussel by 2005.     

Soil carbon dioxide efflux determined from large undisturbed soil cores collected in different soil management systems

The aim of this study was to evaluate a measuring technique for determining soil CO₂ efflux from large soil samples having undisturbed structure under controlled laboratory conditions. Further objectives were to use the developed measuring method for comparing soil CO₂ efflux from samples, collected in three different soil management systems at various soil water content values. The experimental technique was tested and optimised for timing of sampling by taking air samples after 1, 3 and 6 hours of incubation. Based on the results, the incubation time was set to three hours. The CO₂ efflux measured for different soil management systems was the highest in the no-till and the lowest in the ploughing treatment, which was in accordance with measurements on accessible organic carbon for microbes. An increase in CO₂ efflux with increasing soil water content was found in the studied soil water content range. Our results indicate that soil respiration rates, measured directly after tillage operations, can highly differ from those measured long after.     

Climate sensitivity of soil water regime of different Hungarian Chernozem soil subtypes

In this study the possible effects of two predicted climate change scenarios on soil water regime of Hungarian Calcic Chernozem soils has been investigated. Soil profiles classified as Calcic Chernozem — in total 49 — were selected from the MARTHA soil physical database that incorporates soil data at national scale. These profiles were subdivided into three groups (sandy loam, loam and clayey loam) in accordance with their mechanical composition. Soil water retention curves were scaled separately for each of the three textural groups, using similar media scaling in order to represent the variability of soil hydrophysical data with one parameter, the scaling factor (SF). Reference soil profiles were chosen according to the cumulative distribution function of the scaling factor, six for each textural group. Daily downscaled meteorological data from A2 and B2 climate scenarios of the Hadley Centre (2070–2100) and data from a reference period (RF, 1961–1990) were used in this study to characterize different climatic situations. Nine representative years were selected in case of all the three scenarios, using the cumulative probability function of the annual precipitation sum. Scenario analyses were performed, validating the SWAP soil water balance simulation model for the 18 reference soil profiles and 27 representative years in order to evaluate the expected changes in soil water regime under different from the present (RF) climatic conditions (A2 and B2 scenarios). Our results show that the scaling factor could be used as a climate sensitivity indicator of soil water regime. The large climate sensitivity of the majority of Chernozem soil subtypes water regime has been proven.