Purinergic system in cancer stem cells.

Accumulating evidence supports the idea that cancer stem cells (CSCs) are those with the capacity to initiate tumors, generate phenotypical diversity, sustain growth, confer drug resistance, and orchestrate the spread of tumor cells. It is still controversial whether CSCs originate from normal stem cells residing in the tissue or cancer cells from the tumor bulk that have dedifferentiated to acquire stem-like characteristics. Although CSCs have been pointed out as key drivers in cancer, knowledge regarding their physiology is still blurry; thus, research focusing on CSCs is essential to designing novel and more effective therapeutics. The purinergic system has emerged as an important autocrine-paracrine messenger system with a prominent role at multiple levels of the tumor microenvironment, where it regulates cellular aspects of the tumors themselves and the stromal and immune systems. Recent findings have shown that purinergic signaling also participates in regulating the CSC phenotype. Here, we discuss updated information regarding CSCs in the purinergic system and present evidence supporting the idea that elements of the purinergic system expressed by this subpopulation of the tumor represent attractive pharmacological targets for proposing innovative anti-cancer therapies.

Transactivation of receptor tyrosine kinases by purinergic P2Y and adenosine receptors.

Transactivation of receptor tyrosine kinases (RTK) is a crosstalk mechanism exhibited by G-protein-coupled receptors (GPCR) to activate signaling pathways classically associated with growth factors. The discovery of RTK transactivation was a breakthrough in signal transduction that contributed to developing current concepts in intracellular signaling. RTK transactivation links GPCR signaling to important cellular processes, such as cell proliferation and differentiation, and explains the functional diversity of these receptors. Purinergic (P2Y and adenosine) receptors belong to class A of GPCR; in the present work, we systematically review the experimental evidence showing that purinergic receptors have the ability to transactivate RTK in multiple tissues and physiopathological conditions resulting in the modulation of cellular physiology. Of particular relevance, the crosstalk between purinergic receptors and epidermal growth factor receptor is a redundant pathway that participates in multiple pathophysiological processes. Specific and detailed knowledge of purinergic receptor-regulated pathways advances our understanding of the complexity of GPCR signal transduction and opens the way for pharmacologic intervention in the pathological context.

Autocrine and paracrine purinergic signaling in the most lethal types of cancer.

Cancer comprises a collection of diseases that occur in almost any tissue and it is characterized by an abnormal and uncontrolled cell growth that results in tumor formation and propagation to other tissues, causing tissue and organ malfunction and death. Despite the undeniable improvement in cancer diagnostics and therapy, there is an urgent need for new therapeutic and preventive strategies with improved efficacy and fewer side effects. In this context, purinergic signaling emerges as an interesting candidate as a cancer biomarker or therapeutic target. There is abundant evidence that tumor cells have significant changes in the expression of purinergic receptors, which comprise the G-protein coupled P2Y and AdoR families of receptors and the ligand-gated ion channel P2X receptors. Tumor cells also exhibit changes in the expression of nucleotidases and other enzymes involved in nucleotide metabolism, and the concentrations of extracellular nucleotides are significantly higher than those observed in normal cells. In this review, we will focus on the potential role of purinergic signaling in the ten most lethal cancers (lung, breast, colorectal, liver, stomach, prostate, cervical, esophagus, pancreas, and ovary), which together are responsible for more than 5 million annual deaths.

Purinergic signaling in hepatic disease.

Extracellular purines (ATP and adenosine) are ubiquitous intercellular messengers. During tissular damage, they function as damage-associated molecular patterns (DAMPs). In this context, purines announce tissue alterations to initiate a reparative response that involve the formation of the inflammasome complex and the recruitment of specialized cells of the immune system. The present review focuses on the role of the purinergic system in liver damage, mainly during the onset and development of fibrosis. After hepatocellular injury, extracellular ATP promotes a signaling cascade that ameliorates tissue alterations to restore the hepatic function. However, if cellular damage becomes chronic, ATP orchestrates an aberrant reparative process that results in severe liver diseases such as fibrosis and cirrhosis. ATP and adenosine, their receptors, and extracellular ectonucleotidases are mediators of unique processes that will be reviewed in detail.

Creation of optical speckle by randomizing a vortex-lattice.

We investigate the number of vortices embedded in a carrier beam needed to produce a speckle pattern and the necessary conditions in terms of their initial distribution and topological charges. A spatial light modulator is used to imprint arrays of vortices in a Gaussian beam, which is propagated in free space for a given distance and then focused in order to induce interaction among the vortices in the focal region. The resulting optical field is analyzed after propagation up to a transverse plane where the carrier beam would recover its initial size in the absence of vortices. The role of different control parameters for obtaining ordered and disordered patterns is discussed. Our experimental study is complemented with a thorough numerical analysis, from which the statistical properties of the disordered patterns are characterized, and the conditions for obtaining well-developed speckle are determined. We also discuss the creation and annihilation of vortex pairs, depending on the initial conditions.

In Situ Synthesis of Al-Based MMCs Reinforced with AlN by Mechanical Alloying under NH3 Gas.

Aluminum matrix composites (AMCs) reinforced by aluminum nitride were prepared by mechanical alloying followed by a simple press and sintering method. Milling began under vacuum and after a period of between 1 and 4 h, NH3 gas flow (1 cm³/s) was incorporated until the total milling time of 5 h was reached. Results show that in addition to the strain hardening taking place during mechanical alloying, NH3 plays an additional role in powder hardening. Thereby, the properties of the sintered compacts are strongly influenced by the amount of N incorporated into the powders during milling and the subsequent formation of AlN during the consolidation process. The obtained AMC reaches tensile strengths as high as 459 MPa and hardness much higher than that of the as-received aluminum compact.

The influence of yeast on chemical composition and sensory properties of dry white wines.

This study evaluates the impact on two varietal white wines from 'Chardonnay' and 'Verdejo' cultivars of different fermentative strategies: inoculation with Saccharomyces cerevisiae yeast (CT), sequential inoculation (Torulaspora delbrueckii/Saccharomyces cerevisiae) (SI), and spontaneous fermentation (SP). The wines' chemical composition was characterized by oenological parameters, organic acids, metals, major volatile compounds, ester compounds and sensory analyses. The fermentative strategy (CT, SI and SP) was found to be a key factor for assessing different styles of white wines. SI wines showed enhanced 'mature fruit' nuances and a chemical profile characterized by higher content of ethyl propanoate, ethyl isobutyrate and ethyl dihydrocinnamate. Meanwhile, the SP wines presented enhanced "stone fruit" nuances possible related to the higher contents of 2-phenyl acetate and isobutyl acetate. After a chemometric approach the above esters were identified as the markers of each fermentative strategy, independently of the variety.

Cellular Migration Ability Is Modulated by Extracellular Purines in Ovarian Carcinoma SKOV-3 Cells.

Extracellular nucleotides and nucleosides have emerged as important elements regulating tissue homeostasis. Acting through specific receptors, have the ability to control gene expression patterns to direct cellular fate. We observed that SKOV-3 cells express the ectonucleotidases: ectonucleotide pyrophosphatase 1 (ENPP1), ecto-5'-nucleotidase (NT5E), and liver alkaline phosphatase (ALPL). Strikingly, in pulse and chase experiments supplemented with ATP, SKOV-3 cells exhibited low catabolic efficiency in the conversion of ADP into AMP, but they were efficient in converting AMP into adenosine. Since these cells release ATP, we proposed that the conversion of ADP into AMP is a regulatory node associated with the migratory ability and the mesenchymal characteristics shown by SKOV-3 cells under basal conditions. The landscape of gene expression profiles of SKOV-3 cell cultures treated with apyrase or adenosine demonstrated similarities (e.g., decrease FGF16 transcript) and differences (e.g., the negative regulation of Wnt 2, and 10B by adenosine). Thus, in SKOV-3 we analyzed the migratory ability and the expression of epithelium to mesenchymal transition (EMT) markers in response to apyrase. Apyrase-treatment favored the epithelial-like phenotype, as revealed by the re-location of E-cadherin to the cell to cell junctions. Pharmacological approaches strongly suggested that the effect of Apyrase involved the accumulation of extracellular adenosine; this notion was strengthened when the incubation of the SKOV-3 cell with α,ß-methylene ADP (CD73 inhibitor) or adenosine deaminase was sufficient to abolish the effect of apyrase on cell migration. Overall, adenosine signaling is a fine tune mechanism in the control of cell phenotype in cancer. J. Cell. Biochem. 118: 4468-4478, 2017. © 2017 Wiley Periodicals, Inc.

Nucleotides and nucleoside signaling in the regulation of the epithelium to mesenchymal transition (EMT).

The epithelium-mesenchymal transition (EMT) is an important process of cell plasticity, consisting in the loss of epithelial identity and the gain of mesenchymal characteristics through the coordinated activity of a highly regulated informational program. Although it was originally described in the embryonic development, an important body of information supports its role in pathology, mainly in cancerous and fibrotic processes. The purinergic system of inter-cellular communication, mainly based in ATP and adenosine acting throughout their specific receptors, has emerged as a potent regulator of the EMT in several pathological entities. In this context, cellular signaling associated to purines is opening the understanding of a new element in the complex regulatory network of this phenotypical differentiation process. In this review, we have summarized recent information about the role of ATP and adenosine in EMT, as a growing field with high therapeutic potential.

Mechanochemical synthesis in the Li-Mg-N-D system under deuterium gas: a neutron diffraction study.

The Mg(NH2)2/2LiH mixture is considered as one of the most valuable reversible hydrogen storage systems for feeding PEM fuel cells. In this paper, we investigate the mechanochemical synthesis in the Li-Mg-N-H system under deuterium gas, using Li3N and Mg as reactants, and the structural and sorption properties of the intermediate and final products mainly by means of neutron powder diffraction. Mechanochemistry leads to the end formation of amorphous Mg(ND2)2, which crystallizes upon heating above 425 K. During synthesis, a novel cation-mixed nitride/imide phase of simplified composition Li3MgN2D has been unveiled as the intermediate phase. It crystallizes in the cubic disordered anti-fluorite type structure (S.G. Fm3[combining macron]m) with a lattice parameter of 4.996 Å at room temperature. Deuterium absorption in this compound occurs through an original solid solution type mechanism ending with the imide compound ß-Li2MgN2D2. The conjoint use of mechanochemistry under deuterium gas and in situ neutron diffraction techniques offers new avenues for better characterization of the efficient hydrogen storage materials. In particular, this work highlights the unexpected role of intermediate nitride/imide phases in the Li-Mg-N-H system.

Authentication of Italian PDO lard using NIR spectroscopy, volatile profile and fatty acid composition combined with chemometrics.

This study analysed the usefulness of near infrared spectroscopy (NIRS), combined with volatile compound (VOC) and fatty acid (FA) analyses, for the authentication of the unique Italian Valle d'Aosta Arnad Protected Designation of Origin (PDO) lard. Ensuring the authenticity of high value meat products remains an emerging topic within the food sector. This study validated a FA, VOC and NIRS model for use in the authentication of Arnad PDO lard. The model showed a high potential rate to recognize patterns in lard samples. In particular the sensitivity and specificity calibration values were both 100%, and cross-validation models were performed using FAs and VOCs separately. The NIRS model obtained sensitivity and specificity values of 98.2% in the calibration data set, and 94.4% in the cross-validation step. This analytical approach may represent an effective tool to prevent food fraud, which is crucial for meat derived products with a high commercial value.

Effect of management (organic vs conventional) on volatile profiles of six plum cultivars (Prunus salicina Lindl.). A chemometric approach for varietal classification and determination of potential markers.

The volatile profiles of six plum cultivars ('Laetitia', 'Primetime', 'Sapphire', 'Showtime', 'Songold' and 'Souvenir') produced under two management systems (conventional and organic) and harvested in two consecutive years were obtained by HS-SPME-GC-MS. Twenty-five metabolites were determined, five of which (pentanal, (E)-2-heptenal, 1-octanol, eucalyptol and 2-pentylfuran) are reported for the first time in Prunus salicina Lindl. Hexanal stood out as a major volatile compound affected by the management system. In addition, partial least square discriminant analysis (PLS-DA) achieved an effective classification of genotypes based on their volatile profiles. A high classification accuracy model was obtained with a sensitivity of 97.9% and a specificity of 99.6%. Furthermore, the application of a dual criterion, based on a method of variable selection, VIP (variable importance in projection) and the results of a univariate analysis (ANOVA), allowed the identification of potential volatile markers in 'Primetime', 'Showtime' and 'Souvenir' genotypes (cultivars not characterised to date).

Gas-phase synthesis of Mg-Ti nanoparticles for solid-state hydrogen storage.

Mg-Ti nanostructured samples with different Ti contents were prepared via compaction of nanoparticles grown by inert gas condensation with independent Mg and Ti vapour sources. The growth set-up offered the option to perform in situ hydrogen absorption before compaction. Structural and morphological characterisation was carried out by X-ray diffraction, energy dispersive spectroscopy and electron microscopy. The formation of an extended metastable solid solution of Ti in hcp Mg was detected up to 15 at% Ti in the as-grown nanoparticles, while after in situ hydrogen absorption, phase separation between MgH2 and TiH2 was observed. At a Ti content of 22 at%, a metastable Mg-Ti-H fcc phase was observed after in situ hydrogen absorption. The co-evaporation of Mg and Ti inhibited nanoparticle coalescence and crystallite growth in comparison with the evaporation of Mg only. In situ hydrogen absorption was beneficial to subsequent hydrogen behaviour, studied by high pressure differential scanning calorimetry and isothermal kinetics. A transformed fraction of 90% was reached within 100 s at 300 °C during both hydrogen absorption and desorption. The enthalpy of hydride formation was not observed to differ from bulk MgH2.

Kisspeptin is involved in ovarian follicular development during aging in rats.

We have previously reported that kisspeptin (KP) may be under the control of the sympathetic innervation of the ovary. Considering that the sympathetic activity of the ovary increases with aging, it is possible that ovarian KP also increases during this period and participates in follicular development. To evaluate this possibility, we determined ovarian KP expression and its action on follicular development during reproductive aging in rats. We measured ovarian KP mRNA and protein levels in 6-, 8-, 10- and 12-month-old rats. To evaluate follicular developmental changes, intraovarian administration of KP or its antagonist, peptide 234 (P234), was performed using a mini-osmotic pump, and to evaluate FSH receptor (FSHR) changes in the senescent ovary, we stimulated cultured ovaries with KP, P234 and isoproterenol (ISO). Our results shows that KP expression in the ovary was increased in 10- and 12-month-old rats compared with 6-month-old rats, and this increase in KP was strongly correlated with the increase in ovarian norepinephrine observed with aging. The administration of KP produced an increase in corpora lutea and type III follicles in 6- and 10-month-old rats, which was reversed by P234 administration at 10 months. In addition, KP decreased the number and size of antral follicles in 6- and 10-month-old rats, while P234 administration produced an increase in these structures at the same ages. In ovarian cultures KP prevented the induction of FSHR by ISO. These results suggest that intraovarian KP negatively participates in the acquisition of FSHR, indicating a local role in the regulation of follicular development and ovulation during reproductive aging.

The P2RY2 Receptor Induces Carcinoma Cell Migration and EMT Through Cross-Talk With Epidermal Growth Factor Receptor.

Extracellular nucleotides are signaling elements present in the tumor microenvironment; however, their role in tumor growth is not completely understood. In the present study, we asked whether nucleotides regulate cell migration in ovarian carcinoma-derived cells. We observed that 100 µM UTP induced migration in SKOV-3 cells (1.57 ± 0.08 fold over basal), and RT-PCR showed expression of transcripts for the P2RY2 and P2RY4 receptors. Knockdown of P2RY2 expression in SKOV-3 cells (P2RY2-KD) abolished the UTP-induced migration. The mechanism activated by UTP to induce migration involves transactivation of the epidermal growth factor receptor (EGFR) since we observed that the EGFR kinase inhibitor AG1478 and the PI3K inhibitor Wortmannin inhibit this response (to 0.76 ± 0.23 and 0.46 ± 0.14 relative to the control, respectively). In agreement with these observations, UTP was able to modify the phosphorylation state of the EGFR; likewise, the induction of ERK1/2 phosphorylation promoted by UTP was abolished by a 30-60 min treatment with AG1478. Our data also suggested that the enhanced cell migration involves the epithelium to mesenchymal transition (EMT) process, since a 12 h stimulation of SKOV-3 cells with 100 µM UTP showed an increase in vimentin and SNAIL protein levels (459.8 ± 132.4% over basal for SNAIL). Interestingly, treatment with apyrase (10 U/mL) reduces the migration of control cells and induces a considerable enrichment of E-cadherin in the cell-cell contacts, favoring an epithelial phenotype and strongly suggesting that the nucleotides released by tumor cells and acting through the P2RY2 receptor are potential regulators of invasiveness.

Mechanochemistry of lithium nitride under hydrogen gas.

Hydrogen uptake during the mechanochemistry of lithium nitride under 9 MPa hydrogen pressure has been analyzed by means of in situ solid-gas absorption and ex situ X-ray diffraction (XRD) measurements. In situ hydrogenation curves show two H-sorption steps leading to an overall hydrogen uptake of 9.8 wt% H after 3 hours of milling. The milled end-products consist of nanocrystalline (∼10 nm) LiNH2 and LiH phases. The first reaction step comprises the transformation of the polymorph α-Li3N (S.G. P6/mmm) into the ß-Li3N (S.G. P63/mmc) metastable phase and the reaction of the latter with hydrogen to form lithium imide: ß-Li3N + H2→ Li2NH + LiH. Reaction kinetics of the first step is zero-order. Its rate-limiting control is assigned to the collision frequency between milling balls and Li3N powder. In the second absorption step, lithium imide converts to lithium amide following the reaction scheme Li2NH + H2→ LiNH2 + LiH. Reaction kinetics is here limited by one-dimensional nucleation and the growth mechanism, which, in light of structural data, is assigned to the occurrence of lithium vacancies in the imide compound. This study provides new insights into the reaction paths and chemical kinetics of light hydrogen storage materials during their mechanochemical synthesis.

Exploring the high-dimensional structure of muscle redundancy via subject-specific and generic musculoskeletal models.

Subject-specific and generic musculoskeletal models are the computational instantiation of hypotheses, and stochastic techniques help explore their validity. We present two such examples to explore the hypothesis of muscle redundancy. The first addresses the effect of anatomical variability on static force capabilities for three individual cat hindlimbs, each with seven kinematic degrees of freedom (DoFs) and 31 muscles. We present novel methods to characterize the structure of the 31-dimensional set of feasible muscle activations for static force production in every 3-D direction. We find that task requirements strongly define the set of feasible muscle activations and limb forces, with few differences comparing individual vs. species-average results. Moreover, muscle activity is not smoothly distributed across 3-D directions. The second example explores parameter uncertainty during a flying disc throwing motion by using a generic human arm with five DoFs and 17 muscles to predict muscle fiber velocities. We show that the measured joint kinematics fully constrain the eccentric and concentric fiber velocities of all muscles via their moment arms. Thus muscle activation for limb movements is likely not redundant: there is little, if any, latitude in synchronizing alpha-gamma motoneuron excitation-inhibition for muscles to adhere to the time-critical fiber velocities dictated by joint kinematics. Importantly, several muscles inevitably exhibit fiber velocities higher than thought tenable, even for conservative throwing speeds. These techniques and results, respectively, enable and compel us to continue to revise the classical notion of muscle redundancy for increasingly more realistic models and tasks.

GFP labelling and epigenetic enzyme expression of bone marrow-derived mesenchymal stem cells from bovine foetuses.

Mesenchymal stem cells (MSC) are multipotent progenitor cells defined by their ability to self-renew and give rise to differentiated progeny. Since MSC from adult tissues represent a promising source of cells for a wide range of cellular therapies, there is high scientific interest in better understanding the potential for genetic modification and the mechanism underlying differentiation. The main objective of this study was to evaluate the potential for gene delivery using a GFP vector and lipofectamine, and to quantify the expression of epigenetic enzymes during foetal bMSC multilineage differentiation. Proportion of GFP-positive cells achieved (15.7% ± 3.5) indicated moderately low transfection efficiency. Analysis of DNA methyltransferase expression during MSC multilineage differentiation suggested no association with osteogenic and chondrogenic differentiation. However, up-regulation of KDM6B expression during osteogenic differentiation was associated with adoption of osteogenic lineage. Furthermore, increase in epigenetic enzyme expression suggested an intense epigenetic regulation during adipogenic differentiation.

Structure of the set of feasible neural commands for complex motor tasks.

The brain must select its control strategies among an infinite set of possibilities; researchers believe that it must be solving an optimization problem. While this set of feasible solutions is infinite and lies in high dimensions, it is bounded by kinematic, neuromuscular, and anatomical constraints, within which the brain must select optimal solutions. That is, the set of feasible activations is well structured. However, to date there is no method to describe and quantify the structure of these high-dimensional solution spaces. Bounding boxes or dimensionality reduction algorithms do not capture their detailed structure. We present a novel approach based on the well-known Hit-and-Run algorithm in computational geometry to extract the structure of the feasible activations capable of producing 50% of maximal fingertip force in a specific direction. We use a realistic model of a human index finger with 7 muscles, and 4 DOFs. For a given static force vector at the endpoint, the feasible activation space is a 3D convex polytope, embedded in the 7D unit cube. It is known that explicitly computing the volume of this polytope can become too computationally complex in many instances. However, our algorithm was able to sample 1,000,000 uniform at random points from the feasible activation space. The computed distribution of activation across muscles sheds light onto the structure of these solution spaces-rather than simply exploring their maximal and minimal values. Although this paper presents a 7 dimensional case of the index finger, our methods extend to systems with at least 40 muscles. This will allow our motor control community to understand the distributions of feasible muscle activations, providing important contextual information into learning, optimization and adaptation of motor patterns in future research.

Lower extremity dexterity is associated with agility in adolescent soccer athletes.

Agility is important for sport performance and potentially injury risk; however, factors affecting this motor skill remain unclear. Here, we evaluated the extent to which lower extremity dexterity (LED) and muscle performance were associated with agility. Fourteen male and 14 female soccer athletes participated. Agility was evaluated using a hopping sequence separately with both limbs and with the dominant limb only. The LED test evaluated the athletes' ability to dynamically regulate foot-ground interactions by compressing a spring prone to buckling with the lower limb. Muscle performance included hip and knee isometric strength and vertical jump height. Correlation analyses were used to assess the associations between muscle performance, LED, and agility. Multiple regression models were used to determine whether linear associations differed between sexes. On average, the female athletes took longer to complete the agility tasks than the male athletes. This difference could not be explained by muscle performance. Conversely, LED was found to be the primary determinant of agility (double limb: R(2) = 0.61, P < 0.001; single limb: R(2) = 0.63, P < 0.001). Our findings suggest that the sensorimotor ability to dynamically regulate foot-ground interactions as assessed by the LED test is predictive of agility in soccer athletes. We propose that LED may have implications for sport performance, injury risk, and rehabilitation.