Sulfated Polysaccharides from Macroalgae Are Potent Dual Inhibitors of Human ATP-Hydrolyzing Ectonucleotidases NPP1 and CD39

Extracellular ATP mediates proinflammatory and antiproliferative effects via activation of P2 nucleotide receptors. In contrast, its metabolite, the nucleoside adenosine, is strongly immunosuppressive and enhances tumor proliferation and metastasis. The conversion of ATP to adenosine is catalyzed by ectonucleotidases, which are expressed on immune cells and typically upregulated on tumor cells. In the present study, we identified sulfopolysaccharides from brown and red sea algae to act as potent dual inhibitors of the main ATP-hydrolyzing ectoenzymes, ectonucleotide pyrophosphatase/phosphodiesterase-1 (NPP1) and ecto-nucleoside triphosphate diphosphohydrolase-1 (NTPDase1, CD39), showing nano- to picomolar potency and displaying a non-competitive mechanism of inhibition. We showed that one of the sulfopolysaccharides tested as a representative example reduced adenosine formation at the surface of the human glioblastoma cell line U87 in a concentration-dependent manner. These natural products represent the most potent inhibitors of extracellular ATP hydrolysis known to date and have potential as novel therapeutics for the immunotherapy of cancer.     

Cartap activity on the cockroach nervous and neuromuscular transmission

By employing intracellular electrodes on the 6th abdominal ganglion, Cartap hydrochloride 10^?5M caused in all experiments a block of the provoked stimulus transmission and a decrease of the cell membrane resting potential; the giant fiber conduction was not affected. In the experiments with extracellular electrodes Cartap 10^?5M provoked a marked increase of the spontaneous activity followed by block which partially disappeared after washing. The same effects were obtained on spontaneous activity when Cartap 10^?5M was used on the denervated 6th abdominal ganglion and in experiments conducted at 0 Ca²⁺ or at 20 mM Mg²⁺. Cartap 10^?5M did not affect the response to direct or indirect stimulation of cockroach neuromuscular preparation. These results tend to confirm that Cartap affects the postsynaptic region of the ganglionic nervous junction. The possible cause of the resting potential decrease is also discussed.     

Localization of orexin B and receptor 2 for orexins in testicular cytotypes of the camelid alpaca (Vicugna pacos)

The orexins A (OxA) and B (OxB) are two hypothalamic peptides involved in many physiological functions of the mammalian body. They act through the binding of two G‐coupled receptors named receptor 1 (OX₁) and receptor 2 (OX₂) for orexins. The first receptor is specific for OxA, while the second binds both the substances with equal affinity. The orexins and the relative receptors have been traced by means of different techniques also at the periphery of the body and particularly in the adrenals, and in gastrointestinal and genital organs. Aim of this work was to investigate the presence of OxB and OX₂ by means of immunohistochemistry and Western blotting analysis in the testis of the South American camelid alpaca, a species primarily breed in Chile and Ecuador and recently diffused in Europe where the quality of its wool is particularly appreciated. OxB immunoreactivity (IR) was found in the tubular compartment of the testis where spermatogonia (resting), zygotene and pachytene spermatocytes, and spermatids clearly showed differently sized and shaped cytoplasmic positive structures. OX₂‐IR was found both in the interstitial and tubular compartments of the testis and particularly in Leydig cells and round and elongated spermatids. Western blotting analysis of testis lysates showed the presence of a protein band whose molecular weight corresponded to that currently assigned to OX₂. Such findings easily translate the hypothesis that OxB and its receptor 2 play a functional role both in the interstitial and tubular compartments of the alpaca testis.     

Role of secondary metabolites in the biocontrol activity of <Emphasis Type="Italic">Pseudomonas corrugata</Emphasis> and <Emphasis Type="Italic">Pseudomonas mediterranea</Emphasis>

Colletotrichum gloeosporioides is the causal agent of Camellia oleifera anthracnose, mainly infecting fruits and leaves. The fungus secretes degrading enzymes to destroy the cuticle of aerial plant parts and help infect the host successfully. To validate whether a cutinase gene (CglCUT1) was required for cutinase activity and pathogenicity of C. gloeosporioides, the CglCUT1 gene was cloned and analyzed. The characterization of CglCUT1 predicted protein suggests that the cloned DNA encoded a cutinase in C. gloeosporioides affecting C. oleifera. The CglCUT1 showed a high homology to those from C. gloeosporioides causing papaya anthracnose and C. capsici causing pepper anthracnose, as well as those of other ascomycetes. The whole CglCUT1 gene was knocked-out and the knockout mutant (?CglCUT39) was subsequently complemented using Agrobacterium tumefaciens mediated transformation. The knockout transformants exhibited significant decreases in cutinase activity and virulence compared with the wild-type strain. The complemented transformants of the disrupted transformant ?CglCUT39 showed a significant increase in cutinase activity and virulence compared with the disrupted transformant ?CglCUT39. This study suggests that the CglCUT1 gene has a positive effect on fungal virulence of the hemibiotrophic C. gloeosporioides on C. oleifera.     

Hydrolysis and adsorption of cyhalofop-butyl and cyhalofop-acid on soil colloids

A study was undertaken to investigate the stability of cyhalofop-butyl (2 R)-2-[4-(4-cyano-2-fluorophenoxy)phenoxy]butylpropanoate (CyB), an aryloxyphenoxy-propionic herbicide, at different pH values. The hydrolysis of CyB was faster in nonsterile than in sterile water. In sterile medium, CyB degraded only to (2 R)-2-[4-(4-cyano-2-fluorophenoxy)phenoxy]propanoic acid (CyA), whereas in nonsterile water, also the metabolites (2 R)-2-[4-(4-carbamoyl-2-fluorophenoxy)phenoxy]propanoic acid (CyAA) and (2 R)-2-[4-(4-carboxyl-2-fluorophenoxy)phenoxy]propanoic acid (CyD) were detected. The adsorption of CyB onto clays, iron oxide, and dissolved organic matter (DOM), using a batch equilibrium method, was also studied. A lipophilic bond is responsible for CyB adsorption on DOM. CyB was adsorbed on Fe(III)- and Ca-clays through hydrogen bonding between the carbonyl oxygen and water surrounding the exchangeable cations. In the interlayer of K-clay, CyB was hydrolyzed to CyA, which remained adsorbed therein as a monomer. The acid CyA was adsorbed only by the Fe-oxide through complexation. The CyA-Fe-oxide complex was stable and did not undergo degradation.     

Potential Biomedical Applications of Collagen Filaments derived from the Marine Demosponges Ircinia oros (Schmidt, 1864) and Sarcotragus foetidus (Schmidt, 1862)

Collagen filaments derived from the two marine demosponges Ircinia oros and Sarcotragus foetidus were for the first time isolated, biochemically characterised and tested for their potential use in regenerative medicine. SDS-PAGE of isolated filaments revealed a main collagen subunit band of 130 kDa in both of the samples under study. DSC analysis on 2D membranes produced with collagenous sponge filaments showed higher thermal stability than commercial mammalian-derived collagen membranes. Dynamic mechanical and thermal analysis attested that the membranes obtained from filaments of S. foetidus were more resistant and stable at the rising temperature, compared to the ones derived from filaments of I. oros. Moreover, the former has higher stability in saline and in collagenase solutions and evident antioxidant activity. Conversely, their water binding capacity results were lower than that of membranes obtained from I. oros. Adhesion and proliferation tests using L929 fibroblasts and HaCaT keratinocytes resulted in a remarkable biocompatibility of both developed membrane models, and gene expression analysis showed an evident up-regulation of ECM-related genes. Finally, membranes from I. oros significantly increased type I collagen gene expression and its release in the culture medium. The findings here reported strongly suggest the biotechnological potential of these collagenous structures of poriferan origin as scaffolds for wound healing.     

Priority knowledge needs for management of migratory fish species in Cambodia

Despite their economic and ecological importance, migratory fishes of the Lower Mekong Basin (LMB) remain understudied, which hampers effective management to sustain valuable fisheries and address serious threats such as habitat degradation, development and overharvest. From a list of potential knowledge needs, a group of fisheries professionals most frequently identified six top priorities for managing migratory fishes in Cambodia: (1) population abundances and trends, (2) life cycles and life history, (3) migration timing and triggers, (4) migration routes and distances, (5) locations of key habitats and spawning areas, and (6) environmental and habitat requirements. These needs are discussed along with nine relevant methodologies for addressing them, including fisheries-dependent and fisheries-independent sampling, reproductive techniques and captive studies, otolith and genetic analysis tools, and tagging and imaging techniques. A suggested research framework is also presented to inform adaptive management of migratory fishes. While emphasis is given to Cambodia, the analysis is also applicable to other LMB countries, given that migratory fishes occur throughout the basin and migrate across borders. It is suggested that a robust research and monitoring agenda is required to prioritise knowledge needs and select appropriate methodologies to answer questions vital to inform sustainable migratory fish management in Cambodia.     

Finite element analysis of a novel pin-sleeve system for external fixation of distal limb fractures in horses

The transfixation pin cast (TPC) is an external skeletal fixation technique used to treat horses with distal limb fractures, but its use is often associated with pin-loosening and an increased risk of treatment failure. To address implant loosening, the pin sleeve cast system (PSC) was recently designed and consists of a pin-sleeve unit inserted into the bone. Each pin runs through a sleeve placed in the bone, making contact at two fixed points only within the sleeve. Each pin is attached to a ring embedded in a resin cast. In this report, the mechanical performance of a traditional TPC pin arrangement was compared with that of the PSC using validated finite element models of bone substitutes previously tested in vitro. The PSC resulted in a marked reduction in peak strain magnitude around the pins and a more even distribution of strain across the bone cortex. The two systems resulted in comparable proximal fragment displacement and had a similar stress concentration around bone defects during implant removal. The findings suggest that the PSC load transfer mechanism is effective even in geometrically complex structures like equine bones.     

Clinical pharmacokinetics of a dexmedetomidine–methadone combination in dogs undergoing routine anaesthesia after buccal or intramuscular administration

This study aimed to define the pharmacokinetic profiles of dexmedetomidine and methadone administered simultaneously in dogs by either an oral transmucosal route or intramuscular route and to determine the bioavailability of the oral transmucosal administration relative to the intramuscular one of both drugs, so as the applicability of this administration route in dogs. Twelve client‐owned dogs, scheduled for diagnostic procedures, were treated with a combination of dexmedetomidine hydrochloride (10 μg/kg) and methadone hydrochloride (0.4 mg/kg) through an oral transmucosal route or intramuscularly. Oral transmucosal administration caused ptyalism in most subjects, and intramuscular administration caused transient peripheral vasoconstriction. The results showed reduced and delayed absorption of both dexmedetomidine and methadone when administered through an oral transmucosal route, with median (range) C_max values of 0.82 (0.42–1.49) ng/ml and 13.22 (2.80–52.30) ng/ml, respectively. The relative bioavailability was low: 16.34% (dexmedetomidine) and 15.5% (methadone). Intramuscular administration resulted in a more efficient absorption profile, with AUC and C_max values for both drugs approximately 10 times higher. Dexmedetomidine and methadone administered simultaneously by an oral transmucosal route using injectable formulations were not well absorbed through the oral mucosa. Nevertheless, additional studies on these drugs combination using alternative administration routes are recommended.