Isolation of the Bα and Bβ mating-type loci of Schizophyllum commune

The B mating type of the basidiomycete fungus, Schizophyllum commune is determined by two, tightly linked, multi-specificity (also called multi-allelic) loci: B and B. A plasmid library was used in DNA-mediated transformation to obtain transformants that displayed B-directed development. Plasmids that conferred B1 and B1 mating-type specificities were rescued from the transformants. Fragments of DNA from each plasmid hybridized to genomic DNA from the strain used to make the plasmid library; however, they did not hybridize, or hybridized only weakly, to genomic DNA from strains with mating-type specificities different from B1 or B1. The cloned fragments are presumed to correspond to active regions of each B mating-type locus.     

The effect of tumour necrosis factor-α and insulin on equine digital blood vessel function in vitro

Objective and design

Insulin and inflammatory cytokines may be involved in equine laminitis, which might be associated with digital vascular dysfunction. This study determined the effects of TNF-α and insulin on the endothelial-dependent relaxant responses of equine digital blood vessels and on equine digital vein endothelial cell (EDVEC) cGMP production.

Material

Isolated rings of equine digital arteries (EDAs) and veins (EDVs) were obtained and EDVECs were cultured from horses euthanized at an abattoir.

Methods

The effect of incubation with TNF-α (10 ng/ml) and/or insulin (1,000 μIU/ml) for 1.5 h or overnight under hyperoxic and hypoxic conditions on carbachol (endothelium-dependent) induced relaxation was assessed. The time course and concentration dependency of the effect of TNF-α and the effect of insulin (1,000 μIU/ml) on EDVEC cGMP production was determined.

Results

Incubation of EDAs overnight with TNF-α under hypoxic conditions resulted in endothelial-dependent vascular dysfunction. EDVs produced a more variable response. TNF-α increased EDVEC cGMP formation in a time- and concentration-dependent manner. Insulin had no significant effects.

Conclusions

There is a mismatch between the results obtained from isolated vessel rings and cultured endothelial cells suggesting TNF-α may reduce the biological effect of NO by reducing its bioavailability rather than its formation, leading to endothelial cell dysregulation.     

OX40: Structure and function – What questions remain?

OX40 is a type 1 transmembrane glycoprotein, reported nearly 30 years ago as a cell surface antigen expressed on activated T cells. Since its discovery, it has been validated as a bone fide costimulatory molecule for T cells and member of the TNF receptor family. However, many questions still remain relating to its function on different T cell sub-sets and with recent interest in its utility as a target for antibody-mediated immunotherapy, there is a growing need to gain a better understanding of its biology.Here, we review the expression pattern of OX40 and its ligand, discuss the structure of the receptor:ligand interaction, the downstream signalling it can elicit, its function on different T cell subsets and how antibodies might engage with it to provide effective immunotherapy.     

Histo- and immunocytochemical detection of inducible NOS and TNF-α in the locus coeruleus of human opiate addicts

In order to determine the role of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-α (TNF-α) in the pathogenesis of opiate addiction in humans, the expression of both was analyzed in the locus coeruleus (LC) of patients who died from heroin overdose. In control subjects, NADPH-diaphorase (NADPHd) histochemistry was mainly observed in non-noradrenerdic neurons, some glial and endothelial cells. However, in the brain of opiate addicts, NADPHd and iNOS expression was detected in noradrenergic LC cells, correlating with an increase in iNOS and TNF-α expression in glial cells as revealed by immunohistochemical and Western blot analyses. These findings indicate that sustained overproduction of cytokine and NO via iNOS expression may be responsible, at least in part, for some neurochemical changes in the locus coeruleus caused by chronic opiate usage in humans.     

The interplay between nanostructured carbon-grafted chitosan scaffolds and protein adsorption on the cellular response of osteoblasts: Structure–function property relationship

The rapid adsorption of proteins occurs during the early stages of biomedical device implantation into physiological systems. In this regard, the adsorption of proteins is a strong function of the nature of a biomedical device, which ultimately governs the biological functions. The objective of this study was to elucidate the interplay between nanostructured carbon-modified (graphene oxide and single-walled carbon nanohorn) chitosan scaffolds and consequent protein adsorption and biological function (osteoblast function). We compare and contrast the footprint of protein adsorption on unmodified chitosan and nanostructured carbon-modified chitosan. A comparative analysis of cell–substrate interactions using an osteoblast cell line (MC3T3-E1) implied that biological functions were significantly enhanced in the presence of nanostructured carbon, compared with unmodified chitosan. The difference in their respective behaviors is related to the degree and topography of protein adsorption on the scaffolds. Furthermore, there was a synergistic effect of nanostructured carbon and protein adsorption in terms of favorably modulating biological functions, including cell attachment, proliferation and viability, with the effect being greater on nanostructured carbon-modified scaffolds. The study also underscores that protein adsorption is favored in nanostructured carbon-modified scaffolds such that bioactivity and biological function are promoted.     

A normative‐developmental study of executive function: A window on prefrontal function in children

Normative‐developmental performance on a battery of executive function tasks was investigated. Executive function was defined as goal‐directed behavior, including planning, organized search, and impulse control. Measures were drawn from clinical neuropsychology (visual search, verbal fluency, motor sequencing, and Wisconsin Card Sorting Task [WCST]) and from developmental psychology (Tower of Hanoi [TOH] and Matching Familiar Figures Test [MFFT]). A discriminant task, recognition memory, was administered, and IQ scores were available on a subset of the sample. One hundred subjects ranging from 3 to 12 years old participated; an adult group was also studied. Three major results were found: (a) adult‐level performance on different subsets of the executive function tasks was achieved at three different ages—6 years old, 10 years old, and adolescence; (b) the measures clustered into three different factors reflecting speeded responding, set maintenance, and planning; and (c) most of the executive function tasks were uncorrelated with IQ. The implications of these results for our understanding of the development of prefrontal lobe functions are discussed.     

Structure–function relationship of working memory activity with hippocampal and prefrontal cortex volumes

A rapidly increasing number of studies are quantifying the system-level network architecture of the human brain based on structural-to-structural and functional-to-functional relationships. However, a largely unexplored area is the nature and existence of “cross-modal” structural–functional relationships, in which, for example, the volume (or other morphological property) of one brain region is related to the functional response to a given task either in that same brain region, or another brain region. The present study investigated whether the gray matter volume of a selected group of structures (superior, middle, and inferior frontal gyri, thalamus, and hippocampus) was correlated with the fMRI response to a working memory task, within a mask of regions previously identified as involved with working memory. The subjects included individuals with schizophrenia, their siblings, and healthy controls (n = 154 total). Using rigorous permutation testing to define the null distribution, we found that the volume of the superior and middle frontal gyri was correlated with working memory activity within clusters in the intraparietal sulcus (i.e., dorsal parietal cortex) and that the volume of the hippocampus was correlated with working memory activity within clusters in the dorsal anterior cingulate cortex and left inferior frontal gyrus. However, we did not find evidence that the identified structure–function relationships differed between subject groups. These results show that long-distance structural–functional relationships exist within the human brain. The study of such cross-modal relationships represents an additional approach for studying systems-level interregional brain networks.     

Structure and function of Norrin in assembly and activation of a Frizzled 4–Lrp5/6 complex

Norrin is a cysteine-rich growth factor that is required for angiogenesis in the eye, ear, brain, and female reproductive organs. It functions as an atypical Wnt ligand by specifically binding to the Frizzled 4 (Fz4) receptor. Here we report the crystal structure of Norrin, which reveals a unique dimeric structure with each monomer adopting a conserved cystine knot fold. Functional studies demonstrate that the novel Norrin dimer interface is required for Fz4 activation. Furthermore, we demonstrate that Norrin contains separate binding sites for Fz4 and for the Wnt ligand coreceptor Lrp5 (low-density lipoprotein-related protein 5) or Lrp6. Instead of inducing Fz4 dimerization, Norrin induces the formation of a ternary complex with Fz4 and Lrp5/6 by binding to their respective extracellular domains. These results provide crucial insights into the assembly and activation of the Norrin–Fz4–Lrp5/6 signaling complex.     

The immunomodulatory effects of barettin and involvement of the kinases CAMK1α and RIPK2

Abstract

Background: Barettin is a marine natural compound with reported anti-inflammatory and antioxidant properties. The combination of these effects led us to explore barettin further as an inhibitor of atherosclerosis development.

Methods: The effect of barettin on MCP-1 and IL-10 secretion from activated immune cells was detected by ELISA. Determination of cell viability of oxidized low-density lipoprotein (oxLDL) and barettin exposed HUVEC cells were investigated by using CellTiter 96® AQ_ueous One Solution. The kinase inhibition assays were performed using a radioactive (³³P-ATP) filter binding assay at the University of Dundee, UK.

Results: Barettin reduces the secretion of monocyte chemotactic protein-1 (MCP-1) from LPS-stimulated monocytes, but was not able to prevent oxLDL-induced cell death in HUVEC. Barettin has inhibitory activity against two protein kinases related to inflammation, namely the receptor-interacting serine/threonine kinase 2 (RIPK2) and calcium/calmodulin-dependent protein kinase 1α (CAMK1α). We also demonstrate that barettin reduce the production of the anti-inflammatory cytokine interleukin-10 (IL-10) in a dose and time-dependent manner, possibly by inhibiting CAMK1α.

Conclusions: The anti-inflammatory activity of barettin is exerted through the regulation of inflammatory mediators such as MCP-1 and IL-10, possibly via inhibition of kinases.     

The vertebral column—A phylogenetic failure? A theory explaining the function and vulnerability of the human spine

The increase in degenerative diseases of the vertebral column is often attributed to an inadequate adaptation to the upright posture in the human. On the basis of a precise analysis of the motion segments, an attempt has here been made to demonstrate that the design of the vertebrae in both the larger mammals and in humans has resulted in no qualitative differences between the stresses to which either is subjected. In the course of evolution there has certainly been an obvious conflict in aims between the need for essential stability and the desired or necessary mobility. These mutually self-limiting mechanisms are reflected in the highly specialized architecture of the ligamentous apparatus and vertebral joints. We conclude that the human vertebral column seems to be an optimized compromise of evolution. © 1996 Wiley-Liss, Inc.     

Structure–function relationships and source-to-ground distance in electrospun polycaprolactone

The strength of electrospun scaffolds has direct relevance to their function within tissue engineering. We characterized the effects of source-to-ground distance on the mechanical properties of electrospun poly(ε-caprolactone) (PCL). Source-to-ground distances of 10, 15 and 20 cm, solids concentrations of 12 and 18 wt.% and mandrel rotation surface speeds of 0–12 m s^?1 were utilized. Tensile tests evaluated elastic modulus, tensile strength and elongation at failure. Scanning electron microscopy provided morphology and quantified fiber alignment. Increased source-to-ground distance yielded a microstructure allowing greater fiber rearrangement under load, tripling the observed tensile strength. Increases in rotational speed generally increased fiber alignment and strength at high but not low to moderate speeds. As fiber is quickly pulled out of a comparatively gentle falling process, collision with neighboring fibers moving at different speeds and in different directions can occur. The source-to-ground distance influences these collisions and thus has critical implications for microstructure and biocompatibility. In larger diameter (18 wt.% PCL), heavily point-bonded fibers (produced using a shorter, 10 cm source-to-ground distance), elongation at failure in the aligned direction increases dramatically due to severe localized necking. These specimens show only half of the tensile strength (from 2.6 to 4.5 MPa) and a dramatic increase (from 94% to 503%) in elongation at failure vs. a longer 20 cm source-to-ground distance. Strains of several hundred per cent are accompanied by periodic necking of large-diameter fibers in which microstructural failure appears to occur in a sequential manner involving an equilibrium between localized strain in the tensile direction and anisotropic point bonding that locally resists strain.     

Parental Smoking and Adolescent Smoking Stages: The Role of Parents’ Current and Former Smoking, and Family Structure

This study examined the role of parents’ current and former smoking in predicting adolescent smoking acquisition stages. Participants were 7,426 students from 33 schools in the Netherlands. Participants’ survey data were gathered at baseline and at two-year follow-up. Logistic regression models showed that parental smoking status was not only predictive of transitions from never smoking to trying smoking, monthly smoking, or daily smoking, but also of the progression from trying smoking to daily smoking. Further, although parental former smoking was weaker associated with progressive adolescent smoking transitions than current parental smoking, however absence of parental smoking history was most preventive. Compared to the situation in which both parents had never smoked, cessation of parental smoking after the child was born was associated with an increased risk for children to smoke. Adolescents living in a single-parent family were at greater risk of smoking than adolescents living in an intact family with both mother and father. In sum, the role of parental smoking is not restricted to smoking onset and is present throughout different phases of the acquisition process. Results support the delayed modeling hypothesis that parental smoking affects the likelihood for children to smoke even when parents quit many years before. Children living in single-parent families are only exposed to the behaviour of one parent; in two-parent families the behaviour from one parent may magnify or buffer the behaviour of the other parent.     

A ligand-independent fast function of RARα promotes exit from metabolic quiescence upon T cell activation and controls T cell differentiation

Vitamin A metabolites play important roles in T cell activation and differentiation. A conventional model of RARα function relies upon retinoic acid (RA)-liganded RARα binding to specific DNA motifs to regulate gene expression in the nucleus. However, this genomic function fails to explain many of the biological responses of the RA-RARα axis on T cells. We generated a mouse line where RARα is over-expressed in T cells to probe RARα function with unprecedented sensitivity. Using this model together with mice specifically lacking RARα in T cells, we found that RARα is required for prompt exit from metabolic quiescence in resting T cells upon T cell activation. The positive effect of RARα on metabolism is mediated through PI3K and subsequent activation of the Akt and mTOR signaling pathway. This largely non-genomic function of RARα is surprisingly ligand-independent and controls the differentiation of effector and regulatory T cell subsets.     

A study of properties of “micelle-enhanced” polyelectrolyte capsules: Structure,encapsulation and in vitro release

“Micelle-enhanced” polyelectrolyte capsules were fabricated via a layer-by-layer technique, templated on hybrid calcium carbonate particles with built-in polymeric micelles based on polystyrene-b-poly(acrylic acid). Due to the presence of a large number of negatively charged micelles inside the polyelectrolyte capsule, which were liberated from templates, the capsule wall was reconstructed and had properties different to those of conventional polyelectrolyte capsules. This type of capsule could selectively entrap positively charged water-soluble substances. The encapsulation efficiency of positively charged substances was dependent on their molecular weight or size. For some positively charged compounds, such as rhodamine B and lysozyme, the concentration in the capsules was orders of magnitude higher than that in the incubation solution. In addition, in vitro release study suggested that the encapsulated compounds could be released through a sustained manner to a certain degree. All these results point to the fact that these capsules might be used as novel delivery systems for some water-soluble compounds.     

Individualism and immune function: are asthma and allergies partly a function of an overly constricted self?

Relations of individualism/collectivism to asthma and allergies were examined in two studies. I proposed that a narrower psychological identity (individualism) might be associated with an overactive immune system. In Study 1, average individualism levels across 15 countries correlated significantly positively (.50) with national asthma rates. Pollution and crowding levels were unrelated to national asthma rates. In Study 2, higher levels of personality individualism were associated with higher numbers of allergies among college students even with some other individual difference factors controlled. Potential mechanisms behind, and potential implications of, these results are described. Some suggestions for further research on this topic are given.     

The contractile vacuole complex of protists – New cues to function and biogenesis

Abstract

The contractile vacuole complex (CVC) of freshwater protists sequesters the excess of water and ions (Ca²⁺) for exocytosis cycles at the pore. Sequestration is based on a chemiosmotic proton gradient produced by a V-type H⁺-ATPase. So far, many pieces of information available have not been combined to a comprehensive view on CVC biogenesis and function. One main function now appears as follows. Ca²⁺-release channels, type inositol 1,4,5-trisphosphate receptors (InsP₃R), may serve for fine-tuning of local cytosolic Ca²⁺ concentration and mediate numerous membrane-to-membrane interactions within the tubular spongiome meshwork. Such activity is suggested by the occurrence of organelle-specific soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) and Ras-related in brain (Rab) proteins, which may regulate functional requirements. For tubulation, F-Bin–amphiphysin–Rvs (F-BAR) proteins are available. In addition, there is indirect evidence for the occurrence of H⁺/Ca²⁺ exchangers (to sequester Ca²⁺) and mechanosensitive Ca²⁺-channels (for signaling the filling sate). The periodic activity of the CVC may be regulated by the mechanosensitive Ca²⁺-channels. Such channels are known to colocalize with and to be functionally supported by stomatins, which were recently detected in the CVC. A Kif18-related kinesin motor protein might control the length of radial arms. Two additional InsP₃-related channels and several SNAREs are associated with the pore. De novo organelle biogenesis occurs under epigenetic control during mitotic activity and may involve the assembly of γ-tubulin, centrin, calmodulin and a never in mitosis A-type (NIMA) kinase – components also engaged in mitotic processes.