Revascularization and tissue regeneration of an empty root canal space is enhanced by a direct blood supply and stem cells

Background Regenerative endodontics is an innovative treatment concept aiming to regenerate pulp, dentin and root structures. In the diseased or necrotic tooth, the limitation in vascular supply renders successful tissue regeneration/generation in a whole tooth challenging. The aim of this study is to evaluate the ability of vascularized tissue to develop within a pulpless tooth using tissue engineering techniques. Materials and methods A pulpless tooth chamber, filled with collagen I gel containing isolated rat dental pulp cells (DPC) and angiogenic growth factors, was placed into a hole created in the femoral cortex or into its own tooth socket, respectively. The gross, histological and biochemical characteristics of the de novo tissue were evaluated at 4 and 8weeks post-transplantation. Results Tooth revascularization and tissue generation was observed only in the femur group, confirming the important role of vascular supply in tissue regeneration. The addition of cells and growth factors significantly promoted connective tissue production in the tooth chamber. Conclusion Successful revascularization and tissue regeneration in this model demonstrate the importance of a direct vascular supply and the advantages of a stem cell approach. © 2012 John Wiley & Sons A/S.

Induction of epithelial-mesenchymal transition (EMT) in breast cancer cells is calcium signal dependent

Signals from the tumor microenvironment trigger cancer cells to adopt an invasive phenotype through epithelial-mesenchymal transition (EMT). Relatively little is known regarding key signal transduction pathways that serve as cytosolic bridges between cell surface receptors and nuclear transcription factors to induce EMT. A better understanding of these early EMT events may identify potential targets for the control of metastasis. One rapid intracellular signaling pathway that has not yet been explored during EMT induction is calcium. Here we show that stimuli used to induce EMT produce a transient increase in cytosolic calcium levels in human breast cancer cells. Attenuation of the calcium signal by intracellular calcium chelation significantly reduced epidermal growth factor (EGF)- and hypoxia-induced EMT. Intracellular calcium chelation also inhibited EGF-induced activation of signal transducer and activator of transcription 3 (STAT3), while preserving other signal transduction pathways such as Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation. To identify calcium-permeable channels that may regulate EMT induction in breast cancer cells, we performed a targeted siRNA-based screen. We found that transient receptor potential-melastatin-like 7 (TRPM7) channel expression regulated EGF-induced STAT3 phosphorylation and expression of the EMT marker vimentin. Although intracellular calcium chelation almost completely blocked the induction of many EMT markers, including vimentin, Twist and N-cadherin, the effect of TRPM7 silencing was specific for vimentin protein expression and STAT3 phosphorylation. These results indicate that TRPM7 is a partial regulator of EMT in breast cancer cells, and that other calcium-permeable ion channels are also involved in calcium-dependent EMT induction. In summary, this work establishes an important role for the intracellular calcium signal in the induction of EMT in human breast cancer cells. Manipulation of calcium-signaling pathways controlling EMT induction in cancer cells may therefore be an important therapeutic strategy for preventing metastases.

Actinidia DRM1 : an intrinsically disordered protein whose mRNA expression is inversely correlated with spring budbreak in Kiwifruit

Intrinsically disordered proteins (IDPs) are a relatively recently defined class of proteins which, under native conditions, lack a unique tertiary structure whilst maintaining essential biological functions. Functional classification of IDPs have implicated such proteins as being involved in various physiological processes including transcription and translation regulation, signal transduction and protein modification. Actinidia DRM1 (Ade DORMANCY ASSOCIATED GENE 1), represents a robust dormancy marker whose mRNA transcript expression exhibits a strong inverse correlation with the onset of growth following periods of physiological dormancy. Bioinformatic analyses suggest that DRM1 is plant specific and highly conserved at both the nucleotide and protein levels. It is predicted to be an intrinsically disordered protein with two distinct highly conserved domains. Several Actinidia DRM1 homologues, which align into two distinct Actinidia-specific families, Type I and Type II, have been identified. No candidates for the Arabidopsis DRM1-Homologue (AtDRM2) an additional family member, has been identified in Actinidia.

An ancient duplication of apple MYB transcription factors is responsible for novel red fruit-flesh phenotyp

Anthocyanin accumulation is coordinated in plants by a number of conserved transcription factors. In apple (Malus × domestica), an R2R3 MYB transcription factor has been shown to control fruit flesh and foliage anthocyanin pigmentation (MYB10) and fruit skin color (MYB1). However, the pattern of expression and allelic variation at these loci does not explain all anthocyanin-related apple phenotypes. One such example is an open-pollinated seedling of cv Sangrado that has green foliage and develops red flesh in the fruit cortex late in maturity. We used methods that combine plant breeding, molecular biology, and genomics to identify duplicated MYB transcription factors that could control this phenotype. We then demonstrated that the red-flesh cortex phenotype is associated with enhanced expression of MYB110a, a paralog of MYB10. Functional characterization of MYB110a showed that it was able to up-regulate anthocyanin biosynthesis in tobacco (Nicotiana tabacum). The chromosomal location of MYB110a is consistent with a whole-genome duplication event that occurred during the evolution of apple within the Maloideae family. Both MYB10 and MYB110a have conserved function in some cultivars, but they differ in their expression pattern and response to fruit maturity.

There is less to this argument than meets the eye

In this article the authors continue the Artificial Intelligence and the law debate begun with Moles' 1991 article. In it the authors answer the latest criticisms made by Moles and others as they explain and argue the case for the practical benefits to be gained by AI systems involving the law.

School literate repertoires : that was then, this is now

This chapter documents the literacy learning lives of two young Australians now in their early twenties. Both were originally informants in studies of literacy development. The original studies were longitudinal, so in each case there was already a story of change, especially in the young people’s identities as school. The case studies presented here, Tessa and Cruz, tell different stories of learning literacy at school, teaching that makes a difference and raise questions about predictable normative developmental trajectories. Both young people have taken up, albeit selectively, valuable literacy and learner dispositions from school and their wider everyday worlds.

Clinical dyslipidaemia is associated with changes in the lipid composition and inflammatory properties of apolipoprotein-B-containing lipoproteins from women with type 2 diabetes

The aim of this study was to use lipidomics to determine if the lipid composition of apolipoprotein-B-containing lipoproteins is modified by dyslipidaemia in type 2 diabetes and if any of the identified changes potentially have biological relevance in the pathophysiology of type 2 diabetes. VLDL and LDL from normolipidaemic and dyslipidaemic type 2 diabetic women and controls were isolated and quantified with HPLC and mass spectrometry. A detailed molecular characterisation of VLDL triacylglycerols (TAG) was also performed using the novel ozone-induced dissociation method, which allowed us to distinguish vaccenic acid (C18:1 n-7) from oleic acid (C18:1 n-9) in specific TAG species. Lipid class composition was very similar in VLDL and LDL from normolipidaemic type 2 diabetic and control participants. By contrast, dyslipidaemia was associated with significant changes in both lipid classes (e.g. increased diacylglycerols) and lipid species (e.g. increased C16:1 and C20:3 in phosphatidylcholine and cholesteryl ester and increased C16:0 [palmitic acid] and vaccenic acid in TAG). Levels of palmitic acid in VLDL and LDL TAG correlated with insulin resistance, and VLDL TAG enriched in palmitic acid promoted increased secretion of proinflammatory mediators from human smooth muscle cells. We showed that dyslipidaemia is associated with major changes in both lipid class and lipid species composition in VLDL and LDL from women with type 2 diabetes. In addition, we identified specific molecular lipid species that both correlate with clinical variables and are proinflammatory. Our study thus shows the potential of advanced lipidomic methods to further understand the pathophysiology of type 2 diabetes.

Apple skin patterning is associated with differential expression of MYb10

Background Some apple (Malus × domestica Borkh.) varieties have attractive striping patterns, a quality attribute that is important for determining apple fruit market acceptance. Most apple cultivars (e.g. 'Royal Gala') produce fruit with a defined fruit pigment pattern, but in the case of 'Honeycrisp' apple, trees can produce fruits of two different kinds: striped and blushed. The causes of this phenomenon are unknown. Results Here we show that striped areas of 'Honeycrisp' and 'Royal Gala' are due to sectorial increases in anthocyanin concentration. Transcript levels of the major biosynthetic genes and MYB10, a transcription factor that upregulates apple anthocyanin production, correlated with increased anthocyanin concentration in stripes. However, nucleotide changes in the promoter and coding sequence of MYB10 do not correlate with skin pattern in 'Honeycrisp' and other cultivars differing in peel pigmentation patterns. A survey of methylation levels throughout the coding region of MYB10 and a 2.5 Kb region 5' of the ATG translation start site indicated that an area 900 bp long, starting 1400 bp upstream of the translation start site, is highly methylated. Cytosine methylation was present in all three contexts, with higher methylation levels observed for CHH and CHG (where H is A, C or T) than for CG. Comparisons of methylation levels of the MYB10 promoter in 'Honeycrisp' red and green stripes indicated that they correlate with peel phenotypes, with an enrichment of methylation observed in green stripes. Conclusions Differences in anthocyanin levels between red and green stripes can be explained by differential transcript accumulation of MYB10. Different levels of MYB10 transcript in red versus green stripes are inversely associated with methylation levels in the promoter region. Although observed methylation differences are modest, trends are consistent across years and differences are statistically significant. Methylation may be associated with the presence of a TRIM retrotransposon within the promoter region, but the presence of the TRIM element alone cannot explain the phenotypic variability observed in 'Honeycrisp'. We suggest that methylation in the MYB10 promoter is more variable in 'Honeycrisp' than in 'Royal Gala', leading to more variable color patterns in the peel of this cultivar.

Kiwifruit floral gene APETALA2 is alternatively spliced and accumulates in aberrant indeterminate flowers in the absence of miR172

In Arabidopsis, the identity of perianth and reproductive organs are specified by antagonistic action of two floral homeotic genes, APETALA2 (AP2) and AGAMOUS (AG). AP2 is also negatively regulated by an evolutionary conserved interaction with a microRNA, miR172, and has additional roles in general plant development. A kiwifruit gene with high levels of homology to AP2 and AP2-like genes from other plant species was identified. The transcript was abundant in the kiwifruit flower, particularly petal, suggesting a role in floral organ identity. Splice variants were identified, all containing both AP2 domains, including a variant that potentially produces a shorter transcript without the miRNA172 targeting site. Increased AP2 transcript accumulation was detected in the aberrant flowers of the mutant 'Pukekohe dwarf' with multiple perianth whorls and extended petaloid features. In contrast to normal kiwifruit flowers, the aberrant flowers failed to accumulate miR172 in the developing whorls, although accumulation was detected at the base of the flower. An additional role during dormancy in kiwifruit was proposed based on AP2 transcript accumulation in axillary buds before and after budbreak.

What is a good project manager? An Aristotelian perspective

The purpose of this paper is to take a critical look at the question “what is a competent project manager?” and bring some fresh added-value insights. This leads us to analyze the definitions, and assessment approaches of project manager competence. Three major standards as prescribed by PMI, IPMA, and GAPPS are considered for review from an attribute-based and performance-based approach and from a deontological and consequentialist ethics perspectives. Two fundamental tensions are identified: an ethical tension between the standards and the related competence assessment frameworks and a tension between attribute and performance-based approaches. Aristotelian ethical and practical philosophy is brought in to reconcile these differences. Considering ethics of character that rises beyond the normative deontological and consequentialist perspectives is suggested. Taking the mediating role of praxis and phrónêsis between theory and practice into consideration is advocated to resolve the tension between performance and attribute-based approaches to competence assessment.

Red colouration in apple fruit is due to the activity of the MYB transcription factor, MdMYB10

Anthocyanin concentration is an important determinant of the colour of many fruits. In apple (Malus x domestica), centuries of breeding have produced numerous varieties in which levels of anthocyanin pigment vary widely and change in response to environmental and developmental stimuli. The apple fruit cortex is usually colourless, although germplasm does exist where the cortex is highly pigmented due to the accumulation of either anthocyanins or carotenoids. From studies in a diverse array of plant species, it is apparent that anthocyanin biosynthesis is controlled at the level of transcription. Here we report the transcript levels of the anthocyanin biosynthetic genes in a red-fleshed apple compared with a white-fleshed cultivar. We also describe an apple MYB transcription factor, MdMYB10, that is similar in sequence to known anthocyanin regulators in other species. We further show that this transcription factor can induce anthocyanin accumulation in both heterologous and homologous systems, generating pigmented patches in transient assays in tobacco leaves and highly pigmented apple plants following stable transformation with constitutively expressed MdMYB10. Efficient induction of anthocyanin biosynthesis in transient assays by MdMYB10 was dependent on the co-expression of two distinct bHLH proteins from apple, MdbHLH3 and MdbHLH33. The strong correlation between the expression of MdMYB10 and apple anthocyanin levels during fruit development suggests that this transcription factor is responsible for controlling anthocyanin biosynthesis in apple fruit; in the red-fleshed cultivar and in the skin of other varieties, there is an induction of MdMYB10 expression concurrent with colour formation during development. Characterization of MdMYB10 has implications for the development of new varieties through classical breeding or a biotechnological approach.

The binding of nuclear factors to the as-1 element in the CaMV 35S promoter is affected by cytosine methylation in vitro

Transcriptional gene silencing (TCS) is often associated with an increased level of cytosine methylation in the affected promoters. The effect of methylation of the cauliflower mosaic virus (CaMV) 35S promoter sequence on its binding to factors present in the nuclei was analyzed by electrophoretic mobility shift assays using extracts of petunia flowers. Specific DNA-protein interactions were detected in the region of the CaMV 35S promoter that contains the as-1 element and the region between -345 and -208. The binding of protein factor(s) to the as-1 element was influenced by cytosine methylation, whereas the binding to the region between -345 and -208 was unaffected. The results suggest that cytosine methylation of the as-1 element potentially affects the activity of the CaMV 35S promoter. © Georg Thieme Verlag KG Stuttgart.

Phospholipid composition of the rat lens is independent of diet

Dietary fatty acids are known to influence the phospholipid composition of many tissues in the body, with lipid turnover occurring rapidly. The aim of this study was to investigate whether changes in the fatty acid composition of the diet can affect the phospholipid composition of the lens. Male Sprague-Dawley rats were fed three diets with distinct profiles in both essential and non-essential fatty acids. After 8 weeks, lenses and skeletal muscle were removed, and the lenses sectioned into nuclear and cortical regions. In these experiments, the lens cortex was synthesised during the course of the variable lipid diet. Phospholipids were then identified by electrospray ionisation tandem mass spectrometry, and quantified via the use of internal standards. The phospholipid compositions of the nuclear and cortical regions of the lens differed slightly between the two regions, but comparison of the equivalent regions across the diet groups showed remarkable similarity. In contrast, the phospholipid composition of skeletal muscle (medial gastrocnemius) in these rats varied significantly. This study provides the first direct evidence to show that the phospholipid composition of the lens is tightly regulated and thus appears to be independent of diet. As phospholipids determine membrane fluidity and influence the activity and function of integral membrane proteins, regulation of their composition may be important for the function of the lens. Crown Copyright (C) 2008 Published by Elsevier Ltd. All rights reserved.

A rapid transcriptional activation is induced by the dormancy-breaking chemical hydrogen cyanamide in kiwifruit (Actinidia deliciosa) buds

Budbreak in kiwifruit (Actinidia deliciosa) can be poor in locations that have warm winters with insufficient winter chilling. Kiwifruit vines are often treated with the dormancy-breaking chemical hydrogen cyanamide (HC) to increase and synchronize budbreak. This treatment also offers a tool to understand the processes involved in budbreak. A genomics approach is presented here to increase our understanding of budbreak in kiwifruit. Most genes identified following HC application appear to be associated with responses to stress, but a number of genes appear to be associated with the reactivation of growth. Three patterns of gene expression were identified: Profile 1, an HC-induced transient activation; Profile 2, an HC-induced transient activation followed by a growth-related activation; and Profile 3, HC- and growth-repressed. One group of genes that was rapidly up-regulated in response to HC was the glutathione S-transferase (GST) class of genes, which have been associated with stress and signalling. Previous budbreak studies, in three other species, also report up-regulated GST expression. Phylogenetic analysis of these GSTs showed that they clustered into two sub-clades, suggesting a strong correlation between their expression and budbreak across species.

Trait and behavioural motor control is associated with resistance to overconsumption

While overall obesity rates are rising, a minority of individuals appear to resist overconsumption and remain lean in spite of an ‘obesogenic’ environment. Studying the factors hypothesised to underpin behaviours associated with resistance to overconsumption may inform weight management strategies in an adverse environment. Trait (BIS-11) and behavioural (response inhibition, GoStop) self control were assessed in the laboratory. Snack food consumption was measured covertly via a sham taste test. Lack of motor control was positively correlated (r = .32, p <. 05) and successful response inhibition was negatively correlated (r = −.35, p <. 05) with snack food intake. Low motor control was also associated with further food intake when satiated (r = .39, p < .01). These relationships were independent of self-reported palatability and perceived reward value of the food. Motor control may be an important factor implicated in ‘mindless’ eating in an environment abundant in palatable, energy-dense snack foods.