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Human endothelial and foetal femur-derived stem cell co-cultures modulate osteogenesis and angiogenesis. - Stem cell research & therapy
A dynamic vasculature is a prerequisite for bone formation where the interaction of bone cells and endothelial cells is essential for both the development and the healing process of bone. Enhanced understanding of the specific mediators involved in bone cell and endothelial cell interactions offers new avenues for skeletal regenerative applications. This study has investigated the osteogenic and angiogenic potential of co-cultures of human foetal diaphyseal or epiphyseal cells with human umbilical vein endothelial cells (HUVEC) in the presence and absence of vascular endothelial growth factor (VEGF) supplementation.Early osteogenic activities of the co-cultures (Â±VEGF) were assessed by alkaline phosphatase (ALP) activity. Osteogenic and angiogenic gene expression was measured using quantitative polymerase chain reaction. An ex vivo organotypic embryonic chick (E11) femur culture model was used to determine the osteogenic effects of VEGF as determined using micro-computed tomography (Î¼CT) and Alcian blue/Sirius red histochemistry and immunocytochemistry for expression of CD31.ALP activity and gene expression of ALP and Type-1 collagen was enhanced in foetal skeletal/HUVECs co-cultures. In foetal diaphyseal/HUVECs co-cultures, VEGF reduced the levels of ALP activity and displayed a negligible effect on von Willebrand factor (vWF) and VEGF gene expression. In contrast, VEGF supplementation was observed to significantly increase FLT-1 and KDR gene expression in co-cultures with modulation of expression enhanced, compared to VEGF skeletal monocultures. In the organotypic chick model, addition of VEGF significantly enhanced bone formation, which coincided with elevated levels of CD31-positiveâ€‰ cells in the mid-diaphyseal region of the femurs.These studies demonstrate a differential skeletal response of early foetal skeletal cells, when co-cultured with endothelial cells and the potential of co-culture models for bone repair. The differential effect of VEGF supplementation on markers of angiogenesis and osteogenesis in co-cultures and organ cultures, demonstrate the importance of the intricate temporal coordination of osteogenic and angiogenic processes during bone formation and implications therein for effective approaches to bone regenerative therapies.
GLUT3 and PKM2 regulate OCT4 expression and support the hypoxic culture of human embryonic stem cells. - Scientific reports
Human embryonic stem cells (hESCs) have the capacity to differentiate into all cell types and thus have great potential for regenerative medicine. hESCs cultured at low oxygen tensions are more pluripotent and display an increased glycolytic rate but how this is regulated is unknown. This study therefore aimed to investigate the regulation of glucose metabolism in hESCs and whether this might impact OCT4 expression. In contrast to the glucose transporter GLUT1, GLUT3 was regulated by environmental oxygen and localised to hESC membranes. Silencing GLUT3 caused a reduction in glucose uptake and lactate production as well as OCT4 expression. GLUT3 and OCT4 expression were correlated suggesting that hESC self-renewal is regulated by the rate of glucose uptake. Surprisingly, PKM2, a rate limiting enzyme of glycolysis displayed a nuclear localisation in hESCs and silencing PKM2 did not alter glucose metabolism suggesting a role other than as a glycolytic enzyme. PKM2 expression was increased in hESCs cultured at 5% oxygen compared to 20% oxygen and silencing PKM2 reduced OCT4 expression highlighting a transcriptional role for PKM2 in hESCs. Together, these data demonstrate two separate mechanisms by which genes regulating glucose uptake and metabolism are involved in the hypoxic support of pluripotency in hESCs.
Protein acetylation dynamics in response to carbon overflow in Escherichia coli. - Molecular microbiology
In Escherichia coli, acetylation of proteins at lysines depends largely on a non-enzymatic acetyl phosphate-dependent mechanism. To assess the functional significance of this post-translational modification, we first grew wild-type cells in buffered tryptone broth with glucose and monitored acetylation over time by immunochemistry. Most acetylation occurred in stationary phase and paralleled glucose consumption and acetate excretion, which began upon entry into stationary phase. Transcription of rprA, a stationary phase regulator, exhibited similar behavior. To identify sites and substrates with significant acetylation changes, we used label-free, quantitative proteomics to monitor changes in protein acetylation. During growth, both the number of identified sites and the extent of acetylation increased with considerable variation among lysines from the same protein. As glucose-regulated lysine acetylation was predominant in central metabolic pathways and overlapped with acetyl phosphate-regulated acetylation sites, we deleted the major carbon regulator CRP and observed a dramatic loss of acetylation that could be restored by deleting the enzyme that degrades acetyl phosphate. We propose that acetyl phosphate-dependent acetylation is a response to carbon flux that could regulate central metabolism.Â© 2015 John Wiley & Sons Ltd.
Stress distribution and contact area measurements of a gecko toe using a high-resolution tactile sensor. - Bioinspiration & biomimetics
The adhesive systems of geckos have been widely studied and have been a great source of bioinspiration. Load-sharing (i.e. preventing stress concentrations through equal distribution of loads) is necessary to maximize the performance of an adhesive system, but it is not known to what extent load-sharing occurs in gecko toes. In this paper, we present in vivo measurements of the stress distribution and contact area on the toes of a tokay gecko (Gekko gecko) using a custom tactile sensor with 100 Î¼m spatial resolution. We found that the stress distributions were nonuniform, with large variations in stress between and within lamellae, suggesting that load-sharing in the tokay gecko is uneven. These results may be relevant to the understanding of gecko morphology and the design of improved synthetic adhesive systems.
Human climbing with efficiently scaled gecko-inspired dry adhesives. - Journal of the Royal Society, Interface / the Royal Society
Since the discovery of the mechanism of adhesion in geckos, many synthetic dry adhesives have been developed with desirable gecko-like properties such as reusability, directionality, self-cleaning ability, rough surface adhesion and high adhesive stress. However, fully exploiting these adhesives in practical applications at different length scales requires efficient scaling (i.e. with little loss in adhesion as area grows). Just as natural gecko adhesives have been used as a benchmark for synthetic materials, so can gecko adhesion systems provide a baseline for scaling efficiency. In the tokay gecko (Gekko gecko), a scaling power law has been reported relating the maximum shear stress Ïƒmax to the area A: Ïƒmax âˆ A(-1/4). We present a mechanical concept which improves upon the gecko's non-uniform load-sharing and results in a nearly even load distribution over multiple patches of gecko-inspired adhesive. We created a synthetic adhesion system incorporating this concept which shows efficient scaling across four orders of magnitude of area, yielding an improved scaling power law: Ïƒmax âˆ A(-1/50). Furthermore, we found that the synthetic adhesion system does not fail catastrophically when a simulated failure is induced on a portion of the adhesive. In a practical demonstration, the synthetic adhesion system enabled a 70 kg human to climb vertical glass with 140 cm(2) of adhesive per hand.Â© 2014 The Author(s) Published by the Royal Society. All rights reserved.
Partnerships for development: municipal solid waste management in Kasese, Uganda. - Waste management & research : the journal of the International Solid Wastes and Public Cleansing Association, ISWA
Municipal solid waste management systems of many developing countries are commonly constrained by factors such as limited financial resources and poor governance, making it a difficult proposition to break with complex, entrenched and unsustainable technologies and systems. This article highlights strategic partnerships as a way to affect a distributed agency among several sets of stakeholders to break so-called path dependencies, which occur when such unsustainable pathways arise, stabilize and become self-reinforcing over time. Experiences from a North-South collaborative effort provide some lessons in such partnership building: In Uganda and Denmark, respectively, the World Wildlife Fund and the network organization access2innovation have mobilized stakeholders around improving the municipal solid waste management system in Kasese District. Through a municipal solid waste management system characterization and mapping exercise, some emergent lessons and guiding principles in partnership building point to both pitfalls and opportunities for designing sustainable pathways. First, socio-technical lock-in effects in the municipal solid waste management system can stand in the way of partnerships based on introducing biogas or incineration technologies. However, opportunities in the municipal solid waste management system can exist within other areas, and synergies can be sought with interlinking systems, such as those represented with sanitation.Â© The Author(s) 2014.
HIF-2Î± regulates NANOG expression in human embryonic stem cells following hypoxia and reoxygenation through the interaction with an Oct-Sox cis regulatory element. - PloS one
Low O2 tension is beneficial for human embryonic stem cell (hESC) maintenance but the mechanism of regulation is unknown. HIF-2Î± was found to bind directly to predicted hypoxic response elements (HREs) in the proximal promoter of OCT4, NANOG and SOX2 only in hESCs cultured under hypoxia (5% O2). This binding induced an array of histone modifications associated with gene transcription while a heterochromatic state existed at atmospheric O2. Interestingly, an enhanced euchromatic state was found when hESCs were exposed to hypoxia followed by 72 hours reoxygenation. This was sustained by HIF-2Î± which enhanced stemness by binding to an oct-sox cis-regulatory element in the NANOG promoter. Thus, these data have uncovered a novel role of HIF-2Î± as a direct regulator of key transcription factors controlling self-renewal in hESCs but also in the induction of epigenetic modifications ensuring a euchromatic conformation which enhances the regenerative potential of these cells.
Design principles for efficient, repeated jumpgliding. - Bioinspiration & biomimetics
Combined jumping and gliding locomotion, or 'jumpgliding', can be an efficient way for small robots or animals to travel over cluttered terrain. This paper presents functional requirements and models for a simple jumpglider which formalize the benefits and limitations of using aerodynamic surfaces to augment jumping ability. Analysis of the model gives insight into design choices and control strategies for higher performance and to accommodate special conditions such as a slippery launching surface. The model informs the design of a robotic platform that can perform repeated jumps using a carbon fiber spring and a pivoting wing. Experiments with two different versions of the platform agree with predictions from the model and demonstrate a significantly greater range, and lower cost-of-transport, than a comparable ballistic jumper.
Effect of oxygen tension on the amino acid utilisation of human embryonic stem cells. - Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
Human embryonic stem cells (hESCs) are a potential source of cells for treatment of many degenerative diseases, but in culture have a propensity to spontaneously differentiate, possibly due to suboptimal conditions. Culture at low oxygen tensions improves hESC maintenance and regulates carbohydrate metabolism. Hence, a greater understanding of the nutrient requirements of hESCs will allow production of more appropriate culture media. This study aims to investigate the effect of environmental oxygen tension on the amino acid metabolism of hESCs.The production or depletion of amino acids by hESCs cultured at 5% or 20% oxygen in the presence or absence of FGF2 was measured by reversephase HPLC.Atmospheric oxygen, or removal of FGF2 from hESCs cultured at 5% oxygen, perturbed the uptake or release of individual amino acids and the total amino acid turnover compared to hESCs cultured at 5% oxygen. In particular, serine uptake was reduced at 20% oxygen and by removal of FGF2.Highly pluripotent hESCs, cultured at 5% oxygen, demonstrate a greater amino acid turnover than hESCs cultured at 20% oxygen, or without FGF2. These data suggest that amino acid turnover could be used as a measure of the self-renewal capacity of hESCs.
Biliary and plasma copper and zinc in pregnant Simmental and Angus cattle. - The Onderstepoort journal of veterinary research
Three each of 3-year-old Angus and Simmental heifers, surgically modified to collect bile, were used to measure the effects of pregnancy and breed on bile flow, biliary copper and zinc excretion and plasma copper and zinc concentrations. Bile copper excretion was significantly higher at 7-mo of pregnancy when samples from both breeds were pooled. From then onwards it declined to its lowest, one week post-partum. During pregnancy, plasma copper concentration increased slightly, reaching its highest level at 7-mo of pregnancy and then decreased slightly until full term. In pooled samples from both breeds, the correlation between increase in bile copper excretion and plasma copper concentration from 0 to 7-mo of pregnancy was high (r = 0.85) and significant (p < 0.05). Plasma zinc concentration decreased to the lowest level around 6-mo of pregnancy but increased thereafter until full term. In cows that were dried off one week after parturition, major shifts in bile and plasma copper and zinc parameters occurred at one week following and these coincided with a marked decline of bile flow and bile copper and zinc excretion. By 3-mo post-partum, biliary copper and zinc excretion and plasma copper and zinc concentrations had reached levels observed prior to pregnancy. When the data from all samples were pooled, the bile flow and bile copper excretion were significantly (p < 0.05) higher in Simmental, and plasma copper and zinc concentration higher in the Angus.
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