Increased endocytosis in retinal vascular endothelial cells grown in high glucose medium is modulated by inhibitors of nonenzymatic glycosylation

We sought to determine if hyperglycaemia is responsible for increased retinal vascular endothelial-cell (RVEC) endocytosis in diabetes and to assess the role of nonenzymatic glycosylation in mediation of this novel endothelial-cell pathology. RVECs were propagated in media containing either 5 or 25 mmol/l glucose for up to 10 days after which they were exposed to the protein tracer horseradish peroxidase for 30 min. The level of RVEC endocytosis was quantified in intact cell monolayers by electron microscopic stereology, and in cell lysates by a simple spectrophotometric method. The effect of the nonenzymatic glycosylation inhibitors, aminoguanidine and D-lysine, on high-glucose medium induced changes in RVEC endocytosis was tested by inclusion of these agents in the culture medium. RVECs exposed to 25 mmol/l glucose showed a stepwise increase in endocytosis of horseradish peroxidase culminating in a two- to threefold increase after 10 days. Endocytosis returned to normal levels after a further 10 days in 5 mmol/l glucose medium. The increase in RVEC endocytosis was markedly reduced, but not completely normalised, by aminoguanidine and D-lysine. Exposure of cultured RVECs to 25 mmol/l glucose causes an increase in endocytosis of similar magnitude to that experienced by RVEC in early diabetes, and implicates hyperglycaemia in the latter situation. A significant component of the increase in RVEC endocytosis appears to be mediated by nonenzymatic glycosylation.

Molecular dissection of Meis1 reveals 2 domains required for leukemia induction and a key role for Hoxa gene activation

The Hoxa9 and Meis1 genes represent important oncogenic collaborators activated in a significant proportion of human leukemias with genetic alterations in the MLL gene. In this study, we show that the transforming property of Meis1 is modulated by 3 conserved domains, namely the Pbx interaction motif (PIM), the homeodomain, and the C-terminal region recently described to possess transactivating properties. Meis1 and Pbx1 interaction domain-swapping mutants are dysfunctional separately, but restore the full oncogenic activity of Meis1 when cotransduced in primary cells engineered to overexpress Hoxa9, thus implying a modular nature for PIM in Meis1-accelerated transformation. Moreover, we show that the transactivating domain of VP16 can restore, and even enhance, the oncogenic potential of the Meis1 mutant lacking the C-terminal 49 amino acids. In contrast to Meis1, the fusion VP16-Meis1 is spontaneously oncogenic, and all leukemias harbor genetic activation of endogenous Hoxa9 and/or Hoxa7, suggesting that Hoxa gene activation represents a key event required for the oncogenic activity of VP16-Meis1.

Temporal characterization and in vitro comparison of cell survival following the delivery of 3D-conformal, intensity-modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT)

A phantom was designed and implemented for the delivery of treatment plans to cells in vitro. Single beam, 3D-conformal radiotherapy (3D-CRT) plans, inverse planned five-field intensity-modulated radiation therapy (IMRT), nine-field IMRT, single-arc volumetric modulated arc therapy (VMAT) and dual-arc VMAT plans were created on a CT scan of the phantom to deliver 3 Gy to the cell layer and verified using a Farmer chamber, 2D ionization chamber array and gafchromic film. Each plan was delivered to a 2D ionization chamber array to assess the temporal characteristics of the plan including delivery time and 'cell's eye view' for the central ionization chamber. The effective fraction time, defined as the percentage of the fraction time where any dose is delivered to each point examined, was also assessed across 120 ionization chambers. Each plan was delivered to human prostate cancer DU-145 cells and normal primary AGO-1522b fibroblast cells. Uniform beams were delivered to each cell line with the delivery time varying from 0.5 to 20.54 min. Effective fraction time was found to increase with a decreasing number of beams or arcs. For a uniform beam delivery, AGO-1552b cells exhibited a statistically significant trend towards increased survival with increased delivery time. This trend was not repeated when the different modulated clinical delivery methods were used. Less sensitive DU-145 cells did not exhibit a significant trend towards increased survival with increased delivery time for either the uniform or clinical deliveries. These results confirm that dose rate effects are most prevalent in more radiosensitive cells. Cell survival data generated from uniform beam deliveries over a range of dose rates and delivery times may not always be accurate in predicting response to more complex delivery techniques, such as IMRT and VMAT.

Assessing software upgrades, plan properties and patient geometry using intensity modulated radiation therapy (IMRT) complexity metrics

Purpose: The aim of this study is to compare the sensitivity of different metrics to detect differences in complexity of intensity modulated radiation therapy (IMRT) plans following upgrades, changes to planning parameters, and patient geometry. Correlations between complexity metrics are also assessed.

Luminescent Terbium Contrast Agent for Bone Microdamage Detection

The synthesis and photophysical evaluation of a new lanthanide luminescence imaging agent is presented. The agent, a terbium-based cyclen complex can, through the use of an iminodiacetate moiety, bind to damaged bone surface via chelation to exposed Ca(II) sites, enabling the imaging of the damage using confocal fluorescence scanning microscopy.

Self-aligning wireless link using modulated backscatter

A self-tracking (aligning) radio communications system based on the principle of modulated backscatter detection is presented here. Using amplitude-modulated backscatter from a co-operating target, a modified IQ demodulation scheme is used to reproduce automatically and in real time both the received and conjugate of the received phase of an incoming signal. The phase conjugated demodulated IF signal can have incoming backscattered data removed and new data are inserted before being up-converted using IQ single side-band generation so that retro-directive re-transmission can be made to occur. In the absence of the backscattered modulation signal, no phase conjugation and hence no retro-directed re-transmission can occur. The system reported is therefore inherently self-authenticating a facility not readily available from previously reported retro-directive self-tracking systems. © 2010 © The Institution of Engineering and Technology.

Wnt-β-catenin-Tcf-4 signalling-modulated invasiveness is dependent on osteopontin expression in breast cancer.

Background:We have previously demonstrated that Tcf-4 regulates osteopontin (OPN) in rat breast epithelial cells, Rama37. In this report, we have examined the importance of this regulation in human breast cancer.Methods:The regulatory roles of Tcf-4 on cell invasion and OPN expression were investigated. The mRNA expression of Tcf-4 and OPN, and survival of breast cancer patients were correlated.Results:Tcf-4 enhanced cell invasion in both MCF10AT and MDA MB 231 breast cancer cells by transcriptionally activating OPN expression. Osteopontin was activated by Wnt signalling in MDA MB 231 cells. Paradoxical results on Tcf-4-regulated OPN expression in MCF10AT (activation) and Rama37 (repression) cells were shown to be a result of differential Wnt signalling competency in MCF10AT and Rama37 cells. High levels of OPN and Tcf-4 mRNA expression were significantly associated with survival in breast cancer patients. Most importantly, Tcf-4-positive patients had a poorer prognosis when OPN was overexpressed, while OPN-negative patients had a better prognosis when Tcf-4 was overexpressed.Conclusion:Our results suggest that Tcf-4 can act as a repressor or activator of breast cancer progression by regulating OPN expression in a Wnt-dependent manner and that Tcf-4 and OPN together may be a novel prognostic indicator for breast cancer progression.

The effect of familiarity on echolocation in the megachiropteran bat Rousettus aegyptiacus

Much is known about the way bats adjust their echolocation behaviour in response to environmental structure or to locate insect prey. By contrast, little is known about how echolocation calls are modulated in response to familiarity of the environment and objects within it. Here we show that the echolocating Megachiropteran bat Rousettus aegyptiacus produces echolocation signals at the same rate whether an obstacle is predictable or unpredictable in location, but that it has a reduced rate of echolocation signal production in a familiar environment with no obstacle present. This suggests that signal production is reduced in a familiar environment absent of 'clutter' but that probing the environment for maximum information is more important for this species than minimizing any cost of probing the environment in a cluttered space.