high-resolution-time-lapse-imaging-automated-analysis-microtubule

High-resolution Time-lapse Imaging and Automated Analysis of Microtubule Dynamics in Living Human Umbilical Vein Endothelial Cells

The physiological process by which new vasculature forms from existing vasculature requires specific signaling events that trigger morphological changes ...

University of the Sciences in Philadelphia

We present an experimental technique that allows the simultaneous spatial imaging and spectral analysis of falling droplets that exhibit lasing. Single droplet investigations serve as, among other purposes, a preliminary study for spray and combustion researchers. The described setup provides a valuable tool for the evaluation of microdroplet investigations with laser-spectroscopic techniques that rely on laser-induced fluorescence (LIF) or similar spectroscopical phenomena. The emphasis is that both spatial and spectral information are obtained from single-shot images of a falling droplet. Furthermore, combining spatial imaging and a spatially resolving optical multichannel analyzer makes a pointwise rastering of the droplets spectrum possible. This allows for the (almost) unambiguous determination of sources of influence on the spectrum of these droplets-such as geometrical distortion and lasing, nondissolved tracer lumps, and similar phenomena. Although the focus is on the experimental technique itself, we supplement detailed studies of lasing in falling microdroplets. These results were obtained with the aim of developing a system for measuring temperature distributions in droplets and sprays. In the light of these results the practice of calibrating a droplets spectrum by use of a bulk liquid sample needs to be critically reviewed.

Simultaneous spatial and spectral imaging of lasing droplets.

We report on 11 patients with gastric or colonic bleeding or lesions who were treated with a new endoscopic over-the-scope clip (OTSC) system.

A new endoscopic over-the-scope clip system for treatment of lesions and bleeding in the GI tract: first clinical experiences.

Rupture of the hepatic artery caused by clostridial infection has not been reported before.

Sudden death after pancreatic head resection: rupture of hepatic artery caused by clostridial infection.

Most agronomic plant traits result from complex molecular networks involving multiple genes and from environmental factors. One such trait is the enzymatic discoloration of fruit and tuber tissues initiated by mechanical impact (bruising). Tuber susceptibility to bruising is a complex trait of the cultivated potato (Solanum tuberosum) that is crucial for crop quality. As phenotypic evaluation of bruising is cumbersome, the application of diagnostic molecular markers would empower the selection of low bruising potato varieties. The genetic factors and molecular networks underlying enzymatic tissue discoloration are sparsely known. Hitherto there is no association study dealing with tuber bruising and diagnostic markers for enzymatic discoloration are rare.

Association genetics in Solanum tuberosum provides new insights into potato tuber bruising and enzymatic tissue discoloration.

Gastrointestinal stromal tumors (GISTs) are the most frequent mesenchymal tumors of the gastrointestinal tract usually occurring in the 6th to 7th decade of life, while their occurrence in children is rare (1-2%). Carney triad (CT), a non-hereditary association of gastric GIST with pulmonary chondroma and/or extraadrenal paraganglioma, is an even much rarer disease (to date ~120 cases reported worldwide) usually affecting young adult females. Pediatric GISTs differ from CT-associated GISTs solely by the absence of other components of the triad and are completely different from sporadic GISTs of the adult. Both, pediatric and CT-GISTs, metastasize frequently to regional lymph nodes (29%) and are usually wild type (WT) for common KIT-/PDGFRA mutations.

Multifocal gastric gastrointestinal stromal tumors (GISTs) with lymph node metastases in children and young adults: a comparative clinical and histomorphological study of three cases including a new case of Carney triad.

The hyporheic zone in stream ecosystems is a heterogeneous key habitat for species across many taxa. Consequently, it attracts high attention among freshwater scientists, but generally applicable guidelines on sampling strategies are lacking. Thus, the objective of this study was to develop and validate such sampling guidelines. Applying geostatistical analysis, we quantified the spatio-temporal variability of parameters, which characterize the physico-chemical substratum conditions in the hyporheic zone. We investigated eight stream reaches in six small streams that are typical for the majority of temperate areas. Data was collected on two occasions in six stream reaches (development data), and once in two additional reaches, after one year (validation data). In this study, the term spatial variability refers to patch contrast (patch to patch variance) and patch size (spatial extent of a patch). Patch contrast of hyporheic parameters (specific conductance, pH and dissolved oxygen) increased with macrophyte cover (r(2)=0.95, p<0.001), while patch size of hyporheic parameters decreased from 6 to 2 m with increasing sinuosity of the stream course (r(2)=0.91, p<0.001), irrespective of the time of year. Since the spatial variability of hyporheic parameters varied between stream reaches, our results suggest that sampling design should be adapted to suit specific stream reaches. The distance between sampling sites should be inversely related to the sinuosity, while the number of samples should be related to macrophyte cover.

Drivers and spatio-temporal extent of hyporheic patch variation: implications for sampling.

Genotyping is a prerequisite for tyrosine kinase inhibitor therapy in high risk and malignant GIST. About 10% of GISTs are wild-type for KIT but carry PDGFRA mutations. Applying the traditional approach, mutation analysis of these cases is associated with higher costs if all hotspots regions in KIT (exon 9, 11, 13, 17) are performed at first. Our aim was to evaluate the predictive value of a combined histomorphological-immunohistochemical pattern analysis of PDGFRA-mutated GISTs to efficiently direct KIT and PDGFRA mutation analysis.

Value of epithelioid morphology and PDGFRA immunostaining pattern for prediction of PDGFRA mutated genotype in gastrointestinal stromal tumors (GISTs).

Isotopic variation of food stuffs propagates through trophic systems. But, this variation is dampened in each trophic step, due to buffering effects of metabolic and storage pools. Thus, understanding of isotopic variation in trophic systems requires knowledge of isotopic turnover. In animals, turnover is usually quantified in diet-switch experiments in controlled conditions. Such experiments usually involve changes in diet chemical composition, which may affect turnover. Furthermore, it is uncertain if diet-switch based turnover models are applicable under conditions with randomly fluctuating dietary input signals. Here, we investigate if turnover information derived from diet-switch experiments with dairy cows can predict the isotopic composition of metabolic products (milk, milk components and feces) under natural fluctuations of dietary isotope and chemical composition. First, a diet-switch from a C3-grass/maize diet to a pure C3-grass diet was used to quantify carbon turnover in whole milk, lactose, casein, milk fat and feces. Data were analyzed with a compartmental mixed effects model, which allowed for multiple pools and intra-population variability, and included a delay between feed ingestion and first tracer appearance in outputs. The delay for milk components and whole milk was ~12 h, and that of feces ~20 h. The half-life (t½) for carbon in the feces was 9 h, while lactose, casein and milk fat had a t½ of 10, 18 and 19 h. The (13)C kinetics of whole milk revealed two pools, a fast pool with a t½ of 10 h (likely representing lactose), and a slower pool with a t½ of 21 h (likely including casein and milk fat). The diet-switch based turnover information provided a precise prediction (RMSE ~0.2 ‰) of the natural (13)C fluctuations in outputs during a 30 days-long period when cows ingested a pure C3 grass with naturally fluctuating isotope composition.

Forward modeling of fluctuating dietary 13C signals to validate 13C turnover models of milk and milk components from a diet-switch experiment.

Isotopic turnover quantifies the metabolic renewal process of elements in organs and excreta. Knowledge of the isotopic turnover of animal organs and excreta is necessary for diet reconstruction via stable isotope analysis, as used in animal ecology, palaeontology and food authentication. Effects of dietary protein content on the isotopic carbon and nitrogen turnover (i.e. delay, representing the time between ingestion and start of renewal, and half-life) are unknown for most mammalian organs and excreta.

Dietary protein content affects isotopic carbon and nitrogen turnover.

Endoscopic intracavitary pull-through vacuum treatment of an insufficient pancreaticogastrostomy.

Endothelial cells (ECs) migrate directionally during angiogenesis and wound healing by polarizing to extracellular cues to guide directional movement. EC polarization is controlled by microtubule (MT) growth dynamics, which are regulated by MT-associated proteins (MAPs) that alter MT stability. Mitotic centromere-associated kinesin (MCAK) is a MAP that promotes MT disassembly within the mitotic spindle, yet its function in regulating MT dynamics to promote EC polarity and migration has not been investigated. We used high-resolution fluorescence microscopy coupled with computational image analysis to elucidate the role of MCAK in regulating MT growth dynamics, morphology, and directional migration of ECs. Our results show that MCAK-mediated depolymerization of MTs is specifically targeted to the trailing edge of polarized wound-edge ECs. Regulation of MCAK function is dependent on Aurora A kinase, which is regionally enhanced by signaling from the small guanosine triphosphatase, Rac1. Thus, a Rac1-Aurora A-MCAK signaling pathway mediates EC polarization and directional migration by promoting regional differences in MT dynamics in the leading and trailing cell edges.

Rac1 and Aurora A regulate MCAK to polarize microtubule growth in migrating endothelial cells.

The nitrogen isotope composition (δ¹⁵N) of different amino acids carries different dietary information. We hypothesized that transamination and de novo synthesis create three groups that largely explain their dietary information. Rats were fed with ¹⁵N-labeled amino acids. The redistribution of the dietary ¹⁵N labels among the muscular amino acids was analyzed. Subsequently, the labeling was changed and the nitrogen isotope turnover was analyzed. The amino acids had a common nitrogen half-life of ∼20 d, but differed in δ¹⁵N. Nontransaminating and essential amino acids largely conserved the δ¹⁵N of the source and, hence, trace the origin in heterogeneous diets. Nonessential and nontransaminating amino acids showed a nitrogen isotope composition between their dietary composition and that of their de novo synthesis pool, likely indicating their fraction of de novo synthesis. The bulk of amino acids, which are transaminating, derived their N from a common N pool and hence their δ¹⁵N was similar.

Transamination governs nitrogen isotope heterogeneity of amino acids in rats.

Gastrointestinal stromal tumors (GISTs) have distinct gene expression patterns according to localization, genotype and aggressiveness. DNA methylation at CpG dinucleotides is an important mechanism for regulation of gene expression. We performed targeted DNA methylation analysis of 1.505 CpG loci in 807 cancer-related genes in a cohort of 76 GISTs, combined with genome-wide mRNA expression analysis in 22 GISTs, to identify signatures associated with clinicopathological parameters and prognosis. Principal component analysis revealed distinct DNA methylation patterns associated with anatomical localization, genotype, mitotic counts and clinical follow-up. Methylation of a single CpG dinucleotide in the non-CpG island promoter of SPP1 was significantly correlated with shorter disease-free survival. Hypomethylation of this CpG was an independent prognostic parameter in a multivariate analysis compared to anatomical localization, genotype, tumor size and mitotic counts in a cohort of 141 GISTs with clinical follow-up. The epigenetic regulation of SPP1 was confirmed in vitro, and the functional impact of SPP1 protein on tumorigenesis-related signaling pathways was demonstrated. In summary, SPP1 promoter methylation is a novel and independent prognostic parameter in GISTs, and might be helpful in estimating the aggressiveness of GISTs from the intermediate-risk category.

Combined DNA methylation and gene expression profiling in gastrointestinal stromal tumors reveals hypomethylation of SPP1 as an independent prognostic factor.

HIV-1-infected and/or immune-activated microglia and macrophages are pivotal in the pathogenesis of HIV-1-associated neurocognitive disorders (HAND). Glutaminase, a metabolic enzyme that facilitates glutamate generation, is upregulated and may play a pathogenic role in HAND. Our previous studies have demonstrated that glutaminase is released to the extracellular fluid during HIV-1 infection and neuroinflammation. However, key molecular mechanisms that regulate glutaminase release remain unknown. Recent advances in understanding intercellular trafficking have identified microvesicles (MVs) as a novel means of shedding cellular contents. We posit that during HIV-1 infection and immune activation, microvesicles may mediate glutaminase release, generating excessive and neurotoxic levels of glutamate.

Article	Year
Simultaneous spatial and spectral imaging of lasing droplets. Journal of the Optical Society of America. A, Optics, image science, and vision\| PubMed ID: 16211803	2005
A new endoscopic over-the-scope clip system for treatment of lesions and bleeding in the GI tract: first clinical experiences. Gastrointestinal endoscopy\| PubMed ID: 17591492	2007
Sudden death after pancreatic head resection: rupture of hepatic artery caused by clostridial infection. Surgical infections\| PubMed ID: 18171122	2007
Association genetics in Solanum tuberosum provides new insights into potato tuber bruising and enzymatic tissue discoloration. BMC genomics\| PubMed ID: 21208436	2011
Multifocal gastric gastrointestinal stromal tumors (GISTs) with lymph node metastases in children and young adults: a comparative clinical and histomorphological study of three cases includi... Diagnostic pathology\| PubMed ID: 21663639	2011
Drivers and spatio-temporal extent of hyporheic patch variation: implications for sampling. PloS one\| PubMed ID: 22860053	2012
Value of epithelioid morphology and PDGFRA immunostaining pattern for prediction of PDGFRA mutated genotype in gastrointestinal stromal tumors (GISTs). International journal of clinical and experimental pathology\| PubMed ID: 24040448	2013
Forward modeling of fluctuating dietary 13C signals to validate 13C turnover models of milk and milk components from a diet-switch experiment. PloS one\| PubMed ID: 24392000	2013
Dietary protein content affects isotopic carbon and nitrogen turnover. Rapid communications in mass spectrometry : RCM\| PubMed ID: 24591029	2013
Endoscopic intracavitary pull-through vacuum treatment of an insufficient pancreaticogastrostomy. Endoscopy\| PubMed ID: 24806365	2014
Rac1 and Aurora A regulate MCAK to polarize microtubule growth in migrating endothelial cells. The Journal of cell biology\| PubMed ID: 25002679	2014
Transamination governs nitrogen isotope heterogeneity of amino acids in rats. Journal of agricultural and food chemistry\| PubMed ID: 25036536	2014
Combined DNA methylation and gene expression profiling in gastrointestinal stromal tumors reveals hypomethylation of SPP1 as an independent prognostic factor. International journal of cancer\| PubMed ID: 25046773	2015
Glutaminase-containing microvesicles from HIV-1-infected macrophages and immune-activated microglia induce neurotoxicity. Molecular neurodegeneration\| PubMed ID: 26546362	2015

Alexander Braun

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