Development of a Three-Dimensional In Vitro Model for Longitudinal Observation of Cell Behavior: Monitoring by Magnetic Resonance Imaging and Optical Imaging

Purpose: The aim of this study is the development of a three-dimensional multicellular spheroid cell culture model for the longitudinal comparative and large-scale screening of cancer cell proliferation with noninvasive molecular imaging techniques under controlled and quantifiable conditions. Procedures: The human glioblastoma cell line Gli36ΔEGFR was genetically modified to constitutively express the fluorescence protein mCherry, and additionally labeled with iron oxide nanoparticles for high-field MRI detection. The proliferation of aggregates was longitudinally monitored with fluorescence imaging and correlated with aggregate size by light microscopy, while MRI measurements served localization in 3D space. Irradiation with γ-rays was used to detect proliferational response. Results: Cell proliferation in the stationary three-dimensonal model can be observed over days with high accuracy. A linear relationship of fluorescence intensity with cell aggregate size was found, allowing absolute quantitation of cells in a wide range of cell amounts. Glioblastoma cells showed pronounced suppression of proliferation for several days following high-dose γ-irradiation. Conclusions: Through the combination of two-dimensional optical imaging and 3D MRI, the position of individual cell aggregates and their corresponding light emission can be detected. This allows an exact quantification of cell proliferation, with a focus on very small cell amounts (below 100 cells) using high resolution noninvasive techniques as a well-controlled basis for further cell transplantation studies

Ascorbic acid, a vitamin, is observed by in vivo 13C nuclear magnetic resonance spectroscopy of rat liver

The first in vivo detection of a vitamin with nuclear magnetic resonance (NMR) is reported for mammalian liver. Vitamin C, also known as ascorbic acid, was monitored noninvasively in rat liver by "whole body" 13C NMR spectroscopy at high field after infusion of [1,2-13C2]glucose into anesthetized rats. Generally, the carbon resonances of ascorbic acid overlap with those of other highly abundant cellular metabolites, thus precluding their observation in situ. This problem was resolved by taking advantage of the 13C-13C spin couplings introduced by the two covalently bound 13C nuclei in [1,2-13C2]glucose. During glucose metabolism, [5,6-13C2]ascorbic acid was synthesized, which also exhibited characteristic 13C homonuclear spin couplings. This feature enabled the spectral discrimination of ascorbic acid from overlapping singlet resonances of other metabolites. Quantitative analysis of the spin-coupling patterns provided an estimate of the turnover rate of hepatic ascorbic acid in vivo (1.9 +/- 0.4 nmol.min-1.g-1) and a novel approach toward a better understanding of optimal ascorbic acid requirements in humans. The results obtained in vivo were confirmed with high-resolution proton and 13C NMR spectroscopy of liver extracts

Effects of soil tillage and fertilization on resource efficiency and greenhouse gas emissions in a long-term field experiment in Southern Germany.

Two factorial long-term field experiments were carried out at the experimental site of Scheyern, located in southern Germany, 40 km north of Munich (48 degrees 30'0' N, 11 degrees 26'60' E). Here three soil tillage systems were investigated: CT (conventional tillage with moldboard plough, 25 cm plowing depth), RT1 (reduced tillage with chisel plow, 18 cm working depth), and RT2 (reduced tillage with chisel plow, 8 cm working depth). At the same time, three fertilization systems were analyzed (high (N3), medium (N2) and low (N1) mineral N input) with a crop rotation of winter wheat (Triticum aestivum L) - potatoes (Solanum tuberosum L.) - winter wheat-corn (Zea mays L). The long-term effects of tillage and fertilization on yields, soil properties, nitrogen and energy efficiency, as well as greenhouse gas emissions (GGE) were investigated for the period of 1994-2005. On average conventional tillage (CT) produced yields of 8.03 (N1), 8.82 (N2) and 8.88 (N3) GE (grain equivalents) ha(-1) yr(-1); reduced tillage (RT1) yields of 7.82 (N1), 8.54 (N2) and 9.10 (N3) GE ha(-1) yr(-1) and RT2 yields of 6.9 (Ni), 7.82 (N2) and 8.6 (N3) GE ha(-1) yr(-1). The benefit of reduced soil tillage over CT. is a lower consumption of diesel fuel (reduced by 35%) and fossil energy (by 10%), C sequestration and N accumulation in soil. We recorded the highest soil organic carbon (SOC) in the RT2 treatments with the lowest tillage intensity (52.5 Mg ha(-1)) and the lowest SOC reserves in the CT plowed treatments (41.1 Mg ha(-1)). During the reported period, SOC reserves in the plowed treatments decreased by about 300 kg C ha-1 yr-1, whereas they increased by 150-500 kg C ha(-1) yr(-1) in the chiseled treatments. Similar results were achieved with the soil organic nitrogen (SON) reserves based on the type of tillage. This amounted to around 4000 kg ha-1 (CT), 4500 kg ha (RT1) and more than 5000 kg N ha-1 (RT2). The RT1 treatments were marked by high nutrient and energy efficiency. The disadvantage of reduced tillage lies in higher pesticide consumption and stronger soil compaction. The influence of reduced tillage was more pronounced in RT2 than in RT1 (higher SOC and SON content, higher soil dry bulk density, lower consumption of diesel fuel, higher pesticide input). The significant decreases in yield in the RT2 treatments reduced the nitrogen and energy efficiency and raised yield-related greenhouse gas emissions (GGE) in comparison to the RT1 treatments. In the case of reduced tillage combined with high N doses (RT1/N3, RT2/N2, RT2/N3), high N2O emissions of 10 to 12 kg ha(-1) yr(-1) were measured using closed chambers. It was found that as input of mineral N increased, GGE for tillage treatments, both area and yield related also increased. In RT1/N1, negative net GGE were recorded due to high C sequestration combined with moderate N2O and CO2 emissions (-220 kg CO2 (eq) ha(-1) yr(-1), -28 kg CO2 eq GE-(1)), whereas CT/N3 produced the highest net GGE (3587 kg CO2 (eq) ha(-1) yr(-1), 404 kg CO2 eq GE(-1)). (C) 2013 Elsevier B.V. All rights reserved

Oscillatory brain dynamics supporting impaired Stroop task performance in schizophrenia-spectrum disorder.

The Stroop color-word interference task, prompting slower response to color-incongruent than to congruent items, is often used to study neural mechanisms of inhibitory control and dysfunction in schizophrenia-spectrum disorders. Inconsistent findings of an augmented Stroop effect limit identification of relevant dysfunctional mechanism(s) in schizophrenia. The present study sought to advance understanding of normal and impaired neural oscillatory dynamics by distinguishing interference detection and response preparation during the Stroop task in schizophrenia-spectrum disorders via analysis of behavioral performance and 4-7 Hz (theta) and 10-30 Hz (alpha/beta) EEG oscillations in 40 patients (SZ) and 27 healthy comparison participants (HC). SZ responded more slowly and showed less dorsal anterior cingulate (dACC) theta enhancement during INC trials, less enhancement of dACC-sensorimotor cortex connectivity (theta phase synchrony) during INC trials, more alpha/beta suppression though less enhancement of that suppression during INC trials, and slower post-response alpha/beta rebound than did HC. Reaction time distributions showed larger group and Stroop effects during the 25% of trials with the slowest responses. Poorer theta phase coherence in patients indicates impaired communication between regions associated with interference processing (dACC) and response preparation (sensorimotor cortex). Results suggest a failure cascade in which compromised behavioral Stroop effects are driven at least in part by dysfunctional interference processing (less theta power increase) prompting dysfunctional motor response preparation (less alpha/beta power suppression). Inconsistent Stroop effects in past studies of schizophrenia may result from differing task parameters sampling different degrees of Stroop task difficulty