Small Compartments Research Articles

In a rat model of panic, corticotropin responses to dorsal periaqueductal gray stimulation depend on physical exertion

Panic disorder patients are exquisitely and specifically sensitive to hypercapnia. The demonstration that carbon dioxide provokes panic in fear-unresponsive amygdala-calcified Urbach-Wiethe patients emphasizes that panic is not fear nor does it require the activation of the amygdala. This is consonant with increasing evidence suggesting that panic is mediated caudally at midbrain's dorsal periaqueductal gray matter (DPAG). Another startling feature of the apparently spontaneous clinical panic is the counterintuitive lack of increments in corticotropin, cortisol and prolactin, generally considered 'stress hormones'. Here we show that the stress hormones are not changed during DPAG-evoked panic when escape is prevented by stimulating the rat in a small compartment. Neither did the corticotropin increase when physical exertion was statistically adjusted to the same degree as non-stimulated controls, as measured by lactate plasma levels. Conversely, neuroendocrine responses to foot-shocks were independent from muscular effort. Data are consonant with DPAG mediation of panic attacks.

Alternate routes to the cell surface underpin insulin-regulated membrane trafficking of GLUT4

ABSTRACTInsulin-stimulated delivery of glucose transporters (GLUT4, also known as SLC2A4) from specialized intracellular GLUT4 storage vesicles (GSVs) to the surface of fat and muscle cells is central to whole-body glucose regulation. This translocation and subsequent internalization of GLUT4 back into intracellular stores transits through numerous small membrane-bound compartments (internal GLUT4-containing vesicles; IGVs) including GSVs, but the function of these different compartments is not clear. Cellugyrin (also known as synaptogyrin-2) and sortilin define distinct populations of IGV; sortilin-positive IGVs represent GSVs, but the function of cellugyrin-containing IGVs is unknown. Here, we demonstrate a role for cellugyrin in intracellular sequestration of GLUT4 in HeLa cells and have used a proximity ligation assay to follow changes in pairwise associations between cellugyrin, sortilin, GLUT4 and membrane trafficking machinery following insulin-stimulation of 3T3-L1 adipoctyes. Our data suggest that insulin stimulates traffic from cellugyrin-containing to sortilin-containing membranes, and that cellugyrin-containing IGVs provide an insulin-sensitive reservoir to replenish GSVs following insulin-stimulated exocytosis of GLUT4. Furthermore, our data support the existence of a pathway from cellugyrin-containing membranes to the surface of 3T3-L1 adipocytes that bypasses GSVs under basal conditions, and that insulin diverts traffic away from this into GSVs.

Novel coiled-coil proteins regulate exocyst association with cortical microtubules in xylem cells via the conserved oligomeric golgi-complex 2 protein.

Xylem vessel cells develop secondary cell walls in distinct patterns. Cortical microtubules are rearranged into distinct patterns and regulate secondary cell wall deposition; however, it is unclear how exocytotic membrane trafficking is linked to cortical microtubules. Here, we show that the novel coiled-coil proteins vesicle tethering 1 (VETH1) and VETH2 recruit EXO70A1, an exocyst subunit essential for correct patterning of secondary cell wall deposition, to cortical microtubules via the conserved oligomeric Golgi complex (COG) 2 protein. VETH1 and VETH2 encode an uncharacterized domain of an unknown function designated DUF869, and were preferentially up-regulated in xylem cells. VETH1-green fluorescent protein (GFP) and VETH2-GFP co-localized at novel vesicle-like small compartments, which exhibited microtubule plus-end-directed and end-tracking dynamics. VETH1 and VETH2 interacted with COG2, and this interaction promoted the association between cortical microtubules and EXO70A1 These results suggest that the VETH-COG2 complex ensures the correct secondary cell wall deposition pattern by recruiting exocyst components to cortical microtubules.

A dynamic in vitro model to evaluate the intestinal release behaviour of modified-release corticosteroid products

Corticosteroids form an important component of drug-based therapeutic strategies in both Crohn's disease and ulcerative colitis. Here, we investigated the intestinal release behaviour of two commonly-used corticosteroids – prednisolone and budesonide – from various commercial modified-release formulations. We evaluated the release characteristics using a novel in vitro model that simulates the dynamic intestinal environment by employing bicarbonate buffers representative of the conditions and contents of the gastrointestinal lumen. The performance of the model was validated with an in-house enteric dual layer prednisolone formulation (internal standard) where in vitro release behaviour closely correlated with in vivo targeting performance in humans. The commercial prednisolone product coated with the pH responsive polymer polyvinyl acetate phthalate released its payload promptly on transfer to the upper small intestinal compartment of the test. In contrast, the budesonide products, Entocort® and Budenofalk®, although showing differences in product performance and delayed release characteristics, both demonstrated some suggestion of extended release behaviour. The proposed dissolution method provided a realistic and discriminative in vitro assessment of the release of corticosteroids from different oral formulations. These results demonstrate that this model can be used to aid the rational design of products developed to target different sites of the gut.

Two-Track Epigenetic Remodeling and Backtracking to Embryonic Stem Cell Bivalency in B-Cell Acute Lymphoblastic Leukemias

Purpose: We sought to define the drivers and functional consequences of DNA methylation changes in childhood pre-B cell leukemia. A comprehensive analysis of such DNA methylation changes will reveal aspects of the etiology and pathogenesis of leukemias, and suggest potential therapeutic modalities.Background: The epigenome is extensively altered in carcinogenesis, but the proximate causes, functional consequences, and overall patterning of DNA methylation changes in the pediatric leukemias is not well defined.Methods: We bisulfite sequenced at single base pair resolution two common pre-B cell leukemia case DNAs and a pre-B cell control (CD19+/CD34+ pre-B cells from normal marrow), and analyzed DNA methylation by high definition microarray in another 227 subjects. RNA expression was available for 82 samples, including the sequenced pre-B cells and leukemias.Results: Epigenetic alteration of B-ALLs occurred in two tracks: de novo methylation of small functional compartments and demethylation of large inter-compartmental backbones. The de novo methylation occurred preferentially at polycomb targets and binding sites for the transcriptional co-repressor CtBP2. DNA methylation of ETV6-AML1 leukemia was overall equivalent to the control, with high hyperdiploid methylation levels being 4.4% lower. Hierarchical clustering revealed four groups, with ETV6-AML1 and hyperdiploid leukemias enriched within two separate groups. DNA methylation deviations were exaggerated in lamina-associated domains, with differences corresponding to these methylation clusters and/or cytogenetic groups. DNA methylation changes were correlated with gene expression changes, and a key polycomb expression signature was reversed when DNA methylation marks were pharmacologically removed in leukemia cell lines.Discussion: While leukemia cells retain the majority of their developmentally determined DNA methylation patterns, key modifications with leukemogenesis revealed both broad patterns and functional targets with some dependence on known characteristics of pre-B cell leukemias. Our data suggested a pivotal role of polycomb and CTBP2 in de novo methylation, which may be traced back to bivalency status of embryonic stem cells. Driven by these potent epigenetic modulations, suppression of polycomb target genes was observed along with disruption of developmental fate and cell cycle and mismatch repair pathways and altered activities of key upstream regulators. An appreciation and validation of the observed changes will lead to new insights into etiology, pathogenesis, and treatments of childhood ALL. DisclosuresNo relevant conflicts of interest to declare.

Chronic thromboembolic pulmonary hypertension

Chronic thromboembolic pulmonary hypertension (CTEPH) is a rare and potentially fatal disease which occasionally occurs as a complication of acute pulmonary embolism. The mechanisms leading to failure of thrombus resolution are not completely understood with only some risk factors identified. Vascular alterations are present in the large and small vessel compartment. Signs and symptoms of CTEPH are nonspecific, rendering diagnosis challenging. A VQ scan followed by computed tomography, magnetic resonance or conventional pulmonary angiography is mandatory to confirm diagnosis and assess operability by a multidisciplinary team. Pulmonary endarterectomy remains the treatment of choice in operable patients and results in significantly improved haemodynamics and functional capacity. In inoperable patients medical treatment is well defined, and mainly includes the recently introduced pharmacological substances with impact on haemodynamics and functional performance.

Non-invasive imaging techniques to study O2 micro-patterns around pesticide treated lupine roots

The soil–root interface is a highly heterogeneous system, e.g. in terms of O2 and pH distribution. The destructive character of conventional methods disturbs the natural conditions of those biogeochemical gradients. Therefore, experiments aiming to control these influences and study pesticide kinetics under given O2 and pH conditions suffer from a large uncertainty of the “real” O2/pH at a certain position. Our approach with two different imaging techniques will examine the soil–root interface as well as the dissipation of the applied pesticide at a high spatial resolution.The obtained outcomes show directly that the pH has an influence on enantioselective dissipation of the acetanilide fungicide metalaxyl. In areas with high pH from an applied racemic mixture, the R-enantiomer dissipates faster than the S-enantiomer. Moreover, we found significantly reduced oxygen values in the bulk soil and vicinity of metalaxyl treated roots compared to control plant roots. The combination of matrix-assisted laser desorption/ionization mass spectrometry (MALDI) and fluorescence imaging indicated the oxygen-dependent behavior of metalaxyl at the root surface.The results presented here underline the great potential of combining different imaging methods to examine the soil–root interfaces as well as the dissipation of organic pollutants in small soil compartments.

Mechanisms of salt tolerance in habanero pepper plants (Capsicum chinense Jacq.): Proline accumulation, ions dynamics and sodium root-shoot partition and compartmentation.

Despite its economic relevance, little is known about salt tolerance mechanisms in pepper plants. To address this question, we compared differences in responses to NaCl in two Capsicum chinense varieties: Rex (tolerant) and Chichen-Itza (sensitive). Under salt stress (150 mM NaCl over 7 days) roots of Rex variety accumulated 50 times more compatible solutes such as proline compared to Chichen-Itza. Mineral analysis indicated that Na+ is restricted to roots by preventing its transport to leaves. Fluorescence analysis suggested an efficient Na+ compartmentalization in vacuole-like structures and in small intracellular compartments in roots of Rex variety. At the same time, Na+ in Chichen-Itza plants was compartmentalized in the apoplast, suggesting substantial Na+ extrusion. Rex variety was found to retain more K+ in its roots under salt stress according to a mineral analysis and microelectrode ion flux estimation (MIFE). Vanadate-sensitive H+ efflux was higher in Chichen-Itza variety plants, suggesting a higher activity of the plasma membrane H+-ATPase, which fuels the extrusion of Na+, and, possibly, also the re-uptake of K+. Our results suggest a combination of stress tolerance mechanisms, in order to alleviate the salt-induced injury. Furthermore, Na+ extrusion to apoplast does not appear to be an efficient strategy for salt tolerance in pepper plants.

Emotions on the loose: emotional contagion and the role of oxytocin in pigs.

We studied emotional contagion, a simple form of empathy, and the role of oxytocin herein in pigs. Two training pigs per pen (n = 16 pens) were subjected to a positive treatment (pairwise access to a large compartment filled with peat, straw and some chocolate raisins) and a negative treatment (social isolation in a small compartment) in a test room using a within-subjects design. Thereafter, two naive pen mates joined the training pigs in the test room, but were not given access to the treatments. This allowed testing for emotional contagion. Subsequently, the naive pigs, serving as their own controls, were given 24 IU of oxytocin or a placebo intranasally 30 min before accompanying the training pigs, which were exposed to either the negative or positive treatment, to the test room. Behavioral differences found between the positive and negative treatments (e.g., play and “tail wagging” vs. standing alert, urinating, defecating and ears backward) show that the treatments induced a positive and negative emotional state in the training pigs, respectively. Changes in behaviors of the training pigs with and without naive pigs present (e.g., in ears backwards) and of the naive pigs with and without training pigs present (e.g., in standing alert) indicated that emotional contagion occurred, especially during the negative treatment. Oxytocin did not seem to affect the behavior of the treated naive pigs, but did affect behaviors (e.g., defecating) of the training pigs which had not received oxytocin. This suggests a role for oxytocin in pig communication, which merits further research.Electronic supplementary materialThe online version of this article (doi:10.1007/s10071-014-0820-6) contains supplementary material, which is available to authorized users.

다중이용시설 구획 공간에서의 화재 및 연기 확산에 대한 수치해석적 연구

노래방과 같은 다중이용시설에서는 좁은 공간에 많은 사람들이 이용을 한다. 다중이용시설에서의 화재는 화염 및 연기의 확산으로 많은 인명과 재산의 피해가 야기되고 있다. 따라서 수치 해석은 다중이용시설의 시설 설계 특성과 다중이용시설에 설치된 가연물간의 연소 특성을 예측하여서 화재 위험에 노출된 요소를 제거하는데 도움을 줄 수 있다. 이러한 예측의 신뢰도를 검증하기 위해서 실제 화재 실험과 동일한 조건으로 수치해석이 필요하며, 본 연구에서는 컴퓨터 화재 시뮬레이션 프로그램인 FDS를 사용하여 수치해석을 수행하였다. 수행된 해석 결과의 검증을 위해서는 실물실험을 통한 결과와 비교 분석 하였다. 특히, 작은 구획 공간들 사이에서의 연기 유동 해석은 재실자들의 위험을 사전에 점검하고 대처할 수 있게 할 것이다. In multi purpose use facilities, there are a large number of people occupied in a small compartment space. The fire in the multi purpose use facility can be occurred human loss and property damage due to spread of a fire and a smoke in buildings. It is most critical to control flame spread in the early stage. To make the model work for practical fire protection engineering problems, property of combustible materials were predicted so that it could be help to reduced the risk. Computational analysis result from numerical models were compared with a real scale fire test. For computational analysis, the Fire Dynamics Simulator(FDS) was conducted with a Large Eddy Simulation(LED) model for turbulence. Especially, The computational fire modeling can help to mitigate risk and to prevent a fire.

Bistable forespore engulfment in Bacillus subtilis by a zipper mechanism in absence of the cell wall.

To survive starvation, the bacterium Bacillus subtilis forms durable spores. The initial step of sporulation is asymmetric cell division, leading to a large mother-cell and a small forespore compartment. After division is completed and the dividing septum is thinned, the mother cell engulfs the forespore in a slow process based on cell-wall degradation and synthesis. However, recently a new cell-wall independent mechanism was shown to significantly contribute, which can even lead to fast engulfment in 60 of the cases when the cell wall is completely removed. In this backup mechanism, strong ligand-receptor binding between mother-cell protein SpoIIIAH and forespore-protein SpoIIQ leads to zipper-like engulfment, but quantitative understanding is missing. In our work, we combined fluorescence image analysis and stochastic Langevin simulations of the fluctuating membrane to investigate the origin of fast bistable engulfment in absence of the cell wall. Our cell morphologies compare favorably with experimental time-lapse microscopy, with engulfment sensitive to the number of SpoIIQ-SpoIIIAH bonds in a threshold-like manner. By systematic exploration of model parameters, we predict regions of osmotic pressure and membrane-surface tension that produce successful engulfment. Indeed, decreasing the medium osmolarity in experiments prevents engulfment in line with our predictions. Forespore engulfment may thus not only be an ideal model system to study decision-making in single cells, but its biophysical principles are likely applicable to engulfment in other cell types, e.g. during phagocytosis in eukaryotes.

Multi-photon Intracellular Sodium Imaging Combined with UV-mediated Focal Uncaging of Glutamate in CA1 Pyramidal Neurons

Multi-photon fluorescence microscopy has enabled the analysis of morphological and physiological parameters of brain cells in the intact tissue with high spatial and temporal resolution. Combined with electrophysiology, it is widely used to study activity-related calcium signals in small subcellular compartments such as dendrites and dendritic spines. In addition to calcium transients, synaptic activity also induces postsynaptic sodium signals, the properties of which are only marginally understood. Here, we describe a method for combined whole-cell patch-clamp and multi-photon sodium imaging in cellular micro domains of central neurons. Furthermore, we introduce a modified procedure for ultra-violet (UV)-light-induced uncaging of glutamate, which allows reliable and focal activation of glutamate receptors in the tissue. To this end, whole-cell recordings were performed on Cornu Ammonis subdivision 1 (CA1) pyramidal neurons in acute tissue slices of the mouse hippocampus. Neurons were filled with the sodium-sensitive fluorescent dye SBFI through the patch-pipette, and multi-photon excitation of SBFI enabled the visualization of dendrites and adjacent spines. To establish UV-induced focal uncaging, several parameters including light intensity, volume affected by the UV uncaging beam, positioning of the beam as well as concentration of the caged compound were tested and optimized. Our results show that local perfusion with caged glutamate (MNI-Glutamate) and its focal UV-uncaging result in inward currents and sodium transients in dendrites and spines. Time course and amplitude of both inward currents and sodium signals correlate with the duration of the uncaging pulse. Furthermore, our results show that intracellular sodium signals are blocked in the presence of blockers for ionotropic glutamate receptors, demonstrating that they are mediated by sodium influx though this pathway. In summary, our method provides a reliable tool for the investigation of intracellular sodium signals induced by focal receptor activation in intact brain tissue.

Multi-photon intracellular sodium imaging combined with UV-mediated focal uncaging of glutamate in CA1 pyramidal neurons.

Multi-photon fluorescence microscopy has enabled the analysis of morphological and physiological parameters of brain cells in the intact tissue with high spatial and temporal resolution. Combined with electrophysiology, it is widely used to study activity-related calcium signals in small subcellular compartments such as dendrites and dendritic spines. In addition to calcium transients, synaptic activity also induces postsynaptic sodium signals, the properties of which are only marginally understood. Here, we describe a method for combined whole-cell patch-clamp and multi-photon sodium imaging in cellular micro domains of central neurons. Furthermore, we introduce a modified procedure for ultra-violet (UV)-light-induced uncaging of glutamate, which allows reliable and focal activation of glutamate receptors in the tissue. To this end, whole-cell recordings were performed on Cornu Ammonis subdivision 1 (CA1) pyramidal neurons in acute tissue slices of the mouse hippocampus. Neurons were filled with the sodium-sensitive fluorescent dye SBFI through the patch-pipette, and multi-photon excitation of SBFI enabled the visualization of dendrites and adjacent spines. To establish UV-induced focal uncaging, several parameters including light intensity, volume affected by the UV uncaging beam, positioning of the beam as well as concentration of the caged compound were tested and optimized. Our results show that local perfusion with caged glutamate (MNI-Glutamate) and its focal UV-uncaging result in inward currents and sodium transients in dendrites and spines. Time course and amplitude of both inward currents and sodium signals correlate with the duration of the uncaging pulse. Furthermore, our results show that intracellular sodium signals are blocked in the presence of blockers for ionotropic glutamate receptors, demonstrating that they are mediated by sodium influx though this pathway. In summary, our method provides a reliable tool for the investigation of intracellular sodium signals induced by focal receptor activation in intact brain tissue.

Detecting calcium in cardiac muscle: fluorescence to dye for.

THE IMPORTANCE OF Ca 2 as a second messenger in general and as a mediator between electrical excitation and cellular contraction in the heart is well accepted. Accordingly, numerous Ca 2 -sensitive fluorescent dyes are available nowadays, rendering the choice of the appropriate dye for a particular application a challenge. It appears to be important to provide calibrated Ca 2 signals rather than fluorescence transients, especially when comparing data between studies. In case calibrated signals cannot be provided, then the time course of fluorescence transients should be interpreted with caution and such signals should be referred to as fluorescence transients rather than Ca 2 transients. Additionally, we discuss how major properties of the fluorescence dyes should be considered with a special emphasis on their Ca 2 dissociation properties. Furthermore, we highlight putative risks of tripping, especially in the context of multicellular recordings, and how to circumvent them. As an ubiquitous intracellular messenger, Ca 2 ions play an important role in almost all living cells (4). A particularly dynamic role has been assigned to Ca 2 in muscle cells because here they not only control gene expression and other slower signaling processes but serve as a direct transducer of the electrical excitation of the plasma membrane into the activation of the physiological response of the cell, the contraction, by activation of the contractile machinery (5). Almost 50 years ago, Jobsis and O’Connor (10) reported the measurements of intracellular Ca 2 transients in skeletal muscle using the Ca 2 -sensitive dye murexide, which changes its absorbance upon binding of Ca 2 ions. Later, these absorbance dyes were improved and an important member of this family, arsenazo III, was widely used to study intracellular Ca 2 in

Arabinogalactan glycosyltransferases target to a unique subcellular compartment that may function in unconventional secretion in plants.

We report that fluorescently tagged arabinogalactan glycosyltransferases target not only the Golgi apparatus but also uncharacterized smaller compartments when transiently expressed in Nicotiana benthamiana. Approximately 80% of AtGALT31A [Arabidopsis thaliana galactosyltransferase from family 31 (At1g32930)] was found in the small compartments, of which, 45 and 40% of AtGALT29A [Arabidopsis thaliana galactosyltransferase from family 29 (At1g08280)] and AtGlcAT14A [Arabidopsis thaliana glucuronosyltransferase from family 14 (At5g39990)] colocalized with AtGALT31A, respectively; in contrast, N-glycosylation enzymes rarely colocalized (3–18%), implicating a role of the small compartments in a part of arabinogalactan (O-glycan) biosynthesis rather than N-glycan processing. The dual localization of AtGALT31A was also observed for fluorescently tagged AtGALT31A stably expressed in an Arabidopsis atgalt31a mutant background. Further, site-directed mutagenesis of a phosphorylation site of AtGALT29A (Y144) increased the frequency of the protein being targeted to the AtGALT31A-localized small compartments, suggesting a role of Y144 in subcellular targeting. The AtGALT31A localized to the small compartments were colocalized with neither SYP61 (syntaxin of plants 61), a marker for trans-Golgi network (TGN), nor FM4-64-stained endosomes. However, 41% colocalized with EXO70E2 (Arabidopsis thaliana exocyst protein Exo70 homolog 2), a marker for exocyst-positive organelles, and least affected by Brefeldin A and Wortmannin. Taken together, AtGALT31A localized to small compartments that are distinct from the Golgi apparatus, the SYP61-localized TGN, FM4-64-stained endosomes and Wortmannin-vacuolated prevacuolar compartments, but may be part of an unconventional protein secretory pathway represented by EXO70E2 in plants.

Hydrogen concentration variation and examination of PAR installation in reactor containment building during hydrogen release from different direction failure places

Concentration on a hydrogen explosion mitigation strategy increased after the accident in Fukushima. A passive autocatalytic recombiner (PAR) is considered as the most realistic mitigation technology during a severe accident where electric power is lost. Hydrogen distribution and concentration might be accurately predicted because PAR must be installed in a place having a proper amount of hydrogen in order to produce positive performance. In this paper, hydrogen distributions and concentrations were predicted and analyzed during hydrogen release from different direction failure places, and then PAR installation was examined. The result shows that the hydrogen concentrations are randomly changed in the containment overall, and the behavior of hydrogen in the case of upper part failure is completely different from the case of a side part failure. It is expected that PAR works well in the area among small compartments located at the lower part of the containment building; however, the performance of PAR installed on another area is inferior.

High density and ligand affinity confer ultrasensitive signal detection by a guanylyl cyclase chemoreceptor.

Guanylyl cyclases (GCs), which synthesize the messenger cyclic guanosine 3',5'-monophosphate, control several sensory functions, such as phototransduction, chemosensation, and thermosensation, in many species from worms to mammals. The GC chemoreceptor in sea urchin sperm can decode chemoattractant concentrations with single-molecule sensitivity. The molecular and cellular underpinnings of such ultrasensitivity are not known for any eukaryotic chemoreceptor. In this paper, we show that an exquisitely high density of 3 × 10(5) GC chemoreceptors and subnanomolar ligand affinity provide a high ligand-capture efficacy and render sperm perfect absorbers. The GC activity is terminated within 150 ms by dephosphorylation steps of the receptor, which provides a means for precise control of the GC lifetime and which reduces "molecule noise." Compared with other ultrasensitive sensory systems, the 10-fold signal amplification by the GC receptor is surprisingly low. The hallmarks of this signaling mechanism provide a blueprint for chemical sensing in small compartments, such as olfactory cilia, insect antennae, or even synaptic boutons.

Slow and fast lung compartments in cystic fibrosis measured by nitrogen multiple-breath washout.

Imaging studies describe significant ventilation defects across a wide range of cystic fibrosis (CF) related lung disease severity. These are unfortunately poorly reflected by phase III slope analysis-derived Scond and Sacin from multiple-breath washout (MBW). Methodology extending previous two-lung compartment model-based analysis is presented describing size and function of fast- and slow-ventilating lung compartments from nitrogen (N2) MBW and correlation to obstructive lung disease severity. In 37 CF subjects (forced expiratory volume in 1 s [FEV1] mean [SD] 84.8 [19.9] % predicted; abnormal lung clearance index [LCI] in 36/37, range 7.28-18.9) and 74 matched healthy controls, volume and specific ventilation of both fast and slowly ventilated lung compartments were derived from N2-based MBW with commercial equipment. In healthy controls lung emptying was characterized by a large compartment constituting 75.6 (8.4)% of functional residual capacity (FRC) with a specific ventilation (regional alveolar tidal volume/regional lung volume) of 13.9 (3.7)% and a small compartment with high specific ventilation (48.4 [15.7]%). In CF the slowly ventilated lung compartment constituted 51.9(9.1)% of FRC, with low specific ventilation of 5.3 (2.4)%. Specific ventilation of the slowly ventilated lung compartment showed stronger correlation with LCI (r2 = 0.70, P < 0.001) vs. Sacin (r2 = 0.44, P < 0.001) or Scond (no significant correlation). Overventilation of the fast lung compartment was no longer seen in severe CF lung disease. Magnitude and function of under- and overventilated lung volumes can be derived from routine N2 MBW in CF. Reported values agree with previous modelling-derived estimates of impaired ventilation and offer improved correlation to disease severity, compared with SnIII analysis.

Automated quantification and integrative analysis of 2D and 3D mitochondrial shape and network properties.

Mitochondrial morphology and function are coupled in healthy cells, during pathological conditions and (adaptation to) endogenous and exogenous stress. In this sense mitochondrial shape can range from small globular compartments to complex filamentous networks, even within the same cell. Understanding how mitochondrial morphological changes (i.e. “mitochondrial dynamics”) are linked to cellular (patho) physiology is currently the subject of intense study and requires detailed quantitative information. During the last decade, various computational approaches have been developed for automated 2-dimensional (2D) analysis of mitochondrial morphology and number in microscopy images. Although these strategies are well suited for analysis of adhering cells with a flat morphology they are not applicable for thicker cells, which require a three-dimensional (3D) image acquisition and analysis procedure. Here we developed and validated an automated image analysis algorithm allowing simultaneous 3D quantification of mitochondrial morphology and network properties in human endothelial cells (HUVECs). Cells expressing a mitochondria-targeted green fluorescence protein (mitoGFP) were visualized by 3D confocal microscopy and mitochondrial morphology was quantified using both the established 2D method and the new 3D strategy. We demonstrate that both analyses can be used to characterize and discriminate between various mitochondrial morphologies and network properties. However, the results from 2D and 3D analysis were not equivalent when filamentous mitochondria in normal HUVECs were compared with circular/spherical mitochondria in metabolically stressed HUVECs treated with rotenone (ROT). 2D quantification suggested that metabolic stress induced mitochondrial fragmentation and loss of biomass. In contrast, 3D analysis revealed that the mitochondrial network structure was dissolved without affecting the amount and size of the organelles. Thus, our results demonstrate that 3D imaging and quantification are crucial for proper understanding of mitochondrial shape and topology in non-flat cells. In summary, we here present an integrative method for unbiased 3D quantification of mitochondrial shape and network properties in mammalian cells.

EOR Potential from CO2 Captured from Coal-Fired Power Plants in the Upper Cretaceous (Cenomanian) Woodbine Group, East Texas Basin, and Southeastern Texas Gulf Coast, USA

The East Texas Basin and southeastern Texas Gulf Coast contain a variety of co-located CO2 sources and sinks that may facilitate development of new clean-coal facilities. These facilities can be linked to mature oil fields with potential for enhanced oil recovery (EOR) from miscible CO2 floods. Twenty-three reservoirs in the East Texas Basin and southeastern Texas Gulf Coast, assuming a 15% recovery factor of original oil in place (OOIP), have a CO2-EOR potential for recovery of ~9,697,000 m3 [~62.2 million stock tank barrels] of oil. A network of new CO2 pipelines can link these fields to existing power plants near lignite mine mouths in east and southeast Texas. Representative oil fields in the Woodbine Group illustrate fluvial and deltaic facies variability and different sandstone-body architectures with varying controls on potential CO2 capacity. Reservoir heterogeneity, fluid flow, hydrocarbon production, and potential CO2 capacity in the Woodbine Group in the structurally simple East Texas field are mainly a function of facies architecture. The Woodbine Group in East Texas field contains a lower section of narrow and lenticular sandstone bodies in a fluvially dominated deltaic system. Pressure and production data indicate that additional oil recovery in the lower Woodbine deltaic section in East Texas field is low in muddy, low-permeability prodelta, and distal-delta-front facies. In contrast, oil recovery is greater in sandy distributary-channel sandstones. Core-plug data show that distributary-channel and channel-mouth-bar sandstones in the field have high median-permeability values, and therefore a greater potential for CO2 capacity. These deltaic deposits are overlain disconformably by an upper section of continuous, coarse-grained fluvial deposits that favor high CO2 capacity owing to Darcy-class permeability and extensive vertical and lateral reservoir continuity. In contrast, Woodbine fields in southeast Texas represent distal-deltaic facies. CO2 capacity and EOR potential in these fields are limited by thin distributary-channel and crevasse-splay deposits that contain abundant mudstone beds that may serve to segregate-injected CO2 into small reservoir compartments.