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1.
Williams GS Chikando AC Tuan HT Sobie EA Lederer WJ Jafri MS 《Biophysical journal》2011,(6):2370-1296
We present what we believe to be a new mathematical model of Ca2+ leak from the sarcoplasmic reticulum (SR) in the heart. To our knowledge, it is the first to incorporate a realistic number of Ca2+-release units, each containing a cluster of stochastically gating Ca2+ channels (RyRs), whose biophysical properties (e.g., Ca2+ sensitivity and allosteric interactions) are informed by the latest molecular investigations. This realistic model allows for the detailed characterization of RyR Ca2+-release properties, and shows how this balances reuptake by the SR Ca2+ pump. Simulations reveal that SR Ca2+ leak consists of brief but frequent single RyR openings (∼3000 cell−1 s−1) that are likely to be experimentally undetectable, and are, therefore, “invisible”. We also observe that these single RyR openings can recruit additional RyRs to open, due to elevated local (Ca2+), and occasionally lead to the generation of Ca2+ sparks (∼130 cell−1 s−1). Furthermore, this physiological formulation of “invisible” leak allows for the removal of the ad hoc, non-RyR mediated Ca2+ leak terms present in prior models. Finally, our model shows how Ca2+ sparks can be robustly triggered and terminated under both normal and pathological conditions. Together, these discoveries profoundly influence how we interpret and understand diverse experimental and clinical results from both normal and diseased hearts. 相似文献
2.
Background
A reliable extraction technique for resolving multiple spots in light or electron microscopic images is essential in investigations of the spatial distribution and dynamics of specific proteins inside cells and tissues. Currently, automatic spot extraction and characterization in complex microscopic images poses many challenges to conventional image processing methods. 相似文献3.
María Hernndez Mesa Jonas van den Brink William E. Louch Kimberly J. McCabe Padmini Rangamani 《PLoS computational biology》2022,18(6)
Super-resolution imaging techniques have provided a better understanding of the relationship between the nanoscale organization and function of ryanodine receptors (RyRs) in cardiomyocytes. Recent data have indicated that this relationship is disrupted in heart failure (HF), as RyRs are dispersed into smaller and more numerous clusters. However, RyRs are also hyperphosphorylated in this condition, and this is reported to occur preferentially within the cluster centre. Thus, the combined impact of RyR relocalization and sensitization on Ca2+ spark generation in failing cardiomyocytes is likely complex and these observations suggest that both the nanoscale organization of RyRs and the pattern of phosphorylated RyRs within clusters could be critical determinants of Ca2+ spark dynamics. To test this hypothesis, we used computational modeling to quantify the relationships between RyR cluster geometry, phosphorylation patterns, and sarcoplasmic reticulum (SR) Ca2+ release. We found that RyR cluster disruption results in a decrease in spark fidelity and longer sparks with a lower amplitude. Phosphorylation of some RyRs within the cluster can play a compensatory role, recovering healthy spark dynamics. Interestingly, our model predicts that such compensation is critically dependent on the phosphorylation pattern, as phosphorylation localized within the cluster center resulted in longer Ca2+ sparks and higher spark fidelity compared to a uniformly distributed phosphorylation pattern. Our results strongly suggest that both the phosphorylation pattern and nanoscale RyR reorganization are critical determinants of Ca2+ dynamics in HF. 相似文献
4.
PurposeThe high incidence of defective ultrasound transducers in clinical practice has been shown in several studies. Recently, a novel method using only stored images for automatic detection of defective transducers was presented. The method makes it possible to remotely monitor many transducers at the same time and send a notification when a defective transducer is found. The purpose of the present study was to evaluate the novel method and assess how well it performs when compared to an established method as reference.MethodsTo evaluate the novel method, in-air images were collected from 81 transducers in radiologic departments in nine hospitals. Two observers assessed the in-air images and marked the defects. Receiver operating characteristic (ROC)- and alternative free response receiver operating characteristic (AFROC)-curves and their figures of merit (FOM) were calculated for the novel method, using marked defects in the in-air images as reference truth.ResultsThe area under the ROC curve was 0.88 (SD 0.06), and the AFROC FOM was 0.71 (SE 0.07).ConclusionThe result shows that the novel method has a good agreement with the in-air method for detecting defects in ultrasound systems. This indicates that the novel method could be a complement to the normal quality control for early, and automatic detection of defects. 相似文献
5.
Coordination of hormone-induced calcium signals in isolated rat hepatocyte couplets: demonstration with confocal microscopy. 总被引:3,自引:0,他引:3
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Excitable cells often display rapid coordination of hormone-induced intracellular calcium signals. Calcium elevations that begin in a single epithelial cell also may spread to adjacent cells, but coordination of hormone-induced signals among epithelial cells has not been described. We report the use of confocal microscopy to determine the inter- and intracellular distribution of cytosolic calcium in isolated rat hepatocyte couplets, an isolated epithelial cell system in which functional polarity is maintained. Both vasopressin and phenylephrine evoked sequential coordinated calcium signals in the couplets, even during cytosolic calcium oscillations. The coupling was abolished by closure of intercellular gap junction channels by treatment with octanol. These observations demonstrate that hormone-induced intracellular calcium signals are coordinated among hepatocytes and suggest that gap junction channels mediate this intercellular integration of tissue responsiveness. 相似文献
6.
Background
The distribution of chromatin-associated proteins plays a key role in directing nuclear function. Previously, we developed an image-based method to quantify the nuclear distributions of proteins and showed that these distributions depended on the phenotype of human mammary epithelial cells. Here we describe a method that creates a hierarchical tree of the given cell phenotypes and calculates the statistical significance between them, based on the clustering analysis of nuclear protein distributions.Results
Nuclear distributions of nuclear mitotic apparatus protein were previously obtained for non-neoplastic S1 and malignant T4-2 human mammary epithelial cells cultured for up to 12 days. Cell phenotype was defined as S1 or T4-2 and the number of days in cultured. A probabilistic ensemble approach was used to define a set of consensus clusters from the results of multiple traditional cluster analysis techniques applied to the nuclear distribution data. Cluster histograms were constructed to show how cells in any one phenotype were distributed across the consensus clusters. Grouping various phenotypes allowed us to build phenotype trees and calculate the statistical difference between each group. The results showed that non-neoplastic S1 cells could be distinguished from malignant T4-2 cells with 94.19% accuracy; that proliferating S1 cells could be distinguished from differentiated S1 cells with 92.86% accuracy; and showed no significant difference between the various phenotypes of T4-2 cells corresponding to increasing tumor sizes.Conclusion
This work presents a cluster analysis method that can identify significant cell phenotypes, based on the nuclear distribution of specific proteins, with high accuracy.7.
Antoine Blanc Nicolas Tran-Khanh Dominic Filion Michael D Buschmann 《The journal of histochemistry and cytochemistry》2005,53(9):1171-1175
Tissue engineering of articular cartilage requires accurate imaging of the chondrocyte cytoskeleton. Past studies have applied various fixation and permeabilization protocols without optimization of parameters. In this study, we have examined procedures using glutaraldehyde and paraformaldehyde as fixatives and Triton X-100 and Octyl-POE as permeabilizing detergents. A four-color fluorescence confocal method was developed to simultaneously image actin, tubulin, vimentin, and the nucleus. We found optimal preservation and morphology of the chondrocyte cytoskeleton after simultaneous fixation and permeabilization with glutaraldehyde and Triton X-100. These images displayed less cellular shrinkage and higher-resolution filamentous structures than with paraformaldehyde or when permeabilization followed fixation. 相似文献
8.
Monteiro-Leal LH Troster H Campanati L Spring H F Trendelenburg M 《Journal of structural biology》2003,141(3):228-239
This work presents a computerized method to identify, detect, evaluate, and, by colored overlay, present gold particle pairs in electron microscopy (EM), even in wide-field views. Double gold immunolabeled specimens were analyzed in a LEO 912 electron microscope equipped with a 2k x 2k-pixel slow-scan cooled CCD camera connected to a computer with analySIS 3.1 PRO image processing software. The acquisition of a high-resolution and high-dynamic-range image by the camera allowed correct segmentation of the gold particles, separating them from other cell structures and from the substrate. Particle identification was performed by a classification module designed by us. Based on shape and size, the computer recognized the group of small particles and classified them as either singular or clustered and differentiated these from the single bigger type. The final image shows the particle types separated and colored, and indicates the total number of objects encountered in the specific region of interest. Moreover, a montage tool allowed us to obtain final representative images of large microscopic fields, which on analysis by the Gold Finder module provided information on the distribution and localization of antigens comparable to that provided by the wide-field light microscope images. 相似文献
9.
Termination of cardiac Ca(2+) sparks: an investigative mathematical model of calcium-induced calcium release
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A Ca(2+) spark arises when a cluster of sarcoplasmic reticulum (SR) channels (ryanodine receptors or RyRs) opens to release calcium in a locally regenerative manner. Normally triggered by Ca(2+) influx across the sarcolemmal or transverse tubule membrane neighboring the cluster, the Ca(2+) spark has been shown to be the elementary Ca(2+) signaling event of excitation-contraction coupling in heart muscle. However, the question of how the Ca(2+) spark terminates remains a central, unresolved issue. Here we present a new model, "sticky cluster," of SR Ca(2+) release that simulates Ca(2+) spark behavior and enables robust Ca(2+) spark termination. Two newly documented features of RyR behavior have been incorporated in this otherwise simple model: "coupled gating" and an opening rate that depends on SR lumenal [Ca(2+)]. Using a Monte Carlo method, local Ca(2+)-induced Ca(2+) release from clusters containing between 10 and 100 RyRs is modeled. After release is triggered, Ca(2+) flux from RyRs diffuses into the cytosol and binds to intracellular buffers and the fluorescent Ca(2+) indicator fluo-3 to produce the model Ca(2+) spark. Ca(2+) sparks generated by the sticky cluster model resemble those observed experimentally, and Ca(2+) spark duration and amplitude are largely insensitive to the number of RyRs in a cluster. As expected from heart cell investigation, the spontaneous Ca(2+) spark rate in the model increases with elevated cytosolic or SR lumenal [Ca(2+)]. Furthermore, reduction of RyR coupling leads to prolonged model Ca(2+) sparks just as treatment with FK506 lengthens Ca(2+) sparks in heart cells. This new model of Ca(2+) spark behavior provides a "proof of principle" test of a new hypothesis for Ca(2+) spark termination and reproduces critical features of Ca(2+) sparks observed experimentally. 相似文献
10.
Franklin Sedarat Eric Lin Edwin D W Moore Glen F Tibbits 《Journal of applied physiology》2004,97(3):1098-1103
Colocalization of dihydropyridine (DHPR) and ryanodine (RyR) receptors, a key determinant of Ca(2+)-induced Ca2+ release, was previously estimated in 3-, 6-, 10-, and 20-day-old rabbit ventricular myocytes by immunocytochemistry and confocal microscopy. We now report on the effects of deconvolution (using a maximum-likelihood estimation algorithm) on the calculation of colocalization indexes. Clusters of DHPR and RyR can be accurately represented as point sources of fluorescence, which enables a model of their relative distributions to be constructed using images of point spread functions to simulate their fluorescence inside a cell. This model was used to investigate the effects of deconvolution on colocalization as a function of separation distance. Deconvolution resulted in significant improvements in both axial and transverse resolutions, producing significant increases in clarity. Comparisons of intensity profiles (full-width half-maximum) pre- and postdeconvolution showed decreased dispersion of the fluorescent signal and a corresponding decrease in false colocalization as determined by fluorescence modeling. This hypothesis was extended to physiological data previously collected. The number of colocalized voxels was quantified after deconvolution, and the degree of colocalization of DHPR with RyR decreased significantly after deconvolution in all age groups: 3 days (62 +/- 2% before deconvolution, 43 +/- 3 after deconvolution) to 20 days old (79 +/- 1% before deconvolution, 63 +/- 2% after deconvolution). The data demonstrate that confocal images should be deconvolved before any quantitative analysis, such as colocalization index determination, to minimize the detrimental effects of out-of-focus light in coincident voxels. 相似文献
11.
Balasubramanian L Ahmed A Lo CM Sham JS Yip KP 《American journal of physiology. Regulatory, integrative and comparative physiology》2007,293(4):R1586-R1594
Integrins are transmembrane heterodimeric proteins that link extracellular matrix (ECM) to cytoskeleton and have been shown to function as mechanotransducers in nonmuscle cells. Synthetic integrin-binding peptide triggers Ca(2+) mobilization and contraction in vascular smooth muscle cells (VSMCs) of rat afferent arteriole, indicating that interactions between the ECM and integrins modulate vascular tone. To examine whether integrins transduce extracellular mechanical stress into intracellular Ca(2+) signaling events in VSMCs, unidirectional mechanical force was applied to freshly isolated renal VSMCs through paramagnetic beads coated with fibronectin (natural ligand of alpha(5)beta(1)-integrin in VSMCs). Pulling of fibronectin-coated beads with an electromagnet triggered Ca(2+) sparks, followed by global Ca(2+) mobilization. Paramagnetic beads coated with low-density lipoprotein, whose receptors are not linked to cytoskeleton, were minimally effective in triggering Ca(2+) sparks and global Ca(2+) mobilization. Preincubation with ryanodine, cytochalasin-D, or colchicine substantially reduced the occurrence of Ca(2+) sparks triggered by fibronectin-coated beads. Binding of VSMCs with antibodies specific to the extracellular domains of alpha(5-) and beta(1)-integrins triggered Ca(2+) sparks simulating the effects of fibronectin-coated beads. Preincubation of microperfused afferent arterioles with ryanodine or integrin-specific binding peptide inhibited pressure-induced myogenic constriction. In conclusion, integrins transduce mechanical force into intracellular Ca(2+) signaling events in renal VSMCs. Integrin-mediated mechanotransduction is probably involved in myogenic response of afferent arterioles. 相似文献
12.
S. Kawamata 《The Histochemical journal》1992,24(5):275-282
Summary The localization of pyro-antimonate-precipitable Ca2+ in the undecalcified femur and calvaria of neonatal rats was examined. The fixation of bones with pyro-antimonate-glutaraldehyde followed by pyro-antimonate-osmium, (two-step method) resulted in better preservation of tissue and more precise localization of precipitates than did the direct immersion of specimens in pyro-antimonate-osmium solution (one-step method). The precipitate was frequently observed within the endoplasmic reticulum of obsteoblasts. Most vacuoles in osteoclasts contained precipitate. By contrast, the mitochondria in these cells were associated with small amounts of precipitate. There was no evidence of precipitate in the Golgi apparatus. The presence of calcium in the precipitate was verified by EGTA treatment and X-ray microanalysis. This study demonstrated that (1) the two-step pyro-antimonate method is a useful and reliable procedure for visualizing Ca2+, and (2) cellular Ca2– can be successfully localized in undecalcified bone by this method. 相似文献
13.
N. Nagahara Takaaki Ito Hitoshi Kitamura Takeshi Nishino 《Histochemistry and cell biology》1998,110(3):243-250
In our previous study, we found that mercaptopyruvate sulfurtransferase (MST) was evolutionarily related to mitochondrial rhodanese. To elucidate the difference between MST and rhodanese, the tissue, cellular, and subcellular distribution of rat MST was determined biochemically and immunohistochemically by using anti-MST antibody raised in rabbit. In an immunohistochemical study, tetramethyl rhodamine isothiocyanate-conjugated phalloidin against F-actin and fluorescein isothiocyanate-conjugated goat anti-rabbit immunoglobulin as a secondary antibody to the anti-MST antibody were used for double fluorescent staining. They were detected by confocal laser fluorescence microscopy. In the immunoelectron microscopic study of hepatocyte and renal tubular epithelium, a postembedding immunogold method was used. Biochemical studies including western blot analyses of various tissues and subcellular fractions of the liver were also performed. MST was widely distributed in rat tissues but the cellular distribution was found to be different in each tissue. MST was predominantly localized in proximal tubular epithelium in the kidney, pericentral hepatocytes in the liver, cardiac cells in the heart, and neuroglial cells in the brain. This immunocytochemical study also found that MST was localized in both mitochondria and cytoplasm. 相似文献
14.
Fluorescent confocal laser scanning microscopy allows an improved imaging of microscopic objects in three dimensions. However, the resolution along the axial direction is three times worse than the resolution in lateral directions. A method to overcome this axial limitation is tilting the object under the microscope, in a way that the direction of the optical axis points into different directions relative to the sample. A new technique for a simultaneous reconstruction from a number of such axial tomographic confocal data sets was developed and used for high resolution reconstruction of 3D-data both from experimental and virtual microscopic data sets. The reconstructed images have a highly improved 3D resolution, which is comparable to the lateral resolution of a single deconvolved data set. Axial tomographic imaging in combination with simultaneous data reconstruction also opens the possibility for a more precise quantification of 3D data. The color images of this publication can be accessed from http://www.esacp.org/acp/2000/20-1/heintzmann.++ +htm. At this web address an interactive 3D viewer is additionally provided for browsing the 3D data. This java applet displays three orthogonal slices of the data set which are dynamically updated by user mouse clicks or keystrokes. 相似文献
15.
G J Augustine E M Adler M P Charlton M Hans D Swandulla K Zipser 《Journal of Physiology》1992,86(1-3):129-134
Synthetic calcium buffers, including fluorescent calcium indicators, were microinjected into squid 'giant' presynaptic nerve terminals to investigate the calcium signal that triggers neurotransmitter secretion. Digital imaging methods, applied in conjunction with the fluorescent calcium indicator dye fura-2, reveal that transient rises in presynaptic calcium concentration are associated with action potentials. Transmitter release terminates within 1-2 ms after a train of action potentials, even though presynaptic calcium concentration remains at micromolar levels for many seconds longer. Microinjection of the calcium buffer, EGTA, into the presynaptic terminal has no effect on transmitter release evoked by single presynaptic action potentials. EGTA injection does, however, block the change in calcium concentration measured by fura-2. Therefore, the calcium signal measured by fura-2 is not responsible for triggering release. These results suggest that the rise in presynaptic calcium concentration that triggers release must be highly localized to escape detection with fura-2 imaging. Unlike EGTA, microinjection of BAPTA--a calcium buffer with an equilibrium affinity for calcium similar to that of EGTA--produces a potent, dose-dependent, and reversible block of action-potential evoked transmitter release. The superior ability of BAPTA to block transmitter release apparently is due to the more rapid calcium-binding kinetics of BAPTA compared to EGTA. Because EGTA should bind calcium within a few tens of microseconds under the conditions of our experiments, the inability of EGTA to block release indicates that transmitter release is triggered within a few tens of microseconds after the entry of calcium into the presynaptic terminal.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
16.
The detection of airport attracts lots of attention and becomes a hot topic recently because of its applications and importance in military and civil aviation fields. However, the complicated background around airports brings much difficulty into the detection. This paper presents a new method for airport detection in remote sensing images. Distinct from other methods which analyze images pixel by pixel, we introduce visual attention mechanism into detection of airport and improve the efficiency of detection greatly. Firstly, Hough transform is used to judge whether an airport exists in an image. Then an improved graph-based visual saliency model is applied to compute the saliency map and extract regions of interest (ROIs). The airport target is finally detected according to the scale-invariant feature transform features which are extracted from each ROI and classified by hierarchical discriminant regression tree. Experimental results show that the proposed method is faster and more accurate than existing methods, and has lower false alarm rate and better anti-noise performance simultaneously. 相似文献
17.
Examining calcium spark morphology and its relationship to the structure of the cardiac myocyte offers a direct means of understanding excitation-contraction coupling mechanisms. Traditional confocal line scanning achieves excellent temporal spark resolution but at the cost of spatial information in the perpendicular dimension. To address this, we developed a methodology to identify and analyze sparks obtained via two-dimensional confocal or charge-coupled device microscopy. The technique consists of nonlinearly subtracting the background fluorescence, thresholding the data on the basis of noise level, and then localizing the spark peaks via a generalized extrema test, while taking care to detect and separate adjacent peaks. In this article, we describe the algorithm, compare its performance to a previously validated spark detection algorithm, and demonstrate it by applying it to both a synthetic replica and an experimental preparation of a two-dimensional isotropic myocyte monolayer exhibiting sparks during a calcium transient. We find that our multidimensional algorithm provides better sensitivity than the conventional method under conditions of temporally heterogeneous background fluorescence, and the inclusion of peak segmentation reduces false negative rates when spark density is high. Our algorithm is robust and can be effectively used with different imaging modalities and allows spark identification and quantification in subcellular, cellular, and tissue preparations. 相似文献
18.
A shape-based method for automatic and rapid segmentation of roots in soil from X-ray computed tomography images: Rootine 总被引:1,自引:0,他引:1
Gao Wei Schlüter Steffen Blaser Sebastian R. G. A. Shen Jianbo Vetterlein Doris 《Plant and Soil》2019,441(1-2):643-655
Plant and Soil - X-ray computed tomography (CT) is widely recognized as a powerful tool for in-situ quantification of root system architecture (RSA) in soil. However, employing X-ray CT to identify... 相似文献
19.
《Animal : an international journal of animal bioscience》2022,16(10):100646
Mastitis is one of the most common diseases in dairy cows and has a negative impact on their welfare and life, causing significant economic losses to the dairy industry. Many attempts have been made to develop a detection method for mastitis using thermal infrared thermography. However, the use of this detection technique to determine the health of the cow's udder is susceptible to external factors, resulting in inaccurate detection of dairy cow mastitis. Therefore, this study explored a new and comprehensive detection method of dairy cow mastitis based on infrared thermal images. This method combined the left and right udder skin surface temperature (USST) difference detection method with the ocular surface temperature and USST difference detection method with improvements. The effect of external factors on dairy cow USST was effectively reduced. In addition, after comparing different target localisation algorithms, this paper used the You Only Look Once v5 (YOLOv5) deep learning network model to obtain the temperature information of eyes and udders, and mastitis detection of dairy cows was performed. A total of 105 dairy cows passing through a passage were randomly selected from the thermal infrared video and detected by the new and comprehensive detection method, and the results of cow mastitis detection were compared with somatic cell count. The results showed that the accuracy, specificity, and sensitivity of mastitis detection were 87.62, 84.62, and 96.30%, respectively. Using the YOLOv5 deep learning network model to locate the key parts of the cow had a good effect, with an average accuracy of 96.1%, and an average frame rate of 116.3f/s. The detection accuracy of dairy cow mastitis by deep learning technology combined with the detection method in this paper reached 85.71%. The results showed that the new and comprehensive detection method based on infrared thermal images can be used for the detection of dairy cow mastitis with high detection accuracy. This method can reduce the influence of external factors and can be integrated into the automatic identification system of dairy mastitis based on YOLOv5 to realise on-site monitoring of dairy mastitis. 相似文献
20.
Alexandra Zahradníková Ivan Valent Ivan Zahradník 《The Journal of general physiology》2010,136(1):101-116
Cytosolic calcium concentration in resting cardiac myocytes locally fluctuates as a result of spontaneous microscopic Ca2+ releases or abruptly rises as a result of an external trigger. These processes, observed as calcium sparks, are fundamental for proper function of cardiac muscle. In this study, we analyze how the characteristics of spontaneous and triggered calcium sparks are related to cardiac ryanodine receptor (RYR) gating. We show that the frequency of spontaneous sparks and the probability distribution of calcium release flux quanta of triggered sparks correspond quantitatively to predictions of an allosteric homotetrameric model of RYR gating. This model includes competitive binding of Ca2+ and Mg2+ ions to the RYR activation sites and allosteric interaction between divalent ion binding and channel opening. It turns out that at rest, RYRs are almost fully occupied by Mg2+. Therefore, spontaneous sparks are most frequently evoked by random openings of the highly populated but rarely opening Mg4RYR and CaMg3RYR forms, whereas triggered sparks are most frequently evoked by random openings of the less populated but much more readily opening Ca2Mg2RYR and Ca3MgRYR forms. In both the spontaneous and the triggered sparks, only a small fraction of RYRs in the calcium release unit manages to open during the spark because of the limited rate of Mg2+ unbinding. This mechanism clarifies the unexpectedly low calcium release flux during elementary release events and unifies the theory of calcium signaling in resting and contracting cardiac myocytes. 相似文献