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Level set analysis for leukocyte detection and tracking.

We propose a cell detection and tracking solution using image-level sets computed via threshold decomposition. In contrast to existing methods where manual initialization is required to track individual cells, the proposed approach can automatically identify and track multiple cells by exploiting the shape and intensity characteristics of the cells. The capture of the cell boundary is considered as an evolution of a closed curve that maximizes image gradient along the curve enclosing a homogeneous region. An energy functional dependent upon the gradient magnitude along the cell boundary, the region homogeneity within the cell boundary and the spatial overlap of the detected cells is minimized using a variational approach. For tracking between frames, this energy functional is modified considering the spatial and shape consistency of a cell as it moves in the video sequence. The integrated energy functional complements shape-based segmentation with a spatial consistency based tracking technique. We demonstrate that an acceptable, expedient solution of the energy functional is possible through a search of the image-level lines: boundaries of connected components within the level sets obtained by threshold decomposition. The level set analysis can also capture multiple cells in a single frame rather than iteratively computing a single active contour for each individual cell. Results of cell detection using the energy functional approach and the level set approach are presented along with the associated processing time. Results of successful tracking of rolling leukocytes from a number of digital video sequences are reported and compared with the results from a correlation tracking scheme.

Algorithms↗

Processing by factor analysis of dynamic dual isotope studies using 99Tcm and 201Tl within a middle energy band. Evaluation in thyroid nodule malignancy.

Simultaneous investigations with two isotopes are currently restricted because of spectral overlap. The factor analysis of spectral and dynamic structures (FASDS) method is shown to achieve accurate spectral separation. In addition, it estimates underlying dynamic mechanisms. Twenty-six patients were injected simultaneously with 99Tcm-pertechnetate and 201Tl-chloride to assess the malignancy of solitary thyroid nodules. List-mode acquisition of spectral, temporal and spatial coordinates of events allows the reconstruction of an image sequence indexed by time and energy. FASDS proceeds in two steps. First it yields both dynamic and spatial information related to each isotope (99Tcm and 201Tl) and partially removes the scatter component. Then it estimates the underlying kinetics and associated spatial distributions of each isotope. Using the 201Tl component, an index was derived from the uptake ratio between nodules and normal thyroid tissue. Concerning the detection of malignant nodules the method indicated no false negative in our limited group of 26 patients. One false positive result was found which could not be classified by the investigation of the 201Tl dynamic components contained in the reconstructed 201Tl factor sequence.

Factor Analysis, Statistical↗

[The spatial-temporal organization of the membrane-synaptic modifications and of the topograms of the slow fluctuations in the cortical potentials during learning].

This paper deals with an actual problem of specificity of organization of different stages of a conditioned reflex (CR). We suggest to consider this problem at the level of membrane-synaptic modifications comparing them with the indices of changes in the spatial organization of slow potentials in the cortex. The definite sequence of involvement of cellular mechanisms has been shown which can determine different features of the spatial organization of cortical slow potentials. Changes in cellular excitability of cortical neurons are probably connected with the variations of the general level of momentary electroencephalotopograms and, correspondingly, with the level of general spatial synchronization. Involvement of excitatory and inhibitory synaptic connections (with the efficiency corresponding to CR stages) determines the formation of electroencephalotopogram relief. Indices of both levels depend of hypothalamic effects on the cortex.

Animals↗

Independently-addressable micron-sized biosensor elements.

With the continuing development of micro-total analysis systems and sensitive biosensing technologies, it is often desirable to immobilize biomolecules onto a surface in a small well-defined area. A novel method was developed to electrochemically attach DNA probes to micron-sized regions of a gold surface using biotin-LC-hydrazide (BH). Previously, we have found that the radical produced during the oxidation of BH will attach to a wide variety of electroactive surfaces. An array of micron-sized gold band electrodes (75 microm wide) was fabricated onto glass microscope slides and BH was deposited onto each electrode through the application of an oxidizing potential. Subsequent attachment of avidin to the biotinylated surface created the 'molecular sandwich' architecture necessary for further immobilization of biotinylated biomolecules to the surface. In this work, we utilized biotinylated DNA probes of varying sequence to illustrate the specificity of the attachment scheme. The immobilization of avidin, DNA probe, and hybridization of DNA target is visualized with fluorescence tags and the spatially selective attachment and hybridization of unique DNA sequences is demonstrated.

Avidin↗

Secondary-task effects on sequence learning.

With a repeated sequence of stimuli, performance in a serial reaction-time task improves more than with a random sequence. The difference has been taken as a measure of implicit sequence learning. Implicit sequence learning is impaired when a secondary task is added to the serial RT task. In the first experiment, secondary-task effects on different types of sequences were studied to test the hypothesis that the learning of unique sequences (where each sequence element has a unique relation to the following one) is not impaired by the secondary task, while the learning of ambiguous sequences is. The sequences were random up to a certain order of sequential dependencies, where they became deterministic. Contrary to the hypothesis, secondary-task effects on the learning of unique sequences were as strong or stronger than such effects on the learning of ambiguous sequences. In the second experiment a hybrid sequence (with unique as well as ambiguous transitions) was used with different secondary tasks. A visuo-spatial and a verbal memory task did not interfere with the learning of the sequence, but interference was observed with an auditory go/no-go task in which high- and low-pitched tones were presented after each manual response and a foot pedal had to be pressed in response to high-pitched tones. Thus, interference seems to be specific to certain secondary tasks and may be related to memory processes (but most likely not to visuo-spatial and verbal memory) or to the organizations of sequences, consistent with previous suggestions.

Adolescent↗

Sequence-dependent bending of DNA and phasing of nucleosomes.

Conformational analysis has revealed anisotropic flexibility of the B-DNA double helix: it bends most easily into the grooves, being the most rigid when bent in a perpendicular direction. This result implies that DNA in a nucleosome is curved by means of relatively sharp bends ("mini-kinks") which are directed into the major and minor grooves alternatively and separated by 5-6 base pairs. The "mini-kink" model proved to be in keeping with the x-ray structure of the B-DNA dodecamer resolved later, which exhibits two "annealed kinks", also directed into the grooves. The anisotropy of B DNA is sequence-dependent: the pyrimidine-purine dimers (YR) favor bending into the minor groove, and the purine-pyrimidine dinucleotides (RY), into the minor one. The RR and YY dimers appear to be the most rigid dinucleotides. Thus, a DNA fragment consisting of the interchanging oligopurine and oligopyrimidine blocks 5-6 base pairs long should manifest a spectacular curvature in solution. Similarly, a nucleotide sequence containing the RY and YR dimers separated by a half-pitch of the double helix is the most suitable for wrapping around the nucleosomal core. Analysis of the numerous examples demonstrating the specific alignment of nucleosomes on DNA confirms this concept. So, the sequence-dependent "mechanical" properties of the double helix influence the spatial arrangement of DNA in chromatin.

Animals↗

PASS2: an automated database of protein alignments organised as structural superfamilies.

BACKGROUND: The functional selection and three-dimensional structural constraints of proteins in nature often relates to the retention of significant sequence similarity between proteins of similar fold and function despite poor sequence identity. Organization of structure-based sequence alignments for distantly related proteins, provides a map of the conserved and critical regions of the protein universe that is useful for the analysis of folding principles, for the evolutionary unification of protein families and for maximizing the information return from experimental structure determination. The Protein Alignment organised as Structural Superfamily (PASS2) database represents continuously updated, structural alignments for evolutionary related, sequentially distant proteins. DESCRIPTION: An automated and updated version of PASS2 is, in direct correspondence with SCOP 1.63, consisting of sequences having identity below 40% among themselves. Protein domains have been grouped into 628 multi-member superfamilies and 566 single member superfamilies. Structure-based sequence alignments for the superfamilies have been obtained using COMPARER, while initial equivalencies have been derived from a preliminary superposition using LSQMAN or STAMP 4.0. The final sequence alignments have been annotated for structural features using JOY4.0. The database is supplemented with sequence relatives belonging to different genomes, conserved spatially interacting and structural motifs, probabilistic hidden markov models of superfamilies based on the alignments and useful links to other databases. Probabilistic models and sensitive position specific profiles obtained from reliable superfamily alignments aid annotation of remote homologues and are useful tools in structural and functional genomics. PASS2 presents the phylogeny of its members both based on sequence and structural dissimilarities. Clustering of members allows us to understand diversification of the family members. The search engine has been improved for simpler browsing of the database. CONCLUSIONS: The database resolves alignments among the structural domains consisting of evolutionarily diverged set of sequences. Availability of reliable sequence alignments of distantly related proteins despite poor sequence identity and single-member superfamilies permit better sampling of structures in libraries for fold recognition of new sequences and for the understanding of protein structure-function relationships of individual superfamilies. PASS2 is accessible at http://www.ncbs.res.in/~faculty/mini/campass/pass2.html

Amino Acid Sequence↗

Temporal and spatial expression patterns of the small heat shock (hsp16) genes in transgenic Caenorhabditis elegans.

The expression of the hsp16 gene family in Caenorhabditis elegans has been examined by introducing hsp16-lacZ fusions into the nematode by transformation. Transcription of the hsp16-lacZ transgenes was totally heat-shock dependent and resulted in the rapid synthesis of detectable levels of beta-galactosidase. Although the two hsp16 gene pairs of C. elegans are highly similar within both their coding and noncoding sequences, quantitative and qualitative differences in the spatial pattern of expression between gene pairs were observed. The hsp16-48 promoter was shown to direct greater expression of beta-galactosidase in muscle and hypodermis, whereas the hsp16-41 promoter was more efficient in intestine and pharyngeal tissue. Transgenes that eliminated one promoter from a gene pair were expressed at reduced levels, particularly in postembryonic stages, suggesting that the heat shock elements in the intergenic region of an hsp16 gene pair may act cooperatively to achieve high levels of expression of both genes. Although the hsp16 gene pairs are never constitutively expressed, their heat inducibility is developmentally restricted; they are not heat inducible during gametogenesis or early embryogenesis. The hsp16 genes represent the first fully inducible system in C. elegans to be characterized in detail at the molecular level, and the promoters of these genes should find wide applicability in studies of tissue- and developmentally regulated genes in this experimental organism.

Animals↗

Ten families of variant genes encoded in subtelomeric regions of multiple chromosomes of Plasmodium chabaudi, a malaria species that undergoes antigenic variation in the laboratory mouse.

The chromosome ends of human malaria parasites harbour many genes encoding proteins that are exported to the surface of infected red cells, often being involved in host-parasite interactions and immune evasion. Unlike other murine malaria parasites Plasmodium chabaudi undergoes antigenic variation during passage in the laboratory mouse and hence is a model suitable for investigation of switching mechanisms. However, little is known about the subtelomeric regions of P. chabaudi chromosomes and its variable antigens. Here we report 80 kb of sequence from an end of one P. chabaudi chromosome. Hybridization of probes spanning this region to two dimensional pulsed field gels of the genome revealed 10 multicopy gene families located exclusively in subtelomeric regions of multiple P. chabaudi chromosomes, interspersed amongst multicopy intergenic regions. Hence all chromosomes share a common subtelomeric structure, presumably playing a similar role in spatial positioning as the P. falciparum Rep20 sequence. Expression in blood stages, domains characteristic of surface antigens and copy numbers between four and several hundred per genome, indicate a functional role in antigenic variation for some of these families. We identify members of the cir family, as well as novel genes, that although clearly homologous to cir have large low complexity regions in the predicted extracellular domains. Although all families have homologues in other rodent Plasmodium species, four were previously not known to be subtelomeric. Six have homologues in human and simian malarias.

Animals↗

Analysis of distal flanking regions of maize 19-kDa zein genes.

Two genomic fragments from maize, each containing a 19-kDa zein gene with extensive flanking regions, have been sequenced and examined by computer-aided analysis and Southern blotting techniques. Sequence analysis of the distal flanking sequences has revealed interesting sequence motifs, some not seen before. In particular, four nearly identical, G + C-rich, 17 to 21-bp perfect palindromes were found clustered in a 133-bp stretch lying 2 kb upstream from the zein-coding region in the genomic clone pMS2. These palindromic sequences exhibit other interesting features, including a precise spatial organization with respect to each other, and their proximity to several other repeated motifs in the same region. Southern blot analysis indicates that these palindromes, or closely related sequences, are found frequently in the maize genome. Possible secondary structures for the palindrome units are presented, which resemble functionally important sequences found upstream from other eukaryotic genes.

Base Sequence↗

Specific neuro-cognitive impairments associated with Turner (45,X) and Klinefelter (47,XXY) syndromes: a review.

In Turner syndrome, there is a high incidence of neuro-cognitive impairment specific to the mental processing and sequencing of rotational transformations of shapes in the spatial dimensions of left-right, up-down, and back-front. In Klinefelter syndrome, the corresponding neuro-cognitive impairment is specific to the mental processing of sequence and synchrony in the temporal dimension, including the temporal dimension of language.

Cognition Disorders↗

Structure-function of the putative I-domain within the integrin beta 2 subunit.

The central region (residues 125-385) of the integrin beta(2) subunit is postulated to adopt an I-domain-like fold (the beta(2)I-domain) and to play a critical role in ligand binding and heterodimer formation. To understand structure-function relationships of this region of beta(2), a homolog-scanning mutagenesis approach, which entails substitution of nonconserved hydrophilic sequences within the beta(2)I-domain with their homologous counterparts of the beta(1)I-domain, has been deployed. This approach is based on the premise that beta(1) and beta(2) are highly homologous, yet recognize different ligands. Altogether, 16 segments were switched to cover the predicted outer surface of the beta(2)I-domain. When these mutant beta(2) subunits were transfected together with wild-type alpha(M) in human 293 cells, all 16 beta(2) mutants were expressed on the cell surface as heterodimers, suggesting that these 16 sequences within the beta(2)I-domain are not critically involved in heterodimer formation between the alpha(M) and beta(2) subunits. Using these mutant alpha(M)beta(2) receptors, we have mapped the epitopes of nine beta(2)I-domain specific mAbs, and found that they all recognized at least two noncontiguous segments within this domain. The requisite spatial proximity among these non-linear sequences to form the mAb epitopes supports a model of an I-domain-like fold for this region. In addition, none of the mutations that abolish the epitopes of the nine function-blocking mAbs, including segment Pro(192)-Glu(197), destroyed ligand binding of the alpha(M)beta(2) receptor, suggesting that these function-blocking mAbs inhibit alpha(M)beta(2) function allosterically. Given the recent reports implicating the segment equivalent to Pro(192)-Glu(197) in ligand binding by beta(3) integrins, these data suggest that ligand binding by the beta(2) integrins occurs via a different mechanism than beta(3). Finally, both the conformation of the beta(2)I-domain and C3bi binding activity of alpha(M)beta(2) were dependent on a high affinity Ca(2+) binding site (K(d) = 105 microm), which is most likely located within this region of beta(2).

Amino Acid Sequence↗

A low-field MR system in acute traumatological imaging in radiology. Technical note.

PURPOSE: In this paper initial clinical observations with the application of a new low-field MR system in acute musculoskeletal imaging in radiology are presented. MATERIALS AND METHODS: This system is designed as a three-sided open permanent magnet with a revolving table. Main magnetic field strength is 0.2 T, maximal gradient field strength 20 mT/m, and minimal gradient rise time is 800 microseconds. First clinical experiences in 25 patients with acute trauma of peripheral joints were obtained. The following sequences were applied: gradient-echo (GRE) STIR, T1-weighted spin-echo, and PD- and T2-weighted turbo spin-echo. Correlation with the findings of high-field MR (1.5 T), plain radiograms (including stress views), CT or ultrasound was available in each patient confirming the low-field diagnoses. Furthermore, phantom measurements were performed to verify the spatial resolution of the system for some sequences with typical measurement parameters. DISCUSSION: This low-field system has several advantages: fast and economic installation, limited required space, easy operation, and high patient comfort. The medical benefit of such a system integrated in the traumatological suite of a radiological department seems to be evident due to the impact on the early initiation of the correct therapy which avoids additional diagnostic procedures and therefore may reduce costs. Implementation of such a system may be useful, if installation of an additional high-field scanner is not possible due to economic considerations.

Adolescent↗

RF inhomogeneity compensation in structural brain imaging.

Three-dimensional T(1)-weighted magnetization-prepared rapid gradient-echo (MP-RAGE) sequences with centric phase encoding (PE) in the inner loop provide structural brain images with a high spatial resolution and high tissue contrast. A disadvantage of this sequence type is the susceptibility to inhomogeneities of the radiofrequency (RF) coil, which may result in poor image contrast in some peripheral regions. A special excitation pulse is presented which compensates for these effects in both the head/foot and anterior/posterior directions. This pulse has a duration of only 1.3 ms and is thus compatible with the short repetition times (TRs) required for MP-RAGE imaging. It is shown experimentally that images acquired with the compensation pulse may be segmented without using intensity correction algorithms during data postprocessing.

Artifacts↗

Velocity encoding versus acceleration encoding for pressure gradient estimation in MR haemodynamic studies.

Many methods have been proposed to extract pressure gradient maps from magnetic resonance (MR) images. They were based on the resolution of the haemodynamic model of Navier-Stokes and needed the flow acceleration to be known. Most used velocity data acquisition and computed acceleration from temporal and spatial derivatives of the velocity field. However, MR sequences have been developed in order to acquire the acceleration field directly. Here we compared direct MR measurements of acceleration field components with those calculated from MR velocity acquisitions. Two experimental phantoms were used to separately evaluate the inertial and convective components of the acceleration. Mathematical simulation of the convective phantom further explained the origin of the noise generated by the spatial and temporal derivatives of the velocity data, and the misregistration artefacts due to MR sequences. We found that direct measurement of the acceleration field generates less noise and fewer artefacts than calculation from velocity derivatives.

Acceleration↗

Exploring the transcriptional crosstalk between adipose tissue and locoregional recurrence in breast cancer using independent component analysis.

Locoregional recurrence (LRR) poses a persistent clinical challenge in breast cancer, with emerging evidence implicating the tumor-associated adipose tissue in modulating recurrence risk. This study investigates shared transcriptional programs between adipose tissue and breast tumors and examines their association with disease-free survival (DFS), particularly in the context of reconstructive surgery where adipose tissue from different body compartments are commonly used. We analyzed bulk gene expression data from 5,691 breast tumors and 978 human adipose tissue samples from different body compartments using consensus-independent component analysis (c-ICA) to identify transcriptional components (TCs). Gene set enrichment analysis (GSEA) and copy number alteration profiling were used for biological annotation. Associations between TCs and DFS were evaluated through univariate Cox regression. Key findings were validated using spatial transcriptomic and single-cell RNA sequencing datasets. Among the 411 TCs identified, 332 showed biological enrichment, and 35 were significantly associated with DFS. Four DFS-associated TCs (TC257, TC350, TC371, TC400) were enriched for adipogenesis-related genes and exhibited heightened activity in high-grade, triple-negative tumors and in patients with elevated BMI. Notably, TC350 was highly active in adipose tissue from common reconstructive donor sites (abdomen, omentum, subcutis) but not in native breast adipose tissue. Spatial transcriptomic and single-cell analyses confirmed the increased activity of these adipogenesis-related TCs in tumor regions and adipose cells. TC350 included FABP4, a gene previously linked to poor prognosis in breast cancer and considered as a potential new therapeutic target. Adipose tissue-derived transcriptional programs influence breast cancer prognosis and this seems to differ by tissue origin. These findings generate a hypothesis that donor site selection for adipose tissue in reconstructive surgery may impact LRR risk through adipogenesis-associated mechanisms. Further research is warranted to elucidate the biological and clinical implications of adipose-tumor transcriptional interactions.

Humans↗

ROSINA (RSI), a novel protein with DNA-binding capacity, acts during floral organ development in Antirrhinum majus.

Petal and stamen identity of the Antirrhinum majus flower is under the genetic control of the floral homeotic gene DEFICIENS (DEF). To isolate factors involved in the regulation of DEF gene activity, a promoter segment of this B-function gene, containing cis-acting regulatory elements, was used to identify the novel trans-acting factor ROSINA (RSI). RSI does not show an extended similarity with any gene product present in the database. Rather RSI constitutes a protein that contains domains similar to known proteins from organisms of different phyla. The capacity of RSI to bind a sequence element of the DEF promoter, its spatial and temporal expression pattern together with the phenotype of RSI-RNAi interference plants as well as RSI over-expression in Arabidopsis thaliana suggest that RSI is a putative regulator of DEF gene activity in A. majus.

Amino Acid Sequence↗

Flow-independent magnetic resonance venography of the calf.

PURPOSE: To show that flow-independent venography that exploit the intrinsic MR properties of blood to isolate vessels from surrounding structures can be used for depiction of peripheral veins and for detection of deep venous thrombosis (DVT). MATERIALS AND METHODS: Sequence and parameters were first determined on a theoretical basis. The sequence was then optimized in volunteers (N = 4) for the depiction of the peripheral venous vessels. Qualitative evaluation of the normal venous anatomy was performed in five volunteers. The feasibility of diagnosing DVT of the calf with this method was evaluated with preliminary clinical studies. RESULTS: Excellent depiction of the venous anatomy was achieved in all volunteers with the optimized technique. Very small venous structures, such as superficial, muscular, and perforator veins, were clearly depicted because of the high spatial and contrast resolution capacities of the sequences. In all six patients, DVT findings diagnosed by duplex sonography were also seen on MR venography. CONCLUSION: Venous anatomy mapping and detection of lower-limb DVT appear feasible using flow-independent MR venograms.

Humans↗