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PubMed2005

Characterization of vascular protein expression patterns in cerebral ischemia/reperfusion using laser capture microdissection and ICAT-nanoLC-MS/MS.

Cerebral ischemia rapidly initiates structural and functional changes in brain vessels, including blood-brain barrier disruption, inflammation, and angiogenesis. Molecular events that accompany these changes were investigated in brain microvessels extracted using laser-capture microdissection (LCM) from Sprague-Dawley rats subjected to a 20 min transient global cerebral ischemia followed by 1, 6, or 24 h reperfusion. Proteins extracted from approximately 300 LCM captured microvessels (20-100 microm) were ICAT-labeled and analyzed by nanoLC-MS. In-house software was used to identify paired ICAT peaks, which were then sequenced by nanoLC-MS/MS. Pattern analyses using k-means clustering method classified 57 differentially expressed proteins in 7 distinct dynamic patterns. Protein function was assigned using Panther Classification system. Early reperfusion (1 h) was characterized by down-regulation of ion pumps, nutrient transporters, and cell structure/motility proteins, and up-regulation of transcription factors, signal transduction molecules and proteins involved in carbohydrate metabolism. The up-regulation of inflammatory cytokines and proteins involved in the extracellular matrix remodeling and anti-oxidative defense was observed in late reperfusion (6-24 h). The up-regulation of IL-1beta and TGF-1beta in ischemic brain vessels was confirmed by ELISA, quantitative PCR, and/or immunohistochemistry. A biphasic postischemic (1 and 24 h) BBB opening for (3)H-sucrose was evident in the same model. Differentially expressed proteins identified in brain vessels during reperfusion are likely involved in orchestrating functional vascular responses to ischemia, including the observed BBB disruption.

Animals↗

PubMed2002

The microbicide cyanovirin-N expressed on the surface of commensal bacterium Streptococcus gordonii captures HIV-1.

OBJECTIVE: To explore the feasibility of expressing the potent HIV-inactivating protein, cyanovirin-N (CV-N), in the human commensal bacterium Streptococcus gordonii, as a possible approach for local delivery of CV-N to prevent sexual transmission of HIV-1. DESIGN AND METHODS: To express CV-N in S. gordonii, we used the host-vector system we had previously developed. CV-N was expressed as a fusion protein both attached to the bacterial surface and secreted in soluble form in the supernatant of liquid cultures. The soluble form of recombinant CV-N was tested for gp120-binding activity in an enzyme-linked immunosorbent assay, whereas S. gordonii strain expressing CV-N on the surface was analyzed in an in vitro HIV capturing assay. RESULTS: Two recombinant S. gordonii strains secreting or displaying CV-N on the bacterial surface were constructed and the expression of CV-N was confirmed by immunoblot and flow-cytometric analysis. The secreted form of recombinant CV-N exhibited a concentration-dependent binding to the envelope glycoprotein gp120 of HIV-1, whereas CV-N displayed on the bacterial surface was able to capture HIV virions efficiently. CONCLUSION: The anti-HIV protein CV-N in S. gordonii was expressed in a biologically active form. This represents a first step in the development of a system to deliver and maintain an effective concentration of a microbicide in the vaginal mucosa.

Bacterial Proteins↗

PubMed1993

Establishment of accurate incidence rates for head and spinal cord injuries in developing and developed countries: a capture-recapture approach.

Prevention of head and spinal cord injuries is defined as a reduction in the incidence of these disabilities. Accurate incidence data are fundamental to any prevention program. The current approaches toward determining incidence rates for head and spinal cord injuries are summarized. Previous research has focused on passive surveillance systems and population-based registries. An alternative system for monitoring the incidence of head injuries is discussed that uses a surveillance methodology called capture-recapture. This method employs multiple population-based sources to identify cases and uses the cases that overlap between the sources to estimate the degree of undercount in the population. This estimate in turn is used to produce an ascertainment-corrected incidence estimate. Through the use of methods such as capture-recapture, accurate monitoring of the incidence of head and spinal injuries across developing and developed countries is indeed feasible.

Bias↗

PubMed2001

Managed care's attempts to capture aesthetic surgery.

Managed care organizations recently have attempted to add aesthetic surgery to their line of available services. To better understand the challenges posed by these actions, all members of the American Society for Aesthetic Plastic Surgery were surveyed about managed care overtures to aesthetic plastic surgeons, their responses, and the likely responses of their patients. The goal was to examine both the supplier and consumer ends of the aesthetic surgery market to determine the likely effects of managed care's attempts to capture aesthetic surgery. A total of 632 plastic surgeons returned the surveys (response rate, 54.5 percent). Twenty-two percent reported being approached by managed care organizations about joining a panel of aesthetic surgeons. Approximately one-quarter of the plastic surgeons said they would participate in aesthetic surgery panels developed by managed care organizations. Characteristics significantly associated with willingness to participate were solo practice structure, a low percentage of practice revenues from aesthetic surgery, and a very competitive practice environment. Plastic surgeons believed that their colleagues would be even more willing to acquiesce to managed care aesthetic surgery; more than one-third said that 25 to 50 percent of their colleagues would join, and nearly one-third thought that more than half would participate. Plastic surgeons believed that many of their patients would also participate in managed care aesthetic surgery. Twenty-four percent thought that more than half of their patients would choose an aesthetic surgeon through their managed care organization if that organization developed a network for aesthetic surgery. This figure increased to almost 40 percent if the organization would deny coverage for complications resulting from nonpanel surgeons, and to 41 percent if the organization would offer price discounts. This survey shows that most plastic surgeons are against managed care aesthetic surgery. But it also shows that some plastic surgeons will participate, and that most plastic surgeons think many of their colleagues and patients will do likewise. This means that managed care organizations have the potential to make inroads in aesthetic surgery on both the supplier and consumer ends of the market. To prevent managed care from capturing aesthetic surgery, plastic surgeons must anticipate the likely business strategy of managed care. To this end, they must understand the steps involved in the creation of a new service business and offer organized countermeasures against each of them.

Attitude of Health Personnel↗

PubMed2006

Laser capture microdissection of epithelial cancers guided by antibodies against fibroblast activation protein and endosialin.

Transcriptional profiling of cancer biopsies is used extensively to identify expression signatures for specific cancer types, diagnostic and prognostic subgroups, and novel molecular targets for therapy. To broaden these applications, several challenges remain. For example, the integrity of RNA extracted even from small tissue samples has to be insured and monitored. Moreover, total tumor RNA may hide the marked histologic heterogeneity of human cancers. A principle approach to this heterogeneity has been provided by laser capture microdissection performed on antibody-stained tissue sections (immuno-LCM; iLCM). In this study, we have established a procedure to assess the quality of RNA obtained from tissue sections, coupled with immunostaining using antibodies to different tumor stromal markers, and subsequent iLCM to selectively capture the cancer stroma compartments. The procedure was applied to 53 frozen specimens of human epithelial cancers. Sections were stained for histopathological evaluation, and RNA was isolated from adjacent serial sections. RNA quality was assessed by the Agilent-Bioanalyzer (Agilent, Palo Alto, CA) and by multiplex RT-PCR. Two thirds of the specimens were found to yield good to excellent RNA quality. For microdissection of the tumor stroma with reactive fibroblasts and tumor blood vessels, a rapid incubation protocol with antibodies against fibroblast activation protein (FAP) and against endosialin was developed to ensure RNA integrity for subsequent iLCM. Using these procedures, RNA from distinct tumor compartments can be isolated, analyzed, amplified, and used for transcription profiling.

Antibodies, Neoplasm↗

PubMed2005

Methodology for preservation of high molecular-weight RNA in paraffin-embedded tissue: application for laser-capture microdissection.

Laser-capture microdissection techniques have enhanced the ability to perform molecular studies of pure-cell populations. Although many technical factors affect the outcome of the procedure, none is more critical than the appropriate handling of the tissue. Because extraction of intact RNA from paraffin-embedded tissue is a difficult and inconsistent process, frozen sections with their attendant problems are used for this purpose. The major limitation of frozen section is its inferior morphologic quality compared with paraffin-embedded sections that may complicate accurate identification of cells during microdissection. We have developed a procedure that provides both high-quality histomorphology and RNA preservation in paraffin-embedded tissue. It is based on the use of a methanol-based fixative coupled with microwave-assisted rapid tissue processing. This technology in conjunction with a modified hematoxylin-eosin stain and a RNA extraction method allows isolation of high molecular-weight RNA from laser-capture microdissected, hematoxylin and eosin-stained paraffin sections. The high quality of the extracted RNA was confirmed by capillary electrophoresis and RT-PCR. The combination of a methanol-based fixative, rapid microwave tissue processing, and a modified hematoxylin and eosin stain produces paraffin sections that yield high molecular-weight RNA upon microdissection. This methodology opens the door for a wide range of gene expression analyses using paraffin-embedded tissue.

Electrophoresis, Capillary↗

PubMed2025

CUT&TIME captures the history of open chromatin in developing neurons.

Chromatin structure plays a central role in defining cell identity by regulating gene expression. During development, shifts in chromatin structure facilitate changes in gene expression needed to specify distinct cell types. To understand how changes in chromatin structure influence the developmental trajectory of neural progenitor cells, we developed CUT&TIME, a technique that uses a hyperactive 6-methyl adenosine (6mA) methyltransferase pulsed in living cells to map historical chromatin accessibility genome-wide in single cells. We show that CUT&TIME produces a record of the chromatin landscape during neurogenesis in the developing retina, specifically as neural progenitors produce the major projection neuron type, retinal ganglion cells (RGCs). We further show that this method is compatible with single cell profiling technologies, which allows us to visualize and capture the diversity of chromatin states that produce RGCs. Additionally, we identify changes in promoter accessibility associated with the transition from progenitor to RGC. Together, these data demonstrate that CUT&TIME captures a historical record of chromatin structure, which can be used to identify early changes in accessibility associated with cell-fate commitment.

Journal Article↗

PubMed2003

Control of chaos by capture and release.

We propose and demonstrate a different method for the control of flip saddles in dissipative chaotic systems. Due to the dynamics of a flip saddle, the stable manifolds of a target orbit and its perturbation can be modeled as a pair of concentric Möbius bands. Over the period of the target orbit, these bands rotate relative to one another. The method of capture and release (CR) takes advantage of this rotation, and captures a nearby system state in the perturbed stable manifold, releasing it when the rotation of the Möbius bands brings them into alignment. Unlike the method of Ott, Grebogi, and Yorke and most of its variants, CR does not rely on the unstable component of the flow to push the system state onto the stable manifold; the system state is evolving in a stable subspace for the duration of the control perturbation.

Journal Article↗

PubMed2005

Lattice Boltzmann interface capturing method for incompressible flows.

A lattice Boltzmann interface capturing method for incompressible flows is proposed in this paper. The interface is naturally captured by minimizing the free energy functional. It is easily implemented and does not require interface reconstruction as required by most of the traditional interface tracking methods such as the volume of fluid method. Moreover, the method does not require the isotropic property of the fourth order lattice tensor as do other lattice Boltzmann methods. Thus the D2Q5 (D2 means two dimensional, Q5 means five velocity model) discrete velocity model is applied in the method. The interface profile along the flat interface and coexistence curve can be given analytically. The proposed method is validated for some test cases, and compared to the volume of fluid and level set methods. Numerical results show that the present method performs very well and can generate very sharp interfaces.

Journal Article↗

PubMed2001

Neutron capture on (180)Ta(m): clue for an s-process origin of nature's rarest isotope.

The neutron capture cross section of (180)Ta(m) has been measured in the keV range, yielding a stellar average of 1465+/-100 mb at kT = 30 keV. Though the sample contained only 6.7 mg (180)Ta(m) (at an enrichment of 5.5%), the few capture events could be separated from much larger backgrounds by a unique combination of high efficiency, good energy resolution, and high granularity of the Karlsruhe 4 pi BaF(2) detector. A detailed s-process analysis based on this first experimental value indicates that (180)Ta(m) is predominantly of s-process origin.

Journal Article↗

PubMed2001

Direct measurement of the L/K ratio in (7)Be electron capture.

The ratio of L- to K-shell electron captures in light nuclei is particularly sensitive to electron overlap and exchange effects. Calculations of these effects in (7)Be disagree by more than 20%. We report a measurement of the L/K ratio in (7)Be, using a cryogenic microcalorimeter which clearly separates L- and K-shell captures. The obtained L/K ratio of 0.040(6) is less than half that of existing predictions for free (7)Be. The discrepancy is likely due to in-medium effects distorting the L-shell electron orbitals.

Journal Article↗

PubMed2002

Time-dependent close-coupling calculations of dielectronic capture in He.

We report on the first time-dependent close-coupling calculation of dielectronic capture into a doubly excited state of a two-electron atom. An incoming electron is represented by a Gaussian wave packet which collides with singly ionized helium in its ground state. The close-coupling equations describe the propagation of the total compound wave function on a two-dimensional radial lattice. By projecting this wave function onto a doubly excited state of neutral helium, we can determine the probability amplitude for dielectronic capture into one of these states and the subsequent autoionization from it.

Journal Article↗

PubMed2002

Observation of double radiative capture on pionic hydrogen.

We report the first observation of double radiative capture on pionic hydrogen. The experiment was conducted at the TRIUMF cyclotron using the RMC spectrometer and detected gamma-ray coincidences following pi(-) stops in liquid hydrogen. We found the branching ratio for double radiative capture to be [3.05+/-0.27(stat)+/-0.31(syst)]x10(-5). The measured branching ratio and angle-energy distributions support the theoretical prediction of a dominant contribution from the pipi-->gammagamma annihilation mechanism.

Journal Article↗

PubMed2004

Enhanced electron-capture decay rate of 7Be encapsulated in C60 cages.

The decay rate of 7Be electron capture was measured in C60 and Be metal with a reference method. The half-life of 7Be endohedral C60 ((7)Be@C(60)) and 7Be in Be metal (Be metal (7Be)) is found to be 52.68+/-0.05 and 53.12+/-0.05 days, respectively. This amounts to a 0.83% difference in electron-capture decay half-life between (7)Be@C(60) and Be metal (7Be). Our result is a reflection of the different electron wave functions for (7)Be@C(60) inside C60 compared to the situation when 7Be is in a Be metal.

Journal Article↗

PubMed2005

Application of radiative electron capture for the diagnostics of spin-polarized ion beams at storage rings.

It is proposed to apply the radiative electron capture into high-Z projectiles as a probe process for measuring the spin polarization of the hydrogenlike ions at storage rings. We argue that such polarization measurements are possible since the linear polarization of emitted x-ray photons is greatly sensitive to the spin states of incoming ions with nuclear spin I > 1/2. In particular, for K-shell electron capture into the hydrogenlike ions, the linear polarization of light as measured out of the reaction plane is found to be proportional to the degree of ion polarization. Detailed computations for the dependence of the photon polarization on the ion spin states and projectile energies are carried out for the electron recombination into hydrogenlike Bi 82+ ions.

Journal Article↗

PubMed2003

Laser capture microdissection of cells from plant tissues.

Laser capture microdissection (LCM) is a technique by which individual cells can be harvested from tissue sections while they are viewed under the microscope, by tacking selected cells to an adhesive film with a laser beam. Harvested cells can provide DNA, RNA, and protein for the profiling of genomic characteristics, gene expression, and protein spectra from individual cell types. We have optimized LCM for a variety of plant tissues and species, permitting the harvesting of cells from paraffin sections that maintain histological detail. We show that RNA can be extracted from LCM-harvested plant cells in amount and quality that are sufficient for the comparison of RNAs among individual cell types. The linear amplification of LCM-captured RNA should permit the expression profiling of plant cell types.

DNA, Plant↗

PubMed2003

Laser-capture microdissection, a tool for the global analysis of gene expression in specific plant cell types: identification of genes expressed differentially in epidermal cells or vascular tissues of maize.

Laser-capture microdissection (LCM) allows for the one-step procurement of large homogeneous populations of cells from tissue sections. In mammals, LCM has been used to conduct cDNA microarray and proteomics studies on specific cell types. However, LCM has not been applied to plant cells, most likely because plant cell walls make it difficult to separate target cells from surrounding cells and because ice crystals can form in the air spaces between cells when preparing frozen sections. By fixing tissues, using a cryoprotectant before freezing, and using an adhesive-coated slide system, it was possible to capture large numbers (>10,000) of epidermal cells and vascular tissues (vascular bundles and bundle sheath cells) from ethanol:acetic acid-fixed coleoptiles of maize. RNA extracted from these cells was amplified with T7 RNA polymerase and used to hybridize a microarray containing approximately 8800 maize cDNAs. Approximately 250 of these were expressed preferentially in epidermal cells or vascular tissues. These results demonstrate that the combination of LCM and microarrays makes it feasible to conduct high-resolution global gene expression analyses of plants. This approach has the potential to enhance our understanding of diverse plant cell type-specific biological processes.

Aquaporins↗

PubMed2005

Analyzing and capturing articulated hand motion in image sequences.

Capturing the human hand motion from video involves the estimation of the rigid global hand pose as well as the nonrigid finger articulation. The complexity induced by the high degrees of freedom of the articulated hand challenges many visual tracking techniques. For example, the particle filtering technique is plagued by the demanding requirement of a huge number of particles and the phenomenon of particle degeneracy. This paper presents a novel approach to tracking the articulated hand in video by learning and integrating natural hand motion priors. To cope with the finger articulation, this paper proposes a powerful sequential Monte Carlo tracking algorithm based on importance sampling techniques, where the importance function is based on an initial manifold model of the articulation configuration space learned from motion-captured data. In addition, this paper presents a divide-and-conquer strategy that decouples the hand poses and finger articulations and integrates them in an iterative framework to reduce the complexity of the problem. Our experiments show that this approach is effective and efficient for tracking the articulated hand. This approach can be extended to track other articulated targets.

Algorithms↗