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At least 721 records · Page 40Linked to original sources
PubMed2001

Quantitative comparison of algorithms for tracking single fluorescent particles.

Single particle tracking has seen numerous applications in biophysics, ranging from the diffusion of proteins in cell membranes to the movement of molecular motors. A plethora of computer algorithms have been developed to monitor the sub-pixel displacement of fluorescent objects between successive video frames, and some have been claimed to have "nanometer" resolution. To date, there has been no rigorous comparison of these algorithms under realistic conditions. In this paper, we quantitatively compare specific implementations of four commonly used tracking algorithms: cross-correlation, sum-absolute difference, centroid, and direct Gaussian fit. Images of fluorescent objects ranging in size from point sources to 5 microm were computer generated with known sub-pixel displacements. Realistic noise was added and the above four algorithms were compared for accuracy and precision. We found that cross-correlation is the most accurate algorithm for large particles. However, for point sources, direct Gaussian fit to the intensity distribution is the superior algorithm in terms of both accuracy and precision, and is the most robust at low signal-to-noise. Most significantly, all four algorithms fail as the signal-to-noise ratio approaches 4. We judge direct Gaussian fit to be the best algorithm when tracking single fluorophores, where the signal-to-noise is frequently near 4.

Algorithms↗
PubMed1994

Time-resolved delayed luminescence image microscopy using an europium ion chelate complex.

Improvements and extended applications of time-resolved delayed luminescence imaging microscopy (TR-DLIM) in cell biology are described. The emission properties of europium ion complexed to a fluorescent chelating group capable of labeling proteins are exploited to provide high contrast images of biotin labeled ligands through detection of the delayed emission. The streptavidin-based macromolecular complex (SBMC) employs streptavidin cross-linked to thyroglobulin multiply labeled with the europium-fluorescent chelate. The fluorescent chelate is efficiently excited with 340-nm light, after which it sensitizes europium ion emission at 612 nm hundreds of microseconds later. The SBMC complex has a high quantum yield orders of magnitude higher than that of eosin, a commonly used delayed luminescent probe, and can be readily seen by the naked eye, even in specimens double-labeled with prompt fluorescent probes. Unlike triplet-state phosphorescent probes, sensitized europium ion emission is insensitive to photobleaching and quenching by molecular oxygen; these properties have been exploited to obtain delayed luminescence images of living cells in aerated medium thus complementing imaging studies using prompt fluorescent probes. Since TR-DLIM has the unique property of rejecting enormous signals that originate from scattered light, autofluorescence, and prompt fluorescence it has been possible to resolve double emission images of living amoeba cells containing an intensely stained lucifer yellow in pinocytosed vesicles and membrane surface-bound SBMC-labeled biotinylated concanavalin A. Images of fixed cells represented in terms of the time decay of the sensitized emission show the lifetime of the europium ion emission is sensitive to the environment in which it is found. Through the coupling of SBMC to streptavidin,a plethora of biotin-based tracer molecules are available for immunocytochemical studies.

Animals↗
PubMed2002

L-carnitine accelerates the in vitro regeneration of neural network from adult murine brain cells.

The development, growth and regeneration of nerve cells remain an unresolved issue. The up-to-date reported brain repair mechanisms are numerous and evidence suggests that, apart from the required trophism, tropism, microenvironment and specificity of the brain, a plethora of chemical, physiological and immunological compounds can contribute to such events. Among these compounds, we concentrated our interest on L-carnitine (L-Cn), which regulates the beta-oxidation of long chain fatty acids necessary for brain development, myelinization and growth. In contrast to fetal brain cells that grow easily in culture, adult brain cells show limited neurogenesis. Here, using adult brain cells from experimental mice, we show that although L-Cn does not improve their proliferative activity in short-term cultures, it accelerates the growth and differentiation of neurons, astrocytes, oligodendrocytes and ependymal cells from neurospheres in long-term cultures. Thus, the formation of a confluent neural network requires a 2-month period in culture. These observations provide new insights for in vivo use of L-Cn to support brain cell development in cases of injury or brain degenerative diseases.

Animals↗
PubMed1999

How clinicians should use the diagnostic laboratory in a changing medical world.

In developed countries, clinicians are faced with a plethora of diagnostic tests to apply to patients to guide their clinical management. The quality, effectiveness, and efficiency of patient care should be foremost in the clinician's mind. Laboratory directors should make every effort to guide clinicians in appropriate laboratory test ordering, interpretation, and resulting actions. Medicine, being at its center a moral enterprise grounded in a covenant of trust, and laboratory medicine being a subset of medicine, must first care and advocate for the patient, and consider clinical outcomes as most important.

Administrative Personnel↗
PubMed2003

Cold--an underrated risk factor for health.

Cardiovascular diseases (CVD) are responsible for around 20% of all deaths worldwide (approximately 14 million) and are the principal cause of death in all developed countries, accounting for 50% of all deaths. Variations in the annual per capita death rates in different countries are well documented. Less well known are seasonal variations in death rates, with the highest levels occurring during the colder winter months, which have been described in many countries. This phenomenon is referred to as excess winter mortality. CVD-related deaths account for the majority of excess winter deaths (up to 70% in some countries), while about half of the remaining are due to increases in respiratory diseases. Paradoxically, CVD mortality increases to a greater extent with a given fall in temperature in regions with warm winters. While much of the indirect evidence points to the notion that cold is somehow involved in explaining excess winter deaths, the mechanism by which seemingly mild exposure to cold ambient conditions can increase the risk of death remains unclear. The strong indirect epidemiological evidence coupling cold climate to mortality may be related to indoor rather than outdoor climatic conditions (e.g., cold/damp houses versus warm/dry houses) coupled with a plethora of factors including health status, ageing-related deterioration in physiological and behavioral thermoregulation, toxicology, and socioeconomic factors.

Adult↗
PubMed2002

Neuropeptide Y receptors as targets for anti-obesity drug development: perspective and current status.

Neuropeptide Y is a widely distributed neuropeptide that elicits a plethora of physiological effects via interaction with six different receptors (Y(1)-y(6)). Recent attention has focused on the role of neuropeptide Y in the regulation of energy homeostasis. Neuropeptide Y stimulates food intake, inhibits energy expenditure, increases body weight and increases anabolic hormone levels by activating the neuropeptide Y Y(1) and Y(5) receptors in the hypothalamus. Based on these findings, several neuropeptide Y Y(1) and Y(5) receptor antagonists have been developed recently as potential anti-obesity agents. In addition, mice lacking neuropeptide Y, the neuropeptide Y Y(1) receptor or the neuropeptide Y Y(5) receptor have been generated. The data obtained to date with these newly developed tools suggests that neuropeptide Y receptor antagonists, particularly neuropeptide Y Y(1) receptor antagonists, may be useful anti-obesity agents. However, the redundancy of the neurochemical systems regulating energy homeostasis may limit the effect of ablating a single pathway. In addition, patients in whom the starvation response is activated, such as formerly obese patients who have lost weight or patients with complete or partial leptin deficiency, may be the best candidates for treatment with a neuropeptide Y receptor antagonist.

Animals↗
PubMed2000

ATP-dependent regulation of nuclear Ca(2+) levels in plant cells.

Localised alterations in cytoplasmic Ca(2+) levels are an integral part of the response of eukaryotic cells to a plethora of external stimuli. Due to the large size of nuclear pores, it has generally been assumed that intranuclear Ca(2+) levels reflect the prevailing cytoplasmic Ca(2+) levels. Using nuclei prepared from carrot (Daucus carota L.) cells, we now show that Ca(2+) can be transported across nuclear membranes in an ATP-dependent manner and that over 95% of Ca(2+) is accumulated into a pool releasable by the Ca(2+) ionophore A.23187. ATP-dependent nuclear Ca(2+) uptake did not occur in the presence of ADP or ADPgammaS and was abolished by orthovanadate. Confocal microscopy of nuclei loaded with dextran-linked Indo-1 showed that the initial ATP-induced rise in [Ca(2+)] occurs in the nuclear periphery. The occurrence of ATP-dependent Ca(2+) uptake in plant nuclei suggests that alterations of intranuclear Ca(2+) levels may occur independently of cytoplasmic [Ca(2+)] changes.

Adenosine Triphosphate↗
PubMed2001

Nuclear and mitochondrial apoptotic pathways of p53.

In contrast to p53-mediated cell cycle arrest, the mechanisms of p53-mediated apoptosis in response to cellular stresses such as DNA damage, hypoxia and oncogenic signals still remain poorly understood. Elucidating these pathways is all the more pressing since there is good evidence that the activation of apoptosis rather than cell cycle arrest is crucial in p53 tumor suppression. Moreover, the therapeutic interest in p53 as the molecular target of anticancer intervention rests mainly on its powerful apoptotic capability. This puzzling elusiveness suggests that p53 not only engages a plethora of downstream pathways but itself might possess a biochemical flexibility that goes beyond its role as a mere transcription factor. Recent evidence of a direct pro-apoptotic role of p53 protein at mitochondria suggests a synergistic effect with its transcriptional activation function and brings an unexpected new level of complexity into p53 apoptotic pathways.

Animals↗
PubMed2001

Cellular strategies for proteolytic targeting during migration and invasion.

Cell migration over or through the extracellular matrix (ECM) is an integral feature of both physiological and pathological processes. Regulation of the changing cell-ECM interactions involved can be effected by proteolysis and requires strict spatial and temporal targeting of proteinase activity. The versatile use of different proteinase systems, with a variety of localisation mechanisms and cleavage targets, is being revealed by a plethora of studies using in vitro models. This mini review reflects the status of our knowledge of strategies for the localisation of proteolytic activity effected during cell migration.

Animals↗
PubMed2003

The unfolding tale of PECAM-1.

Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) is a member of the immunoglobulin (Ig) superfamily that has distinctive features of an immunoreceptor based upon its genomic structure and the presence of intrinsic immunoreceptor tyrosine inhibitory motifs (ITIMs) in its ligand binding polypeptide. This has lead to its subclassification into the Ig-ITIM superfamily. Its amino-terminal Ig-like domain of PECAM-1 is necessary for its homophilic binding, which plays an important role in cell-cell interactions. Its intracellular ITIMs serve as scaffolds for recruitment of signalling molecules including protein-tyrosine phosphatases to mediate its inhibitory co-receptor activity. Increasing evidence has implicated PECAM-1 in a plethora of biological phenomena, including modulation of integrin-mediated cell adhesion, transendothelial migration, angiogenesis, apoptosis, cell migration, negative regulation of immune cell signalling, autoimmunity, macrophage phagocytosis, IgE-mediated anaphylaxis and thrombosis. In this review, we discuss some of the new developments attributed to this molecule and its unique roles in biology.

Enzymes↗
PubMed1998

Human and mouse proteomic databases: novel resources in the protein universe.

Proteomics is an emerging area of research of the post-genomic era that deals with the global analysis of gene expression using a plethora of techniques to resolve (high resolution two-dimensional polyacrylamide gel electrophoresis, 2D PAGE), identify (peptide sequencing by Edman degradation, mass spectrometry, Western immunoblotting, etc.), quantitate and characterize proteins, as well as to store (comprehensive 2D PAGE databases), communicate and interlink protein and DNA sequence and mapping information from genome projects. Here we review the current status as well as applications of human and mouse proteomic 2D PAGE databases that are being systematically constructed for the global analysis of gene expression in both health and disease (http://biobase.dk/cgi-bin/celis). Furthermore, we discuss the problems one faces when using powerful proteomic technology to study heterogeneous tissue and tumor biopsies, and emphasize the importance of building comprehensive databases that contain a critical mass of information for both known and novel proteins in normal and disease conditions.

Animals↗
PubMed1998

Peptidyl-prolyl-cis/trans-isomerase activity may be necessary for memory formation.

At present, evidence for a plethora of physiological roles for the different classes of peptidyl-prolyl-cis/trans-isomerases (PPIases, EC 5.2.1.8) is emerging. Cyclosporin A (CyA) has been previously reported to disrupt memory formation in a temporally specific manner, when administered intracranially to day-old chicks trained on a single-trial, passive-avoidance task [Bennett, P.C., Zhao, W., Lawen, A. and Ng, K.T. (1996) Brain Res. 730, 107-1171. CyA is known to inhibit both the PPIase activity of cyclophilin and, indirectly, the protein phosphatase activity of calcineurin. Therefore to begin to distinguish between these two functions we studied the effects on memory formation of three non-immunosuppressive CyA analogues, in order to study the involvement of cyclophilins. These drugs retain the capacity to bind to and inhibit the PPIase activity of cyclophilin, but do not bind in the complex with cyclophilin to calcineurin and, therefore, do not inhibit its phosphatase activity. All three drugs exert effects on memory formation comparable to those induced by CyA, significantly inhibiting memory formation when injected intracranially (50 fmol per hemisphere) immediately following training. Brain extracts from chicks treated with [MeVal4]CyA show a strong inhibition of cyclophilin activity. These data show a requirement for the PPIase activity of a cyclophilin for successful memory formation and constitute the first set of data establishing a physiological role for a cyclophilin.

Animals↗
PubMed1984

Human in vitro fertilization and the law.

Physicians who intend to perform in vitro fertilization--or the newer alternative, in vivo fertilization and embryo transfer--should be aware of the plethora of laws that potentially cover their work. In perhaps no other area of medicine are there so many separate statutes and regulations that potentially apply. State fetal research laws, abortion statutes, human subject protection laws, and specific in vitro fertilization statutes can determine whether and how the procedure can be undertaken. When donor gametes or a surrogate carrier is used, additional laws governing artificial insemination, paternity, or adoption may come into play to determine the child's legal status and its relationship to the parties involved. This article is designed to guide physicians through the legal maze.

Advisory Committees↗
PubMed2003

Empowerment or control? An analysis of the extent to which client participation is enabled during health visitor/client interactions using a structured health needs assessment tool.

The demand for explicitness in the way health visitors target their services has given rise to a plethora of different health needs assessment tools (HNATs). This paper describes an in-depth conversational analysis of the use in practice of these structured health needs assessment tools (HNATs) in two different NHS Community Trusts in England. These HNATs aimed to enable clients to participate in the assessment of their own health needs, as well as fulfilling the political requirements of justifying the expenditure of health visitor time where needs are identified. However, conversational analysis of 10 interactions showed that use of the instruments was associated with a failure to either identify needs that are relevant to the client or to enable clients to participate in the process. Use of the structured instrument simultaneously emphasises the significance of a professional lead, instead of client participation, and minimises the importance of inter-personal relationships and communication. In one site, a directly controlling style was apparent in the practice of health visitors who were, themselves, explicitly controlled by their managers. In the other site, professional expertise was emphasised, and a covert assessment style acted to disempower clients. The controlling nature of the interactions, the number of missed cues and the possibility of distress caused by the insensitivity of questioning style are all potentially harmful side effects of using structured instruments to assess needs. The problems seem to stem from the use of a pre-determined list of questions that form the basis for assuming that any family's health promotion needs can be categorised and predicted in advance. In conclusion, therefore, it is recommended that health visitors should use the open, conversational style of needs assessment that has been shown to be effective and acceptable, rather than an approach based on a structured instrument.

Communication↗
PubMed1998

The effects of continuing professional education on the clinical practice of nurses: a review of the literature.

In recent years, nurse education in the United Kingdom has undergone some major organisational changes, not least the move into higher education institutions. Concurrently, developments in the professionalisation of nursing through the UKCC's post registration education and practice reforms have led to nurses being required to identify their own educational needs. Having identified their own needs, nurses are then faced with a plethora of post registration education programmes which are aimed to facilitate their learning and, directly or indirectly, improve patient care. However, do these programmes positively influence nursing practice and the standard of care delivered? This paper reviews the literature regarding the effects that continuing professional education (CPE) programmes have upon the clinical practice of nurses. The review draws on papers primarily from North America and the UK and highlights the methodological approaches used to discover if and how nursing practice is influenced. The impact of CPE on nurses as individuals is detailed along with the effects of CPE on the delivery of patient care.

Clinical Competence↗
PubMed2001

Cellular trafficking in trypanosomatids: a new target for therapies?

Pathogenic trypanosomatids cause a plethora of diseases marked by the lack of efficient vaccines and therapies. As a consequence, studies are being conducted that are geared towards the understanding of basic mechanisms and various biological aspects of these parasites that might be used as targets for new developments in these areas. One such aspect is the understanding of specific cellular trafficking mechanisms that might be attacked with the intention of disease control. In this paper, we give an overview of our current knowledge of cellular targeting mechanisms in trypanosomatids, with special emphasis on our data related to lysosomal targeting of cysteine proteinases in Leishmania.

Animals↗
PubMed1998

Implications of global change and climate variability for vector-borne diseases: generic approaches to impact assessments.

Global change is pervasive and occurring at a dramatic rate. It involves changes in land use, vegetation cover, species translocations and even the climate of the planet. The consequences for the biosphere are uncertain. Past research emphasis has been on the science of climate change as the major driver of policy. The present priority in the global-change community is to define the likely nature and extent of those impacts on biodiversity and the functioning of ecosystems. In addition, increasing consideration is now being given to adaptation measures. The way in which that is being initiated is to develop adaptation measures to respond to medium-term climate variability in the form of altered El Nino and similar cycles, and changes in the frequency of extreme events. Given the large number of stakeholders in agriculture, human health and environment, there is a need for great efficiencies if the scientific community is going to be able to respond in a meaningful way with foreseeable resources. The plethora of problems means that generic approaches are needed. The present situation, with parasitologists each doing their own thing in terms of developing and using software tools, is like the tower of Babel. Parasitologists need common tools and languages to facilitate communication and collaboration. Advances in computing, with object-oriented programming languages and seamless exchange of information between different packages and platforms, are providing some exciting opportunities to overcome these problems.

Adaptation, Biological↗
PubMed1999

Simultaneous quantitation of 15 cytokines using a multiplexed flow cytometric assay.

Several methods have been developed to quantify cytokines and chemokines in biological fluids and tissue culture samples, including bioassays, enzyme-linked immunosorbent assay (ELISA), intracellular staining, ribonuclease protection assay (RPA) and polymerase chain reaction (PCR). However, each of these techniques possesses one or more significant limitations. Here, we describe a new multiplexed assay, using the FlowMetrix system, that can quantify multiple cytokines simultaneously in a small sample volume. This assay was found to be more accurate, sensitive and reproducible than the conventional microtitre ELISA procedure. Furthermore, the time and cost involved are comparable to, or less than, the ELISA. A key feature of the FlowMetrix assay is its ability to multiplex: here, we show that this assay can accurately quantitate 15 cytokines in a 100 microl sample volume while the same analysis by ELISA requires 1.5 ml (100 microl for each cytokine assay). By using this Flow Metrix assay, we could demonstrate that only T helper 1 (T(H)1)-deviated cells produce detectable levels of interleukin (IL)-2, while only T(H)2-deviated cells produce significant amounts of IL-4. Six other cytokines were produced by both T cell subsets, with the T(H)1 population producing more IL-3, granulocyte-monocyte colony stimulating factor (GM-CSF) and interferon (IFN)-gamma, and the T(H)2 population producing more IL-5, IL-10, and IL-13. Seven other cytokines were not produced in detectable amounts. This assay should prove to be a powerful tool in the quantitation of cytokines, or any other soluble product for which antibody pairs are available. It will also provide a more complete picture of the plethora of cytokines secreted during an immune response.

Animals↗