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PubMed2002

Column selectivity in reversed-phase liquid chromatography II. Effect of a change in conditions.

The isocratic retention of 67 widely-different solutes in reversed-phase liquid chromatography (RP-LC) has been investigated as a function of temperature and mobile phase composition (% B) for three different C18 columns. Similar studies were also carried out in a gradient mode, where temperature, gradient time and solvent type were varied. These results show that changes in retention with these conditions are similar for each of these three columns. This suggests that relative column selectivity as defined by experiments for one set of experimental conditions will be approximately applicable for other conditions, with the exception of changes in mobile phase pH-which can affect values of the column parameter C (a measure of silanol ionization). Column selectivity as a function of pH was explored for several columns.

Buffers↗

PubMed1998

Characterization of a receptor subtype-selective lysophosphatidic acid mimetic.

Despite an intriguing cell biology and the suggestion of a role in pathophysiological responses, the mechanism of action of such lipid phosphoric acid mediators as lysophosphatidic acid (LPA) remains obscure, in part because of an underdeveloped medicinal chemistry. We report now the agonist activity of a synthetic phospholipid in which the glycerol backbone of LPA is replaced by L-serine. Like LPA, the L-serine-based lipid mobilizes calcium and inhibits activation of adenylyl cyclase in the human breast cancer cell line MDA MB231. Treatment with LPA desensitizes MDA MB231 cells to subsequent application of the L-serine compound; when the order of application is reversed, however, the L-serine compound does not prevent calcium mobilization by LPA, which might indicate the existence of two LPA receptors in these cells. The analogous D-serine-based phospholipid was distinctly less potent than the L-isomer in those assays; this finding demonstrates stereoselectivity by an LPA receptor. Unlike LPA, the L-serine-based lipid does not evoke a chloride conductance in Xenopus laevis oocytes, but injection of poly(A)+ RNA from HEK 293 cells confers this phenotype on the oocyte. The latter result has practical importance in that it allows use of the frog oocyte for expression cloning of an LPA receptor DNA, an assay system made problematic by the oocyte's strong endogenous response to LPA.

Adenylate Cyclase Toxin↗

PubMed2002

So you want to be a specialist registrar?--What to put in your CV.

Dentists applying to a specialist training programme often receive conflicting advice over what to put in their curriculum vitae (CV). We conducted a survey of the Training Programme Directors of the dental specialties to determine what aspects of CV content and presentation styles are considered important. This has allowed us to construct guidelines for what to put in a CV. Recently, structured application forms have become increasingly popular and may be a more objective way to carry out the shortlisting process. The guidelines presented could also be used as a framework for medical personnel departments if structured application forms eventually replace the CV.

Career Mobility↗

PubMed1988

The uptake and distribution of cadmium in tomato plants as affected by ethylenediaminetetraacetic acid and 2,4-dinitrophenol.

The uptake and distribution of cadmium in tomato plants (Lycopersicon esculentum, Mill, cv. Tiny Tim) were examined with and without the presence of ethylenediaminetetraacetic acid (EDTA) as chelating agent and 2,4-dinitrophenol (DNP) as metabolic inhibitor. Eight-week-old intact and derooted tomato seedlings were used in hydroculture experiments with cadmium applied as (115)Cd(NO(3))(2) in a range of concentrations. Measurements of the (115)Cd content of roots, stems and leaves were carried out by gamma-ray spectroscopy. The data showed that applications of both EDTA and DNP resulted in reduced total Cd accumulation in the plants, but relatively enhanced Cd transport into the above-ground plant parts. The Cd mobility in the transport channels in the shoots was increased by EDTA in both intact and derooted plants. Application of DNP leads to increased relative Cd import to leaves in derooted plants, but a reduced import into leaves of intact plants. These results suggest that Cd-complexes are formed in root cells before root-to-shoot transport. Furthermore, initial Cd uptake may be associated with adsorption on the negative charges of the cell walls of the root system. The high Cd mobility in shoots, in experiments with intact plants and Cd-EDTA application, indicates the possibility of simultaneous uptake of Cd and EDTA, possibly as a Cd-EDTA complex.

Journal Article↗

PubMed2006

Label-free kinase profiling using phosphate affinity polyacrylamide gel electrophoresis.

Herein we describe three applications of label-free kinase profiling using a novel type of phosphate affinity polyacrylamide gel electrophoresis. The phosphate affinity site is a polyacrylamide-bound dinuclear Mn2+ complex that enables the mobility shift detection of phosphorylated proteins from their nonphosphorylated counterpart. The first application is in vitro kinase activity profiling for the analysis of varied phosphoprotein isotypes in phosphorylation status. The activity profiles of six kinds of kinases, glycogen synthase kinase-3beta, cyclin-dependent kinase 5/p35, protein kinase A, mitogen-activated protein kinase (MAPK), casein kinase II, and calmodulin-dependent protein kinase II, were determined using a substrate protein, Tau, which has a number of phosphorylation sites. Each kinase demonstrated characteristic multiple electrophoresis migration bands up-shifted from the nonphosphorylated Tau due to differences in the phosphorylation sites and stoichiometry. The second application is in vivo kinase activity profiling for the analysis of protein phosphorylation involved in intracellular signal transduction. The time course changes in the epidermal growth factor-induced phosphorylation levels of Shc and MAPK in A431 cells were visualized as highly up-shifted migration bands by subsequent immunoblotting with anti-Shc and anti-MAPK antibodies. The third application is in vitro kinase inhibition profiling for the quantitative screening of kinase-specific inhibitors. The inhibition profile of a tyrosine kinase, Abl (a histidine-tagged recombinant mouse Abl kinase), was determined using the substrate Abltide-GST (a fusion protein consisting of a specific substrate peptide for Abl and glutathione S-transferase) and the approved drug Glivec (an ATP competitor). In the kinase assay, the slower migration band, monophosphorylated Abltide-GST, increased time-dependently, whereas the faster migration band, nonphosphorylated Abltide-GST, decreased. The dose-dependent inhibition of Glivec was determined by a change in the ratio of the faster and slower migration bands, which showed an IC50 value of 1.6 microM in the presence of 0.10 mM ATP.

Cell Line, Tumor↗

PubMed2004

HIV/STD risk behaviors and perceptions among rural-to-urban migrants in China.

Data from 2,153 sexually active rural-to-urban migrants in China were analyzed to examine the relationship between the movement of rural-to-urban migration and increased HIV/STD (sexually transmitted disease) risk and the applicability of constructs of a Western-based theory of behavioral change to the study population. Measurements included migrant mobility, sexual risk, and the seven constructs of the protection motivation theory (PMT). Data in the current study suggest that high mobility among rural-to-urban migrants was associated with increased sexual risk. The PMT constructs are applicable in identifying perceptions and attitudes associated with sexual risk behaviors in this culturally distinct population. Increased sexual risk was associated with increased perceptions of extrinsic rewards, intrinsic rewards, and response cost. Also consistent with PMT, increased sexual risk was associated with perceptions of decreased severity, vulnerability, response efficacy, and self-efficacy. After controlling for a number of key confounding factors, all seven PMT constructs were associated with sexual risk in the manner posited by the theory. The association between mobility and sexual risk underscores the importance of effective HIV/STD prevention efforts among this vulnerable population. The social cognitive theories including the PMT may form a logical base for prevention intervention programs targeting rural-to-urban migrants in China.

Adult↗

PubMed2002

Avoiding ischemia in latissimus dorsi muscle grafts: electrical prestimulation versus vascular delay.

BACKGROUND: Surgical mobilization of the latissimus dorsi muscle produces regional ischemic damage that may compromise its function in clinical applications such as cardiomyoplasty. We compared the effectiveness of two procedures designed to maintain blood flow throughout the mobilized muscle. METHODS: Adult pigs were assigned to two experimental groups: an electrically prestimulated group (n = 10) and a vascular delay group (n = 10). In the prestimulated group the left latissimus dorsi muscle was activated in situ at 2 Hz for 24 h/d. In the vascular delay group, the intercostal perforating arteries to the left latissimus dorsi muscle were divided. Two weeks later, hyperemic blood flow was measured by means of fluorescent microspheres immediately before and after mobilizing the latissimus dorsi muscle and again after recovery for a further 2 days. RESULTS: In the prestimulated group, blood flow was not significantly depressed in any region of the muscle immediately after mobilization, and blood flow increased significantly in proximal (p = 0.01), middle (p = 0.02), and distal (p = 0.007) regions following recovery. In muscles subjected to vascular delay the proximal and middle regions showed no significant changes in blood flow after mobilization or recovery, but flow in the distal region was 50% lower after mobilization (p = 0.003), and it remained significantly depressed even after recovery (p = 0.008). CONCLUSIONS: Prestimulation was significantly more effective than vascular delay in preserving distal blood flow. Because it is also less invasive and initiates metabolic transformation before mobilization, this technique should allow cardiac assistance to be introduced at an earlier postoperative stage without compromising the viability of the grafted muscle.

Animals↗

PubMed2006

Self-organization of spiking neural network that generates autonomous behavior in a real mobile robot.

In this paper, we propose self-organization algorithm of spiking neural network (SNN) applicable to autonomous robot for generation of adoptive and goal-directed behavior. First, we formulated a SNN model whose inputs and outputs were analog and the hidden unites are interconnected each other. Next, we implemented it into a miniature mobile robot Khepera. In order to see whether or not a solution(s) for the given task(s) exists with the SNN, the robot was evolved with the genetic algorithm in the environment. The robot acquired the obstacle avoidance and navigation task successfully, exhibiting the presence of the solution. After that, a self-organization algorithm based on a use-dependent synaptic potentiation and depotentiation at synapses of input layer to hidden layer and of hidden layer to output layer was formulated and implemented into the robot. In the environment, the robot incrementally organized the network and the given tasks were successfully performed. The time needed to acquire the desired adoptive and goal-directed behavior using the proposed self-organization method was much less than that with the genetic evolution, approximately one fifth.

Action Potentials↗

PubMed1996

Retroviral transduction of human progenitor cells: use of granulocyte colony-stimulating factor plus stem cell factor to mobilize progenitor cells in vivo and stimulation by Flt3/Flk-2 ligand in vitro.

The clinical application of gene transfer is hindered by the availability of the multipotential stem cells and the difficulty in obtaining efficient retroviral transduction. To assess potential means by which gene transfer into human hemopoietic stem cells might be enhanced, the retroviral transduction efficiency of human bone marrow cells (BM) or peripheral blood progenitor cells (PBPC) was compared at multiple time points after in vivo administration of granulocyte colony-stimulating factor (G-CSF). This was further compared with the transduction efficiency of cells mobilized with G-CSF plus stem cell factor (SCF) in a cohort of patients randomized to receive either one or two growth factors and with normal BM function. Using the LNL6 retrovirus, retroviral transduction efficiencies of up to 19% were observed for both PBPC and BM (n = 26 patients). There was at least a 100-fold increase in PBPC with G-CSF alone and a further 30-fold increase in the total number of progenitor cells available for retroviral transduction using the combination of SCF plus G-CSF. However, pretreatment of patients with G-CSF with or without SCF did not enhance the retroviral infectability of growth factor-mobilized progenitor cells. The effect of the growth factor, Flk-2/Flt3 ligand (FL), was also examined with respect to retroviral transduction efficiency of human progenitor cells. FL plus IL-3 in vitro increased the retroviral transduction efficiency up to eightfold compared with results observed using other combinations of cytokines tested (P < .001). These findings have clinical implications both for increasing the number of target cells for in vivo gene-marking/gene-therapy studies and improving the efficiency of gene transfer.

Cell Cycle↗

PubMed2005

Accelerometers in rehabilitation medicine for older adults.

Recent technological developments have led to the production of inexpensive, miniature accelerometer sensors with potential for use in a clinical setting. These sensors can provide reliable information on mobility and objective measurement of gait. They are currently used mainly in a research setting; however, with recent advances, incorporation into clinical practice is possible. For illustrative purposes this paper describes some current applications of accelerometers in gait and balance evaluation, falls risk assessment and mobility monitoring. Accelerometers provide quantitative measures of gait, they are capable of identifying specific gait changes in older adults and in fallers and can be used to objectively quantify ambulatory activity levels. Accelerometers have many potential uses in monitoring of patients in rehabilitation. They provide an added objective and quantitative dimension to gait analysis when combined with clinical assessment. They have the potential in the future to stratify falls risk facilitating early initiation of appropriate therapeutic intervention, thus reducing further falls. The challenge facing clinicians and biomedical engineers is to further harness this technology making it part of everyday clinical practice.

Acceleration↗

PubMed1986

Noninvasive xenon-133 measurements of cerebral blood flow using stationary detectors compared with dynamic emission tomography.

Repeated bedside measurements of CBF have been made possible by the recent development of a mobile unit with 10 stationary detectors using the intravenous xenon-133 method. To evaluate this technique, comparative CBF studies at rest and following the application of a cerebral vasodilatory stimulus (acetazolamide, 1 g i.v.) were performed with the mobile equipment and with xenon-133 single-photon emission inhalation tomography in patients with cerebrovascular disease. The CBF level and the flow response to acetazolamide as determined with the two methods were well correlated, although at low flow levels the stationary detectors yielded somewhat lower CBF values than did emission tomography. Considering the side-to-side asymmetry, an excellent correlation was obtained. Using the initial slope index, the stationary detectors revealed quantitatively 83% of the interhemispheric asymmetry and 63% of the asymmetry in the middle cerebral artery territory shown with the tomograph. As illustrated by a case history, the nontomographic CBF unit used in this study may provide reliable and useful information in patients with occlusive cerebrovascular disease by performing repeated CBF studies and challenging the cerebral circulation with acetazolamide.

Adult↗

PubMed1999

High level of retrovirus-mediated gene transfer into dendritic cells derived from cord blood and mobilized peripheral blood CD34+ cells.

Dendritic cells (DCs), the most potent antigen-presenting cells, can be generated from CD34+ hematopoietic stem cells and used for generating therapeutic immune responses. To develop immunotherapy protocols based on genetically modified DCs, we have investigated the conditions for high-level transduction of a large amount of CD34+-derived DCs. Thus, we have used an efficient and clinically applicable protocol for the retroviral transduction of cord blood (CB) or mobilized peripheral blood (MPB) CD34+ cells based on infection with gibbon ape leukemia virus (GALV)-pseudotyped retroviral vectors carrying the nls-LacZ reporter gene. Infected cells have been subsequently cultured under conditions allowing their dendritic differentiation. The results show that using a growth factor combination including granulocyte-macrophage colony-stimulating factor plus tumor necrosis factor alpha plus interleukin 4 plus stem cell factor plus Flt3 ligand, more than 70% of DCs derived from CB or MPB CD34+ cells can be transduced. Semiquantitative PCR indicates that at least two proviral copies per cell were detected. Transduced DCs retain normal immunophenotype and potent T cell stimulatory capacity. Finally, by using a semisolid methylcellulose assay for dendritic progenitors (CFU-DCs), we show that more than 90% of CFU-DCs can be transduced. Such a highly efficient retrovirus-mediated gene transfer into CD34+-derived DCs makes it possible to envision the use of this methodology in clinical trials.

Antigens, CD34↗

PubMed2005

alpha-lytic protease can exist in two separately stable conformations with different His57 mobilities and catalytic activities.

alpha-Lytic protease is a bacterial serine protease widely studied as a model system of enzyme catalysis. Here we report that lyophilization induces a structural change in the enzyme that is not reversed by redissolution in water. The structural change reduces the mobility of the active-site histidine residue and the catalytic activity of the enzyme. The application of mild pressure to solutions of the altered enzyme reverses the lyophilization-induced structural change and restores the mobility of the histidine residue and the enzyme's catalytic activity. This effect of lyophilization permits a unique opportunity for investigating the relationship between histidine ring dynamics and catalytic activity. The results demonstrate that His57 in resting enzymes is more mobile than previously thought, especially when protonated. The histidine motion and its correlation to enzyme activity lend support to the reaction-driven ring flip hypothesis.

Catalysis↗

PubMed2001

Spleen enlargement in healthy donors during G-CSF mobilization of PBPCs.

BACKGROUND: Recombinant human G-CSF is widely used to mobilize PBPCs in healthy donors for allogeneic transplantation. There have been concerns about donor safety because of splenic ruptures during G-CSF application. To address this problem, changes in splenic size in 91 healthy donors during G-CSF mobilization of allogeneic PBPCs were investigated. STUDY DESIGN AND METHODS: For mobilization, G-CSF in a dosage of 7.5 microg per kg per day was administered for 5 days and PBPC collection started Day 5. Splenic size was determined by ultrasound before G-CSF application was started and on the day of the first apheresis. RESULTS: The mean increase in splenic length was 11 mm (range, 0-28 mm; p<0.0001), whereas a mean increase of 5 mm in width (range, 0-14 mm; p<0.0001) was measured. No major side effects could be observed. There was no significant correlation between the increase in splenic size and the hematologic values, or the age and body-mass index. In a multivariant analysis, no independent risk factor for the development of a spleen enlargement over 19 mm in length and 9 mm in thickness was found in 20 percent of investigated donors. CONCLUSION: In this prospective trial, a significant spleen enlargement was observed in healthy donors during G-CSF mobilization of allogeneic PBPCs. Further investigations are needed to define the degree of spleen enlargement with higher G-CSF dosages to improve donor safety.

Blood Donors↗

PubMed2005

E-cadherin tethered to micropatterned supported lipid bilayers as a model for cell adhesion.

Cell-cell adhesion is a dynamic process requiring recruitment, binding, and reorganization of signaling proteins in the plane of the plasma membrane. Here, we describe a new system for investigating how this lateral mobility influences cadherin-based cell signaling. This model is based on tethering of a GPI-modified E-cadherin protein (hEFG) to a supported lipid bilayer. In this report, membrane microfluidics and micropatterning techniques are used to adopt this tethered protein system for studies with the anchorage-dependent cells. As directly formed from proteoliposomes, hEFG exhibits a diffusion coefficient of 0.6 +/- 0.3 microm(2)/s and mobile fraction of 30-60%. Lateral structuring of the supported lipid bilayer is used to isolate mobile proteins from this mixed mobile/immobile population, and should be widely applicable to other proteins. MCF-7 cells seeded onto hEFG-containing bilayers recognize and cluster this protein, but do not exhibit cell spreading required for survival. By micropatterning small anchors into the supported lipid bilayer, we have achieved cell spreading across the bilayer surface and concurrent interaction with mobile hEFG protein. Together, these techniques will allow more detailed analysis of the cellular dynamics involved in cadherin-dependent adhesion events.

Cadherins↗

PubMed2000

Validity of HAMIS: a test of hand mobility in scleroderma.

OBJECTIVE: Hand Mobility in Scleroderma (HAMIS) is a hand function test for persons who have systemic sclerosis (scleroderma). The purpose of HAMIS is to obtain an estimation of the hand mobility that is precise enough to detect limitation of motion at the same time as it indicates the ability to use the hand in daily occupations. The aim of this study was to test psychometric properties of the HAMIS, and the following aspects of HAMIS were examined: 1) the applicability of HAMIS, 2) concurrent validity, and 3) discriminating ability. METHODS: Forty-five patients with scleroderma were assessed for range of motion (ROM), HAMIS, and skin thickness. In addition, 15 healthy individuals completed HAMIS. RESULTS: The applicability of HAMIS was good for items assessing finger and thumb mobility and moderate for items assessing mobility of the wrist and the forearm. The relationships of HAMIS to ROM and skin score were statistically significant for all items except for pronation and supination of the forearm. There were also statistically significant differences between the patients and the healthy individuals for all items except these two. CONCLUSION: HAMIS has a demonstrated concurrent validity compared with ROM and skin score, and it showed a good ability to discriminate between healthy individuals and persons with scleroderma, although a lack of variation in the items measuring pronation and supination inferred worse psychometric properties for these two items.

Activities of Daily Living↗

PubMed1992

Concave Ferguson plots of DNA fragments and convex Ferguson plots of bacteriophages: evaluation of molecular and fiber properties, using desktop computers.

A desktop computer program evaluating physical properties of DNA and bacteriophages is presented. The analysis is based on data obtained from capillary and submarine-type agarose electrophoresis. Native molecular/particle properties and properties of the gel (or polymer) medium can be derived from electrophoresis at several gel concentrations. This is done conveniently by a computerized evaluation of the semi-logarithmic plot of mobility vs. gel concentration, designated the Ferguson plot. In application to most proteins, this plot is linear and computer programs exist to evaluate it. However, nonlinear Ferguson plots have assumed great importance in view of the fact that the plots are concave for DNA. Similarly, convex plots are important since they prevail in the electrophoresis of large particles in agarose. The computer program reported here is the first to (i) address concave Ferguson plots and (ii) allow for the evaluation of both cases using a desktop computer. Program ELPHOFIT version 2.0, a Macintosh application, is available upon request.

Bacteriophages↗

PubMed1995

Healing determinants in older people with leg ulcers.

A descriptive correlational design was used to identify the physiologic, therapeutic, and psychosocial determinants of leg ulcer healing in community-residing older people with venous and venous-arterial disease (N = 156). A regression model with healing rate as the dependent variable explained 49% of the variance. Increased pain when mobilizing, increased hours with limbs horizontal to the torso, and moderate and severe liposclerosis (hardening and induration of the skin) were associated with poorer healing rates. Higher wound status scores were associated with more rapid healing rates. Self-efficacy beliefs and social support were not significant factors. The findings suggest the need for early detection and management of limb pain that interferes with normal mobility. Limb position and edema assessment prior to the application of compression bandages is recommended, with bandages that provide clients with high compression when ambulant and low compression when resting being potentially beneficial.

Aged↗