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At least 145 records · Page 8Linked to original sources
PubMed1985

Evidence for two calcium-dependent steps and a sodium-dependent step in the mechanism of cell killing by calcium ions in the presence of ionophore A23187.

Elevated intracellular Ca2+ appears to play an important role in the mechanism of cell killing in certain pathologic states such as ischemia. The authors have examined aspects of the biochemical mechanism of cell killing by elevated intracellular Ca2+ using as a model system cultured fibroblasts treated with ionophore A23187 in Ca2+-containing medium. Evidence has been obtained for two Ca2+-mediated steps and a Na+-mediated step in the cell killing process. The first Ca2+-mediated step occurs in low extracellular Ca2+ concentrations (1-100 microM) and exhibits a variety of characteristics in common with the arachidonic acid release response stimulated under the same conditions. These results are consistent with the arachidonic acid release response constituting or closely monitoring the initial injury process. The second Ca2+-mediated process is achieved at near physiologic extracellular Ca2+ concentrations in the absence of A23187. Killing of cells injured by the two Ca2+-dependent steps requires extracellular Na+ ions at half or more the physiologic concentration.

Animals↗
PubMed1977

Preservation of intraerythrocytic forms of malarial parasites by one-step and two-step cooling procedures.

Ring, trophozoite, and schizont stages of Plasmodium knowlesi were cooled in dimethyl sulfoxide either by direct immersion in liquid nitrogen or by a two-step method in which the cells were held at temperatures slightly below 0 degrees C for different lengths of time before they were cooled to -196 degrees C. After the direct plunge treatment, thawed trophozoites and schizonts were found to be extensively damaged. Their survival was markedly increased by holding them at -31 degrees C for 30 min before plunging them into liquid nitrogen. Freeze-substitution showed that cells cooled by the two-step procedure were grossly shrunken and had relatively few intracellular ice cavities. Large amounts of ice formed in trophozoites and schizonts preserved by direct immersion in liquid nitrogen. The two-step protocols investigated did not improve the survival of ring-stage parasites, 25-50% of which survived rapid cooling to -196 degrees C. Infected cell agglutination tests were carried out with frozen and thawed schizonts. Variant specificity was demonstrated with cells that had been plunged directly to -196 degrees C, but cells cooled by the two-step method tended to agglutinate spontaneously.

Animals↗
PubMed1993

The real-time resolution of proton-related transient-state steps in an enzymatic reaction. The early steps in the oxidative deamination reaction of bovine liver glutamate dehydrogenase.

We introduce a novel transient-state kinetic approach which can resolve proton and product time courses into a series of individual steps that comprise the reaction path. We have applied this approach to the oxidative deamination reaction catalyzed by bovine liver glutamate dehydrogenase, measuring both the product (NADPH) and proton time courses at various pH values. The global treatment (over all pH values) resolves the very early portion of this reaction quantitatively and provides a continuous time course for each of the six protonic species. We propose the following mechanism: L-glutamate binds to an open conformation of the enzyme-NADP complex, forming salt bridges between its alpha- and gamma-carboxyl groups and the protonated forms of enzyme lysine residues 114 and 90, respectively. In this position, the alpha-H atom of the substrate is too far from the nicotinamide ring for hydride transfer to occur. In the next step, three events occur in a concerted manner: lysine 126 loses a proton and acquires a single water molecule; the active site cleft closes; bulk water is expelled; the substrate and coenzyme are forced closer together and remain in a nonaqueous environment during the ensuing chemical events, returning to an open conformation only in time to allow the product release steps to occur. Thus, substrate binding accomplishes a number of important tasks which are themselves an integral part of the catalytic mechanism. Combining the novel transient state approach developed here with steady-state kinetic information can produce a detailed mechanistic resolution of otherwise hidden steps.

Amino Acid Sequence↗
PubMed1997

Value of a therapeutic trial to diagnose gastroesophageal reflux disease: step up versus step down therapy.

The 1992 Canadian Association of Gastroenterology consensus conference on gastroesophageal reflux disease (GERD) recommended a therapeutic trial of H2 receptor antagonists as a cost effective approach to the management of patients with typical symptoms of GERD. Omeprazole, with its increased potency and efficacy of acid suppression, appears to be the ideal diagnostic test therapy for GERD. However, there is little evidence in the literature to support this approach. Omeprazole has the potential to mask other disorders such as peptic ulcer disease, thereby delaying appropriate testing for and eradication of Helicobacter pylori. Therefore, omeprazole therapeutic trials should be used with caution in the diagnosis of GERD. The conventional step up therapy is the least costly and reasonably effective approach to treat the majority of patients with mild to moderate symptoms of GERD. For patients who fail therapy or have complications of GERD, the recently proposed step down therapy is probably more effective and appropriate. As physicians become more comfortable with the long term use of proton pump inhibitors, step down therapy may well replace the more conservative step up approach to therapy.

Gastroesophageal Reflux↗
PubMed2004

Ultramorphological analysis of resin-dentin interfaces produced with water-based single-step and two-step adhesives: nanoleakage expression.

This study evaluated the nanoleakage patterns in bonded interfaces using two single-step, self-etching adhesives (Adper Prompt-AD, and One-up Bond F-OB), two two-step, self-etching primers (Clearfil SE Bond-CF, and Unifil Bond-UB), and one two-step, total-etch adhesive (Single Bond-SB). Dentin surfaces were bonded with the adhesive systems and stored in water at 37 degrees C for 1 week and 6 months. After storage periods, teeth were sectioned into 0.8 mm-thick slabs, coated with nail varnish except for the bonded interfaces, and immersed in ammoniacal AgNO(3) for 24 h. After immersion in photodeveloping solution, bonded sections were prepared and observed under a SEM using the backscattered electron mode. Undemineralized, unstained, epoxy resin-embedded sections were prepared for TEM. Nanoleakage patterns were qualitatively compared between periods. Nanoleakage was observed in all bonded specimens at both periods. CF and UB presented silver deposits predominantly restricted to the thin (0.5 microm) hybrid layer (HL) at both periods. Although no evident differences were observed in the nanoleakage pattern of UB at 7 days and 6 months, CF presented enlarged areas of silver impregnation after 6 months. SB presented accumulation of silver particles mostly within the HL at 7 days, which was intensified after 6 months. AD and OB presented massive silver accumulation within the HL and the overlying adhesive layer. No evident differences were noticed between storage periods. Silver impregnation increased for all adhesive systems from 7 days to 6 months, except for UB.

Adhesives↗
PubMed2002

fMRI retinotopic mapping--step by step.

fMRI retinotopic mapping provides detailed information about the correspondence between the visual field and its cortical representation in the individual subject. Besides providing for the possibility of unambiguously localizing functional imaging data with respect to the functional architecture of the visual system, it is a powerful tool for the investigation of retinotopic properties of visual areas in the healthy and impaired brain. fMRI retinotopic mapping differs conceptually from a more traditional volume-based, block-type, or event-related analysis, in terms of both the surface-based analysis of the data and the phase-encoded paradigm. Several methodological works related to fMRI retinotopic mapping have been published. However, a detailed description of all the methods involved, discussing the steps from stimulus design to the processing of phase data on the surface, is still missing. We describe here step by step our methodology for the complete processing chain. Besides reusing methods proposed by other researchers in the field, we introduce original ones: improved stimuli for the mapping of polar angle retinotopy, a method of assigning volume-based functional data to the surface, and a way of weighting phase information optimally to account for the SNR obtained locally. To assess the robustness of these methods we present a study performed on three subjects, demonstrating the reproducibility of the delineation of low order visual areas.

Algorithms↗
PubMed1985

Time trends and key factors in the choice of one-step or two-step biopsy and surgery for breast cancer.

The relationship between type of biopsy-mastectomy procedure and four sets of independent variables (physician, patient, hospital and tumor characteristics) was examined for 993 locally-staged breast cancer patients diagnosed between 1974 and 1981 in 13 counties of western Washington State. Time trends were also investigated. Cases were selected from records of a population-based cancer registry. The frequency of the two-step procedure, in which biopsy and surgery occur on different days, increased from 28 to 57% during the 8-year study period. Use of the two-step procedure was associated with younger age of the patient, less suspicious symptoms or mammogram results, younger physician cohorts and hospitals with government and health maintenance organization proprietorship. These relationships remained unchanged after controlling for potentially confounding variables. There were also substantial differences between individual hospitals in the frequency of the two-step procedure, suggesting differing schools of thought in different locales.

Adult↗
PubMed1995

A unified model for cell-killing by heat: interpretation of continuous, step-down, step-up and split heating.

We have already proposed a model for the lethal effects of heat on cells. In the model, a three-step process for cellular inactivation was hypothesized: 1) heating produces sublethal damage; 2) the damage is repaired in a certain time period; 3) the cell is inactivated when it undergoes cumulative sublethal damage to the extent that it cannot be repaired. Using the previous model, we investigated the modifications of the survival curve in 1) continuous heating, 2) step-down heating, 3) step-up heating, and 4) recovery from the thermal injury by split heating. These four phenomena were explained well with the mathematical model.

Animals↗
PubMed2003

Step-by-step mark-up of medical guideline documents.

Approaches to formalization of medical guidelines can be divided into model-centric and document-centric. While model-centric approaches dominate in the development of clinical decision support applications, document-centric, mark-up-based formalization is suitable for application tasks requiring the 'literal' content of the document to be transferred into the formal model. Examples of such tasks are logical verification of the document or compliance analysis of health records. The quality and efficiency of document-centric formalization can be improved using a decomposition of the whole process into several explicit steps. We present a methodology and software tool supporting the step-by-step formalization process. The knowledge elements can be marked up in the source text, refined to a tree structure with increasing level of detail, rearranged into an XML knowledge base, and, finally, exported into the operational representation. User-definable transformation rules enable to automate a large part of the process. The approach is being tested in the domain of cardiology. For parts of the WHO/ISH Guidelines for Hypertension, the process has been carried out through all the stages, to the form of executable application, generated automatically from the XML knowledge base.

Artificial Intelligence↗
PubMed2002

Total synthesis of (+)-alpha-onocerin in four steps via four-component coupling and tetracyclization steps.

A remarkably short (four steps, 31% overall yield) enantioselective synthesis of the structurally unique C2-symmetric tetracyclic triterpene (+)-alpha-onocerin (1) has been developed. The brevity of this mechanism depends on the assembly of four fragments (two molecules of a chiral epoxy ketone and two molecules of vinyllithium) to generate the chiral bis-epoxide 4 in one step, and on the efficient formation of all four carbocyclic rings in one step by cation-olefin tetracyclization. New and general methodology for the conversion of vinyl tert-butyldimethylsilyl ethers to vinyl triflates with retention of E or Z olefinic geometry also was utilized in the synthesis of 1. A short enantioselective synthesis of a non-C2-symmetric diastereomer of 1 is also described which uses the new methodology.

Cyclization↗
PubMed2002

Functional assembly of the foreign carotenoid lycopene into the photosynthetic apparatus of Rhodobacter sphaeroides, achieved by replacement of the native 3-step phytoene desaturase with its 4-step counterpart from Erwinia herbicola.

Photosynthetic organisms synthesize a diverse range of carotenoids. These pigments are important for the assembly, function and stability of photosynthetic pigment-protein complexes, and they are used to quench harmful radicals. The photosynthetic bacterium Rhodobacter sphaeroides was used as a model system to explore the origin of carotenoid diversity. Replacing the native 3-step phytoene desaturase (CrtI) with the 4-step enzyme from Erwinia herbicola results in significant flux down the spirilloxanthin pathway for the first time in Rb. sphaeroides. In Rb. sphaeroides, the completion of four desaturations to lycopene by the Erwinia CrtI appears to require the absence of CrtC and, in a crtC background, even the native 3-step enzyme can synthesize a significant amount (13%) of lycopene, in addition to the expected neurosporene. We suggest that the CrtC hydroxylase can intervene in the sequence of reactions catalyzed by phytoene desaturase. We investigated the properties of the lycopene-synthesizing strain of Rb. sphaeroides. In the LH2 light-harvesting complex, lycopene transfers absorbed light energy to the bacteriochlorophylls with an efficiency of 54%, which compares favourably with other LH2 complexes that contain carotenoids with 11 conjugated double bonds. Thus, lycopene can join the assembly pathway for photosynthetic complexes in Rb. sphaeroides, and can perform its role as an energy donor to bacteriochlorophylls.

Antioxidants↗
PubMed1997

P-R-A-C-T-I-C-A-L: a step-by-step model for conducting the consultation in general practice.

BACKGROUND: It has been shown that when patients are unable to express all their major concerns, they are less likely to follow the physician's prescribed treatment plan and they are less satisfied. On the other hand, the GP has a limited amount of time to elicit all the appropriate information and must ask certain questions about the biological aspects of the illness in order to carry out her professional responsibilities. By acting in a patient-centred way, first enabling the patient to express himself, the GP can make maximum use of patients' ability for problem formulation and solution. METHODS: We describe a model, for which the mnemonic, P-R-A-C-T-I-C-A-L, will help the practitioner to remember its nine steps. The model uses a chronological succession of strategies during the consultation that balances the voices of medicine and the lifeworld. In overview, the GP takes the patient, step by step, first through an exploration and clarification of his views of the illness, then expands the problem by further examination (e.g. the physical examination), a negotiation about the final model of the illness that includes both diagnosis and management, a discussion of the treatment plan, and finally a moment of reflection to prepare for the next visit.

Abbreviations as Topic↗
PubMed1992

Comparison of a one-step and a two-step polymerase chain reaction with degenerate general primers in a population-based study of human papillomavirus infection in young Swedish women.

The prevalence of human papillomavirus (HPV) infection in cervical cell scrapes from young women was determined by polymerase chain reaction (PCR) by using general primer pairs localized within the L1 region. With a one-step general PCR, 5.9% (35 of 590) of young women in a population-based study were found to contain HPV DNA. The proportion of HPV-positive women increased with age, from 1.4% (1 of 69) among women aged 19 years to 9.2% (13 of 142) among women aged 25 years. Among the cervical scrapes from women with normal cytology, 5.6% (30 of 539) harbored HPV DNA. A total of 5 of 19 (26.3%) of the women with pathological signs were positive for HPV DNA. By a two-step PCR, using nested general primers, 20.3% (118 of 581) of all women were shown to contain HPV DNA. The proportion of HPV-positive women also increased with age, from 17.4% (12 of 69) among women aged 19 years to 31.9% (43 of 135) among women aged 25 years, when the two-step PCR was used. Some 19.2% (102 of 530) of cervical scrapes from women with normal cytology contained HPV DNA. Among the women with pathological signs, 16 of 19 (84.2%) were positive for HPV DNA. The HPV DNA-positive specimens were demonstrated to contain HPV type 6, 11, 16, 18, 31, 33, 35, 39, 40, 45, 55, or 56. The most prevalent HPV types were 6 (2.0%) and 16 (2.7%). More than one type was found in 16 specimens. Sixty HPV-positive samples could not be typed.

Adult↗
PubMed1973

Replication of bacteriophage SH-133 in the facultative autotroph Hydrogenomonas facilis. II. Bacteriophage synthesis and hydrogenase induction under step-up and step-down growth conditions.

Autotrophically grown infected cells are able to replicate phage SH-133 after being switched to a heterotrophic environment (step-up growth). The effect of step-down growth on phage replication varies with the choice of organic substrate. Phage replication and the induction of cellular hydrogenase occur under step-down growth from acetate but not peptone broth. The requirement of a continued source of energy for phage replication in either heterotrophically or autotrophically grown cells could be uniquely demonstrated in this phage-host system by the deletion of hydrogen from the growth medium.

Bacteriophages↗
PubMed1975

A comparative study of horseradish peroxidase conjugates prepared with a one-step and a two-step method.

In this study we compared horseradish peroxidase (HRP)-labeled rabbit antihuman immunoglobulin G (IgG) conjugates, prepared by a one-step and a two-step method. Glutaraldehyde was used as a cross-linking agent. Two methods were used for removing unconjugated HRP: Sephadex G-200 gel chromatography and ammonium sulfate precipitation. The conjugates were characterized immunologically, immunochemically and enzymatically. The immunohistoenzymic properties of the conjugates were tested on unfixed cryostat sections of the skin of patients with chronic discoid lupus erythematosus. The influence of the presence of unconjugated HRP and unconjugated IgG was studied. Optimal results were obtained with conjugates prepared by a two-step method. Removing unconjugated HRPimproved the immunohistoenzymic properties of the conjugates. Conjugated and unconjugated IgG could be separated by Sephadex G-200 gel chromatography.

Animals↗
PubMed1986

Nucleoside diphosphokinase, an enzyme with step changes in activity during the cell cycle of the fission yeast Schizosaccharomyces pombe. I. Persistence of steps after a block to the DNA-division cycle.

In confirmation of earlier results, nucleoside diphosphokinase is shown to be a 'step' enzyme in Schizosaccharomyces pombe with a sharp doubling in activity at the beginning of the cell cycle. These doubling steps occur at the same time in the cycle in the smaller cells of the mutant wee1.6. An important result is that the activity steps persist with normal cell cycle timing after a block to the DNA-division cycle imposed by the cycle mutants cdc2.33 and cdc2.33wee1.6. This is clear proof that oscillatory controls of some cell cycle events can persist after the main periodic events of the DNA-division cycle have been abolished.

Cell Cycle↗
PubMedDate not supplied

Climbing a ladder: a step-by-step approach to understanding the concept of agroecosystem health.

Population and individual health is linked to agroecosystem health. To comprehend the concept of agroecosystem health, one should climb a ladder consisting of several successive steps, each rung presenting a certain degree of instability (conceptual difficulty and uncertainty) in an advisable but not inevitable order. Here we suggest a ladder consisting of the following concepts: ecosystem, agroecosystem, biodiversity, sustainability, ecosystem health, and agroecosystem health. Although these concepts are to a certain extent well understood and grasped by scientists, politicians, natural resource managers, and environmentalists, some steps are still highly debatable, unclear, and present a considerable degree of reluctance to be defined and understood. Consequently, much empirical and theoretical effort must be made to construct solid conceptual ladders made up of such steps. In this enterprise, a traditional reductionistic approach confining interpretations to narrow scientific disciplines is unadvisable. Holistic, transdisciplinary approaches are required to reach the desired goal.

Agriculture↗
PubMed2004

Electroporation of murine embryonic stem cells: a step-by-step guide.

The manipulation of embryonic stem (ES) cells to generate targeted mutations via homologous recombination has proved an invaluable resource for researchers from fields as diverse as embryology, immunology, physiology, and biophysics. The derivation and culture of these cells has been described in many places, but there is no substitute for hands-on experience. In lieu of that, here we provide a step-by-step guide for the technique of electroporation, complete with micrographs of the cells at each step of the process. This is intended to allow the first time ES cell worker, as well as the more experienced researcher, to view the morphology of ES cells under ideal conditions as well as when the cells are suboptimal and should be discarded.

Animals↗