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At least 811 records · Page 45Linked to original sources

PubMed1996

CREB-1 and CREB-2 immunoreactivity in the rat brain.

This study is focused to learn about the cellular localization of transcription factors binding to the cAMP response element-CREBs-in the brain of normal rats and in animals subjected to excitotoxic cell damage. For this purpose, CREB-1 and CREB-2 immunoreactivity is examined in the developing and adult rat brain under physiological conditions, and following systemic kainic acid (KA) injection at convulsant doses in the adult, as a validated experimental model of excitotoxic injury. CREB-1 immunoreactivity is constitutively expressed in periventricular glia and Bergmann glia, and appears in reactive astrocytes following KA-induced excitotoxic cell damage. In contrast, CREB-2 is constitutively expressed in all neurons of the cerebrum, cerebellum and brain stem in the developing and adult brain. CREB-2 immunoreactivity is not increased following KA excitotoxic cell damage. These results demonstrate that CREB-1 and CREB-2 in the brain of the rat are localized in separate cellular compartments and that their expression is differentially regulated in pathologic states.

Animals↗

PubMed1984

An improved method for determination of active and total renin concentration in human plasma using an excess of sheep substrate.

A method for measurement of renin concentration (PRC) in human plasma has been developed and validated experimentally and theoretically. Like most of the previous methods, the present method is based on a radioimmunoassay of angiotensin I generated during incubation of plasma with an excess of renin substrate. In the present study we used, as the substrate, sheep angiotensinogen partially purified from anephric sheep plasma by ammonium sulfate fractionation and pepstatin-aminohexyl-agarose chromatography. The advantage of using sheep substrate is that it has an exceptionally high affinity for human renin. The partially purified substrate, which contained no detectable renin activity, improved the sensitivity, allowing quantification of renin in low-renin plasma. Possible interfering influences of plasma proteins on the radioimmunoassay were eliminated by the introduction of a simple deproteinization step. For determination of total renin concentration (TRC), inactive renin was activated by exposing plasma to low pH or trypsin. Normal values of PRC and TRC after careful selection of assay conditions were 2.9 +/- 0.4 and 33.9 +/- 6.1 ng angiotensin I X ml-1 X h-1, respectively. As an illustrative example of usefulness of the assay, PRC determination of a patient with a ectopic renin-secreting tumor is presented.

Adult↗

PubMed1989

A large scale, high resolution, automated system for rat sleep staging. I. Methodology and technical aspects.

An automatic rat sleep classification system is described which records and analyses bioelectrical signals from 32 rats over extended periods of time. At present this system is used routinely for the screening of drug effects on sleep. The analysis is based on 3 signals, the parieto-occipital EEG, nuchal EMG and a movement indicator signal. The on-line analysis is done per epoch of 2 sec and involves power spectral analysis of the EEG and rectification and integration of the EMG and movement signals. The automatic sleep staging into 6 stages (active and quiet waking; quiet, deep, pre-REM and REM sleep) is performed off-line. Parameters derived from a discriminant analysis of visually scored tracings of individual rats constitute the basis for the automatic scoring procedure. The movement index is used to discriminate between active and quiet waking, while the use of the EMG level improves the separation of waking and REM sleep. After the construction of hypnograms from these computer scorings a set of parameters can be extracted which characterizes the sleep-waking behavior of each individual rat. These parameters are then used to compare statistically the 2-4 treatment groups which make up each experiment of 32 rats. Experimental validation of the system is reported in an accompanying paper.

Analog-Digital Conversion↗

PubMed1992

Biomechanical consequences of callus development in Hoffmann, Wagner, Orthofix and Ilizarov external fixators.

A theoretical analysis by a finite elements model (FEM) of some external fixators (Hoffmann, Wagner, Orthofix and Ilizarov) was carried out. This study considered a logarithmic progress of callus elastic characteristics. A standard configuration of each fixator was defined where design and application characteristics were modified. A comparison among standard configurations and influence of every variation was made with regard to displacement and load transmission at the fracture site. An experimental evaluation of standard configurations was performed with a testing machine. After experimental validation of the theoretical model was achieved, an application of physiological loads which act on a fractured limb during normal gait was analysed. A minimal contribution from an external fixator to the total rigidity of the bone-callus-fixator system was assessed when a callus showing minimum elastic characteristics had just been established. Insufficient rigidity from the fixation devices to assure an adequate immobilization during the early stages of fracture healing was verified. However, regardless of the external fixator, callus development was the overriding element for the rigidity of the fixator-bone system.

Biomechanical Phenomena↗

PubMed1981

A simplified two-dimensional electrophoretic technique.

The two-dimensional electrophoretic O'Farrell technique can be simplified in terms of effort and equipment. This simplification derives mainly from the fact that the first dimension (isoelectric focusing) is run in a vertical slab gel. Since all tracks are cast in a single template, a reproducible and uniform first dimension is obtained, allowing easy and accurate comparison among tracks after standard staining and/or radioautographic procedures. After the first dimension, the focused tracks are cut and, following equilibration, simply laid on top of the second dimensional slab gel, whose spacers are 0.1 mm thicker than those used in the first separation. Thus, basically the same equipment can be used for both dimensions. This modified technique has been experimentally validated since it has allowed the detection of structural variants of two proteins related to the mouse complement system, namely, the fourth component and the sex-limited protein.

Animals↗

PubMed1994

Typing of the HLA-DRB3 gene by temperature gradient gel electrophoresis. Prediction of the resolution of four allelic fragments by computational simulation of DNA melting.

Four alleles are currently recognised at the HLA-DRB3 locus (DRB3*0101, DRB3*0201, DRB3*0202 and DRB3*0301). We studied whether 271 bp fragments of the polymorphic second exon, which were prepared using the polymerase chain reaction, could be typed using temperature gradient gel electrophoresis. Thermal stability curves for the allelic DNA molecules were calculated by computer simulation and the results were validated experimentally. The DRB3*0201 and DRB3*0202 derived homoduplexes were predicted to have identical thermal stability. Thus, only three denaturation and relative mobility curves were obtained for the four homoduplex fragments DRB3*0101, DRB3*0201, DRB3*0202 and DRB3*0301. Computational analysis predicted that DRB3*0201 and DRB3*0202 could be distinguished by electrophoresis of artificially generated heteroduplexes. When verified experimentally, the results of the theoretical analyses were confirmed. We conclude, that computational simulation of the melting behaviour of DNA molecules permits the resolution of allelic sequences to be predicted by temperature gradient gel electrophoresis. We also demonstrate that temperature gradient gel electrophoresis is a powerful tool for the assessment of HLA genotypes. It may have wide application in transplantation immunology and in the study of disease associations of allelic variation at the HLA loci.

Alleles↗

PubMed1991

Optimal coordination and control of posture and locomotion.

This paper presents a theoretical model of stability and coordination of posture and locomotion, together with algorithms for continuous-time quadratic optimization of motion control. Explicit solutions to the Hamilton-Jacobi equation for optimal control of rigid-body motion are obtained by solving an algebraic matrix equation. The stability is investigated with Lyapunov function theory, and it is shown that global asymptotic stability holds. It is also shown how optimal control and adaptive control may act in concert in the case of unknown or uncertain system parameters. The solution describes motion strategies of minimum effort and variance. The proposed optimal control is formulated to be suitable as a posture and stance model for experimental validation and verification. The combination of adaptive and optimal control makes this algorithm a candidate for coordination and control of functional neuromuscular stimulation as well as of prostheses.

Algorithms↗

PubMed1994

Glucose absorption and production following oral glucose: comparison of compartmental and arteriovenous-difference methods.

Both whole-body models and the regional arteriovenous (AV)-difference method have been used to calculate systemic glucose rates of appearance (Ras) under non-steady-state conditions. Although whole-body models have been experimentally validated in the dog, direct comparison of the whole-body and regional-balance approach has not been made in man. We reanalyzed published data obtained by combining the double-tracer technique ([3H]glucose infusion with ingestion of a [14C]glucose-labeled load) with hepatic vein catheterization. Steele's monocompartmental model underestimated the Ra of oral glucose ([RaO] by 12%, P < .0001) and overestimated the Ra of endogenous glucose ([RaE] by 19%, P < .0001) in comparison to a two-compartment (2-c) model calculation. Splanchnic balance data were used to compute the total glucose Ra (RaT) with either steady-state or non-steady-state equations (the latter by estimating splanchnic transit times). Except for one early time point (15 minutes), the two calculations agreed well with one another. The glucose RaT by the balance method was well correlated with that calculated by the 2-c whole-body model (r = .72, P < .0001 on all data points); however, the former significantly underestimated the latter by 0.62 mg.min-1.kg-1 (or 13%, P < .01) on average throughout the absorptive period. This bias was small in comparison to the estimated random error affecting the calculation of glucose RaT by the model, which averaged 1.4 mg.min-1.kg-1 (corresponding to a variation coefficient of approximately 30%).(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗

PubMed1990

A simple model for prediction of oxygen transport rates by flowing blood in large capillaries.

A simple model has been developed for simulation of oxygen transport to and from blood flowing in conduits of the diameter of arterioles and larger (greater than or equal to 20 microns). The basis is the large capillary model (LCAP) of P.K. Nair, et al. 1989 which has been validated experimentally. Detailed calculations of the oxygen concentration distribution reveal that the dominant resistance to oxygen transport is distributed in the plasma. Relatively little resistance is present within or in the immediate vicinity of the red cells. On the basis of these findings, LCAP was simplified from four simultaneous nonlinear partial differential equations (PDEs) to one PDE by (1) assuming chemical equilibrium within the red blood cells, (2) neglecting intracellular and extracellular boundary layer resistances, and (3) incorporating transport in the cell-free region adjacent to the capillary wall into the boundary conditions. The simplified model is much easier to apply mathematically to new situations. A comparison between LCAP and the simpler model shows that they give virtually the same predictions, and the predictions agree well with experimental measurements. The model is predictive in that all the parameters are determined from the literature or from independent measurements. Thus it should be useful in studies of physiological significance, as well as in design and analysis of extracorporeal blood oxygenators.

Biological Transport, Active↗

PubMed1994

TCR gene segments from at least one third of V alpha subfamilies rearrange at the delta locus.

Using PCR and an experimentally validated V alpha subfamily-specific oligonucleotide panel (V alpha 1-w29), we have investigated whether the TCR delta chain may increase its combinatorial diversity by using V genes considered as alpha chain-specific. We show that at least 10 distinct human V alpha segments rearrange at the J delta locus, leading to scrambling of the two V gene repertoires. Fifty-five per cent of the V alpha/J delta transcripts characterized here were in frame. The 17 V alpha/C delta chains analysed included an extended CDR3 region with up to 18 aa encoded by the junctional region. In addition, a new J delta segment (J delta 4) has been characterized. Together, these findings demonstrate that combinatorial diversity in the human delta locus is larger than previously thought.

Amino Acid Sequence↗

PubMed1991

Innovative developments in isotachophoresis (displacement electrophoresis).

This Mini Review is aimed at characterizing the innovative developments in isotachophoresis (ITP) during the past few years, discussing in turn new theoretical, analytical, preparative and applicative aspects of this unique separation method. Examples given from our laboratory include the study of the detailed dynamics of the ITP separation of four components by computer simulation and experimental validation in a capillary-type instrument with multiple sensors along the separation trough; the anionic ITP analysis in presence of a strong cathodic electroosmotic flow using an open-tubular fused-silica capillary with on-column multiwavelength detection, and the fractionation of proteins in a screen-segmented, rotating column as well as by recycling ITP.

Electrophoresis↗

PubMed1992

Structural analysis of 2',3'-dideoxyinosine, 2',3'-dideoxyadenosine, 2',3'-dideoxyguanosine and 2',3'-dideoxycytidine by 500-MHz 1H-NMR spectroscopy and ab-initio molecular orbital calculations.

Solution structure of anti-AIDS drug, 2',3'-dideoxyinosine (ddI) has been assessed by NMR spectroscopy and pseudorotational analysis in conjunction with its analogues: 2',3'-dideoxyadenosine (ddA), 2',3'-dideoxyguanosine (ddG) and 2',3'-dideoxycytidine (ddC). The absence of 3'-hydroxyl groups in these compounds has prompted us to establish the relationship between proton-proton and corresponding endocyclic torsion angles in the 2',3'-dideoxyribofuranose moiety on the basis of five available crystal structures of 2',3'-dideoxynucleosides. A subsequent pseudorotational analysis on ddI (1), ddA (2), ddG (3) and ddC (4) shows that the twist C2'exo-C3'-endo forms of sugar are overwhelmingly preferred (75-80%) over the C2'-endo envelope forms. The phase angles (P) for North and South conformers with the corresponding puckering amplitude (psi m) for ddI (1), ddA (2) and ddG (3) are as follows: PN = 0.1 degrees, PS = 161 degrees and psi m = 34.1 degrees for ddI (1); PN = 1.4 degrees, PS = 160 degrees and psi m = 34.2 degrees for ddA (2) and PN = 2.4 degrees, PS = 163 degrees and psi m = 33.6 degrees for ddG (3). The predominant North conformer of ddC (4) is intermediate between twist C2'-exo-C3'-endo and C3'-endo envelope (P = 10.9 degrees) with a psi m of 34.7 degrees. Note that these preponderant North-sugar structures (approx. 75-80%) found in the solution studies of ddI (1), ddA (2), dG (3) and ddC (4) are not reflected in the X-ray crystal structures of 2',3'-dideoxyadenosine and 2',3'-dideoxycytidine. The constituent sugar residues in both of these crystal structures denosine and 2',3'-dideoxycytidine. The constituent sugar residues in both of these crystal structures are found to be in the South-type geometry (ddA crystalizes in C3'-exo envelope form, while ddC adopts the form intermediate between the C3'-exo envelope and C3'-endo-C4'-exo twist form). This means that X-ray structures of ddA (2) and ddC (4) only represent the minor conformer of the overall pseudorotamer population in solution. An assumption that the structure of the pentofuranose sugar (i.e. P and psi m) participating in conformational equilibrium described by the two-state model remains unchanged at different temperatures has been experimentally validated by assessing five unknown pseudorotational parameters with eight unique observables (3J1'2', 3J1'2", 3J2'3', 3J2'3", 3J2"3', 3J2"3", 3J3'4' and 3J3"4') for 2',3'-dideoxynucleosides.(ABSTRACT TRUNCATED AT 400 WORDS)

Didanosine↗

PubMed1989

Peristaltic versus syringe pumps for push-pull perfusion: tissue pathology and dopamine recovery in rat neostriatum.

The characteristics of performance were compared for two types of pump, peristaltic and infusion-withdrawal, commonly used for localized push-pull perfusion of brain tissue. Guide cannulae were stereotaxically implanted bilaterally in the caudate-putamen complex of the rat at homologous sites in the coronal plane. Consecutive 5.0 min push-pull perfusions were carried out simultaneously with one pump perfusing ipsilaterally while the other pump perfused the contralateral caudate nucleus. 3H-dopamine (DA) diluted with an artificial CSF was used as the radiotracer and was perfused at a rate of 25 microliters/min. Following the collection of 7-8 samples, the brain was fixed, removed, and histological sections taken through the sites of perfusion. Microscopic examination of the perfusion site showed that the circumscribed lesion at the tip of the cannula connected to either pump was generally indistinguishable. A comparison of the values of 3H-DA recovery in samples obtained with the peristaltic and infusion-withdrawal pumps demonstrated that: (1) the respective uptakes of the catecholamine into caudate tissue paralleled one another in most experiments, and (2) the content of 3H-DA in push-pull perfusates recovered from sample to sample varied independently of the pump used. Based on the results of histopathological examination and DA radioactivity values, it is concluded that the peristaltic pump as well as the syringe-driven push-pull pump can yield valid experimental observations which are comparable to one another.

Animals↗

PubMed1988

Detecting functional neurotoxicity in the course of safety evaluations with laboratory animals.

In the course of routinely performed subchronic toxicity studies with laboratory rodents, functional neurotoxicity, i.e. behavioral changes are usually noted first during the daily cage side observations of all animals. Observation and subsequent neurologic examination of a few key animals provide a primary tentative diagnosis. Additional automated testing might include: motor activity, startle response, hurdle stepping and maze behavior. Automated testing provides further refinement of the diagnosis and serves to determine the no observable effect level under conditions of blind testing. Behavioral changes should be assessed as early as possible after initiation of testing, i.e. during acute tests for mortality and the following subchronic range-finding studies. Subsequent subchronic organ toxicity studies are then carried out by following a valid experimental protocol for implementation of automated testing procedures and appropriate neuropathologic evaluation.

Abnormalities, Drug-Induced↗

PubMed1985

Turnover of nascent proteins in HeLa-S3 cells and the quasi-linear incorporation kinetics of amino acids.

The turnover of nascent proteins in HeLa S-3 cells was estimated simultaneously with synthesis by the difference between the expected accumulation of radioactivity, based on rate determinations from very short pulse treatments, from that observed by continuous exposure for 100 min. The results were similar to those from conventional pulse-chase analysis, but not identical for reasons which are discussed. They confirm that extensive protein turnover accompanies synthesis in the growing cell, and therefore net synthesis is much lower than gross synthesis. Cells retained all proteins synthesised during heat treatment at 45 degrees C, indicating the complete inhibition of protein degradation, cells retaining all proteins synthesised. This particular perturbation experimentally validated the deficit method, and its potential usefulness in future turnover studies.

Amino Acids↗

PubMed1996

A focal stress gradient-dependent mass transfer mechanism for atherogenesis in branching arteries.

A new arterial wall permeability function, based on the local wall shear stress gradient, has been developed and employed to simulate enhanced low density lipoprotein transfer across the endothelium. the atherosclerotic model used is that of the aorto-celiac junction of rabbits. The experimentally validated computer simulation model for convection mass transfer provides further evidence that the wall shear stress gradient is a reliable predictor of critical atherogenic sites in branching arteries. Some of the underlying biological aspects of atherogenesis due to locally significant and sustained wall shear stress gradient values are briefly discussed.

Animals↗

PubMed1996

In vivo assessment of bone mechanical properties by vibration and ultrasonic wave propagation analysis.

Vibration analysis and ultrasonic wave propagation analysis were evaluated as noninvasive techniques for the in vivo assessment of bone mechanical properties. The relation between the resonant frequencies, obtained by vibration analysis, and geometrical and material properties of long bones is explained using a simple beam model. This simple beam model was validated experimentally in previous work on excised animal bones. In vitro measurements were performed on human and animal excised bones from specific osteopenic cases and control groups. Using specific protocols for in vivo vibration and ultrasound measurements of the tibia, a population of osteoporotic patients and age-matched controls were tested. From these measurements, it was concluded that the bending rigidity, calculated from the resonant frequencies, in osteoporotic tibiae had decreased as compared to the control group. Also the ultrasound velocity in the tibial cortex was lower in the osteoporotic group. The latter indicates a change in the bone tissue material properties. On the other hand, immobilization osteoporosis appeared to lead to a decrease in bending rigidity without an observable change in bone tissue material properties. By the combination of vibration analysis and ultrasound velocity measurements, the whole bone's mechanical characteristics as well as the bone tissue properties can be assessed in vivo. Since both techniques are noninvasive, they can be used in longitudinal studies for the assessment of bone response on physical loading.

Aged↗

PubMed2005

Activation of chondrocytes calcium signalling by dynamic compression is independent of number of cycles.

Mechanical loading is necessary for the development and maintenance of healthy articular cartilage through the control of extracellular matrix synthesis and catabolism. However, the underlying process of chondrocyte mechanotransduction remains unclear. This study examined the influence of cyclic compression on intracellular calcium (Ca(2+)) signalling within isolated articular chondrocytes cultured in agarose constructs. A validated experimental system was developed for applying controlled cyclic cell deformation. Cell-agarose constructs were subjected to 1Hz cyclic compression between 0 and 10% gross strain for 1, 10, 100 or 300 cycles. The cells were subsequently visualised for 300s in the unstrained state using confocal microscopy and the Ca(2+) indicator, Fluo-4 AM. Within unloaded control constructs, a sub-population of approximately 50% of chondrocytes exhibited characteristic spontaneous Ca(2+) transients each lasting approximately 40-60s. Cyclic compression, for only 1 cycle, significantly up-regulated the percentage of cells exhibiting Ca(2+) transients in the subsequent 5min period (p<0.05). Increasing the number of cycles to 10 or 100 had no additional effect. The up-regulated Ca(2+) signalling was maintained for up to 5min before returning to basal levels. By contrast, 300 cycles were followed by Ca(2+) signalling that was not significantly different from that in unloaded controls. However, this response was shown to be due to the increased time following the start of compression. In conclusion, this study indicates that chondrocyte Ca(2+) signalling is stimulated by dynamic compression, probably mediated by cyclic cell deformation. The overall response appears to be independent of the number of cycles or duration of cyclic compression. The sustained up-regulation of Ca(2+) signalling after 1, 10 or 100 cycles suggests the involvement of an autocrine-paracrine signalling mechanism. Furthermore, the reduced response following 300 cycles indicates a possible receptor desensitisation mechanism. Therefore, Ca(2+) signalling may be part of a mechanotransduction pathway through which chondrocyte populations can modulate their metabolic activity in response to changing mechanical stimuli.

Animals↗