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Biomedical subjects

R Weinberger

Publications and source records attributed to R Weinberger.

At least 19 recordsLinked to original sources
PubMed2001

Differential expression of the actin-binding proteins, alpha-actinin-2 and -3, in different species: implications for the evolution of functional redundancy.

The alpha-actinins are a multigene family of four actin-binding proteins related to dystrophin. The two skeletal muscle isoforms of alpha-actinin (ACTN2 and ACTN3) are major structural components of the Z-line involved in anchoring the actin-containing thin filaments. In humans, ACTN2 is expressed in all muscle fibres, while ACTN3 expression is restricted to a subset of type 2 fibres. We have recently demonstrated that alpha-actinin-3 is absent in approximately 18% of individuals in a range of human populations, and that homozygosity for a premature stop codon (577X) accounts for most cases of true alpha-actinin-3 deficiency. Absence of alpha-actinin-3 is not associated with an obvious disease phenotype, raising the possibility that ACTN3 is functionally redundant in humans, and that alpha-actinin-2 is able to compensate for alpha-actinin-3 deficiency. We now present data concerning the expression of ACTN3 in other species. Genotyping of non-human primates indicates that the 577X null mutation has likely arisen in humans. The mouse genome contains four orthologues which all map to evolutionarily conserved syntenic regions for the four human genes. Murine Actn2 and Actn3 are differentially expressed, spatially and temporally, during embryonic development and, in contrast to humans, alpha-actinin-2 expression does not completely overlap alpha-actinin-3 in postnatal skeletal muscle, suggesting independent function. Furthermore, sequence comparison of human, mouse and chicken alpha-actinin genes demonstrates that ACTN3 has been conserved over a long period of evolutionary time, implying a constraint on evolutionary rate imposed by continued function of the gene. These observations provide a real framework in which to test theoretical models of genetic redundancy as they apply to human populations. In addition we highlight the need for caution in making conclusions about gene function from the phenotypic consequences of loss-of-function mutations in animal knockout models.

Actinin↗
PubMed2001

Capillary electrophoresis of venoms and toxins.

A review of capillary electrophoresis of venoms and toxins is presented. Emphasis is placed on the analysis of real samples in complex matrices. The structures of some of the complex toxins are presented to illustrate the remarkable diversity and complexity of these materials.

Animals↗
PubMed2001

Use of dynamically coated capillaries for the routine analysis of methamphetamine, amphetamine, MDA, MDMA, MDEA, and cocaine using capillary electrophoresis.

A rapid, accurate, precise, reproducible, economical, and environmentally gentle method using capillary electrophoresis (CE) is presented for the routine analysis of methamphetamine, amphetamine, MDA, MDMA, MDEA, and cocaine in seized drugs. The methodology uses a 32 cm by 50 microm capillary (length to detector 23.5 cm) with a commercially available buffer kit and diode array UV detection. Dynamic coating of the capillary surface is accomplished by flushing with base for 1 min, a proprietary polycation for 1 min, and then a proprietary polyanion for 2 min. This approach provides a relatively high and stable electroosmotic flow (EOF), even at low pHs. The background electrolyte (BGE) contains 75 mM phosphate buffer (pH 2.5) with the same polyanion as above. Using this methodology, amphetamine, methamphetamine, MDA, MDMA, MDEA, and an internal standard (n-butylamphetamine) are baseline resolved in less than 5 min. The run-to-run migration time %RSDs and peak area %RSDs are typically <0.3% and <2.1%, respectively. The day-to-day and capillary-to-capillary migration time %RSDs are <1.5% and <2.1%, respectively. The %RSDs of the relative migration times compared with the internal standard on a day-to-day and capillary-to-capillary basis are <0.2% and <0.06%, respectively. The linear dynamic range using peak areas range from 0.003 to 0.10 mg/mL. The correlation coefficients are >0.9998, with all calibration curves passing at or near the origin. Similar data are obtained for cocaine and its internal standard henyltoloxamine. None of the compounds usually encountered in illicit samples interfere with the target compound (e.g., methamphetamine and cocaine) or the internal standard. Quantitative results for synthetic mixtures and seized exhibits are in good agreement with actual values, and also with results obtained from other techniques. The relatively high EOF for the dynamically coated capillary system allows for the screening of basic, acidic, and neutral adulterants in drug seizures; identification is facilitated by the use of automated UV library searches.

Journal Article↗
PubMed1999

Specific isoforms of actin-binding proteins on distinct populations of Golgi-derived vesicles.

Golgi membranes and Golgi-derived vesicles are associated with multiple cytoskeletal proteins and motors, the diversity and distribution of which have not yet been defined. Carrier vesicles were separated from Golgi membranes, using an in vitro budding assay, and different populations of vesicles were separated using sucrose density gradients. Three main populations of vesicles labeled with beta-COP, gamma-adaptin, or p200/myosin II were separated and analyzed for the presence of actin/actin-binding proteins. beta-Actin was bound to Golgi cisternae and to all populations of newly budded vesicles. Centractin was selectively associated with vesicles co-distributing with beta-COP-vesicles, while p200/myosin II (non-muscle myosin IIA) and non-muscle myosin IIB were found on different vesicle populations. Isoforms of the Tm5 tropomyosins were found on selected Golgi-derived vesicles, while other Tm isoforms did not colocalize with Tm5 indicating the association of specialized actin filaments with Golgi-derived vesicles. Golgi-derived vesicles were shown to bind to F-actin polymerized from cytosol with Jasplakinolide. Thus, newly budded, coated vesicles derived from Golgi membranes can bind to actin and are customized for differential interactions with microfilaments by the presence of selective arrays of actin-binding proteins.

Actins↗
PubMed1998

Creating intracellular structural domains: spatial segregation of actin and tropomyosin isoforms in neurons.

Actin microfilaments play a direct role in a variety of cell processes. Distinct populations of microfilaments are associated with different cellular compartments, such as growth cones, filipodia, stress fibers, and lamellipodia. It is becoming clear that these different populations are often composed of different isoforms of the two core microfilament components, actin and tropomyosin. This is particularly true in neurons, where actin and tropomyosin isoforms are segregated into different intracellular compartments which correspond to functionally distinct regions of the neuron. Developmental regulation of this isoform sorting suggests a specific role for some isoforms in growth and for others in stabilization of neuronal structure. This provides a mechanism by which a neuron can create and independently regulate intracellular domains composed of microfilaments with different functional properties.

Actin Cytoskeleton↗
PubMed1998

Isoform sorting and the creation of intracellular compartments.

The generation of isoforms via gene duplication and alternative splicing has been a valuable evolutionary tool for the creation of biological diversity. In addition to the formation of molecules with related but different functional characteristics, it is now apparent that isoforms can be segregated into different intracellular sites within the same cell. Sorting has been observed in a wide range of genes, including those encoding structural molecules, receptors, channels, enzymes, and signaling molecules. This results in the creation of intracellular compartments that (a) can be independently controlled and (b) have different functional properties. The sorting mechanisms are likely to operate at the level of both proteins and mRNAs. Isoform sorting may be an important consequence of the evolution of isoforms and is likely to have contributed to the diversity of functional properties within groups of isoforms.

Actin Cytoskeleton↗
PubMed1997

Planetary nebulae: understanding the physical and chemical evolution of dying stars.

Planetary nebulae are one of the few classes of celestial objects that are active in every part of the electromagnetic spectrum. These fluorescing and often dusty expanding gaseous envelopes were recently found to be quite complex in their dynamics and morphology, but refined theoretical models can account for these discoveries. Great progress was also made in understanding the mechanisms that shape the nebulae and the spectra of their central stars. In addition, applications for planetary nebulae have been worked out; for example, they have been used as standard candles for long-range distances and as tracers of the enigmatic dark matter.

Astronomical Phenomena↗
PubMed1997

Actin and tropomyosin isoforms in morphogenesis.

The major components of the actin microfilament system, actin and tropomyosin (Tm), are encoded by multigene families. There are at least 6 actin and over 20 Tm isoforms in mammals. The observation that isoforms are expressed in a tissue-specific manner has encouraged the hypothesis that they contribute to the formation of cell type-specific structures. Recent studies have indicated that certain specific isoforms do play unique structural roles. One nonmuscle actin isoform, beta, is implicated in the regulation of cell spreading and membrane organisation. The intracellular location of beta-actin mRNA has been shown to be regulated by growth factor stimulation of signal transduction pathways. Actin isoforms have also been shown to differ in their contractile properties in both muscle and non-muscle cells. Tropomyosins have been found to show isoform specific regulation in response to cell transformation. This has correlated with the view that some isoforms of tropomyosin promote filament stability whereas others are associated with more dynamic structures. Neuronal development and maturation are accompanied by dynamic spatial sorting of tropomyosin isoforms into different cellular compartments. It is now apparent that isoforms of these proteins perform different structural tasks. The challenge is now to link the significance of spatial sorting to the different physicochemical properties of these isoforms.

Actin Cytoskeleton↗
PubMed1996

The molecular composition of neuronal microfilaments is spatially and temporally regulated.

The actin-based microfilament system is thought to play a critical role in neuronal development. We have determined specific changes in the composition of microfilaments accompanying neuronal morphogenesis. By using specific antibodies against the isoforms for tropomyosin (Tm) (Tm-5 and TmBr-1/-3) and actin (beta- and gamma-actin), we found that during early morphogenesis in vivo immature growing axons contain beta- and gamma-actin and Tm-5. In particular, Tm-5 is exclusively located in the immature axonal processes relative to the neuronal cell body. In contrast, beta-actin and Tm-5 are absent in mature, quiescent axons. This developmental loss from axons is associated with an approximately twofold downregulation of beta-actin and Tm-5 levels in the brain; gamma-actin levels do not change, and this molecule is widely distributed throughout neurons during development. The loss of beta-actin and Tm-5 from axons is accompanied by a progressive appearance of TmBr-1/-3. This apparent replacement of Tm-5 with TmBr-1/-3 occurs over a 2 d time period during rat embryonic hindbrain development and is conserved in evolution between birds and mammals. The loss of Tm-5 from axons involves a redistribution of this molecule to the cell soma and dendrites. These findings suggest that specialized microfilament domains are associated with the development and maintenance of neuronal polarity. We conclude that these Tm isoforms and beta-actin are subject to specific patterns of segregation associated with axonal development and neuronal differentiation. This provides a potential molecular basis for the temporal and spatial specificity of microfilament function during neuronal differentiation.

Actin Cytoskeleton↗
PubMed1991

Teaching the elderly stress reduction.

1. Psychological or physiological stressors cause changes within the body through the action of the hypothalamus. 2. There are several techniques for stress management, including progressive muscle relaxation and guided imagery. 3. The nurse can teach stress management techniques to the elderly using teaching modifications to respond to changes due to aging. 4. The nurse can teach stress management in various settings to clients who have a nursing diagnosis such as anxiety, alteration in comfort, or sleep pattern disturbance.

Aged↗
PubMed1990

Post-column continuous-flow analysis combined with reversed-phase liquid chromatography and computer-aided detection for the characterisation of peptides.

Peptide mapping is a key technique for structural identification of new proteins or the products of recombinant gene technology. The recognition of oligopeptides, separated by reversed-phase liquid chromatography, is limited by the conventional reliance on the correlation of retention times with standards, supported by dual-wavelength chromatograms. It has been reported that the recognition of phenolic compounds can be achieved by a novel technique, based on computer-aided photodiode-array detection of the pH-shifted solutes after post-column continuous-flow analysis. This work describes how the generation of the pH-shifted difference spectra for dipeptides, containing a tyrosyl residue, may be used to enhance peak recognition, when used in conjunction with absorbance ratios.

Absorptiometry, Photon↗
PubMed1990

Capillary electrophoresis of urinary porphyrins with absorbance and fluorescence detection.

Urinary porphyrins are separated in a 72 cm x 50 microns I.D. fused-silica capillary by micellar electrokinetic capillary chromatography with 100 mM sodium dodecyl sulfate and 20 mM 3-(cyclohexylamino)-1-propanesulfonic acid at pH 11. Detection is accomplished by absorbance at 400 nm or fluorescence with excitation at 400 nm and emission at wavelengths above 550 nm. Substantial trace enrichment is found for porphyrins in urine samples or for porphyrin standards prepared without surfactant in the injection buffer. Limits of detection are in the 100 pmol/ml concentration range with an optimized fluorescence system. The method is shown suitable for the determination of porphyrins in clinical urine specimens. Comparisons are made between electrophoretic and chromatographic methods for the separation and detection of urinary porphyrins.

Chromatography, Liquid↗
PubMed1990

Optimization of multidimensional high-performance liquid chromatography for the determination of drugs in plasma by direct injection, micellar cleanup and photodiode array detection.

Improvements in a multidimensional liquid chromatography system for the direct determination of drug substances in blood plasma are reported. The system employs an on-line micellar chromatographic cleanup followed by a reversed-phase analytical separation. The limit of detection of propranolol is improved by a factor 10 compared to previously reported work. The technique is applied towards the determination of a multicomponent mixture of tricyclic anti-depressants in blood plasma. A protocol for optimization is described.

Chromatography, High Pressure Liquid↗
PubMed1987

Determination of reducing and non-reducing carbohydrates in food products by liquid chromatography with post-column catalytic hydrolysis and derivatization. Comparison with refractive index detection.

Post-column catalytic hydrolysis combined with 4-aminobenzoic acid hydrazide derivatization is employed for the determination of both reducing and non-reducing carbohydrates in a variety of complex sample matrices such as dairy products, processed foods and tobaccos. Comparison with refractive index detection shows the post-column method to be superior from the standpoint of selectivity, sensitivity and simplicity of sample preparation. Limits of detection are in the low-nanogram range. Quantitative results are presented for the determination of sugars in potato extracts. The method is also applicable for the separation and detection of carbohydrate oligomers.

Beverages↗
PubMed1984

Determination of trace levels of steroids in blood plasma by liquid chromatography with peroxyoxalate chemiluminescence detection.

A highly sensitive and specific liquid chromatographic procedure with peroxyoxalate chemiluminescence detection has been developed for the determination of fluocortin butyl, a 3 alpha-ketocorticosteroid, in blood plasma. The technique employs dansylation of the steroid to provide a highly chemiluminescent derivative. After separation by reversed-phase liquid chromatography, reagents necessary for chemiluminescence are added, followed by detection in a conventional fluorimetric detector in which the excitation source is deactivated. The precision is 2.5% relative standard deviation at the 10 ng/ml level, and the response is linear up to at least 4 ng injected steroid. The procedure requires only 1 ml blood plasma and has a limit of detection of 100 pg/ml or 7.5 pg injected steroid. The system is reliably used for routine pharmacokinetic studies and with modifications, is applicable to other steroids as well.

Biological Availability↗