Analysis of TGFbeta3 gene expression and protein levels in human bone and serum.

Recent data indicate that TGFbeta3, one member of the TGFbeta-isoforms, has an important role in bone remodeling. Up to date little is known about the expression and regulation of TGFbeta3 in man. We established a highly specific ELISA for quantitative measurement of TGFbeta3 in bone and blood samples and a RT-PCR in combination with HPLC for detection and quantification of TGFbeta3 mRNA in 89 human bone samples.

Proposal of abolition of the skin sensitivity test before equine rabies immune globulin application.

An epizootic outbreak of rabies occurred in 1995 in Ribeirão Preto, SP, with 58 cases of animal rabies (54 dogs, 3 cats and 1 bat) confirmed by the Pasteur Institute of São Paulo, and one human death. The need to provide care to a large number of people for the application of equine rabies immune globulin (ERIG) prevented the execution of the skin sensitivity test (SST) and often also the execution of desensitization, procedures routinely used up to that time at the Emergency Unit of the University Hospital of the Faculty of Medicine of Ribeirão Preto, University of São Paulo (EU-UHFMRP-USP), a reference hospital for the application of heterologous sera. In view of our positive experience of several years with the abolition of SST and of the use of premedication before the application of antivenom sera, we used a similar schedule for ERIG application.

Molecular determinants of inactivation in voltage-gated Ca2+ channels.

Evolution has created a large family of different classes of voltage-gated Ca2+ channels and a variety of additional splice variants with different inactivation properties. Inactivation controls the amount of Ca2+ entry during an action potential and is, therefore, believed to play an important role in tissue-specific Ca2+ signalling. Furthermore, mutations in a neuronal Ca2+ channel (Ca(v)2.

Air quality measurements from the Fresno Supersite.

The Fresno Supersite intends to 1) evaluate non-routine monitoring methods, establishing their comparability with existing methods and their applicability to air quality planning, exposure assessment, and health effects studies; 2) provide a better understanding of aerosol characteristics, behavior, and sources to assist regulatory agencies in developing standards and strategies that protect public health; and 3) support studies that evaluate relationships between aerosol properties, co-factors, and observed health end-points. Supersite observables include in-situ, continuous, short-duration measurements of 1) PM2.5, PM10, and coarse (PM10 minus PM2.

Modulation of slow inactivation in class A Ca2+ channels by beta-subunits.

beta-subunit modulation of slow inactivation of class A calcium (Ca2+) channels was studied with two-microlectrode voltage clamp after expression of the alpha1A- (BI-2) together with beta1a-, beta2a-, beta3- or beta4-subunits in Xenopus oocytes. On- and off-rates of slow inactivation were estimated from the kinetics of recovery from slow inactivation. Ca2+ channels with an alpha1A/beta-subunit composition inducing the slower rate of fast inactivation displayed the faster rate of slow inactivation.

Mitochondrial DNA in the Central European population. Human identification with the help of the forensic mt-DNA D-loop-base database.

Sequencing of mtDNA is an advanced method for the individualisation of traces. Disadvantages of this method are expensive and time-consuming analysis and evaluation procedures as well as the necessary stock of population-genetic data which is still insufficient. Central European institutes of forensic medicine from Germany, Austria, and Switzerland have been working together since the beginning of 1998 to establish a mtDNA database.

Development of the X-linked tetrameric microsatellite marker DXS9898 for forensic purposes.

HumDXS9898 also known as CHLC x GATA 126G01 is a tetrameric microsatellite marker located at the Xq21.33 pericentromeric region. In kinship testing HumDXS9898 is suitable for concomitant use with HumHPRTB and HumDXS6807 which are separated from HumDXS9898 by genetic map distance of 150 and 80 cM, respectively.

Analysis and simulation of wintertime light scattering by the urban aerosol in Dallas-Fort Worth.

Wintertime atmospheric light scattering in Dallas, TX, was estimated through the use of aerosol models. Input data for the aerosol models were provided by measurements of aerosol chemistry, physical particle size distributions, and distributions of particulate sulfur by particle size, and by predictions by an atmospheric simulation model. Light scattering measurements provided a basis for testing the aerosol models.

High affinity interaction of mibefradil with voltage-gated calcium and sodium channels.

Mibefradil is a novel Ca(2+) antagonist which blocks both high-voltage activated and low voltage-activated Ca(2+) channels. Although L-type Ca(2+) channel block was demonstrated in functional experiments its molecular interaction with the channel has not yet been studied. We therefore investigated the binding of [(3)H]-mibefradil and a series of mibefradil analogues to L-type Ca(2+) channels in different tissues.

Molecular mechanism of calcium channel block by isradipine. Role of a drug-induced inactivated channel conformation.

The role of the inactivated channel conformation in the molecular mechanism of Ca(2+) channel block by the 1,4-dihydropyridine (DHP) (+)-isradipine was analyzed in L-type channel constructs (alpha(1Lc); Berjukow, S., Gapp, F., Aczel, S.

Inactivation determinant in the I-II loop of the Ca2+ channel alpha1-subunit and beta-subunit interaction affect sensitivity for the phenylalkylamine (-)gallopamil.

1. The role of calcium (Ca2+) channel inactivation in the molecular mechanism of channel block by phenylalkylamines (PAAs) was analysed in a PAA-sensitive rabbit brain class A Ca2+ channel mutant (alpha1A-PAA). Use-dependent barium current (IBa) inhibition of alpha1A-PAA by (-)gallopamil and Ca2+ channel recovery from inactivation and block were studied with two-microlectrode voltage clamp after expression of alpha1A-PAA and auxiliary alpha2-delta- and beta1a- or beta2a-subunits in Xenopus oocytes.

The Magnitude of Bias in the Measurement of PM25 Arising from Volatilization of Particulate Nitrate from Teflon Filters.

Because the Federal Reference Method for PM25 specifies the collection of ambient particles on Teflon filters, we have examined the loss of a known volatile species, particulate nitrate, during sampling. Data are presented from two studies in southern California for which parallel samples were collected by different methods. Differences in collected nitrate are modeled using an evaporation model based on the work of Zhang and McMurry.

Sequence differences between alpha1C and alpha1S Ca2+ channel subunits reveal structural determinants of a guarded and modulated benzothiazepine receptor.

The molecular basis of the Ca2+ channel block by (+)-cis-diltiazem was studied in class A/L-type chimeras and mutant alpha1C-a Ca2+ channels. Chimeras consisted of either rabbit heart (alpha1C-a) or carp skeletal muscle (alpha1S) sequence in transmembrane segments IIIS6, IVS6, and adjacent S5-S6 linkers. Only chimeras containing sequences from alpha1C-a were efficiently blocked by (+)-cis-diltiazem, whereas the phenylalkylamine (-)-gallopamil efficiently blocked both constructs.

TNFa and b microsatellites in Germany.

Allele frequencies of TNFa and TNFb microsatellites were determined from 315 healthy unrelated Germans (mothers and putative fathers) by means of PCR. They were stably inherited and segregated in a Mendelian way. New mutations were not observed.

Mechanism of voltage- and use-dependent block of class A Ca2+ channels by mibefradil.

1. The action of mibefradil was studied on wild type class A calcium (Ca2+) channels and various class A/L-type channel chimaeras expressed in Xenopus oocytes. The mechanism of Ca2+ channel block by mibefradil was evaluated with two microelectrode voltage clamp.

Molecular mechanism of diltiazem interaction with L-type Ca2+ channels.

Benzothiazepine Ca2+ antagonists (such as (+)-cis-diltiazem) interact with transmembrane segments IIIS6 and IVS6 in the alpha1 subunit of L-type Ca2+ channels. We investigated the contribution of individual IIIS6 amino acid residues for diltiazem sensitivity by employing alanine scanning mutagenesis in a benzothiazepine-sensitive alpha1 subunit chimera (ALDIL) expressed in Xenopus laevis oocytes. The most dramatic decrease of block by 100 microM diltiazem (ALDIL 45 +/- 4.

Molecular basis of drug interaction with L-type Ca2+ channels.

Different types of voltage-gated Ca2+ channels exist in the plasma membrane of electrically excitable cells. By controlling depolarization-induced Ca2+ entry into cells they serve important physiological functions, such as excitation-contraction coupling, neurotransmitter and hormone secretion, and neuronal plasticity. Their function is fine-tuned by a variety of modulators, such as enzymes and G-proteins.

Small-volume and rapid extracellular solution exchange around Xenopus oocytes during voltage-clamp recordings.

A novel method for micro extracellular perfusion of Xenopus oocytes has been designed in order to minimise the amount of test solution required for drug applications during two-microelectrode voltage-clamp experiments. Voltage-clamp experiments on oocytes are performed in a glass-covered microbath without continuous perfusion. The microelectrodes access the oocytes via two small openings in the glass cover.

Structural basis of drug binding to L Ca2+ channels.

At least five different types of voltage-gated Ca2+ channels exist in electrically excitable mammalian cells. Only one type, the family of L-type Ca2+ channels (L channels), contains high-affinity binding domains within their alpha 1-subunits for different chemical classes of drugs (Ca2+ channel antagonists; exemplified by isradipine, verapamil and diltiazem). Their stereoselective, high-affinity binding induces block of channel-mediated Ca2+ inward currents in heart and smooth muscle, resulting in antihypertensive, cardiodepressive and antiarrhythmic effects.

Familial hemiplegic migraine mutations change alpha1A Ca2+ channel kinetics.

Missense mutations in the pore-forming human alpha1A subunit of neuronal P/Q-type Ca2+ channels are associated with familial hemiplegic migraine (FHM). The pathophysiological consequences of these mutations are unknown. We have introduced the four single mutations reported for the human alpha1A subunit into the conserved rabbit alpha1A (R192Q, T666M, V714A, and I1819L) and investigated possible changes in channel function after functional expression of mutant subunits in Xenopus laevis oocytes.

Molecular mechanism of use-dependent calcium channel block by phenylalkylamines: role of inactivation.

The role of channel inactivation in the molecular mechanism of calcium (Ca2+) channel block by phenylalkylamines (PAA) was analyzed by designing mutant Ca2+ channels that carry the high affinity determinants of the PAA receptor site [Hockerman, G. H., Johnson, B.

Nine L-type amino acid residues confer full 1,4-dihydropyridine sensitivity to the neuronal calcium channel alpha1A subunit. Role of L-type Met1188.

Pharmacological modulation by 1,4-dihydropyridines is a central feature of L-type calcium channels. Recently, eight L-type amino acid residues in transmembrane segments IIIS5, IIIS6, and IVS6 of the calcium channel alpha1 subunit were identified to substantially contribute to 1,4-dihydropyridine sensitivity. To determine whether these eight L-type residues (Thr1066, Gln1070, Ile1180, Ile1183, Tyr1490, Met1491, Ile1497, and Ile1498; alpha1C-a numbering) are sufficient to form a high affinity 1,4-dihydropyridine binding site in a non-L-type calcium channel, we transferred them to the 1, 4-dihydropyridine-insensitive alpha1A subunit using site-directed mutagenesis.

Calcium channel block by (-)devapamil is affected by the sequence environment and composition of the phenylalkylamine receptor site.

The pore-forming alpha 1 subunit of L-type calcium (Ca2+) channels is the molecular target of Ca2+ channel blockers such as phenylalkylamines (PAAs). Association and dissociation rates of (-)devapamil were compared for a highly PAA-sensitive L-type Ca2+ channel chimera (Lh) and various class A Ca2+ channel mutants. These mutants carry the high-affinity determinants of the PAA receptor site in a class A sequence environment.