Electrocardiographic effects of diagnostic imaging agents.
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Action Potentials↗
Biomedical subjects
R Lehr
Publications and source records attributed to R Lehr.
Inhibition of transforming growth factor (TGF)-beta1-induced extracellular matrix with a novel inhibitor of the TGF-beta type I receptor kinase activity: SB-431542.
Transforming growth factor beta1 (TGF-beta1) is a potent fibrotic factor responsible for the synthesis of extracellular matrix. TGF-beta1 acts through the TGF-beta type I and type II receptors to activate intracellular mediators, such as Smad proteins, the p38 mitogen-activated protein kinase (MAPK), and the extracellular signal-regulated kinase pathway. We expressed the kinase domain of the TGF-beta type I receptor [activin receptor-like kinase (ALK)5] and the substrate, Smad3, and determined that SB-431542 is a selective inhibitor of Smad3 phosphorylation with an IC50 of 94 nM. It inhibited TGF-beta1-induced nuclear Smad3 localization. The p38 mitogen-activated protein kinase inhibitors SB-203580 and SB-202190 also inhibit phosphorylation of Smad3 by ALK5 with IC50 values of 6 and 3 microM, respectively. This suggests that these p38 MAPK inhibitors must be used at concentrations of less than 10 microM to selectively address p38 MAPK mechanisms. However, the p38 MAPK inhibitor SB-242235 did not inhibit ALK5. To evaluate the relative contribution of Smad signaling and p38 MAPK signaling in TGF-beta1-induced matrix production, the effect of SB-431542 was compared with that of SB-242235 in renal epithelial carcinoma A498 cells. All compounds inhibited TGF-beta1-induced fibronectin (FN) mRNA, indicating that FN synthesis is mediated in part via the p38 MAPK pathway. In contrast, SB-431542, but not the selective p38 MAPK inhibitor SB-242235, inhibited TGF-beta1-induced collagen Ialpha1 (col Ialpha1). These data indicate that some matrix markers that are stimulated by TGF-beta1 are mediated via the p38 MAPK pathway (i.e., FN), whereas others seem to be activated via ALK5 signaling independent of the p38 MAPK pathway (i.e., col Ialpha1).
Activin Receptors, Type I↗
Identification and initial characterization of four novel members of the interleukin-1 family.
Interleukin-1 (IL-1), fibroblast growth factors (FGFs), and their homologues are secreted factors that share a common beta-barrel structure and act on target cells by binding to cell surface receptors with immunoglobulin-like folds in their extracellular domain. While numerous members of the FGF family have been discovered, the IL-1 family has remained small and outnumbered by IL-1 receptor homologues. From expressed sequence tag data base searches, we have now identified four additional IL-1 homologues, IL-1H1, IL-1H2, IL-1H3, and IL-1H4. Like most other IL-1/FGFs, these proteins do not contain a hydrophobic leader sequence. IL-1H4 has a propeptide sequence, while IL-1H1, IL-1H2, and IL-1H3 encode only the mature protein. Circular dichroism spectra and thermal stability analysis suggest that IL-1H1 folds similarly to IL-1ra. The novel homologues are not widely expressed in mammals. IL-1H1 is constitutively expressed only in placenta and the squamous epithelium of the esophagus. However, IL-1H1 could be induced in vitro in keratinocytes by interferon-gamma and tumor necrosis factor-alpha and in vivo via a contact hypersensitivity reaction or herpes simplex virus infection. This suggests that IL-1H1 may be involved in pathogenesis of immune mediated disease processes. The addition of four novel IL-1 homologues suggests that the IL-1 family is significantly larger than previously thought.
Amino Acid Sequence↗
The structure of ribonuclease P protein from Staphylococcus aureus reveals a unique binding site for single-stranded RNA.
Ribonuclease P (RNaseP) catalyses the removal of the 5'-leader sequence from pre-tRNA to produce the mature 5' terminus. The prokaryotic RNaseP holoenzyme consists of a catalytic RNA component and a protein subunit (RNaseP protein), which plays an auxiliary but essential role in vivo by binding to the 5'-leader sequence and broadening the substrate specificity of the ribozyme. We determined the three-dimensional high-resolution structure of the RNaseP protein from Staphylococcus aureus (117 amino acid residues) by nuclear magnetic resonance (NMR) spectroscopy in solution. The protein has an alphabeta-fold, similar to the ribonucleoprotein domain. We used small nucleic acid molecules as a model for the 5'-leader sequence to probe the propensity for generic single-stranded RNA binding on the protein surface. The NMR results reveal a contiguous interaction site, which is identical with the previously identified leader sequence binding site in RNaseP holoenzyme. The conserved arginine-rich motif does not bind single-stranded RNA. It is likely that this peptide segment binds selectively to double-stranded sections of P RNA, which are conformationally more rigid. Given the essentiality of RNaseP for the viability of the organism, knowledge of the S. aureus protein structure and insight into its interaction with RNA will help us to develop RNaseP and RNaseP protein as targets for novel antibiotics against this pathogen.
Amino Acid Sequence↗
Damnacanthal is a highly potent, selective inhibitor of p56lck tyrosine kinase activity.
Damnacanthal, an anthraquinone isolated from a plant extract, was found to be a potent, selective inhibitor of p56lck tyrosine kinase activity. The structure, potency, and selectivity of damnacanthal were confirmed by independent synthesis and testing. Damnacanthal exhibited an IC50 of 17 nM for inhibition of p56lck autophosphorylation and an IC50 of 620 nM for phosphorylation of an exogenous peptide by p56lck. Damnacanthal had > 100-fold selectivity for p56lck over the serine/threonine kinases, protein kinase A and protein kinase C, and > 40-fold selectivity for p56lck over four receptor tyrosine kinases. It also demonstrated modest (7-20-fold), but highly statistically significant, selectivity for p56lck over the homologous enzymes p60src and p59fyn. Mechanistic studies demonstrated that damnacanthal was competitive with the peptide binding site, but mixed noncompetitive with the ATP site. Although damnacanthal contains a potentially reactive aldehyde moiety, equilibrium dialysis experiments demonstrated that significant amine formation between damnacanthal and amines occurred only at high concentrations of reactants. However, damnacanthal appeared to bind nonspecifically to membrane lipids and was not active in whole cell tyrosine kinase assays. Damnacanthal is the most potent, selective inhibitor of p56lck tyrosine kinase activity described to date and may represent the starting point for the identification of novel, selective inhibitors of p56lck which are active in whole cell as well as in cell-free systems.
Alkaloids↗
p56lck interactions accelerate autophosphorylation and enhance activity toward exogenous substrates.
p56lck, which is a lymphoid-restricted tyrosine kinase, plays a significant role both in activation of mature T lymphocytes and in T cell development. Recombinant human p56lck was expressed in a baculovirus system and purified to > 95% purity. Purified recombinant p56lck was found to have enzymatic characteristics indistinguishable from those of the native enzyme expressed in the Jurkat human T lymphocytic cell line. The comparisons of recombinant and native p56lck were performed using an assay in which the enzyme was immobilized with antibodies onto the wells of a microtiter plate, enabling in situ purification of the native enzyme. Further characterization of the purified recombinant p56lck revealed significantly different enzyme concentration curves for peptide phosphorylation by immobilized and soluble p56lck. In particular, there was very low activity at low concentrations of enzyme assayed in solution. The activity at low enzyme concentrations could be substantially increased by preincubation of high concentrations of enzyme with ATP prior to dilution for the peptide phosphorylation reaction. We examined the relationship between autophosphorylation and activation for peptide phosphorylation. As for peptide phosphorylation, nonlinear enzyme concentration curves were also observed for autophosphorylation in solution, with very low levels of autophosphorylation at low enzyme concentrations. There was a correlation between the enzyme concentration dependency for autophosphorylation and for preincubation with ATP for maximal enhancement of subsequent peptide phosphorylation. The results suggest that autophosphorylation activates p56lck for phosphorylation of exogenous substrates and that high enzyme concentrations accelerate intermolecular autophosphorylation and enzyme activation.
Adenosine Triphosphate↗
Sixteen S-squared over D-squared: a relation for crude sample size estimates.
I suggest for memorization an equation for calculating approximate sample size requirements intended only for a specific set of values (80 per cent power for a two-tailed alpha = 0.05 test) which seems to occur often in biopharmaceutical research. After presenting the formula in terms of variance estimate s2 and effect size d, I derive a few alternative forms and then discuss the accuracy of the approximation and other properties as well as examples of its use.
Drug Evaluation↗
[Freeze preservation of swine corneas with combinations of intra- and extracellular cryoprotective agents].
Clinically employed methods of corneal cryopreservation usually use the intracellular cryoprotectant dimethyl sulfoxide (DMSO). However, it has been demonstrated that extracellular cryoprotectants such as chondroitin sulfate (ChS) also display effective cryoprotection. The purpose of our study was to investigate the effect of combinations of intra- and extracellular cryoprotectants in corneal cryopreservation. Porcine corneas were cryopreserved in a cryopreservation medium consisting of MEM-medium containing 20% fetal calf serum and 2% chondroitin sulfate. The medium was varied by the addition of 2%, 4% and 8% DMSO. Sixty corneas were cryopreserved at -196 degrees C and thawed at 37 degrees C in a water bath. Morphometric evaluation was not performed directly after thawing but after a 24-h storage period in organ culture. Cryopreservation in medium without DMSO revealed the best results concerning endothelial cell density (2581 cells/mm2). Addition of 2% or 4% DMSO revealed no significant changes in endothelial cell density. Addition of 8% DMSO, however, resulted in a significant decrease (1312 +/- 319 cells/mm2) combined with a significantly higher amount of necrotic areas in the central corneal surface. We conclude that combining intra- and extracellular cryoprotectants does not enhance endothelial cell density after corneal cryopreservation. Higher concentrations of DMSO added to the cryopreservation medium appear to have a negative impact on endothelial cell viability.
Animals↗
Privacy vs HHA employees.
Employers throughout the country are concerned about the apparent epidemic of employee alcohol or substance abuse. The risks to an organization inherent when employees are under the influence of alcohol or drugs are significant. However, those risks, including accidents, carelessness, absenteeism and lack of reliability, are even more critical to health care employers. With the ever increasing focus on medical malpractice issues, no health care employer can afford the risk of an employee being involved in alcohol or drug use or possession during the course of the working day.
Civil Rights↗
Exceptional carcinogenic activity of benz[a]anthracene 3,4-dihydrodiol in the newborn mouse and the bay region theory.
Benz[a]anthracene and the five metabolically possible trans-dihydrodiols of benz[a]anthracene were tested for carcinogenicity in newborn Swiss-Webster mice. Four hundred, 800, and 1600 nmoles hydrocarbon i.p. were sequentially injected on Days 1, 8, and 15 of life. The mice were killed at 22 weeks of age. Of the mice treated with trans-3,4-dihydroxy-3, 4-dihydrobenz[a]anthracene, 24% developed malignant lymphoma, whereas 4% of the animals treated with benz[a]anthracene had malignant lymphoma. None of the animals treated with the trans-1,2-dihydroxy-1,2-dihydrobenz[a]anthracene, trans-5,6-dihydroxy-5,6-dihydrobenz[a]anthracene, trans-8,9-dihydroxy-8,9-dihydrobenz[a]anthracene, or trans-10,11-dihydroxy-10,11-dihydrobenz[a]anthracene had malignant lymphoma. trans-3,4-Dihydroxy-3,4-dihydrobenz[a]anthracene caused about 35-fold more pulmonary adenomas than did benz[a]anthracene, whereas the trans-1,2-dihydroxy-1,2-dihydrobenz[a]anthracene, trans-5,6-dihydroxy-5,6-dihydrobenz[a]anthracene, trans-8,9-dihydroxy-8,9-dihydrobenz[a]anthracene, and trans-10, 11-dihydroxy-10,11-dihydrobenz[a]anthracene had little or no activity. The exceptionally high carcinogenicity of trans-3,4-dihydroxy-3,4-dihydrobenz[a]anthracene is consistent with the metabolism of this compound to either or both of the diastereomeric bay region 3,4-dihydroxy-1,2-epoxy-1,2,3,4-tetrahydrobenz[a]anthracenes, and the data support the bay region theory of polycyclic hydrocarbon carcinogenesis.
Adenoma↗
Effect of reinforcement schedules on reward shifts.
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Animals↗
Strength of alternative responses and subsequent choices.
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Animals↗
Pharmacokinetics and bioavailability of ergoloid mesylates.
The pharmacokinetics and bioavailability of ergoloid mesylates following single administrations of various dose levels (3-9 mg), dosage forms (oral swallow and sublingual tablets, solution) and under different dosing conditions (fasted, with meal) were studied in young healthy volunteers. Male and female subjects showed a similar rate and extent of bioavailable ergoloid after drug treatment. The absorption of ergoloid using either the tablet dosage forms or the drug administered as a solution was rapid, with peak levels of about 60-80 pg ml-1 mg-1 dose achieved after 0.6 to 1.3 h. The elimination half-life for ergoloid in plasma was 2-5 h. Administration of drug with food had no effect on the extent of absorption (AUC) but lowered the absorption rate. This resulted in a reduction of (by 25 per cent) and delay in (by 1 h) achieving peak levels (Cmax). Increasing the ergoloid dose caused a proportional increase in the AUC, but a smaller than proportional increase for Cmax. The tablet formulations provided similar AUCs as the solution; the objective of the sublingual tablet formulation to provide improved bioavailability over the swallow tablet via circumvention of first-pass metabolism was therefore not realized. Transient decreases in blood pressure after ergoloid treatment paralleled the plasma level profiles. Higher ergoloid levels were paired with the larger pressure decreases.
Administration, Oral↗
Distribution of fluorouracil-2-C14 in hypothermic animals with normothermic tumors.
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Adenocarcinoma↗
Hydrophobic interactions and the design of receptor mimetic peptides.
The possibility that hydrophobic interactions may be used as a basis for the design of receptor mimetic peptides for small peptide hormones that lack the potential to adopt amphiphilic secondary structures was tested by designing and characterizing receptor mimetic peptides for gamma-endorphin. The receptor mimetic peptides were designed to exhibit a pattern of hydrophobic surfaces in an antiparallel orientation matching that of the peptide hormone in an extended conformation. An ELISA-based assay was used to determine the relative binding affinities of receptor mimetic peptides, control peptides and antisense peptides to gamma-endorphin immobilized on a surface. The inhibition constant for the best gamma-endorphin receptor mimetic peptide was 1.6 microM. No binding was detected for scrambled control peptides or the antisense-derived peptide mimetic to the limit of their respective solubilities. Sera from rabbits immunized with a gamma-endorphin receptor mimetic peptide were used to immunopurify the ligand-binding domain of the human opiate receptor and were cross-reactive with purified bovine opiate receptor. These results suggest that patterns of hydrophobicity can provide a rational basis for designing receptor mimetic peptides and may provide an explanation for the ability of some antisense peptides to bind to their cognate hormones and to elicit antibodies cross-reactive with hormone receptors.
Amino Acid Sequence↗