Ira S. Richards

Hydrogen peroxide: A potent activator of dioxygenase activity of soybean lipoxygenase

Hydrogen peroxide, an ubiquitous biologically occurring peroxide, was found to stimulate the dioxygenase activity of soybean lipoxygenase at the physiologically attainable concentration. The increase in enzyme specific activity was directly proportional to hydrogen peroxide concentration up to 0.5 nM. A decrease in the stimulation of dioxygenase activity was observed at higher concentrations. At low enzyme concentration up to 28-fold stimulation was noted when the formation of lipid hydroperoxide was monitored spectrophotometrically. The stimulation was further confirmed by increased oxygen uptake. It is proposed that the mechanism for in vivo activation involves hydrogen peroxide.

Current status of endocrinologic effects of vasectomy

Experimental investigation into possible endocrine consequences of vasectomy has been carried out since 1927. Early results seemed to indicate that vasectomy had both positive and negative consequences. This is a review of the recent literature from animal studies and both short- and long-term studies in man. Examinations of interstitial tissue following vasectomy have shown the tissue to be unchanged. Testosterone production and the sensitivity of the testes to the action of gonadotropins is not seen to be impaired within the 1st 3 months postvasectomy. In addition vasectomy does not seem to be associated with longterm changes in testosterone FSH or LH levels in the peripheral circulation or with alterations in testosterone concentration in seminal plasma. Species differences make transposition of animal data to the human situation difficult. Further research of the following types is still needed: 1) prospective and retrospective studies of more than 5 years duration; 2) in vitro controlled studies of steroid metabolism by testis preparations; 3) nonhuman primate studies to correlate better the morphologic and biochemical effects of vasectomy on the pituitary-testis relationship; and 4) studies of the possible effects of vasectomy on other endocrine glands.

Florida red-tide toxins (brevetoxins) produce depolarization of airway smooth muscle

Crude preparations of brevetoxin (PBTX) produce airway contraction; however, it is not known if this toxin-induced mechanical response is coupled to changes in airway smooth muscle membrane potential. Membrane potentials and contractility of in vitro canine trachealis smooth muscle preparations were simultaneously measured with a microelectrode and microforce transducer before and during exposure to either the crude toxin (0.01-1.2 micrograms/ml), or the purified fractions PBTX-2 or PBTX-3 (0.01-0.07 micrograms/ml). Membrane potentials in cultured airway smooth muscle-reaggregate preparations were similarly studied. Toxins produced concentration-dependent depolarizations and contractions in in vitro preparations. These responses were not obtained in the presence of either the muscarinic blocking agent atropine, the sodium channel blocker tetrodotoxin (TTX), 0 mM extracellular Ca2+, or the Ca2+ channel blocker verapamil. The toxins were without effect in cultured cells, whereas acetylcholine produced depolarizations which were blocked in the presence of atropine, but not TTX. This suggested the presence of functional cholinergic receptors in cultured cells, and the PBTX-induced release of endogenous acetylcholine from peripheral nerve endings in the in vitro airway smooth muscle response.

Ethanol-induced bronchodilatation in TEA-treated canine tracheal smooth muscle is mediated by a β-adrenoceptor-dependent mechanism

The effects of moderate concentrations of ethanol (8-34 mM) on the electromechanical activity of airway smooth muscle cells of canine trachealis, stimulated by the spasmogen tetraethylammonium (TEA), are described for in vitro and cultured reaggregate preparations. Ethanol produced a concentration-dependent hyperpolarization, and suppression of action potentials in smooth muscle preparations, in vitro, whereas it was without effect in cultured airway smooth muscle cells. In the presence of the beta-adrenoceptor antagonist propranolol (1 microM), ethanol had no effect on in vitro preparations. Isoproterenol (0.1 microM) produced hyperpolarization and suppression of action potentials in airway smooth muscle of both preparations. These effects were not observed when propranolol was additionally present. This suggests that both in vitro, and cultured airway smooth muscle preparations maintained their beta-receptors, and that ethanol caused the release of endogenous catecholamine from adrenergic nerve endings which apparently remained intact in in vitro, but not in cultured airway smooth muscle preparations.

Azelastine and desmethylazelastine suppress acetylcholine-induced contraction and depolarization in human airway smooth muscle

We examined the effects of a new anti-asthmatic drug, azelastine, and its principal metabolite, desmethylazelastine, on the in vitro electromechanical response of human airway smooth muscle during cholinergic stimulation. Membrane potential and isometric force were simultaneously measured using an intracellular microelectrode and a microforce transducer. Desmethylazelastine significantly suppressed acetylcholine-induced depolarization and contraction at 10(-6) M, whereas azelastine produced similar results at 10(-4) M, suggesting that the metabolite may be the principal compound acting upon the airway smooth muscle cell.

Ethanol Potentiates the Depressant Effects of Cocaine in Human Fetal Myocardium In Vitro

BACKGROUND Cocaine and ethanol use is widespread in our society and is not uncommon in pregnant women. Previous work has demonstrated that acute exposure to cocaine produced significant effects on the configuration of human fetal myocardial action potentials and contractility in vitro. It has been hypothesized that these target-specific effects of cocaine may provide a plausible mechanism to account for fetal arrhythmia or sudden fetal death in utero. OBJECTIVE This study was conducted to determine if treatment with a low concentration of ethanol (200 mg/L) would predispose human fetal myocardium to cocaine-induced toxicity in vitro. METHODS Fetal hearts (12-14 weeks) were obtained at the time of elective abortion and transported to the laboratory in cold physiological salt solution. The force of muscle contractions and transmembrane potentials of ventricular walls were studied during external electrical stimulation in a specially constructed superfusion chamber. RESULTS When a concentration of cocaine (200 micrograms/L physiological salt solution) that singly produced only modest myocardial depression was used in combination with a concentration of ethanol which alone produced nonsignificant changes, marked depression and block of action potentials and contractility resulted. CONCLUSION The combined use of ethanol and cocaine produces fetal myocardial depression which is greater than that predicted from the effects of these chemicals individually and may have significant in utero implications.

Metabolism of 1,2-dibromoethane in the human fetal liver

1. Toxicity of 1,2-dibromoethane requires bioactivation via glutathione S-transferase. Since this enzyme is undetectable in the fetus of several laboratory animal species during early gestation, in vitro studies were carried out with human fetal liver to assess potential fetotoxicity. 2. Glutathione S-transferase occurs abundantly in the human fetal liver cytosol and its titer is equal to or exceeds that found in adult human liver when estimated using 1-chloro-2,4-nitrobenzene as the second substrate. 3. Human fetal liver cytosolic glutathione S-transferase metabolized 1,2-dibromoethane with a high efficiency (mean +/- SD specific activity of 3.10 +/- 0.83 nmol/min/mg protein). This reaction was enzymatic in nature and the rate of conjugation was proportional to the concentration of reduced glutathione, 1,2-dibromoethane and the enzyme present in the reaction medium. 4. A significant bioactivation with a possibility of only limited detoxication via cytochrome P-450-dependent oxidation suggests that human fetus may be at greater risk from 1,2-dibromoethane toxicity than adult.

Red tide toxin produces in vitro depolarization of human airway smooth muscle

Background. Brevetoxin (PbTx), taken from the earlier species name Ptychodiscus brevis, is a red algae toxin. It has been associated with clinically observed bronchoconstriction in nonasthmatics. In asthmatics, similar exposures may produce severe transient effects, sometimes requiring emergency treatment, thus suggesting that asthmatics are more sensitive to this toxin. As such, we have investigated potential mechanisms in vitro. Methods. Membrane potentials of in vitro airway smooth muscle (ASM) preparations were measured with a microelectrode before, during, and after the exposure to PbTx (0.01–1.2 μg/mL) in strip preparations (SPs) and cultured ASM reaggregate preparations. The latter preparation results in the disruption of normal peripheral nervous ASM associations through enzymatic dissociation of cells. Results. We observed an increased level of depolarization in asthmatic preparations at the same level of exposure. Exposure to PbTx produced concentration-dependent depolarization in both nonasthmatic and asthmatic in vitro SPs. In the former, responses did not occur in the presence of the blocking agents such as atropine or tetrodotoxin (TTX). In asthmatic SPs, atropine and TTX produced little effect, whereas verapamil blocked the PbTx-induced depolarization. The toxin was without effect in nonasthmatic cultured cells, whereas acetylcholine produced depolarization that was blocked in the presence of atropine, but not TTX or verapamil. In contrast, the toxin produced significant depolarization in cultured asthmatic ASM cells, which were unaffected by either atropine or TTX but were blocked by verapamil. Conclusions. We propose that PbTx directly affects asthmatic ASM whereas the effect is neurally mediated in nonasthmatics.

Gender-specific differences in the urinary expression of aldosterone, IL-1α and IL-1β.

AIMS This pilot investigation examined the possibility of using urine specimens to explore the difference between the expression of several biomarkers based on gender. These biomarkers include several associated with cardiac damage, oxidative stress and inflammation. MATERIALS & METHODS Urine specimens were assayed for total protein, aldosterone, high-sensitivity C-reactive protein, myeloperoxidase and IL-1α and -1β using ELISA. RESULTS We observed significant differences between the sexes for aldosterone and IL-1α and -1β. CONCLUSION The presence of gender-based differences in the urinary expression of these biomarkers may be important for establishing normal baseline values in males and females, and may prove to be of value in the development of rapid noninvasive ways to assess inflammatory and oxidative injury during routine urinalysis.

Dose, time and route dependency of the induction of rat hepatic ornithine decarboxylase by 12-tetradecanoylphorbol 13-acetate.

Ornithine decarboxylase (ODC) activity, the rate limiting enzyme in polyamine biosynthesis, was determined after 12-O-tetradecanoylphorbol 13-acetate (TPA) administration to female Sprague-Dawley rats. The extent of induction depended on the dose, exposure, time and route of administration. The most effective dose for ODC induction by the intraperitoneal route was 40 ug TPA/kg which caused 3-5 fold ODC induction. Maximal ODC induction occurred in a narrow time band 5 hours after TPA administration. TPA had no adverse effects on hepatic DNA (measured by alkaline elution), cytochrome P-450 content and reduced glutathione content or serum alanine aminotransferase (SGPT) activity.