The Effects of Iron-Mediated Oxidative Stress in Isolated Renal Cortical Brush Border Membrane Vesicles at Normothermic and Hypothermic Temperatures

Experiments on renal cortical brush border membrane vesicles have been undertaken in order to assess the involvement of iron in oxidative stress at physiological temperatures and under conditions of hypothermia. A decrease in temperature stimulated iron-induced lipid peroxidation. The results are discussed in relation to the role of the oxidation state of the iron and iron(II)/iron(III) ratios in the initiation of peroxidative events.     

Effects of treadmill exercise on fuel metabolism in hepatic cirrhosis

We studied whole body and regional fuel metabolism before, during, and after 90 min of treadmill exercise at 50% of maximal aerobic capacity (VO2max) in four subjects with hepatic cirrhosis and in four normal volunteers. Rates of endogenous glucose production (EGP) were measured using D-[6-3H]glucose infusions and fuel oxidation using indirect calorimetry. In the basal state, cirrhotic subjects had similar rates of EGP compared with controls. Forearm release of alanine and lactate was significantly greater in cirrhotic subjects (P less than 0.05), suggesting increased basal rates of gluconeogenesis. During exercise, EGP increased 2- to 2.5-fold in control subjects (P less than 0.01) but did not increase in cirrhotic subjects. Despite lower glucose concentrations in cirrhotic subjects, progressive hypoglycemia did not occur during exercise, probably because cirrhotic subjects demonstrated increased plasma concentrations of fat-derived substrates and derived a greater percentage of total energy requirement from fat oxidation than did controls (P less than 0.05) and because forearm muscle glucose extraction was significantly lower in cirrhotic subjects compared with controls (0.5 vs. 3.6%, respectively; P less than 0.05). During recovery, control subjects demonstrated significant increases in EGP rates compared with both the basal and exercise periods, but cirrhotic subjects showed no increase. In conclusion, cirrhotic subjects failed to demonstrate the normal increase in EGP during and after exercise. Significant hypoglycemia during exercise did not occur, possibly because of the increased availability of fat-derived fuels, which may spare the requirement for circulating glucose as an oxidative fuel for exercising muscle tissues.     

Change in cough reflex after treatment with enalapril and ramipril.

OBJECTIVE--To find out whether enalapril or ramipril causes the sensitivity of the cough reflex to change or symptomatic cough to develop in patients with hypertension. DESIGN--Prospective, placebo controlled, double blind, randomised crossover study. SETTING--Academic units of clinical pharmacology and medicine. PATIENTS--20 Patients (nine men and 11 women) who needed to take angiotensin converting enzyme inhibitors to control hypertension. INTERVENTIONS--All patients received enalapril 10 mg daily, ramipril 10 mg daily, or placebo daily for one week in random order, with a washout period of at least one week between treatments. For assessment of sensitivity of the cough reflex the patients inhaled various concentrations of capsaicin solution in random order. MAIN OUTCOME MEASURES--Measurement of the doses of capsaicin required to cause two or more and five or more coughs or the development of a symptomatic cough. RESULTS--Blood pressure, symptoms of cough, and the sensitivity of the cough reflex to inhaled capsaicin were recorded at the start of the study and before and at the end of each treatment period. Plasma urea and creatinine concentrations and angiotensin converting enzyme activity were measured at the start of the study and the end of each treatment period. Data were analysed by two way analysis of variance. Mean blood pressure was 159/97 mm Hg at the start of the study and 152/92, 143/88, and 147/86 mm Hg after treatment with placebo, enalapril, and ramipril respectively. Mean (SE) plasma angiotensin converting enzyme activity was 2.2 (0.2) mmol/l/h after treatment with placebo and fell significantly to 1.3 (0.1) mmol/l/h and to 0.4 (0.1) mmol/l/h after treatment with enalapril and ramipril respectively. No patient complained of cough while taking placebo but three women complained of cough when taking both enalapril and ramipril. The mean (95% confidence interval) lowest dose of capsaicin causing two or more coughs was 2.4 (1.5 to 4.0), 1.8 (1.12 to 2.82), and 2.2 (1.7 to 3.0) nmol after treatment with placebo, enalapril, and ramipril respectively; none of these changes were significant. The lowest dose of capsaicin causing five or more coughs was 18.9 (13.9 to 25.8), 14.4 (8.4 to 24.5), and 15.3 (10.8 to 21.2) nmol respectively; none of these changes were significant. The three patients who complained of cough had normal sensitivity to capsaicin after treatment with placebo but had a considerably increased sensitivity after treatment with enalapril and ramipril. CONCLUSIONS--Both enalapril and ramipril increase the sensitivity of the cough reflex appreciably in patients who complain of cough during treatment, but they do not change the se     

Genetic and morphological divergence among sympatric canids

Numerous studies have suggested that the extent of character divergence observed between two sympatric species reflects the intensity of competition for resources or space. However, the influence of time on divergence is often overlooked. We examined the relationship between time and character divergence in two groups of congeneric, sympatric canids on two continents: South American foxes and African jackals. Character divergence was assessed from measurements of body mass and dental and cranial shape. Divergence time was estimated from data on mitochondrial DNA restriction site polymorphisms. Our findings indicate that African jackals are morphologically similar despite having diverged more than 2 million years ago. By contrast, South American foxes differ substantially in both size and morphology after only 250,000 years of evolution. Thus, the lack of character divergence among the African jackals cannot be explained as a result of very recent common ancestry.     

Somatostatin gene expression in the thymus gland

A complex pattern of interactions appears to exist between the immune and neuroendocrine systems. Recently, vasopressin, oxytocin and vasoactive intestinal peptide have been isolated from the thymus. Using a rat somatostatin antisense RNA probe we have demonstrated expression of the somatostatin gene in the rat thymus. Furthermore, we have shown that the levels of thymic somatostatin mRNA exhibit a bell-shaped response to dexamethasone administration. Lipocortin I and II antisense RNA probes have been used as a positive control for the effects of the dexamethasone. We would suggest that somatostatin acts in the thymus in a paracrine mode to modulate T lymphocyte development.     

Hormonal stimulation of tyrosinase activity in human foreskin organ cultures

Summary A human foreskin organ culture system has been developed to study the response of human skin to hormonal stimulation. Foreskins are maintained in culture on floating plastic supports which allows the epidermal surface to be exposed to air while the dermis is bathed in nutrient medium. Both black and white human foreskins can be maintained in organ culture for at least 1 wk with no change in the tissue structure or cell viability as determined by histochemical staining and by dopa reaction staining. Tyrosinase activity in both black and white human foreskin cultures decays markedly during the first 2 d of culture to a new steady state level which remains stable throughout the culture period. Both black and white foreskin cultures consistently demonstrate 2- to 10-fold increases in tyrosinase activity when treated with theophylline (1 mM). Approximately 90% of all skin cultures examined showed an increase in enzyme activity when treated with this phosphodiesterase inhibitor. Dibutyryl cAMP (0.1 mM) and [Nle⁴, D-phe⁷]-alpha MSH (10⁻⁸ M), were also found to markedly stimulate tyrosinase activity in some skin cultures, whereas alpha-MSH and prostaglandin E₁ produced only an inconsistent and small increase in the activity of the enzyme. Histamine (1 μM), vitamin D₃ (1 μM), and retinoic acid (1μM) failed to stimulate tyrosinase activity in either white or black foreskin cultures. This hormone-responsive organ culture system can be utilized to characterize the molecular processes responsible for the regulation of tyrosinase and pigmentation in human skin. This work was supported by a research contract from the Oklahoma Center for the Advancement of Science and Technology (OCAST) and by a research grant from the Presbyterian Health Foundation.     

Ventilatory effects of substance P, vasoactive intestinal peptide, and nitroprusside in humans.

Animal studies suggest that the neuropeptides, substance P and vasoactive intestinal peptide (VIP), may influence carotid body chemoreceptor activity and that substance P may take part in the carotid body response to hypoxia. The effects of these peptides on resting ventilation and on ventilatory responses to hypoxia and to hypercapnia have been investigated in six normal humans. Infusions of substance P (1 pmol.kg-1.min-1) and of VIP (6 pmol.kg-1.min-1) were compared with placebo and with nitroprusside (5 micrograms.kg-1.min-1) as a control for the hypotensive action of the peptides. Both peptides caused significantly less hypotension than nitroprusside. Substance P and nitroprusside caused significantly greater increases in ventilation and in the hypoxic ventilatory response than VIP. No changes were seen in hypercapnic sensitivity. The stimulation of ventilation and the differential effects on ventilatory chemosensitivity that accompanied hypotension are consistent either with stimulation of carotid body chemoreceptor activity or with an interaction with peripheral chemoreceptor input to the respiratory center, as is seen in animals. The similar cardiovascular but different ventilatory effects of the peptides suggest that substance P may also stimulate the carotid body in a manner independent of the effect of hypotension. This is consistent with a role of substance P in the hypoxic ventilatory response in humans.     

Allele-specific suppression of a defective trans-Golgi network (TGN) localization signal in Kex2p identifies three genes involved in localization of TGN transmembrane proteins.

Kex2 protease (Kex2p) and Ste13 dipeptidyl aminopeptidase (Ste13p) are required in Saccharomyces cerevisiae for maturation of the alpha-mating factor in a late Golgi compartment, most likely the yeast trans-Golgi network (TGN). Previous studies identified a TGN localization signal (TLS) in the C-terminal cytosolic tail of Kex2p consisting of Tyr-713 and contextual sequences. Further analysis of the Kex2p TLS revealed similarity to the Ste13p TLS. Mutation of the Kex2p TLS results in transport of Kex2p to the vacuole by default. When expression of a GAL1 promoter-driven KEX2 gene is shut off in MAT(alpha) cells, the TGN becomes depleted of Kex2p, resulting in a gradual decline in mating competence which is greatly accelerated by TLS mutations. To identify the genes involved in localization of Kex2p, we isolated second-site suppressors of the rapid loss of mating competence observed upon shutting off expression of a TLS mutant form of Kex2p (Y713A). Seven of 58 suppressors were allele specific, suppressing point mutations at Tyr-713 but not deletions of the TLS or entire C-terminal cytosolic tail. By linkage analysis, the allele-specific suppressors defined three genetic loci, SOI1, S0I2, and S0I3. Pulse-chase analysis demonstrated that these suppressors increased net TGN retention of both Y713A Kex2p and a Ste13p-Pho8p fusion protein containing a point mutation in the Ste13p TLS. SOI1 suppressor alleles reduced the efficiency of localization of wild-type Kex2p to the TGN, implying an impaired ability to discriminate between the normal TLS and a mutant TLS. soi1 mutants also exhibited a recessive defect in vacuolar protein sorting. Suppressor alleles of S0I2 were dominant. These results suggest that the SOI1 and S0I2 genes encode regulators or components of the TLS recognition machinery.     

Developmental and Tissue-Specific Structural Alterations of the Cell-Wall Polysaccharides of Arabidopsis thaliana Roots

The plant cell wall is a dynamic structure that plays important roles in growth and development and in the interactions of plants with their environment and other organisms. We have used monoclonal antibodies that recognize different carbohydrate epitopes present in plant cell-wall polysaccharides to locate these epitopes in roots of developing Arabidopsis thaliana seedlings. An epitope in the pectic polysaccharide rhamnogalacturonan I is observed in the walls of epidermal and cortical cells in mature parts of the root. This epitope is inserted into the walls in a developmentally regulated manner. Initially, the epitope is observed in atrichoblasts and later appears in trichoblasts and simultaneously in cortical cells. A terminal [alpha]-fucosyl-containing epitope is present in almost all of the cell walls in the root. An arabinosylated (1->6)-[beta]-galactan epitope is also found in all of the cell walls of the root with the exception of lateral root-cap cell walls. It is striking that these three polysaccharide epitopes are not uniformly distributed (or accessible) within the walls of a given cell, nor are these epitopes distributed equally across the two walls laid down by adjacent cells. Our results further suggest that the biosynthesis and differentiation of primary cell walls in plants are precisely regulated in a temporal, spatial, and developmental manner.