Reactive oxygen species: from health to disease.

Upon reaction with electrons, oxygen is transformed into reactive oxygen species (ROS). It has long been known that ROS can destroy bacteria and destroy human cells, but research in recent decades has highlighted new roles for ROS in health and disease. Indeed, while prolonged exposure to high ROS concentrations may lead to non-specific damage to proteins, lipids, and nucleic acids, low to intermediate ROS concentrations exert their effects rather through regulation of cell signalling cascades. Biological specificity is achieved through the amount, duration, and localisation of ROS production. ROS have crucial roles in normal physiological processes, such as through redox regulation of protein phosphorylation, ion channels, and transcription factors. ROS are also required for biosynthetic processes, including thyroid hormone production and crosslinking of extracellular matrix. There are multiple sources of ROS, including NADPH oxidase enzymes; similarly, there are a large number of ROS-degrading systems. ROS-related disease can be either due to a lack of ROS (e.g., chronic granulomatous disease, certain autoimmune disorders) or a surplus of ROS (e.g., cardiovascular and neurodegenerative diseases). For diseases caused by a surplus of ROS, antioxidant supplementation has proven largely ineffective in clinical studies, most probably because their action is too late, too little, and too non-specific. Specific inhibition of ROS-producing enzymes is an approach more promising of clinical efficacy.

Airway geometry models of children's lungs for use in dosimetry modeling.

Single-path whole-lung and lobar models of the lungs of 11 children between 3 mo and 21 yr of age were developed based on a combination of cast data and published information on distal airway dimensions. The cast data used to generate these models were taken from one of the largest databases of actual measurements in children. The methods used to develop the children's models were based on techniques that have been used to develop adult single-path airway geometry models. Model dimensions for the conducting airways, as well as the estimated dead space, for all children fell within the range of the limited published information. Thus, the method for estimating airway dimensions in adults may be successfully applied to develop estimates of airway dimensions in children. The predicted total lung capacity (TLC) for the older children (aged 8 to 21 yr) fell within or near the range arising from published scaling equations. The assumptions used to generate the gas exchange region for children 8 yr and older produced results that were reasonably consistent with available physiological data. However, these assumptions do not result in a physiologically consistent gas exchange region for children 3 yr of age and younger; also, to maintain physiologically reasonable relationships between dead space and alveolar volume, the models for children 3 yr of age and younger resulted in predicted TLCs well below those predicted using published scaling equations. These discrepancies may be reflective of dysanaptic growth, in which the alveolar region is growing more rapidly than the airways. The results for children 3 yr of age and under suggest the need for a greater understanding of lung development during this critical period. This is particularly important considering the increasing evidence that exposure to pollutants and other toxicants and allergens during the first 2 yr of life may have long-term consequences on respiratory disease outcomes. Our results suggest that the geometry model airway dimensions for all ages are appropriate for use with dosimetry models, but dosimetry modelers need to assess carefully the reasonableness of TLC and functional residual capacity volumes to which airway dimensions are scaled for children 3 yr of age and under.

I = 0 (Information has no intrinsic meaning)

It helps identify some contradictions that can be found in the notion of knowledge management though its application is wider-ranging, and suggests that knowledge can scarcely be understood and managed even by ourselves, much less by means of sophisticated information and communications technologies.

Modeling age-related particle deposition in humans.

Inhalation of airborne material poses a potential health risk to various subpopulations one of which is children. Little is known about the fate of particles in the respiratory tracts of children. Modeling efforts have been limited due largely to the lack of adequate information on lung geometry during growth. Lung morphometry measurements in children and adults between 3 months and 21 years of age were used to create 5-lobe lung geometries. Each lobe had a dichotomous, symmetric branching structure and was structurally different from the other lobes. The lung geometries were used in a multiple-path particle deposition model to calculate particle deposition fractions in different regions, lobes and airway generations of the lungs. Simulated breathing patterns were representative of resting breathing. Age-dependent, semi-empirical expressions of particles losses in the nasal airways, which were based on fits to the available experimental measurements, showed larger nasal deposition in adults than in children. Predicted tracheobronchial deposition patterns were similar among different ages for a given particle size. In the alveolar region, the predicted deposition fraction varied with age such that a clear trend could not be identified. Deposition fraction in a lobe was proportional to the volume of air going to that lobe. Deposition fractions in the lower left and right lobes were similar but higher than those in the other lobes for a given particle diameter. Lobar deposition fraction adjusted for lobar lung volume or lung deposition fraction adjusted for lung volume was found to be a unique property for an individual and presented a means for age-dependent deposition comparisons. The adjusted tracheobronchial and pulmonary deposition fractions were greatest for infants and decreased with age. A similar trend was also observed for deposition fraction per unit area as a function of airway generation. The distribution of particle deposition fraction per unit surface area varied with particle size for an individual, with ultrafine particles being more uniformly distributed throughout the lungs and coarse particles depositing primarily in the first few tracheobronchial airways. The trend of particle deposition with age indicates that children, particularly infants, may be at a greater health risk from exposure to airborne particulate matter and noxious materials all other conditions being equal. The age-dependent predicted deposition fraction pattern per unit area of different size particles has implications in the calculation of inhaled reference concentrations as well as site-specific delivery of drugs and other therapeutic compounds to the lungs of patients.

Dose response for formaldehyde-induced cytotoxicity in the human respiratory tract.

Human studies of the sensory irritant effects of formaldehyde are complicated by the subjective nature of some clinical endpoints. This limits the usefulness of such studies for quantitative noncancer risk assessment of airborne formaldehyde. Objective measures of the noncancer effects of formaldehyde, such as the rate of regenerative cellular proliferation (RCP) secondary to cytolethality, can be obtained from laboratory animals but present the challenge of interspecies extrapolation of the data. To the extent that uncertainties associated with this extrapolation can be reduced, however, dose-response data obtained in laboratory animals are a viable alternative to clinical studies. Here, we describe the extrapolation of dose-response data for RCP from F344 rats to humans. Rats inhaled formaldehyde (0, 0.7, 2.0, 6.0, 10, and 15 ppm) 6 h/day, 5 days/week for up to 2 years. The dose response for RCP was J-shaped, with the rates of RCP at 0.7 and 2.0 ppm below but not statistically different from control, while the rates at the higher concentrations were significantly greater than control. Both the raw J-shaped data and a hockey-stick-shaped curve fitted to the raw data were used for predicting the human dose response for RCP. Cells lining the nasal airways of F344 rats and rhesus monkeys are comparably sensitive to the cytolethal effects of inhaled formaldehyde, suggesting that the equivalent human cells are also likely to be comparably sensitive. Using this assumption, the challenge of rat-to-human extrapolation was reduced to accurate prediction of site-specific flux of formaldehyde from inhaled air into the tissue lining the human respiratory tract. A computational fluid dynamics model of air flow and gas transport in the human nasal airways was linked to a typical path model of the human lung to provide site-specific flux predictions throughout the respiratory tract. Since breathing rate affects formaldehyde dosimetry, cytotoxicity dose-response curves were predicted for three standard working levels. With the most vigorous working level, the lowest concentrations of formaldehyde predicted to exert any cytotoxic effects in humans were 1.0 and 0.6 ppm, for the J-shaped and hockey-stick-shaped RCP curves, respectively. The predicted levels of response at the lowest effect concentrations are smaller than can be measured clinically. Published literature showing that the cytotoxicity of inhaled formaldehyde is related to exposure level rather than to duration of exposure suggests that the present analysis is a reasonable basis for derivation of standards for continuous human exposure.

Calcium-activated potassium channels mask vascular dysfunction associated with oxidized LDL exposure in rabbit aorta.

Endothelium-dependent vasodilation is impaired in atherosclerosis. Oxidized low density lipoprotein (ox-LDL) plays an important role, possibly through alterations in G-protein activation. We examined the effect of acute exposure to ox-LDL on the dilator responses of isolated rabbit aorta segments. We sought also to evaluate the specificity of this dysfunction for dilator stimuli that traditionally operate through a Gi-protein mechanism. Aortic segments were prepared for measurement of isometric tension. After contraction with prostaglandin F2alpha, relaxation to thrombin, adenosine diphosphate (ADP), or the endothelium-independent agonists, sodium nitroprusside (SNP) or papaverine was examined. Maximal relaxation to thrombin was impaired in the presence of ox-LDL (17.7+/-3.7% p<0.05) compared to control (no LDL) (52.6+/-4.0%). Ox-LDL did not affect maximal relaxation to ADP or SNP. However, in the presence of charybdotoxin (CHTX: calcium-activated potassium channel inhibitor) ox-LDL impaired relaxation to ADP (17.4+/-3.2%). CHTX did not affect control (no LDL) responses to ADP (69.6+/-5.0%) or relaxation to thrombin or papaverine. In conclusion, ox-LDL impairs relaxation to thrombin, but in the case of ADP, calcium-activated potassium channels compensate to maintain this relaxation.

Dosimetry modeling of inhaled formaldehyde: binning nasal flux predictions for quantitative risk assessment.

Interspecies extrapolations of tissue dose and tumor response have been a significant source of uncertainty in formaldehyde cancer risk assessment. The ability to account for species-specific variation of dose within the nasal passages would reduce this uncertainty. Three-dimensional, anatomically realistic, computational fluid dynamics (CFD) models of nasal airflow and formaldehyde gas transport in the F344 rat, rhesus monkey, and human were used to predict local patterns of wall mass flux (pmol/[mm(2)-h-ppm]). The nasal surface of each species was partitioned by flux into smaller regions (flux bins), each characterized by surface area and an average flux value. Rat and monkey flux bins were predicted for steady-state inspiratory airflow rates corresponding to the estimated minute volume for each species. Human flux bins were predicted for steady-state inspiratory airflow at 7.4, 15, 18, 25.8, 31.8, and 37 l/min and were extrapolated to 46 and 50 l/min. Flux values higher than half the maximum flux value (flux median) were predicted for nearly 20% of human nasal surfaces at 15 l/min, whereas only 5% of rat and less than 1% of monkey nasal surfaces were associated with fluxes higher than flux medians at 0.576 l/min and 4.8 l/min, respectively. Human nasal flux patterns shifted distally and uptake percentage decreased as inspiratory flow rate increased. Flux binning captures anatomical effects on flux and is thereby a basis for describing the effects of anatomy and airflow on local tissue disposition and distributions of tissue response. Formaldehyde risk models that incorporate flux binning derived from anatomically realistic CFD models will have significantly reduced uncertainty compared with risk estimates based on default methods.

Dosimetry modeling of inhaled formaldehyde: the human respiratory tract.

Formaldehyde (HCHO), which has been shown to be a nasal carcinogen in rats and mice, is used widely and extensively in various manufacturing processes. Studies in rhesus monkeys suggest that the lower respiratory tract may be at risk and some epidemiologic studies have reported an increase in lung cancer associated with HCHO; other studies have not. Thus, an assessment of possible human risk to HCHO exposure based on dosimetry information throughout the respiratory tract (RT) is desirable. To obtain dosimetry estimates for a risk assessment, two types of models were used. The first model (which is the subject of another investigation) used computational fluid dynamics (CFD) to estimate local fluxes in a 3-dimensional model of the nasal region. The subject of the present investigation (the second model) applied a 1-dimensional equation of mass transport to each generation of an adult human symmetric, bifurcating Weibel-type RT anatomical model, augmented by an upper respiratory tract. The two types of modeling approaches were made consistent by requiring that the 1-dimensional version of the nasal passages have the same inspiratory air-flow rate and uptake during inspiration as the CFD simulations for 4 daily human activity levels. Results obtained include the following: (1) More than 95% of the inhaled HCHO is predicted to be retained by the RT. (2) The CFD predictions for inspiration, modified to account for the difference in inspiration and complete breath times, are a good approximation to uptake in the nasal airways during a single breath. (3) In the lower respiratory tract, flux is predicted to increase for several generations and then decrease rapidly. (4) Compared to first pulmonary region generation fluxes, the first few tracheobronchial generations fluxes are over 1000 times larger. Further, there is essentially no flux in the alveolar sacs. (5) Predicted fluxes based on the 1-dimensional model are presented that can be used in a biologically based dose-response model for human carcinogenesis. Use of these fluxes will reduce uncertainty in a risk assessment for formaldehyde carcinogenicity.

Nitric oxide synthase inhibitors decrease coronary sinus-free radical concentration and ameliorate myocardial stunning in an ischemia-reperfusion model.

OBJECTIVES: Our objective was to determine the effect of a nitric oxide synthase inhibitor, NG-nitro-L-arginine (L-NNA) on free radical generation and myocardial contractility after ischemia-reperfusion. BACKGROUND: Cardiotoxic free radicals are generated by ischemia-reperfusion sequences. Nitric oxide reacts with superoxide radical to form peroxynitrite, which generates additional free radicals. Our hypothesis was that by inhibiting NO production, free radical formation will be diminished, which should be cardioprotective. METHODS: We studied 32 dogs. Coronary occlusion-reperfusion (20 min each) sequences were created by intracoronary balloon angioplasty inflation-deflation. Using electron paramagnetic resonance, we monitored the coronary sinus concentration of ascorbate free radical (Asc*-), a measure of total oxidative flux. The L-NNA (4.8 mg/kg total) was infused intravenously during occlusion-reperfusion; control dogs received saline. Immunohistochemical staining demonstrated the peroxynitration product nitrotyrosine. RESULTS: In the control dogs Asc*- rose from 3.2 +/- SD 0.5 nmol/l to 4.8 +/- 1.1 nmol/l with reperfusion, a 50% rise. With L-NNA the Asc*- rose from 3.2 +/- 0.9 nmol/l to 4.0 +/- 1.2 nmol/l, a 25% rise (p < 0.01, L-NNA vs. control). Echocardiographic left ventricular fractional area shortening (FAS) in the control dogs declined from 38 +/- 19% (baseline) to 26 +/- 14% (ischemia), and to 22 +/- 11% with reperfusion (p < 0.01 vs. baseline). With L-NNA, FAS declined from 36 +/- 13% (baseline) to 27 +/- 12% (ischemia) but then rose to 33 +/- 14 with reperfusion (p = NS vs. baseline). Nitrotyrosine was present in the myocardium subjected to ischemia-reperfusion, but almost absent in dogs receiving L-NNA. Myocardial perfusion was not altered by L-NNA. CONCLUSIONS: The NO synthase inhibitors decrease coronary sinus free radical concentration and ameliorate myocardial stunning after ischemia-reperfusion.

H(2)O(2)-induced O(2) production by a non-phagocytic NAD(P)H oxidase causes oxidant injury.

Non-phagocytic NAD(P)H oxidases have been implicated as major sources of reactive oxygen species in blood vessels. These oxidases can be activated by cytokines, thereby generating O(2), which is subsequently converted to H(2)O(2) and other oxidant species. The oxidants, in turn, act as important second messengers in cell signaling cascades. We hypothesized that reactive oxygen species, themselves, can activate the non-phagocytic NAD(P)H oxidases in vascular cells to induce oxidant production and, consequently, cellular injury. The current report demonstrates that exogenous exposure of non-phagocytic cell types of vascular origin (smooth muscle cells and fibroblasts) to H(2)O(2) activates these cell types to produce O(2) via an NAD(P)H oxidase. The ensuing endogenous production of O(2) contributes significantly to vascular cell injury following exposure to H(2)O(2). These results suggest the existence of a feed-forward mechanism, whereby reactive oxygen species such as H(2)O(2) can activate NAD(P)H oxidases in non-phagocytic cells to produce additional oxidant species, thereby amplifying the vascular injury process. Moreover, these findings implicate the non-phagocytic NAD(P)H oxidase as a novel therapeutic target for the amelioration of the biological effects of chronic oxidant stress.

Percutaneous sclerotherapy for congenital venous malformations in the extremities.

Between January 1991 and January 1998, a total of 15 patients underwent direct injection sclerotherapy for painful peripheral venous malformations. Duplex ultrasonography or venography was used in all cases for the detection and localization of tortuous venous structures. Direct injection with absolute ethyl alcohol was performed in 12 patients, and Sotradecol or sodium morrhuate was used in 5 patients. Provocative lidocaine testing was used in 2 patients in whom major nerves were in proximity to the malformations. All patients underwent follow-up in the clinic with duplex examination after each sclerotherapy. Clinical symptoms of all patients improved during average follow-up of 2.5 years (range: 3 months to 6 years.) Duplex examination was useful in detecting the venous component including the size and course of veins, which were often less well seen on magnetic resonance imaging. Duplex study was helpful in follow-up after sclerotherapy in all patients. Direct injection sclerotherapy is an acceptable treatment modality for venous malformations. Complications are manageable, and regular follow-up with Duplex is helpful.

Requirement of a centrosomal activity for cell cycle progression through G1 into S phase.

Centrosomes were microsurgically removed from BSC-1 African green monkey kidney cells before the completion of S phase. Karyoplasts (acentrosomal cells) entered and completed mitosis. However, postmitotic karyoplasts arrested before S phase, whereas adjacent control cells divided repeatedly. Postmitotic karyoplasts assembled a microtubule-organizing center containing gamma-tubulin and pericentrin, but did not regenerate centrioles. These observations reveal the existence of an activity associated with core centrosomal structures-distinct from elements of the microtubule-organizing center-that is required for the somatic cell cycle to progress through G1 into S phase. Once the cell is in S phase, these core structures are not needed for the G2-M phase transition.

Inhalation toxicology of urban ambient particulate matter: acute cardiovascular effects in rats.

Wistar rats were exposed for 4 hours by nose-only inhalation to clean air, resuspended Ottawa ambient particles (EHC-93*, 48 mg/m3), the water-leached particles (EHC-93L, 49 mg/m3), diesel soot (5 mg/m3), or carbon black (5 mg/m3). Continuous data for physiologic endpoints (heart rate, blood pressure, body temperature, animal's activity) were captured by telemetry before and after exposure. Blood was sampled from jugular cannulas 1 to 3 days before exposure and at 2 and 24 hours after exposure, and by heart puncture on termination at 32 hours (histology group) or 48 hours (telemetry group) after exposure. Lung injury was assessed by 3H-thymidine autoradiography after the rats were killed. We measured endothelins (plasma ET-1, big ET-1, ET-2, ET-3) to assess the vasopressor components; nitric oxide (NO)-related metabolites (blood nitrate, nitrite, nitrosyl compounds, and plasma 3-nitrotyrosine) to assess the vasodilator components; and catecholamines (epinephrine, norepinephrine, L-DOPA, dopamine) and oxidative stressors (m- and o-tyrosine) for additional insight into possible stress components. Lung cell labeling was uniformly low in all treatment groups, which indicates an absence of acute lung injury. Inhalation of EHC-93 caused statistically significant elevations (P < 0.05) of blood pressure on day 2 after exposure, plasma ET-1 at 32 hours after exposure, and ET-3 at 2, 32, and 48 hours after exposure. In contrast, the modified EHC-93L particles, from which soluble components had been extracted, did not affect blood pressure. The EHC-93L particles caused early elevation (P < 0.05) of the plasma levels of ET-1, ET-2, and ET-3 at 2 hours after exposure, but the endothelins returned to basal levels 32 hours after exposure. Exposure to diesel soot, but not carbon black, caused an elevation (P < 0.05) of plasma ET-3 at 36 hours after exposure; blood pressure was not affected by diesel soot. Our results indicate that inhalation of the urban particles EHC-93 can affect blood levels of ET-1 and ET-3 and cause a vasopressor response in Wistar rats without causing acute lung injury. Furthermore, the potency of the particles to influence hemodynamic changes appears to be modified by removing polar organic compounds and soluble elements. Because the pathophysiologic significance of elevated endothelins has been clinically established in humans, our observations suggest a novel mechanism by which inhaled particles may cause cardiovascular effects. These findings in rats contribute to the weight of evidence in favor of a biologically plausible epidemiologic association between ambient particulate matter and cardiovascular morbidity and mortality in human populations.

Haber's rule: a special case in a family of curves relating concentration and duration of exposure to a fixed level of response for a given endpoint.

The concept that the product of the concentration (C) of a substance and the length of time (t) it is administered produces a fixed level of effect for a given endpoint has been ascribed to Fritz Haber, who was a German scientist in the early 1900s. He contended that the acute lethality of war gases could be assessed by the amount of the gas in a cubic meter of air (i.e. the concentration) multiplied by the time in min that the animal had to breathe the air before death ensued (i.e. C x t=k). While Haber recognized that C x t=k was applicable only under certain conditions, many toxicologists have used his rule to analyze experimental data whether or not their chemicals, biological endpoints, and exposure scenarios were suitable candidates for the rule. The fact that the relationship between C and t is linear on a log-log scale and could easily be solved by hand, led to early acceptance among toxicologists, particularly in the field of entomology. In 1940, a statistician named Bliss provided an elegant treatment on the relationships among exposure time, concentration, and the toxicity of insecticides. He proposed solutions for when the log-log plot of C and t was composed of two or more rectilinear segments, for when the log-log plot was curvilinear, and for when the slope of the dosage-mortality curve was a function of C. Despite the fact that Haber's rule can underestimate or overestimate effects (and consequently risks), it has been used in various settings by regulatory bodies. Examples are presented from the literature of data sets that follow Haber's rule as well as those that do not. Haber's rule is put into perspective by showing that it is simply a special case in a family of power law curves relating concentration and duration of exposure to a fixed level of response for a given endpoint. Also shown is how this power law family can be used to examine the three-dimensional surface relating C, t, and varying levels of response. The time has come to move beyond the limited view of C and t relationships inferred by Haber's rule to the use of the broader family of curves of which this rule is a special case.

Clinical manifestations in a large hereditary hemorrhagic telangiectasia (HHT) type 2 kindred.

HHT type 2 (HHT 2) is a multi-system vascular dysplasia caused by a mutation in the ALK-1 gene, but the phenotype has not been well defined. We report on 51 members of an HHT 2 kindred with an ALK-1 gene mutation shown to be associated with the disorder. This ALK-1 mutation was detected in 38 kindred members who were evaluated systematically for associated vascular abnormalities. Pulmonary arteriovenous malformations (AVMs) were found in 6% of those screened, cerebral AVM in 7%, hepatic AVM in 17%, and spinal AVM in 3%. We discuss these and other findings in the 38 affected kindred members, as well as findings in the 13 kindred members in whom the mutation was not detected. This study shows that pulmonary, cerebral, spinal, and hepatic AVMs can all occur in HHT 2. It also adds to the evidence suggesting that pulmonary AVMs are more common in HHT 1 than in HHT 2. We identify a higher prevalence of hepatic AVMs than previously reported in either HHT 1 or 2. This may be specific to the mutation in this kindred, but probably reflects the lack of routine screening for this manifestation. Even in this family in which all affected individuals have the same mutation, the clinical manifestations of HHT and their severity varied tremendously. Intrafamilial variation in expression of HHT is clearly significant, emphasizing the difficulty in establishing the diagnosis in individuals and in sub-typing families when DNA testing is not available.

Activation of protein kinase C alters p34(cdc2) phosphorylation state and kinase activity in early sea urchin embryos by abolishing intracellular Ca2+ transients.

The p34(cdc2) protein kinase, a universal regulator of mitosis, is controlled positively and negatively by phosphorylation, and by association with B-type mitotic cyclins. In addition, activation and inactivation of p34(cdc2) are induced by Ca(2+) and prevented by Ca(2+) chelators in permeabilized cells and cell-free systems. This suggests that intracellular Ca(2+) transients may play an important physiological role in the control of p34(cdc2) kinase activity. We have found that activators of protein kinase C can be used to block cell cycle-related alterations in intracellular Ca(2+) concentration ([Ca(2+)](i)) in early sea urchin embryos without altering the normal resting level of Ca(2+). We have used this finding to investigate whether [Ca(2+)](i) transients control p34(cdc2) kinase activity in living cells via a mechanism that involves cyclin B or the phosphorylation state of p34(cdc2). In the present study we show that the elimination of [Ca(2+)](i) transients during interphase blocks p34(cdc2) activation and entry into mitosis, while the elimination of mitotic [Ca(2+)](i) transients prevents p34(cdc2) inactivation and exit from mitosis. Moreover, we find that [Ca(2+)](i) transients are not required for the synthesis of cyclin B, its binding to p34(cdc2) or its destruction during anaphase. However, in the absence of interphase [Ca(2+)](i) transients p34(cdc2) does not undergo the tyrosine dephosphorylation that is required for activation, and in the absence of mitotic [Ca(2+)](i) transients p34(cdc2) does not undergo threonine dephosphorylation that is normally associated with inactivation. These results provide evidence that intracellular [Ca(2+)](i) transients trigger the dephosphorylation of p34(cdc2) at key regulatory sites, thereby controlling the timing of mitosis entry and exit.