Direct monitoring of UV-induced free radical generation in HaCaT keratinocytes.

BACKGROUND: Ultraviolet radiation (UVR) is one of the most important aetiological factors in the development of skin cancer, with an estimated 100,000 new cases of nonmelanoma skin cancer (NMSC) diagnosed each year in the UK. To date, little work has been carried out to investigate the role of UVR in the increased production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) following exposure of skin cells to simulated solar UVR. AIM: To monitor directly the effects of simulated solar UVR on ROS and RNS generation in HaCaT keratinocytes. METHODS: This study reports the use of electrochemical monitoring techniques for the direct, real-time detection of two highly reactive free radical species, superoxide (O2-) and nitric oxide (NO), from HaCaT keratinocyte cells that had been exposed to a source of UVR designed to simulate the doses of UVA and UVB found in solar light. RESULTS: An increase in both O2- and NO generation was observed in HaCaT cells that had been exposed to UVR. No detectable increase in either species was observed in cells that had not been exposed to UVR. The specificity of the electrochemical methods for O2- or NO was confirmed through the scavenging or inhibition of these species. CONCLUSION: The findings of this study demonstrated that exposure of HaCaT cells to relatively low doses of UVR resulted in the immediate generation of both O2- and NO, therefore potentially leading to the downstream generation of highly damaging metabolites and the development of a number of pathologies, including cancer.

Using the fast orthogonal search with first term reselection to find subharmonic terms in spectral analysis.

The fast orthogonal search (FOS) algorithm has been shown to accurately model various types of time series by implicitly creating a specialized orthogonal basis set to fit the desired time series. When the data contain periodic components, FOS can find frequencies with a resolution greater than the discrete Fourier transform (DFT) algorithm. Frequencies with less than one period in the record length, called subharmonic frequencies, and frequencies between the bins of a DFT, can be resolved. This paper considers the resolution of subharmonic frequencies using the FOS algorithm. A new criterion for determining the number of non-noise terms in the model is introduced. This new criterion does not assume the first model term fitted is a dc component as did the previous stopping criterion. An iterative FOS algorithm called FOS first-term reselection (FOS-FTR), is introduced. FOS-FTR reduces the mean-square error of the sinusoidal model and selects the subharmonic frequencies more accurately than does the unmodified FOS algorithm.

Immune response to an 18-kilodalton outer membrane antigen identifies lipoprotein 20 as a Helicobacter pylori vaccine candidate.

Experiments were performed using the standardized murine model of Helicobacter pylori infection to determine the immunogenicity of H. pylori outer membrane vesicles in immune protection. These vesicles, which are naturally shed from the surface of the bacterium, induce a protective response when administered intragastrically to mice in the presence of cholera holotoxin, despite the absence of the urease enzyme and associated Hsp54 chaperonin. Immunoblotting identified a specific serum immunoglobulin G (IgG) response to an 18-kDa outer membrane protein in a significant number of immunized animals. This commonly expressed, immunodominant protein was subsequently identified as lipoprotein 20 (Lpp20). Hybridoma backpacks secreting an IgG1 subclass monoclonal antibody to Lpp20 were generated in H. pylori-infected mice and were found to significantly reduce bacterial numbers, providing evidence that this surface-exposed antigen is a true vaccine candidate and not merely an antigenic marker for successful, protective immunization.

[Langerhans cell histiocytosis and myelomonocytic leukemia: a non-fortuitous association]. / Histiocytose langerhansienne et leucémie de la lignée myélomonocytaire: une association non fortuite.

BACKGROUND: Langerhans cell histiocytosis is an uncommon clonal disorder. Its reactional or genetic nature is debated. CASE REPORT: Three patients aged 71, 75 and 73 years with Langerhans cell histiocytosis also developed myelomoncytic leukemia (AML4, AML5, and chronic myelomonocytic leukemia). In two cases, there was no prior treatment which could potentially induce leukemia. In the third case, AML4 occurred shortly (10 months) after initiation of a vinblastin treatment. DISCUSSION: This pathogenic association suggests a common origin for these two conditions, corresponding to an anomalous pluripotent stem cell line. These cases provide further evidence favoring the hypothesis of a genetic origin rather than a reactional origin for Langerhans cell histiocytosis.

The characterization of ovine genes for atrial, brain, and C-type natriuretic peptides.

The natriuretic peptides (NPs) play an important role in the homeostasis of blood pressure and sodium balance in all mammals studied to date. Their combined actions on the vasculature, kidneys, and adrenals reduce blood pressure and intravascular volume. In order to provide sequence information about the ovine NP genes for our physiological studies in sheep, we have determined the genomic DNA sequence of each of the NPs; atrial NP, brain NP (BNP), and C-type NP using an ovine genomic library. Strong homology with other species was found for ovine peptide and genomic sequences of atrial NP and for C-type NP. Further, despite previous reports of poor conservation of BNP across species, the peptide sequence for ovine BNP was found to be identical to both the 26 amino acid-residue porcine BNP, and the 35 amino acid peptide known as bovine aldosterone secretion-inhibitory factor. This data also revealed strong homology of BNP mature forms in dog, cow, pig, and sheep, thus permitting the use of porcine antisera to study BNP-level changes in sheep models of cardiac failure. This conservation of the BNP gene sequence suggests that BNP, like the other NPs, plays an important role in mammalian physiology.

Amino-terminal proBNP in ovine plasma: evidence for enhanced secretion in response to cardiac overload.

We have recently identified a novel amino-terminal fragment of pro-brain natriuretic peptide (NT-proBNP) in the circulation of humans, the concentration of which increases progressively as the left ventricle fails. To clarify the origins of NT-proBNP in experimental animals, we have developed an RIA for NT-proBNP based on residues 52-71 of ovine proBNP-(1-103) and used it to study cardiac processing, secretion, and metabolism of BNP in sheep with cardiac overload induced by coronary artery ligation (CAL) or rapid left ventricular pacing (rLVP). The concentration of NT-proBNP in left atrial plasma extracts drawn from normal control sheep was threefold that of mature BNP. Size-exclusion and reverse-phase HPLC analyses of plasma extracts coupled to RIA revealed a single peak of immunoreactive (ir) NT-proBNP [ approximately 8,000 relative molecular weight (Mr)], quite distinct from a single peak of ir-mature BNP ( approximately 3,000 Mr). In contrast, ovine cardiac tissue contained only a single immunoreactive peak of high-molecular-weight BNP ( approximately 11,000 Mr), consistent in size with proBNP-(1-103). Sampling from the cardiac coronary sinus in normal control sheep (n = 5) and sheep with CAL (n = 5) revealed that the molar ratio of NT-proBNP to mature BNP was similar. There was a significant gradient of both mature and NT-proBNP across the heart in normal sheep, whereas after CAL the gradient was significant for mature BNP only. In both forms of cardiac overload (CAL and rLVP), left atrial plasma levels of NT-proBNP were significantly increased above normal levels, in contrast with mature BNP levels, which were raised only in the rLVP group of animals. Blockade of natriuretic peptide metabolism in sheep with heart failure (induced by rLVP) raised mature BNP levels threefold but did not affect levels of NT-proBNP. In conclusion, these studies show that NT-proBNP is formed from proBNP stores during secretion and, compared with mature BNP, accumulates in plasma because metabolism of NT-proBNP appears to differ from that of mature BNP. Although its function, if any, remains unclear, plasma NT-proBNP may prove to be a sensitive marker of cardiac overload and/or decompensation.

Nutrition and diabetes: putting guidelines into practice.

Dietary recommendations for the management of diabetes have changed over the past 10 years. There is now a reduced emphasis on the importance of carbohydrate in the diabetic diet. The first move away from carbohydrate restriction came in 1982 when the British Diabetic Association published its 'Dietary recommendations for diabetics for the 1980s'. This document suggested that the diet for people with diabetes should be based on a high intake of complex carbohydrate and fibre, with a restriction of fat intake. This article discusses the impact of these recommendations on different nutrients, and demonstrates, through the use of a case study, how these can be put into practice.

Ovine brain natriuretic peptide in cardiac tissues and plasma: effects of cardiac hypertrophy and heart failure on tissue concentration and molecular forms.

Whereas numerous studies have examined the cardiac tissue content and secretion of atrial natriuretic peptide (ANP), the response of brain natriuretic peptide (BNP) in states of experimental cardiac overload is less well documented. Our recent partial cloning of the ovine BNP gene has enabled us to study changes in cardiac tissue concentration, together with tissue and circulating molecular forms of ANP and BNP, in response to cardiac overload induced by rapid ventricular pacing (n = 7) and aortic coarctation (n = 6). In normal sheep, although highest levels of BNP were found in atrial tissue (15-fold those of the ventricle), the BNP/ANP concentration ratio in the ventricles was 10- to 20-fold higher than the ratio calculated for atrial tissue. Compared with normal sheep, significant depletion of both ANP and BNP concentrations within the left ventricle occurred after rapid ventricular pacing. Size exclusion and reverse phase HPLC analysis of atrial and ventricular tissue extracts from normal and overloaded sheep showed a single peak of high molecular weight BNP consistent with the proBNP hormone. In contrast, immunoreactive BNP extracted from plasma drawn from the coronary sinus was all low molecular weight material. Further analysis of plasma BNP using ion exchange HPLC disclosed at least 3 distinct immunoreactive peaks consistent with ovine BNP forms 26-29 amino acid residues in length. These findings show that BNP is stored as the prohormone in sheep cardiac tissues and that complete processing to mature forms occurs at the time of secretion. The capacity to process the prohormone at secretion is not impaired by chronic heart failure.

The sites of gene expression of atrial, brain, and C-type natriuretic peptides in mouse fetal development: temporal changes in embryos and placenta.

Atrial (ANP), brain (BNP), and C-type natriuretic peptide (CNP) belong to a family of hormones important in blood pressure and sodium homeostasis. Expression of ANP has been reported in embryo hearts, but BNP and CNP expression during development has not been described. We used in situ hybridization to identify the sites of gene expression of ANP, BNP, and CNP during development in mouse embryos at daily intervals from midgestation. Very intense expression of ANP and BNP was visible in the heart from 9.5 days gestation; levels of expression of both peptides in the ventricle exceeded those in atria throughout gestation. There was a major peak of atrial and ventricular ANP and BNP expression at 12.5 days, attaining levels similar to those in adult heart and then declining until birth. Cardiac expression of CNP was undetectable. Expression of ANP and CNP was also observed in distinct sites in the brain, and all three peptides were expressed in the spinal cord. In mouse placenta, strong CNP expression was seen in the decidua basalis around the large maternal blood vessels, and BNP message was detected at the peripheral margin of the decidual layer. This pattern of expression indicates that ANP and CNP are present during development of the mouse central nervous system and suggests that CNP and BNP participate in regulating the maternal blood supply to the developing embryo.

Increased accuracy and precision of heparin and protamine dosing reduces blood loss and transfusion in patients undergoing primary cardiac operations.

Individual aspects of heparin or protamine dosing have been better controlled than previously as useful tests have become available. Although many variables including drug potency, drug source, and individual patient response have been separately identified, there has not been an attempt to integrate them into a single management strategy. This study was undertaken to learn whether more precise control of drug variables and patient response would affect blood loss and transfusion requirements. Adult patients having primary cardiac operations were prospectively randomized into two groups. A control group received heparin and protamine by conventional methods. The test group received heparin and protamine according to in vitro predictive tests integrating drugs, tests, and patient response. Supplemental protamine was given in this group only if heparin was specifically found by testing. Anticoagulation in all patients was maintained at an activated coagulation time greater than 400 seconds, and any other treatment for bleeding was at the discretion of the clinical team caring for the patients. Testing and treatment for both groups followed routine practice after patient arrival in the intensive care unit. Test patients received slightly more heparin and a markedly lower dose of protamine than the control patients. Testing identified patients with decreased heparin sensitivity (preoperative heparin therapy) and correctly predicted the effective heparin dose. Supplemental protamine was given twice as often to control patients and frequently when no heparin was detectable (retrospectively). Test patients exhibited less 24-hour chest tube drainage (671 ml versus 1298 ml) and fewer patients received transfusion (9/22 versus 18/24) with fewer donor exposures (22/22 versus 101/24). The management strategy used for heparin and protamine added accuracy and precision, which was associated with improved hemostasis. Although the observation is valid, the mechanism or mechanisms are not completely clear. Nevertheless, it is reasonable to apply basic pharmacologic principles and establishment of consistent, predictable protocols that are beneficial. It is against this background that the efficacy of additional drugs or equipment should be assessed. It is quite possible that only marginal if any improvement in hemostasis may be found in patients having primary, uncomplicated cardiac operation with the addition of more costly drugs or equipment.

Area-adaptive simulated annealing for image reconstruction and restoration.

Simulated annealing (SA) is a robust, stable, but computationally costly method for solving ill-posed image-restoration problems. We describe the use of a backprojection operator that identifies those regions of an object estimate that have the greatest likelihood of being in error at each step of the SA process. This reduces computational time by concentrating the computing effort of SA on those pixels most effective in reducing the reconstruction error. The performance of an area-adaptive SA algorithm is evaluated for the restoration of images blurred by a simple pillbox space-invariant and a biconical space-variant point-spread function typical of a depth-measuring optical system.

Prediction of atmospherically induced wave-front degradations.

Accurate prediction of the short-term future behavior of atmospherically distorted wave fronts would permit the elimination of delays inherent in current adaptive-optics systems. It is shown by using astronomical image data that atmospherically induced wave-front distortions as represented by time series of wave-front tips and tilts measured in the visible and piston values measured in the infrared are predictable to a degree that would be useful in an adaptive-optics system. Adaptive linear predictors as well as predictors based on the back-propagation neural network are employed in this study.

Evidence of a chaotic attractor in star-wander data.

The observed motion of stellar-image centroids is shown to have a chaotic attractor with a correlation dimension of ~6. The existence of a chaotic attractor in star wander, or equivalently in wave-front tilts, indicates that the atmospheric processes that cause image degradation may be more accurately described as chaotic, not so random as is usually assumed. This new result has important implications for the accurate modeling of atmospheric processes, the operation of adaptive optics systems, and the processing of stellar images.