Molecular evidence that oral supplementation with lycopene or lutein protects human skin against ultraviolet radiation: results from a double-blinded, placebo-controlled, crossover study.

BACKGROUND: Increasing evidence suggests photoprotection by oral supplementation with ß-carotene and lycopene. OBJECTIVES: To examine the capacity of lycopene-rich tomato nutrient complex (TNC) and lutein, to protect against ultraviolet (UV)A/B and UVA1 radiation at a molecular level. METHODS: In a placebo-controlled, double-blinded, randomized, crossover study two active treatments containing either TNC or lutein were assessed for their capacity to decrease the expression of UVA1 the radiation-inducible genes HO1, ICAM1 and MMP1. Sixty-five healthy volunteers were allocated to four treatment groups and subjected to a 2-week washout phase, followed by two 12-week treatment phases separated by another 2 weeks of washout. Volunteers started either with active treatment and were then switched to placebo, or vice versa. At the beginning and at the end of each treatment phase skin was irradiated and 24 h later biopsies were taken from untreated, UVA/B- and UVA1-irradiated skin for subsequent reverse transcriptase polymerase chain reaction analysis of gene expression. Moreover, blood samples were taken after the washout and the treatment phases for assessment of carotenoids. RESULTS: TNC completely inhibited UVA1- and UVA/B-induced upregulation of heme-oxygenase 1, intercellular adhesion molecule 1 and matrix metallopeptidase 1 mRNA, no matter the sequence (anova, P < 0·05). In contrast, lutein provided complete protection if it was taken in the first period but showed significantly smaller effects in the second sequence compared with TNC. CONCLUSIONS: Assuming the role of these genes as indicators of oxidative stress, photodermatoses and photoageing, these results might indicate that TNC and lutein could protect against solar radiation-induced health damage.

Prevalence of nosocomial pathogens in German ambulances: the SEKURE study.

OBJECTIVE: The increasing prevalence of multidrug resistant bacteria is a problem in the inpatient care setting, and in the emergency care system. The aim of this observational, cross-sectional study was to evaluate the prevalence of pathogens on well-defined surfaces in German ambulances that have been designated as 'ready for service'. METHODS: After informed consent was obtained, ambulance surfaces were sampled with agar plates for microbiological examination during an unannounced visit. A standardised questionnaire was used to obtain information regarding the disinfection protocols used at each rescue station. RESULTS: Methicillin resistant staphylococcus aureus contamination was present in 18 sampling surfaces from 11 out of 150 ambulance vehicles (7%) that were designated as ready for service. Contact surfaces directly surrounding patients or staff were most frequently contaminated with pathogens. However, bacterial contamination was not related to annual missions, methods or frequency of disinfection. CONCLUSIONS: In accordance with previous studies, disinfection and cleaning of areas with direct contact to patients or staff seem to be the most challenging. This should also be reflected in disinfection guidelines and the related continuing education.

Conformational analysis of green apple flavour: the gas-phase structure of ethyl valerate validated by microwave spectroscopy.

We report on the microwave spectrum of ethyl valerate, C(4)H(9)-COO-C(2)H(5), observed by molecular beam Fourier transform microwave spectroscopy (MB-FTMW). Highly accurate rotational and centrifugal distortion constants of the two most abundant conformers were determined. Their structure was investigated by comparison of the experimental rotational constants with those obtained by quantum chemical calculations. MP2/ 6-311++G(d,p) calculations failed to give a proper description of the observed conformers in the supersonic jet. Therefore, also other basis sets were tried using Hartree-Fock, second-order Møller-Plesset (MP2), and B3LYP methods to predict the observed gas-phase structures of the molecule. The quantum chemical results guided the conformer assignment of the rotational constants, obtained from the microwave experiment. One of the two observed conformers has C(s) symmetry, while the most abundant conformer has C(1) symmetry. The main conformer possesses a strong spectrum with high intensities. Additionally, harmonic frequency calculations at different levels of theory where carried out and a low lying vibration of the entire n-butyl group against the rest of the molecule was identified. The symbiotic interplay of microwave gas-phase investigations and quantum chemical calculations becomes evident in our results.

Plasma levels of vitamin E and carotenoids are decreased in patients with Nonalcoholic Steatohepatitis (NASH).

OBJECTIVES: Oxidative stress is suggested to play an important role in the pathogenesis of nonalcoholic steatohepatitis (NASH). The present study was aimed to compare plasma levels of antioxidants in patients suffering from NASH and healthy controls. METHODS: Plasma levels of the antioxidants α-tocopherol, γ-tocopherol, lutein, zeaxanthin, ß-cryptoxanthin, lycopene, α-carotene ß-carotene were determined in 57 patients with biopsy-proven NASH and 40 healthy controls. RESULTS: Levels of α-tocopherol (22.4 vs. 26.8 nmol/ ml; p<0.01), lutein (0.19 vs. 0.33 nmol/ml; p<0.0001), zeaxanthin (0.04 vs. 0.08 nmol/ml; p<0.0001), lyco?pene (0.15 vs. 0.42 nmol/ml; p<0.0001), α-carotene (0.03 vs. 0.06 nmol/ml; p<0.005) and ß-carotene (0.25 vs. 0.39 nmol/ml; p<0.01) were significantly decreased in NASH patients compared to controls. Age, aminotransferase status (ALT, AST) and BMI were not correlated with the levels of tocopherols or caro?tenoids. CONCLUSIONS: Given the decreased levels supplementation of lipophilic antioxidants might be a rational treatment option for patients with NASH.

Supplementation of flaxseed oil diminishes skin sensitivity and improves skin barrier function and condition.

BACKGROUND: Skin sensitivity is a common problem in the Western population correlated with changes of skin properties like skin barrier function, hydration and skin physiology. Skin properties can be modulated by dietary fatty acids (FA), especially poly-unsaturated FA. The present study was performed to evaluate the effect of daily supplementation with flaxseed oil and safflowerseed oil on healthy volunteers with sensitive skin. METHODS: The study was designed as a randomized, double-blind 12-week intervention with 2 female treatment groups (n = 13). Plasma FA profile, skin sensitivity, skin hydration, transepidermal water loss (TEWL) and skin surface were evaluated on day 0, week 6 and week 12. RESULTS: Supplementation with flaxseed oil led to significant decreases in sensitivity (after nicotinate irritation), TEWL, skin roughness and scaling, while smoothness and hydration were increased. Concomitantly, the ratio of n-6/n-3 FA in plasma decreased. Upon supplementation with safflowerseed oil, only a significant improvement in skin roughness and hydration was observed; however, the effects were less pronounced and determined at a later point in time than with flaxseed oil. The plasma n-6/n-3 FA ratio increased. CONCLUSION: The data provide evidence that daily intake of flaxseed oil modulates skin condition.

[Cardioprotection by thoracic epidural anesthesia? : meta-analysis]. / Kardioprotektion durch thorakale Periduralanästhesie? : Metaanalyse.

BACKGROUND: Thoracic epidural analgesia (EDA) is thought to provide cardioprotective effects in patients undergoing noncardiac surgery. The results of two previous meta-analysis showed controversial conclusions regarding the impact of EDA on perioperative survival. The purpose of the present meta-analysis was to evaluate, whether thoracic EDA has the potential to reduce perioperative cardiac morbidity or mortality on the basis of available randomized controlled trials. PATIENTS AND METHODS: A systematic literature search was conducted in medical databases (Med-Line, EBM-Reviews, Embase, Biosis and Biological Abstracts) and relevant clinical trials including patients undergoing noncardiac surgery were evaluated by two independent investigators. All randomized controlled trials investigating the effects of thoracic EDA on perioperative outcome, published from 1980 up to the end of 2008 were included into this quantitative systematic review. Calculations were performed using the statistics program Review Manager 4.1 using a fixed-effects model. RESULTS: Nine studies with a total of 2,768 patients were included in the meta-analysis. Thoracic EDA did not reduce perioperative mortality [odds ratio (Peto OR): 1.08; 95% confidence interval (CI) 0.74-1.58]. Patients receiving thoracic EDA demonstrated a tendency to a lower rate of perioperative myocardial infarction. However, this effect of thoracic EDA did not reach statistical significance (Peto OR: 0.65; 95% CI 0.4-1.05). CONCLUSIONS: The present meta-analysis did not prove any positive influence of thoracic EDA on perioperative in-hospital mortality in patients undergoing noncardiac surgery. Furthermore, it remains questionable if thoracic EDA has the potential to reduce the rate of perioperative myocardial infarction.

Carbon monoxide - beyond toxicity?

Carbon monoxide (CO) intoxication is one of the most frequent causes of accidental poisoning, mechanistically related to the inhibition of oxygen transport following blockage of the oxygen binding site of hemoglobin. However, it has become evident that CO is also a gaseous signaling molecule like nitric oxide and capable to trigger cellular stress responses in complex organisms. Endogenously, CO is synthesized upon degradation of heme by heme oxygenases (HOs) of which two enzymatically active isoenzymes are known in mammals; the stress-inducible HO-1 and the constitutively expressed HO-2. Among other pathways, HO-1 expression is stimulated by the Nrf2/Keap1 system which senses electrophilic compounds including alkylating agents and reactive oxygen species (ROS) such as superoxide or hydrogen peroxide. In context with ROS, HO-1 expression has been associated with antioxidant defense related to the heme-metabolite redox pair biliverdin/bilirubin. Studies on CO signaling were facilitated by the introduction of so called "CO releasing molecules" (CORMs), which allow for the controlled release of the compound in biological systems. Obviously, major biological targets of CO comprise intracellular heme-proteins such as cytochrome c oxidase of the respiratory chain, cytochrome P450-dependent monooxygenases (CYPs), or NADPH oxidases. From toxicological studies it is known that exposure to high amounts of CO provokes an inhibition of mitochondrial respiration and increased generation of ROS. In contrast, biological response to low amounts of CO comprises moderate mitochondrial uncoupling (proton leakage) due to the activation of channels including phosphate carrier (PiC), adenine nucleotide translocase (ANT) or large-conductance Ca2+-activated K+ channels (BKCa). Uncoupling of mitochondrial respiration from ATP production is accompanied by a loss of mitochondrial membrane potential - a key sensor and regulator of mitochondrial quality control and mitophagy. Inhibitory effects of CO on mitochondrial respiration are compensated by an increased glycolysis. However, on a short term, utilization of glucose is shifted to the pentose phosphate pathway, to provide NADPH for detoxification. It is notable that endogenous CO production is associated with the physiological response against exogenous electrophilic insult like Nrf2-dependent expression of phase II enzymes or glutathione synthesis. In contrast phase I enzymes such as CYPs which usually generate more electrophiles are inhibited by CO. Together with direct and indirect transient effects on energy metabolism and mitochondrial quality control CO may be an important regulator in cellular stress response.

Fine structural localization of potassium-stimulated rho nitrophenylphosphatase activity in denrites of the cerebral cortex.

A histochemical technique for the demonstration of K+-rho-nitrophenylphosphatase (K+-rhoNPPase) activity, a component of the Na+,K+-ATPase, has been applied at the fine structural level in the somatosensory cortex of the rat. Reaction product was consistenly found in dendrites and in association with the cytoplasmic aspect of the dendritic plasmalemma. Reaction product often filled portions of the tubular smooth endoplasmic reticulum in these processes. The results of these studies are interpreted to indicate that enzymatic activity is associated with large-and small-diameter dendrites. No convincing evidence of high activity was found in glial profiles. The importance of neurons and their dendrites in active transport of sodium and potassium ions in the cerebral cortex may be more significant than indicated by studies with isolated neurons and glia.

Origin of the gamma polypeptide of the Na+/K+-ATPase.

The Na+/K+-ATPase purified from lamb kidney contains a gamma polypeptide fraction which is a collection of fragments derived from the alpha and beta polypeptides of the enzyme. This fraction has the solubility characteristics of a proteolipid and was isolated either by high performance liquid chromatography (size exclusion chromatography) in 1% sodium dodecyl sulfate or by sequential organic extraction of purified lamb kidney Na+/K+-ATPase. Formation of gamma polypeptide(s) from detergent solubilized holoenzyme was accelerated by sulfhydryl containing reagents and was unaffected by addition of inhibitors of proteolytic enzymes. Treatment of the holoenzyme with the photoaffinity reagent N-(2-nitro-4-azidophenyl)[3H]ouabain ([3H]NAP-ouabain) labeled the alpha polypeptide and the gamma polypeptide fraction but not the beta polypeptide. Amino acid sequence analysis of one gamma polypeptide preparation revealed homology of one component of this fraction with the N-terminus of the beta subunit of the Na+/K+-ATPase. Amino acid analysis of two preparations of proteolipid showed similar amino acid compositions with a peptide derived from the alpha subunit. The insolubility and complexity of the gamma polypeptide(s)/proteolipid fraction appears to preclude a conclusive sequence analysis of all components of this fraction.

Incorporation of cis-parinaric acid, a fluorescent fatty acid, into synaptosomal phospholipids by an acyl-CoA acyltransferase.

The cis-isomer of parinaric acid, a naturally occurring C-18 polyene fatty acid, was incubated with brain subcellular fractions and the polarization of fluorescence increased in a time dependent manner. Greatest increases occurred in synaptosomal and microsomal membranes. This increase in polarization of fluorescence was found with the cis, but not the trans, isomer of parinaric acid and required Mg2+ or Ca2+ and was stimulated by coenzyme A and ATP. Synaptosomes were incubated with cis-parinaric acid and lipids were extracted and examined by high performance liquid chromatography. The highest incorporations of cis-parinaric acid were found in phosphatidylcholine (71%) and phosphatidylethanolamine (20%) while only traces were found in phosphatidylserine and phosphatidylinositol. [3H]Oleic acid was also incorporated into membrane phospholipids and unlabeled oleic acid blocked incorporation of cis-parinaric acid. It is proposed that cis-parinaric acid, like fatty acids normally found in brain, is incorporated into membrane phospholipids by an acyl-CoA acyltransferase. The presence of this enzyme in nervous tissue may make it possible to easily introduce fluorescent fatty acid probes into membrane phospholipids and to thereby facilitate study of membrane-mediated processes.

Interaction of a new fluorescent reagent with sulfhydryl groups of the (Na+ + K+)-stimulate ATPase.

The (Na+ + K+)-ATPase enzyme of rat brain microsomes can be reversibly inhibited by a new fluorescent sulfhydryl (SH) probe, dimethylaminoaphthalenecysteine-Hg+ (Dn-cys-Hg+). This reagent is more reactive than N-ethylamaleimide (MalNEt) toward membrane sulfhydryl groups. A procedure using the two SH reagents sequentially seems to permit a more selective labelling of the SH groups involved in the (Na+ + K+)-ATPase than is possible by using MalNEt alone. Brain microsomes treated by this method incorporate the fluorescent label within or near the active site of the enzyme. When the probe was bound a blue shift of its fluorescence emission maximum (from 540 to 495 nm) and a 20-fold increase in relative fluorescence occurred. This indicates that the Dn moiety is within a very non-polar region of the membrane.

Conformational changes of purified (Na+ + K+)-ATPase detected by a sulfhydryl fluorescence probe.

The fluorescent sulfhydryl reagent S-mercuric-N-dansyl cysteine (Dn-Cys-Hg+) has been used to label a purified preparation of the (Na+ + K+)-ATPase obtained from the electric organ of Electrophorous electricus. The labelled (Na+ +K+)-ATPase (ATP phosphohydrolase, EC 3.6.1.3), although reversibly inhibited, was capable of undergoing conformational changes associated with the active enzyme that could be monitored fluorometrically. The presence of ligands (Na+ + Mg2+ + ATP or Mg2+ + Pi) which are known to convert the enzyme from the E-1 state to the E-2-P state brought about large (97--100%) increases in fluorescence of the dimethylaminonaphthalene sulfonyl (Dn) label. An E-2 state could be achieved by the addition of Mg2+ which caused only a 32.3% increase in fluorescence over the E-1 state. Neither AMP nor TTP with or without Mg2+ or Na+ or Pi added without Mg2+ had any effect on the Dn fluorescence. If the enzyme was denatured, no fluorescence changes were observed. Small changes in the polarization of fluorescence of the Dn moiety were observed under all the conditions used. These small polarization changes and the large increases in the fluorescence intensity suggest that the enzyme can change conformational states in the presence of appropriate ligands and these conformational changes may take place in a relatively limited region of the protein's structure.

The insect brain (Na+ + K+)-ATPase. Binding of ouabain in the hawk moth, Manduca sexta.

(1) A quantitative study has been made of the binding of ouabain to the (Na+ + K+)-ATPase in homogenates prepared from brain tissue of the hawk moth, Manduca sexta. The results have been compared to those obtained in bovine brain microsomes. (2) The insect brain (Na+ + K+)-ATPase will bind ouabain either in the presence of Mg2+ and Pi, ('Mg2+, Pi' conditions) or in the presence of Na+, Mg2+, and an adenine nucleotide ('nucleotide' conditions) as is the case for the bovine brain (Na+ + K+)-ATPase. The binding conditions did not alter the total number of receptor sites measured at high ouabain concentrations in either tissue. (3) Potassium ion decreases the affinity (increases the KD) of ouabain to the M. sexta brain (Na+ + K+)-ATPase under both binding conditions. However, ouabain binding is more sensitive to K+ inhibition under the nucleotide conditions. In bovine brain ouabain binding is equally sensitive to K+ inhibition under both conditions. (4) The enzyme-ouabain complex has a rate of dissociation that is 10-fold faster in the M. sexta preparation than in the bovine brain preparation. Because of this, the M. sexta (Na+ + K+)-ATPase has a higher KD for ouabain binding and is less sensitive to inhibition by ouabain than the bovine brain enzyme. (5) This data supports the hypothesis that two different conformational states of the M. sexta (Na+ + K+)-ATPase can bind ouabain.

Immunoreactivity of subunits of the (Na+ + K+)-ATPase. Cross-reactivity of the alpha, alpha + and beta forms in different organs and species.

The immunologic cross-reactivity of the alpha and alpha + forms of the large subunit and the beta subunit of the (Na+ + K+)-ATPase from brain and kidney preparations was examined using rabbit antiserum prepared against the purified holo lamb kidney enzyme. As previously reported by Sweadner ((1979) J. Biol. Chem. 254, 6060-6067) phosphorylation of the large subunit of the (Na+ + K+)-ATPase in the presence of Na+, Mg2+, and [gamma-32P]ATP revealed that dog and, very likely, rat brain contain two forms of the large subunit (designated alpha and alpha +) while dog, rat, and lamb kidney contain only one form (alpha). The cross-reactivity of the alpha and alpha + forms in these preparations was investigated by resolving the subunits by SDS-polyacrylamide gel electrophoresis. The separated polypeptides were transferred to unmodified nitrocellulose paper and reacted with rabbit anti-lamb kidney serum, followed by detection of the antigen-antibody complex with 125I-labeled protein A and autoradiography. By this method, the alpha and alpha + forms of rat and dog brain, as well as the alpha form found in kidney, were shown to cross-react. In addition, membranes from human cerebral cortex were shown to contain two immunoreactive bands corresponding to the alpha and alpha + forms of dog brain. In contrast, the brain of the insect Manduca sexta contains only one immunoreactive polypeptide with a molecular weight intermediate to the alpha and alpha + forms of dog brain. The beta subunit from lamb, dog and rat kidney and from dog and rat brain cross-reacts with anti-lamb kidney (Na+ + K+)-ATPase serum. The mobility of the beta subunit from dog and rat brain on SDS-polyacrylamide electrophoresis gels is greater than the mobility of the beta subunit from lamb, rat or dog kidney.

Antioxidant defense: vitamins E and C and carotenoids.

Reactive oxygen species are thought to be implicated in the pathogenesis of various human diseases. They are generated endogenously under physiological and pathological conditions but also upon exposure to exogenous challenge. The organism maintains defense systems against reactive oxygen species, including enzymes and low-molecular-weight antioxidants. Important antioxidants such as vitamins E and C and carotenoids are provided from the diet. Vitamin E, as the major chain-breaking antioxidant, inhibits lipid peroxidation, thus preventing membrane damage and modification of low-density lipoproteins. It is regenerated by the water-soluble vitamin C. Carotenoids efficiently scavenge singlet molecular oxygen and peroxyl radicals. There is increasing evidence from epidemiological studies, animal experiments, and in vitro investigations that an increased intake of antioxidants is associated with a diminished risk for several diseases.

Macular carotenoids: lutein and zeaxanthin.

The yellow color of the macula lutea is due to the presence of the carotenoid pigments lutein and zeaxanthin. In contrast to human blood and tissues, no other major carotenoids including Beta-carotene or lycopene are found in this tissue. The macular carotenoids are suggested to play a role in the protection of the retina against light-induced damage. Epidemiological studies provide some evidence that an increased consumption of lutein and zeaxanthin with the diet is associated with a lowered risk for age-related macular degeneration, a disease with increasing incidence in the elderly. Protecting ocular tissue against photooxidative damage carotenoids may act in two ways: first as filters for damaging blue light, and second as antioxidants quenching excited triplet state molecules or singlet molecular oxygen and scavenge further reactive oxygen species like lipid peroxides or the superoxide radical anion.

Localization of insulin receptor mRNA in rat brain by in situ hybridization.

Insulin receptor mRNA was demonstrated in rat brain slices by in situ hybridization with three 35S-oligonucleotide probes and contact film autoradiography. Specificity was confirmed by showing that (a) excess unlabeled probe abolished the signal, (b) an oligonucleotide probe for rat neuropeptide Y mRNA showed a different distribution of hybridization signal, and (c) the distribution of insulin receptor binding was consistent with the distribution of insulin receptor mRNA. Insulin receptor mRNA was most abundant in the granule cell layers of the olfactory bulb, cerebellum and dentate gyrus, in the pyramidal cell body layers of the pyriform cortex and hippocampus, in the choroid plexus and in the arcuate nucleus of the hypothalamus.

Vitamin D influences gap junctional communication in C3H/10T 1/2 murine fibroblast cells.

Vitamin D3, cholecalciferol, induces cell-cell communication via gap junctions in murine fibroblasts (C3H/10T 1/2 cells) at concentrations between 0.01 and 1.0 microM, as assayed by the dye transfer method using Lucifer yellow CH. The extent of induction is similar to that obtained with the positive controls, canthaxanthin or retinoic acid, applied at 10 and 1 microM, respectively. Vitamin D2 also induces cell-cell communication. At elevated concentrations of vitamin D3 (5 microM) there is a suppression of gap junctional communication, reversible upon exposure to all-trans retinoic acid (1 microM) after removal of vitamin D3 from the medium. Conversely, communication between cells prestimulated with retinoic acid (1 microM) rapidly decreases when the retinoid is replaced by vitamin D3 (5 microM). The data demonstrate a role for vitamin D in the regulation of intercellular communication. This novel property of vitamin D may contribute to the antiproliferative effects of vitamin D exhibited in some types of cancer.

Profiles of antioxidants in human plasma.

The profile of antioxidants in biological fluids and tissues may be helpful in assessing oxidative stress in humans. Plasma antioxidants can be decreased as compared to established normal values, in abnormal or subnormal conditions, for instance as a consequence of disease-related free radical production. Alternatively, plasma antioxidants may be below the normal range due to insufficient dietary supply. Therefore, the profile of antioxidants can be of use only in conjunction with other parameters of the oxidative stress status. This article examines the profiles of plasma antioxidants in oxidative stress-related conditions, e.g., diabetes and some other diseases, as well as smoking and smoking cessation.

Carotenoid mixtures protect multilamellar liposomes against oxidative damage: synergistic effects of lycopene and lutein.

Antioxidant activity of carotenoids in multilamellar liposomes assayed by inhibition of formation of thiobarbituric acid-reactive substances was in the ranking: lycopene> alpha-tocopherol > alpha-carotene > beta-cryptoxanthin > zeaxanthin = beta-carotene > lutein. Mixtures of carotenoids were more effective than the single compounds. This synergistic effect was most pronounced when lycopene or lutein was present. The superior protection of mixtures may be related to specific positioning of different carotenoids in membranes.