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Vasogenic edema and VEGF expression in a rat two-vein occlusion model.

Vasogenic edema plays an important etiologic role in the pathogenesis of cerebral venous circulation disturbances (CVCDs). Since vascular endothelial growth factor (VEGF) is a major mediator in angiogenesis and vascular permeability, including induction of vasogenic edema, the present study was undertaken to investigate whether it has any relevance to CVCDs. Male Wistar rats (n = 15) were used. Two adjacent cortical veins were occluded photochemically using rose bengal dye and fiberoptic illumination, with evaluation 24 hours thereafter by magnetic resonance imaging (MRI). Each brain was removed from the skull immediately after MRI and processed for hematoxylin-eosin staining (H&E staining) of sections for histopathology and comparison with MRI data. VEGF expression as demonstrated immunohistochemically appeared to coincide with vasogenic edema, diagnosed as high intensity areas on apparent diffusion coefficient of water (ADCw) maps. On the basis of these data, we conclude that VEGF is related to formation of vasogenic edema in the acute stage of CVCD.

Animals↗

Extinction of cells of cyanobacterium Anabaena circinalis in the presence of humic acid under illumination.

Laboratory experiments targeting the effect of humic acid (HA) on the cell lysis of cyanobacterium Anabaena circinalis have been performed. Light irradiation was found to be an important factor for the cell lysis phenomenon, whereas intracellular hydrogen peroxide (H2O2) might be a chemical factor for the process. An exogenous H2O2 concentration of 1.0 mg l(-1) was determined as the threshold for cell survival. Our results indicated that HA or its possible product(s) of photochemical reaction can induce damage to intracellular catalase under artificial illumination, which leads intracellular H2O2 to be accumulated to an abnormally high concentration, eventually resulting in cell death. Moreover, H2O2 released into the culture from dead cells can damage other cells, which in turn brings about the population extinction.

Anabaena↗

Transient thiyl radicals in yeast copper(I) thionein.

In an EPR study employing yeast copper(I) thionein, GSH and Cu-GSH it was shown that thiyl radicals could be successfully generated from the thiolate sulfur via oxidation by photochemically formed superoxide at 77 K. The g-value was 2.036. Essentially no EPR detectable copper(II) was monitored under the experimental conditions, indicating that the oxidation reduction process is restricted to the thiolate sulfur. The Cu(I)-thiolate chromophores remained fully intact as deduced from chiroptical and luminescence measurements. Thus, copper thionein is supposed to be actively involved in the scavenging of oxygen free radicals by a reversible thiolate oxidation reduction cycle. The coordinated Cu(I) seems to serve as a prominent candidate to stabilize the transiently formed thiyl radical.

Carrier Proteins↗

Thermal activation of photoactivatable urocanase from Pseudomonas putida.

The dark inactivation of urocanase from Pseudomonas putida is caused by the formation of a sulfite adduct of the tightly bound coenzyme, nicotinamide adenine dinucleotide. Photodissociation of this adduct by UV radiation restores the enzyme activity. Based on cold exhaustive dialysis the modification reaction appeared to be irreversible. However, we now report that sulfite modification of urocanase is reversible at higher temperatures. An Arrhenius plot of the thermal activation is linear (20-38 degrees C). The activation energy for the enzyme activation is 114 kJ mol-1. The substance that is photodissociated from inactive urocanase reacts with urocanase to reform the modified enzyme indicating that sulfite is not oxidized, or otherwise changed through these processes. Nucleophiles (sulfite, hydroxylamine, hydride, cyanide) are known to inhibit urocanase by forming adducts with nicotinamide adenine dinucleotide. Urocanase inactivated by hydride or cyanide is not reactivated thermally or photochemically. Urocanase inactivated by hydroxylamine and by glycylglycine can be reactivated by a thermal reaction. In conclusion, sulfite-modified urocanase, which is formed in cells, can be reactivated not only by sunlight but also at physiological temperatures.

Enzyme Activation↗

Excimer laser surgery of the cornea.

The excimer laser, which produces light in the far-ultraviolet portion of the spectrum, allows precise removal of corneal tissue through a photochemical laser-tissue interaction. This interaction is not thermal and does not involve optical breakdown; rather, it directly breaks organic molecular bonds without tissue heating. We used this process of ablative photodecomposition to remove corneal tissue in a series freshly enucleated cow eyes. Applying the far-ultraviolet light in short intense pulses permitted us to control the depth of the incision with great precision. We found that 1 joule/cm2 ablates corneal tissue to a depth of 1 micron. Adjacent tissue suffered no thermal damage and the stromal lamellae adjacent to the incision showed no evidence of disorganization.

Animals↗

Photochemical electron transport in photosynthetic reaction centers from Rhodopseudomonas spheroides. I. Kinetics of the oxidation and reduction of P-870 as affected by external factors.

Photosynthetic reaction centers from Rhodopseudomonas spheroides were prepared with the detergent lauryl dimethylamine oxide (LDAO). In contrast to reaction centers made with Triton X-100, these contained no cytochromes and little or no ubiquinone (UQ). The reduction of P-870, after its photochemical oxidation, was studied in these materials with the following results. In reaction centers made with Triton X-100, slow kinetic components (seconds to minutes) could be attributed to secondary electron acceptors or traps. In reaction centers made with LDAO the kinetics were predominantly fast (half-times, 100 msec or less); slower components could be introduced by adding UQ. Added UQ appeared to become bound to reaction centers made with LDAO, but the binding might have meant only that both components were trapped within detergent micelles. Ferricyanide could retard the reduction of oxidized P-870, apparently by capturing electrons from the reducing side of the photochemical system. Under conditions in which the participation of secondary electron acceptors seemed to have been eliminated, the recovery of P-870 was mainly by a first-order process with a half-time of about 60 msec at room temperature and 20-30 msec at about -80 degrees C and below. The transition with decreasing temperature suggested the presence of a mixed population, exhibiting both the 60 and 20 msec components, but variations in the absorption spectra with temperature did not suggest the presence of a mixed population. Absorption difference spectra in the ultraviolet were compatible with the idea that UQ added to reaction centers became reduced in the light.

Detergents↗

Lens refilling and endocapsular polymerization of an injectable intraocular lens: in vitro and in vivo study of potential risks and benefits.

Endocapsular cataract removal and injection of a liquid artificial lens has several advantages, including restitution of accommodation, small corneoscleral incision, a more physiological position of the intraocular lens, and a reduced rate of secondary opacification. Our technique consists of bimanual phacofragmentation followed by injection of a fluid monomeric material that can be polymerized inside the capsular bag by short light exposure. Our study assessed the potential risks of the technique (e.g., heat damage to ocular tissue, light damage to the retina) and investigated the technique in vivo. We performed in vitro experiments on porcine cadaver eyes and an in vivo study on 15 rabbits. After a clinical follow-up of at least 12 weeks, the eyes were investigated histopathologically. During the polymerization process, the highest temperature measured at the posterior lens capsule was 45.1 degrees Celsius for a few seconds. The measured irradiance (0.065 watts per cm2) and the risk of photochemical damage to the retina during 20 seconds of polymerization were comparable to that caused by 1.5 minutes of standard coaxial illumination with the operating microscope. In vivo there were no serious inflammatory reactions except in four cases in which there had been intraoperative problems. The rate of secondary opacification appeared less than in conventional intraocular lens implantation in rabbits, especially when the capsule refilled completely. Retinal damage could not be detected histopathologically. In conclusion, refilling techniques may be successful once appropriate refilling materials become available.

Animals↗

Photodynamic therapy enhanced by hyperbaric oxygen in acute endoluminal palliation of malignant bronchial stenosis (clinical pilot study in 40 patients).

OBJECTIVES: Photodynamic tumor therapy (PDT) is based upon a photochemical reaction that is limited by the availability of molecular oxygen in the target tissue. The use of hyperbaric oxygenation (HBO) increases the amount of oxygen available for the process may thereby enhance the efficacy of PDT. We proved in a prospective, non-randomized clinical pilot study the acute effects on malignant bronchial stenosis and the technical feasibility of combined PDT/HBO. METHODS: Forty patients (29 males, 11 females, mean age: 64.3 years; range 39-82 years) with inoperable, advanced malignant bronchial tumor stenosis were studied prospectively. Photosensitization was carried out using a hematoporphyrin-derivative 2 mg/kg bw 48 h prior to PDT. The light dose was calculated as 300 J/cm fiber tip. The assessment of outcome 1 and 4 weeks after PDT/HBO was done by endoscopy, chest X-ray, spirometry, laboratory parameters, subjective report of dyspnea and Karnofsky performance status. RESULTS: At 1 and 4 weeks after the treatment the patients felt a significant improvement of dyspnea and hemoptysis alongside with an objective subsiding of poststenotic pneumonia, though spirometric parameters revealed no significant difference. A significant reduction of tumor stenosis (P<0.05) and an improvement of the Karnofsky performance status (P<0.05) were documented 1 and 4 weeks after PDT/HBO. No therapy related complications were observed. CONCLUSION: Although the small number of patients does not allow to draw definitive conclusions, the results suggest that combined PDT/HBO represents a new, safe and technically feasible approach. It enables efficient and rapid reduction of the endoluminal tumor load and helps conditioning the patient for further treatment procedures.

Adenocarcinoma↗

Piperazine additions to C60--a facile approach to fullerene substitution.

A range of fullerene monoadducts can be generated via the photochemical reaction of piperazine derivatives with C60. Addend functionality can also be efficiently incorporated by transformation of the hydroxyl-substituted adduct prepared in this fashion. Reaction yields and process simplicity compete with current standard procedures for fullerene mono-functionalisation.

Journal Article↗

Exploratory monitoring of air pollutants for mutagenicity activity with the Tradescantia stamen hair system.

The Tradescantia genetic system developed by the late Dr. Arnold H. Sparrow for the study of effects of ionizing radiation is applicable to chemical mutagen detection. Early radiobiological data demonstrated that the stamen hairs were sensitive to as little as 0.25 rad of x-rays and that the number of cells showing a phenotypic change in pigmentation from blue to pink plateaus after approximately 21 days of chronic, low-level irradiation. Exposures to the air pollutants SO(2), NO(2), and O(3) and to vapors of mutagens such as 1,2-dibromoethane (DBE) and ethyl methanesulfonate (EMS) demonstrated the usefulness of the system as a detector of chemical mutagens. A significant number of phenotypic changes was observed following exposures to as little as 0.14 ppm of DBE. The maximum sensitivity of the system is obtained with long-term or chronic exposures because the response increases linearly in proportion to the duration of exposure up to 21 days. To monitor industrial sites for atmospheric mutagens a mobile laboratory was designed to support plant culture in the field. Environment-controlled growth chambers were installed in a trailer so that both ambient air fumigations and concurrent clean-air control exposures could be made. Sites monitored by the mobile laboratory were: Elizabeth, N. J.; Charleston, W. Va.; Birmingham, Ala.; Baton Rouge, La.; Houston, Tex.; Upland, Calif.; Magna, Utah; and Grand Canyon, Ariz. The latter site at Grand Canyon served as a clean air control study. Atmospheric contaminants from petroleum and chemical processing plants generated a significant number of phenotypic pigment changes that were 17 to 31% above the control levels; contaminants from steel and copper smelters, automotive combustion products and photochemical compounds were negative. Chemical analyses are underway to identify the atmospheric mutagens at the sites that showed a positive response.

Air Pollutants↗

Phototriggered formation and repair of DNA containing a site-specific single strand break of the type produced by ionizing radiation or AP lyase activity.

DNA strand breaks are produced by a variety of agents and processes such as ionizing radiation, xenobiotics, oxidative metabolism, and enzymatic processing of DNA base damage. One of the major types of strand breaks produced by these processes is a single nucleotide gap terminating in 5'- and 3'-phosphates. Previously, we had developed a method for sequence-specifically producing such phosphate-terminated strand breaks in an oligodeoxynucleotide by way of two photochemically activated (caged) building blocks placed in tandem. We now report the design and synthesis of a single caged building block consisting of 1,3-(2-nitrophenyl)-1,3-propanediol, for producing phosphate-terminated strand breaks, and its use producing such a break at a specific site in a double-stranded circular DNA vector. To produce the site-specific break in a duplex vector, a primer containing the caged single strand break was extended opposite the single strand form of a circular DNA vector followed by enzymatic ligation and purification. The single strand break could then be formed in quantitative yield by irradiation of the vector with 365 nm light. In contrast to a previous study, it was found that the strand break can be repaired by Escherichia coli DNA polymerase I and E. coli DNA ligase alone, though less efficiently than in the presence of the 3'-phosphate processing enzyme E. coli endonuclease IV. Repair in the absence of endonuclease IV could be attributed to hydrolysis of the 3'-phosphate in the presence of dNTP and to a lesser extent to exonucleolytic removal of the 3'-phosphate-bearing terminal nucleotide by way of the 3' --> 5' exonuclease activity of polymerase I. This work demonstrates that specialized 3'-end processing enzymes such as endonuclease IV or exonuclease III are not absolutely required for repair of phosphate-terminated gaps. In addition to preparing single strand breaks, the caged building block described should also be useful for preparing double strand breaks and multiply damaged sites that might otherwise be difficult to prepare by other methods due to their lability.

Bacteriophage T4↗

Human pigmentation genes and their response to solar UV radiation.

Identification and characterisation of the genes involved in melanin pigment formation, together with the study of how their action is influenced by exposure to UV radiation, is providing a molecular understanding of the process of skin photoprotection through tanning. The mechanisms underlying this change in epidermal melanin involve both a transcriptional response of the pigmentation genes and post-translational control of the melanin biosynthetic pathway. UV rays are known to interact with numerous molecules within cells, and among these the photochemical reactions involving lipids and DNA are implicated in modulating melanogenesis. The combination of DNA damage, the formation of diacylglycerol, and the action of the melanocyte stimulating hormone receptor are all likely to be involved in UV-induced tanning.

Animals↗

Direct generation of superoxide anions by flash photolysis of human oxyhemoglobin.

The results presented in this report suggest that human oxyhemoglobin can directly form methemoglobin and superoxide anion when flashed with low intensity (38 joules) white light. The effect only occurred in quartz but not glass (cut off lambda approximately equal to 300 nm) cuvettes. The formation of O2 was established by observing the reduction of oxidized cytochrome c concomitant with MetHb formation at pH 9, and by showing that superoxide dismultase and catalse inhibit cytochrome c reduction at that pH. The inhibition of cytochrome c reduction by catalase led us to explore the possibility that H2O2 might reduce oxidized cytochrome c at pH 9. We show that H2O2 does reduce oxidized cytochrome c at that pH but not at pH 7. Furthermore, catalase but not superoxide dismutase, almost completely inhibited this reduction process. These experiments serve to confirm our interpretation of the effect of catalase on the reduction of oxidized cytochrome c in the photolytic experiments, thus establishing that H2O2 was also formed. In addition, we were able to identify the production of O2 and H2O2 due to the photolysis of water in agreement with the results of McCord and Fridovich ((1973) Photochem. Photobiol. 17, 115-121). Production of O2 from this source was considerably less than that observed when HbO2 was present. Addition of MetHb to aerated solutions of oxidized cytochrome c did not cause additional reduction, unlike addition of HbO2. The production of MetHb was found to have at least two components. One component was the primary photolytic process, and the second was a strongly pH-dependent reattack of HbO2 by O2. Addition of superoxide dismutase inhibited this second component, but did not significantly effect the primary photolytic process.

Cytochrome c Group↗

Photochemical-biological treatment of a real industrial biorecalcitrant wastewater containing 5-amino-6-methyl-2-benzimidazolone.

5-amino-6-methyl-2-benzimidazolone (AMBI), used in the manufacture of dyes, was characterised as a biorecalcitrant compound by means of different biodegradability tests. In order to enhance the biodegradability of this important pollutant, the application of Advanced Oxidation Process (AOPs) as a pretreatment was explored. Some experiments were addressed to find the most efficient AOP. The systems H2O2/hv, TiO2/H2O2/hv, Fe3+/hv, Fe3+/H2O2 and Fe3+/H2O2/hv were compared. The photo-Fenton system was the most efficient and the optimal conditions (AMBI, Fe3+, H2O2 concentrations) for the degradation of AMBI were found. During the photo-Fenton degradation, experiments were also made to obtain information concerning the evolution of: (a) organic carbon and initial compound concentration; (b) the oxidation state; (c) the toxicity; (d) the biodegradability; and (e) the chemical nature of the intermediates. These analyses show that the solution resulting from the treatment of AMBI is biologically compatible and complete mineralisation can be performed by biological means. A combined photochemical (Fenton) and biological flow reactor for the degradation of AMBI was successfully operated in continuous mode at laboratory scale. 100% of the initial concentration of AMBI and 80.3% of Dissolved Organic Carbon (DOC) were removed in 3.5 hours of total residence time. Finally, some field experiments under direct sunlight carried out at the Plataforma Solar de Almeria, Spain, demonstrated that this solar catalytic system is an effective treatment for this kind of industrial wastewater.

Benzimidazoles↗

Mechanism for formation of the lightstruck flavor in beer revealed by time-resolved electron paramagnetic resonance.

Time-resolved electron paramagnetic resonance (TREPR) data collected during the photodegradation of iso-a-acids (isohumulones), the principal bittering agents from hops in beer, are presented and discussed, and, from the data, the photophysics leading to free-radical production as the primary step in the photodecomposition of iso-alpha-acids towards the development of "skunky" beer are explained. During laser flash photolysis of iso-alpha-acids at 308 nm in toluene/methylcyclohexane (1:1), TREPR spectra exhibit net emissive signals that are strongly spin polarized by the triplet mechanism of chemically induced electron spin polarization. From two potential photochemically active sites, the TREPR data show that although the first site, an enolized beta-triketone, is the primary light-absorbing chromophore, an uphill intramolecular triplet energy transfer process leads to Norrish type I alpha-cleavage at a second site, an alpha-hydroxycarbonyl. The energy transfer mechanism is supported by additional TREPR experiments with chemically modified hop compounds. Structural parameters (hyperfine coupling constants, g factors, line widths) for the observed free radicals, obtained from computer simulations, are presented and discussed.

Beer↗

Chemical and Photochemical Dual Polymerization in a Mixed Langmuir Monolayer of Diacetylene Derivatives and Octadecyltrimethoxysilane.

A chemical-and-photochemical-induced dual polymerization at the air-water interface was performed for the first time, by using a ternary mixture of a octadecyltrimethoxysilane (C18TMS), 10,12-pentacosadiynoic acid (PDA), and N-(11-O-alpha-d-mannopyranosyl-3,6,9-trioxa) undecyl 10,12-pentacosa-diynamide (PDTM). Different dual polymerization behaviors of the mixed monolayers of PDA/PDTM/C18TMS (7:1:8 molar ratio) at different subphase pH were monitored by surface pressure-area isotherms and UV-vis spectra. At pH 1, the C18TMS formed a long linear polymer network by strong intermolecular condensation. The PDA/PDTM was isolated to small domains by this network and can only be polymerized weakly upon UV irradiation. At pH 2.6, the weak condensation of C18TMS formed short linear polymers, which caused phase separation between C18TMS and PDA/PDTM. The strong photochemical polymerization of the PDA/PDTM rich domain was similar to the PDA/PDTM binary mixed monolayer. At pH 4.0, the partially hydrolyzed C18TMS was miscible with PDA/PDTM and slowed the polymerization process of PDA/PDTM upon UV irradiation. These results show geometry and kinetic controls of the PDA/PDTM polymer in 2D by using the C18TMS polymer network. Copyright 1998 Academic Press.

Journal Article↗

An immunofluorescence study of T and B lymphocytes in ozone-induced pulmonary lesions in the mouse.

Ozone is a photochemical oxidant which reacts with a variety of biological molecules. In experimental animals the toxicity of ozone results from damage to cells in the lung and is associated with the process of reactive repair and an influx of inflammatory cells. In this work we used an indirect immunofluorescence technique to study the effect of ozone on T and B lymphocytes with special reference to their presence in ozone-induced pulmonary lesions. BALB/c mice were exposed to ozone at a concentration of 0.7 ppm for 20 hr per day, 7 days per week, for 4 or 14 days. Control mice were housed in comparable chambers lacking ozone. Frozen sections of lung were prepared and stained for the surface markers Thy-1.2 (T lymphocyte) and sIgM (B lymphocyte). These experiments showed that T lymphocytes infiltrate the lung lesions during ozone inhalation, and increase their presence as ozone exposure continues. They were present in the centroacinar region and tended to occur in clusters within the ozone-induced lesions. In contrast, the infiltration of B lymphocytes was virtually nonexistent with few or no sIgM positive cells present in the lesions after either 4 or 14 days of exposure to ozone. These results show that T lymphocytes occur within the sites of ozone-induced damage and support that this cell type plays a role in the reactive host response to ozone inhalation.

Animals↗

Functional characterization of sensory rhodopsin II from Halobacterium salinarum expressed in Escherichia coli.

Sensory rhodopsin II (SRII) from Halobacterium salinarum is heterologously expressed in Escherichia coli with a yield of 3-4 mg of purified SRII per liter cell culture. UV/Vis absorption spectroscopy display bands characteristic for native SRII. The resonance Raman spectrum provides evidence for a strongly hydrogen-bonded Schiff base like in mammalian rhodopsin but unlike to the homologous pSRII from Natronobacterium pharaonis. Laser flash spectroscopy indicates that SRII in detergent as well as after reconstitution into polar lipids shows its typical photochemical properties with prolonged photocycle kinetics. The first functional heterologous expression of SRII from H. salinarum provides the basis for studies with its cognate transducer HtrII to investigate the molecular processes involved in phototransduction as well as in chemotransduction.

Electrophoresis↗