Cáncer de testículo asociado a microlitiasis testicular / Association between testicular cancer and microlithiasis

Introducción: En la actualidad no existe evidencia que determine de forma concreta la relación entre microlitiasis testicular sola o en relación con otros factores como elemento de riesgo para el desarrollo de tumores testiculares, además de no existir recomendaciones claras sobre el seguimiento de esta condición. El objetivo de esta revisión es aportar con una guía para el seguimiento de estos pacientes basado en la evidencia de la literatura. Métodos: Se realizó una revisión de literatura durante diciembre de 2021 en PubMed, base de datos Cochrane y TRIP Database, la selección de los artículos se realizó por medio de las recomendaciones de PRISMA 2020. Resultados: Un total de 4 revisiones sistemáticas fueron seleccionadas para el trabajo final. Se logró determinar que la asociación de microlitiasis testicular a otros factores de riesgo incrementa aún más el riesgo de desarrollo de cáncer, sin embargo, en ausencia de estos factores el riesgo el riesgo de cáncer testicular es similar al de la población general. Conclusiones: En pacientes con riesgo de desarrollo de cáncer testicular se recomienda un seguimiento individualizado dependiendo de la edad, de los factores de riesgo asociados, de la infertilidad y del síndrome de disgenesia testicular, para poder determinar la necesidad de seguimiento versus realización de biopsia testicular. (AU)

Introduction: Currently, no evidence determines the relationship between testicular microlithiasis by itself, or in relation with other factors, as a risk factor for the development of testicular tumors. There are no clear recommendations regarding the follow-up of this medical condition. Therefore, this review aims to provide a guide to monitoring these patients, supported by the literature. Methodology: A literature review was carried out in December 2021 in PubMed, Cochrane, and TRIP Database, and the selection of the articles was made following the PRISMA 2020 recommendations. Results: Overall, the four systematic reviews chosen to conduct the final study determined that the combination of microlithiasis testicular with other risk factors further increased cancer development. However, the likelihood of testicular cancer risk is similar to that of the general population. Conclusions: Patients at risk of developing testicular cancer should undergo personalized monitoring according to their age, associated risk factors, infertility, and testicular dysgenesis syndrome to determine their follow-up needs or perform a testicular biopsy. (AU)

Association between testicular cancer and microlithiasis.

INTRODUCTION: Currently, no evidence determines the relationship between testicular microlithiasis by itself, or in relation with other factors, as a risk factor for the development of testicular tumors. There are no clear recommendations regarding the follow-up of this medical condition. Therefore, this review aims to provide a guide to monitoring these patients, supported by the literature. METHODOLOGY: A literature review was carried out in December 2021 in PubMed, Cochrane, and Trip databases, and the selection of the articles was made following the PRISMA 2020 recommendations. RESULTS: Overall, the four systematic reviews chosen to conduct the final study determined that the combination of microlithiasis testicular with other risk factors further increased cancer development. However, the likelihood of testicular cancer risk is similar to that of the general population. CONCLUSIONS: Patients at risk of developing testicular cancer should undergo personalized monitoring according to their age, associated risk factors, infertility, and testicular dysgenesis syndrome to determine their follow-up needs or perform a testicular biopsy.

[Interventions to promote social communication in children with autism spectrum disorders]. / Intervenciones para promover la comunicacion social en niños con trastornos del espectro autista.

INTRODUCTION: The difficulties encountered when it comes to social communication are one of the core disorders experienced by persons with autism spectrum disorders (ASD). This problem leads to feelings of loneliness and social exclusion, which negatively affect the quality of life. AIMS: To review the characteristics of the main interventions in this field in high-functioning ASD. DEVELOPMENT: Strategies employed include social stories, comic-strip conversations or videomodelling, and interventions mediated by peers and multicomponent treatments. CONCLUSIONS: The design of the programmes used today has evolved towards more ecological approaches that take the family, teachers and companions into account in the treatment. The most recent literature reviews have found evidence of significant improvements following their implementation, which has been reflected in better social competence and lesser feelings of loneliness. The social communication intervention must combine individualised instruction with consideration of the child's environment and the motivation towards communicative interaction.

TITLE: Intervenciones para promover la comunicacion social en niños con trastornos del espectro autista.Introduccion. Las dificultades en la comunicacion social constituyen una de las alteraciones nucleares que experimentan las personas con trastorno del espectro autista (TEA). Esta problematica provoca sentimientos de soledad y exclusion social, que repercuten negativamente en la calidad de vida. Objetivo. Revisar las caracteristicas de las principales intervenciones en este ambito en el TEA de alto funcionamiento. Desarrollo. Se sintetizan estrategias como las historias sociales, comic-strip conversations o videomodelaje, e intervenciones mediadas por pares y tratamientos multicomponente. Conclusiones. El diseño de los actuales programas ha evolucionado hacia enfoques mas ecologicos, considerandose a la familia, a los profesores y a los compañeros en el tratamiento. Las ultimas sintesis de la bibliografia han evidenciado mejoras significativas tras su implementacion, lo que se ha manifestado en una mejor competencia social y un menor sentimiento de soledad. La intervencion en comunicacion social debe combinar la instruccion individualizada con la consideracion del entorno del niño y de la motivacion hacia la interaccion comunicativa.

Characterization of nonstarch polysaccharides content from different edible organs of some vegetables, determined by GC and HPLC: comparative study.

Content and composition of dietary fiber as nonstarch polysaccharides (NSP) was determined in vegetables belonging to different types of edible organs, using GC and HPLC. Samples analyzed were subterranean organs (radish and leek), leaves (celery, swiss chard, and lettuce), stalks (celery, swiss chard, and asparagus), inflorescence (broccoli), and fruits (tomato, green pepper, and marrow). The results indicate that though the monomeric profile is similar in all these samples quantitative differences were found for neutral sugars and uronic acids among samples of the same type of vegetal organ. The NSP values determined using CG method were in good agreement with HPLC method (R(2) = 0.9005). However, arabinose, mannose, and galactose plus rhamnose are more influenced by the analytical method used than the rest of the monomers in nearly all the samples analyzed. Final values of NSP depend on the method used in celery stalks, broccoli, and green pepper.

Reductively activated cleavage of DNA mediated by o, o'-diphenylenehalonium compounds.

o,o'-Diphenylenehalonium (DPH) cations represent a novel class of DNA-affinic compounds characterized by binding constants within the range of 10(5)-10(6) M(-1). The maximum binding capacity of 2-2.5 base pairs per DPH cation and about 30% hypochromic reduction in the optical absorption of DPH cations upon binding to DNA suggest intercalation as a likely binding mode. In a DNA-bound form, DPH cations induce strand breaks upon reduction by radiation-produced electrons in aqueous solutions. In keeping with this mechanism, the cleavage is strongly inhibited by oxygen and is not affected by OH radical scavengers in the bulk. The yields of DPH-mediated base release significantly exceed the yield of base release caused by hydroxyl radical (in the absence of scavenger) in anoxic solutions. The yields are weakly dependent on DNA loading within the range from 5 to 50 base pairs per intercalator, which indicates the ability of excess electrons in DNA to react with a scavenger separated by tens of base pairs from the electron attachment site. The question regarding the mechanism by which the distant reactants reach each other in DNA remains unanswered, although it most likely involves electron hopping rather than a single-step long-distance tunneling. The latter conclusion is based on our finding that the electron affinity of DPH cations does not affect their properties as electron scavengers in DNA as would be expected if the direct long-distance tunneling is involved.

Postharvest storage of white asparagus (Asparagus officinalis l.): changes in dietary fiber (Nonstarch polysaccharides).

Changes occurring in the content and composition of the dietary fiber of white asparagus during storage in different conditions were studied (2 degrees C; 2 degrees C in polyethylene bags with air; 2 degrees C in polyethylene bags with a selected gas mixture). The neutral sugars and uronic acid composition of dietary fiber was determined by gas chromatography and by a spectrophotometric method. The modifications observed in the dietary fiber of the asparagus stored at 2 degrees C were more rapid and pronounced than those in polyethylene bags. The most important changes corresponded to xylose and glucose from insoluble dietary fiber and galactose from soluble dietary fiber. Statistical analysis indicated that the modifications were significantly affected by the type of storage and time.

Modification of the reductive pathway in gamma-irradiated DNA by electron scavengers: targeting the sugar-phosphate backbone.

Several electron scavengers that irreversibly form potential hydrogen-abstracting species upon one-electron reduction have been tested as agents for conversion of reductive damage to DNA bases into damage to the sugar-phosphate backbone. Electron spin resonance spectroscopy is employed to follow the production of radicals and transformations after irradiation. The scavengers tested included neutral (acrylamide, iodoacetamide) and cationic [triphenylsulfonium (Ph3S+), o,o'-diphenylenebromonium (DPB) and o,o'-diphenyleneiodonium (DPI)] compounds. Modification of reductive radiation damage in DNA is found to occur by scavenging of initial mobile electrons at low temperatures as well as thermally activated electron transfer from DNA electron-gain centers upon annealing. Electron transfer from the bases to hydrogen-bonded acrylamide has the smallest activation energy among other scavengers but produces a secondary alkyl radical incapable of abstracting hydrogen from the sugar-phosphate backbone. A primary alkyl radical generated from iodoacetamide has been shown to abstract preferentially from the thymine methyl group but not from deoxyribose moieties. Aryl radicals generated from aromatic onium salts such as Ph3S+, and especially DPI and DPB, are found to be the agents which best abstract hydrogen atoms from the deoxyribose portion of DNA. The use of DPB and DPI as radiation modifiers allows the elimination of undesirable side reactions of aryl radicals and through hydrogen abstraction results in high yields of a species identified as the DNA C1'. sugar radical. The second reaction pathway found for DPI and DPB in DNA is addition of an aryl radical to the thymine 5,6 double bond. Cysteamine is shown to preferentially eliminate sugar radicals upon annealing and to have little impact on the thermal stability of the thymine adduct radical.

TEMPOL protects against lens DNA strand breaks and cataract in the x-rayed rabbit.

PURPOSE: To investigate the ability of the nitroxide free radical and superoxide dismutase mimic 4-hydroxy-2,2,6,6-tetramethylpiperidine-n-oxyl (TEMPOL) to protect against x-ray-induced lens DNA damage and cataract formation in the rabbit. METHODS: Eleven gray (Gy) x-rays was delivered twice, with a 48-hour interval, to the same eye of 5-week-old rabbits. Fifteen minutes before each x-ray, 150 microliters aqueous humor was removed from the anterior chamber and replaced with 150 microliters citrate buffer containing 0 mM or 100 mM TEMPOL. The development of cataract was classified into seven stages by slit-lamp examination. DNA strand breaks were measured in the lens epithelium of x-rayed rabbits using a single-cell gel electrophoresis method. RESULTS: The level of total TEMPOL in the aqueous humor of rabbits at 15 minutes after intracameral injection of the compound was 21 mM with 84% present in the oxidized form (determined by electron paramagnetic resonance spectroscopy). At 19 weeks after x-ray, rabbits irradiated without TEMPOL showed either stage 5 (complete posterior subcapsular opacity) or stage 6 (mature) cataracts, whereas the animals that had first been injected with TEMPOL developed only stage 2 to stage 4 cataracts (the difference between the two groups was significant at P < 0.01). Intracameral injection of TEMPOL resulted in a decrease of nearly 70% in the level of DNA strand breaks produced by a single 11-Gy dose of x-ray. In vitro studies showed that TEMPOL was reduced rapidly by ascorbic acid but not by reduced glutathione. Oxidized but not reduced TEMPOL (TEMPOL-H) was an effective radioprotector in cultured rabbit lenses, and TEMPOL was nearly completely bioreduced in the plasma and aqueous humor of animals that were fed the compound in drinking water. CONCLUSIONS: TEMPOL was effective in protecting against lens epithelial DNA damage and cataract formation in x-rayed rabbits. Although a number of mechanisms are possible, the protective effect may be associated with the ability of TEMPOL to protect against radiation-produced peroxides by acting as a superoxide dismutase mimic and to oxidize Fe2+ bound to DNA, thus preventing formation of the hydroxyl radical and subsequent DNA damage through the Haber-Weiss mechanism.

Secondary radical attack on DNA nucleotides: reaction by addition to DNA bases and abstraction from sugars.

In this work radicals generated by dissociative electron attachment to iodoacetamide (H2NCOCH2.) and 6-chloromethyluracil (U5CH2.) are suggested to react with DNA nucleotides in frozen aqueous solutions via either hydrogen abstraction or addition to the double bonds of the bases. Methyl hydrogens of TMP are the preferential sites of the attack by H2NCOCH2. radical. For dCMP the C1' site on the sugar group is found to be the preferred site of hydrogen abstraction by H2NCOCH2. while for dGMP and to a lesser extent dAMP attack at the C8 position of the purine ring is found to be competitive with sugar attack. In general allylic U6CH2. and U5CH2. radicals are found to be poor hydrogen abstractors and the only reaction pathway found is the addition to double bonds at C6 in thymine and C8 in adenine and guanine. Whereas, the cytosine 5,6 double bond appears to be unreactive towards addition at low temperatures. Some evidence is found for sugar radical addition to the adenine C8 position.

Electron spin resonance of DNA irradiated with a heavy-ion beam ([16]O[8+]): evidence for damage to the deoxyribose phosphate backbone.

The free radicals produced from the irradiation of hydrated DNA with a heavy-ion beam have been investigated by ESR spectroscopy. The dominant free radical species formed after 60 MeV/nucleon (16)O(8+) ion-beam irradiations at low temperatures (77 K) are the same as those previously identified from studies using low-LET radiation, pyrimidine electron-gain radicals and purine electron-loss radicals; however, greater relative amounts of neutral carbon-centered radicals are found with the higher-LET radiation, and a new phosphorus-centered radical is identified. The fraction of neutral carbon radicals is also found to increase along the ion-beam track with the highest amounts found in the Bragg peak. The neutral carbon-centered radicals likely arise in part from the sugar moiety. The G values found for total trapped radicals at 77 K are significantly smaller for the (16)O(8+) ion beam than those found for low-LET radiation. The new phosphorus-centered radical species is identified by its large 31P parallel hyperfine coupling of about 780 G as a phosphoryl radical. This species is produced linearly with dose and is not found in significant amounts in DNA irradiated with low-LET radiation. The phosphoryl radical must be produced by the fragmentation of a P-O bond and suggests the possibility of a direct strand break. The yield of phosphoryl species is small (about 0.1% of all radicals); however, it clearly indicates that new mechanisms of damage which are not significant for low-LET radiation must be considered for high-LET radiation.

Yields of OH in gamma-irradiated DNA as a function of DNA hydration: hole transfer in competition with OH formation.

In this work, we report the yields of hydroxyl radicals, as G values and "destruction constants," in the DNA hydration shell as a function of the level of hydration. Electron spin resonance spectroscopy of gamma-irradiated DNA at low temperatures is employed for detection of the hydroxyl radical. Due to the glassy nature of the DNA hydration layer at low temperature, the hydroxyl radical gives a broad ESR resonance which is easily distinguished from the hydroxyl radical in a polycrystalline ice phase; thus .OH in both glassy and ice regions is quantified. Three regimes of radiological behavior for waters of hydration in DNA are found. For the first approximately 9 waters/nucleotide (which are glassy), no significant amounts of .OH are found, suggesting hole transfer to DNA. The second regime of hydration waters comprises up to about 12 additional glassy waters/nucleotide (gamma = 21). In this regime, substantial amounts of glassy .OH are found, suggesting that only a few holes which escape recombination in spurs charge-transfer to the DNA. In these two glassy regimes no trapped electrons are found, which is in accord with previous work that has reported that all electrons which do not undergo recombination in spurs transfer to DNA. The third regime of hydration water is comprised of bulk (or bulk-like) polycrystalline ice which forms when levels of hydration over 21 waters/nucleotide are reached. These waters form a separate phase from the DNA/glassy-water system, and neither hole nor substantial electron transfer to the DNA occurs; .OH in this ice phase is observed with G values that vary slightly with the amount of water in the ice phase, but which are close to the values found for pure ice.

Reactions of sulphonyl peroxyl radicals with DNA and its components: hydrogen abstraction from the sugar backbone versus addition to pyrimidine double bonds.

The reactions of sulphonyl peroxyl radicals with DNA and its components at low temperatures are investigated through use of electron spin resonance spectroscopy. Radiolysis of cysteine sulphinic acid solution in oxygenated aqueous 5 mol dm-3 LiCl glass has been employed to generate sulphonyl peroxyl radicals, CysSO2OO.. The mechanism of production is a two-step process involving the oxidation of the sulphinate (CysSO2-) to the sulphonyl radical (CysSO2.) by radiolytically produced Cl2- followed by oxygen addition to CysSO2.. Sulphonyl peroxyls are shown to be reactive toward DNA and nucleotides in low temperature glasses, giving rise to carbon-based peroxyl radicals. However, of the DNA bases only thymine was found to react readily. The thymine methyl group contains a relatively weak C-H bond which makes it easily abstractable. DNA nucleosides and nucleotides are likely attacked by CysSO2OO. by abstraction from the sugar C-H bonds. The C(1) position of deoxyribose ring of model nucleosides is suggested to be a preferred site for attack at low temperatures. The reaction with double-stranded DNA at higher temperatures may occur at the more accessible C(4) position. CysSO2OO. is also shown to react with a number of methyl substituted uracils, producing the highly persistent 5-oxouracil-6-yl radicals. The reaction mechanism proposed involves the addition of CysSO2OO. to pyrimidine 5,6-double bonds followed by elimination of cysteine sulphonic acid.

Elucidation of primary radiation damage in DNA through application of ab initio molecular orbital theory.

This review summarizes the results of recent ab initio molecular orbital calculations performed on DNA constituents that attempt to further our understanding of the radiation-induced damage to DNA. The results reviewed include calculations performed on the four individual DNA bases, the base pairs in gas phase and modelled aqueous phase, the deoxyribose moiety, and a portion of the sugar-phosphate backbone. The emphasis is on the electron affinities and ionization potentials of the radical species calculated under various conditions (i.e. gas phase, aqueous phase, proton transfer, base stacking), as it has been shown that the initial ion radical distribution is largely a function of these two properties. Theoretical studies of the electronic excited states of the individual bases and radioprotection of the biomolecule by various thiol compounds are also reviewed. Finally, a summary is provided to allow for further elaboration of the current model for radiation damage to DNA and to show the present advantages and limitations of ab initio theory in the investigation of such processes.

Reaction of cysteamine with individual DNA base radicals in gamma-irradiated nucleotides at low temperature.

An ESR investigation of the individual DNA base radicals produced by gamma-irradiation of frozen solutions of the nucleotides TMP, dCMP, dGMP and dAMP and their reactions with cysteamine upon annealing is reported. The results show that water radicals in bulk ice do not lead to the formation of DNA or cysteamine radicals. Radicals from the oxidation pathway which include the DNA base one electron oxidized radicals and their successors, G(C8)OH., A(C8)OH. and thymine dimers (.Tdi) and/or T(C6)OH; readily react with cysteamine to form RS. and ultimately RSSR-. Reactions of dGMP and dCMP radicals from the oxidation pathway with cysteamine occur at lower temperatures than those of dAMP and TMP, suggesting hole migration. Both T(C6)H. and C(N3)H. react with cysteamine to form RS. and diamagnetic products, but G(C8)H. and A(C8)H. do not. Subtraction of the anion radical T-. and its proton adduct T(C6)H. from the total radical yield of TMP (with or without cysteamine) suggests that somewhat less than half of the total TMP radicals found are a result of the oxidative pathway. Similar results are found in the other nucleotides. The total spectral intensity derived from the radicals from the oxidative pathway such as G(C8)OH., A(C8)OH. and .Tdi/T(C6)OH. are somewhat less than that for the protonated anion radicals. Only one non-base radical is identified, a sugar radical at the C(1)' site on the deoxyribose portion of dAMP. This species, S(A)., is also found to react with cysteamine or its disulfide radical anion. Analyses performed in the presence and absence of a thiol are found to allow for a clear separation of oxidative and reductive pathways.

ESR detection at 77 K of the hydroxyl radical in the hydration layer of gamma-irradiated DNA.

Previous ESR reports of gamma-irradiated DNA at low temperatures have suggested that hydroxyl radicals are not formed in the first hydration layer of DNA. In this report we show that hydroxyl radicals are produced in low yield. Due to the glassy nature of this hydration layer at low temperature, the hydroxyl radical gives a broad ESR resonance which is not easily detected. Low-field ESR spectra of hydroxyl radicals in an irradiated 6 M CsF aqueous glass are shown to be nearly identical to those found in DNA; however, the yields in the aqueous glass (G = 0.087 to 0.13 mumol/J) are found to be greater than those in DNA's first hydration layer (G = 0.035 +/- 0.02 mumol/J). A large kappa value for destruction of the OH in DNA's hydration layer limits the yield of OH at high doses. The yield of H2O2 (which likely results from hydroxyl radical recombinations that occur both during irradiation and upon annealing) is found to 0.0035 mumol/J in the dose range 65 kGy to 195 kGy. The amount of H2O2 formed corresponds to most of that expected from recombination of the OH trapped at 77 K at the equivalent dose. The low yield of trapped OH radicals in the first hydration layer has implications for possible hole transfer to DNA.

Influence of oxygen on the repair of direct radiation damage to DNA by thiols in model systems.

Here the reactions of thiols with DNA primary radical intermediates formed after gamma-irradiation of frozen (77 K) anoxic and oxic solutions of DNA/thiol mixtures are investigated. Through analysis of the experimental composite spectra at each annealing temperature, the relative concentrations of individual radicals present are estimated and reaction sequences inferred. In all samples the primary DNA radical anions and cations (DNA.+ and DNA.-) are suggested to be the predominant radicals at low temperatures. In anoxic samples, TH. (5,6-dihydrothym-5-yl radical), .RSSR.- and, in glutathione samples, .GSH [gamma-glu-NHC(CH2SH)CO-gly] radicals are observed as the temperature is increased. The presence of oxygen efficiently suppresses the formation of RSSR.- and .GSH; instead, in oxic samples, O2.-, DNAOO., RSOO. and RSO. are observed at higher temperatures. The photolytic conversion of RSOO. to RSO2. is used to verify the presence of RSOO. in gamma-irradiated DNA/thiol systems and confirm that the computer analysis employed yields reasonable estimates of the relative DNAOO. and RSOO. concentrations. From the relative concentrations of radicals present, it is clear that the radicals observed at higher temperatures originate from the radical reactions of the primary DNA.+ and DNA.- radicals. Based on the reaction sequences inferred and previous work with thiols alone, it is concluded that TH., DNAOO. and RSOO. (in part) originate largely with DNA.-, whereas RSSR.-, .GSH and RSOO. (in part) originate largely with DNA.+. The possible roles of DNAOO., RSOO., RSO., RSO2. and .OOGSH in the chemical oxygen enhancement effect at biologically realistic temperatures are discussed.

Protonation of nucleobase anions in gamma-irradiated DNA and model systems. Which DNA base is the ultimate sink for the electron?

Electron spin resonance (ESR) spectroscopy has been used to investigate irreversible protonations of the nucleobase anions in gamma-irradiated frozen aqueous solutions of dGMP-dCMP, polyG-polyC, poly[dGdC].poly[dGdC], dAMP-dTMP, poly[dAdT].poly-[dAdT] and DNA itself. Analysis of the ESR spectra at a dose of 22 kGy shows that fractional conversion of total radicals to carbon-protonated species on annealing is in the order: dAMP-dTMP (43%) > pdAdT = DNA (23%) > dGMP.dCMP (15%) > poly-dGdC.polydGdC (6%) > polyG.polyC (3%). Two hydrogen addition radicals make contributions to the polyG.polyC, poly-[dGdC].poly[dGdC] and dGMP.dCMP spectra in H2O on annealing. They are those formed by protonations at C6 of the cytosine anion radical, C(C6)H., and at C8 of the guanine anion radical, G(C8)H.. Computer analysis reveals that anion protonation reaction in dGMP.dCMP results in mainly C(C6)H., whereas protonation reaction in polyG.polyC and poly[dGdC].poly-[dGdC] yields mainly G(C8)H.. In dAMP.dTMP and poly[dAdT].poly[dAdT] as in DNA itself, the only DNA base found to undergo an irreversible protonation at a carbon site is thymine, resulting in T(C6)H.. The conversion of DNA anion to T(C6)H. is found to be dependent on dose. At low doses (5 kGy), about 30% conversion to T(C6)H. is found, whereas at high doses (94 kGy), only 13% conversion is found. The dose dependence is ascribed in part to ion radical recombinations whose probabilities are increased at high doses. A consideration of the rates of protonations of the purine and pyrimidine anion radicals as well as the differences in electron affinities suggest carbon protonation reactions of DNA base anions in irradiated stacked double-strand DNA at 37 degrees C would be predominantly at thymine and perhaps guanine, whereas in single-strand DNA all bases would contribute.

The influence of hydration on the absolute yields of primary free radicals in gamma-irradiated DNA at 77 K. II. Individual radical yields.

In this report we present an electron spin resonance (ESR) investigation of the yields of the individual free radicals formed in gamma-irradiated frozen DNA as a function of hydration and dose at 77 K. Analysis of the ESR spectra taken at low hydration shows that the ion radical composition remains nearly constant with dose and that few secondary radicals are formed even at high doses (above about 50 kGy). For fully hydrated samples, the radical composition changes dramatically with dose. Thymine anion radical (T-.) is found in abundance at low doses but nearly disappears at higher doses, with a corresponding increase in the N-3 deuterated cytosine anion radical (CD.). Guanine cation radical (G+.) decreases at high doses, with a concomitant increase in secondary radical species (S.). Analysis of the dose-response data for G values (the yields in microM/J), k values (the destruction constants) and k' values (a new constant that characterizes the change in G with dose) was performed for each of the DNA base free radicals present at 77 K. The G value for each of the base radicals increases with the hydration level. The k values for CD. and G+. increase slightly with hydration; however, that of T-. increases substantially. Destruction constants for neutral radicals such as TH. and CD. are found to be substantially smaller than those for ion radicals and provide an indication of the radical charge state. A negative k' value for T-. and a positive k' value for CD. are explained in terms of radiation effects that result in the formation of a deuterated cytosine base, i.e., C(N3)D+, which greatly increases cytosine's electron affinity. The ratio of anion radical to cation radical concentrations is found to be about 1.6 and is invariant with hydration. A speculation on the imbalance based on hole-hole combinations in spurs is presented.

The influence of hydration on the absolute yields of primary ionic free radicals in gamma-irradiated DNA at 77 K. I. Total radical yields.

An ESR (electron spin resonance) investigation of the effect of hydration water (D2O) on the absolute yields (G) of ion radicals in gamma-irradiated DNA at 77 K is reported. The total DNA radical yield is found to increase by over fourfold on addition of the primary hydration layer (20 water molecules/nucleotide) to DNA. At hydrations greater than ca. 20 waters/nucleotide the excess water freezes into a separate and apparently radiologically independent bulk ice phase. Ice formation steals ca. 5 waters from the hydration layer which causes a proportionate (by weight) drop in the total yield of DNA radical ions. Hydroxyl radicals (.OD) are not observed even at great signal amplification in the primary hydration layer but are found only in the ice phase, which suggests efficient transfer of holes and electrons from the hydration layer to the DNA; as a consequence, DNA and its associated hydration water form the target mass for radiation damage. These results show that water of hydration is critical to radical formation and stabilization in DNA; however, the ice surrounding DNA does not contribute to direct DNA damage and is found to have the same properties as bulk ice. At 18 waters per nucleotide (gamma = 18) about one-fourth of all ion radicals produced by the radiation are trapped at 77 K. The k value for destruction is found to vary only slightly with hydration. Based on the measured G and k values for hydrated DNA, a trapped radical ion cluster size is estimated to be about 6 nm. For hydrated DNA the cluster size suggests electron migration distances of about 17 bases (for migration along the DNA strand).

The superoxide dismutase mimic TEMPOL protects cultured rabbit lens epithelial cells from hydrogen peroxide insult.

The superoxide dismutase mimic, 4-hydroxy TEMPO (TEMPOL), was used to investigate the mechanism by which H2O2 damages cultured rabbit lens epithelial cells and to identify some of the targets of H2O2 insult. Most studies aimed at determining the mechanism by which H2O2 exerts its cytotoxic effect have used iron chelators to prevent the generation of the damaging hydroxyl radical. Since TEMPOL does not chelate transition metals, we were afforded an additional means of investigating the mechanism by which H2O2 exerts its cytotoxicity. Cells at low or high density were cultured in MEM containing 5 mM TEMPOL and exposed to a single sub-lethal dose of 0.05 or 0.5 mM H2O2, respectively. Analysis of EPR spectra indicated that TEMPOL was stable in MEM, did not destroy H2O2 and penetrated the intracellular fluid. TEMPOL prevented or curtailed the H2O2-induced inhibition of cell growth, blebbing of the cell membrane, the decrease in NAD+, the activation of poly ADP-ribose polymerase, an enzyme involved in DNA repair, and limited the induction of single strand breaks in DNA normally brought about by H2O2. TEMPOL did not prevent the H2O2-induced decrease in reduced glutathione, lactate production, and the activity of glyceraldehyde 3-phosphate dehydrogenase, or the H2O2-induced increases in oxidized glutathione and hexose monophosphate shunt activity. Addition of TEMPOL 1-15 min after exposure of cells to H2O2 offered partial protection from the inhibition of cell division. TEMPOL at 5 mM did not inhibit cell growth. These results, coupled with our other findings suggest that some of the H2O2-induced damage in cultured rabbit LECs is mediated by intracellular redox-active metals involved in the Haber-Weiss cycle. Cellular changes not protected by TEMPOL, including attack of H2O2 on the thiol groups of GSH (mediated through glutathione peroxidase) and G3PDH, are likely brought about by H2O2 itself and not by reactions of oxygen free-radicals generated from H2O2.