Postnatal hypoxic preconditioning attenuates lung damage from hyperoxia in newborn mice.

BACKGROUND: Preterm infants frequently require oxygen supplementation at birth. However, preterm lung is especially sensible to structural and functional damage caused by oxygen free radicals. METHODS: The adaptive mechanisms implied in the fetal-neonatal transition from a lower to a higher oxygen environment were evaluated in a murine model using a custom-designed oxy-chamber. Pregnant mice were randomly assigned to deliver in 14% (hypoxic preconditioning group) or 21% (normoxic group) oxygen environment. Eight hours after birth FiO2 was increased to 100% for 60 min and then switched to 21% in both groups. A control group remained in 21% oxygen throughout the study. RESULTS: Mice in the normoxic group exhibited thinning of the alveolar septa, increased cell death, increased vascular damage, and decreased synthesis of pulmonary surfactant. However, lung histology, lamellar bodies microstructure, and surfactant integrity were preserved in the hypoxic preconditioning group after the hyperoxic insult. CONCLUSION: Postnatal hyperoxia has detrimental effects on lung structure and function when preceded by normoxia compared to controls. However, postnatal hypoxic preconditioning mitigates lung damage caused by a hyperoxic insult. IMPACT: Hypoxic preconditioning, implemented shortly after birth mitigates lung damage caused by postnatal supplemental oxygenation. The study introduces an experimental mice model to investigate the effects of hypoxic preconditioning and its effects on lung development. This model enables researchers to delve into the intricate processes involved in postnatal lung maturation. Our findings suggest that hypoxic preconditioning may reduce lung parenchymal damage and increase pulmonary surfactant synthesis in reoxygenation strategies during postnatal care.

Exacerbated response to oxidative stress in the Retinitis Pigmentosa CerklKD/KO mouse model triggers retinal degeneration pathways upon acute light stress.

The retina is particularly vulnerable to genetic and environmental alterations that generate oxidative stress and cause cellular damage in photoreceptors and other retinal neurons, eventually leading to cell death. CERKL (CERamide Kinase-Like) mutations cause Retinitis Pigmentosa and Cone-Rod Dystrophy in humans, two disorders characterized by photoreceptor degeneration and progressive vision loss. CERKL is a resilience gene against oxidative stress, and its overexpression protects cells from oxidative stress-induced apoptosis. Besides, CERKL contributes to stress granule-formation and regulates mitochondrial dynamics in the retina. Using the CerklKD/KO albino mouse model, which recapitulates the human disease, we aimed to study the impact of Cerkl knockdown on stress response and activation of photoreceptor death mechanisms upon light/oxidative stress. After acute light injury, we assessed immediate or late retinal stress response, by combining both omic and non-omic approaches. Our results show that Cerkl knockdown increases ROS levels and causes a basal exacerbated stress state in the retina, through alterations in glutathione metabolism and stress granule production, overall compromising an adequate response to additional oxidative damage. As a consequence, several cell death mechanisms are triggered in CerklKD/KO retinas after acute light stress. Our studies indicate that Cerkl gene is a pivotal player in regulating light-challenged retinal homeostasis and shed light on how mutations in CERKL lead to blindness by dysregulation of the basal oxidative stress response in the retina.

The antioxidant l-Ergothioneine prevents cystine lithiasis in the Slc7a9-/- mouse model of cystinuria.

The high recurrence rate of cystine lithiasis observed in cystinuria patients highlights the need for new therapeutic options to address this chronic disease. There is growing evidence of an antioxidant defect in cystinuria, which has led to test antioxidant molecules as new therapeutic approaches. In this study, the antioxidant l-Ergothioneine was evaluated, at two different doses, as a preventive and long-term treatment for cystinuria in the Slc7a9-/- mouse model. l-Ergothioneine treatments decreased the rate of stone formation by more than 60% and delayed its onset in those mice that still developed calculi. Although there were no differences in metabolic parameters or urinary cystine concentration between control and treated mice, cystine solubility was increased by 50% in the urines of treated mice. We also demonstrate that l-Ergothioneine needs to be internalized by its transporter OCTN1 (Slc22a4) to be effective, as when administrated to the double mutant Slc7a9-/-Slc22a4-/- mouse model, no effect on the lithiasis phenotype was observed. In kidneys, we detected a decrease in GSH levels and an impairment of maximal mitochondrial respiratory capacity in cystinuric mice that l-Ergothioneine treatment was able to restore. Thus, l-Ergothioneine administration prevented cystine lithiasis in the Slc7a9-/- mouse model by increasing urinary cystine solubility and recovered renal GSH metabolism and mitochondrial function. These results support the need for clinical trials to test l-Ergothioneine as a new treatment for cystinuria.

Sustainable Biocatalytic Procedure for Obtaining New Branched Acid Esters.

Biocatalytic synthesis of 2-ethylhexyl 2-methylhexanoate is described in this work for the first time. This branched-chain ester is suitable for use at low temperatures in numerous applications. The immobilized lipase Novozym® 435 has demonstrated its ability to catalyze the ester synthesis from 2-ethylhexanol and 2-methylhexanoic acid in a solvent-free medium. The high reaction times that are required result in a loss of alcohol by evaporation, which must be compensated for with an excess of this substrate if high conversions are to be achieved. Therefore, two strategies are established: 70 °C with a 10% excess of alcohol, which requires a longer operating time and provides conversions of 97%, and 80 °C with a 20% excess of alcohol, which allows for the achievement of a 99% conversion in a shorter time. The optimal reaction conditions have been chosen based on reusability of the enzyme, process productivity, green metrics and preliminary economic study. When the synthesis is carried out under the best conditions (70 °C, 10% molar excess of alcohol and six uses of the immobilized enzyme) a productivity of 203.84 kg product × kg biocatalyst-1 is attained. The biocatalytic procedure matches many of the objectives of "green chemistry" and is suitable to be scaled up and used in industrial manufacturing.

Analysis of Lipid Peroxidation by UPLC-MS/MS and Retinoprotective Effects of the Natural Polyphenol Pterostilbene.

The loss of redox homeostasis induced by hyperglycemia is an early sign and key factor in the development of diabetic retinopathy. Due to the high level of long-chain polyunsaturated fatty acids, diabetic retina is highly susceptible to lipid peroxidation, source of pathophysiological alterations in diabetic retinopathy. Previous studies have shown that pterostilbene, a natural antioxidant polyphenol, is an effective therapy against diabetic retinopathy development, although its protective effects on lipid peroxidation are not well known. Plasma, urine and retinas from diabetic rabbits, control and diabetic rabbits treated daily with pterostilbene were analyzed. Lipid peroxidation was evaluated through the determination of derivatives from arachidonic, adrenic and docosahexaenoic acids by ultra-performance liquid chromatography coupled with tandem mass spectrometry. Diabetes increased lipid peroxidation in retina, plasma and urine samples and pterostilbene treatment restored control values, showing its ability to prevent early and main alterations in the development of diabetic retinopathy. Through our study, we are able to propose the use of a derivative of adrenic acid, 17(RS)-10-epi-SC-Δ15-11-dihomo-IsoF, for the first time, as a suitable biomarker of diabetic retinopathy in plasmas or urine.

Pterostilbene Prevents Early Diabetic Retinopathy Alterations in a Rabbit Experimental Model.

Oxidative stress generated by diabetes plays a key role in the development of diabetic retinopathy (DR), a common diabetic complication. DR remains asymptomatic until it reaches advanced stages, which complicate its treatment. Although it is known that good metabolic control is essential for preventing DR, knowledge of the disease is incomplete and an effective treatment with no side effects is lacking. Pterostilbene (Pter), a natural stilbene with good antioxidant activity, has proved to beneficially affect different pathologies, including diabetes. Therefore, our study aimed to analyse the protective and/or therapeutic capacity of Pter against oxidant damage by characterising early retinal alterations induced by hyperglycaemia, and its possible mechanism of action in a rabbit model of type 1 diabetes mellitus. Pter reduced lipid and protein oxidative damage, and recovered redox status and the main activities of antioxidant enzymes. Moreover, the redox regulation by Pter was associated with activation of the PI3K/AKT/GSK3ß/NRF2 pathway. Our results show that Pter is a powerful protective agent that may delay early DR development.

7,8-hydroxy-2'-deoxyguanosine/2'-deoxiguanosine ratio determined in hydrolysates of brain DNA by ultrachromatrography coupled to tandem mass spectrometry.

7,8-hydroxy-2'-deoxyguanosine (8-OHdG) is an abundant DNA lesion formed by oxidation of the nucleoside 2'-deoxyguanosine (2-dG) and one of the most studied and accepted oxidative stress biomarkers. 8-OHdG has a strong carcinogenic potential, and prolonged oxidative stress heightens pathological conditions and especially cancer risk. Our aim was to develop, validate and apply a reliable method to assess DNA oxidation in genomic cellular DNA of sensible target organs such as brain. A procedure to isolate and digest the DNA of brain tissue properly for further detection of 8-OHdG and 2-dG by Ultra Performance Liquid Chromatography-tandem Mass Spectrometry (UPLC-MS/MS) was optimized. The UPLC-MS/MS was validated following the American Food and Drug Administration (FDA) Guidelines using mice pups' brain samples. To demonstrate the applicability of the UPLC-MS/MS method, 8-OHdG/2-dG ratio was determined in brain tissue of 1day old newborn mice pups (P1) in a model of hypoxia pre-conditioning during fetal-to-neonatal transition. We found that hypoxia at birth (FiO2 0.14) and for 8h thereafter induced lower levels of DNA oxidation in mice pups and rendered even protective against a postnatal asphyxia/reoxygenation insult compared with fetal to neonatal transition in room air. We conclude that the UPLC-MS/MS method developed has proven suitable for the analysis of DNA oxidation biomarker 8-OHdG/2-dG ratio in tissue samples from newborn mice pups. We aim to apply this method in future studies aiming to provide a deeper insight into the mechanisms of oxidation DNA caused during neonatal asphyxia and resuscitation.

Protein-bound tyrosine oxidation, nitration and chlorination by-products assessed by ultraperformance liquid chromatography coupled to tandem mass spectrometry.

BACKGROUND: Free radicals cause alterations in cellular protein structure and function. Oxidized, nitrated, and chlorinated modifications of aromatic amino acids including phenylalanine and tyrosine are reliable biomarkers of oxidative stress and inflammation in clinical conditions. OBJECTIVE: To develop, validate and apply a rapid method for the quantification of known hallmarks of tyrosine oxidation, nitration and chlorination in plasma and tissue proteins providing a snapshot of the oxidative stress and inflammatory status of the organism and of target organs respectively. MATERIAL AND METHODS: The extraction and clean up procedure entailed protein precipitation, followed by protein re-suspension and enzymatic digestion with pronase. An Ultra Performance Liquid Chromatography-tandem Mass Spectrometry (UPLC-MS/MS) method was developed to quantify protein released ortho-tyrosine (o-Tyr), meta-tyrosine (m-Tyr), 3-nitrotyrosine (3NO2-Tyr) and 3-chlorotyrosine (3Cl-Tyr) as well as native phenylalanine (Phe) and tyrosine (p-Tyr) in plasma and tissue from a validated hypoxic newborn piglet experimental model. RESULTS: In plasma there was a significant increase in the 3NO2-Tyr/p-Tyr ratio. On the other hand m-Tyr/Phe and 3Cl-Tyr/p-Tyr ratios were significantly increased in liver of hypoxic compared with normoxic animals. Although no significant differences were found in brain tissue, a clear tendency to increased ratios was observed under hypoxic conditions. CONCLUSIONS: UPLC-MS/MS has proven suitable for the analysis of plasma and tissue samples from newborn piglets. The analysis of biomarkers of protein oxidation, nitration and chlorination will be applied in future studies aiming to provide a deeper insight into the mechanisms of oxidation-derived protein modification caused during neonatal asphyxia and resuscitation.

Role of Natural Stilbenes in the Prevention of Cancer.

Natural stilbenes are an important group of nonflavonoid phytochemicals of polyphenolic structure characterized by the presence of a 1,2-diphenylethylene nucleus. Stilbenes have an extraordinary potential for the prevention and treatment of different diseases, including cancer, due to their antioxidant, cell death activation, and anti-inflammatory properties which associate with low toxicity under in vivo conditions. This review aims to discuss various approaches related to their mechanisms of action, pharmacological activities in animal models and humans, and potential chemoprevention in clinical studies. The biological activity of natural stilbenes is still incompletely understood. Furthermore, after administration to animals or humans, these molecules are rapidly metabolized. Thus pharmacokinetics and/or activities of the natural structures and their metabolites may be very different. Novel drug formulations have been postulated in order to improve stability and bioavailability, to minimize side effects, and to facilitate interaction with their domains in target proteins. These pharmacological improvements should lead stilbenes to become effective candidates as anticancer drugs.

Topical treatment with pterostilbene, a natural phytoalexin, effectively protects hairless mice against UVB radiation-induced skin damage and carcinogenesis.

The aim of our study was to investigate in the SKH-1 hairless mouse model the effect of pterostilbene (Pter), a natural dimethoxy analog of resveratrol (Resv), against procarcinogenic ultraviolet B radiation (UVB)-induced skin damage. Pter prevented acute UVB (360 mJ/cm(2))-induced increase in skin fold, thickness, and redness, as well as photoaging-associated skin wrinkling and hyperplasia. Pter, but not Resv, effectively prevented chronic UVB (180 mJ/cm(2), three doses/week for 6 months)-induced skin carcinogenesis (90% of Pter-treated mice did not develop skin carcinomas, whereas a large number of tumors were observed in all controls). This anticarcinogenic effect was associated with (a) maintenance of skin antioxidant defenses (i.e., glutathione (GSH) levels, catalase, superoxide, and GSH peroxidase activities) close to control values (untreated mice) and (b) an inhibition of UVB-induced oxidative damage (using as biomarkers 8-hydroxy-2'-deoxyguanosine, protein carbonyls, and isoprostanes). The molecular mechanism underlying the photoprotective effect elicited by Pter was further evaluated using HaCaT immortalized human keratinocytes and was shown to involve potential modulation of the Nrf2-dependent antioxidant response.

Assessment of oxidative damage to proteins and DNA in urine of newborn infants by a validated UPLC-MS/MS approach.

The assessment of oxidative stress is highly relevant in clinical Perinatology as it is associated to adverse outcomes in newborn infants. This study summarizes results from the validation of an Ultra Performance Liquid Chromatography-tandem Mass Spectrometry (UPLC-MS/MS) method for the simultaneous quantification of the urinary concentrations of a set of endogenous biomarkers, capable to provide a valid snapshot of the oxidative stress status applicable in human clinical trials, especially in the field of Perinatology. The set of analytes included are phenylalanine (Phe), para-tyrosine (p-Tyr), ortho-tyrosine (o-Tyr), meta-tyrosine (m-Tyr), 3-NO2-tyrosine (3NO2-Tyr), 3-Cl-tyrosine (3Cl-Tyr), 2'-deoxyguanosine (2dG) and 8-hydroxy-2'-deoxyguanosine (8OHdG). Following the FDA-based guidelines, appropriate levels of accuracy and precision, as well as adequate levels of sensitivity with limits of detection (LODs) in the low nanomolar (nmol/L) range were confirmed after method validation. The validity of the proposed UPLC-MS/MS method was assessed by analysing urine samples from a clinical trial in extremely low birth weight (ELBW) infants randomized to be resuscitated with two different initial inspiratory fractions of oxygen.

Pterostilbene: Biomedical applications.

Resveratrol and its naturally dimethylated analog, pterostilbene, show similar biological activities. However, the higher in vivo bioavailability of pterostilbene represents a fundamental advantage. The main focus of this review is on biomedical applications of pterostilbene. The metabolism and pharmacokinetics of this stilbene in inflammatory dermatoses and photoprotection, cancer prevention and therapy, insulin sensitivity, blood glycemia and lipid levels, cardiovascular diseases, aging, and memory and cognition are addressed. Safety and toxicity, as well as recommendations for future research and biomedical uses, are discussed. This review includes comparisons between pterostilbene and other polyphenols, with particular emphasis on resveratrol. Potential benefits of using combinations of different polyphenols are considered. Based on present evidences we conclude that pterostilbene is an active phytonutrient and also a potential drug with multiple biomedical applications.

Stress hormones promote growth of B16-F10 melanoma metastases: an interleukin 6- and glutathione-dependent mechanism.

BACKGROUND: Interleukin (IL)-6 (mainly of tumor origin) activates glutathione (GSH) release from hepatocytes and its interorgan transport to B16-F10 melanoma metastatic foci. We studied if this capacity to overproduce IL-6 is regulated by cancer cell-independent mechanisms. METHODS: Murine B16-F10 melanoma cells were cultured, transfected with red fluorescent protein, injected i.v. into syngenic C57BL/6J mice to generate lung and liver metastases, and isolated from metastatic foci using high-performance cell sorting. Stress hormones and IL-6 levels were measured by ELISA, and CRH expression in the brain by in situ hybridization. DNA binding activity of NF-κB, CREB, AP-1, and NF-IL-6 was measured using specific transcription factor assay kits. IL-6 expression was measured by RT-PCR, and silencing was achieved by transfection of anti-IL-6 small interfering RNA. GSH was determined by HPLC. Cell death analysis was distinguished using fluorescence microscopy, TUNEL labeling, and flow cytometry techniques. Statistical analyses were performed using Student's t test. RESULTS: Plasma levels of stress-related hormones (adrenocorticotropin hormone, corticosterone, and noradrenaline) increased, following a circadian pattern and as compared to non-tumor controls, in mice bearing B16-F10 lung or liver metastases. Corticosterone and noradrenaline, at pathophysiological levels, increased expression and secretion of IL-6 in B16-F10 cells in vitro. Corticosterone- and noradrenaline-induced transcriptional up-regulation of IL-6 gene involves changes in the DNA binding activity of nuclear factor-κB, cAMP response element-binding protein, activator protein-1, and nuclear factor for IL-6. In vivo inoculation of B16-F10 cells transfected with anti-IL-6-siRNA, treatment with a glucocorticoid receptor blocker (RU-486) or with a ß-adrenoceptor blocker (propranolol), increased hepatic GSH whereas decreased plasma IL-6 levels and metastatic growth. Corticosterone, but not NORA, also induced apoptotic cell death in metastatic cells with low GSH content. CONCLUSIONS: Our results describe an interorgan system where stress-related hormones, IL-6, and GSH coordinately regulate metastases growth.

Amniotic fluid oxidative and nitrosative stress biomarkers correlate with fetal chronic hypoxia in diabetic pregnancies.

BACKGROUND: In spite of improvement in obstetrical care, pregnancy in women with type 1 diabetes mellitus is associated with increased perinatal morbidity and mortality. Hyperglycemia during pregnancy causes excessive fetal growth and chronic fetal hypoxia as reflected in increased erythropoietin (EPO) levels in amniotic fluid (AF). OBJECTIVES: We hypothesized that the degree of fetal hypoxia would correlate with fetal oxidative and nitrosative stress as evidenced ty the concentration of specific biomarkers in AF. MATERIAL AND METHODS: 19 pregnant women with type 1 or insulin-treated gestational diabetes mellitus were studied. AF samples were collected and processed for EPO, meta-tyrosine, nitro-tyrosine and 8-hydroxy-2-deoxiguanosine by chemiluminescent immunoassay and high-performance liquid chromatography coupled to tandem mass spectrometry methods, respectively. RESULTS: The mean (SD) of the last HbA1c concentration before delivery was 7.7% (1.1). Median gestational age was 258 days (range 231-268). Birth weight was 3,868 ± 695 g with a z-score >2 SD in 47% of the cases. A significant correlation was found between the concentrations of AF EPO and meta-tyrosine/phenylalanine ratio (p < 0.001), nitro-tyrosine (p < 0.01) and 8-oxo-dG/2dG ratio (p < 0.001). CONCLUSIONS: We confirmed that fetuses of type 1 diabetes or insulin-treated gestational diabetes pregnancies experience chronic hypoxia as reflected by increased EPO concentrations in AF near term. Moreover, EPO levels significantly correlated with the concentration of oxidative and nitrosative stress biomarkers in AF. This pro-oxidant status may predispose newborn infants to poor postnatal adaptation and early neonatal complications.

Obese rats exhibit high levels of fat necrosis and isoprostanes in taurocholate-induced acute pancreatitis.

BACKGROUND: Obesity is a prognostic factor for severity in acute pancreatitis in humans. Our aim was to assess the role of oxidative stress and abdominal fat in the increased severity of acute pancreatitis in obese rats. METHODOLOGY: Taurocholate-induced acute pancreatitis was performed in lean and obese Zucker rats. Levels of reduced glutathione, oxidized glutathione, L-cysteine, cystine, and S-adenosylmethionine were measured in pancreas as well as the activities of serine/threonine protein phosphatases PP1 and PP2A and tyrosin phosphatases. Isoprostane, malondialdehyde, triglyceride, and free fatty acid levels and lipase activity were measured in plasma and ascites. Lipase activity was measured in white adipose tissue with and without necrosis and confirmed by western blotting. FINDINGS: Under basal conditions obese rats exhibited lower reduced glutathione levels in pancreas and higher triglyceride and free fatty acid levels in plasma than lean rats. S-adenosyl methionine levels were markedly increased in pancreas of obese rats. Acute pancreatitis in obese rats led to glutathione oxidation and lower reduced glutathione levels in pancreas together with decreased activities of redox-sensitive phosphatases PP1, and PP2A. S-adenosyl methionine levels decreased but cystine levels increased markedly in pancreas upon pancreatitis. Acute pancreatitis triggered an increase in isoprostane levels in plasma and ascites in obese rats. Free fatty acid levels were extremely high in pancreatitis-associated ascitic fluid from obese rats and lipase was bound with great affinity to white adipose tissue, especially to areas of necrosis. CONCLUSIONS: Our results show that oxidative stress occurs locally and systemically in obese rats with pancreatitis favouring inactivation of protein phosphatases in pancreas, which would promote up-regulation of pro-inflammatory cytokines, and the increase of isoprostanes which might cause powerful pulmonary and renal vasoconstriction. Future studies are needed to confirm the translational relevance of the present findings obtained in a rat model of taurocholate-induced pancreatic damage and necrosis.

Glutathione and Bcl-2 targeting facilitates elimination by chemoradiotherapy of human A375 melanoma xenografts overexpressing bcl-xl, bcl-2, and mcl-1.

BACKGROUND: Bcl-2 is believed to contribute to melanoma chemoresistance. However, expression of Bcl-2 proteins may be different among melanomas. Thus correlations among expression of Bcl-2-related proteins and in vivo melanoma progression, and resistance to combination therapies, was investigated. METHODS: Human A375 melanoma was injected s.c. into immunodeficient nude mice. Protein expression was studied in tumor samples obtained by laser microdisection. Transfection of siRNA or ectopic overexpression were applied to manipulate proteins which are up- or down-regulated, preferentially, during melanoma progression. Anti-bcl-2 antisense oligonucleotides and chemoradiotherapy (glutathione-depleting agents, paclitaxel protein-binding particles, daunorubicin, X rays) were administered in combination. RESULTS: In vivo A375 cells down-regulated pro-apoptotic bax expression; and up-regulated anti-apoptotic bcl-2, bcl-xl, and mcl-1, however only Bcl-2 appeared critical for long-term tumor cell survival and progression in vivo. Reduction of Bcl-2, combined with partial therapies, decreased melanoma growth. But only Bcl-2 targeting plus the full combination of chemoradiotherapy eradicated A375 melanoma, and led to long-term survival (> 120 days) without recurrence in 80% of mice. Tumor regression was not due to immune stimulation. Hematology and clinical chemistry data were within accepted clinical toxicities. CONCLUSION: Strategies to target Bcl-2, may increase the effectiveness of antitumor therapies against melanomas overexpressing Bcl-2 and likely other Bcl-2-related antiapoptotic proteins.

Natural polyphenols in cancer therapy.

Natural polyphenols are secondary metabolites of plants involved in defense against different types of stress. Extracts containing these compounds have been used for thousands of years in traditional eastern medicine. Polyphenols act on multiple targets in pathways and mechanisms related to carcinogenesis, tumor cell proliferation and death, inflammation, metastatic spread, angiogenesis, or drug and radiation resistance. Nevertheless, reported effects claimed for polyphenols are controversial, since correlations between in vitro effects and in vivo evidence are poorly established. The main discrepancy between health claims versus clinical observations is the frequent use of nonphysiologically relevant concentrations of these compounds and their metabolites in efficacy and mechanistic studies. The present review will discuss how in vivo administration correlates with polyphenol metabolism, toxicity, and bioavailability. Analysis of the general application of polyphenols in cancer therapy will be complemented by potential applications in the therapy of specific tumors, including melanoma, colorectal and lung cancers. Possible pharmaceutical formulations, structural modifications, combinations, and delivery systems aimed to increase bioavailability and/or biological effects will be discussed. Final remarks will include recommendations for future research and developments.

Intertissue flow of glutathione (GSH) as a tumor growth-promoting mechanism: interleukin 6 induces GSH release from hepatocytes in metastatic B16 melanoma-bearing mice.

B16 melanoma F10 (B16-F10) cells with high glutathione (GSH) content show high metastatic activity in vivo. An intertissue flow of GSH, where the liver is the main reservoir, can increase GSH content in metastatic cells and promote their growth. We have studied here possible tumor-derived molecular signals that could activate GSH release from hepatocytes. GSH efflux increases in hepatocytes isolated from mice bearing liver or lung metastases, thus suggesting a systemic mechanism. Fractionation of serum-free conditioned medium from cultured B16-F10 cells and monoclonal antibody-induced neutralization techniques facilitated identification of interleukin (IL)-6 as a tumor-derived molecule promoting GSH efflux in hepatocytes. IL-6 activates GSH release through a methionine-sensitive/organic anion transporter polypeptide 1- and multidrug resistance protein 1-independent channel located on the sinusoidal site of hepatocytes. Specific siRNAs were used to knock down key factors in the main signaling pathways activated by IL-6, which revealed a STAT3-dependent mechanism. Our results show that IL-6 (mainly of tumor origin in B16-F10-bearing mice) may facilitate GSH release from hepatocytes and its interorgan transport to metastatic growing foci.

Antenatal steroids and antioxidant enzyme activity in preterm infants: influence of gender and timing.

Antenatal steroids have improved the survival of preterm infants; however, the mechanism of action is not fully understood. We aimed to establish an association between antenatal steroids and antioxidant activity and postnatal oxidative stress. In a prospective cohort study, extremely preterm neonates receiving antenatal steroids (CORT) or not (NOCORT) were enrolled. An association between antenatal steroids and activities of antioxidant enzymes and glutathione cycle enzymes in cord blood was found. In addition, reduced oxidative stress (GSH/GSSG ratio, CORT vs. NOCORT, 35.68 + or - 12.20 vs. 28.38 + or - 9.92; p < 0.01) and, decreased oxidation of proteins (ortho-tyrosine/phenylalanine ratio, CORT vs. NOCORT, 8.66 + or - 2.45 vs. 12.55 + or - 4.41; p < 0.01) and DNA (8oxodG/2dG ratio, CORT vs. NOCORT, 6.73 + or - 2.18 vs. 9.53 + or - 3.83; p < 0.01) also was found. Antenatal steroids were associated with reduced oxygen supplementation, mechanical ventilation, and conditions such as bronchopulmonary dysplasia, intra-periventricular hemorrhage, or retinopathy of prematurity. The maximal effectiveness was when steroids were administered 2-4 days before delivery. Female preterm infants had less oxidative stress and increased antioxidant activity and better clinical outcomes than did male infants, independent of receiving or not antenatal steroids. Antenatal steroids are accompanied by a reduction in postnatal oxidative-stress-derived conditions and increased antioxidant enzyme activity. Both these effects seem to be influenced by specific timing and female gender.

Preterm resuscitation with low oxygen causes less oxidative stress, inflammation, and chronic lung disease.

OBJECTIVE: The goal was to reduce adverse pulmonary adverse outcomes, oxidative stress, and inflammation in neonates of 24 to 28 weeks of gestation initially resuscitated with fractions of inspired oxygen of 30% or 90%. METHODS: Randomized assignment to receive 30% (N = 37) or 90% (N = 41) oxygen was performed. Targeted oxygen saturation values were 75% at 5 minutes and 85% at 10 minutes. Blood oxidized glutathione (GSSG)/reduced glutathione ratio and urinary o-tyrosine, 8-oxo-dihydroxyguanosine, and isoprostane levels, isofuran elimination, and plasma interleukin 8 and tumor necrosis factor alpha levels were determined. RESULTS: The low-oxygen group needed fewer days of oxygen supplementation (6 vs 22 days; P < .01) and fewer days of mechanical ventilation (13 vs 27 days; P < .01) and had a lower incidence of bronchopulmonary dysplasia at discharge (15.4% vs 31.7%; P < .05). GSSG/reduced glutathione x 100 ratios at day 1 and 3 were significantly higher in the high-oxygen group (day 1: high-oxygen group: 13.36 +/- 5.25; low-oxygen group: 8.46 +/- 3.87; P < .01; day 3: high-oxygen group: 8.87 +/- 4.40; low-oxygen group: 6.97 +/- 3.11; P < .05). Urinary markers of oxidative stress were increased significantly in the high-oxygen group, compared with the low-oxygen group, in the first week after birth. GSSG levels on day 3 and urinary isofuran, o-tyrosine, and 8-hydroxy-2'-deoxyguanosine levels on day 7 were correlated significantly with development of chronic lung disease. CONCLUSIONS: Resuscitation of preterm neonates with 30% oxygen causes less oxidative stress, inflammation, need for oxygen, and risk of bronchopulmonary dysplasia.