Association between fluid balance and mortality in patients with septic shock: a post hoc analysis of the TRISS trial.

BACKGROUND: Several studies have shown an association between a positive fluid balance and increased mortality in patients with septic shock. This may have led to a more restrictive use of intravenous fluids. The association between fluid accumulation and mortality in the setting of a more restrictive use of intravenous fluids, however, is uncertain. We therefore aimed to investigate the association between a cumulative fluid balance 3 days after randomization and 90-day mortality in a recent Nordic multicentre cohort of patients with septic shock. METHODS: A post hoc analysis of patients from the Transfusion Requirements in Septic Shock (TRISS) trial treated for 3 days or more in the ICU after randomization. The patients were categorized into four groups depending on their weight-adjusted cumulative fluid balance after 3 days. We performed multivariable Cox regression analysis, adjusting for important prognostics (study site, age, chronic cardiovascular and chronic lung disease, haematologic malignancy, chronic dialysis, source of infection, baseline SOFA score and plasma lactate). RESULTS: The median cumulative fluid balance of the 841 included patients was 2480 ml (IQR 47-5045). The median time from ICU admission to inclusion in the trial was 22 h. The overall 90-day mortality was 52%. There was no statistically significant association between fluid balance 3 days from inclusion and 90-day mortality after the adjustment for the prognostics (P = 0.37). CONCLUSION: In our cohort of patients with septic shock and a comparably low cumulative fluid balance, there was no association between fluid balance and mortality. However, the study design and the limited power preclude strong conclusions. There is an urgent need for high-quality trials assessing the benefit and harm of different fluid volume strategies in patients with septic shock.

Urinary neutrophil gelatinase-associated lipocalin to hepcidin ratio as a biomarker of acute kidney injury in intensive care unit patients.

BACKGROUND: Labile iron is important in the pathogenesis of acute kidney injury (AKI). Neutrophil gelatinase-associated lipocalin (NGAL) and hepcidin control iron metabolism and are upregulated during renal stress. However, higher levels of urinary NGAL are associated with AKI severity whereas higher urinary hepcidin levels are associated with absence of AKI. We aimed to investigate the value of combining both biomarkers to estimate the severity and progression of AKI in intensive care unit (ICU) patients. METHODS: Urinary NGAL and hepcidin were quantified within 48 hours of ICU admission in patients with the systemic inflammatory response syndrome and early kidney dysfunction (oliguria for ≥ 2 hours and/or a 25 µmol/L creatinine rise from baseline). Diagnostic and prognostic characteristics were assessed by logistic regression and receiver operating characteristics (ROC) analysis. RESULTS: Of 102 patients, 26 had mild AKI and 28 patients had severe AKI on admission. Sepsis (21%), cardiac surgery (17%) and liver failure (9%) were primary admission diagnoses. NGAL increased (P=0.03) whereas hepcidin decreased (P=0.01) with increasing AKI severity. The value of NGAL/hepcidin ratio to detect severe AKI was higher than when NGAL and hepcidin were used individually and persisted after adjusting for potential confounders (adjusted OR 2.40, 95% CI 1.20-4.78). The ROC areas for predicting worsening AKI were 0.50, 0.52 and 0.48 for NGAL, 1/hepcidin and the NGAL/hepcidin ratio. CONCLUSION: The NGAL/hepcidin ratio is more strongly associated with severe AKI than the single biomarkers alone. NGAL and hepcidin, alone or combined as a ratio, were unable to predict progressive AKI in this selected ICU cohort.

Photophysics and ex vivo biodistribution of ß-cyclodextrin-meso-tetra(m-hydroxyphenyl)porphyrin conjugate for biomedical applications.

Low aqueous solubility of porphyrin-based photosensitizers hampers their clinical use in photodynamic therapy because of complex delivery. In this study, we explore meso-tetra(m-hydroxyphenyl)-21,23H-porphyrin (mTHPP), a potent photosensitizer, covalently attached to ß-cyclodextrin (CD-mTHPP) with a focus on topical delivery and cellular uptake. The photophysical properties of CD-mTHPP were examined using steady-state fluorescence and lifetime measurements verifying increased aqueous solubility. Confocal and fluorescence lifetime imaging microscopy on human squamous carcinoma cells (A431) evidenced a cytoplasmic uptake of CD-mTHPP in predominantly monomeric form. CD-mTHPP was also delivered to human skin ex vivo and the skin penetration was assessed using two-photon fluorescence microscopy. The results indicated that CD-mTHPP exhibits improved skin distribution compared to mTHPP alone using aqueous vehicles. Thus the CD-mTHPP conjugate demonstrates improved biodistribution ex vivo compared to mTHPP and is a promising multimodal system for photodynamic therapy.

Novel biomarkers of acute kidney injury and failure: clinical applicability.

Acute kidney injury (AKI) has a number of triggers, including ischaemia, nephrotoxins, radiocontrast, and bacterial endotoxins. It occurs in around one-third of patients treated in intensive care unit (ICU) and is even more prevalent in cardiac surgery patients. There is a higher mortality in patients with AKI compared with non-AKI counterparts, and in severe AKI requiring renal support, the 6 month mortality is >50%. Unlike the progressive development of biomarkers in cardiology, there have been few changes in kidney diagnostic markers. Creatinine is still used as an indicator of kidney function but not of the parenchymal kidney injury. Serum creatinine (sCr) concentration does not change until around 50% of kidney function is lost, and varies with muscle mass, age, sex, medications, and hydration status. The lag time between injury and loss of function, risks missing a therapeutic opportunity, and may explain the high associated mortality. Novel biomarkers of AKI- and failure include neutrophil gelatinase-associated lipocalin, N-acetyl-ß-d-glucosaminidase, kidney injury molecule-1, interleukin-18, and cystatin C. The pathophysiology associated with accumulation of these markers in plasma and urine is not clear, but a common denominator is inflammation. Some of these new AKI biomarkers may have clinical applicability in anaesthesia and intensive care in the future. It is possible that a 'kidney biomarker panel' will become standard before and after major surgery. If elevated or positive, the anaesthetist must take special care to optimize the patients after operation on the surgical wards or ICU to avoid further nephrotoxic insults and initiate supplementary care.

Enhanced intersystem crossing in donor/acceptor systems based on zinc/iron or free-base/iron porphyrins.

The deactivation pathways of the singlet excited state of a series of zinc or free-base donor porphyrins covalently linked by a bridge to a paramagnetic iron(III) chloride porphyrin acceptor have been studied. These donor-bridge-acceptor systems all share a similar geometry (25 A donor-acceptor center-to-center distance), but the bridges vary in electronic structure. In previously reported investigations of zinc/iron porphyrin systems, the fluorescence quenching of the donor has predominantly been assigned to electron transfer. However, for the porphyrin systems studied in this paper, we show that the dominant deactivation channels are enhanced intersystem crossing and singlet energy transfer. In both series, the intersystem crossing rate (S1-->T1) of the donor moiety is almost doubled in the presence of a paramagnetic high-spin metal-porphyrin acceptor. The significant spectral overlap of the donor fluorescence and acceptor absorption in both series allows for efficient singlet energy transfer (Forster mechanism). Furthermore, the bridging chromophores mediate energy transfer and the enhancement is inversely dependent upon the energy gap between the donor and bridge excited states. Although Marcus theory predicts thermodynamically favorable electron transfer to occur in the systems investigated, the quenching rate constants were found to be independent of solvent polarity, and no charge-separated state could be detected, indicating very small electronic coupling for electron transfer.

Patients with implantable cardioverter-defibrillators and their conceptions of the life situation: a qualitative analysis.

The implantable cardioverter-defibrillator (ICD) is today widely used for the treatment of sudden cardiac near-death episodes as a result of malignant ventricular dysrhythmia. After examining the literature, only four descriptive studies, all carried out in the USA, with a qualitative analysis based on ICD-patients' own perspectives on their life situation have been found. The aim of this study was to describe how patients living with an ICD-device in south-western Sweden conceive their life situation. As the focus was on patients' conceptions seen from a holistic perspective, an analysis inspired by phenomenography was employed on a strategic sample of 15 ICD-patients. Six categories emerged: a feeling of safety, a feeling of gratitude, a feeling of being, having a network, having a belief in the future, and gaining awareness. Although the findings cannot be generalized because of the descriptive research design, they illuminate the beneficial as well as intrusive effects of such a device, and emphasize the need for support groups for patients and families as well as further education for personnel in hospital and primary health care.

Glutathione metabolism in newborns: evidence for glutathione deficiency in plasma, bronchoalveolar lavage fluid, and lymphocytes in prematures.

Respiratory distress in premature newborns is associated with deficiency of surfactant in the bronchoalveolar lining fluid; this may be influenced by a local deficiency of antioxidants. Severe L-buthionine-S,R-sulfoximine-induced depletion of glutathione (GSH, a major antioxidant) in rodents is associated with lung type 2 cell lamellar body damage and decreased concentrations in lung and bronchoalveolar lavage fluid (BALF) of phosphatidyl choline (a major component of surfactant). At birth, prematurely born newborns (30-34 weeks) had lower peripheral venous plasma GSH concentrations than term (> 36 weeks) babies; these levels decreased further with increasing prematurity (< 27 weeks, with respiratory distress). On day 2, the peripheral venous plasma GSH concentrations reached a nadir, and the lowest levels were found in the most premature newborns. Lymphocyte GSH concentrations were lowest on day 2 and day 7, and in prematures (< 27 weeks, with respiratory distress) remained below adult lymphocyte GSH levels for at least 4 weeks. At birth, prematures (< 27 weeks, with respiratory distress) had a central plasma arterio-venous (A-V) GSH gradient across the lung (an estimate of lung uptake of GSH) of 0.72 +/- 0.15 (mean +/- SD) mumol/L; on day 2, the A-V gradient did not change significantly (0.49 +/- 0.09 mumol/L). At birth, these prematures had markedly decreased BALF GSH concentrations (compared with adult levels), and they were not significantly changed during the first 4 weeks of life. These results suggest that GSH deficiency is present in prematures and that it increases with the degree of prematurity. At birth, GSH deficiency will compromise the lungs' defense against oxidative stress injury. Oxidative stress is likely to increase if hyperoxic treatment is given for respiratory distress in these infants.

The chemical nature, purity and stability of 99mTc-N-(2,6-Diethyl-3-iodo-phenylcarbamoylmethyl)-iminodiacetic acid.

The chemical purity and stability of a radio pharmaceutical 99mTc-N-(2,6-diethyl-3-iodo-phenylcarbamoylmethyl)-iminodiacetic acid (IODIDA) has been studied using HPLC. The HPLC results indicate that IODIDA decomposed into at least three species after 99mTc labelling, and the amount of these varied with time. Mass spectra obtained by negative ion FABMS of no-carrier-added (NCA) Tc-labelled IODIDA showed, in addition to the molecular ion and the sodium adduct of N-(2,6-diethyl-3-iodo-phenylcarbamoylmethyl)-iminodiacetic acid, peaks corresponding to sodium containing dimers, trimers and tetramers. Several peaks were also found that may be assigned to 99Tc(I) containing dimers, trimers and tetramers.

L-2-oxothiazolidine-4-carboxylate, a cysteine precursor, stimulates growth and normalizes tissue glutathione concentrations in rats fed a sulfur amino acid-deficient diet.

The efficiency of L-2-oxothiazolidine-4-carboxylate, a cysteine precursor, in stimulating glutathione synthesis and growth was evaluated in growing rats. Animals were fed a sulfur amino acid-deficient diet (0.25% L-methionine and no cysteine) supplemented with L-2-oxothiazolidine-4-carboxylate (0.35%) for 3 wk and compared with age-matched animals receiving the sulfur amino acid-deficient diet alone. Rats fed the sulfur amino acid-deficient diet had lower glutathione concentrations in bronchoalveolar lining fluid, lung, lymphocytes, and liver than rats fed a sulfur amino acid-deficient diet supplemented with L-2-oxothiazolidine-4-carboxylate. Rats fed the supplemented diet had normal tissue and bronchoalveolar lining fluid glutathione levels. Central venous plasma glutathione concentrations, mostly reflecting liver excretion, were less affected by L-2-oxothiazolidine-4-carboxylate supplementation. Rats fed L-2-oxothiazolidine-4-carboxylate supplementation had normal weight gain compared with a much lower weight gain in animals fed the sulfur amino acid-deficient diet alone. Thus, L-2-oxothiazolidine-4-carboxylate increased tissue glutathione concentrations and stimulated growth in rats. The lung glutathione status of the rats was reflected by glutathione concentrations in lymphocytes and the bronchoalveolar lining fluid, but not by the central venous plasma glutathione concentrations.

Liver function and halothane-diethyl-ether azeotrope anaesthesia. Re-evaluation of an obsolete drug with special reference to early postoperative effects.

A general anaesthetic drug that fulfils requirements for use under difficult circumstances is the inhalation agent halothane-diethyl-ether (HE) azeotrope. Although both halothane and diethyl ether have been described in detail, their effect on the liver when given together as an azeotrope has not been systematically characterised. The effect on liver function was evaluated and compared with the effects of halothane anaesthesia (H) and spinal anaesthesia with tetracaine (S), the last named serving as controls. The series consisted of 33 healthy men (ASA 1-2) receiving no medication and scheduled for inguinal hernia repair. The patients were randomly allocated to receive HE, H or S. The following parameters were estimated the day before surgery and on the first postoperative day: liver cell metabolism (bile acids, unconjugated bilirubin), cell integrity (aminotransferases), synthesizing capacity (Prothrombin complex), cholestasis (conjugated bilirubin, alkaline phosphatase, gamma-glutamyl transpeptidase), and global liver function (chenodeoxycholic load test). No major differences emerged between the groups. Unconjugated bilirubin was increased in all groups. Prothrombin complex activity was reduced in all groups. Conjugated bilirubin was increased in the H group. The oral bile acid load test and the fasting bile acid were unaltered by anaesthesia in all groups. No major impact on liver cell function was seen in the early post-operative period after HE azeotrope anaesthesia. The findings support our view that HE azeotrope could be considered as an alternative anaesthetic agent under field conditions.

Effect of ascorbate or N-acetylcysteine treatment in a patient with hereditary glutathione synthetase deficiency.

A 45-month-old girl with 5-oxoprolinuria (pyroglutamic aciduria), hemolysis, and marked glutathione depletion caused by deficiency of glutathione synthetase was followed before and during treatment with ascorbate or N-acetylcysteine. High doses of ascorbate (0.7 mmol/kg per day) or N-acetylcysteine (6 mmol/kg per day) were given for 1 to 2 weeks without any obvious deleterious side effects. Ascorbate markedly increased lymphocyte (4-fold) and plasma (8-fold) levels of glutathione. N-Acetylcysteine also increased lymphocyte (3.5-fold) and plasma (6-fold) levels of glutathione. After these treatments were discontinued, lymphocyte and plasma glutathione levels decreased rapidly to pretreatment levels. Ascorbate treatment was extended for 1 year, and lymphocyte (4-fold) and plasma (2- to 5-fold) glutathione levels remained elevated above baseline. In parallel, the hematocrit increased from 25.4% to 32.6%, and the reticulocyte count decreased from 11% to 4%. The results demonstrate that ascorbate and N-acetylcysteine can decrease erythrocyte turnover in patients with hereditary glutathione deficiency by increasing glutathione levels.

Glutathione ester delays the onset of scurvy in ascorbate-deficient guinea pigs.

Previous studies showed that administration of ascorbate to glutathione (GSH)-deficient newborn rats and guinea pigs prevented toxicity and mortality and led to increased tissue and mitochondrial GSH levels; ascorbate thus spares GSH. In the present work, we tried to answer the converse question: Does administration of GSH spare ascorbate? Because administered GSH is not well transported into most cells, we gave GSH monoethyl ester (which is readily transported and converted into GSH intracellularly) to guinea pigs fed an ascorbate-deficient diet. We found that treatment with GSH ester significantly delays appearance of the signs of scurvy and that this treatment spares ascorbate; thus, the decrease of tissue levels of ascorbate was delayed. The findings support the conclusions that (i) GSH is essential for the physiological function of ascorbate because it is required in vivo for reduction of dehydroascorbate and (ii) there is metabolic redundancy and overlap of the functions of these antioxidants. The sparing effect of GSH in scurvy may be mediated through an increase in the reduction of dehydroascorbate (which would otherwise be degraded) and to antioxidant effects of GSH that are also produced by ascorbate. Other studies indicate that GSH deficiency in adult mice stimulates ascorbate synthesis in liver. During this work we found that administration of GSH itself is highly toxic to ascorbate-deficient guinea pigs when given in divided i.p. doses totaling 3.75 mmol/kg daily.

Glutathione deficiency increases hepatic ascorbic acid synthesis in adult mice.

Glutathione deficiency, induced in adult mice by administering buthionine sulfoximine (an inhibitor of glutathione synthesis), led to a rapid and substantial increase in ascorbate in the liver. This effect was apparent 2-4 hr after giving the inhibitor; subsequently, the level of ascorbate decreased and that of dehydroascorbate increased markedly, supporting the conclusion that glutathione functions physiologically to keep ascorbate in its reduced form. In kidney and lung also, ascorbate levels decreased, and dehydroascorbate increased. Increased synthesis of ascorbate in glutathione-deficient adult mice seems to protect against tissue damage. In contrast, newborn rats, which (like guinea pigs and humans) apparently do not synthesize ascorbate, suffer severe damage to liver and other organs; previous studies showed that administration of ascorbate prevents such tissue damage. The findings support the view that the antioxidant actions of glutathione and ascorbate are closely linked and involve a mechanism in which decrease of the glutathione level, perhaps associated with an oxidative event, stimulates ascorbate synthesis.

Ascorbic acid prevents oxidative stress in glutathione-deficient mice: effects on lung type 2 cell lamellar bodies, lung surfactant, and skeletal muscle.

Glutathione deficiency in adult mice leads to lung type 2 cell lamellar body and mitochondrial damage; as reported here, these effects are associated with marked decrease of the levels of phosphatidylcholine (the main component of lung surfactant) in the lung and the bronchoalveolar lining fluid. Severe mitochondrial damage was also found in skeletal muscle. Treatment with ascorbate (1-2 mmol per kg of body weight per day), which led to greatly increased (approximately 2-fold) levels of lung and muscle mitochondrial glutathione, prevented damage to lamellar bodies and mitochondria as well as the decline of phosphatidylcholine levels in lung and alveolar lining fluid. The findings indicate that glutathione deficiency leads to depletion of lung surfactant and that this can be prevented with ascorbate. Administration of ascorbate spares glutathione and prevents cellular damage. Lamellar body degeneration in glutathione deficiency appears to be associated with oxidative damage to the perilamellar membrane, which contains the enzymes required for phosphatidylcholine synthesis. It is notable that although severe glutathione deficiency is lethal to newborn rats, which apparently do not synthesize ascorbate, adult mice are better able to survive such a deficiency because they can synthesize ascorbate. The present studies, which suggest that high doses of ascorbate may be of therapeutic value, emphasize that ascorbate and glutathione have actions in common and that they function together in a physiologically significant antioxidant system.

Sulfur amino acid metabolism in hepatobiliary disorders.

Sulfur amino acid metabolism was studied in patients with mild to severe forms of liver dysfunction and compared with that of healthy controls. Patients with mild liver dysfunction (for example, Gilbert's syndrome) had a normal sulfur amino acid metabolism. With increased inflammatory activity and cirrhosis (for example, chronic active hepatitis, alcohol-induced cirrhosis, and hepatic coma) a decreased ability to metabolize methionine (to cysteine, with cystathionine accumulation) and cysteine (to inorganic sulfate, with thiosulfate and N-acetylcysteine accumulation) was found. In contrast, transaminative metabolism of sulfur amino acids was preserved in patients with advanced forms of liver dysfunction, suggesting that transamination of sulfur amino acids is performed not only in the liver but also in extrahepatic tissues. Some implications of these findings are discussed.

Mitochondrial glutathione in hypermetabolic rats following burn injury and thyroid hormone administration: evidence of a selective effect on brain glutathione by burn injury.

Cerebral cortex, heart, skeletal muscle, and liver mitochondrial glutathione (GSH) levels in severely burned rats are decreased to between approximately 50% to 70% of sham-operated, normally fed controls. In semistarved rats, weight-matched with burned rats, mitochondrial GSH levels in these tissues are decreased to between approximately 70% to 91% of those in sham-operated rats. Total GSH levels in peripheral tissues and brain are decreased to approximately 60% to 65% of control levels in rats with burn injury and food restriction, suggesting a higher mitochondrial GSH turnover in burned rats than in semistarved rats, probably because of higher "stress hormone" levels in burned rats than in semistarved rats. Cerebral cortex mitochondrial GSH levels are unaffected by variations in thyroid hormone status, but liver mitochondrial GSH levels are decreased by triiodothyronine and increased by propylthiouracil. The present results suggest that mitochondrial GSH is not only regulated by the rate of GSH synthesis in the cytosol, but seems to be under hormonal influence as well; stress hormones and triiodothyronine may decrease mitochondrial GSH by increasing mitochondrial oxygen consumption with increased reactive oxygen species formation or by increasing GSH exchange between mitochondria and the cytosol. These findings may be of importance therapeutically in increasing antioxidative defenses to limit oxidative stress injury in hypermetabolic patients.

Inhibition of glutathione synthesis in the newborn rat: a model for endogenously produced oxidative stress.

A model for oxidative stress is described in which glutathione (GSH) synthesis is selectively blocked in newborn rats by administration of L-buthionine-(S,R)-sulfoximine (BSO). In this model, the normal endogenous physiological formation of reactive oxygen species is largely unopposed, and therefore oxidative tissue damage occurs; because GSH is used for reduction of dehydroascorbate, tissue ascorbate levels decrease. In lung there are decreased numbers of lamellar bodies and decrease of intraalveolar surfactant. Proximal renal tubular, hepatic, and brain damage also occur. A diastereoisomer of BSO that does not inhibit GSH synthesis, L-buthionine-R-sulfoximine, does not produce toxicity; this control experiment renders it unlikely that the observed effects of BSO are produced by the sulfoximine moiety itself. There is correlation between the decrease of mitochondrial GSH levels and mitochondrial and cell damage. Oxidative stress as evaluated by mitochondrial damage and mortality can be prevented by treatment with GSH esters or ascorbate. There is apparent linkage between the antioxidant actions of GSH and ascorbate. This model, which may readily be applied to evaluation of the efficacy of other compounds in preventing oxidative stress, offers an approach to study of other effects of GSH deficiency (e.g., on lipid metabolism, hematopoiesis), and closely resembles oxidative stress that occurs in certain human newborns and in other clinical states.

Glutathione deficiency decreases tissue ascorbate levels in newborn rats: ascorbate spares glutathione and protects.

Glutathione deficiency in newborn rats, produced by administration of L-buthionine-(S,R)-sulfoximine, a transition-state inactivator of gamma-glutamylcysteine synthetase, decreases ascorbate levels of kidney, liver, brain, and lung. These tissues, especially their mitochondria, undergo severe damage and the animals die within a few days. When glutathione levels are markedly decreased, ascorbate levels decrease leading to formation of dehydroascorbate, which is degraded. Ascorbate has high antioxidant activity, but it (and other antioxidants such as alpha-tocopherol) must be maintained in reduced forms. These studies show in vivo that an important function of glutathione is to maintain tissue ascorbate. Administration of large doses of ascorbate (but not of dehydroascorbate) to buthionine sulfoximine-treated newborn rats decreases mortality, leads to normal levels of ascorbate, and spares glutathione. Newborn rats given lower doses of buthionine sulfoximine develop cataracts that, as shown previously, can be prevented by giving glutathione monoester; as found here, such cataracts can be partially prevented by administration of high doses of ascorbate or dehydroascorbate. Ascorbate spares glutathione indicating that these compounds have similar antioxidant actions. Ascorbate may have reductive functions that are not efficiently performed by glutathione. Although glutathione normally functions to maintain ascorbate, alpha-tocopherol, and other cellular components in reduced states, ascorbate can serve as an essential antioxidant in the presence of severe glutathione deficiency.

Glutathione deficiency leads to mitochondrial damage in brain.

Glutathione deficiency induced in newborn rats by giving buthionine sulfoximine, a selective inhibitor of gamma-glutamylcysteine synthetase, led to markedly decreased cerebral cortex glutathione levels and striking enlargement and degeneration of the mitochondria. These effects were prevented by giving glutathione monoethyl ester, which relieved the glutathione deficiency, but such effects were not prevented by giving glutathione, indicating that glutathione is not appreciably taken up by the cerebral cortex. Some of the oxygen used by mitochondria is known to be converted to hydrogen peroxide. We suggest that in glutathione deficiency, hydrogen peroxide accumulates and damages mitochondria. Glutathione, thus, has an essential function in mitochondria under normal physiological conditions. Observations on turnover and utilization of brain glutathione in newborn, preweaning, and adult rats show that (i) some glutathione turns over rapidly (t 1/2, approximately 30 min in adults, approximately 8 min in newborns), (ii) several pools of glutathione probably exist, and (iii) brain utilizes plasma glutathione, probably by gamma-glutamyl transpeptidase-initiated pathways that account for some, but not all, of the turnover; thus, there is recovery or transport of cysteine moieties. These studies provide an animal model for the human diseases involving glutathione deficiency and are relevant to oxidative phenomena that occur in the newborn.

High-affinity transport of glutathione is part of a multicomponent system essential for mitochondrial function.

Glutathione, an essential cellular antioxidant required for mitochondrial function, is not synthesized by mitochondria but is imported from the cytosol. Rat liver mitochondria have a multicomponent system that underlies the remarkable ability of mitochondria to take up and retain glutathione. At external glutathione levels of less than 1 mM, glutathione is transported into the mitochondrial matrix by a high-affinity component (Km, approximately 60 microM; V max, approximately 0.5 nmol/min per mg of protein), which is saturated at levels of 1-2 mM and stimulated by ATP. Another component has lower affinity (Km, approximately 5.4 mM; Vmax, approximately 5.9 nmol/min per mg of protein) and is stimulated by ATP and ADP. Both components are inhibited by carbonylcyanide p-(trifluoromethoxy)phenylhydrazone (FCCP), glutamate, and ophthalmic acid. Increase of extramitochondrial glutathione promotes uptake and exchange; the intermembranous space seems to function as a recovery zone that promotes efficient recycling of matrix glutathione. The findings are in accord with in vivo data showing that (i) rapid exchange occurs between mitochondrial and cytosolic glutathione, (ii) lowering of cytosolic glutathione levels (produced by administration of buthionine sulfoximine) decreases export of glutathione from mitochondria to cytosol, and (iii) administration of glutathione esters increases glutathione levels in mitochondria more than those in the cytosol.