Brain, Vol. 126, No. 5, 1224-1230,
May 2003
© 2003 Guarantors of Brain
doi: 10.1093/brain/awg116
Non protein bound iron as early predictive marker of neonatal brain damage
Department of Paediatrics, Obstetrics and Reproductive Medicine, University of Siena, Italy
Correspondence to: Professor Giuseppe Buonocore, MD, Department of Paediatrics, Obstetrics and Reproductive Medicine, University of Siena, Policlinico ‘Le Scotte’, V. le Bracci 36, 53100 Siena, Italy E-mail: buonocore{at}unisi.it
Received September 29, 2002. Revised December 12, 2002. Accepted December 12, 2002.
 |
Summary |
|---|
 Top Summary IntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Of the approximately 130 million births worldwide each year, four million infants will suffer from birth asphyxia and, of these, one million will die and a similar number will develop serious sequelae. Before being able to develop effective interventions, a better understanding of the pathophysiological mechanisms leading to brain injury and an early identification of babies at high risk for brain injury are required. This study tests the predictivity of traditional and new markers of foetal oxidative stress in relation to neurodevelopmental outcome in 384 newborn infants. The results indicate plasma non protein bound iron as the best early predictive marker of neurodevelopmental outcome, with 100% sensitivity and 100% specificity for good neurodevelopmental outcome at 0–1.16 µmol/l, and for poor neurodevelopmental outcome at values >15.2 µmol/l. The number of children with values between 1.16 and 15.2 were 195. Common use of this predictive marker in neonatology units will improve the ability of clinicians to identify those newborn babies who will develop neurodisability.
Keywords: iron; free radicals; brain injury; infant; newborn
Abbreviations: AOPP= advanced oxidative protein products; Hx = Hypoxanthine, NPBI = non protein bound iron; NRBC = nucleated red blood cell; TH = total hydroperoxides; ROC curve = receiver operating characteristic curve
|
Introduction |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Some progress has been made in treating high-risk term newborn babies and small preterm infants. This has resulted in major reductions in neonatal mortality and neurological handicaps due to perinatal injury, but certain pre- and post-natal conditions causing brain damage are still difficult to identify and treat. As a consequence, the risk of permanent damage remains unacceptably high: 2–6 infants per 1000 births develop hypoxic-ischaemic encephalopathy, resulting in sequelae of differing severity (World Health Organization, 1991
). Efforts to understand and prevent neonatal cerebral injury are therefore worthwhile.
To develop effective preventive measures, early identification of babies at high risk for brain injury is necessary. Although several methods (scoring systems, markers, EEG, cerebral function monitoring, etc.) are developed for early identification of neonates who may benefit from intervention, these indexes are reported to have a limited predictive value for death or survival with abnormal neurodevelopmental outcome (Goldenberg et al., 1984; Low et al., 1985; Ruth and Raivio, 1988; International Neonatal Network, 1993; Greisen, 1994).
Recently, we demonstrated the damaging effect of free radicals in preterm hypoxic babies in perinatal period (Buonocore et al., 2000, 2002a). Reactions involving free radical toxicity in neonates have a high potential for tissue damage, and particularly brain damage because fast-growing tissues are especially sensitive to free radicals (Goplerud et al., 1992; Sorg et al., 1997; Palmer et al., 1999). Free radical release is followed by endothelial cell damage, haemostatic abnormalities, inflammatory reactions, brain cell damage due to astrocyte dysfunction, n-methyl-D-aspartate receptor impairment and synaptosome structural damage (Dammann and Leviton, 1997; Bracci, 1999; Mishra and Delivoria-Papadopoulos, 1999; Akhter et al., 2001). Free radicals may be generated by several sources including phagocyte activation, catecholamine metabolism, mitochondrial dysfunction, arachidonic acid cascade and Fenton’s reaction driven by non protein bound iron (NPBI). Indirect markers of increased free radical release and perinatal brain injury have recently emerged with reports of increased advanced oxidative protein products (AOPP) and NPBI in erythrocytes and plasma of hypoxic newborns (Dorrepaal et al., 1996; Buonocore et al., 1998a,b, 2000, 2001). Recently, we found that increased nucleated red blood cell (NRBC) count at birth is helpful in identifying perinatal hypoxia and in predicting neurodevelopmental outcome (Buonocore et al., 1999) Although experimental studies have demonstrated the key role of NPBI and free radicals in the development of hypoxic-ischaemic brain damage, the relationships between plasma and red cell markers of oxidative stress and brain damage remain unclear.
The present study was undertaken to evaluate the predictive role of certain new and conventional markers of intrauterine oxidative stress in cord blood for neurodevelopmental outcome.
|
Material and methods |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Patients
Four hundred of the 2902 babies born in the Neonatology Division (University of Siena, Italy) between 1st June 1997 and 31st May 2000 were enrolled in this prospective study according to a random algorithm using the Minitab statistical software package (Mintab Inc., State College, PA, USA). A sample size of 400 was judged sufficiently numerous for meaningful statistical evaluation of new items (with unknown median and range of normal values) calculating a defection <10% at follow-up.
Babies not born in the clinic or with major congenital malformations, inherited errors of metabolism, blood group incompatibility, evidence of congenital infections, multiple gestation, anaemia or diabetic mothers were excluded. The study was masked throughout.
The number lost to follow-up during the study period was 16: one very low birth weight infant died; eight did not present for neurodevelopmental examination or did not complete it; three could not be traced; and four declined the invitation because the infant was well and supervised regularly by a paediatrician.
The final cohort consisted of 384 newborns with gestational ages from 24 to 42 weeks: 225 >36 weeks, 90 from 32 to 36 weeks and 69 <32 weeks and birth weight <1500 g (birth weight ranged from 490 to 5300 g). Birth was accomplished by: normal delivery 129; elective caesarean section 178; and emergency caesarean section 77.
The study was approved by the Human Ethics Committee of the Medical Faculty, University of Siena and written informed consent was obtained from parents.
Methods
Heparinized blood samples were drawn from the umbilical vein after cord clamping immediately after delivery. Complete blood cell count and gas analysis were performed.
Variables selected for statistical evaluation were: gestational age; Apgar–1 min; Apgar–5 min; birth weight; pH; base deficit; hypoxanthine (Hx, a recognized reliable biochemical marker of hypoxia); NRBC; NPBI; total hydroperoxides (TH); and AOPP.
Total white blood cell count was initially determined in the laboratory of our neonatal section. NRBC count was estimated by light microscope examination of May–Grunwald–Giemsa-stained blood smears. NRBC count was expressed as absolute erythroblast count (NRBC per mm3) (Buonocore et al., 1999). Blood gas analysis was performed with a Radiometer ABL 505 analyser (Radiometer, Copenhagen, Denmark) immediately after blood sampling. The blood was centrifuged and plasma and buffy coats were removed. To avoid storage effects, all analyses (including Hx, NPBI, AOPP and TH) were carried out in plasma within 2 h of blood sampling.
Hx levels were evaluated using a Varian Vista 5500 high performance liquid chromatograph equipped with a variable wavelength UV detector (Varian, model 4290, Palo Alto, CA, USA) as described previously (Buonocore et al., 2000).
AOPP were measured as described by Witko-Sarsat and colleagues using spectrophotometry on a microplate reader (Witko-Sarsat et al., 1996). The instrument was calibrated with chloramine-T solutions, which absorb at 340 nm in the presence of potassium iodide. In test wells, 200 µl of plasma diluted 1:5 in phosphate-buffered saline (PBS) was distributed on a 96-well microtitre plate, and 20 µl of acetic acid was added. In standard wells, 10 µl of 1.16M potassium iodide was added to 200 µl of chloramine-T solution (0–100 µmol/l) followed by 20 µl of acetic acid. The absorbance of the reaction mixture was read immediately at 340 nm on the microplate reader against a blank containing 200 µl of PBS, 10 µl of potassium iodide and 20 µl of acetic acid. Since the absorbance of chloramine-T at 340 nm is linear up to 100 µmol/l, AOPP concentrations were expressed as µmol/ml chloramine-T equivalents.
TH production was measured with the d-ROMs Kit (Diacron Srl, Grosseto, Italy) expressed in conventional arbitrary units, called Carr units (U.CARR) as described previously (Buonocore et al., 2000).
NPBI plasma levels were detected by HPLC using the method described by Kime and colleagues, but with some modifications (Kime et al., 1996). The system was operated isocritically at a pressure of 
115 Bar and flow of 0.75 ml/min. The detection wavelength was 450 nm with a reference wavelength at 620 nm. A low affinity ligand, disodium nitryloacetic acid, was used first to complex all low molecular weight iron and iron non-specifically bound to serum proteins such as albumin. Because it does not remove iron bound to transferrin or ferritin, a two-step filtration process was used: filtration with a 100 kDa molecular weight cut-off Whatman ultracentrifuge filter (Whatman, Maidstone, Kent, UK) was followed by filtration with one having a cut-off at 20 kDa. The filtrate was analysed by direct injection into a reverse phase liquid chromatography system, utilizing precolumn derivatization with the high-affinity iron chelator CP22 (3-hydroxyl-1-propyl-2-methyl-pyridin-4-one hydrochloride).
Hypoxic-ischaemic brain injury was ascertained by pulsed Doppler sonograms of the cerebral arteries and brain ultrasonography performed within 48 h of birth. Neonatal neurodevelopmental examinations were performed at a mean postmenstrual age of 38 weeks according to the criteria of Allen and Capute (Allen and Capute, 1989). Follow-up visits were scheduled at age 1, 2, 4, 12 and 24 months. Physical and neurological examinations were performed by the same neonatal neurologist. Motor development was assessed in the areas of muscle tone, primitive reflexes, automatic reactions, and head and trunk tone. The evaluation included quantitative changes and quality of performance in developmental patterns and milestones. Three to four transfontanellar cranial ultrasound examinations were performed within the first 6–8 months of follow-up by one experienced neonatologist, blind to outcome. Hearing tests included free-field behavioural audiometry and timpanography, and were carried out by an audiologist. Children were also seen for formal assessment by a paediatric ophthalmologist. Formal developmental testing was performed with the Bayley Scales of Infant Development (Bayley, 1969), an age-adjusted standardized test that compares motor development and cognitive skills to established nomograms with a psychomotor developmental index and a mental developmental index.
Fifty-one of the 384 babies (13.3%) showed persistent abnormal neurodevelopmental status at 24 months. This percentage was similar to that observed in whole population of newborns (n = 2902) in the study period.
Maternal/pregnancy variables capable of affecting the oxidation status, e.g. pre-eclampsia, drug use, smoking and alcohol abuse were considered to exclude independent effects on neurological outcome.
Statistical analysis
Summary statistics of data were expressed as mean ± SD, 95% CI (confidence interval) for the mean, median and 25th–75th percentiles (interquartile range). The Kolmogorov–Smirnov test was performed on populations and subpopulations to check normal distribution of data. The Mann–Whitney U test for unpaired data was used for continuous variables.
The effects of potential confounding factors (such as gestational age and birth weight) on relationships between follow-up and other variables or their associations, and the identification of subsets of independent variables that could be good predictors of the dichotomous follow-up variable were checked by multivariate logistic regression models with forward stepwise conditional selection of variables.
The above tests were performed using the SPSS V.6.1 for Windows statistical package (SPSS Inc, Chicago, IL, USA). Receiver operating characteristic (ROC) curve data were analysed using the STATA 6 statistical package (StataCorp, College Station, Texas, USA).
|
Results |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
The results of descriptive analysis and differences between babies with normal and abnormal outcome at 24 months are reported in Table 1. No differences in maternal/pregnancy variables were observed between babies with normal and abnormal outcome at 24 months.
|
The Mann–Whitney U test for unpaired data showed significantly higher plasma levels of Hx, NRBC, NPBI, TH and AOPP in cord blood of babies with abnormal development than in babies with normal neurodevelopmental outcome at 24 months. Conversely, birth weights, gestational ages and Apgar–1 min and Apgar–5 min scores were significantly lower in babies with abnormal development than in babies with normal neurodevelopmental outcome at 24 months.
After codification of neurodevelopmental outcome (dependent variable) as 0 for normal and 1 for neurological impairment, all independent variables (gestational age, Apgar–1 min and Apgar–5 min, birth weight, pH, base deficit, Hx, NRBC, NPBI, TH and AOPP) checked by univariate analysis were considered in a multivariate logistic regression analysis with forward stepwise conditional selection of variables. The cut-off value of significance chosen for each variable was 0.05 to enter in the model and 0.1 for removal.
Variables meeting the above criteria were in order of decreasing significance: NPBI (model 
2 = 79.7, P = 0.0000), gestational age (model 2 = 117.8, P = 0.0000), pH (model 2 = 125.4, P = 0.0000), TH (model 2 = 137.2, P = 0.0000) Apgar–5 min (model 2 = 144.2, P = 0.0000), NRBC (model 2 = 148.2, P = 0.0000), Hx (model 2 = 153.2, P = 0.0000); base deficit, AOPP, Apgar–1 min and birth weight were not significant.
Partial correlation between the dependent and each independent variable (R) with relative estimated coefficients (B), significance (P) and exponential or odds ratio Exp(B) showed that the logistic model correctly predicted 92.68% of cases (Table 2).
|
All ROC curves enabled impairment and normal neurodevelopment to be distinguished; the largest area was that of NPBI showing 100% sensitivity and 100% of true negative fraction for a good neurological outcome with ranges between 0 to 1.16 µmol/l. Conversely, 100% of poor neurodevelopmental outcome was observed for NPBI values >15.2 µmol/l. The NPBI plotted curve indicated 3.83 µmol/l as the best predictive threshold with a sensitivity of 88.2% (95% CI 76.1–95.5) and specificity of 79.3% (95% CI 74.5–83.5) (Table 3 and Fig. 1).
|
|
|
Discussion |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Tissue damage has been demonstrated when NPBI concentrations are high due to iron overload and transferrin saturation (Berger et al., 1995
; McNamara et al., 1999). Plasma NPBI is reported to be higher in human newborns than at any other age (Dorrepaal et al., 1996; Buonocore et al., 2001). Adult plasma does not contains NPBI: transferrin saturation is only 20–35% and conversion of ferrous to ferric iron which binds to transferrin is facilitated by ferroxidase in plasma proteins (Lindeman et al., 1992). In the present study, NPBI levels in cord blood were significantly higher in neonates with a poor outcome than in those with normal neurodevelopmental outcome. The highest sensitivity and specificity of plasma NPBI strongly indicate NPBI at birth as a reliable index of brain damage. It is worthy to note that no babies with a normal value of NPBI (range within 0 and 1.16 µmol/l) subsequently exhibited neurological abnormalities.
NPBI in plasma causes release of hydroxyl radicals by superoxide and hydrogen peroxide possibly via iron–oxygen complexes (Schafer et al., 2000; Buonocore and Perrone, 2002b).
On the other hand, newborn infants, especially preterm ones, have high transferrin saturation. This condition can become crucial during acidosis, which is known to induce removal of iron from safe sites (Schafer and Buettner, 2000). Acidosis during cerebral ischaemia potentiates oxidative neuronal death resulting from impaired antioxidant enzyme functions. Increased intracellular free iron levels in rat brain in vivo and neuronal death resulting from iron-induced free radical release and reduced activity of antioxidant enzyme have been described (Waterfall et al., 1996; Ying et al., 1999). Enhanced proteolytic activity occurring in injured tissue also releases iron from storage proteins (Rothman et al., 1992). When NPBI gains access to the extracellular space, its uptake by cells is enhanced by intracellular calcium and, paradoxically, by increased levels of intracellular iron (Kaplan et al., 1991).
It is still unclear whether NPBI is only the consequence of removal of iron by transferrin due to acidosis or increased redox iron released by other sources. Some authors suggest that iron may be removed from storage protein because of the very low plasma and tissue pH typical of asphyxiating foetuses and newborns (Dorrepaal et al., 1996). The complexity of iron-ferritin reactions makes this hypothesis difficult to demonstrate.
In our previous papers, we reported increased red cell NPBI content in hypoxic newborns (Buonocore et al., 1998, 2001). These data are in line with recent reports of increased haemoglobin auto-oxidation during hypoxia and the subsequent demonstration of increased NPBI release from newborn erythrocytes during incubation under hypoxic conditions (Dorrepaal et al., 1996). Furthermore, our preliminary studies on red cells incubated under hypoxic conditions suggest that redox iron is released by red cells irrespective of haemolysis (Ciccoli et al., 2001). Thus, we speculated that besides acidosis, NPBI may be due to iron released from red cells stressed by hypoxia.
The brain may be especially at risk from free radical-mediated injury because neuronal membranes are rich in polyunsaturated fatty acids (Halliwell, 1992). Moreover, the iron-binding capacity of CSF is low (low transferrin concentrations), and most of the iron could be in its active ferrous form because of the high concentrations of vitamin C and low concentrations of ceruloplasmin in CSF (Gutteridge, 1992a, b).
It is, therefore, possible that plasma NPBI is a predictive marker of free radical-mediated brain damage. In testing the predictivity of NPBI for neurodevelopmental outcome against other possible perinatal variables in newborn infants (gestational age, birth weight, Apgar–1 min, Apgar–5 min, pH, base deficit, AOPP, Hx, TH, NRBC and NPBI) by univariate analysis, we found that all variables except pH and base deficit showed significant statistical differences between babies with normal and abnormal outcome at 24 months, This suggests their possible utility for recognizing brain damage.
Of the subset of independent variables tested by multivariate analysis and multivariate logistic regression analysis for predictivity of neurodevelopmental outcome, NPBI ranked first in the hierarchy followed by pH, gestational age, Apgar–5 min, TH, NRBC and Hx. This result strongly supports the role of these factors, and particularly NPBI, in predicting brain damage. To verify the reliability of this hypothesis, we used cut-off values obtained by ROC analysis.
When ROC curves were used to test the performance, diagnostic accuracy, practical clinical value and usefulness of the above measures for predicting neurodevelopmental outcome, they indicated reliable sensitivity and specificity thresholds for each value of NPBI. NPBI area showed better predictivity for babies with poor outcome than those with normal neurological outcome. It was the first marker included in the logistic regression model and showed the best partial correlation coefficient and a greater area under the ROC curve than the other variables.
In conclusion, plasma NPBI appears to be a reliable new early indicator of intrauterine oxidative stress and brain injury. Its use in neonatology units will improve the ability of clinicians to predict those newborn babies who will develop neurodisability. Reliable prediction is essential if therapeutic research is to be accurately targeted.
|
Acknowledgements |
|---|
This study was supported by grants from the Italian Ministry for the University and Scientific-Technological Research (MIUR 2001: identification of etiopathogenetic factors characteristics of newborn at high risk of brain damage in perinatal period. Clinical and experimental study). The authors had no financial arrangements nor were there other factors that could compromise the objectivity of the research.
|
References |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Akhter W, Ashraf QM, Zanelli SA, Mishra OP, Delivoria-Papadopoulos M. Effect of graded hypoxia on cerebral cortical genomic DNA fragmentation in newborn piglets. Biol Neonate 2001; 79: 187–93.[CrossRef][ISI][Medline]
Allen MC, Capute AJ. Neonatal neurodevelopmental examination as a predictor of neuromotor outcome in premature infants. Pediatrics 1989; 83: 498–506.
Bayley N. Bayley scales of infant development. New York: Psychological Corporation; 1969.
Berger HM, Mumby S, Gutteridge JMC. Ferrous ions detected in iron-overloaded cord blood plasma from preterm and term babies: implications for oxidative stress. Free Radic Res 1995; 22: 555–9.[ISI][Medline]
Bracci R. Free radicals in neonatal brain disorders. Possible benefits of antioxidants. In: Cosmi EV, Marinoni E, Di Iorio R, editors. New Technologies in reproductive medicine, neonatology and gynaecology. New York: Parthenon Publishing; 1999. pp. 131–9.
Buonocore G, Zani S, Perrone S, Caciotti B, Bracci B. Intraerythrocyte nonprotein-bound iron and plasma malondialdehyde in the hypoxic newborn. Free Radic Biol Med 1998a; 25: 766–70.[CrossRef][ISI][Medline]
Buonocore G, Zani S, Sargentini I, Gioia D, Signorini C, Bracci R. Hypoxia-induced free iron release in the red cells of newborn infants. Acta Paediatr 1998b; 87: 77–81.[CrossRef][ISI][Medline]
Buonocore G, Perrone S, Gioia D, Gatti MG, Massafra C, Agosta R, et al. Nucleated red blood cell count at birth as an index of perinatal brain damage. Am J Obstet Gynecol 1999; 181: 1500–5.[CrossRef][ISI][Medline]
Buonocore G, Perrone S, Longini M, Terzuoli L, Bracci R. Total hydroperoxide and advanced oxidation protein products in preterm hypoxic babies. Pediatr Res 2000; 47: 221–4.[ISI][Medline]
Buonocore G, Perrone S, Carlini M, Bagnoli F, Gatti MG, Paffetti P, et al. Non protein bound iron plasma levels in critically ill newborns [abstract]. Biol Neonate 2001; 80: 93–4.
Buonocore G, Perrone S, Longini M, Vezzosi P, Marzocchi B, Paffetti P, et al. Oxidative stress in preterm neonates at birth and on the seventh day of life. Pediatr Res 2002a; 52: 46–9.[CrossRef][ISI][Medline]
Buonocore G, Perrone S. Iron: a potent prooxidant. In: Raiha NC, Rubaltelli FF, editors. Infant formula: closer to the reference. Philadelphia: Lippincott Williams and Wilkins; Nestlè Nutrition workshop series. Pediatric Program; Vol. 47 Suppl. 2002b. pp. 85–96.
Ciccoli L, Rossi V, Signorini C, Leoncini S, Buonocore G, Paffetti P, et al. Iron release in newborn and adult erythrocytes exposed to hypoxia-reoxygenation [abstract]. Biol Neonate 2001; 80: 102.
Dammann O, Leviton A. Maternal intrauterine infection, cytokines and brain damage in the preterm newborn. Pediatr Res 1997; 42: 1–8.[ISI][Medline]
Dorrepaal CA, Berger HM, Benders MJN, van Zoeren-Grobben D, Van de Bor M, Van Bel F. Non protein-bound iron in postasphyxial reperfusion injury of the newborn. Pediatrics 1996; 98: 883–9.
Goldenberg RL, Huddleston JF, Nelson KG. Apgar scores and umbilical arterial pH in preterm newborn infants. Am J Obstet Gynecol 1984; 149: 651–4.[ISI][Medline]
Goplerud JM, Mishra OP, Delivoria-Papadopoulos M. Brain cell membrane dysfunction following acute asphyxia in newborn piglets. Biol Neonate 1992; 61: 33–41.[CrossRef][ISI][Medline]
Greisen G. Tape-recorded EEG and the cerebral function monitor: amplitude-integrated, time-compressed EEG. J Perinat Med 1994; 22: 541–6.[ISI][Medline]
Gutteridge JMC. Ferrous ions detected in cerebrospinal fluid by using bleomycin and DNA damage. Clin Sci 1992a; 82: 315–320[Medline]
Gutteridge JMC. Iron and oxygen radicals in brain. Ann Neurol 1992b; 32 Suppl: S16–21.
Halliwell B. Reactive oxygen species and the central nervous system. J Neurochem 1992; 59: 1609–23. [ISI][Medline]
International Neonatal Network. The CRIB (clinical risk index for babies) score: a tool for assessing initial neonatal risk and comparing performance of neonatal intensive care units. Lancet 1993; 342: 193–8. [CrossRef][ISI][Medline]
Kaplan J, Jordan I, Sturrock A. Regulation of the transferrin-independent iron transport system in cultured cells. J Biol Chem 1991; 266: 2997–3004.
Kime R, Gibson A, Yong W, Hider R, Powers H. Chromatographic method for the determination of non-transferrin-bound iron suitable for use on the plasma and bronchoalveolar lavage fluid of preterm babies. Clin Sci 1996; 91: 633–8.[Medline]
Lindeman JHN, Houdkamp E, Lentjes EGW, Poorthuis BJH, Berger HM. Limited protection against iron-induced lipid peroxidation by cord blood plasma. Free Radic Res Commun 1992; 16: 285–94.[ISI][Medline]
Low JA, Galbraith RS, Muir DW, Killen HL, Pater EA, Karchmar EJ. The relationship between perinatal hypoxia and newborn encephalopathy. Am J Obstet Gynecol 1985; 152: 256–60.[ISI][Medline]
McNamara L, MacPhail AP, Mandishona E, Bloom P, Paterson AC, Rouault TA, et al. Non-transferrin-bound iron and hepatic dysfunction in African dietary iron overload. J Gastroenterol Hepatol 1999; 14: 126–32.[CrossRef][ISI][Medline]
Mishra OP, Delivoria-Papadopoulos M. Cellular mechanisms of hypoxic injury in the developing brain. Brain Res Bull 1999; 48: 233–8.[CrossRef][ISI][Medline]
Palmer C, Menzies SL, Roberts RL, Pavlick G, Connor JR. Changes in iron histochemistry after hypoxic-ischemic brain injury in the neonatal rat. J Neurosci Res 1999; 56: 60–71.[CrossRef][ISI][Medline]
Rothman RJ, Serroni A, Farber JL. Cellular pool of transient ferric iron, chelatable by deferoxamine and distinct from ferritin, that is involved in oxidative cell injury. Mol Pharmacol 1992; 42: 703–10.[Abstract]
Ruth VJ, Raivio KO. Perinatal brain damage: predictive value of metabolic acidosis and the Apgar score. BMJ 1988; 297: 24–7.[ISI][Medline]
Schafer FQ, Buettner GR. Acidic pH amplifies iron-mediated lipid peroxidation in cells. Free Radic Biol Med 2000; 28: 1175–81.[CrossRef][ISI][Medline]
Schafer FQ, Qian SY, Buettner GR. Iron and free radical oxidations in cell membranes. Cell Mol Biol (Noisy-le-grand) 2000; 46: 657–62.[ISI][Medline]
Sorg O, Horn TFW, Yu NC, Gruol DL, Bloom FE. Inhibition of astrocyte glutamate uptake by reactive oxygen species: role of antioxidant enzymes. Mol Med 1997; 3: 431–40.[ISI][Medline]
Waterfall AH, Singh G, Fry JR, Marsden CA. Acute acidosis elevates malonaldehyde in rat brain in vivo. Brain Res 1996; 712: 102–6.[CrossRef][ISI][Medline]
Witko-Sarsat V, Friedlander M, Capeillere-Blandin C, Nguyen-Khoa T, Nguyen AT, Zingraff J, et al. Advanced oxidation protein products as a novel marker of oxidative stress in uremia. Kidney Int 1996; 49: 1304–13.[ISI][Medline]
World Health Organization. Child health development: health of the newborn. Geneva: World Health Organization; 1991.
Ying W, Han S-K, Miller JW, Swanson RA. Acidosis potentiates oxidative neuronal death by multiple mechanisms. J Neurochem 1999; 73: 1549–56.[CrossRef][ISI][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  S. E. Juul, R. J. McPherson, L. A. Bauer, K. J. Ledbetter, C. A. Gleason, and D. E. Mayock A Phase I/II Trial of High-Dose Erythropoietin in Extremely Low Birth Weight Infants: Pharmacokinetics and Safety Pediatrics, August 1, 2008; 122(2): 383 - 391. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||