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Pediatrics International (2014) 56, 709–713
doi: 10.1111/ped.12332
Original Article
Oxidative stress early in infancy and neurodevelopmental outcome in very low-birthweight infants Hiromichi Shoji, Naho Ikeda, Mariko Hosozawa, Natsuki Ohkawa, Nobuaki Matsunaga, Hiroki Suganuma, Ken Hisata, Kyoko Tanaka and Toshiaki Shimizu Department of Pediatrics, Juntendo University Faculty of Medicine, Tokyo, Japan Abstract
Background: Reactive oxygen species may be involved in serious diseases in premature infants. The objective of this study was to assess the relationship between neurodevelopmental outcome and oxidative stress marker level in the urine of very low-birthweight (VLBW) infants. Methods: Spot urine samples were collected from 35 VLBW infants. Urinary excretion of 8-hydroxy-2″deoxyguanosine (8-OHdG), a marker of oxidative DNA damage, and 8-iso-prostaglandin F2α (8-isoPGF), a marker of lipid peroxidation, was measured at 1, 2, 4, and 6 weeks of age. Neurodevelopmental outcome at 18 months’ corrected age was assessed using the Bayley Scales of Infant Development (BSID)-II. Results: Significant correlations were found between urinary 8-OHdG at 2 and 4 weeks and the Mental Development Index of the BSID-II. No significant correlation was found between urinary 8-isoPGF and indices of the BSID-II. Conclusions: In VLBW infants, urinary 8-OHdG level correlated with mental development rather than psychomotor development at 18 months’ corrected age; urinary 8-OHdG might be a predictive marker of neurodevelopmental outcome in VLBW infants.
Key words 8-hydroxy-2″-deoxyguanosine, 8-iso-prostaglandin F2α, Bayley Scales of Infant Development, oxidative stress, very low birthweight.
Advances in perinatal and neonatal intensive care have led to a dramatic increase in the survival of preterm infants, but this improvement has resulted in a greater risk for long-term adverse neurodevelopmental problems.1,2 The morbidities associated with prematurity have been linked to oxidative stress and free radical-mediated cell and tissue injury.3,4 Premature infants are less prepared to cope with the oxygen-rich environment of extrauterine life because their antioxidant mechanisms are underdeveloped when compared with infants born at term.4,5 Many mechanisms lead to free radical overproduction in the perinatal period, including ischemia– reperfusion, the arachidonic acid cascade, free iron, the nitric oxide cascade, phagocyte activation, hypoxia, and hyperoxia.3,6 Indeed, reactive oxygen species (ROS) may be involved in serious diseases in premature infants, including chronic lung disease7,8 and retinopathy of prematurity.8 Prediction of long-term neurodevelopmental outcomes in very low-birthweight (VLBW) infants remains challenging because access to biological samples is difficult in the neonatal period. 8-Hydroxy-2″-deoxyguanosine (8-OHdG) is produced by oxidation of the nucleoside, deoxyguanosine, and is excreted directly Correspondence: Hiromichi Shoji, MD, Department of Pediatrics, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421, Japan. Email: [email protected] Received 9 June 2013; revised 22 January 2014; accepted 4 February 2014.
© 2014 Japan Pediatric Society
into the urine. Urinary 8-OHdG excretion has been identified as a sensitive marker for oxidative DNA damage.9 Isoprostanes, among which 8-iso-prostaglandin F2α (8-isoPGF) is the major compound, are prostaglandin-like compounds formed in vivo from free radical-initiated peroxidation of arachidonic acid without involvement of cyclo-oxygenase, and are excreted in urine.10 8-isoPGF is also an important oxidative stress marker for assessment of oxidative stress directed against phospholipids.11 Recent studies have identified a relationship between prematurity and urinary 8-OHdG or 8-iso PGF concentration in low-birthweight infants.12,13 Limited information is available describing the influence of oxidative DNA damage and lipid peroxidation on morbidity in VLBW infants. Therefore, this study investigated the relationship between the levels of 8-OHdG and 8-isoPGF in the urine of VLBW infants and neurodevelopmental outcome at 18 months’ corrected age.
Methods Subjects
A pilot study was conducted in a six-bed neonatal intensive care unit (NICU) of Juntendo University Hospital, Tokyo, Japan. A total of 35 preterm infants (19 girls, 16 boys) with birth weight 100 mL/kg/day (days) Period of parenteral nutrition (days) Lipid emulsion Antenatal steroids IUGR Apgar score at 5 min ≤7 Intracranial hemorrhage (III, IV) Necrotizing enterocolitis Photocoagulation for ROP Period of ventilation (days) Oxygen supply For 28 days At CA 36 weeks 18 months’ CA Bodyweight (g) Body length (cm) Head circumference (cm)
Mean ± SD or n (%) 19/16 28.3 ± 2.7 969 ± 314 35.0 ± 3.9 25.3 ± 2.7 11.9 ± 5.3 19.2 ± 11.7 25 (71) 20 (57) 14 (40) 4 (11) 2 (6) 0 (0.0) 8 (23) 37 ± 40 25 (71) 15 (43) 9197 ± 772 78.8 ± 2.6 46.3 ± 1.2
CA, corrected age; IUGR, intrauterine growth restriction; ROP, retinopathy of prematurity; VLBW, very low birthweight.
Oxidative damage in VLBWI (a)
140
711
(b)
120
MDI
100 80 60 40 20 0 0
40
80
120
0
40
80
120
160
8-OHdG (ng/mg Cr)
Fig. 1 Urinary 8-hydroxy-2″-deoxyguanosine (8-OHdG) at (a) 2 or (b) 4 weeks and Mental Development Index (MDI) of the Bayley Scales of Infant Development-II.
MDI and 8-OHdG at 2 and 4 weeks of age. Furthermore, mean 8-OHdG in the normal MDI group at 2 weeks of age was significantly lower than that in the subnormal MDI group, but there was no correlation between 8-isoPGF and the BSID-II scales. Survival of VLBW infants has increased considerably in recent decades. Although developmental outcome varies with the age of the child, long-term follow-up studies show that preterm infants are at a greater risk for developmental problems, including poor cognitive function, and are at greater risk for behavioral problems, such as attention-deficit–hyperactivity disorder and learning disorders.18 In the brain, oxidative stress can trigger apoptosis,19 and excessive neural apoptosis may result in longterm brain dysfunction. Fukuda et al. reported that the levels of urinary and cerebrospinal fluid 8-OHdG in children with hypoxic–ischemic encephalopathy were significantly higher than those of control subjects.20 No detailed neurodevelopmental evaluations, however, have been conducted in survivors at 18 months’ corrected age. Isoprostanes are non-invasive biomarkers of lipid peroxidation, which occurs when oxygen free radicals disrupt lipid cell membranes of various organs, such as the brain, making their measurement particularly suitable for studies in infants.21 Cord blood 8-isoPGF is elevated in oxidative stress-related diseases in premature infants.22 Back et al. investigated brains from human autopsies and found that isoprostane level was significantly increased when periventricular white matter injury had
occurred in preterm infants.23 We think it is difficult, however, to predict neurodevelopmental outcome by lipid peroxidation measurement in preterm infants during the early postnatal period because of low fatty acid body stores and consumed lipids from milk. The mother’s breast milk is considered the ideal food for infants, and its importance in neurodevelopmental processes is clearly recognized.24 In addition, breast milk from mothers of term and preterm infants has important antioxidant properties when compared with formula milk.25 We previously compared urinary 8-OHdG level in 29 preterm infants, and found that 8-OHdG in breast-fed infants at 14 and 28 days of age was significantly lower than in formula-fed infants.26 No significant differences were observed, however, in the proportion of breastfed infants between the normal and subnormal MDI groups. The reasons for the positive relationship between urinary 8-OHdG and MDI of the BSID-II at 2 and 4 weeks but not at 1 and 6 weeks are unknown. We propose, however, that neurodevelopmental outcome can be improved with appropriate management, such as breast-feeding and avoidance of high oxygen saturation, to decrease exposure of VLBW infants to oxidative stress. The present study had limitations. The sample size was relatively small. It is unclear to what extent the MDI at 18 months of age is predictive of later cognitive function among VLBW infants, although MDI is the most widely used measure of cognitive function in preterm infants. Hack et al. reported that the
Table 2 8-OHdG, 8-isoPGF and indices of the BSID-II 1 week MDI PDI
r r
8-OHdG 0.05 −0.02
8-isoPGF 0.13 0.16
2 weeks 8-OHdG −0.39* −0.19
8-isoPGF −0.18 0.06
4 weeks 8-OHdG −0.44* −0.36
8-isoPGF −0.17 −0.38
6 weeks 8-OHdG −0.06 0.12
8-isoPGF −0.02 0.13
*P < 0.05. r, Spearman’s rank correlation coefficient. 8-OHdG, 8-hydroxy-2″-deoxyguanosine; 8-isoPGF, 8-iso-prostaglandin F2α; BSID, Bayley Scales of Infant Development; MDI, Mental Development Index; PDI, Psychomotor Development Index.
© 2014 Japan Pediatric Society
712
H Shoji et al.
Table 3 Subject characteristics vs MDI
Gestational age Bodyweight
8-OHdG (ng/mg Cr)
8-isoPGF (ng/mg Cr)
AST (IU/L) ALT (IU/L) Hb (g/dL) Cr (mg/dL) Human milk/enteral nutrition (%)
0 days 2 weeks 4 weeks 6 weeks CA18 months 1 weeks 2 weeks 4 weeks 6 weeks 1 weeks 2 weeks 4 weeks 6 weeks 2 weeks 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks
MDI
Pediatrics International (2014) 56, 709–713
doi: 10.1111/ped.12332
Original Article
Oxidative stress early in infancy and neurodevelopmental outcome in very low-birthweight infants Hiromichi Shoji, Naho Ikeda, Mariko Hosozawa, Natsuki Ohkawa, Nobuaki Matsunaga, Hiroki Suganuma, Ken Hisata, Kyoko Tanaka and Toshiaki Shimizu Department of Pediatrics, Juntendo University Faculty of Medicine, Tokyo, Japan Abstract
Background: Reactive oxygen species may be involved in serious diseases in premature infants. The objective of this study was to assess the relationship between neurodevelopmental outcome and oxidative stress marker level in the urine of very low-birthweight (VLBW) infants. Methods: Spot urine samples were collected from 35 VLBW infants. Urinary excretion of 8-hydroxy-2″deoxyguanosine (8-OHdG), a marker of oxidative DNA damage, and 8-iso-prostaglandin F2α (8-isoPGF), a marker of lipid peroxidation, was measured at 1, 2, 4, and 6 weeks of age. Neurodevelopmental outcome at 18 months’ corrected age was assessed using the Bayley Scales of Infant Development (BSID)-II. Results: Significant correlations were found between urinary 8-OHdG at 2 and 4 weeks and the Mental Development Index of the BSID-II. No significant correlation was found between urinary 8-isoPGF and indices of the BSID-II. Conclusions: In VLBW infants, urinary 8-OHdG level correlated with mental development rather than psychomotor development at 18 months’ corrected age; urinary 8-OHdG might be a predictive marker of neurodevelopmental outcome in VLBW infants.
Key words 8-hydroxy-2″-deoxyguanosine, 8-iso-prostaglandin F2α, Bayley Scales of Infant Development, oxidative stress, very low birthweight.
Advances in perinatal and neonatal intensive care have led to a dramatic increase in the survival of preterm infants, but this improvement has resulted in a greater risk for long-term adverse neurodevelopmental problems.1,2 The morbidities associated with prematurity have been linked to oxidative stress and free radical-mediated cell and tissue injury.3,4 Premature infants are less prepared to cope with the oxygen-rich environment of extrauterine life because their antioxidant mechanisms are underdeveloped when compared with infants born at term.4,5 Many mechanisms lead to free radical overproduction in the perinatal period, including ischemia– reperfusion, the arachidonic acid cascade, free iron, the nitric oxide cascade, phagocyte activation, hypoxia, and hyperoxia.3,6 Indeed, reactive oxygen species (ROS) may be involved in serious diseases in premature infants, including chronic lung disease7,8 and retinopathy of prematurity.8 Prediction of long-term neurodevelopmental outcomes in very low-birthweight (VLBW) infants remains challenging because access to biological samples is difficult in the neonatal period. 8-Hydroxy-2″-deoxyguanosine (8-OHdG) is produced by oxidation of the nucleoside, deoxyguanosine, and is excreted directly Correspondence: Hiromichi Shoji, MD, Department of Pediatrics, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421, Japan. Email: [email protected] Received 9 June 2013; revised 22 January 2014; accepted 4 February 2014.
© 2014 Japan Pediatric Society
into the urine. Urinary 8-OHdG excretion has been identified as a sensitive marker for oxidative DNA damage.9 Isoprostanes, among which 8-iso-prostaglandin F2α (8-isoPGF) is the major compound, are prostaglandin-like compounds formed in vivo from free radical-initiated peroxidation of arachidonic acid without involvement of cyclo-oxygenase, and are excreted in urine.10 8-isoPGF is also an important oxidative stress marker for assessment of oxidative stress directed against phospholipids.11 Recent studies have identified a relationship between prematurity and urinary 8-OHdG or 8-iso PGF concentration in low-birthweight infants.12,13 Limited information is available describing the influence of oxidative DNA damage and lipid peroxidation on morbidity in VLBW infants. Therefore, this study investigated the relationship between the levels of 8-OHdG and 8-isoPGF in the urine of VLBW infants and neurodevelopmental outcome at 18 months’ corrected age.
Methods Subjects
A pilot study was conducted in a six-bed neonatal intensive care unit (NICU) of Juntendo University Hospital, Tokyo, Japan. A total of 35 preterm infants (19 girls, 16 boys) with birth weight 100 mL/kg/day (days) Period of parenteral nutrition (days) Lipid emulsion Antenatal steroids IUGR Apgar score at 5 min ≤7 Intracranial hemorrhage (III, IV) Necrotizing enterocolitis Photocoagulation for ROP Period of ventilation (days) Oxygen supply For 28 days At CA 36 weeks 18 months’ CA Bodyweight (g) Body length (cm) Head circumference (cm)
Mean ± SD or n (%) 19/16 28.3 ± 2.7 969 ± 314 35.0 ± 3.9 25.3 ± 2.7 11.9 ± 5.3 19.2 ± 11.7 25 (71) 20 (57) 14 (40) 4 (11) 2 (6) 0 (0.0) 8 (23) 37 ± 40 25 (71) 15 (43) 9197 ± 772 78.8 ± 2.6 46.3 ± 1.2
CA, corrected age; IUGR, intrauterine growth restriction; ROP, retinopathy of prematurity; VLBW, very low birthweight.
Oxidative damage in VLBWI (a)
140
711
(b)
120
MDI
100 80 60 40 20 0 0
40
80
120
0
40
80
120
160
8-OHdG (ng/mg Cr)
Fig. 1 Urinary 8-hydroxy-2″-deoxyguanosine (8-OHdG) at (a) 2 or (b) 4 weeks and Mental Development Index (MDI) of the Bayley Scales of Infant Development-II.
MDI and 8-OHdG at 2 and 4 weeks of age. Furthermore, mean 8-OHdG in the normal MDI group at 2 weeks of age was significantly lower than that in the subnormal MDI group, but there was no correlation between 8-isoPGF and the BSID-II scales. Survival of VLBW infants has increased considerably in recent decades. Although developmental outcome varies with the age of the child, long-term follow-up studies show that preterm infants are at a greater risk for developmental problems, including poor cognitive function, and are at greater risk for behavioral problems, such as attention-deficit–hyperactivity disorder and learning disorders.18 In the brain, oxidative stress can trigger apoptosis,19 and excessive neural apoptosis may result in longterm brain dysfunction. Fukuda et al. reported that the levels of urinary and cerebrospinal fluid 8-OHdG in children with hypoxic–ischemic encephalopathy were significantly higher than those of control subjects.20 No detailed neurodevelopmental evaluations, however, have been conducted in survivors at 18 months’ corrected age. Isoprostanes are non-invasive biomarkers of lipid peroxidation, which occurs when oxygen free radicals disrupt lipid cell membranes of various organs, such as the brain, making their measurement particularly suitable for studies in infants.21 Cord blood 8-isoPGF is elevated in oxidative stress-related diseases in premature infants.22 Back et al. investigated brains from human autopsies and found that isoprostane level was significantly increased when periventricular white matter injury had
occurred in preterm infants.23 We think it is difficult, however, to predict neurodevelopmental outcome by lipid peroxidation measurement in preterm infants during the early postnatal period because of low fatty acid body stores and consumed lipids from milk. The mother’s breast milk is considered the ideal food for infants, and its importance in neurodevelopmental processes is clearly recognized.24 In addition, breast milk from mothers of term and preterm infants has important antioxidant properties when compared with formula milk.25 We previously compared urinary 8-OHdG level in 29 preterm infants, and found that 8-OHdG in breast-fed infants at 14 and 28 days of age was significantly lower than in formula-fed infants.26 No significant differences were observed, however, in the proportion of breastfed infants between the normal and subnormal MDI groups. The reasons for the positive relationship between urinary 8-OHdG and MDI of the BSID-II at 2 and 4 weeks but not at 1 and 6 weeks are unknown. We propose, however, that neurodevelopmental outcome can be improved with appropriate management, such as breast-feeding and avoidance of high oxygen saturation, to decrease exposure of VLBW infants to oxidative stress. The present study had limitations. The sample size was relatively small. It is unclear to what extent the MDI at 18 months of age is predictive of later cognitive function among VLBW infants, although MDI is the most widely used measure of cognitive function in preterm infants. Hack et al. reported that the
Table 2 8-OHdG, 8-isoPGF and indices of the BSID-II 1 week MDI PDI
r r
8-OHdG 0.05 −0.02
8-isoPGF 0.13 0.16
2 weeks 8-OHdG −0.39* −0.19
8-isoPGF −0.18 0.06
4 weeks 8-OHdG −0.44* −0.36
8-isoPGF −0.17 −0.38
6 weeks 8-OHdG −0.06 0.12
8-isoPGF −0.02 0.13
*P < 0.05. r, Spearman’s rank correlation coefficient. 8-OHdG, 8-hydroxy-2″-deoxyguanosine; 8-isoPGF, 8-iso-prostaglandin F2α; BSID, Bayley Scales of Infant Development; MDI, Mental Development Index; PDI, Psychomotor Development Index.
© 2014 Japan Pediatric Society
712
H Shoji et al.
Table 3 Subject characteristics vs MDI
Gestational age Bodyweight
8-OHdG (ng/mg Cr)
8-isoPGF (ng/mg Cr)
AST (IU/L) ALT (IU/L) Hb (g/dL) Cr (mg/dL) Human milk/enteral nutrition (%)
0 days 2 weeks 4 weeks 6 weeks CA18 months 1 weeks 2 weeks 4 weeks 6 weeks 1 weeks 2 weeks 4 weeks 6 weeks 2 weeks 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks
MDI
