IMPAIRED ENERGY METABOLISM IN BRAINS OF NEWBORN INFANTS WITH INCREASED CEREBRAL ECHODENSITIES

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IMPAIRED ENERGY METABOLISM IN BRAINS OF NEWBORN INFANTS WITH INCREASED CEREBRAL ECHODENSITIES PATRICIA A. HAMILTON P. L. HOPE E. B. CADY D. T. DELPY E. O. R. REYNOLDS J. S. WYATT

TABLE I-CLINICAL DETAILS OF INFANTS WITH INCREASED ECHODENSITIES

Departments of Paediatrics and Medical Physics and Bioengineering, University College London School of Medicine, Rayne Institute, London WC1 Intracellular energy metabolism was studied by phosphorus magnetic resonance spectroscopy in the brains of 27 preterm and term infants with increased echodensities consistent with hypoxic-ischaemic injury and 18 comparable In the normal infants the infants. normal

Summary

phosphocreatine (PCr)/inorganic orthophosphate (Pi) ratio increased significantly from 0·77±0·24 (95% confidence limits) at a gestational plus postnatal age of 28 weeks to 1·09±0·24 at 42 weeks. 9 of the 15 infants with increased echodensities whose PCr/Pi ratios fell below the normal range died; in all 6 survivors cerebral atrophy developed (cysts in brain tissue or microcephaly). In contrast, all 12 infants with increased echodensities whose PCr/Pi ratios remained within the normal range survived, although cerebral atrophy developed in 3 with ratios towards the lower limit of normal. Introduction

AREAS of increased echodensity on ultrasound scans of the brain in newborn infants are often attributed to hypoxic-ischaemic injury.l6 In some cases these areas reflect severe damage leading to death. In surviving infants they may evolve into cystic peri ventricular. leucomalacia or other forms of cerebral atrophy, or they may disappear, apparently without adverse sequelae .7,11 No non-invasive method has hitherto been available to explore intracellular metabolic events associated with increased echodensities or to distinguish infants with good and poor prognoses. We have previously detected abnormalities consistent with abnormal energy metabolism by phosphorus magnetic resonance spectroscopy in the brains of term infants who had been asphyxiated during delivery.9,10 We have now studied whether evidence of impaired energy metabolism can be found in the brains of preterm and term infants with increased echodensities, and whether measurements of the energy state of cerebral tissue give useful prognostic information. Subjects and Methods Normal

Infants

18 normal infants

(13 male, 5 female) were studied. 15 were born at University College Hospital and 3 were referred to the neonatal unit because of preterm birth. The infants were born at 28-42 (median 32) weeks of gestation, weighing 1115-3860 g (median 1738 g). No infant had any evidence of birth asphyxia or neurological abnormality and ultrasound scans of their brains (Diasonics DS1 scanner with 6 and 7-5 MHz probes) were normal. Magnetic resonance spectroscopy was carried out without sedation when the infants were 1-61 (median 5) days old at a gestational plus postnatal age of 28-42 (median 35) weeks.

In 29, the echodensities were very clearly delineated and confluent with intraventricular haemorrhage; they were therefore thought to be due mainly or entirely to haemorrhage. In 62 of the other 118 infants the increased echodensities were mild and transient, but in the remaining 56 they were pronounced, persisted for at least 48 h in surviving infants, and appeared consistent with hypoxic-ischaemic injury.I-4 Magnetic resonance spectroscopy was carried out on 27 of the latter infants. The echodensities were diffuse and bilateral in 14, born at 27-42 (median 40) weeks’ gestation, 13 of whom had been severely asphyxiated during delivery (median base excess in arterial blood shortly afterwards - 21 mmol/1). In 10 infants, born at 30-41 (median 34) weeks’ gestation, the echodensities were largely confined to the periventricular region, were bilateral in all but 1 infant, and were thought to be due to periventricular leucomalacia. In the other 3 infants, born at 34-35 weeks’ gestation, they were unilateral and consistent with infarction in the distribution of the middle cerebral artery plus, in 1 infant, the posterior cerebral artery. The principal diagnoses of the infants are given in table i. Magnetic resonance spectroscopy was first carried out when the infants were 8 h to 23 days (median 3 days) old, 0-8 (median 1) days after the echodensities were first noted and while these remained clearly visible. Further studies were carried out on 19 of the infants at intervals of a few days. The total number of studies was 64 (median 2, range 1-6, in each infant). Sedation was not given specifically, though 17 infants had been treated with phenobarbitone to prevent convulsions.

Magnetic Resonance Spectroscopy Spectroscopy was carried out with

an Oxford Research TMR 32-200 spectrometer operating at a field Systems strength of 1 -89 Tesla. 10,12 The infant’s head lay on a 5 cm or 7-4 cm diameter magnetic resonance surface coil which could be tuned to the resonance frequencies of both phosphorus-31 (32.5 MHz) and hydrogen-1 (80-3 MHz). The coil supplied the exciting radiofrequency pulse and also detected the magnetic resonance signal returning from the sensitive volume of the magnet, which was centred in the infant’s adjacent temporoparietal cortex. The pulse duration was 80 ps for the 5 cm coil and 100 us for the 7-4 cm coil; the flip angle at the centre of the coil was between 90° and 100°, and the pulse interval was 2-256 s. The 31p spectra were usually produced as the sum of three consecutive 10 min accumulations (768 pulses) bracketed by 1H spectra. The accumulated free induction decays were processed by means of 12 Hz exponential line broadening to improve the signal/noise ratio and an enhancement technique to increase resolution and to remove the broad spectral feature due to phospholipid and bone.13 In infants with unilaterally increased echodensities both hemispheres were studied. The relative concentrations of the phosphorus metabolites were calculated from the integrals of the gated spectral peaks after correction for the effects of saturation.9 Intracellular pH was estimated from the difference in chemical shift between the phosphocreatine (PCr) and inorganic phosphate (Pi)

resonances. 14,15

Infants with Increased Echodensities

Follow-up

33-month period increased parenchymal echodensities were detected by routine ultrasound scanning of the brain in 147 of the 979 infants admitted to the neonatal unit.

was

During

a

Ultrasound scanning of the brains of the surviving infants continued weekly or more frequently until they were discharged home. Scanning was always done at the equivalent

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Fig I-Phosphorus spectra from 2 normal infants A: born at 28 weeks’ gestation, studied 7 days after birth; PCr/Pi = 0 74. B: born at 40 weeks’-gestation and studied aged 1 day; PCr/Pi = 19. of term either in the neonatal unit or in the follow-up clinic, where further scans were carried out as indicated. Infants with increased echodensities were scanned until they were 4--36 (median 12) weeks old. At each visit neurodevelopmental assessments were carried out.16 This study was approved by the University College London and University College Hospital ethics committee.

sensitive index of the energy state of the tissue, but significant changes were found in the other ratios or in intracellular pH. Ultrasound scans in the normal infants remained normal, and none showed any evidence of neurodevelopmental abnormality at ages between 2 and 32 months (median 8 months).

most no

Results Normal

Infants

Whatever the gestational age at birth, the 31P. spectrum (fig 1) always showed peaks at the characteristic chemical shiftsr’ of phosphomonoesters, Pi, phosphodiesters, PCr, and the y, ot, and P phosphorus nucleotide of nuclei magnesium-complexed triphosphates, mainly ATP. The relative concentrations of some of the metabolites seemed to change with age. To define normal values, linear regressions and 95 % confidence limits were therefore calculated for metabolite ratios on gestational plus postnatal age (table TABLE II—CHANGES IN PHOSPHORUS METABOLITE CONCENTRATION RATIOS AND INTRACELLULAR pH WITH GESTATIONAL PLUS POSTNATAL AGE IN 18 NORMAL INFANTS —————————.——————————————————————————.——————————

Fig 2-Relation between PCr/Pi and gestational plus postnatal age in 18 normal infants (A) and data from 27 infants with increased cerebral echodensities (B). Regression line and 95% confidence limits from normal infants are shown. 0 = survived without cerebral atrophy; j= survived with cerebral atrophy; V died. =

Mean values +95% confidence limits from the regressions

are

given.

PME=phosphomonoesters; PDE=phosphodiesters; pH,=intracellular pH. *p<0001.

always calculated from the P resonance (righthand peak, fig 1). There was a highly significant (p < 0-001) increase in PCr/Pi (fig 2A), the

n). Values for ATP

were

Infants with Increased Echodensities Fig 3 shows ultrasound scans and 31p spectra from an infant with increased echodensities consistent with periventricular leucomalacia. The spectrum 3 days after birth was abnormal, with a low PCr/Pi ratio; subsequently, periventricular cysts appeared in brain

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Fig 3-Spectra and parasagittal ultrasound scans from an infant born at 33 weeks’ gestation. A: 3 days after birth; PCr/Pi=0 60; echodensities consistent with peri ventricular leucomalacia. B: 10 days (spectrum) and 27 days (scan) after birth; PCr/Pi 0-85; cysts in brain tissue. =

tissue, although the spectrum had by then become normal. 2B shows minimum values for PCr/Pi in all 27 infants with increased echodensities. 15 were below the 95% confidence limits for normal infants. In the 3 infants with unilateral cerebral artery infarctions PCr/Pi was always lower in the affected hemisphere than in the normal one, though only 1 value from an abnormal hemisphere was below the 95% confidence limits; PCr/Pi was low in both hemispheres of the infant thought to have unilateral periventricular leucomalacia.

Fig

9 of the 15 infants with low PCr/Pi values died, aged 12 h-10 days, predominantly because of the effects of their cerebral pathology. In 1 infant (PCr/Pi 0-02) periventricular and subcortical cysts developed in brain tissue before death aged 10 days. Of the 6 infants examined at necropsy, 2 who died early had severe 1 cerebral softening, had microcysts in the periventricular zones, and 3 loss or shrinkage of cortical

All 6 survivors had cerebral atrophy: cysts developed in 4 infants and microcephaly in the other 2,

neurons.

whose head circumferences were 2-5 cm and 5 cm below the third centile at 26 and 30 months of age. Serial magnetic resonance spectroscopy on 5 of the survivors showed that the spectra always returned to normal as cerebral atrophy became established (fig 3B). In contrast, all 12 infants with minimum PCr/Pi ratios within the normal range survived. In 3 evidence of cerebral atrophy developed; 2 had cysts in brain tissue and 1 microcephaly, with a head circumference 55 cm below the third centile aged 18 months. The difference in adverse outcome (death or cerebral atrophy) between infants with normal and low PCr/Pi ratios was highly Fisher’s exact test). All infants significant (p<0-001, with cerebral atrophy were neurodevelopmentally abnormal at follow-up, aged 3-30 (median 20) months, whereas only 3 of the 9 infants without atrophy were thought to be abnormal aged 16-21 months (1 of the 3 had kernicterus).

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Fig 4-Spectra from phosphorus ratios.

2 infants with diffuse increased echodensities and reduced

ATP/total

A: infant born at 38 weeks’ gestation studied 2 days after birth. She had been severely asphyxiated during delivery: ATP/total P 007; Pcr/Pi 029. Died. B: infant bom

at

episode aged 5 days:

27 weeks’ gestation studied at 8 days. He had suffered ATP and Per were virtually absent. Died.

Abnormalities of other metabolite ratios and intracellular pH were less obvious. However, ATP/total phosphorus was below the 95% confidence limits in 7 infants (fig 4) whose PCr/Pi ratios were all extremely low (median 0-23, range 002-035): 5 died and the other 2 survived with severe cerebral atrophy. Changes in PCr/ATP and Pi/ATP were difficult to interpret, because of the reductions in ATP/total phosphorus; PCr/ATP was below the 95% confidence limits in 2 infants and Pi/ATP above them in 13. Intracellular pH was very low ( < 6 7) in 2 terminally ill infants who died soon after the observations were made. Otherwise, it tended to be raised; values from 19 of the 27 infants were above the regression line for normal infants and in 6 were above the 95 % confidence limits.

Discussion In the normal infants the

PCr/Pi

ratio increased

greatly with maturation of the brain, apparently because of both a rise in PCr and a fall in Pi (since PCr/ATP tended to rise and Pi/ATP to fall). It is likely that the phosphorylation potential or energy state of the brain tissue increased with age, as occurs in the newborn rat.18 No significant changes were detected in the other metabolite ratios or in intracellular pH. The trend towards a fall in phosphomonoesters/ATP was, however, expected, since this ratio falls with age in the newborn rat and guineapigl8 and is low in adult brain tissue in man and other species.l8n9 The main constituent of the phosphomonoester peak is thought to be phosphoethanolamine, which is a major precursor of membrane phospholipid and myelin and is often present in high concentration in rapidly growing tissue.20,21 Most of the infants who had increased cerebral echodensities were moderately or severely ill, requiring mechanical ventilation and other forms of intensive care.

an

extremely

severe

asphyxial

3 were, however,

symptom-free and their echodensities (indicating periventricular leucomalacia in 2 infants and unilateral middle cerebral artery infarction in the other) were detected as a result of routine ultrasound scanning. Increased echodensities

are

known

to

be associated with

hypoxic-ischaemic injury to the brain,1-4 so it was not surprising that many of the infants studied had been asphyxiated during delivery or were diagnosed as having hyaline membrane disease (table i). The number who were small for gestational age or had polycythaemia was striking. Both these disorders carry an increased risk of neurodevelopmental disorders;22,23 a prospective investigation of the energy state of the brain in similar infants seems warranted. Our necropsy studies of infants who die with increased echodensities show various histological features, ranging from normality to frank necrosis of brain tissue (S. J. Gould et al, unpublished); follow-up of surviving infants demonstrates that increased echodensities do not necessarily signify a poor neurodevelopmental outcome.7,8 The major purpose of our study was, therefore, to find out whether the presence of increased echodensities was associated with evidence of impaired energy metabolism and, if so, whether measurements of the energy state of cerebral tissue distinguished infants with good and bad prognoses. The PCr/Pi ratio was often low in infants with increased echodensities (fig 2B). A reduction in ATP was detected less frequently and only when was greatly reduced (fig 4). These findings are consistent with ATP levels being maintained by the creatine kinase

PCr/Pi

reaction

in the face of impaired phosphorylation, as previously reported asphyxiated infants.lo

oxidative in birth-

of the PCr/Pi ratio was highly of outcome, since all 15 infants with values predictive below the normal range either died or showed clear

Measurement

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evidence of loss of brain tissue at follow-up (fig 2B). By comparison, all 12 infants whose PCr/Pi ratios remained within the normal range survived, although cerebral atrophy developed in 3 with ratios towards the lower limit of normal. A description of the longer-term neurodevelopmental outcome of the infants will be given when they are old enough for firm conclusions to be reached. We thank Prof D. R. Wilkie, FRS, for advice and encouragement; Dr S. J. Gould for necropsy reports; Mr R. Aldridge for technical assistance; the staff of the neonatal unit, the department of medical physics and’ bioengineering, and Oxford Research Systems for their help; and Mrs G. Harris and Mrs J. Baldwyn for preparing the typescript. This work was supported by grants from the Medical Research Council, Action Research for the Crippled Child, the Muscular Dystrophy Group, the Wellcome Trust, and the special trustees of University College Hospital.

Correspondence should be addressed to E. 0. R. R., Department of’ Paediatrics, University College London School of Medicine, Rayne Institute, University Street, London WC1E 6JJ. REFERENCES 1. Nwaesei CG, Pape KE, Martin DJ, Becker LE, Fitz CR. Periventricular infarction diagnosed by ultrasound: a postmortem correlation. J Pediatr 1984; 105: 106-10. 2. Fawer C-L, Calame A, Perentes E, Anderegg A. Periventricular leukomalacia a correlation study between real-time ultrasound and autopsy findings. Neuroradiology 1985; 27: 292-30. 3. Rushton DI, Preston PR, Durbin GM. Structure and evolution of echo dense lesions in the neonatal brain. Arch Dis Child 1985; 60: 798-808. 4. Levene MI, Williams JL, Fawer C-L. Ultrasound of the infant brain. Clinics in developmental medicine, no 92. Oxford: Blackwell, for Spastics International Publications, 1985: 76-92. 5. de Vries LS, Dubowitz LMS, Dubowitz V, et al. Predictive value of cranial ultrasound in the newborn baby: a reappraisal. Lancet 1985; ii: 137-40. 6. Sinha SK, Davies JM, Sims DG, Chiswick ML. Relation between periventricular haemorrhage and ischaemic brain lesions diagnosed by ultrasound in very preterm infants. Lancet 1985; ii: 1154-56. 7. McMenamin JB, Shackelford GD, Volpe JJ. Outcome of neonatal intraventricular hemorrhage with periventricular echodense lesions. Ann Neurol 1984; 15: 285-90. 8. Stewart AL, Hope PL, Hamilton PA, Baudin J, Wyatt JS, Reynolds EOR. Increased periventricular echodensities in very preterm infants and prediction of early neurodevelopmental outcome. Pediatr Res 1985; 19: 1084 (abstr). 9. Cady EB, Costello AM de L, Dawson MJ, et al. Non-invasive investigation of cerebral metabolism in newborn infants by phosphorus nuclear magnetic resonance spectroscopy. Lancet 1983; i: 1059-62. 10. Hope PL, Costello AM de L, Cady EB, et al. Cerebral energy metabolism studied with phosphorus NMR spectroscopy in normal and birth-asphyxiated infants. Lancet 1984; ii: 366-70. 11. Lubchenco LO, Hansman C, Dressler M, Boyd E. Intrauterine growth as estimated from live born birth weight data at 24 to 42 weeks of gestation. Pediatrics 1963; 32: 793-800. 12. Hope PL, Reynolds EOR. Investigation of cerebral energy metabolism in newborn infants by phosphorus nuclear magnetic resonance spectroscopy. Clin Perinatol 1985; 12: 261-75. 13. Gordon RE, Hanley PE, Shaw D. Topical magnetic resonance. Prog Nucl Mag Res Spectr 1982; 15: 1-47. 14. Cady EB, Wilkie DR. Estimation of cerebral intracellular pH by 31P NMR spectroscopy. In: Rolfe P, ed. Fetal and neonatal physiological measurements. London: Butterworths (in press). 15. Petroff OAC, Prichard JW, Behar KL, Alger JR, den Hollander JA, Shulman RG Cerebral intracellular pH by 31P nuclear magnetic resonance spectroscopy. Neurology 1985; 35: 781-88. 16. Stewart AL, Thorburn RJ, Hope PL, Goldsmith M, Lipscomb AP, Reynolds EOR. Ultrasound appearance of the brain in very preterm infants and neurodevelopmental outcome at 18 months of age. Arch Dis Child 1983; 58: 598-604. 17. Gadian DG. Nuclear magnetic resonance and its applications to living systems. Oxford Clarendon Press, 1982. 18 Tofts P, Wray S. Changes in brain phosphorus metabolites during the post-natal development of the rat. J Physiol 1985; 359: 417-29. 19. Bottomley PA, Hart HR, Edelstein WA, et al. Anatomy and metabolism of the normal human brain studied by magnetic resonance at 1·5 tesla. Radiology 1984; 150: 441-46. 20 Gyulai L, Bolinger L, Leigh JS, Barlow C, Chance B. Phosphorylethanolaminethe major constituent of the phosphomonoester peak observed by 31P-NMR in developing dog brain. FEBS Lett 1984, 178: 137-42. 21. Maris JM, Evans AE, McLaughlin AC, et al. 31P Nuclear magnetic resonance spectroscopic investigation of human neuroblastoma in situ. N Engl J Med

1985, 312: 1500-05. panorama of cerebral palsy in Sweden 1954-1970. III. The importance of fetal deprivation of supply. Acta Paediatr Scand 1976; 65: 403-08. 23 Black VD, Lubchenco LO, Koops BL, Poland RL, Powell DP. Neonatal hyperviscosity: randomized study of effect of partial plasma exchange transfusion on long-term outcome Pediatrics 1985; 75: 1048-53.

22.

Hagberg B, Hagberg G, Olow I. The changing

EFFECT OF AGE AT ORCHIDOPEXY ON RISK OF TESTICULAR CANCER M. C. PIKE M.

CLAIR CHILVERS

J. PECKHAM

Imperial Cancer Research Fund Cancer Epidemiology Unit, Radcliffe Infirmary, Oxford; and Institute of Cancer Research, Sutton, Surrey, UK

(9·5%) of 724 patients with testicular first seen at the Royal Marsden over the 10-year period 1975 to 1984 had a Hospital of history cryptorchidism. 11 patients had an Summary

69

cancer

uncorrected undescended testis when their testicular cancer was diagnosed, and 58 had had an orchidopexy or an orchidectomy for undescended testis. There are no general population data with which to compare the 15·9% (11/69) of testicular cancer patients with uncorrected cryptorchidism. However, the distribution of age at orchidopexy (or orchidectomy) of the 58 patients treated for undescended testis was almost identical to that expected on the basis of national rates. The age at treatment of undescended testis appears to have no effect on the risk of testicular cancer.

Introduction

OVER the past 20 years there has been a steady trend towards early operation for undescended testis; Hospital In-patient Enquiry data for England and Wales show that in 1981 the orchidopexy rate at ages 0-4 years was about 10 times the rate in 1962.’Many orchidopexies are done before the age of 1 year. The reason for early operation is the belief that fertility is more likely to be preserved2 and that the risk of cancer of the testis may be reduced.3 There is, however, little evidence to support the former4 and no data with which to investigate the latter supposition.34 The Royal Marsden Hospital (RMH) has had a particular interest in treating patients with testicular cancer over the past 15 years and has kept detailed records of these patients. With the use of these records we investigated whether orchidopexy at an early age reduces the risk of testicular cancer.

Patients and Methods The records of all patients who attended the RMH for the first time with a diagnosis of testicular cancer between Jan 1, 1975, and Dec 31, 1984, were reviewed. Almost all of these patients had their orchidectomy elsewhere and were referred soon afterwards for detailed evaluation and, if necessary, further treatment. Patients with bilateral tumours were included only if they attended the RMH with their first tumour within this period. Only patients with germ-cell tumours were eligible; patients with lymphomas and sarcomas were excluded, as were patients whose tumour was not positively identified as being a testicular primary. The case-notes of the patients were abstracted by one of two research nurses onto a standard form. The information abstracted included details of the histology of the tumour, history of undescended testis (UDT) and its treatment, and other testicular problems such as late descent and retractility. Tumours were classified as seminomas (pure seminoma with no other elements present) or teratomas (all others). All living patients identified as having a definite history or a possible