Abnormal Cerebral Structure Is Present at Term in Premature Infants
Terrie E. Inder, MD*‡§; Simon K. Warfield, PhD㛳; Hong Wang‡; Petra S. Hu¨ppi, MD§¶; and Joseph J. Volpe, MD§
ABSTRACT. Background. Long-term studies of the outcome of very prematurely born infants have clearly documented that the majority of such infants have sig-nificant motor, cognitive, and behavioral deficits. How-ever, there is a limited understanding of the nature of the cerebral abnormality underlying these adverse neuro-logic outcomes.
Aim. The overall aim of this study was to define quantitatively the alterations in cerebral tissue volumes at term equivalent in a large longitudinal cohort study of very low birth weight premature infants in comparison to term-born infants by using advanced volumetric 3-di-mensional magnetic resonance imaging (MRI) tech-niques. We also aimed to define any relationship of such perinatal lesions as white matter (WM) injury or other potentially adverse factors to the quantitative structural alterations. Additionally, we wished to identify the rela-tionship of the structural alterations to short-term neuro-developmental outcome.
Methods. From November 1998 to December 2000, 119 consecutive premature infants admitted to the neonatal intensive care units at Christchurch Women’s Hospital (Christchurch, New Zealand) and the Royal Women’s Hospital (Melbourne, Australia) were recruited (88% of eligible) after informed parental consent to undergo an MRI scan at term equivalent. Twenty-one term-born in-fants across both sites were recruited also. Postacquisi-tion advanced 3-dimensional tissue segmentaPostacquisi-tion with 3-dimensional reconstruction was undertaken to estimate volumes of cerebral tissues: gray matter (GM; cortical and deep nuclear structures), WM (myelinated and un-myelinated), and cerebrospinal fluid (CSF).
Results. In comparison to the term-born infants, the premature infants at term demonstrated prominent re-ductions in cerebral cortical GM volume (premature in-fants [meanⴞSD]: 178ⴞ41 mL; term infants: 227ⴞ26 mL) and in deep nuclear GM volume (premature infants: 10.8 ⴞ 4.1 mL; term infants: 13.8ⴞ 5.2 mL) and an in-crease in CSF volume (premature infants: 45.6ⴞ22.1 mL; term infants: 28.9 ⴞ 16 mL). The major predictors of altered cerebral volumes were gestational age at birth and the presence of cerebral WM injury. Infants with
significantly reduced cortical GM and deep nuclear GM volumes and increased CSF volume volumes exhibited moderate to severe neurodevelopmental disability at 1 year of age.
Conclusions. This MRI study of prematurely born in-fants further defines the nature of quantitative cerebral structural abnormalities present as early as term equiva-lent. The abnormalities particularly involve cerebral neu-ronal regions including both cortex and deep nuclear structures. The pattern of cerebral alterations is related most significantly to the degree of immaturity at birth and to concomitant WM injury. The alterations are fol-lowed by abnormal short-term neurodevelopmental outcome.Pediatrics2005;115:286–294;magnetic resonance imaging, prematurity, neurodevelopmental outcome.
ABBREVIATIONS. VLBW, very low birth weight; WM, white matter; GM, gray matter; SPGR, spoiled gradient recalled; MR, magnetic resonance; DE, spin-echo sequence; CSF, cerebrospinal fluid; ICV, intracranial volume; IVH, intraventricular hemorrhage; IUGR, intrauterine growth restriction.
T
hroughout the last decade there has been an increasing prevalence of surviving very low birth weight (VLBW) premature infants.1Theimprovements in survival have been greatest in the most immature infants but have been accompanied by an increasing awareness of subsequent neurode-velopmental deficits. Approximately 10% of VLBW infants (⬃4000 yearly in the United States) develop cerebral palsy, and fully 50% (⬃20 000 yearly) have impaired academic achievement and/or behavioral disorders requiring additional educational resourc-es.2–6 The nature of the cerebral abnormalities that
underlie these common and serious developmental disabilities is not entirely understood. To improve outcome for these infants, it is crucial to understand both the nature of the cerebral abnormalities and any modifiable perinatal factors involved in pathogene-sis.
Studies based on neuropathological analysis and conventional brain imaging of premature infants have provided evidence primarily for injury to cere-bral white matter7–9(WM). The principal lesions
in-clude periventricular hemorrhagic infarction and periventricular leukomalacia. The latter of these 2 forms of WM injury seems to be more common and more important in the genesis of the subsequent neurologic deficits. Periventricular leukomalacia at its most severe results in focal necroses with sub-sequent cyst formation, readily detected by cranial ultrasonography.7 The clinical importance of the
finding of cystic WM injury by neonatal cranial ul-From the *Murdoch Children’s Research Institute, Royal Women’s and
Royal Children’s Hospitals, University of Melbourne, Melbourne, Australia; ‡Howard Florey Institute, University of Melbourne, Melbourne, Australia; 㛳Department of Radiology, Brigham and Women’s Hospital, Harvard Med-ical School, Boston, Massachusetts; ¶Department of Pediatrics, University of Geneva, Geneva, Switzerland; and §Department of Neurology, Chil-dren’s Hospital, Harvard Medical School, Boston, Massachusetts. Accepted for publication Jul 9, 2004.
doi:10.1542/peds.2004-0326 No conflict of interest declared.
Address correspondence to Terrie E. Inder, MD, Department of Neurology, Royal Children’s Hospital, 2nd Floor, Flemington Rd, Parkville, Victoria 3052, Australia. E-mail: terrie.inder@rch.org.au
trasonography is illustrated by the frequent demon-stration of a greatly increased risk of subsequent neurologic deficits observable on follow-up.10
How-ever, recent studies by conventional magnetic reso-nance imaging (MRI) have shown that noncystic WM injury, manifested by signal abnormalities in cerebral WM, often accompanied by ventricular dilation, is much more common than cystic WM injury.8,11–13In
our preliminary study of this cohort, we documented in 94 premature infants that diffuse cerebral WM injury was common and associated with proven sep-sis and inotrope use.14 Unfortunately, the principal
method for imaging the premature brain, cranial ul-trasonography, has a low sensitivity for detection of noncystic WM injury identified by either MRI12,13or
neuropathological analysis.15,16The clinical correlate
of the noncystic WM abnormality has not been es-tablished yet, although it is noteworthy that the high frequency of this noncystic disorder is similar to the high frequency (ie, ⬃50%) of later cognitive and behavioral deficits in VLBW infants. Taken together, these data lead to the postulate that the relatively high frequency of cognitive and related neurodevel-opmental deficits documented on follow-up of pre-mature infants who do not exhibit overt WM abnor-malities (ie, cystic injury) on neonatal cranial ultrasonography17,18 is related to noncystic WM
in-jury.
Recent observations in the premature infant sug-gest that the basis for the cognitive and related def-icits may not relate directly to the WM injury per SE. Qualitative appearances of alteration in gray matter (GM) development have been undertaken in this cohort and shown to have a relationship between WM injury and delayed GM gyral development.14
However, qualitative appreciation of both cortical and deep nuclear GM development are extremely limited in comparison to more advanced MRI anal-ysis tools such as 3-dimensional volumetric assess-ment. Thus, several studies of prematurely born in-fants evaluated by volumetric MRI in late childhood and adolescence have shown reductions in either regional total cerebral volumes (GM and WM) or cortical GM volumes and, importantly, correlations of these reductions with impaired IQ,19 memory,20
calculation,21 and global cognitive functioning.22 Of
particular note is that the affected structures have been principally GM, especially cerebral cortical GM.19–23A recent small study of premature infants
reported regional reductions in GM and WM vol-umes that correlated with measures of neurodevel-opment at 18 months.24 However, this small set of
premature infants may not be truly representative, because 9 of 10 had bronchopulmonary dysplasia and 6 of 10 received postnatal steroids. Additionally, the relationship of the presence of neonatal cerebral injury to WM, GM, or both to the subsequent cortical GM alterations was not assessed. The possibility that cerebral cortical changes could reflect alterations in cortical development directly related to the presence of noncystic WM injury was suggested by our recent volumetric MRI study of premature infants at term.25
However, the number of infants studied without WM injury in that series was small (n⫽10), and the
issue of the relation of reduced cerebral cortical GM volume at term to neonatal factors other than WM injury was not resolved conclusively. Resolution of this issue is critical, because detection of reductions in cortical GM as early as term has implications for pathogenesis of the disturbance and, perhaps, for early detection of infants at risk for subsequent cog-nitive deficits.
Thus, the aim of this study was, first, to utilize quantitative volumetric 3-dimensional MRI at term in a large sample of premature infants to determine the presence of alterations in specific cerebral GM and WM structures reflecting altered cerebral struc-tural development. Second, we aimed to define the relationship of any such alterations to such perinatal lesions as WM injury and to other potentially ad-verse perinatal factors. Third, we also aimed to de-fine the relationship of any cerebral volumetric alter-ations to neurodevelopmental outcome at 1 year of age.
METHODS Subjects
One hundred nineteen premature infants with birth weight ⬍1500 g and gestation ⱕ32 weeks’ postconceptional age were recruited into the study to undergo an MRI scan at term equiva-lent. These infants were recruited from 2 geographical sites: Christchurch Women’s Hospital neonatal intensive care unit (New Zealand) and the Royal Women’s Hospital neonatal intensive care unit (Melbourne, Australia). One hundred consecutive premature infants were recruited from Christchurch between November 1998 and March 2001 (90% of those eligible), and 29 consecutive pre-mature infants were recruited from Melbourne between July 2001and September 2001 (80% of those eligible). With the recruit-ment of the premature infants, 21 healthy term-born infants were recruited from the 2 sites (10 from Christchurch and 11 from Melbourne) and were also scanned within 1 week of due date. The term infants were recruited randomly from the labor-and-delivery wards of the same public hospitals in which the premature infants were delivered. The recruitment rate for term infants was 38% of those eligible. An extensive perinatal database was collected re-garding the pregnancy, labor, and neonatal course of the infants. The characteristics of the premature infants are shown in Table 1. Informed parental consent was obtained for all infants. The study was approved by the relevant ethics committees (ie, the Canterbury Health Ethics Committee and the Royal Women’s Hospital Ethics Committee).
MRI
MRI Analysis
For quantitative volumetric analyses, postacquisition process-ing was conducted on workstations (Sun Microsystems, Mountain View, CA). For the 129 infants that were recruited, there were 10 infants with images that could not be processed due to motion artifact (n⫽5), sequence errors (n⫽3), or registration difficulties (n⫽2). Thus, 119 (92%) infants were able to have image process-ing and volumetric analysis. A sequence of established image-processing algorithms was used to segment each of the MRI slices into separate tissue classes: cerebral cortical GM, deep nuclear GM, unmyelinated WM, myelinated WM, and cerebrospinal fluid (CSF) (including ventricular and extra-axial CSF; Fig 1).26,27These
algorithms were designed to reduce imaging-system noise, iden-tify a linear transformation, and resample the DE spin-echo im-ages according to this transform, to align the axial DE spin-echo images with the coronal SPGR images to form a 3-channel data set, classify tissue types on the basis of the MR intensity in the 3 channels, and identify tissue-class surfaces for 3-dimensional vi-sualization. Absolute volumes of each cerebral tissue were deter-mined, and these absolute volumes also were referenced to the total intracranial volume (ICV) and thereby reported as relative percentage of each cerebral tissue within the intracranial cavity. These segmentation and volumetric analyses for this study were undertaken by a single investigator (H.W.) and repeated on every fifth case in a systematic fashion, with⬍5% variance in volumes. For qualitative structural assessment, MR images were graded for WM abnormality in a blinded manner by an experienced
neuroradiologist and by a second independent scoring (T.E.I.) for WM and GM abnormality and development. The scoring system was adapted from previously published systems.28,29 WM was
graded with a score between 1 and 3 for 5 variables including WM signal abnormality (shortening on T1-weighted imaging), qualita-tive reduction in WM volume, apparent cystic abnormality, lateral ventricular size, and the combination of corpus callosum size and maturation of cerebral myelination, particularly in the posterior limb of the internal capsule. WM abnormality then was catego-rized further by the composite scores of these 5 variables into no WM abnormality, mild WM abnormality, moderate-severe non-cystic WM abnormality, or moderate-severe non-cystic WM abnormal-ity.12,14Concordance between assessors was 95%, with 5%
requir-ing consensus on second review. A repeat blinded readrequir-ing resulted in identical scores for 19 of 20 cases.
Neurodevelopmental Evaluation
At 12 months’ corrected age, 112 of 119 premature study infants underwent an evaluation that included a pediatric physical exam-ination by an experienced neurologist or neonatologist and/or a developmental examination based on observational data from parent report and examination with the Denver Developmental Screening tool. Children were classified as exhibiting severe dis-ability if they had clinical evidence of severe abnormality on neurologic motor examination (eg, marked spasticity, weakness, and developmental delay of⬎6 months); moderate disability if they had clinical evidence of moderate abnormality on neurologic
Fig 1. Tissue segmentation (A) from the coronal T2-weighted (B) and coronal SPGR (C) MR images. The tissues are segmented into cortical GM (gray), unmyelinated WM (red), myelinated WM (yellow), deep nuclear GM (white), and CSF (blue).
TABLE 1. Characteristics of the Total Cohort of Premature Infants
Characteristics No. of Infants in Cohort
(% Total Cohortn⫽119 Infants)
Birth weight, g, mean⫾SD (range) 1040⫾272 (440–1505)
Gestational age, wk, mean⫾SD (range) 27.7⫾2.2 (23–32)
Male/female 59:60
Singletons 91 (76%)
IUGR* 26 (22%)
Prolonged rupture of membranes† 26 (22%)
Antenatal steroids administered 103 (87%)
Positive pressure ventilation, d, mean⫾SD (range) 5.7⫾12 (0–62)
Oxygen therapy, d, mean⫾SD (range) 26⫾37 (0–138)
Bronchopulmonary dysplasia (oxygen requirement at 36 wk) 25 (21%)
Pneumothorax 5 (4%)
Postnatal steroid therapy‡ 3 (2%)
Patent ductus arteriosus (confirmed by echocardiography) 38 (32%) Age of full enteral feeding, wk, mean⫾SD (range) 10.3⫾9.2 (0–50)
IVH, any grade 20 (17%)
IVH, grade III/IV§ 7 (6%)
*zscore⬎2 SD below weight for gestational age. † Proven rupture of membranes for⬎24 hours.
motor examination and developmental delay between 4 and 6 months; or mild disability if they had only mild spasticity and/or motor deficit and developmental delay of 2 and 4 months.
Statistical Analysis
Statistical analyses were undertaken with SPSS for Windows (SPSS Inc, Chicago, IL) to compare the volume of cerebral tissues between the premature and the term-born infant groups by using Student’s independentttest with unequal variance assumed. For group analysis within the premature infant groups of the effect of perinatal factors, such as grade of intraventricular hemorrhage (IVH) or WM injury, an analysis of variance (1-way) with pairwise multiple comparison procedures was used. To isolate the group or groups that differ from the others, a multiple comparison proce-dure using the Bonferronittest or Dunn’s method for nonpara-metric data was performed. Correlations were performed between continuous variables of outcome (cerebral volumes) and predic-tors (perinatal facpredic-tors such as length of gestation and birth weight). Among the premature infants, cerebral volumes and relative cerebral volumes were analyzed for potential confound-ing variables by multiple linear regressions. Because of the colin-earity in certain variables of interest, a single variable was selected based on its strength of primary significance with the dependent variable (ie, for gestational age and birth weight, gestational age was selected; for ventilation days and days of oxygen therapy, days of oxygen therapy was selected). The major variables in-cluded in the multiple regression models with cerebral volumes were gestational age, days of oxygen therapy, grade of IVH, grade of WM injury, and intrauterine growth restriction (IUGR).
RESULTS Subjects
One hundred nineteen premature infants were in-cluded with a mean (⫾SD) gestational age of 27.7⫾ 2.2 weeks (range: 23–32 weeks) and a mean (⫾SD) birth weight of 1040⫾272 g (range: 440 –1505 g). The characteristics of the population of premature infants are described in Table 1. There were 21 term-born infants recruited over the same study period. The premature and term infants had no history of prena-tal illicit drug use and did not differ in maternal socioeconomic status or highest level of education. There was no significant difference in head circum-ference (term infants [mean ⫾ SD]: 35.6 ⫾ 0.9 cm; premature infants: 34.9⫾1.3 cm [P⬎ .1]) or weight (term infants: 3.5⫾0.5 kg; premature infants: 3.05⫾ 0.6 kg [P ⬎ .1]) at MRI between the premature and term-born infants.
Cerebral Tissue Volumes of Premature Versus Term-Born Infants
Prematurely born infants had prominently de-creased volumes of cerebral tissue classes at term equivalent in comparison to term-born infants. Thus, when compared with the term-born infants, prema-ture infants had significantly reduced absolute cere-bral volumes of cortical GM (P⫽.001), deep nuclear GM (P⫽.02), and myelinated WM (P ⫽.02) (Table 2). The absolute volumes of both cortical GM and deep nuclear GM were reduced by 22% and the absolute volume of myelinated WM was reduced by 35% for premature infants in comparison to term-born infants. Notably the absolute volumes of my-elinated WM at this age are relatively small. There was a prominent increase in the volume of CSF in the premature versus the term-born infants (P ⫽ .02) associated with the clear reduction in the total abso-lute volume of cerebral tissue within the cranial
cav-ity (Table 2). The absolute cerebral tissue volumes are represented as the percentage of the total ICV in both premature and term infants (Table 2), demon-strating significant reductions in percentage of corti-cal GM (P ⫽ .01), myelinated WM (P ⫽ .05), and brain tissue (P ⫽ .001) and significant increase in percentage of CSF volume (P ⫽.001).
Perinatal Factors Associated With Altered Cerebral Tissue Volumes
Cerebral Injury
Twenty-one infants (18% of all premature infants) exhibited qualitative MRI evidence for moderate or severe WM injury. Of these infants, only 5 had cystic WM injury, and 16 had noncystic diffuse WM injury. The latter consisted principally of diffuse signal changes in the cerebral WM with moderate reduction in WM volume and ventriculomegaly (n⫽8) or with marked reduction in WM volume with ventriculo-megaly, delayed cerebral myelination, and thinning of the corpus callosum (n⫽8).14Between premature
infants with and without WM injury, there was no significant difference in birth weight (WM injury [mean⫾SD]: 1007⫾219; no WM injury: 1081⫾288; P⬎.1) or gestational age (WM injury [n⫽21] 27.7⫾ 2.4; no WM injury [n⫽ 98]: 28.5⫾2.4;P⬎.1).
Premature infants with WM injury had significant reductions in absolute and relative percent ICVs of cortical GM (WM injury: 152⫾32 mL ICV and 33.4% ⫾ 6.8% ICV; no WM injury: 182 ⫾ 41 mL ICV and 40.2% ⫾ 7.6% ICV; P ⬍ .001) and myelinated WM (WM injury: 12.5 ⫾ 4.9 mL ICV and 2.7% ⫾ 1.0% ICV; no WM injury: 16.7 ⫾6.7 mL ICV and 3.7%⫾ 1.6% ICV;P⫽.01), with increased CSF volume (WM injury: 61.1⫾21 mL ICV and 13.4%⫾3.8% ICV; no WM injury: 40.9⫾18.7 ICV and 9.0%⫾3.4% ICV;P ⬍.001). Thus, there was a 17% reduction in absolute cortical GM volume, a 25% reduction in absolute myelinated WM volume, and a 49% increase in ab-solute CSF volume in the presence of WM injury (Fig 2). Expression of the data as a percentage of ICV on TABLE 2. Cerebral Tissue Volumes (Mean⫾SD as absolute volume关mL兴and relative to ICV) for All Premature (n⫽119) and Term-Born (n⫽21) Infants
Tissue Class Premature Infants (n⫽119)
Term-Born Infants (n⫽21)
PValue (tTest)
Cortical GM
Absolute volume, mL 178⫾41 227⫾26 .001
% ICV 39.4⫾7.6 48.2⫾9.5 .01
Deep nuclear GM
Absolute volume, mL 10.8⫾4.2 13.8⫾5.2 .02
% ICV 2.4⫾1.0 2.9⫾0.9 .09
Myelinated WM
Absolute volume, mL 13.5⫾5.8 20.8⫾12 .02
% ICV 3.4⫾1.1 4.2⫾2.2 .05
Unmyelinated WM
Absolute volume, mL 202⫾41 206.5⫾78 .9
% ICV 42.6⫾5.9 41.4⫾9.9 .9
Cerebral tissue, total
Absolute volume, mL 406⫾57 457⫾67 .003
% ICV 89.9⫾4.4 94.1⫾3.1 .001
Cerebrospinal fluid
Absolute volume, mL 45.6⫾22.1 28.9⫾16.8 .01
the day of the MR scan did not alter the significance of the reductions in cortical GM and myelinated WM volumes or the increase in CSF volume.
There were 20 infants with IVH. Ten infants had grade I IVH, 3 infants had grade II IVH, 5 infants had grade III IVH, and 2 infants had grade IV IVH (with parenchymal hemorrhagic infarction). There was over-lap between the presence of IVH and cerebral WM injury, with 3 of 13 (23%) infants with grades I/II IVH displaying moderate-severe WM injury in comparison to 3 of 7 (43%) infants with grades III/IV IVH (Fisher’s
exact test: P ⬍ .05). However, on univariate analysis, CSF volume was the only cerebral volume significantly altered with IVH. Absolute CSF volume was increased significantly (IVH [mean ⫾SD]: 51.9⫾ 23.6; no IVH: 41.6⫾19.6 mL;P⫽ .04), although CSF volumes rela-tive to total ICV were not significantly different be-tween premature infants with IVH and premature in-fants without IVH. The higher grades of IVH (grades III and IV) had a trend toward higher CSF volumes in comparison to the lower grades of IVH, but this did not reach statistical significance.
Fig 2. Cortical GM and CSF volumes at term in premature infants with (n⫽21) and without (n⫽98) evidence of mod-erate-severe WM injury (expressed as medians, with 25th/75th centile box, 10th/90th centile error bars, and outli-ers).
Immaturity
Alterations in cerebral tissue volumes in prema-ture infants at term were found in the most immaprema-ture infants. The infants were subdivided into 2 gesta-tional age groups based on our previous qualitative MRI observations of the impact of immaturity.14On
analysis of the infants into 2 groups (ie, extreme immaturity [ⱕ26 weeks’ gestational age at birth] and more mature [ⱖ27 weeks’ gestational age at birth]), the extremely immature infants exhibited significant reductions in deep nuclear GM (Fig 3) and total cerebral tissue volumes and an increase in CSF vol-umes (Table 3). By contrast, there was no significant influence of immaturity on cerebral myelinated or unmyelinated WM volumes. For percent ICV there remained a significant reduction in deep nuclear GM in the more immature infants (immature infants: 1.6%⫾ 0.7% ICV; mature infants: 2.7%⫾0.9% ICV; P⬍.01), although the statistical significance in rela-tion to CSF volumes and cerebral tissue volumes was lost (P⫽ .1 [not significant]).
Respiratory Illness
There was a significant relationship between the severity of respiratory illness and the deep nuclear GM volumes and percent ICV deep nuclear GM vol-umes. Thus, there was a negative correlation be-tween absolute deep nuclear GM volumes and the number of days of ventilator support (r ⫽ ⫺0.312; P⫽.005) and the number of days of oxygen therapy (r⫽ ⫺0.272;P⫽.01). These findings were similarly significant for volumes of deep nuclear GM relative to ICV (P⬍.01 for all).
Other Influences on Cerebral Tissue Volumes
There was no effect of gender, multiple pregnancy, assisted conception, inotrope use, or infant sepsis on cerebral tissue volumes at term equivalent. Antenatal steroid therapy had no effect on cerebral tissue vol-umes, although no mother received⬎2 doses of ste-roids. Only 3 infants received postnatal steroids, and thus there was an insufficient number of infants for
analysis. IUGR resulted in a significant reduction in the absolute volumes of cerebral cortical GM only (IUGR [mean ⫾ SD] [n ⫽ 26]: 177.2 ⫾ 44 mL; no IUGR (n ⫽87): 192.6 ⫾31.9 mL; P⫽.01).
Multivariate Model
The major variables included in the multiple re-gression models with cerebral tissue volumes were gestational age, days of oxygen therapy, IVH grade, grade of WM injury, and IUGR. After adjusting the relationship of gestational age at birth in the prema-ture infants with volumes of deep nuclear GM and of cortical GM for days of oxygen therapy, IVH grade, grade of WM injury, and IUGR, there persisted a significant effect of immaturity on prediction of vol-umes of both deep nuclear GM (r⫽0.29;R2⫽0.130;
P ⫽ .02), and CSF (r ⫽ ⫺0.21; R2⫽ 0.11; P ⫽ .04).
With this model, the gestational age regression coef-ficient (B) was 0.18.
Neurodevelopmental Outcome at 1 Year
Neurodevelopmental outcome was assessed in 112 of the 119 premature infants (94%) at 1 year of age (2 infants died before follow-up at 1 year of age, and 7 were unable to be tracked). Of these infants, 11 ex-hibited moderate to severe disability. The infants who had moderate to severe disability did not differ in gestational age in comparison to those without (Table 4). However, of these 11 infants, 7 (64%) were classified on qualitative MRI as having severe WM injury. Premature infants with moderate-severe disability had significant reductions in GM volumes, both cortical GM and deep nuclear GM volumes (Table 4). CSF volumes were significantly increased in the infants with moderate-severe dis-ability (Table 4).
DISCUSSION
This study used advanced MR techniques to doc-ument in vivo impaired cerebral development, present by term equivalent, in a large unselected cohort of prematurely born infants. The impairments TABLE 3. Absolute Cerebral Tissue Volumes for Premature Infants ⱕ26 Weeks (n ⫽ 37) in
Comparison With Premature Infantsⱖ27 Weeks (n⫽82) Tissue Class Premature Infantsⱕ26 wk (Range: 23–26 wk;n⫽37)
Premature Infantsⱖ27 wk (Range: 27–32 wk;n⫽82)
PValue (tTest)
Cortical GM, mL 166⫾42 178⫾41 .9
Deep nuclear GM, mL 7.4⫾3.7 12.3⫾4.2 ⬍.01
Cerebrospinal fluid, mL 53.2⫾26.1 38.8⫾18.8 .01
Total ICV, mL 448⫾61 463⫾59 .7
Cerebral tissues, total, mL 386⫾60 419⫾57 .06
TABLE 4. Absolute Cerebral Tissue Volumes for Premature Infants With Moderate-Severe Disability (n⫽11) Versus Premature Infants With No or Minimal Disability (n⫽107 at 1 Year of Age)
Tissue Class Premature Infants With
Moderate-Severe Disability (n⫽11)
Premature Infants With No or Minor Disability
(n⫽101)
PValue (Kruskal-Wallis)
Cortical GM, mL 157⫾41 181⫾41 .04
Deep nuclear GM, mL 8.4⫾3.6 11.3⫾4.2 .03
Cerebrospinal fluid, mL 59.3⫾28.8 44.3⫾21.1 .04
Gestational age at birth, wk 27.1⫾1.6 28.0⫾2.2 NS
in cerebral development include, particularly, highly significant reductions in cerebral cortical and deep nuclear GM volumes in comparison to volumes in term-born infants. These reductions in cerebral GM are accompanied by a near doubling of the ICV of CSF. These cerebral structural changes would not be detectable by conventional cranial ultrasonography and would be difficult to discern by qualitative MR techniques. The decrease in cerebral cortical GM vol-ume was strongly related to the presence of cerebral WM injury, confirming our previous findings with a smaller and selected series of premature infants.25
However, in contrast to this previous study, in this much larger cohort we demonstrate that just being born prematurely, without any evidence of overt cerebral WM injury, also results in a significant al-teration in cerebral structure observable by term equivalent. We also document that these alterations in cerebral structure are related to adverse neurode-velopmental outcome by 1 year of age.
Qualitative MR and Quantitative MR Techniques Qualitative MR studies of prematurely born in-fants in both the neonatal period and childhood have not defined a consistent relationship between the presence of qualitative WM abnormalities defined by conventional MRI and the more common cognitive deficits identified subsequently in these infants.30
This inconsistency may reflect the lack of sensitivity of qualitative MR for the more subtle alterations in cerebral volumetric structure that may underlie these deficits. We previously reported 94 infants from this cohort with qualitative analysis of the nature of ce-rebral abnormality in the WM and GM.14This study
assisted in the definition of cerebral injury, particu-larly that of diffuse cerebral WM injury, alongside the risk factors of proven sepsis and inotrope sup-port. In contrast, qualitative MR analysis is limited in the detection of subtle alterations in the sequence of cerebral structural development, particularly in structures such as the deep nuclear GM. Postacqui-sition MR-processing techniques, using tissue-seg-mentation methods with 3-dimensional renderings, have been applied by our group to quantitate the major cerebral tissue classes (cortical GM, deep nu-clear GM, unmyelinated WM, myelinated WM, and CSF) in the neonatal period and to thereby define normal maturation of these structures between 29 and 41 weeks’ gestation.31The work has also shown
the impact of postnatal dexamethasone administra-tion and periventricular leukomalacia on cerebral volumetric development by term in relatively small selected groups of infants.25,32 More recent studies
using volumetric cerebral analysis in adolescence and later childhood of subjects born prematurely have demonstrated reductions in specific regional cerebral volumes that correlate with impaired cogni-tive functions.19–22A recent smaller study of a group
of sick premature infants (n⫽10) showed reductions in cerebral WM and GM volumes.24 Notably, these
infants had severe respiratory illness with significant postnatal steroid exposure, and comparisons of MRI between premature and term infants were based on somewhat different time points for MRI (premature
infants at 35 weeks versus term infants at 41 weeks). Nevertheless, our current findings in a much larger and more representative group of premature infants are consistent with these pilot findings and support the conclusion that important alterations in cerebral structure in the premature infant occur in the cere-bral GM, involving both cortical and deep nuclear GM, and are present by term equivalent. The in-volvement of GM structures in our study is consis-tent with the GM involvement documented by volu-metric MRI at later ages by others (see earlier), but our findings suggest that potential interventions must be applied before term, perhaps during the neonatal intensive care unit course, to optimize long-term cerebral structural development.
Insights Into the Nature and Pathogenesis of the Cerebral Lesions Associated With Prematurity
This study suggests that disruption of cerebral GM structures is a major feature in prematurely born infants and occurs in relation to 2 critical factors: cerebral WM injury and immaturity. Although the precise nature of the neuropathological abnormali-ties associated with this reduced GM volume is un-known, the major possibilities are either neuronal loss or impaired neuronal differentiation with a re-duction in dendritic and axonal development.
Relationship to Cerebral WM Injury
A role for cerebral WM injury in the genesis of the reductions in cerebral cortical GM at term is clear from the current data. We previously documented in a small case series of premature infants with periven-tricular leukomalacia a significant reduction in cere-bral GM volumes by term.25The reduction in cortical
GM volumes in premature infants with cerebral WM injury could reflect blunted neuronal differentiation caused by destruction of ascending and descending axons (corticopetal, corticofugal, and corticocortical association fibers) in WM with resulting input depri-vation and output isolation of the overlying cortical GM. As a consequence of this cortical GM isolation, there may be impaired GM differentiation.33 These
conclusions are based on the work by Marin-Pa-dilla33with newborn brain using Golgi stains of
au-topsy or neurosurgical specimens of the cortical GM overlying destructive WM lesions; the neuronal changes included impaired development of neuropil. Supportive of the notion of axonal injury in cerebral WM injury are the immunocytochemical findings of axonal injury (-amyloid precursor protein immuno-reactivity) within and surrounding areas of necrosis in periventricular leukomalacia.34–36 However,
whether prominent axonal injury is present in non-cystic WM injury is unknown. Diffusion-imaging studies in vivo in the premature infant have shown that there seems to be an alteration of axonal fiber development in association with periventricular leu-komalacia,37whether cystic or noncystic in nature.8
Relationship to Immaturity
Our data also show a potent effect of gestational age at birth on volumetric measures of cerebral deep nuclear GM at term equivalent. Controlling for many other potential confounders did not alter the signif-icance of this association. There are several potential pathogenetic pathways that may be interactively re-sponsible for the association of immaturity with al-tered deep nuclear GM volumes including direct injury to neurons of cortex and deep nuclear struc-tures or secondary atrophy after sublethal axonal injury, or injury to the vulnerable subplate neurons. The subplate neuronal cell population is particularly abundant in superficial WM of human brain from 22 to 34 weeks of gestation39,40and is crucial for
devel-opment of overlying corticothalamic connectivi-ty,39–47 including the proper guidance of the
“wait-ing” thalamocortical afferents to their final cortical layer (layer IV) targets.48–50 Injury within the
sub-plate zone during this early period therefore could have a highly significant effect on subsequent con-nectivity and development of the deep nuclear GM.51
Relationship to Other Perinatal Factors
There are many potentially adverse perinatal med-ical, nutritional, and environmental factors that could be postulated to cause altered cerebral devel-opment in the premature infant.2,52 In our cohort,
severity of respiratory illness seemed to have a con-sistent impact on deep nuclear GM structure. Inter-estingly, infants with bronchopulmonary dysplasia have been noted to exhibit a movement disorder,53
with neuropathological evidence of neuronal loss in the caudate, putamen, and globus pallidus.
It is interesting that many other factors that we had hypothesized a priori to be of potential importance, such as gender, multiple pregnancy, fertility-assisted conception, poor postnatal growth, patent ductus arteriosus and its treatment, neonatal sepsis, and other markers of severity of medical illness had little or no influence on cerebral structures at term equiv-alent. It is important to note that our sample, al-though the largest published series, may lack ade-quate power to detect smaller independent effects from such perinatal variables.
High-dose postnatal dexamethasone therapy has been shown by our group to be associated with significantly reduced cortical GM and cerebral tissue volumes.32This observation is consistent with
clini-cal data from longitudinal follow-up studies that show an adverse neurodevelopmental outcome after such therapy, and together these findings have led to a reduced use of this potentially harmful drug. This change in practice is reflected in our cohort, because too few infants received postnatal steroids to analyze this effect further. Repeated courses of antenatal ste-roids administered to women at risk of premature delivery also have been hypothesized to have a det-rimental effect on brain development, including a reduction in cerebral gyral development.54 Our
study documented no adverse effect of a partial or completed course of antenatal steroids on cerebral structure. It is important to note that no woman
received ⬎2 completed courses of steroids, but in this large unselected series our findings provide sup-port for a lack of any adverse effect from a single or double prenatal steroid course.
Relationship of Altered Cerebral Volumes to Neurodevelopmental Outcome
The relationship of our measures to functional per-formance in our children at 1 year of age suggests that the MR findings are clinically relevant. Thus, the reduction in cerebral cortical GM and deep nuclear GM volumes seemed sensitive as predictive markers for disability at 1 year. Neurologic outcome at 1 year is acknowledged to be of limited predictive capacity, and thus we will be assessing the functional perfor-mance in our cohort more extensively at later ages. Additionally, regional brain volumetric data com-bined with the potency of later MR evaluation with functional MR55 may also provide critical insights
into the structural bases for plasticity and changing cognitive abilities in these prematurely born in-fants.56
CONCLUSIONS
This study has further defined alterations in cere-bral structure in the largest published cohort of pre-maturely born infants at term equivalent by using in vivo advanced MR techniques. Cerebral cortical and deep nuclear GM were particularly affected. The na-ture of these changes cannot be visualized by cranial ultrasound or conventional qualitative MR tech-niques. The major risk factors for the alterations in cerebral structure were immaturity and MR-defined cerebral WM injury. These findings of altered cere-bral structure were associated with adverse short-term neurodevelopmental outcome. This study shows that alterations in cerebral structure, similar to those found in volumetric studies in older premature infants, are present by term equivalent. Thus, atten-tion must be focused on this critical neonatal period to optimize cerebral structural development.
ACKNOWLEDGMENTS
This study was funded by Lottery Health (New Zealand), the Neurologic Foundation of New Zealand, the Jack Brockhoff Foun-dation (Australia), and the Royal Women’s Hospital (Melbourne, Australia).
We acknowledge the enormous time and efforts of Carole Spencer and Merilyn Bear in assisting with coordinating the stud-ies; Drs Nicola Austin and Lianne Woodward for assistance with the 1-year evaluations; and the families and children of the cohort.
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DOI: 10.1542/peds.2004-0326
2005;115;286
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Terrie E. Inder, Simon K. Warfield, Hong Wang, Petra S. Hüppi and Joseph J. Volpe
Abnormal Cerebral Structure Is Present at Term in Premature Infants
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