THE EFFECT OF DISEASE AND INJURY ON THEADRENAL CORTEX OF STILLBORN AND NEWBORN

INFANTSBY

F. A. LANGLEY and J. C. BURNEFrom the Departnents of Obstetrics and Gynaecolgy and ofPathology, University of Mawchester

(REcEIvED FOR PUBLICATIoN NOVEMBER 4, 1954)

The activity of the adrenal cortex in the newbornhas usually been studied biochemically or by theresponse to stimuli which are adequate in the adult(Bongiovanni, 1951; Heller, 1954). Such investiga-tions have been made on normal, healthy infants.The effect of systemic disease on the adrenal cortexof children, from birth to 15 years, has been studiedmorphologically by Stoner, Whiteley and Emery(1953). Changes in the adrenal cortex of the foetusand newborn infant may be (a) specific, i.e., peculiarto the disease or injury suffered by the infant, suchas the occurrence ofhaemopoietic foci in the adrenalsin haemolytic disease, or (b) non-specific, i.e., notpeculiar to the disease or injury suffered by theinfant. The non-specific response is usually recog-nized by variations in the lipid content of the adult(definitive) zone of the cortex.

This paper records an investigation into the non-specific response to disease and injury of the defini-tive zone of the adrenal cortex in stillborn foetusesand infants dying during the first week of life.

Material and MetbodsThe material for the study of the non-specific response

of the adrenal cortex consisted of a sample of 158 casesdrawn from a pool of about 1,900 necropsies performedon stillborn foetuses and newborn infants. The samplewas unselected except that macerated foetuses wereexcluded and none surviving seven days or more wasincluded. The sections of the glands were stainedroutinely by haematoxylin and eosin and, for lipid, bySudan IV and haemalum.

As_eamnent of Chages in Lipid Content of AhultCortex. Although in adrenals of this age the definitivecortex cannot be divided into the three classical zones,yet there is an ill-defined architectural difference betweenthe sub-capsular region and that adjacent to the foetalcortex. This difference is reflected in the lipid contentof the definitive cortex, the outer (sub-capsular) thirdusually containing less lipid than the inner two-thirdsduring the first week of life. In the first place the lipid

content of the two parts was assessed separately. It wasthen found that the changes in the outer third ran closelyparallel to the changes in the inner two-thirds and thusthe state of the definitive cortex could be assessed as awhole.For the sake of brevity and clarity, and without pre-

judice to any other meaning, the following terminologywas adopted. The cortex was consid to be 'resting'if it contained abundant sudanophilic droplets (Fig. 1),

4-A

I.

FK;. 1.-Resting adrenal cortex. The definitive cortex is packedwith lipid. From a full-term female infant, aged I day, dying from

congenital morbus cordis. Sudan IV and haemalum; x 90.

141

ARCHIVES OF DISEASE IN CHILDHOODwhich in the inner two-thirds were large; to be 'active'when the number and size of the droplets had diminis(Fig. 2); and 'exhausted' when there was complete lossof sudanophilia in the definitive cortex. When thediminution in sudanophilia was slight it was frequentlypatchy.

the number of cases in which the adrenals areresting, active or exhausted, respectively. Apartfrom Fig. 3, the active and exhausted adrenals aregrouped together as 'diminished' sudanophilia.

SUDANOPHILIA FULL mns)Lu IPji,zn M

* i i v^-~~~~~Lamiizi. .; vzEXHAUSTED

U.. CASES 64 24 11 26 25

A'-_A'E SB Cbhrs D2Iws <24hrs 1<3days 3<7dcrysJ~-P. FIG. 3.-The relation between the sudanophilia of the definitive cortex

and the age at death in 158 infants.

(4;^^- ff Asphyxia. The upper part of Fig. 4 shows thatamong asphyxiated infants there was no change infrequency of resting glands in infants who survivedcompared with those who were stillborn. The term

~' 'pure' asphyxia is used to indicate that in theseinfants neither injury, infection nor other disease wasfound at the post-mortem examination. In none ofthese 33 cases was there evidence of focal necrosis orarteriolar changes in the adrenal.

SUDANOPH ILI A.

CASE S

FULL m DIMi'NS-ED14 4 Is

PURE

FiG. 2.-Active adrenal cortex. In the definitive cortex there is ageneral dimution of sudanophila, more obv in the sub-capsular zone. The distnrbution is patchy. A haemopoaetic focuscan be seen (X). From a full-term male infant, aged 36 hours,

dying from haemolytic disease. Sudan IV and haemalum; x 90.

ResultsVartio in Sudanophlia with Age. The changes

in lipid content of the adult cortex with age areshown in Fig. 3. The proportion of stillborninfants with resting adrenals was just under half(28 out of 64) and diminished to a minimum in thosedying between 12 and 24 hours, after which it roseto about the level in the stillborn group. This fallin the frequency of resting glands was not statisticallysignificant, as tested by a 2 x 2 contingency table,but it suggested that further analysis might beinteresting.

In this figure and in those that follow the area ofeach circle represents the number of cases in thecorresponding sub-group and the areas of the sectors

AGE

CONGENITALANOMALI ES

CASES

SB O'bhrs bhrs <7days

10 2FiG. 4.-The effect of 'pure' asphyxia and congenital anomalies on

the lipid content (sudanophilia) of the definitive cortex.

Congenl Anomaly. The lower part of Fig. 4shows that in stillborn infants the frequency of rest-ing glands was not signifintly different in thosedying of congenital anomalies from that in thosedying of asphyxia and of injury; there was noevidence that survival reduced this frequency. Thisgroup of congenital anomalies was small because ofthe exclusion of those dying with any other detect-able lesion.

142

i

I

EFFECT OF DISEASE AND INJURY ON ADRENAL CORTEX OF INFANTS 143

Birth Injury. The effect of birth injury was

examined both in respect of cases of 'pure' birthinjury, i.e., infants dying with no complication, andalso those in whom the injury was combined withasphyxia. These two groups gave essentially thesame result; because of this and because asphyxia,by itself, had no effect on the incidence of restingglands, the results have been combined in Fig. 5.

SUDNOPHILIA

but such cases are infrequent and do not invalidatethese findings.

Hlaemolytic DMase. Eighteen infants out of thesample of 158 suffered from haemolytic disease(Fig. 6). Of those who were stillborn or died

SUDANOPHILIA:

FULL DIMNISHED a

FULLDIMINISHED

CASES 21

AGE SB

7 10 10

0<bhrs 6<72hrs 3<7daysFIG. 5.-The effect of birth injury on the lipid content (sudanophilia)

of the defitive cortex.

Both in stillborn infants and in infants dyingwithin six hours of birth the frequency of restingadrenals was essentially the sane as in the groups of'pure' asphyxia and congenital anomalies. Noneof the infants dying between six and 72 hours afterbirth had resting glands, all showed diminution ofadrenal cortical sudanophilia and two were ex-

hausted. These differences are statistically signi-ficant (see Table 1). After three days four of 10infants had resting glands.

TABLE 1

THE EFFECT OF BIRTH INJURY

Stillborn or Dying betweenState of Surviving Less 6 and 72 Hours Total

Definitive Cortex than 6 Hours

Resing .. .. 14 0 14

Active or exhausted 14 10 24

Total . .. 28 10 38

Whence P is less than 0D005. This indicates that when infants diefrom birth injury active or exhausted glands are significantly more

conmon in infants dying between six and 72 hours than in stillborninfants and those survvmg kss than six hours.

It should be emphasized that this was an un-

selected sample drawn from a much larger popula-tion. Another sample would be expected to havethe same general pattern but to differ in detail. Forexample, infants are occasionally found dyingbetween six and 72 hours after birth in whom thedefinitive adrenal cortex is in the resting condition,

37

CASES 4

AGE SB

7 7

O<hrs bhrs<7daysFiG. 6.-The effect of haemolytic disease on the lipid content

(sudanophilia) of the definitive cortex.

within six hours, only one out of 11 had adrenals ina resting state. Compared with the effects ofasphyxia, congenital anomaly and birth injury thisis statistically significant (Table 2). Likewise there

TABLE 2COMPARISON OF HAEMOLYTIC DISEASE WITH OTHER

CAUSES OF DEATH IN STILLBORN INFANTS ANDINFANTS SURVIVING LESS THAN SIX HOURS

State of Other HaemolyticAdult Cortex Causes Disease Total

Resting .. .. 29 1 30

Active or exhausted.. 29 10 39

Total . .. 58 11 69

Whence P is 0-0113. This indicates that when infants die fromhaemolytic disease active or exhausted glands are significantly morecommon than in infants dying from other causes at this age.

was a marked difference between those dying ofhaemolytic disease between six hours and seven daysand those dying of asphyxia and congenital anomaly.

Pneumonia. Pneumonia was present in 11 infantsand was usually associated with a complicatinglesion such as asphyxia or birth injury. This groupis too small for a detailed analysis. Of the fiveinfants who were stillborn or who died within sixhours of birth, three showed some loss of sudano-philia in the definitive cortex and one complete loss.This pattern is similar to that found in haemolyticdisease. Of those who survived more than sixhours, only three out of six showed diminution ofsudanophilia.

ARCHIVES OF DISEASE IN CHILDHOOD

Weight oftheAdeumii Reition to Lipid CouteL adrenals of premature infants as in those born atSelye (1936a, 1936b) showed that in the mature rat full term.the adrenal responds to noxious stimuli in the phase Dcisionof the 'alarm' reaction by loss of lipid and increase Infants who suffered birth injury showed diminu-in weight. In Fig. 7 the straight line is the regression tion of lipid in the definitive cortex six to 72 hours

14-

t2.

-J

z S.

3r 4wix

'I

- Regression line for I9209.,-rsting adrenals

---95% limits forrsting adrenals

,b _

x Adrenals exhausteor nearly so

0 iBODY WEIGHT (Kg)

2 3 4 5

FIG. 7.-The relton betwee adrenal weight (both gands together) and body weight.

line of adrenal weight on body weight for those

infants in whom the adrenals were in the restingstate. The curved, interrupted lines indicate thelimits within which 95% of the resting adrenals lie.When the weights of the adrenals showing diminu-tion in lipid are superimposed on this frameworkthere is no evidence of increase in weight with loss oflipid from the adult cortex; most of the points lyingwithin the 95% range and being symmeuically dis-tributed about the regression line. Cases ofhaemolytic disease have been excluded from thisanalysis because the adrenal is affected by changesspecific to that disease such as oedema (Burne and

Langley, 1955).

Premaity. In the preceding analysis no con-

sideration has been given to the effects ofprematurity,full-term and premature infants being grouped to-gether. The adult cortex of live-born infants showedless sudanophilia in premature than in full-terminfants (Fig. 8). This difference (which is notstatisticaly signiint) might be attributed to lackof development of sudanophilic granules in pre-

mature adrenals. This was not so, since in stillborninfants full sudanophilia was as frequent in the

after birth; the loss of lipidoccurred less ftequenly inthose who survived morethan thre days. Twointerpretations of this arepossible. First, that be-tween six and 72 hours theadreal response to injuryreaches a maximum to befollowed by a recoveryphase, or secondly, thatadrenal failure plays a partin causing death in the firstthree days of life. Sincethis is not an animal experi-ment there is no way ofdeciding between these twopossibilities. The patternis clearly similar to that inadult rats exposed tonoxious agents (Selye,1936a; 1936b) in which,after a latent period of sixhours, an 'alarm' reactionoccurs which lasts up to 48

hours and is characterized by loss of adrenal corticallipid and by adrenal hypertrophy. Adrenal hyper-trophy was not detected in this series (see Fig. 7)either because it did not occur or because of therelatively large size of the foetal zone maskingchanges in the adult zone.

Haemolytic disease of the foetus and newborn,in contrast to birth injury, is a chronic disease whichstarts to affect the infant before birth and the res-ponse of the adrenal differs. Both in the stillborngroup and in infants who survived up to six daysthere was, as a group, diminished cortical lipid.This corresponds to Selye's third stage of the stresssyndrome which occurs when the noxious stimulusis slight and prolonged. Stoner et al. (1953) foundlittle response to haemolytic disease, five out of sixcases in their series being in the resting phase.The adrenal response to pneumonia in stillborninfants and in infants surviving less than six hourswas similar to that in haemolytic disease. Perhapsthis is because infection may precede delivery bysome hours, e.g., when the membranes have beenruptured 48 hours, or owing to the cumulativeeffects of the multiple lesions which are usuallypresent.

144

td

EFFECT OF DISEASE AND INJURY ON ADRENAL CORTEX OF INFANTS 145

Although the adrenal response of neonatal infantsresembles the stress reaction described by Selye inadult rats there are a number of differences. Theresponse to stress in rats affects the fasciculate zone(Greep and Deane, 1949). In the alarm reactionthis zone suffers lipid depletion and enlargement.In the newborn infant there is no recognizable zonafasciculata; the lipid depletion affects the wholewidth of the definitive cortex and no enlargementcan be recognized by increase of weight or thickness.In the adrenals of rats an increase in sudanophiliaoccurs in the 'resistance' phase with a markedincrease in birefringence, which is maintained afterthe secondary fall in sudanophilia begins. Thispresent study of neonatal adrenals is based onmaterial obtained at routine post-mortem examina-tions over a period of years and no refined histo-chemical tests have been used. Thus it has notbeen possible to distinguish between a resting glandand one in the resistance phase, and glands showingfull sudanophilia in infants dying from four to sixdays after birth may be in the phase of 'resistance'.Since the functional roles of the zones in the humanadrenal are not necessarily the same as in the rats(Bergner and Deane, 1948), and since we are con-sidering the response of an immature adrenal, thecomparison between the adult rat and the infantmust not be pressed too far.The infant adrenal appears to show no response

to asphyxia in terms of changes in sudanophilia ofthe definitive cortex. Gruenwald (1951) describedother changes in the adrenal, namely, focal necrosisand haemorrhage. We have seen such necrosis ofthe adrenals but not in this series and not associatedwith uncomplicated asphyxia. Thus two cases wereof haemolytic disease dying at 20 min. and four daysrespectively, and another in which there was aclinical history of asphyxia in a stillborn infant, butno evidence of such at necropsy. It may well bethat necrosis is a response to chronic stress, asdescribed by Selye and Stone (1950), rather than toasphyxia. Haemorrhage occurs in many parts ofthe body in asphyxia and there is nothing especiallyodd in its occurring in the adrenal. It is probablethat the foetus is adapted metabolically to anoxia,by use of anaerobic mechanisms (Wilson, Reardonand Murayama, 1948), and thus asphyxia is not anadequate stimulus to set going the stress reaction.Wolman (1952) showed that the eosinophil

response to A.C.T.H. was the same in premature asin full-term infants. Jailer, Wong and Engle (1951)tested adrenal function by the effect of injectedadrenalin on the eosinophil count and found thatthere was less fall in the count the more prematurethe infant. Since response of the adrenal cortex to

adrenalin is mediated through the pituitary, whilstthat to A.C.T.H. is a direct response of the adrenalit may be supposed that the observations of Jaileret al. reveals immaturity of the pituitary-adrenalaxis in premature infants. If the reaction of theadrenal to birth injury and haemolytic disease isa stress reaction in Selye's sense then it is mediatedby the pituitary-adrenal axis. Using the yardstick ofdiminished cortical lipid we have failed to demon-strate immaturity of this mechanism.

SUDANOPHILIA: FULL -

CASES

DIMINISHEDa

39 14 8

STILLBORN I I

CROWN-HEEL >47LENGTH (cm)

40<47 <40

SURVIVING UPTO 7 DAYS

CASES 34 27 34FIG. 8.-The effect of prematurity on the lipid content (sudanophilia)

of the definitive cortex.

The results of Stoner et al. (1953) differ from oursin a number of ways, probably because their methodof assessment of the morphological changes isdifferent. They write, 'At birth the [lipid] dropletsare fairly uniformly distributed between the surfaceand the junction of the adult [definitive] cortex withthe X-zone. A few days after birth they tend tobecome concentrated in the inner half of the adultcortex', and quote Sarason (1943) in support of thisdescription. In our observations this appearanceis well recognized, but is associated with a generalfall in sudanophilia and is considered to representadrenal activity; Stoner et al. finding it in theircontrol series consider it 'normal'. Their controlsare described as instances of sudden death, 'still-birth, asphyxia, road accident, etc.'. It seemsunlikely that more than a few of their 23 controlinfants died in the first week as a result of roadaccidents or that stillbirth or asphyxia can be con-sidered normal. We have taken the view that noinfants dying under seven days can be used as normalcontrols; the changes in adrenal morphology canbest be assessed by comparing groups of infants in

146 ARCHIVES OF DISEASE IN CHILDHOOD

relation to the disease or injury causing death and theduration of survival.

SemmaryThe definitive adrenal cortex of stillborn and

newborn infants showed no response to asphyxia.Birth injury caused a diminution of cortical lipidsix to 72 hours after birth; the diminution occurredless frequently in those who survived more thanthree days. Haemolytic disease, in contrast tobirth injury, is a chronic disease starting beforebirth, and the response differed; both in the still-born group and in infants who survived up to sixdays there was diminished cortical lipid. Neonatalpneumonia, beginning before birth, also caused areduction in cortical lipid.

It is concluded that the definitive adrenal cortexat birth is responsive to disease and injury. The

relationship of this to Selye's stress syndrome isdiscussed.

We should like to thank Professor A. C. P. Campbellfor reviewing the manuscript and Mr. P. Stuart for thephotomicrographs and assistance with the charts.

REFERECSBergner, G. E. and Deane, H. W. (1948). Endocrinology, 43, 240.Bongiovanni, A. M. (1951). Amer. J. med. ScL, 222, 710.Burne, J. C. and Langley, F. A. (1955). In prepaaton. theGreep, R. 0. and Deane, H. W. (1949). AnnL N. Y. Acad. Sci.,

50. 596.Gruenwald, P. (1951). Amer. J. Path., 27, 722.Heller, H. (1954). Et. s, 3, 31.Jailer,J. W., Wong, A. S. H. and Engle, E. T. (1951). J. clin. Endocr.,

11, 186.Saason, E. L. (1943). Arch. intern- Med., 71, 702.Selye, H. (1936a). Brit. J. exp. Path.. 17, 234.

(1936b). Nature, Load., 13X, 32.and Stone, H. (1950). On the Experimental Morphology cf theAdrenal Cortex. (Amer. Lec. Ser. no. 74.) Spring_elllnoi

Stoner. H. B., Whiteley, H. J. and Emery, J. L. (1953). J. Path. Boct.,,174.

Wilson, J. L., Reardon, H. S. and Murayama, M. (1948). Pediatrics,1, 581.

Wolman, B. (1952). Archives of Disease in Childhood, 27, 283.