Journal of Medical Genetics (1970). 7, 27.

Congenital Adrenal Hypoplasia-An X-linked DiseaseLESTER WEISS and RAYMOND C. MELLINGER

From the Department of Pediatrics and the Division of Endocrinology, The Henry Ford Hospital,Detroit, Michigan, U.S.A.

Dacou and DiGeorge (1968) reported in abstractform their findings from a study ofthree male infantswith adrenal hypoplasia. These infants by historyhad a common mother and two different fathers. Atabout the same time Bickham, Silvestre, andMellinger (1968) reported on two affected male sibswho had a common mother and different fathers.Both authors suggested X-linked inheritance in thisdisorder, but genetic data documenting the pater-nity of these individuals were not presented ineither instance.The present report describes an additional

affected male born to the mother of the patientsstudied by Bickham, and includes genetic datadocumenting differing paternity and pathologicalconfirmation of adrenal hypoplasia.

Case ReportThe patient was first seen at the Henry Ford Hospital

at 8 days of age. His mother became concerned whenhe began persistent vomiting two days before admission,because two of three older children, who were known tohave adrenal hypofunction, had experienced similarsymptoms.The patient was a male infant born after an uncom-

plicated 40-week pregnancy. The only medicationstaken during pregnancy were iron and vitamins. Hisbirthweight was 4082 g. He had lost weight continu-ously since birth and there was gradually increasinglethargy.The family pedigree obtained from the mother is

indicated in Fig. 1. In two marriages she had bornefour children, all male. The first child (II.1) has adre-nal hypofunction and is under treatment. The secondchild (II.2), who was born in a second marriage, isnormal. The next child (II.3) had adrenal hypoplasiaand is under treatment, and the proband is shown asII.4. There were no miscarriages. The first child wasdiagnosed as having adrenal hypoplasia at the age of 3years 4 months. However, he had not gained weightduring the previous two years and weighed only 11-8 kg.Growth and development have been normal since be-ginning corticosteroid therapy. The diagnosis in the

Received 5 August 1969.

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second affected child (II.3) was made at 14 days of agewhen he was admitted to hospital because of weight loss,dehydration, and lethargy. His growth and develop-ment have also been normal since the institution ofspecific therapy.The initial physical examination when the patient was

8 days old revealed a thin, dehydrated, lethargic whitemale infant. His pulse was 160 a minute and his bloodpressure was 64 mm. Hg systolic. He weighed 3515 g.and was 53 cm. long. The anterior fontanelle wassunken and the oral mucous membranes were dry. Thelungs were clear and the heart had no murmurs. Nointra-abdominal masses were palpable. A reducibleright inguinal hernia was present. The genitalia werethose of a normal male. There were no abnormalitiesof the extremities and no apparent abnormality of skinpigmentation.

Laboratory studies. The haemoglobin was 9-0 g./100 ml., WBC 16,000/cu. mm.; the differential count,urinalysis, blood urea nitrogen, and creatinine werenormal. The serum sodium was 125, potassium 8-6,chloride 100 and CO2 13 mEq/l. Calcium was 9-7 andphosphorus 4-2 mg./100 ml. Radiological examinationof the chest was normal.The patient was treated with intravenous fluids and

electrolytes, desoxycorticosterone acetate, and cortisone.After his condition was stabilized he was given hydro-cortisone and fluorhydrocortisone orally and additionalsodium chloride in his formula. He was discharged at6 weeks of age weighing 4450 g. and was well until 3months of age when he was readmitted for the repair of aright inguinal hernia. Local anaesthesia was used forthe operation, and 50 mg. of cortisone acetate was givenintramuscularly the evening before and the morning andevening of the operation. During operation unexpecteddifficulty was encountered in reducing the hernia.

NormalAdrenal hypoplasia

II I:v(i: (9< Green,RedFIG. 1 jtheqamily pedigree,obtained scolour blindFIG. 1. The family pedigree, obtained by history.

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TABLE IACTH STIMULATION TEST

Creatinine 17-OHCS 17-KS 17-KGS. PregnanetriolPatient Age (yr.) Test Day (mg./24 hr.) (mg./24 hr.) (mg./24 hr.) (mg./24 hr.) (mg./24 hr.)

1.2 Adult Control 10-3 7-1ACTH (50 U x 3-day) 41 6 18 7

II.1 34/12 Control 118 03284ACTH (40 U b.i.d.) 148 0-8 3-5

Control 157 2-3 1-8 4-511.2 5 Control 138 2-7 2-7 3-2

ACTH (20 U b.i.d.) 173 6-1 1-7 11-6

Control 63 0-5 0 4 0-8 0-1Control 40 0-6 0-6 0 4 0-111.3 2 wk. ACTH (20 U b.i.d.) 54 0 5 0 4 1-0ACTH (20 U b.i.d.) 69 1-1 0 3 2-3 0-15

Control 24 1-4 0-2 2-4

II.4* < 2 wk. Control 34 129 3 1ACTH (40 U b.i.d.) 43 2-2 41 001ACTH (40 U brivd.) 48 2f7400

*Patient was receiving fluorhydrocortisone 0-1 mg. b.i.d.

Manipulation resulted in haemorrhage into the wall ofthe intestine, and a suture was required to control thisbleeding. On the following day the patient developedfever, tachycardia, and vomiting. Coffee-groundmaterial was noted in the vomitus. In spite of cortisone,desoxycorticosterone acetate, and intravenous fluids andelectrolytes the infant died the next day.

Endocrine studies. Urinary 17-hydroxycorticos-teroids were determined by a modification of the methodof Porter and Silber (1950), the urinary 17-ketosteroidsby the procedure of Sobel et al. (1958), 17-ketogenicsteroids by the method of Metcalf (1963), and pregane-triol by a modification of the procedure of Bongiovanniand Clayton (1954), using gas chromatography for finalidentification and quantitation. The results of thesedeterminations on control days and after administrationof ACTH are shown in Table I. Data were obtainedfrom the mother and all four of her children. In themother (I.2) and the unaffected child (II.2) urinaryadrenal steroid levels were normal and rose more thantwofold after ACTH administration. In the otherthree children, levels of corticoid metabolites were lowand failed to increase after ACTH. In patient II.4 theapparent increase in 17-hydroxycorticosteroid excretionafter ACTH represents only a more complete collection

I.1I.21.3HA.1II.2II.3II.4

ABO C D

A1 Cc DO cc DB cc DA1 cc DB cc DO cc DO Cc D

E MN

eeEeEE

eeEEEEEe

MNMNNNMM

MNMNINN

of urine, as indicated by a proportional increase increatinine level (24 mg. vs. 48 mg.). These corticoiddata in conjunction with the low 17-ketosteroid and preg-nanetriol values established the diagnosis of primaryadrenal hypofu.nction.

Genetic studies. The results of blood group,colour vision, and glucose-6-phosphate dehydrogenasedeterminations in this family are shown in Table II.The data indicate different paternity for each of thethree adrenal deficient children. The blood groups ofthe affected individual II.1 are compatible with those ofthe mother I.2 and the putative father 1.1. The bloodgroups of affected individual II.3 and the unaffectedchild II.2 are compatible with the putative parents I.2and I.3. The blood groups of the 4th child (II.4) bornto the mother are considered incompatible with paternityof either I.1 or I.3. The paternity of I.1 is ruled out bythe Fya findings. The paternity of I.3 is ruled out by Cand Fya. These findings were confirmed by repeatedspecimens in two different laboratories.

All 7 subjects had normal glucose-6-phosphate de-hydrogenase and all those tested except the child withnormal adrenal function had normal colour vision. Thischild had mild red-green colour blindness of an un-

classified type as determined by the Hardy-Rand-Rittler

LE II

N = Normal ND = No data D = Red-green colour blind.

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pseudo-isochromatic plates. The infant II.4 was nottested for colour vision.The Xga blood group was negative in all family mem-

bers. No grandparents were available for study; thematernal grandfather was not available for colour visiontesting.

Necropsy findings. The necropsy was conductedunder the direction of Dr. R. P. Patton, Department ofLaboratories. There was a recent surgical incision inthe right inguinal area, 5 cm. in length. Haematomaswere found in the subcutaneous tissues in the area ofoperation and in the underlying peritoneum. Therewere several haematomas in the mesentery of the distal15 cm. of the ileum. The thymus weighed 6 g. Thethyroid weighed 1-5 g. and revealed no abnormalities.The heart was normal but both lungs were very con-gested and there were multiple subpleural haemorrhages.The bronchi contained a small amount of bloody mucoidmaterial. Frothy, bloody material exuded from the cutsurface of the parenchyma of the lungs. The spleenweighed 14-5 g. and was otherwise unremarkable. Theliver, which weighed 200 g., the gall-bladder, and biliarysystem were all normal. The oesophagus was normalbut the stomach contained blood-tinged mucoid materialand there were several areas of superficial ulceration ofthe mucosa. Mucosal petechiae were noted in theduodenum and jejunum. Haemorrhage was present inthe wall of the distal 15 cm. of the ileum, in addition tothe peritoneal and mesentery haematomata. The colonand rectum were normal. The pancreas weighed 7 g.the right kidney 30 g., and the left 25 g., and the testiclesweighed 2-5 g. each. All were normal. Careful exami-nation of the suprarenal region and the para-aorticregions disclosed no evidence of adrenal glands. Thebrain was moderately oedematous with congestion of theleptomeninges. The pituitary gland appeared normal.

Microscopical examination revealed abnormalitiesonly in the following organs: the right and left lung hadinterstitial infiltration of polymorphonuclear leucocytesand mononuclear cells, chiefly leucocytes and histio-cytes. The alveolar spaces were lined with hyalinemembrane and contained patchy haemorrhages. In thedistal part of the ileum, diffuse congestion and haemor-rhage were seen in the mucosa, submucosa, between themuscle layers, and in focal areas of the subserosa arealayer. The mucosal epithelium was almost completelyautolysed. A small area of interstitial haemorrhage waspresent in the right testicle. No ectopic adrenal corticaltissue could be seen in the sections of either epididymis.The thyroid gland, thymus, and parathyroid glands werenormal on microscopical examination. Serial sectionswere made of perirenal fat and tissue from both para-lumbar areas. Minute adrenal glands were found inthe perirenal fat in the normal location. They measured0-7 x 0-2 mm. in size, and the estimated combined weightwas less than 0-5 g. There was complete disarray of thenormal architecture of the adrenal cortex, with the cellsarranged in a compact mass without normal zone or cordformation. Two types of cells were present. The pre-dominant cells had eosinophilic granular cytoplasm with-

out vacuoles and small round hypochromatic nuclei,some of which were pycnotic. Cellular limits were illdefined. The other cells were small groups of largeeosinophilic cells which have been described as charac-teristic of congenital adrenal hypoplasia (Fig. 2 and 3).Autonomic ganglia were identified. The pituitary glandshowed no remarkable changes. Staining with PiercePAS stain demonstrated normal distribution of thecellular components of the hypophysis. Sections of thecerebrum, brain-stem, and cerebellum showed diffusecongestion and oedema without any focal lesions.

DiscussionThis report documents adrenal cortical insuf-

ficiency in the newborn son of a mother who hadborne three other male children, two with adrenalhypofunction. Blood group determinations estab-lished a different father for each of the affectedchildren. The occurrence of adrenal hypoplasia inthree males with the same mother and differentfathers is strong evidence indicating that thiscongenital anomaly is due to an X-linked gene.McKusick's (1966) extensive catalogue ofMendelianinheritance in man lists hypoadrenal corticalism inthe group of recessive disorders. An erroneousconclusion in this reference work could result insignificant errors in genetic counselling. Thepossibility that this condition is due to a maternalnon-genetic cause is not absolutely excluded but isconsidered extremely unlikely.

Unfortunately, we obtained no information re-garding linkage to Xga or glucose-6-phosphatedehydrogenase, because all family members wereXga negative and had normal G6PD. Colourvision was normal in all the family members withthe exception of the single unaffected child. Theproband was too young to be tested. Therefore,colour vision cannot be used in this family to showcrossing-over between the locus for adrenal hypo-plasia and that for colour vision.

In order to substantiate further congenital adrenalhypoplasia as an X-linked disease we reviewed thereported cases of congenital hypoadrenalism. Wedid not exclude cases in which the pituitary was notmentioned nor those in which the patient was alivewhen reported. We did exclude three male sibsreported by Roberts initially in 1949 and subse-quently in detail in July 1952 (Roberts and Baehren),because in the latter report it is apparent that thesechildren had congenital adrenal hyperplasia. Theresults of this review support our contention thatcongenital adrenal hypoplasia is X-linked, and that,as suggested by Dacou and DiGeorge (1968), thepathological anatomy in affected female infantsdiffers from that found in male infants. We found28 cases of congenital hypoadrenalism (including

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_

a

Fl w t lN f x

''J- e(t tj' et 9*

FIG. 2. The right adrenal gland showing the complete lack of organization of the cortex. ( x 175.)

--

.

aa_.FIG. 3. The right adrenal gland showing both the small compact cells and the groups of large eosinophilic cells. ( x 830)

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those in this family) not associated with tuberculosis,adrenal haemorrhage, or pituitary abnormalities.Twenty-three of these occurred in male infants(Dacou and DiGeorge, 1968; Bickham et al. 1968;Zondek and Zondek, 1968; Boyd and MacDonald,1960; Stempfel and Engel, 1960; Mitchell andRhaney, 1959; Gardner, 1957; MacMahon, Wagner,and Weiner, 1957; Harlem and Myhre, 1957;Denys, Corbeel, and Malbrain, 1955; Deamer andSilver, 1950; Provenzano, 1950; Geppert, Spencer,and Richmond, 1950; and Sikl, 1948) and five infemales (Winquist, 1961; Shepard, Landing, andMason, 1959; Williams and Robinson, 1956; Weensand Golden, 1955). Of the affected males, 15 hadaffected male sibs, but none had an affected femalesib. Of the reported female patients there wasonly one sib pair (Shepard et al., 1959). In thisfemale sibship, the onset of symptoms was in thesecond year of life while in the males the onset wasusually in the first weeks of life. One of the girlsdied, and at necropsy the adrenal glands were de-scribed as 'shell-like and resembled connectivetissue of the surrounding area'. No cells could befound in the zona fasciculata and reticularis, butoccasional clumps of cortical cells were described inthe zona glomerulosa.The adrenal glands of the female patient reported

by Williams and Robinson (1956) as Case 4 had awell-formed definitive cortex with little evidence offetal cortex. This tissue is similar to that found inanencephalic infants, though the pituitary glandwas not mentioned in this report. The adrenalhistology in the female patient described by Win-quist (1961) is reported as, 'histologically normalglomerular and fascicular zones and a slightly moreconspicuous 'permanent' reticular zone. There wasno fetal reticular zone'. The adrenal cortex ofCase 2, reported by Weens and Golden (1955), wasdescribed as having a thin but orderly zona fascicu-lata, slight evidence of the zona glomerulosa, and anormally involuting zona reticularis. Thus, his-tology in these female infants is distinctly differentfrom that described for affected males, in whomthere is disorganization of the cortex with scatteredgroups of large eosinophilic cells in haphazardarrangements. This description is consistent in allexcept three reports of male infants. Provenzano(1950) did not mention large cells, and described a'mature type differentiation into glomerular andfascicular zones'. Stempfel and Engel (1960)found 'no adrenal tissue'. Gardner (1957) does notdescribe the histology of the adrenal gland. In ourpatient and presumably in his similarly affected sibsthe histological picture is very similar to that foundin most of the previous cases. Adrenal hypoplasia

with complete disorganization of the cortex, theabsence of the normal formation of cords of cellsand the presence of large eosinophilic cells is there-fore characteristic of X-linked adrenal hypoplasia.This histological picture and the clinical appearanceof adrenal hypofunction in the first few months oflife suggests the failure of an inducer substancewhich normally is responsible for the formation ofthe adult cortex as the fetal cortex undergoes in-volution. A recessive gene located on a maternalX-chromosome is undoubtedly responsible for thefailure of adrenal maturation and function.

SummaryAdrenal hypoplasia not associated with abnor-

malities of the pituitary gland has been consideredto be either an autosomal recessive disease or asporadic event. We have studied a family inwhich the gene for this abnormality was inherited asan X-linked recessive. A summary of previouscases supports this observation.Three of four male sibs had adrenal hypofunction

shown by clinical and biochemical determinationsand failure of adrenal response to ACTH stimula-tion. One infant died after hernia repair, and hypo-plasia of the adrenal glands in the presence of a nor-mal pituitary gland was confirmed at necropsy.Blood group determinations established differentfathers for each of the affected children. X-linkedinheritance is therefore believed to be establishedin this family.Of 25 additional reported cases of adrenal hypo-

plasia not associated with tuberculosis, adrenalhaemorrhage, or pituitary abnormalities, 20 weremale and 5 were female. Fifteen affected maleshad affected male sibs, while only one femalesibship is reported. The characteristic adreno-cortical histology found in males, lack of organiza-tion of the adult cortex into cords, and the presenceof clumps of large pale staining cells, has not beendescribed in females. These findings support thecontention that this distinctive form of adrenalhypoplasia is inherited as an X-linked recessive.The histology suggests the failure of an inducer sub-stance which normally stimulates formation ofadult cortex as the fetal cortex undergoes involution.

We are grateful to Dr. S. M. Saeed for performing theblood group determinations.

REFERENCESBickham, R. E., Silvestre, D., and Mellinger, R. C. (1968). Addi-

son's disease in half-siblings. Henry Ford Hospital MedicalJournal, 16, 157-162.

Bongiovanni, A. M., and Clayton, G. W., Jr. (1954). A simplifiedmethod for the routine determination of pregnanediol and preg-nanetriol in urine. Bulletin of the Johns Hopkins Hospital, 94,180-186.

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Weiss and MellingerBoyd, J. F., and MacDonald, A. M. (1960). Adrenal cortical

hypoplasia in siblings. Archives of Disease in Childhood, 35, 561-568.

Dacou, C., and DiGeorge, A. M. (1968). Evidence for an X-linkedform of congenital adrenal hypoplasia. The Society for PediatricsResearch, 38th Annual Meeting, Atlantic City, New Jersey,U.S.A.

Deamer, W. C., and Silver, H. K. (1950). Abnormalities in thesecretion of the adrenal cortex during early life. Journal ofPediatrics, 37, 490-516.

Denys, P., Corbeel, L., and Malbrain, H. (1955). Hypocorticismesimple congenital. Acta Paediatrica Belgica, 9, 169-182.

Gardner, L. I. (1957). Newer knowledge of adrenocortical dis-turbances. Pediatric Clinics ofNorth America, 4, 889-904.

Geppert, L. J., Spencer, W. A., and Richmond, A. M. (1950).Adrenal insufficiency in infancy. A clinical classification, review,and a report of a case. J7ournal of Pediatrics, 37, 1-22.

Harlem, 0. K., and Myhre, E., (1957). Congenital adrenal hypo-plasia. American_Journal of Diseases of Children, 94, 696-701.

McKusick, V. A. (1966). Mendelian Inheritance in Man. Catalogsof Autosomal Dominant, Autosomal Recessive, and X-linked Pheno-types. Johns Hopkins Press, Baltimore.

MacMahon, H. E., Wagner, R., and Weiner, D. B. (1957). Acuteadrenal insufficiency due to congenital defect. American Journalof Diseases of Children, 94, 282-285.

Metcalf, M. G. (1963). A rapid method for measuring 17-hydroxy-corticosteroids in urine. J7ournal of Endocrinology, 26, 415-423.

Mitchell, R. G., and Rhaney, K. (1959). Congenital adrenal hypo-plasia in siblings. Lancet, 1, 488-492.

Porter, C. C., and Silber, R. H. (1950). A quantitative color reac-

tion for cortisone and related 17, 21-dehydroxy-20-ketosteroids.J7ournal of Biological Chemistry, 185, 201-207.

Provenzano, R. W. (1950). Adrenocortical hypoplasia in the new-born infant. Report of a case with replacement therapy. NewEngland Journal of Medicine, 242, 87-89.

Roberts, M. H. (1949). Emergencies encountered in the neonatalperiod. Journal of the American Medical Association, 139, 439-444.

-, and Baehren, P. F. (1952). Adrenocortical insufficiency,observed in three male infants in one family. J'ournal of the Medi-cal Association of Georgia, 41, 299-304.

Shepard, T. H., Landing, B. H., and Mason, D. G. (1959). FamilialAddison's disease. Case reports of two sisters with corticoiddeficiency unassociated with hypoaldosteronism. AmericanJournal of Diseases of Children, 97, 154-162.

Sikl, H. (1948). Addison's disease due to congenital hypolasia ofthe adrenals in an infant aged 33 days. Journal of Pathology andBacteriology, 60, 323-324.

Sobel, C., Golub, 0. J., Henry, R. J., Jacobs, S. L., and Basu, G. K.(1958). Study of the Norymberski methods for determination of17-ketogenic steroids (17-hydroxycorticosteroids) in urine.Journal of Clinical Endocrinology and Metabolism, 18, 208-221.

Stempfel, R. S., Jr., and Engel, F. L. (1960). A congenital familialsyndrome of adrenocortical insufficiency without hypoaldosteron-ism. Journal of Pediatrics, 57, 443-451.

Weens, H. S., and Golden, A. (1955). Adrenal cortical insufficiencyin infants simulating high intestinal obstruction. AmericanJournal of Roentgenology, 74, 213-219.

Williams, A., and Robinson, M. J. (1956). Addison's disease ininfancy. Archives of Disease in Childhood, 31, 265-269.

Winquist, P. G. (1961). Adrenal hypolasia. Archives of Pathology,71, 324-329.

Zondek, L. H., and Zondek, T. (1968). Congenital adrenal hypo-plasia in two infants. Acta Paediatrica Scandatica, 57, 250-254.

AddendumAfter this manuscript was prepared a report appeared

documenting two additional male sibships with thehistological characteristics (Uttley, 1968).

REFERENCEUttley, W. S. (1968). Familial congenital adrenal hypoplasia.

Archives of Disease in Childhood, 43, 724-730.

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