American Journal of Medical Genetics 134A:430–433 (2005)
Congenital Diaphragmatic Hernia in WAGR Syndrome
D.A. Scott,1,2* M.L. Cooper,1P. Stankiewicz,1A. Patel,1L. Potocki,1,2and S.W. Cheung1
1Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas
2Texas Children’s Hospital, Houston, Texas
Wilms tumor, aniridia, genitourinary anomalies,
and mental retardation (WAGR) syndrome is a
contiguous gene deletion syndrome involving the
Wilms tumor 1 gene (WT1), the paired box gene 6
(PAX6), and possibly other genes on chromosome
genitourinary system and the high incidence of
Wilms tumor and genitourinary anomalies found
in patients with WAGR are attributed to haploin-
sufficiency of this gene. It has been hypothesized
that WT1 also plays an important role in the
development of the diaphragm. During mamma-
lian embryonic development, WT1 is expressed in
the pleural and abdominal mesothelium that
forms part of the diaphragm. Furthermore, mice
that are homozygous for a deletion in the mouse
homolog of WT1 have diaphragmatic hernias.
Case reports describing congenital diaphrag-
matic hernias in infants with Denys–Drash and
Frasier syndromes, both of which can be caused
by mutations in WT1, provide additional support
for this hypothesis. We report an infant with
aniridia, bilateral cryptorchidism, vesicoureteral
reflux, and a right-sided Morgagni-typediaphrag-
matic hernia. G-banded chromosome analysis
revealed a deletion of 11p12-p15.1. Breakpoint
regions were refined by fluorescence in situ
hybridization (FISH) and deletion of the WAGR
critical region, including WT1, was confirmed. A
review of the medical literature identified a
second patient with a deletion of 11p13, a left-
sided Bochdalek-type diaphragmatic hernia, and
anomalies that suggest a diagnosis of WAGR
bilateral cryptorchidism, and a hypoplastic scro-
tum. These cases demonstrate that congenital
diaphragmatic hernia can be associated with
WAGR syndrome and suggest that deletions of
WT1 may predispose individuals to develop con-
genital diaphragmatic hernia.
? 2005 Wiley-Liss, Inc.
KEY WORDS:WAGR; congenital diaphragmatic
hernia; chromosome 11p13; WT1
Wilms tumor, aniridia, genitourinary anomalies, and men-
tal retardation (WAGR) syndrome (OMIM 194072) is a
1gene (WT1), thepaired box 6gene (PAX6), andpossibly other
genes on chromosome 11p13. The risk for Wilms tumor
approaches 50% and is attributable to deletion of WT1. WT1
is also required for normal formation of the genitourinary
system and deletion of this gene is the most likely cause of the
genitourinary anomalies seen in individuals with WAGR
[Pritchard-Jones et al., 1990; Kreidberg et al., 1993]. Aniridia
and associated anterior chamber defects are caused by
haploinsufficiency of PAX6 which encodes a transcriptional
regulator involved in oculogenesis [Hanson and Van Heynin-
gen, 1995]. Mental retardation in WAGR ranges from border-
line to severe. However, individuals with normal intelligence
have been reported [Huff, 1994].
We report an infant with aniridia, bilateral cryptorchidism,
vesicoureteral reflux, and a right-sided Morgagni-type dia-
deletion of 11p12-p15.1. Breakpoint regions were refined by
fluorescence in situ hybridization (FISH) and deletion of the
WAGR critical region, including WT1, was confirmed. This is
the first report of congenital diaphragmatic hernia in a child
with a clinical diagnosis of WAGR syndrome. This case adds to
the growing body of evidence, in both humans and mice, that
WT1 plays an important role in diaphragm development and
that abnormalities in WT1 may predispose individuals to
develop congenital diaphragmatic hernia.
Our patient is a 2-month-old Caucasian male born at
28 weeks gestation to a 37-year-old mother and a 42-year-old
father. He is this couple’s first child; however, his father had
three healthy children from a previous marriage and his
mother had two healthy children and two early miscarriages
from a previous marriage. The remainder of family history is
The pregnancy was complicated by preterm labor and
premature rupture of membranes approximately 1 week prior
to delivery. Routine prenatal testing, including several
alities. The child was born via spontaneous vaginal delivery in
the breech position.
At delivery, he was intubated and chest compressions were
given secondary to low heart rate. Apgar scores were two at
1 min and six at 5 min. Birth weight was 965 g (25th centile).
Physical examination revealed bilateral cryptorchidism. The
testes remained impalpable at 2 months of age. Minor
anomalies that became apparent at 2 months of age include a
unilateral cup-shaped ear and a squared appearance to the
After birth, a radiological evaluation forrespiratory distress
revealed evidence of a moderately sized right-sided Morgagni-
type congenital diaphragmatic hernia with a significant
amount of liver protruding into the chest cavity. The patient
Grant sponsor: Baylor Child Health Research Center (NIH);
Grant number: K12 HD41648.
*Correspondence to: Daryl A. Scott, 633E, One Baylor Plaza,
Houston, TX 77030-3498. E-mail: firstname.lastname@example.org
Received 9 November 2004; Accepted 2 February 2005
? 2005 Wiley-Liss, Inc.
underwent primary closure of this hernia at 6 weeks of age. A
renal ultrasound revealed no structural abnormalities but a
reflux on the right with a possible small ureterocele. An
echocardiogram wasnormal. Head ultrasound showed agrade
II intraventricular hemorrhage on the left and grade IV
intraventricular hemorrhage on the right.
Ophthalmologic abnormalities included hazy corneas, small
bilateral anterior polar cataracts with iris strands, increased
intraocular pressure, bilateral anterior segment dysgenesis,
large pupil diameters, and irregular iris boarders consistent
with a diagnosis of aniridia. The patient did not have
MATERIALS AND METHODS
High-resolution G-banded metaphase chromosome analysis
was performed on peripheral blood lymphocytes using stan-
dard protocols. Metaphase spreads for FISH were prepared
from peripheral blood lymphocytes using a standard protocol.
FISH was carried out using BAC clones selected based on
mapping information from both the UCSC Browser (http://
genome.ucsc.edu) and the National Center of Biotechnology
clones B2.1, P60, and FAT5 from the WAGR critical region
TABLE I. Summary of Fluorescence In Situ Hybridization (FISH) Analysis Defining the
Centromeric and Telomeric Breakpoints on Chromosome 11p
map position (Mb)a
Signals on deleted
aThe approximate map position and cytogenetic location is based on information from both the UCSC browser
segment of chromosome 11 is indicated.
Partial karyotype from high-resolution chromosome analysis
Diaphragmatic Hernia in WAGR 431
clones were provided by the Baylor Human Genome Sequenc-
ing Center and a commercial probe was used to identify the
chromosome 11 centromere (Vysis, Downers Grove, IL). BAC
clones were grown in media containing 20 mg/ml chloramphe-
nicol and DNA was isolated using a commercially available kit
(Eppendorf, Hamburg, Germany). The DNA was labeled with
biotin-14-dUTP or digoxigenin-11-dUTP by standard nick
translation and used for FISH analysis using standard
protocols [Trask, 1991]. Signals were detected by FITC-avidin
and rhodamine-conjugated anti-digoxigenin antibodies, and
the slides analyzed using a Zeiss Axiophot fluorescent micro-
scope equipped with both single-band pass filters and a triple-
band pass filter. Digital images were captured by a Power
Macintosh G3 system and MacProbe version 4.3 (Applied
Imaging, Santa Clara, CA).
G-banded chromosome analysis revealed a cytogenetically
visible deletion of chromosome 11p12-p15.1 (Fig. 1). Meta-
phase FISH analysis confirmed that PAX6 and WT1 were
deleted and further defined the breakpoints (Table I). One
signal was seen with probes RP11-583F24 and RP11-361M6
and RP11-159D8. This pattern is consistent with the patient’s
maximal deletion spans approximately 19.2 Mb. The patient’s
parents did not consent to chromosome analysis despite
The eye and genitourinary anomalies seen in our patient,
combined with a cytogenetic evidence of a deletion of WT1,
PAX6, and other markers within 11p13, substantiate a
diagnosis of WAGR. Although, congenital diaphragmatic
hernia has not been reported previously in a child with a
clinical diagnosis of WAGR syndrome, a review of the
literature revealed a report of congenital diaphragmatic
hernia associated with a deletion of chromosome 11p13
[Gustavson et al., 1984]. The child in this report died shortly
after birth and an autopsy revealed a large left-sided
Bochdalek-type diaphragmatic hernia with intrathoracic
herniation of a large portion of the intestines and the left lobe
of the liver. Additional findings included large fontanelles,
wide sutures, bilateral microphthalmia, low-set dysplastic
ears, redundant nuchal folds, hepatomegaly, small penis,
bilateral cryptorchidism, and a hypoplastic scrotum. No iris
anomalies were noted; however, a dedicated ophthalmologic
examination was not performed. Chromosome analysis
revealed an interstitial deletion of 11p13 and a possible
deletion of 11p14. Further delineation of the chromosomal
breakpoints was not reported and deletion of PAX6 and WT1
was not documented. Clinical findings of microphthalmia,
cryptorchidism, and hypoplastic scrotum in conjunction with
an 11p13 deletion are highly suggestive of WAGR syndrome.
Congenital diaphragmatic hernia has been described in
other individuals with abnormalities of WT1 (Table II).
Devriendt et al. [1995, 1996] described a child with Denys–
Drash syndrome (OMIM 194080)—male pseudohermaphro-
ditism, nephropathy, and Wilms tumor—and a left-sided
Bochdalek-type diaphragmatic hernia who harbored a hetero-
zygous Arg366His mutation in WT1. Denamur et al. 
ism, and gonadoblastoma—and a left-sided diaphragmatic
hernia who harbored a heterozygous 1228þ4 C!T alter-
native splice site mutation in WT1.
Abnormalities in WT1 are not always associated with CDH.
CDH occurs in only a small portion of individuals with WAGR
syndrome, and Arg366His and 1228þ4 C!T mutations in
Frasier syndrome, who do not have diaphragmatic defects
[Pelletier et al., 1991; Baird et al., 1992; Barbaux et al., 1997].
Although, it is possible that the diaphragmatic hernias seen in
individuals with these syndromes are not associated with the
presence of an abnormality in WT1, it is more likely that
heterozygous mutations/deletions of WT1 increase the like-
lihood that an individual will develop CDH. Environmental
factors and differences in genetic background may also play a
role in determining whether an individual with a WT1
abnormality develops CDH. Although, disruption of another
gene(s) on 11p could play a role in the development of
diaphragmatic hernia in patients with chromosome 11p13
Arrows indicate the deleted chromosome 11. A: FISH shows hybridization
signals with probes RP11-583F24 (red) and the chromosome 11 centromere
(green). B: FISH shows hybridization signals with probe RP11-361M6 (red)
and the chromosome 11 centromere (green).
Fluorescence in situ hybridization (FISH) results for patient.
432 Scott et al.
deletions,involvementofasecondgeneintheregionisunlikely Download full-text
in the patients with Denys–Drash and Frasier syndrome who
have point mutations in WT1.
Studies in mice provide a second line of evidence suggesting
that disruption of WT1 function may lead to the development
of diaphragmatic hernias. Homozygous WT1 null-mice die
between the 13 and 15 day of gestation. Examination of
homozygous embryos revealed defects in urogenital develop-
ment and diaphragmatic hernias [Kreidberg et al., 1993]. A
role for WT1 in diaphragm development is further supported
mesothelium both of which contribute to diaphragm develop-
ment [Pritchard-Jones et al., 1990].
Although, WT1 may play an important role in diaphragm
development, abnormalities in WT1 do not appear to be a
common cause of isolated diaphragmatic hernia. In a study of
21 children with isolated congenital diaphragmatic hernia,
to have mutations or rearrangements in WT1 [Nordenskjo ¨ld
et al., 1996]. This finding is not unexpected since all of
individuals with congenital diaphragmatic hernia associated
with WT1 described to date have had genitourinary tract
two individuals with both congenital diaphragmatic hernia
and genitourinary anomalies but examination of a larger
number of individuals will be required to determine if WT1 is
responsible some cases in this subpopulation.
her helpful discussion.
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mutations in the WT1 gene in patients with Denys Drash syndrome.
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Elion J, Gubler MC, Fellous M, Niaudet P, Loirat C. 2000. WT1 splice-
and segmental glomerulosclerosis. Kidney Int 57:1868–1872.
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TABLE II. Summary of Clinical Findings in Patients With Abnormalities of WT1 and Congenital Diaphragmatic Hernia
diaphragmatic herniaAdditional findings
This report WAGR syndrome11p12-p15.1,
WT1 and PAX6
Craniofacial, unilateral cup-shaped ear;
genitourinary, cryptorchidism, unilateral grade
II–III vesicoureteral reflux, possible small
ureterocele; ophthalmologic, aniridia; other,
squaring of fingertips
Craniofacial, large fontanelles, wide cranial
sutures, low-set, dysplastic ears; genitourinary,
small penis, bilateral cryptorchidism,
hypoplastic scrotum; ophthalmologic, bilateral
microphthalmia, narrow palpebral fissures;
other, hepatomegaly, redundant nuchal skin
Craniofacial, hypoplastic skull bones, diastasis
of cranial sutures, large fontanelle,
epicanthal folds; genitourinary, male
pseudohermaphroditism, double vagina,
dysgenic ovaries, focal mesangial proliferative
glomerulosclerosis, irregular mesangial
sclerosis; other, hypoplastic lungs
Genitourinary, proximal hypospadias with
chordee, right testicular ectopia,
steroid-resistant nephrotic syndrome, focal
and segmental glomerulosclerosis
Gustavson et al.
WAGR syndrome? 11p13-11p14?Large, left-sided,
Devriendt et al.
Denamur et al.
Left-sided, type not
Diaphragmatic Hernia in WAGR 433