Congenital diaphragmatic hernia and chromosome 15q26: determination of a candidate region by use of fluorescent in situ hybridization and array-based comparative genomic hybridization

Abstract

Congenital diaphragmatic hernia (CDH) has an incidence of 1 in 3,000 births and a high mortality rate (33%-58%). Multifactorial inheritance, teratogenic agents, and genetic abnormalities have all been suggested as possible etiologic factors. To define candidate regions for CDH, we analyzed cytogenetic data collected on 200 CDH cases, of which 7% and 5% showed numerical and structural abnormalities, respectively. This study focused on the most frequent structural anomaly found: a deletion on chromosome 15q. We analyzed material from three of our patients and from four previously published patients with CDH and a 15q deletion. By using array-based comparative genomic hybridization and fluorescent in situ hybridization to determine the boundaries of the deletions and by including data from two individuals with terminal 15q deletions but without CDH, we were able to exclude a substantial portion of the telomeric region from the genetic etiology of this disorder. Moreover, one patient with CDH harbored a small interstitial deletion. Together, these findings allowed us to define a minimal deletion region of approximately 5 Mb at chromosome 15q26.1-26.2. The region contains four known genes, of which two--NR2F2 and CHD2--are particularly intriguing gene candidates for CDH.

Figure 1

Array CGH results. a, Patient 1, with CDH and del(15) interstitial deletion. b, Patient 3, with CDH and r(15)(p11q26). c, Patient 4, with CDH and der(15)t(3;15)(q29;q26.1). d, Patient 7, with CDH and del(15)(q25q26.3). e, Patient 8, without CDH and with r(15)(p11.1q26.3).

Figure 2

FISH results. a, Patient 1: partial metaphase, probe D15Z4 (red signal) at chromosome 15 centromeric locus and probe RP11-114I24 (green signal) at 15q26.3. b, Patient 2: interphase, probe RP11-369K8 (red signal) and RP11-253B9 (green signal) near the chromosome 5 centromeric region at 5p13.2. c, Patient 3: partial metaphase and interphase, deletion probe RP11-143C19 (green signal) and normal probe RP11-64K10 (red signal) at 15q23. d, Patient 4: metaphase spread, gain of chromosome 3q29; probe RP1-196F4 (red signal) (3qtel) present on der(15) and normal signal probe D15Z4 (yellow/red signal) at the centromeric region of chromosome 15. The der(15) contains both signals. e, Patient 4: partial metaphase, deletion probe RP11-183E24 (green signal) at 15q26.2 and normal probe D15Z4 (yellow/red signal). f, Patient 6: interphase, deletion probe RP11–57P19 (red signal) and normal probe D15Z4 (green signal). Patients 1–6 all have CDH.

Figure 3

Schematic representation of the critical CDH region, with partial ideogram of chromosome 15q. Patients (Pt) 1–7 have CDH; patients 8 and 9 do not have CDH. BAC clones that were tested by array CGH and FISH are listed. The black dots inside boxes indicate that probes have been tested only on the array and not by FISH. The smallest common overlapping deletion interval involved in CDH is denoted by the large red square.