Short 9q interstitial deletion in a neonate with lethal non-immune hydrops

� 2008 Wiley-Liss, Inc. American Journal of Medical Genetics Part A 9999:1–4 (2008)

Research Letter

Short 9q Interstitial Deletion in a Neonate WithLethal Non-Immune Hydrops

Maria Sellitto,1 Rita Genesio,2 Anna Conti,2 Floriana Fabbrini,2 Lucio Nitsch,2

Maria D’Armiento,3 Letizia Capasso,1 Roberto Paludetto,1 and Francesco Raimondi1*1NeonatologiaQ1, Dipartimento di Pediatria, Universita ‘‘Federico II’’, Napoli, Italy

2Dipartimento Assistenziale di Patologia Clinica, Universita ‘‘Federico II’’, Napoli, Italy3Sezione di Anatomia Patologica, Dipartimento di Scienze Biomorfologiche e Funzionali, Universita ‘‘Federico II’’, Napoli, Italy

Received 29 February 2008; Accepted 9 March 2008

How to cite this article: Sellitto M, Genesio R, Conti A, Fabbrini F, Nitsch L, D’Armiento M, Capasso L,Paludetto R, Raimondi F. 2008. Short 9q interstitial deletion in a neonate with lethal non-immune

hydrops. Am J Med Genet Part A 9999:1–4.

To the Editor:

Hydrops fetalis is characterized by diffuse sub-cutaneous edema and fluid collections in some or allserous cavities. Historically, Rhesus isoimmunizationwas the leading cause of hydrops in the newborn.With the institution of passive maternal immuniza-tion and the development of intrauterine fetaltransfusions over the last few decades, non-immunehydrops (NIHF) has become the most common type.Incidence of NIHF is variable, according to differentauthors, between 1:1,400 and 1:3,500 [Keeling, 1990;Maidam et al., 1980]. NIHF carries a limited prognosisand a mortality rate ranging from 50% to 98%[Bukowski and Saade, 2000]. Several hypothesesregarding the pathophysiologic events that lead tofetal hydrops have been suggested. The basicmechanism involved in the development of NIHF isrelated to abnormal fluid transportation betweenplasma and the tissues. This is mainly due to theincrease of hydrostatic capillary pressure and capil-lary permeability and reduction of plasma osmoticpressure or lymphatic flow. Fluid accumulation inthe fetus can result from congestive heart failure,obstructed lymphatic flow, or decreased plasmaosmotic pressure. The fetus is particularly suscep-tible to interstitial fluid accumulation because ofits greater capillary permeability, compliant inter-stitial compartments, and vulnerability to venouspressure on lymphatic return. The causes ofNIHF include congenital infections, genetic syn-dromes, cardiovascular malformations and arrhyth-mias, thoracic masses, renal disorders, renal veinthrombosis, and idiopathic forms. Genetically trans-mitted conditions account for more than 35% of thefetal disorders associated with NIHF. The mostfrequently identified genetic abnormalities are chro-

mosomal disorders in approximately 15% of cases[Jauniaux et al., 1990; Van Maldergem et al., 1992],mostly monosomy X and trisomy 21 [Boyd andKeeling, 1992]. Interstitial deletion of the long armof chromosome 9 is an uncommon cytogeneticabnormality. We report the case of a neonate withprenatal diagnosis of de novo interstitial deletion ofchromosome 9q32-q33 and fetal hydrops. Chromo-some analysis was performed by standard resolution(400–550 bands for set aploid) and GTG banding.The following samples were analyzed: amniocytes(15 clones), fetal lymphocytes of the proband(20 cells) and peripheral blood lymphocytes of bothparents (50 cells each).

Genomic DNA of the patient was investigated bycomparative genomic hybridization using normalmale reference DNA as a control. DNA was isolatedusing standard methods. Briefly, genomic DNA sam-ples were differently labeled by nick translation withSpectrumGreen1-dUTP (Vysis, Downers Grove, IL,test DNA) and SpectrumOrange1-dUTP (Vysis, refer-ence DNA). Test and reference DNAs were mixedtogetherwithunlabeledCot-1DNA(LifeTechnologies,Inc., Gaithersburg, MD), denatured, and hybridized tonormal male human metaphase chromosomes (Vysis).Chromosomes were counterstained with 4,6-diami-dino-2-phenylindole (Vysis). Separate digitized graylevel images of DAPI, Spectrum green, and Spectrumred fluorescence were acquired with a CCD camera

AJMA-08-0164(32350)

*Correspondence to: Francesco Raimondi, M.D., Division of Neona-tology, Department of Pediatrics, Universita ‘‘Federico II’’, Naples, Italy,Via Pansini 5, 80131 Napoli, Italy. E-mail: [email protected]

Published online 00 Month 2008 in Wiley InterScience(www.interscience.wiley.com)

DOI 10.1002/ajmg.a.32350

coupled to an Olympus BX60 microscope. Imageprocessing was carried out using Cytovision 3.52 sof-tware from Applied Imaging. Average green/redfluorescence ratio profiles were calculated for eachchromosome from 8 to 10 metaphases.

FISH analysis was performed on metaphasespreads of the patient using standard techniques.Commercial DNA probes, whole chromosomepainting 9 (Vysis), subtelomeric region 9q (Vysis).Bacterial Artificial Chromosomes (BAC) for regions9q32-q33, RP11-402G3, and RP11-2C9, kindly donat-ed by Dr M. Rocchi, University of Bari, were used toidentify and define the derivative chromosome. TheBACs were extracted using standard methodand labeled by nick translation with Cy3-dUTP(Amersham Pharmacia Biotech Italia, MilanQ2).

A female infant with a birth weight of 2.150 kg wasborn at 30 weeks’ gestation by cesarean delivery. Themother was gravida 1 and para 1, blood group 0 Rhpositive, Coombs test negative. Her first pregnancyhad been normal. She had no family history ofhydrops fetalis, thalassemia, or other congenitalanomalies. She had no underlying medical condi-tion. No abnormal symptoms were reported duringthe pregnancy. Ultrasound examination at 21 weeks’gestation demonstrated a single viable fetus withbilateral pleural effusion (13 mm� 4 mm). Nostructural abnormalities and dysmorphic featureswere found. Following ultrasound examinationsremained unchanged. Amniocentesis performed at24 weeks’ gestation revealed a karyotype of 46, XX,del (9)(q32-q33). Parental karyotypes were normal.

At birth, the infant was intubated in the deliveryroom and required positive pressure ventilation with100% oxygen. Apgar scores were 1 at 1 min, 3 at5 min, 4 at 10 min, 4 at 20 min. In delivery room,pleural effusions were managed with thoracentesesand ascites was also treated with taps. She respondedto resuscitation, was placed on a ventilator andquickly transferred to the neonatal intensive careunit. On physical examination, pale skin color andmarked subcutaneous edema were evident (Fig. 1).She had a thick nuchal fold and rather low-set ears.

Auscultation on the chest confirmed reduced airentry. The abdomen was distended with fluid whileno organomegaly was detectable. Ventilation andcentral vascular access were rapidly secured andshortly thereafter the neonate received a single-volume exchange transfusion with packed ‘‘O’’ Rhnegative red blood cells (blood group 0 Rh positive,Coombs test negative). Chest radiograms showedincreased opacity of lung fields, compatible withhyaline membrane disease, with widening of pleuraland subcutaneous spaces.

The laboratory evaluation revealed leucocytosis(WBC 46.280 cell/mm3), anemia and hypoalbu-minemia. Blood gases analysis showed severe mixedacidosis.

Pleural effusion and ascites increased over thefollowing 12 hr and her clinical condition continuedto deteriorate despite thoracenteses, paracentesesand albumin infusions. At 24 hr of life she presentedwith a desaturation event and bradycardia. Afterepinephrine administration there was a rapid lifeparameters resumption. At 30 hr of life circa, she diedafter a new critical episode. Macroscopic examina-tion revealed a hydropic female newborn withshort neck, thick nuchal fold, absent nipples, andlow-set ears. Heart examination showed a lowinterventricular septal defect. Significant ascites,mildcerebellar hypoplasia with normal brain structure.Both kidneys appeared of slightly reduced dimen-sions. On microscopic examination, nuchal skinshowed edema and lymphatic vessels dilation.

Analysis of metaphases of the proband revealeda male karyotype with a deletion on the q arm ofchromosome 9 de novo.

CGH analysis of 10 metaphase spreads revealeda loss of red fluorescent signal on the q arm ofchromosome 9, which indicated a 6 Mb deletionspanning from 9q32.1 to 9q33.1 in the test DNA(Fig. 2). Chromosome imbalance of the test DNA wasconfirmed by FISH using locus specific probes.Chromosomal abnormality is the most commoncause of genetic NIHF. To date, few reports ofrelatively large interstitial deletions spanning 9q32-q33 have been described. The present case repre-sents the first report of prenatal diagnosis of short

FIG. 1. Image of the patient: note the severe, hard, subcutaneous edema, theshort neck, athelia and low-set ears. [Color figure can be viewed in the onlineissue, which is available at www.interscience.wiley.com.]

FIG. 2. CGH ideogram shows the loss of fluorescent intensity in the deleted9q32-q33 region, as visible from the GTG banding. [Color figure can be viewedin the online issue, which is available at www.interscience.wiley.com.]

2 SELLITTO ET AL.

American Journal of Medical Genetics Part A

interstitial deletion of 9q in association with neonatalhydrops. The karyotype after amniocentesis showeda de novo deletion 9q32-q33. Interestingly, thedeleted region includes the COL27A1 gene. Fibrillarcollagens, such as COL27A1, compose one of themost ancient families of extracellular matrix mole-cules [Boot-Handford et al., 2003]. A modified com-position of the extracellular matrix may alter the fluidbalance between intra- and the extravascular com-partments hence concurring in generating fetalhydrops. A similar mechanism has been proposedfor type II achondrogenesis, which presents withfetal hydrops [Saldino, 1971] and is caused by ausually de novo dominant mutation in COL2A1 gene.

The short span of our deletion may explain theabsence of characteristic features previouslyreported for larger deletions including the 9qsubtelomeric region.

Among the more of 20 cases of 9q interstitialdeletions reported [Wisniewski et al., 1977; Turleauet al., 1978; Sekhon et al., 1982; Ying et al., 1982;Worsham et al., 1989; Farrell et al., 1991; Park et al.,1991; Kroes et al., 1994; Schimmenti et al., 1994;Shimkets et al., 1996; Ayyash et al., 1997; Kleymanet al., 1997; Paoloni-Giacobino et al., 2000; Sasakiet al., 2000Q3; Midro et al., 2004; Chen et al., 2005],five overlap the deletion of our case [Turleau et al.,1978; Farrell et al., 1991; Park et al., 1991; Paoloni-Giacobino et al., 2000; Midro et al., 2004]. Turleau[1978]Q4 described a deletion 9q32-q34 in a 5-month-old boy with craniofacial dysmorphism and absenceof triradii. Farrell [1991]Q5 reported three cases andreviewed the literature on seven other patients withchromosome 9 interstitial deletions. In the secondcase of his series, 9q deletion was not diagnosed onfirst karyotype analyses: it was initially reported asnormal and then it revealed deletion of ‘‘either9q22q32 or 9q32q34.’’ Farrell could not identify acharacteristic phenotype related to the 9q deletedregion. However common features were observed,including developmental delay and malformed ears.In a 28-year-old man with mental retardation andschizophrenia a de novo interstitial deletion 9q32-q34.1 was found. Additional phenotypic abnormal-ities included short stature, a shortwebbedneckwitha low posterior hairline, dysmorphic facies, a narrowpalate with an inverted V soft palate, and taperedfingers with bilateral short fifth metacarpals [Parket al., 1991].

Cystic hygroma and hydrops fetalis have beendescribed in a case with deletion 9q22-q34.1[Paoloni-Giacobino et al., 2000]. Interestingly kar-yotype analysis during choriocentesis did not revealanomalies and deletion was found only whenblood and skin samples were postnatally analyzed.Pregnancy was terminated at the 19th week ofgestation.

Midro [2004]Q6 described an interstitial deletion9q22.32-q33.2 associated with translocation (9;17)

(q34.11;p11.2). This complex chromosomal ano-maly produced the clinical features of ‘‘basal cellnevus syndrome’’ together with someone of theNail-patella syndrome.Cytogenetic analysis revealedinvolvement of PTC and LMX1B genes.

Determining the underlying cause of hydrops isimportant for prognosis and riskof recurrence. Theseresults confirm the need of systematic chromosomeanalysis in fetuses with NIHF. We found thatthe interstitial 9q deletion represents a risk factorfor development of severe fetal hydrops.

The shorter span of deletion with phenotypiccharacteristics similar to a previously describedcase [Paoloni-Giacobino et al., 2000], reduces thechromosomal zone implicated in hydrops develop-ment. This further delineation of the 9q deletionphenotype may so aid diagnosis and geneticcounseling.

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Cindy Chen HE. 2005. Prenatal diagnosis of de novo proximalinterstitial deletion of 9q and review of the literature ofuncommon aneuploidies associated with increased nuchaltranslucency. Prenat Diagn 25:383–389.

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Softproofing for advanced Adobe Acrobat Users - NOTES toolNOTE: ACROBAT READER FROM THE INTERNET DOES NOT CONTAIN THE NOTES TOOL USED IN THIS PROCEDURE.

Acrobat annotation tools can be very useful for indicating changes to the PDF proof of your article.By using Acrobat annotation tools, a full digital pathway can be maintained for your page proofs.

The NOTES annotation tool can be used with either Adobe Acrobat 4.0, 5.0 or 6.0. Other annotation tools are also available in Acrobat 4.0, but this instruction sheet will concentrateon how to use the NOTES tool. Acrobat Reader, the free Internet download software from Adobe,DOES NOT contain the NOTES tool. In order to softproof using the NOTES tool you must havethe full software suite Adobe Acrobat 4.0, 5.0 or 6.0 installed on your computer.

Steps for Softproofing using Adobe Acrobat NOTES tool:

1. Open the PDF page proof of your article using either Adobe Acrobat 4.0, 5.0 or 6.0. Proofyour article on-screen or print a copy for markup of changes.

2. Go to File/Preferences/Annotations (in Acrobat 4.0) or Document/Add a Comment (in Acrobat6.0 and enter your name into the “default user” or “author” field. Also, set the font size at 9 or 10point.

3. When you have decided on the corrections to your article, select the NOTES tool from theAcrobat toolbox and click in the margin next to the text to be changed.

4. Enter your corrections into the NOTES text box window. Be sure to clearly indicate where thecorrection is to be placed and what text it will effect. If necessary to avoid confusion, you canuse your TEXT SELECTION tool to copy the text to be corrected and paste it into the NOTEStext box window. At this point, you can type the corrections directly into the NOTES textbox window. DO NOT correct the text by typing directly on the PDF page.

5. Go through your entire article using the NOTES tool as described in Step 4.

6. When you have completed the corrections to your article, go to File/Export/Annotations (inAcrobat 4.0) or Document/Add a Comment (in Acrobat 6.0).

7. When closing your article PDF be sure NOT to save changes to original file.

8. To make changes to a NOTES file you have exported, simply re-open the original PDFproof file, go to File/Import/Notes and import the NOTES file you saved. Make changes and re-export NOTES file keeping the same file name.

9. When complete, attach your NOTES file to a reply e-mail message. Be sure to include yourname, the date, and the title of the journal your article will be printed in.

Short 9q interstitial deletion in a neonate with lethal non-immune hydrops

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