709: Gene expression profiling of lymphoblastic cell lines from monosomy 1p36 patients reveals differential regulation (expression) of cardiac and neurologic relevant genes

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www.AJOG.org ances in a single cell can be detected by oligo-based array CGH when there is enough high-density coverage in the targeted regions. 0002-9378/$ – see front matter • doi:10.1016/j.ajog.2009.10.725

709 Gene expression profiling of lymphoblastic cell lines from monosomy 1p36 patients reveals differential regulation (expression) of cardiac and neurologic relevant genes Avishai Alkalay1, Melanie Babcock2, Anna Bergsmedh2, Dennis Monks2, Mary E. King1, Lisa Shaffer3, Bernice Morrow1 1

Albert Einstein College of Medicine/Montefiore Medical Center, Bronx, New York, 2Albert Einstein College of Medicine, Bronx, New York, 3 Signature Genomic Laboratories, Spokane, Washington

OBJECTIVE: Genetic interactions are significant determinants of 1p36 deletion syndrome (1p36DS). However, identification of causal genes and downstream genetic pathways has been hampered by genetic and phenotypic heterogeneity. Objective #1: Analyze the whole-genome mRNA expression profile from lymphoblastoid cell lines (LCLs) from patients with 1p36DS and verify this approach in this population. Objective #2: Identify common genetic pathways that may be responsible for the phenotype variation of patients with this syndrome, which include neurologic impairment, cardiovascular anomalies, hypothyroidism, and auditory and ophthalmologic dysmorphism. STUDY DESIGN: A sample of nine patients with 1p36DS confirmed by FISH analysis was compared to five unaffected controls. Affymetrix GeneST arrays were used to analyze gene expression levels. Genespring was used to analyze the fold changes between cases and controls. The 1p36 region was investigated to determine whether there was reduced expression of genes within the deleted interval. Ingenuity Pathway Analysis was then used to explore important common pathways among the differentially expressed genes elsewhere in the genome. RESULTS: Out of 97 genes within the largest sized deletion, 31 genes were found to be consistently down regulated and none were up regulated, confirming the functional deletion of genes within this region. There was a total of 332 genes dysregulated outside of the 1p36 region. The most common pathways affected by the dysregulated genes include those for neurologic development, endocrine function, and cardiovascular development. CONCLUSION: Our results provide evidence that candidate genes for 1p36DS are differentially expressed in LCLs. In addition, molecular profiling of LCLs can identify common pathways downstream of genes deleted within the 1p36 region. Finally, neurologic development, endocrine function, and cardiovascular development have been identified as the most common pathways affected by the dysreguated genes. This is consistent with physical defects commonly seen in patients with 1p36DS. 0002-9378/$ – see front matter • doi:10.1016/j.ajog.2009.10.726

710 Fetal cells detection by endocervical sampling at first gestational trimester Stavros Sifakis1, Apostolos Zaravinos1, Satish Ghatpande2, Antti Seppo2, Elena Vakonaki1, Michael W Kilpatrick2, Triantaphyllos Tafas2, Petros Tsipouras2 1

University Hospital of Heraklion, Crete, Department of Obstetrics & Gynecology, Heraklion, Crete, Greece, 2 Ikonisys Inc, New Haven, Connecticut

OBJECTIVE: Fetal trophoblasts are present in the uterine cervix from 5-15 weeks and can been obtained with a variety of methods. Our previous work demonstrated that dual FISH labeling combined with automated microscopy is an efficient and sensitive method to detect rare cells. Herein, we apply this approach to fetal cells obtained from the uterine cervix during the first trimester of pregnancy. STUDY DESIGN: Pregnant women at 6-13 weeks of gestation were enrolled in the study after informed consent. The samples were obtained with the use of a cytobrush after insertion through the external os to a depth of 1.5 cm following by one full turn similar to obtaining a Pap smear. After Carnoy fixation the slides were hybridized with two Y-

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specific and one X-chromosome specific FISH probes and scanned using an automated system developed specifically for rare cell detection and analysis RESULTS: In 18 confirmed male pregnancies, the detection rate of fetal nuclei was 67%, with a mean of detected nuclei 2.6 and a range of 1-11 nuclei per case. Almost 50,000 nuclei were examined per case by the automated FISH analysis. The determination of fetal gender was carried out sonographically during the second trimester of the pregnancy, and confirmed either at birth or by cytogenetic analysis in some cases that an invasive test was performed. Although spotting after sampling was present at 35% of examined women no one returned for medical evaluation. CONCLUSION: Fetal cells are present at the external segment of the endocervical canal and they can be collected safely by the use of a cytobrush at 6-13 wks of pregnancy. An automated FISH analysis system can recognize trophoblasts from male pregnancies with a detection rate of 67%. To determine the proper gestational week of sampling for obtaining the greater number of trophoblasts, a larger sample size is required. These encouraging results support the launch of additional studies to evaluate the possible diagnostic or screening efficacy of this approach in the non-invasive prenatal diagnosis of fetal chromosomal abnormalities. 0002-9378/$ – see front matter • doi:10.1016/j.ajog.2009.10.727

711 Genetic variation in the GLUT1 gene of patients affected with meningomyelocele Clint Cormier1, Hope Northrup1, Kit Sing Au1 1

University of Texas Health Science Center at Houston, Houston, Texas

OBJECTIVE: Recent studies in our laboratory showed association be-

tween the glucose transporter 1 (GLUT1) gene and risk for meningomyelocele (MM). The current study sought to examine the diversity and extent of sequence variations in GLUT1 in patients with MM and to identify variations conferring risk of MM. STUDY DESIGN: Sequences of the 10 exons of GLUT1 for 96 MM patients (48 Caucasian and 48 Hispanic) were determined by direct sequencing of DNA. Amplified DNA was sequenced using BigDye terminator method. Sequences from each sample were compared to GenBank reference sequences (RefSeq) and to one another using gene alignment software and manual analysis. Frequencies of known single nucleotide polymorphisms (SNPs) were compared based on patient ethnicity and MM lesion level. Variants not catalogued in the RefSeq represent potential novel variants. Variants without population frequencies reported by public database were examined and comparisons were made between our MM patients and Caucasian and Hispanic controls. RESULTS: Allele frequencies for 7 known SNPs were determined: rs1385129 (c⬎t, .82/.18), rs11537641 (c⬎t, .84/.16), rs2229682 (g⬎a, .85/.15), rs2229681 (c⬎t,1.0/0), rs2306662 (a⬎g, 1.0/0), rs2236574 (c⬎t, 1.0/0), and rs13306458 (c⬎a, 1.0/0). Among MM patients, the rs1385129 variant C allele was more frequent in Caucasians than Hispanics (82% vs. 67%, P⫽0.02). Comparative PCR analysis of a 10bp deletion within intron 9 showed it to be more common in Caucasian MM patients than in Caucasian controls (P⫽ 0.02). Additionally, Hispanics with MM occurring at and above L1 were more likely to have the deletion variant than individuals with MM below L1 (73% vs 46%, P⫽0.03). CONCLUSION: We describe genotype frequencies of several known SNPs and a 10bp deletion polymorphism in the GLUT1 gene in our population of MM patients. In our population, presence of the 10bp deletion in intron 9 is associated with risk of MM for Caucasian MM patients. 0002-9378/$ – see front matter • doi:10.1016/j.ajog.2009.10.728

Supplement to DECEMBER 2009 American Journal of Obstetrics & Gynecology

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