Quantification of the placental epigenetic signature of the SERPINB5 gene in maternal plasma of pregnancies complicated by small for gestational age.

INTRODUCTION: To investigate the association between pregnancies with small for gestational age (SGA) neonates and the concentration of cell-free fetal DNA or cell-free total DNA in maternal plasma during the first and second trimesters using tissue-specific epigenetic characteristics of the SERPINB5 gene. METHODS: A nested case-control study was conducted with maternal plasma collected at 11 to 26 gestational weeks from 51 women with SGA neonates and 102 controls. We performed a real-time quantitative methylation-specific PCR to quantify concentrations of unmethylated-SERPINB5 (U-SERPINB5) as a cell-free fetal DNA marker and methylated-SERPINB5 (M-SERPINB5) as a cell-free total DNA marker. RESULTS: A positive correlation was observed between U-SERPINB5 and M-SERPINB5 concentrations in both control (r = 0.363, p < 0.001) and SGA groups (r = 0.548, p < 0.001). Moreover, the concentration of U-SERPINB5 or M-SERPINB5 was significantly positive correlated with gestational age at sampling in both controls (U-SERPINB5: r = 0.397, p < 0.001; M-SERPINB5: r = 0.275, p = 0.005) and SGA (U-SERPINB5: r = 0.274, p = 0.052; M-SERPINB5: r = 0.439, p = 0.001). However, the concentration of U-SERPINB5 or M-SERPINB5 was not correlated with birthweight. At 11-14 weeks, U-SERPINB5 and M-SERPINB5 concentrations in SGA did not differ significantly from those of controls. There were also no statistically significant differences in the concentrations of U-SERPINB5 and M-SERPINB5 between SGA and controls at 15-26 weeks of gestation. DISCUSSION: Our findings suggest that U-SERPINB5 and M-SERPINB5 concentrations in maternal plasma during early pregnancy are not associated with pregnancies who delivered SGA neonates.

Genome-wide microRNA expression profiling in placentas of fetuses with Down syndrome.

INTRODUCTION: Down syndrome (DS) is the most common aneuploidy, caused by an extra copy of all or part of chromosome 21 (chr21). Differential microRNA (miRNA) expression is involved in many human diseases including DS. However, the genome-wide changes in miRNA expression in DS fetal placentas have yet to be determined, and the function of these changes is also unclear. METHODS: We profiled genome-wide miRNA expression in placenta samples from euploid or DS fetuses by using microarray technology and predicted the functions of differentially expressed miRNAs using bioinformatics tools. RESULTS: Thirty-four miRNAs were significantly differentially expressed in the DS placenta compared with the normal placenta (16 up-regulated and 18 down-regulated). However, expression of chr21-derived miRNAs did not change. Predicted target genes included 7434 genes targeted by up-regulated miRNAs and 6071 genes targeted by down-regulated miRNAs. Seventy-six of these target genes were located on chr21 (10 genes controlled by down-regulated miRNAs and 34 genes by up-regulated miRNAs, and 32 genes by both). Target genes on chr21 were significantly associated with DS and DS-related disorders, such as mental retardation, neurobehavioral manifestations, and congenital abnormalities. DISCUSSION: To our knowledge, this is the first genome-wide study to comprehensively survey placental miRNAs in DS fetuses. Our results provide new insight into miRNA expression in placentas of fetuses with DS. Additionally, our findings indicate that the differentially expressed miRNAs in the DS placenta may potentially affect various pathways related to DS pathogenesis.

Unusual maternal uniparental isodisomic x chromosome mosaicism with asymmetric y chromosomal rearrangement.

Infertile men with azoospermia commonly have associated microdeletions in the azoospermia factor (AZF) region of the Y chromosome, sex chromosome mosaicism, or sex chromosome rearrangements. In this study, we describe an unusual 46,XX and 45,X mosaicism with a rare Y chromosome rearrangement in a phenotypically normal male patient. The patient's karyotype was 46,XX[50]/45,X[25]/46,X,der(Y)(pter→q11.222::p11.2→pter)[25]. The derivative Y chromosome had a deletion at Yq11.222 and was duplicated at Yp11.2. Two copies of the SRY gene were confirmed by fluorescence in situ hybridization analysis, and complete deletion of the AZFb and AZFc regions was shown by multiplex-PCR for microdeletion analysis. Both X chromosomes of the predominant mosaic cell line (46,XX) were isodisomic and derived from the maternal gamete, as determined by examination of short tandem repeat markers. We postulate that the derivative Y chromosome might have been generated during paternal meiosis or early embryogenesis. Also, we suggest that the very rare mosaicism of isodisomic X chromosomes might be formed during maternal meiosis II or during postzygotic division derived from the 46,X,der(Y)/ 45,X lineage because of the instability of the derivative Y chromosome. To our knowledge, this is the first confirmatory study to verify the origin of a sex chromosome mosaicism with a Y chromosome rearrangement.

Fetal and neonatal outcomes in women taking domperidone during pregnancy.

The safety of domperidone in pregnancy remains unknown. Therefore, the study aimed to prospectively evaluate the fetal outcomes of women who were taking domperidone during pregnancy. In a prospective cohort study design, 120 1st- trimester pregnant women who were taking domperidone for controlling gastrointestinal tract symptoms and 212 age-matched pregnant women not exposed to any potential teratogenic agent, were followed-up until delivery. In the case group, domperidone was indicated for control of functional gastrointestinal disorders in 59.2%, the maximum dose was 30 mg/day and exposure occurred between 2(+4) and 20 weeks' gestation. Fetal outcomes including gestational age at birth, birth weight and length, head circumference at birth, and 1- and 5-min Apgar score were similar in the two study groups. There were three babies born with malformations in each group (OR = 0.6; 95% CI 0.1, 2.8). In conclusion, domperidone does not appear to be a major human teratogen. However, our findings require further confirmation in larger studies.

Association of fetal-derived hypermethylated RASSF1A concentration in placenta-mediated pregnancy complications.

OBJECTIVES: To assess whether fetal-derived hypermethylated RASSF1A concentrations in maternal plasma during pregnancy are altered in pregnancies associated with placental dysfunction manifested by intrauterine growth restriction (IUGR), preeclampsia (PE), or placental previa (PP) and whether this alteration can be detected in susceptible subjects before the onset of clinical disease. METHODS: We performed a real-time quantitative polymerase chain reaction to quantify RASSF1A concentrations before and after methylation-sensitive restriction digestion in maternal plasma at 7-41 gestational weeks of normal pregnancies (n = 161), IUGR (n = 43), PE (n = 22), PP (n = 14) and non-pregnant women (n = 20). RESULTS: A positive correlation was observed between fetal-derived hypermethylated RASSF1A concentration and gestational age for all study groups (r = 0.624, p < 0.001 for IUGR; r = 0.381, p = 0.042 for PE; r = 0.697, p < 0.001 for PP; r = 0.560, p < 0.001 for controls). The concentration of hypermethylated RASSF1A was relatively high at 7-14 gestational weeks in all patient groups. Hypermethylated RASSF1A concentration at 15-28 weeks was significantly higher in patients who subsequently developed IUGR (p = 0.002), PE (p < 0.001) or PP (p < 0.001) than in controls. CONCLUSION: We first demonstrated increased concentration of fetal-derived hypermethylated RASSF1A sequences according to advancing gestation and before the onset of the clinical manifestation of pregnancy complications secondary to placental dysfunction, such as IUGR, PE and PP. Hypermethylated RASSF1A in maternal plasma may be useful as a potential biomarker to detect placental-mediated pregnancy complications, regardless of fetal gender and polymorphism.

Foetal and neonatal outcomes in first-trimester pregnant women exposed to abdominal or lumbar radiodiagnostic procedures without administration of radionucleotides.

BACKGROUND/AIM: Little is known about exposures to low radiation doses in the first trimester of pregnancy and deterministic adverse effects in the offspring, and risks are extrapolated from catastrophic events or from exposures to radiotherapy. The study aimed to assess the foetal and neonatal outcomes of pregnant women exposed to radiodiagnostic procedures with abdominal or lumbar irradiation. METHODS: In a prospective cohort design, we studied the foetal and neonatal outcomes in 115 singleton pregnant women who required abdominal or lumbar radiodiagnostic procedures without the administration of radionucleotides, and in 527 age-matched (± 2 years) control pregnant women. RESULTS: In the exposed group, lumbar spine radiography (33.9%), plain abdominal radiography (16.5%) and upper gastrointestinal tract radiography with abdominal irradiation (15.7%) were the most common radiodiagnostic procedures. Major congenital malformations were identified in two (1.9%) babies born in the exposed group and in two (0.4%) babies born in the control group (odds ratio = 4.7; 95% confidence interval 0.7-33.6; P = 0.15). The rest of the foetal and neonatal outcomes was similar in the two groups except by a marginally higher rate of admissions to the neonatal intensive care unit among babies born to exposed women (odds ratio = 2.9; 95% confidence interval 1.0-9.4; P = 0.06). CONCLUSION: Our results indicate that X-ray and computed tomography scan exposure involving abdominal irradiation without the administration of radionucleotides is not associated with adverse foetal and neonatal deterministic outcomes. Efforts are required to reduce the use of radiodiagnostic procedures for general check-ups in childbearing age women.

Molecular and clinical characteristics of 26 cases with structural Y chromosome aberrations.

Structural abnormalities include various types of translocations, inversions, deletions, duplications and isochromosomes. Structural abnormalities of the Y chromosome are estimated to affect less than 1% of the newborn male population and are particularly hazardous for male reproductive function. The objective of this study was to characterize a group of patients with structural abnormalities of the Y chromosome. All patients who visited our laboratory between 2007 and 2010 underwent cytogenetic investigations. Among these, we detected 26 patients with structural abnormalities of the Y chromosome. To confirm the structural Y chromosome alterations, we used special bandings, FISH and multiplex PCR to detect Y chromosome microdeletions. Of the 26 patients presented here, 11 had an isodicentric Y chromosome, 7 had an inversion, 3 had a translocation, 2 had a derivative, 2 had a Yqs and 1 had a deletion. Sixteen were diagnosed with azoospermia, 8 as normal fertile males and 1 as a man who was unable to donate semen due to mental retardation. One of the patients having 45,X/46,X,idic(Y) was reported to be phenotypically female with primary amenorrhea and without uterus. Deletions of the AZFbc region were correlated with the sperm concentration (p < 0.05), but no correlation with the levels of FSH, LH, testosterone, prolactin and estradiol were found. The present report shows that the precise identification of structural Y chromosome aberrations may be clinically important for genetic counseling and assisted reproductive technology treatment.

Pregnancy outcome after exposure to the probiotic Lactobacillus in early pregnancy.

The present study prospectively assessed pregnancy outcome of women taking probiotics during the periconceptional period. A group of 104 women who had taken Lactobacillus in early pregnancy and 200 age- and parity-matched control pregnant women exposed to non-teratogenic agents were also recruited into the study and followed-up prospectively. Median gestational age of women exposed to Lactobacillus was 5.2 (range: 1.9-17.6) weeks. Exposure was at a mean dose of 510 mg/day for a median of 4.0 days (range: 1-90 days). In the exposed group, pregnancy outcomes included 96 live births and eight spontaneous abortions versus 187 live births and 21 spontaneous abortions in the non-exposed group. There was no statistical difference in adverse pregnancy outcomes, including the number of spontaneous abortions, pre-term births as well as a low birth weight between the two groups (p > 0.05). In the exposed group, there were two (2.1%) major congenital malformations in comparison with five (2.7%) in the comparison group (p = 0.7). In conclusion, no association was identified between ingestion of Lactobacillus in early pregnancy for a limited period of time and adverse pregnancy outcomes. However, rare pregnancy outcomes may have been missed due to the limited sample size included in the study.

Hypoxia inducible factor-1α gene polymorphisms in Korean patients with pre-eclampsia.

BACKGROUND: Placental hypoxia has been implicated in the pathogenesis of pre-eclampsia. Hypoxia inducible factor-1α (HIF-1α) is activated by low oxygen tension and is a key regulator of genes involved in the cellular responses to hypoxia. AIM: We determined whether maternal blood c.1722C>T (Pro582Ser) and c.1790G>A (Ala588Thr) polymorphisms in exon 12 of the HIF-1α gene are associated with pre-eclampsia. SUBJECTS AND METHODS: Subjects included 163 pre-eclamptic patients (48 mild and 115 severe preeclampsia) and 194 healthy pregnant women. Polymorphisms were genotyped by PCR and direct DNA sequencing. RESULTS: There were no significant differences in genotype or allele frequencies of the c.1772C>T and c.1790G>A polymorphisms of the HIF-1α gene among the study groups. Moreover, subgroup analysis according to pre-eclampsia severity revealed no significant differences in genotype or allele frequencies of the HIF-1α c.1772C>T and c.1790G>A polymorphism in mild pre-eclamptic compared to severe pre-eclamptic group. In addition, there were no significant differences in the frequencies of 3 haplotypes (C-G,-G, T-G, and C-A) between the control and pre-eclamptic groups. CONCLUSIONS: Our results suggest that the HIF-1α gene polymorphisms are not associated with the development of pre-eclampsia in the studied Korean women population.

Pregnancy outcome after 1st-trimester inadvertent exposure to barium sulphate as a contrast media for upper gastrointestinal tract radiography.

Despite barium being used as a contrast media for decades, the specific assessment of its safety in pregnant women is scarce. We are reporting the favourable pregnancy outcome in women who were inadvertently exposed to barium swallow and associated ionising radiation, early in pregnancy. A control group of age- and gravidity-matched unexposed pregnant women was also included. There were 32 live-born babies in the exposed group and 94 in the control group. Women had undergone diagnostic upper gastrointestinal tract (UGT) fluoroscopic examination at 3.3 ± 1.5 weeks' gestation. Estimated maternal radiation dose secondary to barium swallow varied widely, the maximum dose was estimated to be 2.45 mSv. Similar pregnancy outcomes were observed between the groups. The number of babies born with major malformations was not significantly different (p = 1.0) between cases and controls: one (3.1%) vs three (3.2%), respectively. In conclusion, our small prospective cohort study of women suggests no association between inadvertent exposure to ionising radiation and barium sulphate during fluoroscopic barium swallow and adverse fetal outcomes.

Interleukin-18, transforming growth factor-ß, and vascular endothelial growth factor gene polymorphisms and susceptibility to primary glomerulonephritis.

Several studies have showed an association of gene polymorphisms with the development of glomerulonephritis (GN). We investigated the effects of gene polymorphisms on the development of GN by analyzing polymorphisms in the interleukin (IL)-18, transforming growth factor (TGF)-ß, and vascular endothelial growth factor (VEGF) genes in Korean patients with primary GN. The study included 146 normal subjects (controls) and 100 patients diagnosed with primary GN by kidney biopsy. The gene polymorphisms A-607C and G-137C in IL-18, C-509T and T869C in TGF-ß1, and C-2578A and C405G in VEGF were investigated in DNA extracted from peripheral blood. Significant differences were observed between the GN and control groups in the genotype and allele frequencies of A-607C IL-18 and C405G VEGF. The frequencies of the IL-18-607CC genotype [P = 0.001, odds ratio (OR) = 2.473] and the VEGF 405GG genotype (P = 0.001, OR = 2.473) were significantly increased in the GN group. The combination of IL-18-607CC+ and VEGF 405GG+ genotypes had a higher risk for developing GN in comparison with the combination of IL-18-607CC- and VEGF 405GG- genotypes (P < 0.001, OR = 8.642). In the haplotype analysis of the IL-18 gene, the CG haplotype was significantly more frequent in the GN group than the control group (61.5% vs 46.9%, P = 0.002). These results show that the -607CC genotype of the IL-18 gene and the 405GG genotype of the VEGF gene are associated with susceptibility to and the development of primary GN.

Prenatal multivitamins containing folic acid do not decrease prevalence of depression among pregnant women.

Several studies have reported an association between depression and folic acid deficiency. We investigated whether intake of prenatal multivitamins containing folic acid (MVandFA) was associated with decreased rates of depression among pregnant women. A questionnaire was given to 1,314 low-risk pregnant women. Of them, 1,277 (97.2%) women completed the questionnaire. The overall prevalence of depression was 8.1%. Of 652 participants who did not take MVandFA, 9.4% had depression, whereas 6.9% of 624 women who had MVandFA had depression (p = 0.11). In a multivariate logistic regression analysis, family history of depression (adjusted OR 3.7; 95% CI 1.9-7.3) and premenstrual syndrome (adjusted OR 3.0, 95% CI 1.8-4.8) were identified as risk factors for depression during pregnancy. In conclusion, intake of MVandFA was not associated with lower rates of depression during pregnancy whereas family history of depression and personal history of premenstrual syndrome were significant risk factors.

The 769G>A variant of the inhibin-alpha gene in Korean patients with preeclampsia.

OBJECTIVE: Increased inhibin A serum and plasma levels in the second trimester are significantly associated with the development of preeclampsia. The measurement of inhibin A during early pregnancies may be helpful to predict those at risk of this disorder. The purpose of this study was to determine whether the 769G>A variant of the inhibin alpha (INHalpha) gene was associated with preeclampsia. PATIENTS AND METHODS: We screened the 769G>A variation in 162 preeclamptic patients and in 202 normal pregnancies. The 769G>A variant of the INHalpha gene was determined by the PCR-based restriction fragment length polymorphism analysis and DNA sequencing. RESULTS: We found no variation between the normal subjects and the preeclamptic patients. CONCLUSION: The 769G>A variant of the INHalpha gene may be rare in Korean patients with preeclampsia.

Pregnancy outcome after exposure to oral contraceptives during the periconceptional period.

To evaluate whether periconceptional exposure to oral contraceptives (OCs) increased adverse pregnancy outcomes, 136 pregnant women taking OCs within the periconceptional period were identified at the Korean Motherisk Program. Of them, 120 pregnant women accepted to participate in their study and were followed up until completion of the pregnancy. A control group of 240 age- and gravidity-matched pregnant women exposed to non-teratogen drugs for at least 1 month before pregnancy was also included. The median gestational age at delivery was 39.1 (27.0-41.0) weeks in the exposed group and 39.3 (27.4-42.0) weeks in the control group (P = 0.19). In the exposed group, 7.1% of babies were born with low birth weight versus 2.6% in the control group (P = 0.068). The number of preterm deliveries or babies born large for gestational age did not differ between the two groups. In the exposed group, the rate of birth defects was 3.2% (n = 3/99) versus 3.6% (n = 7/193) in the control group (P = 1.0). There were 15 women who took high doses of progesterone (emergency contraception) and no adverse fetal outcomes were observed. In conclusion, periconceptional exposure to OCs does not appear to increase the risk for adverse pregnancy outcomes.

Teratogen risk counselling by internet: a prospective cohort study.

Although the internet may play a role in providing proper teratogen-risk counselling for pregnant women, the experience with this type of service has not been reported. We aimed to compare the pregnancy outcome of women counselled by the internet to women that received typically in-clinic teratogen risk counselling in the clinic. In a prospective cohort design, 1,011 patients were counselled in the clinic and 235 patients were counselled via the internet. Teratogen risk counselling was provided according with the information obtained from medical literature and specialised software. Information about pregnancy outcome (delivery, spontaneous abortion, or termination of pregnancy and major malformations) was obtained from 903 (89.3%) patients from the clinic group and 141 (60%) from the internet group. The amount of alcohol, cigarettes smoked, and millirads of X-ray were greater ( p < 0.05) in patients counselled by internet than in clinic. The rate of deliveries, ongoing pregnancies, pregnancy terminations, and spontaneous abortions were similar between clinic and internet (chi2 = 1.32; p = 0.7). Of 498 babies born to mothers counselled in the clinic, major malformations were identified in 3.6%. Of 67 babies born to mothers counselled by internet, major malformations were present in 1.5% ( p = 0.6). Teratogen-risk counselling by internet and clinic seems to have a similar efficacy in pregnancy outcome.

Molecular cytogenetic characterization of ring chromosome 4 in a female having a chromosomally normal child.

We present the clinical and molecular findings of mosaic ring chromosome 4. The patient was referred to us for infertility and short stature. Results of three repeated cytogenetic analyses from lymphocytes showed a similar mosaic karyotype with multiple cell-lines [46,XX,r(4)/45,XX,-4/46,XX,dic r(4)/47,XX,r(4),+r(4)/46,XX]. FISH showed deletion of the 4p subtelomeric region and the 4q telomeric region from the ring chromosome 4. The breakpoints were mapped using molecular analysis. Parental karyotypes were normal. During the course of this study, the patient became pregnant without assisted reproductive technology. The result of amniocentesis performed at 16 weeks gestation showed a normal karyotype. Delivery was uncomplicated. This is the first report, to our knowledge, of the presence of ring chromosome 4 having various mosaic conditions in a female having a chromosomally normal fetus.

Identification of new dinucleotide-repeat polymorphisms in factor VIII gene using fluorescent PCR.

Haemophilia A is an X-linked inherited bleeding disorder. Linkage diagnosis using polymorphic markers in the factor VIII gene is used to archive the carrier detection and prenatal diagnosis. The objective of this study was to establish the allele frequency and heterozygosity rate (HR) of two new intragenic markers (Intron 1 and 24) and other markers (Intron 13 and 22) using fluorescent PCR. Five hundred unrelated healthy women were screened and haemophilic family was studied for carrier detection and prenatal diagnosis. We observed five different alleles of Intron 1, 10 of Intron 24, nine of Intron 13 and six of Intron 22. The observed HR for Intron 1, 24, 13 and 22 were 34.0, 35.2, 53.0 and 42.6%, while the expected HR were 33.6, 36.3, 50.1 and 44.3%, respectively. Heterozygosity rate with the combined use of all four intragenic markers was 76.6% (383/500). In prenatal diagnosis of a haemophilic family, a pregnant woman was heterozygous with three intragenic (Intron 1, 13 and 22) and one extragenic St14 VNTR (DXS52) markers. She was considered to be a carrier, and she carried a male foetus by AMXY PCR and chromosome analysis of amniocytes. Foetus did not have mutant haplotype as his uncle, suggesting a normal male status. Our study demonstrates the utility of two new intragenic markers in FVIII gene for carrier detection and prenatal diagnosis of haemophilic families.

Mesenchymal progenitor cells in the human umbilical cord.

Mesenchymal progenitor or stem cells (MPCs) isolated from fetal blood, liver, and bone marrow are a population of multipotential cells that can proliferate and differentiate into multiple mesodermal tissues including bone, cartilage, muscle, ligament, tendon, fat, and stroma. The objective of this study was to isolate and characterize MPCs in the human umbilical cord. The suspensions of endothelial and subendothelial cells in cord vein were collected and cultured in M199 supplemented with 10% fetal bovine serum (FBS). Of 50 umbilical cord samples, 3 had numerous fibroblastoid cells morphologically distinguishable from endothelial cells. Fibroblastic cells displayed lack of expression of vWF, Flk-1, and PECAM-1, indicating the endothelial cell-specific marker. To investigate the differentiation potentials, the cells were cultured in adipogenic or osteogenic medium for 2 weeks. Fibroblast-like cells treated with adipogenic supplementation showed Oil red O-positive staining and expressed adipsin, FABP4, LPL, and PPARgamma2 genes by reverse transcriptase polymerase chain reaction (RT-PCR). In osteogenic differentiation, alkaline phosphatase activity and calcium accumulation were detected. RT-PCR studies determined that Cx43, osteopontin, and Runx2 genes were expressed in the osteogenic cultures. Among three cell lines cultured continuously for passage 10, two had normal karyotypes; however, one retained a karyotype of mos 46,XY[19]/47,XY,+mar[3]. These observations suggest that MPCs are present in human umbilical cord and possess several typical traits of MPCs.

Prenatal diagnosis of a familial complex chromosomal rearrangement involving chromosomes 5, 10, 16 and 18.

We report one case of a familial complex chromosomal rearrangement (CCR) involving four different chromosomes 5, 10, 16 and 18. The CCR was detected prenatally at 20 weeks' gestation because of advanced maternal age and history of recurrent miscarriages. Cytogenetic analysis of cultured amniotic fluid cells with GTG banding showed a 46,XX,t(5;16;10;18)(q13;q22;q11.2;q21) karyotype. Parental cytogenetic study revealed that the mother has the same CCR. RBG banding, high resolution banding and fluorescence in situ hybridization (FISH) were used to characterize further and confirm the conventional banding data. No physical abnormalities were shown in the targeted fetal ultrasonography examination. The parents decided to continue the pregnancy. The child is now 2 years old and has neither congenital anomalies nor evidence of delayed psychomotor development. The fetal targeted ultrasound and FISH analysis helped us reassure fetal status.

Increased chromosome X, Y, and 18 nondisjunction in sperm from infertile patients that were identified as normal by strict morphology: implication for intracytoplasmic sperm injection.

OBJECTIVE: To determine the incidence of nondisjunction for chromosomes X, Y, and 18 using fluorescence in situ hybridization (FISH) on morphologically normal sperm from infertile men who are candidates for ICSI. DESIGN: After standard hematoxylin staining, sperm with normal morphology were identified using Kruger's strict morphology criteria. The location of each normal-appearing sperm was recorded using an electronic microstage locator. Slides were subsequently subjected to FISH for detection of chromosomes X, Y, and 18 (control probe). Nuclei were relocated and analyzed under the fluorescent microscope. SETTING: University-affiliated IVF and intracytoplasmic sperm injection program. PATIENT(S): Men classified as infertile on the basis of abnormal strict morphology (<4% by Kruger's criteria). For controls, normal fertile men (n=6) were also analyzed. INTERVENTION(S): Semen smears were obtained retrospectively from infertile (n=8) and fertile (n=6) men. MAIN OUTCOME MEASURE(S): Ploidy of each cell was determined according to the number of signals detected for each probe. RESULT(S): Approximately 100-150 morphologically normal sperm were identified and located in each case. Subsequent FISH analysis of these normal sperm showed aneuploidy to range from 1.8% to 5.5% in the infertile group as compared with 0 to 2.6% among the control fertile group. Statistically significant differences in the incidence of aneuploidy for the sex chromosomes as well as for all three (X, Y, and 18) chromosomes was observed. CONCLUSION(S): Although 95% to 98% of the sperm were found to be normal for X, Y, and 18, our findings show that infertile couples undergoing ICSI are likely to be at an increased risk for having a genetically abnormal conceptus as compared with the fertile controls. These results demonstrate that normal morphology is not an absolute indicator for the selection of genetically normal sperm. Hence, observed pregnancy failures among ICSI patients may in part be due to the selection of aneuploid sperm.