Paternal Blood Research Articles

The Born in Guangzhou Cohort Study (BIGCS).

The Born in Guangzhou Cohort Study (BIGCS) is a large-scale prospective observational study investigating the role of social, biological and environmental influences on pregnancy and child health and development in an urban setting in southern China. Pregnant women who reside in Guangzhou and who attend Guangzhou Women and Children's Medical Center (GWCMC) for antenatal care in early pregnancy (<20weeks' gestation) are eligible for inclusion. Study recruitment commenced in February 2012, with an overall participation rate of 76.3%. Study recruitment will continue until December 2018 to achieve the target sample size of 30,000 mother-child pairs. At 30 April 2016, a total of 75,422 questionnaires have been collected, while 14,696 live births have occurred with planned follow-up of cohort children until age 18years. During the same period a total of 1,053,000 biological samples have been collected from participants, including maternal, paternal and infant blood, cord blood, placenta, umbilical cord, and maternal and infant stool samples. The dataset has been enhanced by record linkage to routine health and administrative records. We plan future record linkage to school enrolment and national examination records.

Influence of Maternal Gestational Hypertensive Disorders on Microvasculature in School-Age Children: The Generation R Study.

Gestational hypertensive disorders may lead to vascular changes in the offspring. We examined the associations of maternal blood pressure development and hypertensive disorders during pregnancy with microvasculature adaptations in the offspring in childhood. This study was performed as part of the Generation R Study in Rotterdam, the Netherlands (2002-2012), among 3,748 pregnant mothers and their children for whom information was available on maternal blood pressure in different periods of pregnancy and gestational hypertensive disorders. Childhood retinal arteriolar and venular calibers were assessed at the age of 6 years. We found that higher maternal systolic and diastolic blood pressures in early pregnancy were associated with childhood retinal arteriolar narrowing (P < 0.05). Higher maternal systolic blood pressure in late pregnancy, but not in middle pregnancy, was associated with childhood narrower retinal venular caliber (standard deviation score per standardized residual increase in systolic blood pressure: -0.05; 95% confidence interval: -0.08, -0.01). Paternal blood pressure was not associated with childhood retinal vessel calibers. Children of mothers with gestational hypertensive disorders tended to have narrower retinal arteriolar caliber (standard deviation score: -0.13, 95% confidence interval: -0.27, 0.01). Our results suggest that higher maternal blood pressure during pregnancy is associated with persistent microvasculature adaptations in their children. Further studies are needed to replicate these observations.

Associations of Maternal and Paternal Blood Pressure Patterns and Hypertensive Disorders during Pregnancy with Childhood Blood Pressure.

BackgroundHypertensive disorders in pregnancy may affect the cardiovascular risk of offspring. We examined the associations of maternal blood pressure throughout pregnancy and hypertensive disorders in pregnancy with childhood blood pressure of offspring. Specific focus was on the comparison with paternal blood pressure effects, the identification of critical periods, and the role of birth outcomes and childhood body mass index in the observed associations.Methods and ResultsThis study was embedded in a population‐based prospective cohort study among 5310 mothers and fathers and their children. We measured maternal blood pressure in each trimester of pregnancy and paternal blood pressure once. Information about hypertensive disorders in pregnancy was obtained from medical records. We measured childhood blood pressure at the median age of 6.0 years (95% range 5.7–8.0 years). Both maternal and paternal blood pressure were positively associated with childhood blood pressure (all P<0.05), with similar effect estimates. Conditional regression analyses showed that early, mid‐, and late‐pregnancy maternal blood pressure levels were all independent and positively associated with childhood blood pressure, with the strongest effect estimates for early pregnancy. Compared with children of mothers without hypertensive disorders in pregnancy, children of mothers with hypertensive disorders in pregnancy had higher diastolic blood pressure by a standard deviation score of 0.13 (95% CI 0.05–0.21). The observed associations were not materially affected by birth outcomes and childhood body mass index.ConclusionsBoth maternal and paternal blood pressure affects childhood blood pressure, independent of fetal and childhood growth measures, with the strongest effect of maternal blood pressure in early pregnancy.

LB03.02

Objective: To investigate the association between paternal circulatory health in middle age and offspring blood pressure across childhood and adolescence in order to examine the importance of inter-generational influences on blood pressure trajectories and the potential development of hypertension in later adult life. Design and method: We used data from 1,969 paternal-offspring pairs recruited as part of the Avon Longitudinal Study of Parents and Children (ALSPAC) birth cohort. Paternal pressures and arterial stiffness (systolic blood pressure (SBP), diastolic blood pressure (DBP), pulse wave velocity (PWV) and augmentation index (AI)) were measured at the Focus on Fathers clinic between 2011 to 2013. This was linked to 6 prior measures of offspring SBP and DBP taken at research clinics when the ALSPAC participants were approximately 7, 9, 10, 11, 15 and 17 years old. We used both multivariable linear regression and 2 level linear-spline models to model the associations between a one standard deviation increase (z-score) in paternal circulatory measures and offspring SBP and DBP at each time point as well as the intercept (BP at age 7) and rate of blood pressure change from linear slopes between ages 7–11, 11–15 and 15–17 from our multi-level model. Results: A positive association was found between paternal SBP and male offspring systolic and diastolic blood pressures in simple and adiposity adjusted analyses (SBP +1.48 mmHg; CI 0.82 - 2.14 mmHg, p < 0.0001 at age 7 for 1 z-score increase in paternal SBP). These associations remained similar throughout adolescence. No associations were seen with paternal PWV or AI and male offspring pressures. Far weaker or null associations were seen for female blood pressures (p-value for gender interaction=0.01 at 7 years). Conclusions: This study demonstrates sexual dimorphism for associations between paternal and offspring blood pressures which require replication from other studies. The associations were strongest in the pre-pubertal period and, if anything, weakened with adolescence suggesting other environmental factors may be more important in determining age-related increases in blood pressure. Future follow-up is required to test whether associations reappear in later life.

Birth outcomes and background exposures to select elements, the Longitudinal Investigation of Fertility and the Environment (LIFE)

Evidence suggests that trace exposures to select elements may increase the risk for adverse birth outcomes. To investigate further, we used multiple regression to assess associations between preconception parental exposures to Pb, Cd, and total Hg in blood, and 21 elements in urine, with n=235 singleton birth outcomes, adjusted for confounders and partner's exposure. Earlier gestational age at delivery (GA) was associated with higher tertiles of urine maternal W (−1.22 days) and paternal U (−1.07 days), but GA was later for higher tertiles of maternal (+1.11 days) and paternal (+1.30 days) blood Hg. Additional analysis indicated shorter GA associated with higher paternal urine Ba, W, and U, and with higher maternal blood Pb for boys, but GA was longer in association with higher maternal urine Cr. Birth weight (BW) was lower for higher tertiles of paternal urine Cs (−237.85g), U (−187.34g), and Zn (−209.08g), and for higher continuous Cr (P=0.021). In contrast, BW was higher for higher tertiles of paternal urine As (+194.71g) and counterintuitively for maternal blood Cd (+178.52g). Birth length (BL) was shorter for higher tertiles of urine maternal W (−1.22cm) and paternal U (−1.10cm). Yet, higher tertiles of maternal (+1.11cm) and paternal (+1.30) blood Hg were associated with longer BL. Head circumference at delivery was lower for higher tertiles of paternal urine U (−0.83cm), and for higher continuous Mo in boys (−0.57cm). Overall, associations were most consistently indicated for GA and measures of birth size with urine W and U, and paternal exposures were more frequently associated than maternal. Though limited by several factors, ours is the largest multi-element investigation of prospective couple-level trace exposures and birth outcomes to date; the novel observations for W and U merit further investigation.

Autosomal dominant polycystic kidney disease caused by somatic and germline mosaicism

Autosomal dominant polycystic kidney disease (ADPKD) is a heterogeneous genetic disorder caused by loss of function mutations of PKD1 or PKD2 genes. Although PKD1 is highly polymorphic and the new mutation rate is relatively high, the role of mosaicism is incompletely defined. Herein, we describe the molecular analysis of ADPKD in a 19-year-old female proband and her father. The proband had a PKD1 truncation mutation c.10745dupC (p.Val3584ArgfsX43), which was absent in paternal peripheral blood lymphocytes (PBL). However, very low quantities of this mutation were detected in the father's sperm DNA, but not in DNA from his buccal cells or urine sediment. Next generation sequencing (NGS) analysis determined the level of this mutation in the father's PBL, buccal cells and sperm to be ∼3%, 4.5% and 10%, respectively, consistent with somatic and germline mosaicism. The PKD1 mutation in ∼10% of her father's sperm indicates that it probably occurred early in embryogenesis. In ADPKD cases where a de novo mutation is suspected because of negative PKD gene testing of PBL, additional evaluation with more sensitive methods (e.g. NGS) of the proband PBL and paternal sperm can enhance detection of mosaicism and facilitate genetic counseling.

Newborns and low to moderate prenatal environmental lead exposure: might fathers be the key?

This study is part of the BioMadrid Project, a bio-monitoring study designed to assess pollutants in the environment surrounding children born in the Madrid region. Our aim in this report is to evaluate the association between prenatal lead exposure and fetal development using three biological samples (maternal and paternal blood lead at around 34 weeks of gestation as well as cord blood lead levels), three biomarkers of effect in cord blood peripheral lymphocytes (micronucleus in binucleated cells, nucleoplasmic bridges, and nuclear buds), and different anthropometrical characteristics at birth. Maternal and cord blood lead were not associated with newborn measurements or genotoxicity biomarkers. In contrast, increases in father blood lead were coupled with lower weight (mean difference (MD), -110.8 g; 95% confidence intervals (95%CI), -235.6 to 6.00; p < 0.10) and shorter abdominal (MD, -0.81 cm; 95%CI, -1.64 to 0.00; p < 0.05) and cephalic (MD, -0.32 cm; 95%CI, -0.65 to 0.00; p < 0.05) circumferences at birth as well as with the presence of nucleoplasmic bridges (odds ratio, 1.03; 95%CI, 1.00 to 1.06; p < 0.05) and nuclear buds (odds ratio, 1.02; 95%CI, 0.99 to 1.04; p < 0.10). These associations were mainly confined to female babies, in whom paternal lead was also inversely associated with length. Our results support the hypothesis that paternal lead exposure may be affecting the development of newborns.

Role Of Maternal Immunization Against Low Frequency Human Platelet Antigens (LFHPA) In Neonatal Alloimmune Thrombocytopenia (NAIT)

Neonatal alloimmune thrombocytopenia (NAIT) is largely caused by maternal immunization against Human Platelet Antigens (HPA) inherited by a fetus from its father. In about 70% of apparent NAIT cases, however, maternal HPA antibodies are not detected by standard serologic testing. At least 24 low frequency HPAs (LFHPA) have now been implicated in NAIT. Fathers are not ordinarily typed for these antigens and the corresponding antibodies can be difficult to detect. We studied paternal and maternal blood samples from suspected NAIT cases to investigate the extent to which maternal immunization against LFHPAs causes “antibody-negative” NAIT.In Phase 1, we studied paternal DNA in 244 NAIT cases not resolved by standard laboratory testing in which there was compelling clinical and/or serologic evidence for NAIT. LFHPAs were identified by full length sequencing of relevant exons encoding platelet glycoproteins (GP). Sixteen LFHPAs (6.6% of cases) were identified. HPA-4b, 6bw, 11bw, 12bw, 13bw, 19bw, 20bw, 21bw, 22bw, and 23bw were found in single cases and HPA-9bw in 6 cases. In 15 of the 16 cases, a maternal antibody reactive with paternal platelets and/or a platelet GP was identified.Cases studied in Phase 1 were highly selected. To obtain an unbiased estimate of the extent to which LFHPAs trigger NAIT, we determined the prevalence of 22 recognized LFHPAs in fathers of 1067 consecutive “antibody-negative” NAIT cases using high throughput screening methods (Phase 2). Mothers of 1338 cases were similarly typed to assess the normal prevalence of LFHPAs in a population racially/ethnically similar to the fathers. Of the 1067 fathers, 28 (2.6%) were positive for a LFHPA. LFHPAs identified were HPA-4b (1 case), 6bw (2), 8bw (1), 9bw (17), 12bw (4) and 21bw (3). Of the 1338 mothers, 27 (2.0%) were positive for a LFHPA. Maternal LFPAs identified were 4b (2 cases), 6bw (5), 9bw (12), 12bw (4), 13bw (3) and 21bw (1). Of the LFHPAs identified, only HPA-9bw was significantly more common in fathers than in mothers (P=0.02). Serologic evaluations done in the Phase 2 cases provided evidence for maternal antibodies against paternal GPIIb/IIIa in only 2 of 23 instances in which the paternal low frequency antigen was located on this GP complex.Findings made in Phase 1, show that efforts to identify a paternal LFHPA in unresolved NAIT cases can be rewarding when maternal serum reacts with paternal platelets and the clinical picture strongly suggests NAIT. The Phase 2 studies of consecutive unresolved NAIT cases show that, with the exception of HPA-9b, the prevalence of LFHPAs is about the same in mothers and fathers of apparent NAIT cases not attributable to maternal immunization against “common” HPAs. It is clear that maternal immunization against an LFHPA can account for only a small fraction of the apparent “diagnostic gap” in serologic diagnosis of NAIT. HPA-9bw is by far the most important immunogen among the recognized LFHPAs and consideration should be given to typing fathers of suspected NAIT cases routinely for this antigen. Recent studies suggest that “low avidity” HPA antibodies not detected using conventional serology may account for an unknown fraction of the “antibody-negative” NAIT cases. Whether HLA antibodies cause NAIT more often than has been thought also deserves re-examination. Disclosures:No relevant conflicts of interest to declare.

In utero Hematopoietic Stem Cell Transplantation in Canines: Exploring the Gestational Age Window of Opportunity to Maximize Engraftment

Objective: In utero hematopoietic stem cell transplantation (IUHSCT) is a promising therapy for a variety of congenital disorders. Our objective was to determine the optimal time in gestation for IUHSCT in a canine model. Methods: IUHSCT was performed in day 31-50 (term 63) fetal canines with CD34+ cells isolated from paternal bone marrow at doses of 0.09-3.4 × 10⁹ CD34+ cells/kg and T cells (CD3+/CD5+) from paternal blood at 0.11-1.1 × 10⁹ cells/kg. Engraftment was assayed using PCR-based chimerism analysis (SRY gene detection for female recipients, and unique microsatellite loci for both sexes). Results: Microchimerism and chimerism were present in multiple recipients across most gestational ages at transplant. Maximal engraftment was obtained in hematopoietic tissues in transplants performed at 42 days. At extremes of recipient gestational age, minimal to no engraftment was seen. Conclusion: Fetal age at the time of IUHSCT plays an important role in achieving engraftment in our canine model.

Cigarette smoke‐induced transgenerational alterations in genome stability in cord blood of human F1 offspring

The relevance of preconceptional and prenatal toxicant exposures for genomic stability in offspring is difficult to analyze in human populations, because gestational exposures usually cannot be separated from preconceptional exposures. To analyze the roles of exposures during gestation and conception on genomic stability in the offspring, stability was assessed via the Comet assay and highly sensitive, semiautomated confocal laser scans of γH2AX foci in cord, maternal, and paternal blood as well as spermatozoa from 39 families in Crete, Greece, and the United Kingdom. With use of multivariate linear regression analysis with backward selection, preconceptional paternal smoking (% tail DNA: P>0.032; γH2AX foci: P>0.018) and gestational maternal (% tail DNA: P>0.033) smoking were found to statistically significantly predict DNA damage in the cord blood of F1 offspring. Maternal passive smoke exposure was not identified as a predictor of DNA damage in cord blood, indicating that the effect of paternal smoking may be transmitted via the spermatozoal genome. Taken together, these studies reveal a role for cigarette smoke in the induction of DNA alterations in human F1 offspring via exposures of the fetus in utero or the paternal germline. Moreover, the identification of transgenerational DNA alterations in the unexposed F1 offspring of smoking-exposed fathers supports the claim that cigarette smoke is a human germ cell mutagen.

Evaluation of single‐nucleotide polymorphisms as internal controls in prenatal diagnosis of fetal blood groups

Determination of fetal blood groups in maternal plasma samples critically depends on adequate amplification of fetal DNA. We evaluated the routine inclusion of 52 single-nucleotide polymorphisms (SNPs) as internal reference in our polymerase chain reaction (PCR) settings to obtain a positive internal control for fetal DNA. DNA from 223 plasma samples of pregnant women was screened for RHD Exons 3, 4, 5, and 7 in a multiplex PCR including 52 SNPs divided into four primer pools. Amplicons were analyzed by single-base extension and the GeneScan method in a genetic analyzer. Results of D screening were compared to standard RHD genotyping of amniotic fluid or real-time PCR of fetal DNA from maternal plasma. The vast majority of all samples (97.8%) demonstrated differences in maternal and fetal SNP patterns when tested with four primer pools. These differences were not observed in less than 2.2% of the samples most probably due to an extraction failure for adequate amounts of fetal DNA. Comparison of the fetal genotypes with independent results did not reveal a single false-negative case among samples (n = 42) with positive internal control and negative fetal RHD typing. Coamplification of 52 SNPs with RHD-specific sequences for fetal blood group determination introduces a valid positive control for the amplification of fetal DNA to avoid false-negative results. This new approach does not require a paternal blood sample. It may also be applicable to other assays for fetal genotyping in maternal blood samples.

Multiplex PCR for 17 Y-Chromosome Specific Short Tandem Repeats (STR) to Enhance the Reliability of Fetal Sex Determination in Maternal Plasma

The aim of the study was to demonstrate the influence of target gene and amplification product length on the performance of fetal gender determination systems using maternal plasma. A total of 40 pairs of plasma DNA samples from pregnant women and genomic DNA samples from maternal blood, amniotic fluid and paternal blood were isolated for gender determination by amplification of the amelogenin gene and 17 Y-chromosome STR loci, using three different commercial kits. The gender of the fetuses was confirmed by cytogenetic analysis or phenotype at birth. Both the AmpFℓSTR-Identifiler amplification kit and the Mini-STR Amplification kit for amelogenin gene detection were reliable in determining fetal gender (92.0% and 96.0%, respectively), but false negatives were present in both systems. AmpFℓSTR-Yfiler was found to be fully reliable as it amplified Y-STR in all cases of pregnancies with male fetuses and thus was 100% correct in determining fetal gender. The results demonstrated that multiple fluorescent PCR for 17 Y-STR loci was more reliable than AMELY gene testing in fetal sex determination with maternal plasma. We also found that the shorter amplification products could improve the performance of fetal gender determination systems.

Offspring Birth Weight and Cardiovascular Risk in Parents—A Population-based HUNT 2 Study

Low birth weight is associated with increased risk of cardiovascular disease and type 2 diabetes in later life. The fetal insulin hypothesis suggests that shared genetic factors partly explain this association. If fetal genes predispose to both low birth weight and cardiovascular disease in adulthood, fathers of offspring with low birth weight should display an unfavorable profile of cardiovascular risk factors. To study this, the authors linked data on more than 14,000 parents, collected from the second Health Study of Nord Trøndelag County, Norway (HUNT 2, 1995-1997), to offspring data from the Norwegian Medical Birth Registry (1967-2005). Linear regression was used to study associations of offspring birth weight for gestational age with the parents' body mass index, waist circumference, blood pressure, glucose, and serum lipids. All analyses were adjusted for shared environment by means of the socioeconomic measures, lifestyle, and cardiovascular risk factors of the partner. The authors found that low offspring birth weight for gestational age was associated with increased paternal blood pressure, body mass index, waist circumference, and unfavorable levels of glucose and lipids. For mothers, associations similar to those for fathers were found for blood pressure, whereas associations in the opposite direction were found for glucose, lipids, and body mass index. The paternal findings strengthen the genetic hypothesis.

Implementing non-invasive RHD genotyping on cell-free foetal DNA from maternal plasma: the Pavia experience.

The occurrence of cell-free foetal DNA in maternal circulation opens new possibilities in non-invasive antenatal diagnosis. Real-time polymerase chain reaction (PCR) analysis is an useful approach to foetal RhD blood group determination, thus being important in the prevention of haemolytic disease of foetus and new-born (HDFN). Using real-time PCR assays we typed 20 samples of plasma, provided in a blinded fashion, from the International Reference Laboratory, two plasma samples sent by the "2010 Workshop on Molecular Blood Group Genotyping"; seven samples from D-negative mothers at the 16th week of gestation provided by our Hospital as prospective validation cases, and two plasma samples received from the "1(st) Collaborative study establishing the sensitivity standard for non-invasive prenatal determination of foetal RHD genotype". To confirm the RHD typing of the seven prospective samples, PCR with sequence specific primers (PCR-SSP) was applied on genomic DNA from amniocytes (5 cases) and paternal peripheral blood (2 cases). The results for the 31 investigated samples showed 100% concordance. Our measurable benefits were: confidence with a new technology, awareness of having gained the European standard level and increased self-assurance regarding the introduction of this typing technique in prenatal diagnostics. These results demonstrate the feasibility and accuracy of our validation protocol. RHD typing on cell-free foetal DNA is a procedure which requires particular care and a great degree of expertise for diagnostic use. International collaborations are essential for monitoring the performance of laboratories in the absence of specific quality control programmes.

Paternal Contribution to Small for Gestational Age Babies: A Multicenter Prospective Study

Our aims were to investigate whether men who fathered small for gestational age (SGA) infants themselves had lower birthweight, were more likely to be obese, have central adiposity and elevated blood pressure in adult life compared with men who fathered non-SGA infants. A total of 2,002 couples participating in the Screening for Pregnancy Endpoints (SCOPE) study were enrolled in early pregnancy and pregnancy outcome data collected prospectively. SGA was defined as birthweight <10th customized centile, obesity as BMI ≥30 kg/m(2), central adiposity as waist circumference >102 cm. Logistic regression was used to compare rates of obesity, and central adiposity between men who fathered SGA infants compared with those with non-SGA infants and the final model was adjusted for maternal and paternal confounders. The men who fathered an SGA infant (209 (10.4%)) themselves had lower mean birthweight (3,291 (530) g vs. 3,472 (584) g, P < 0.0001), were more likely to be obese (50 (24.8%) vs. 321 (18.3%), adjusted odds ratio (OR) 1.50, 95% confidence interval 1.05-2.16, adjusted for maternal and paternal factors) and to have central adiposity (52 (25.1%) vs. 341 (19.2%), adjusted OR 1.53, 95% confidence interval 1.06-2.20) compared with men who fathered a non-SGA infant. Elevated paternal blood pressure was not associated with SGA. In conclusion, we report a novel relationship between paternal obesity/central adiposity and birth of an SGA infant, which appears to be independent of maternal factors associated with fetal growth restriction.

Detection of fetal RHD pseudogene (RHDΨ) and hybrid RHD-CE-Ds from RHD-negative pregnant women with a free DNA fetal kit

Hemolytic disease of the newborn is a clinical condition in which maternal and paternal Rh blood group antigens are incompatible and the mother is negative for the antigen whereas the father is positive. Analysis of fetal cells recovered from maternal plasma can provide a highly sensitive prenatal diagnosis. The fetal RHD gene in plasma DNA is detected by real-time PCR amplification of two different segments of the RHD gene (exons 7 and 10). Each amplicon is revealed with specific probes. We examined 40 female blood samples to verify the specificity of RHD exons (7 and 10) amplified by real-time PCR. Thirty fetuses were predicted to be RHD-positive based on analysis of plasma DNA. Seven fetuses were predicted to be RHD-negative. One fetus was negative for RHD on exon 10, and positive for RHD on exon 7 (early gestation age); two fetuses were RHD-negative on exon 7, and RHD-positive on exon 10 (RHD-CE-D(s) or RHDΨ), indicative of a maternal RHD allele. We conclude that it is necessary to analyze at least two exon regions in the RHD gene.

Single nucleotide polymorphisms of folate metabolism pathway genes in central chinese families with neural tube defects

Objective To investigate the contribution of single nucleotide polymorphisms (SNP)variation in folate metabolism pathway genes and its interaction with environmental risk factors to the etiology of NTD. Methods In 275 families from central China, a total of 278 aborted fetal tissues or blood samples were collected from NTD individuals, 478 maternal and/or paternal blood samples were also obtained as controls. Folate supplementation, maternal diabetes mellitus and medication before pregnancy and during the first trimester of pregnancy were investigated. SNP analyses of all samples were performed by CEQ 8800. Case-parent control study and transmission/disequilibrium tests (TDT) were performed according to environmental cofactors stratification to evaluated 28 SNP in 12 folate pathway genes associated with human NTD. Results Only gene MTHFR rs1801133 was significantly associated with NTD, and synergistic effects of environmental risk factors (no folate supplementation and maternal diabetes) were shown on the occurrence of NTD. Linkage disequilibrium between BHMT rs3733890 and NTD existed in case of no folate supplementation,whereas the genotype alone did not contribute to the etiology of NTD. Other SNP were not significantly associated with NTD. Conclusions MTHFR rs1801133 is a risk factor of NTD, but BHMT rs3733890 is not an independent risk factor. Further investigations in folate and methionine cycle genes are requird in larger scale to enclose the interactions between gene and gene, or gene and environmental factors. Key words: Neural tube defects; Folic acid; Polymorphism,single nucleotide; Methylenetetrahydrofolate reductase (NADPH2); Betaine-homxysteine S-methyltransferase

Loss of Non-Inherited Maternal MHC and Materno-Fetal Transmission of p190 Type BCR-ABL Leukemia.

Abstract 980Poster Board I-2 Introduction:Materno-fetal transmission of malignant clone is one of the rare causes of leukemia in infants. It has been recorded some 20 times over the past 100 years but definitive, genetic evidence for a maternal origin has been lacking. Here we present a detailed genetic analysis of materno-fetal transmission in a case with p190 type BCR-ABL leukemia. Patients course:A mother was found to have p190 type BCR-ABL positive acute lymphoblastic leukemia (ALL) one month after delivery of a baby, and died during induction therapy by uncontrolled bacillus cereus encephalitis. Her baby developed p190 type BCR-ABL positive B-precursor lymphoblastic lymphoma (B-LBL stage3 jaw tumor and pleural effusion) at 11-month old. Non-Hodgkin's lymphoma type chemotherapy according to Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG) ALB-NHL03, followed by Hyper-CVAD imatinib therapy without stem cell transplantation were given to her, leading to complete remission for more than 31 months. Material and methods:We first cloned the BCR-ABL genomic breakpoint region from the infant's pleural effusion (PE). The breakpoint was designated as a fusion between BCR intron 1 and ABL intron1. Then, We conducted Q-PCR by using BCR-ABL genomic breakpoint specific primer and sequencing. Fifteen polymorphic STR markers were amplified in the paternal blood DNA and Patient's PE and PB DNA samples using the Powerplex-16 system. HLA serotyping was by microdroplet lymphocyte cytotoxity. Genotyping was carried out using a reversed SSO HLA DNA typing method. The infant jaw tumor was further analyzed by Affymetrix high resolusion (250K) SNP arrays. Results:Short tandem repeat profiles indicated that the infant jaw tumor was, unambiguously, of maternal origin. Amplification of BCR-ABL genomic breakpoints on maternal leukemic cells and infant lymphoma cells revealed that these leukemia clones shared the same unique BCR-ABL genomic sequence, indicating a shared, single cell origin. Positivity of BCR-ABL specific PCR on Guthrie card blood spot also indicated that the leukemic clone was present in the blood at birth. Additionally the infant, maternal-derived leukemic cells had uniparental disomy of chromosome 6p with a deletion of non-inherited maternal MHC allele (NIMA), suggesting a mechanism for immune evasion in the baby. Conclusions:This is the first case with definite evidence of in utero materno-fetal transmission of mother's leukemic cells. This case also suggests that materno-feto transfer of leukemic cells is more common than as reflected in the frequency of clinically diagnosed cases, where the immuno-surveillance, predominantly directed against major HLA locus antigens, may be the principal constraint. Disclosures:No relevant conflicts of interest to declare.

Osmoregulatory role of the paternal brood pouch for two Syngnathus species

Even though sea horses, sea dragons, and pipefishes of the teleost family Syngnathidae have been studied for over a century, the physiological significance of the paternal brood pouch is not well defined. Here we document the regulation of brood pouch osmolality during embryonic development for Syngnathus floridae and Syngnathus fuscus, particularly during the middle brood stages or around hatching. S. fuscus brood pouch osmolality was significantly lower than non-brooding conspecifics for all brood stages, but early and late stage S. floridae displayed pouch osmolality comparable to non-brooding males, suggesting brood pouch osmoregulation at these stages may be important for S. fuscus embryonic development. Quantification of Na +, K +, Ca 2+, and Mg 2+ in paternal blood plasma, pouch fluid, and embryos indicated that regulation of these ions contributes to pouch osmoregulation and furthermore, that ions in the brood pouch are most likely derived from the environment and not paternal blood. While both species displayed significant increases in dry mass and changes in embryonic ion concentrations during development, net uptake of paternally-derived ions was not documented. Overall, this examination furthers our understanding of syngnathid brood pouch physiology and offers insight into the evolution of paternal care in these fishes.

Diagnosis and management of non‐anti‐D red cell antibodies in pregnancy

Anti‐D prophylaxis has reduced the incidence of haemolytic disease of the newborn. A variety of non‐anti‐D red cell antibodies can cause a degree of neonatal haemolysis. The frequency of antibody testing should be individualised. Management of non‐anti‐D alloimmunisation should be aimed at minimising perinatal morbidity. Learning objectives: To understand the causes and risk factors for maternal non‐anti‐D antibodies. To learn when to initiate invasive testing in the antenatal period. To know when to deliver the baby to maximise perinatal outcome. To learn about which antibodies can cause fetal hydrops and intrauterine haemolysis. Ethical issues: When is it necessary to perform paternal blood tests to determine red cell antibody status? When should the mother be delivered in cases where the red cell antibody detected has a weak association with neonatal haemolysis? Please cite this article as: Gajjar K, Spencer C. Diagnosis and management of non‐anti‐D red cell antibodies in pregnancy. The Obstetrician &amp; Gynaecologist 2009;11:89–95.