Prenatal diagnosis of isovaleric acidaemia by enzyme and metabolite assay in the first and second trimesters

Isovaleric acidaemia (IVA) is caused by a deficiency of isovaleryl CoA dehydrogenase. The diagnosis can be established biochemically by the demonstration of increased levels of isovalerylglycine (IVG) and 3-hydroxyisovaleric acid in urine and by the deficiency of incorporation of radiolabel from [¹⁴C]isovaleric acid in macromolecules in cultured fibroblasts. This paper reports a consecutive series of 24 prenatal diagnoses in pregnancies at high risk, using both methods-metabolite and indirect enzyme assay. Affected fetuses were diagnosed in four pregnancies: three in the second trimester and one recent case in the first trimester. The latter represents the first reported case of a first-trimester diagnosis of IVA by direct analysis of chorionic villi. We also report the first demonstration of strongly accumulated IVG in the amniotic fluid in the 12th week of an affected pregnancy.     

Are low maternal estriol levels a predictor for pro-opiomelanocortin (POMC) deficiency caused by POMC mutation during pregnancy?

Isolated adrenocorticotropic hormone deficiency is a rare cause of adrenocortical insufficiency, especially in children, and may be an underestimated cause of neonatal death. Low estriol levels are usually correlated with compromised uteroplacental perfusion and associated with fetal death. A 30-years old woman applied for pregnancy follow-up. Ultrasonographic evaluation and karyotype of the fetus are normal. Low estriol level 0.34 MoM (% 0.24) was detected in maternal triple screening test. Amniocentesis was performed, and chromosomal disorders, steroid sulfatase deficiency, and Smith–Lemli–Opitz syndrome (SLOS) were excluded with karyotype, fluorescence in situ hybridization (FISH), and molecular analysis of SLOS, respectively. As their first child had pro-opiomelanocortin (POMC) deficiency, POMC gene analysis was performed from both amniotic fluid and ethylene diamine tetra aceticacid (EDTA) blood sample of affected previous child, and homozygote mutation was detected. Fetus is diagnosed as POMC deficiency. We are presenting this case to discuss possible relationship of low maternal E3 levels and fetal POMC deficiency. © 2013 John Wiley & Sons, Ltd.     

Prenatal diagnosis of pseudo arylsulphatase a deficiency

Prenatal diagnosis was performed on a pregnancy at risk for metachromatic leukodystrophy (MLD) in a family with the pseudo arylsulphatase A deficiency trait. Extracts of cultured amniotic fluid cells were deficient in arylsulphatase A indicating that the fetus was either affected with MLD or had the benign pseudodeficiency trait. In the cerebroside sulphate loading test, the at risk cells hydrolysed sulphatide like control cultured amniotic fluid cells implying that the fetus had pseudodeficiency. The pregnancy was carried to term and a male child was delivered. Placenta, urine and fibroblasts had very low activities of arysulphatase A. However, no sulphatide could be detected in urine and growing fibroblasts responded normally in the cerebroside sulphate loading test, suggesting pseudodeficiency. At 29 months, the infant is healthy and shows no stigmata of MLD. The prediction based on the results of the cerebroside sulphate loading test on cultured amniotic fluid cells appeared to be borne out.     

Prenatal diagnosis of congenital human cytomegalovirus infection

Fifteen fetuses at risk of congenital human cytomegalovirus (HCMV) infection underwent prenatal diagnosis at 16–30 weeks' gestation by a combination of amniocentesis and fetal blood sampling. HCMV was isolated from the amniotic fluid in six patients, but HCMV-specific IgM was detected in only three of them. Two of the nine neonates, who were delivered following a negative prenatal diagnosis, had congenital HCMV infection diagnosed by virus isolation in the urine. The interval from infection to prenatal testing was 3 and 4 weeks in the two false-negative cases and ⩾ 7 weeks in the true-positive cases. Although timely testing for HCMV infection allows the option of termination of pregnancy, it may be flawed by false-negative results.     

南方某幼儿园儿童尿液中菲单羟基代谢物浓度分析

为了解我国南方某城市儿童尿液中单羟基菲的含量,于2007年11月采集该市一幼儿园80名4-6岁儿童尿样。尿液经过酶解、固相萃取(SPE)富集、净化,采用高效液相色谱-荧光检测器检测,共检测出4种单羟基菲,且以9-羟基菲为主。除4-羟基菲的检出率为53.75％外,其余单羟基菲的检出率近乎100％。各单羟基菲的中位数分别为...     

Detection of Y chromosome-specific DNA in the plasma and urine of pregnant women using nested polymerase chain reaction

The present study was undertaken to evaluate a nested polymerase chain reaction (PCR) for detection of Y chromosome-specific fetal DNA in maternal plasma and urine of pregnant women during different gestational stages. DNA isolated from plasma and urine samples of 80 pregnant women (between 7 and 40 weeks' gestation) underwent amplification for Y chromosome-specific 198 bp DNA by nested PCR. The postpartum analysis of fetal gender showed that 55 women carried male and 25 female fetuses. Among the 55 women bearing male fetuses, Y chromosome-specific signals were detected in 53 (96%) plasma and 21 (38%) urine samples. Moreover, out of 25 women bearing female fetuses, 3 (12%) and 1 (4%) women had Y chromosome-specific signal in plasma and urine, respectively. Analysis of results with respect to gestational age revealed that there was no significant difference in the detection of Y chromosome-specific DNA between different trimesters in maternal plasma of women bearing male fetuses. These results showed that fetus-specific DNA was detected with high sensitivity (96%) and specificity (88%) in the maternal plasma by nested PCR, and therefore the method could be useful as a non-invasive procedure for fetal sex determination and prenatal diagnosis. Copyright © 2001 John Wiley & Sons, Ltd.     

Evaluation of a fetus at risk for dihydropteridine reductase deficiency by direct mutation analysis using denaturing gradient gel electrophoresis

Dihydropteridine reductase (DHPR) is an enzyme involved in the recycling of tetrahydrobiopterin (BH₄), which is an obligate co-factor of the aromatic amino acid hydroxylases. DHPR deficiency is a rare, autosomal recessive disorder caused by mutations in the QDPR gene. DHPR-deficient patients are diagnosed by a lack of response to a low phenylalanine diet and by severe neurological symptoms. Final diagnosis is made by measurements of neurotransmitters and pterin metabolites in cerebrospinal fluid (CSF) and urine, in addition to DHPR enzyme activity, which can be assessed in whole red blood cells. Treatment of DHPR deficiency can be difficult and the outcome is not always satisfying, even if all treatment strategies are followed. Therefore prenatal diagnosis is of great importance in affected families. Prenatal diagnosis is possible by measuring DHPR activity in different cell types but this is time consuming. More than 25 different mutations have to date been identified in the QDPR gene and direct identification of a mutation in a fetus would be easy and rapid. We have developed a method based on denaturing gradient gel electrophoresis (DGGE) for the analysis of the QDPR gene. The method is useful for rapid and simultaneous scanning of all exons and flanking intronic sequences of the QDPR gene. We describe the first prenatal diagnosis conducted using this method. Copyright © 2001 John Wiley & Sons, Ltd.     

Prenatal diagnosis of NADH:ubiquinone oxidoreductase deficiency

NADH:ubiquinone oxidoreductase (complex I of the mitochondrial respiratory chain) deficiency is a severe disorder with an often early fatal outcome. Prenatal diagnosis for complex I defects currently relies mainly on biochemical assays of complex I in fetal tissues such as chorionic villi (CV), and is only in a minority of cases possible by means of mutational analysis of nuclear-encoded genes of complex I. We report on our experience to date with prenatal diagnosis in pregnancies at risk for complex I deficiency. We measured complex I activity in native CV and/or cultured CV in 23 pregnancies in 15 families. In accordance with the results of the investigations in CV, 15 children were born clinically unaffected. Two prenatally diagnosed unaffected fetuses and two prenatally diagnosed affected fetuses were lost prematurely with spontaneous or provoked abortions, respectively. Two affected children were born (prenatally found to be affected). In two pregnancies a discrepancy between native and cultured cells was found. We conclude that prenatal diagnosis for complex I deficiency can be reliably performed. Pitfalls were encountered in using cultured CV as a result of maternal cell contamination (MCC). Future research on pathogenic nuclear mutations underlying complex I deficiency will extend the possibilities for prenatal diagnosis at the molecular level. Copyright © 2001 John Wiley & Sons, Ltd.     

Occurrence of perfluoroalkyl substances in matched human serum, urine, hair and nail

The purpose of this study was to determine perfluoroalkyl substances(PFASs) in human serum, urine, hair and nail from general populations, and to investigate the possibility for human urine, hair and nail used as the biomonitoring sample for PFASs exposure. We detected the concentrations of PFHx A, PFOA, PFNA, PFDA, PFUn DA, PFDo A, PFHx S and PFOS in 39 matched human serum, urine, hair and nail samples from Shenzhen in China. The detection frequency and the median level of PFOS were all higher than that of the other PFASs in four matrices. The median concentration of PFOS in serum, urine, hair, and nail were 9.24 ng/mL, 13.96 ng/L, 0.58 ng/g and 0.63 ng/g, respectively. The results of spearman correlation test indicated that nail was an ideal matrix for biomonitoring PFOS rather than human urine and hair in general populations for the non-invasive sampling.     

尿液收集储存过程中的性质变化及其资源化利用

尿液收集和储存过程中,其营养元素的变化会受到多种环境因素和生物活动的影响,基于源分离厕所理念,模拟尿液收集过程,检测了新鲜尿液收储过程中的性质变化,测定了实验过程中3批次新鲜尿液以及1份经密封储存＞60 d的陈尿的各项关键指标.结果 表明:不同尿液性质不同,陈尿ρ(TN)为7.6 g/L,ρ(TP)为286 mg/L....     

Evaluation of a prenatal screening procedure for β-thalassaemia carriers in a chinese populationbased on the mean corpuscular volume (MCV)

Haemoglobin A₂ (HbA₂) levels were determined on 25 β-thalassaemia carriers by the microcolumn method and were found to range from 4.5–7.2 per cent (mean 5.2±0.82 S.D.). The haemoglobin level (Hb), mean corpuscular volume (MCV), plasma ferritin and HbA₂ levels were measured on a further 299 cconsecutive Chinese pregnant women at a gestation of less than 24 weeks. 18 patients (6 per cent) had HbA₂ level greater than 4.5 per cent and were diagnosed to be β-thalassaemic carriers. It was observed that all these patients had a MCV below 75 fl. If this level is selected in a screening procedure based on measurement of MCV alone all β-thalassaemia carriers could be detected and 11 per cent of the population screened would require HbA₂ estimation. At a lower cut-off level of 70 fl, 8 per cent of the population screened wouid require HbA₂ measurement (a decrease of 27 per cent) but the detection rate will be lowered considerably (83 per cent). The high false positive rate at all cut-off levels of MCV was largely due to the prevalence of iron deficiency anaemia in the population. Estimation of plasma ferritin level in patients with low MCV will reduce this false positive rate, but there will be a considerable delay in diagnosis in patients with concomitant iron deficiency and β-thalassaemia. The presence of iron deficiency in β-thalassaemia carriers did not reduce their HbA₂ level below the diagnostic range in this study. Measurement of Hb level did not appear to be useful as a screening method since one third of the β-thalassaemia carriers had a Hb level over 11 g/dl. The validity of the MCV cut-off levels derived from the first part of the study was assessed in screening a larger population. 61 β-thalassaemia carriers (6 per cent) were detected out of 1166 patients screened. This incidence was not significantly different from the first part of the study. All these 61 patients had a MCV less than 75 ml. It was concluded that a two-step screening policy, based on MCV measurement followed by HbA₂ estimation when the MCV value is less than 75 fl, is suitable for our population. It is efficient, straight forward with excellent sensitivity and required less time and effort for both laboratory staff and clinicians.     

Prenatal diagnosis of molybdenum cofactor deficiency and isolated sulfite oxidase deficiency

Molybdenum cofactor deficiency and isolated sulfite oxidase deficiency are autosomal recessive inborn errors of metabolism with severe neurological symptoms resulting from a lack of sulfite oxidase activity. The deficiencies can be diagnosed prenatally by monitoring sulfite oxidase activity in chorionic villus sampling (CVS) tissue. In those families in which the specific defects have been identified, diagnosis can be achieved by mutation analysis or linkage studies directed at affected genes. These include MOCS1, MOCS2 or GEPH, in cases of molybdenum cofactor deficiency, or SUOX in patients with isolated sulfite oxidase deficiency. Copyright © 2002 John Wiley & Sons, Ltd.     

Prenatal diagnosis of infantile GM 2 gangliosidosis type II (sandhoff disease) by detection of N-acetylglucosaminyl-oligosaccharides in amniotic fluid with high-performance liquid chromatography

Prenatal diagnosis of Sandhoff disease (infantile onset) at 16 weeks gestation has been made by detection and analysis of N-acetylglucosaminyl-oligosaccharides in amniotic fluid using high performance liquid chromatography. The elution profile for the branched chain oli-gosaccharides was identical with that obtained with neonatal and infantile Sandhoff urine. The concentration of the oligosaccharides in the fluid was 1/100th that of urine but when calculated relative to creatinine the levels were similar. No oligosaccharides were detected in normal control amniotic fluids (10 patients) at a similar gestational age. Based on the levels of the amniotic fluid oligosaccharides and the sensitivity limits of the assay, prenatal diagnosis of patients with the juvenile onset form of the disease may also be possible with this technique.     

Prenatal diagnosis of purine nucleoside phosphorylase deficiency in the first and second trimesters of pregnancy

Prenatal diagnosis was performed in two successive pregnancies of a mother with a previous child with purine nucleoside phosphorylase (PNP) deficiency. In one pregnancy, an affected fetus was diagnosed in the 18th week of gestation after the demonstration of PNP deficiency in cultured amniotic fluid cells. Also an abnormal purine nucleoside profile was found in the amniotic fluid. The diagnosis of an affected fetus was confirmed by the analysis of cultured fetal skin fibroblasts and placental villi. The complete deficiency of PNP activity in placental villi confirms that the prenatal diagnosis of this disorder is possible by the direct investigation of chorionic villi. In the subsequent pregnancy, a heterozygous fetus was predicted in the tenth week of pregnancy by using chorionic villi.     

Fetal presentation of morquio disease type A

A fetus with mucopolysaccharidosis type IV A (Morquio type A) is described. The family had one affected child exhibiting symptoms of classical Morquio A disease, and late in the subsequent pregnancy prenatal diagnosis was requested. At 23 weeks' gestation, moderate ascites was detected by detailed ultrasound scan and keratan sulphate was found in the amniotic fluid. The pregnancy was terminated by prostaglandin induction and the diagnosis of mucopolysaccharidosis type IV A was confirmed by demonstration of a deficiency of N-acetylgalactosamine-6-sulphate (GalNac-6-S) sulphatase in cultured amniotic cells and in post-mortem fibroblast cultures. The activities of β-galactosidase and arylsulphatase A were normal, ruling out Morquio disease type B and multiple sulphatase deficiency. These results indicate that mucopolysaccharidosis IV A (a disease that predominantly affects the skeletal system) may produce ascites in the fetus to such an extent that it can be detected by ultrasound.     

壬基酚和辛基酚暴露的生物标志物——尿液5-HT

为了探讨壬基酚和辛基酚对大鼠体内(5-HT)代谢的影响,为壬基酚和辛基酚暴露标志物的鉴定提供依据.选取健康雄性Sprague Dawley大鼠35只,随机分为对照组,壬基酚、辛基酚及其联合染毒低剂量组(50 mg·kg-1·d-1),壬基酚、辛基酚及其联合染毒高剂量组(150 mg·kg-1·d-1),每组5只,连续7d灌胃染毒,建立染毒动物模型.灌胃次日,利用高效液相色谱法(HPLC)检测各组大鼠尿液中5-羟色胺(5-HT)含量.结果表明,染毒组大鼠尿液中5-HT浓度均显著高于正常对照组(p＜0.01);高剂量染毒组大鼠尿液中5-HT浓度均显著高于其低剂量染毒组(p＜0.01);染毒第7天大鼠尿液中5-HT浓度均显著高于其染毒第3天(p＜0 01)尿液中5-HT浓度随染毒剂量和染毒时间的增加而升高,有剂量-毒性效应和时间-毒性效应关系;尿液5-HT可作为评估壬基酚和辛基酚暴露的潜在生物标志物,应用于大规模人群暴露风险的监测.     

Further predictors of renal dysplasia in fetal obstructive uropathy: Bladder pressure and biochemistry of ‘fresh’ urine

Urine was aspirated on two consecutive days from the dilated bladder of nine fetuses with lower urinary tract obstruction. Gestational age ranged from 17 to 35 weeks. Renal dysplasia was diagnosed histologically in four fetuses, whereas the other five had normal renal histology or only partial dysplasia. Urinary sodium (Na⁺) and osmolality (Osm) decreased significantly in the second urine sample 1 day after bladder emptying (median decrease: Na⁺ = −11.3 per cent; Osm= −13.3 per cent). Although there were no significant differences between fetuses with or without renal dysplasia, normalization of an initially raised urine Na ⁺ concentration occurred at the second sample in a fetus with partially normal renal histology, thus correcting a false-positive diagnosis of dysplasia. Bladder pressure was measured at the time of the first urine sampling in seven fetuses and in a further eight with bladder outlet obstruction undergoing a single urine aspiration at 18–28 weeks. Bladder pressure was increased above the reference range in 8 of 15 fetuses with urinary obstruction, but there was no correlation between pressure and the degree of impairment of renal function. Although no conclusive clinical guidelines can be drawn from this study for the evaluation of fetal renal function, these findings suggest that, in lower urinary tract obstruction, tubular reabsorption is impeded by the standing pressure in the urinary tract and that improvement of renal function may occur following relief of obstruction.     

Multiple sulphatase deficiency: Prenatal diagnosis using chorionic villi

Multiple sulphatase deficiency was diagnosed in the first trimester of pregnancy by demonstrating markedly reduced activities of arylsulphatases and heparin sulphamidase by direct assays on chorionic villi (CV). The diagnosis was confirmed by assays on cell cultures of villi and fetal skin fibroblasts. Two further pregnancies of this mother were monitored similarly and predicted to be unaffected; one produced a normal healthy infant, the other miscarried shortly after CV sampling.     

β-Glucuronidase deficiency as a cause of prenatally diagnosed non-immune hydrops fetalis

We describe a case of β-glucuronidase deficiency presenting as a non-immune hydrops fetalis diagnosed at 26 weeks of gestation. The deficiency was disclosed on cultured amniotic fluid cells and in fetal plasma and was confirmed post-abortion. In a second pregnancy, a normal β-glucuronidase activity was found in extracts of chorionic villi obtained at 10 weeks of gestation. The pregnancy is continuing uneventfully. We conclude that it is of great importance to verify the presence of metabolic disease whenever the major causes of hydrops fetalis have been excluded.     

邻苯二甲酸酯暴露的大鼠尿液生物标志物研究

建立了检测尿液中邻苯二甲酸酯类化合物(PAEs)共同水解产物邻苯二甲酸(PA)的分析方法;通过雄性成年SD大鼠代谢试验,检测了邻苯二甲酸二丁酯(DBP)和邻苯二甲酸二(2-乙基己基)酯(DEHP)单独暴露、以及联合暴露条件下大鼠尿液中PA总量,并分析了PA与暴露总量之间的关系.结果显示,随着暴露剂量的增大,尿液中排出的PA总量也不断增大;DBP单独暴露、DEHP单独暴露以及两者联合暴露时尿液PAEs代谢物平均排泄总量占摄入量的百分比分别为65.04%、55.00%和38.35%;大鼠尿液中PA的摩尔总量与DBP、DEHP、以及两者联合暴露的摩尔量之间的相关系数分别为0.92(P<0.01)、0.56(P<0.01)、0.93(P<0.01).尿液水解后的PA能够反映PAEs类物质的总暴露水平,将来可能作为一种潜在的暴露生物标志物用于人群的PAEs暴露监测.