Biomedical subjects
Publications and source records attributed to W Holzgreve.
We performed prenatal testing to predict the inheritance of choroideremia (CHM) using a linked polymorphic DNA marker, DXS95. DNA analysis of chorionic villi at the 12th week of pregnancy indicated that the allele at risk had not been passed from the heterozygous mother to the fetus. This prenatal exclusion of choroideremia was confirmed by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) analysis.
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We have reviewed the follow-up of almost 3000 completed pregnancies in the Münster CVS Program and identified 4 children with distal limb deficiencies. Two cases involved only minor anomalies of distal digital phalanges. One child had a Hanhart anomaly (hypoglossia hypodactylia). We also reviewed 24 cases of limb defects following CVS reported previously. With the exception of a single series, neither the overall incidence of affected children nor cases reported from larger series provide unambiguous evidence of an increased risk of distal limb deficiency caused by CVS.
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In a multicenter trial we retrospectively evaluated the clinical and sonographic data of 49 neonatal ovarian cysts, 44 of which were detected prenatally and 5 on the first day after delivery. Of the 44 prenatally detected cysts 39 were purely cystic, 5 echogenic or had a mixed pattern. In 20 patients the cystic appearance changed during delivery from purely cystic to a mixed pattern being independent on the size of the cyst. 26 of the 44 cysts were treated surgically. Salpingotorsion was found in 8 and was independent on the size of the cyst. In 15 a salpingo-oophorectomy or oophorectomy was performed, in 11 the ovary was saved. 23 patients were followed sonographically: 15 cysts showed complete resolution within 14 months without correlation to the sonographic pattern. The volume of these cysts varied between 5 and 71 ml. Neonatal ovarian cysts disappear spontaneously frequently and rarely cause severe symptoms. The authors recommend follow-up by ultrasound as the primary modality. Surgical intervention is recommended only if the cyst is space-occupying and percutaneous puncture can not be performed or in the case of emergency.
OBJECTIVE: We wanted to test whether the recently described method of using the transferrin receptor system for fluorescence-activated cell-sorter enrichment of nucleated red blood cells can be used for prenatal diagnosis from maternal blood. STUDY DESIGN: Instead of the laborious, expensive fluorescence-activated cell-sorter system, we used the newly described magnetic-activated cell sorter. RESULTS: An effective enrichment could be achieved with separation of lymphocyte subsets. With the transferrin receptor, however, the enrichment was very inefficient because of the poor specificity of the antibody itself. Even in umbilical cord blood only 25% of nucleated red blood cells were labeled as demonstrated by immunogold silver enhancement of transferrin receptor-labeled cells. CONCLUSION: In spite of the availability of a fast and effective separation method (magnetic-activated cell sorter) the use of the transferrin receptor antigen alone is not likely to enable a reliable identification of fetal cells in maternal circulation.
OBJECTIVE: This series was designed to assess in a pilot study the feasibility of using fluorescence in situ hybridization on chorionic villi. STUDY DESIGN: We constructed probes derived from specific subregions of human chromosomes 21, 18, 13, X, and Y that give a single copylike signal when used in conjunction with suppression hybridization. RESULTS: In a blind series of 47 samples all, including one trisomy 21, were correctly identified. The samples were correctly classified as disomic for five chromosomes. CONCLUSIONS: The combination of chromosome-specific probe sets composed primarily of cosmid contigs and optimized hybridization and detection allowed accurate chromosome enumeration in uncultured human chorionic villi; these results are consistent with those obtained by traditional cytogenetic analysis and suggest a use for fluorescence in situ hybridization as an adjunct to karyotyping when rapid results are needed.
OBJECTIVE: Intrauterine growth retardation associated with fetal chromosome anomalies is usually documented on ultrasonography late in the second trimester. However, we believe and attempt to document here that the impact of aneuploidy on fetal growth is evident much earlier (i.e., the aneuploid fetus may appear smaller than dates on ultrasonography even in the first trimester). STUDY DESIGN: For the population referred to our center for chorionic villus sampling from January 1988 to July 1991, we compared gestational age as calculated from the last menstrual period to that derived from fetal size as measured by crown-rump length. A cutoff of 7 days was chosen to select the study group. The remainder of our chorionic villus sampling population in which fetal size was expected was used as controls. We also divided those chorionic villus sampling patients by when a fetal death was observed by size. RESULTS: In the study period 3194 chorionic villus sampling procedures were performed and in 277 (8.7%) fetal length was smaller than expected by at least 7 days. Sixty (1.9%) chromosome anomalies were diagnosed by first trimester chorionic villus sampling in the study period. The frequency of chromosome anomalies was 4.3% in the study group and 1.7% in controls (p < 0.004). The more aberrant the karyotype on "postmortem chorionic villus sampling," the greater the growth retardation tended to be. CONCLUSIONS: In our chorionic villus sampling population a fetal crown-rump length smaller than dates is associated with a significant increase in risk of chromosome anomalies. Moreover, the larger the size-dates discrepancy, the higher the possibility that the aneuploidy affecting that pregnancy is of the severe or lethal type.
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The polymerase chain reaction (PCR) was used for detecting parvovirus B19 DNA in clinical specimens. A pair of oligonucleotide primers spanning the PstI-fragment of the B19 virus genome was used for PCR, and a PCR product of 727 bp was amplified. B19 virus DNA was detected in all sera (n = 26) of individuals in the incubation period and acute phase of infection. PCR was useful for detecting viral B19 DNA in amniotic fluid and fetal blood of hydropic fetuses, confirming fetal B19 virus infection.
OBJECTIVE: To investigate whether the ratio of fetal cells in the maternal circulation differs before and after the blood passes through the maternal lung. METHODS: We performed polymerase chain reaction-based Y-sequence analysis of DNA derived from antecubital vein blood obtained before and 2 hours after cesarean delivery, and from uterine vein blood obtained during cesarean of 14 women carrying male fetuses. RESULTS: Fetal DNA was detected in 17 tested specimens and, as estimated by comparison with parallel dilution series, the fetal-maternal DNA ratio was 1:10(5) to 1:10(6). However, there was no significant difference in the amount of Y-chromosomal DNA detectable between uterine vein blood and peripheral blood after polymerase chain reaction and Southern hybridization. In DNA derived from peripheral blood after delivery, the intensity of Y-specific fetal DNA sequences was also not significantly increased. CONCLUSION: Our results argue against the often-stated hypothesis of different ratios of fetal to maternal DNA in the uterine vein versus peripheral blood, and indicate that even delivery does not seem to increase the flow of fetal cells into the maternal circulation.
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The analysis of fetal cells from the maternal circulation would be the least invasive method of prenatal diagnosis. Potential fetal cell types to enter the maternal circulation are lymphocytes, trophoblast cells and nucleated erythrocytes. With conventional methods, such as cytology and interphase or metaphase cytogenetics, the ratio of fetal to maternal cells was overestimated in the past. Currently most groups use polymerase chain reaction-based Y-sequence analysis for the detection of fetal cells in pregnancies with male fetuses, either with or without prior enrichment of fetal cells. For fetal cell separation, fluorescence-activated cell sorting and immunomagnetic beads have been applied, and recently our group has used discontinuous density gradient centrifugation for this purpose. We have shown that the transferrin receptor antigen alone is not sufficient for enrichment of fetal nucleated erythrocytes. Despite some initial promising results with fluorescence in situ hybridization, the reproducibility and reliability of the techniques are still limited, mainly due to the lack of very specific cell markers and the very low and variable concentrations of fetal cells among numerous maternal cells.
Eighty-nine newborns and infants with congenital urinary tract malformations were treated in the childrens' hospital of the Westfälische Wilhelms-University from 1986 to 1989. Twenty patients of this group (22.5%) developed severe hypertension requiring treatment within the first year of life. Mean age of diagnosis of hypertension was 5 months (range 0.5-12 months). Median values for blood pressure at time of diagnosis were 138 mmHg (range 120-170) for systolic and 92 mmHg (range 80-110) for diastolic values. Six patients showed characteristic symptoms for hypertension such as restlessness, sweating and sleep disorders. Plasma levels of renin were obtained in 12 of 20 patients. Five patients had raised plasma renin levels. All patients with a severe hypertension were treated with one to several antihypertensive drugs. Risk factors for the development of severe renal hypertension in early infancy are cystic renal malformation, vesico-ureteral reflux, obstructive uropathy and to our experience also short term percutaneous nephrostomy in obstructive uropathy in particular in connection with pyelonephritis. Hypertension can still appear after the successful surgical correction of urinary obstruction. We describe the group of patients with severe hypertension in our study group; diagnostic principles and our therapeutic approach are explained. We conclude that early diagnosis of severe hypertension and consecutive treatment are important in infants with congenital urinary malformations.
A 37-year-old gravida was referred for CVS because of advanced maternal age. A trisomy 21 was present in all cells after short-term incubation (direct processing (DP)) and long-term culture. According to our policy, a retap was offered for confirmation of the result during the legally required 3-day waiting period between communication of the result and termination of pregnancy. Unexpectedly all cells after DP showed a normal male chromosome complement. Further investigations revealed mosaicism in trophoblast tissue and a normal karyotype in amniotic fluid cells and fetal blood (50 mitoses each). The parents elected to continue the pregnancy after extensive ultrasound examinations did not show suspicious findings. After the birth of a healthy child, cell cultures from ten different placental sites confirmed mosaicism. Four out of 100 mitoses from a lymphocyte culture showed an additional chromosome 21. The child had no dysmorphic features and the development was normal at the age of 10 weeks. This case demonstrates the restricted validity of prenatal cytogenetic analysis in the presence of true fetal mosaicism. It also stresses the benefit of our policy to offer a retap in cases with abnormal cytogenetic results prior to termination of pregnancy which is considered unnecessary by many cytogeneticists.
Discrepant chromosome findings in placenta and fetus (false negative and false positive) after chorionic villus sampling (CVS) are mainly due to confined mosaicism. Non-mosaic normal or abnormal chromosome counts after direct preparation and culture nearly always correctly reflect the fetal chromosome constitution. False-negative results have almost exclusively been restricted to cytotrophoblast cells not representing a fetal chromosome abnormality. Diagnosis of placental mosaicism definitely requires an adequate follow-up by amniocentesis, fetal blood sampling, or sonography before a pregnancy is terminated. When direct preparations and cultured cells are used for cytogenetic diagnoses and placental mosaicism is not taken as proof for a chromosomal abnormality in the fetus, CVS is an accurate diagnostic tool.
Varicella zoster virus infection acquired in pregnancy is reported to cause fetal damage in 5% to 10% of cases. We used polymerase chain reaction to attempt molecular diagnosis of fetoplacental varicella zoster virus infection in two patients. Tissue obtained by chorionic villus sampling in the second trimester was analyzed by polymerase chain reaction with a varicella zoster virus-specific primer, ORF-63, and was found to be positive in both patients. Viral cultures were negative. One patient elected pregnancy termination at 23 weeks. Southern blot hybridization of neonatal brain tissue for varicella zoster virus was negative. The second patient carried the pregnancy to term and was delivered of a normal infant. Varicella zoster virus immunoglobulin M and viral cultures were negative. The presence of viral deoxyribonucleic acid sequences in placental tissue does not correlate with fetal disease.
An ultrasound evaluation of the pregnancy in the first trimester is very helpful because no other method can better answer questions about the viability of the embryo, the true gestational age, the presence of an ectopic pregnancy and developmental anomalies at this early stage. Especially the introduction of vaginal sonography meant considerable progress in this area. Now there are normal growth curves available based on longitudinal measurements for the yolk sac, the amniotic and chorionic cavity and the crown-rump length. Significant deviations from these normal curves are suggestive of genetic anomalies of the developing child. Of these anomalies, detectable in the first trimester, especially the nuchal edema is associated with a high rate of chromosomal abnormalities. The fact that the gestational age can better be determined in the first trimester than in the later course of pregnancy is a strong argument for the introduction of routine ultrasound screening within the first twelve weeks of pregnancy.