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Association between confined placental trisomy, fetal uniparental disomy, and early intrauterine growth retardation
1284
LETTERS to the EDITOR
Association between confined
placental trisomy, fetal uniparental disomy, and early intrauterine growth retardation SIR,-As an example of genomic imprinting, deletion of chromosome 15ql 1-13 results in the Prader-Willi syndrome when inherited from the father but in the Angehnan syndrome when inherited from the mother.1 Detection of uniparental disomy, the inheritance of a pair of homologous chromosomes from the same parent, requires analysis of DNA polymorphisms, although cases have been identified after father-to-son transmission of haemophilia A and cystic fibrosis in a baby with only one carrier parent.1 In mice, uniparental disomy may be associated with growth disorder.2 Patients with cystic fibrosis due to uniparental disomy 7 have intrauterine growth retardation although babies with cystic fibrosis are usually of normal birthweight. Isolated placental chromosomal mosaicism is detected in 1-2% of chorionic villus biopsy specimens.3 We postulate that isolated placental mosaicism may be associated with early profound growth disorder when the fetus has uniparental disomy but with relatively normal growth when the fetus is normal. In the first case, a primigravid pregnancy, ultrasonography at 13 and 16 weeks showed no fetal abnormalities and appropriate growth. At 23 weeks, fetal growth had fallen dramatically from the 25th centile to below the 5th centile. No structural abnormalities were detected and umbilical artery waveforms were normal. Chorionic villus biopsy and rapid karyotyping revealed trisomy 16. The parent elected for termination. The structurally normal fetus weighed 350 g below the 3rd centile. Culture of placental trophoblast showed mosaic trisomy 16/normal in the ratio 65/35. Fetal brain, ovary, kidney, liver, and lung were all karyotypically normal. The second case was also a primigravid pregnancy, first seen at 19 weeks when ultrasound measurements were appropriate for gestational age and no fetal abnormalities were detected. Fetal growth continued to be normal at 25,27, and 29 weeks. At 34 weeks, fetal growth velocity had slowed with estimated fetal weight now on the 10th centile. Ultrasound showed placental haematoma at the cord insertion, the umbilical cord itself was oedematous, and there was absence of flow at end-diastole on umbilical artery doppler. A normal 16 kg girl, 10th centile for gestation, was delivered by caesarean section. Umbilical blood gases were normal. No fetal abnormalities were detected but examination of the placenta confirmed the haematoma and oedema of the cord. The body had a normal karyotype but direct examination and long-term culture of the placenta showed trisomy 16 in all cells. DNA analysis was done on parental blood and fetal tissues in the first case and on parental and fetal blood in the second case with two chromosome 16, variable number of tandem repeat (VNTR) probes: p79-2-23 (16q22-24) and pMS625 (16 pter). In the first case the mother was homozygous for p79-2-23 and heterozygous for pMS625, and the father was heterozygous for p79-2-23 and homozygous for pMS625 (figure). The placenta showed two maternal and one paternal allele in each case. All the fetal tissues
showed only maternal alleles, which indicates maternal uniparental disomy for chromosome 16. In the second case both parents were heterozygous at both loci. Fetal blood showed two alleles at each locus, one maternal and one paternal. Therefore this fetus did not have uniparental disomy for chromosome 16. The placenta showed three alleles for p79-2-23, two maternal and one paternal, but only two alleles for pMS625, indicating that there had been a recombination event between these two loci in one of the non-disjoined maternal chromosomes. Growth retardation in our first case was less likely to be due to inadequate placental function but more to underexpression of imprinted growth-related genes on chromosome 16. This phenomenon has been seen in the mouse.4Mouse maternally derived, uniparental disomy for distal chromosome 7 is associated
Case One
Case Two
p79-2-23 (16q22-q24)
pMS625 (16pter) Southern blots of fetal and parental DNA. to
Digestion with restriction enzymes Taq 1 and Mbo 1 and hybridisation chromosome 16 VNTR probes p79-2-23 and pMS625, respectively.
1285
with growth retardation. Paternally derived uniparental disomy for distal chromosome 7 is lethal. The late growth retardation in our second case was associated with structural placental abnormality and significant changes in the umbilical artery blood flow characteristics, which suggests a placental cause. Haig and Graham’s mode}5 predicts the evolution of genomic imprinting for growth-related genes when one female may bear offspring by more than one male and where postnatal care is mostly maternal. Evolution should favour expression of paternal alleles to increase the size of offspring whilst maternal gene expression should reduce size. Our two cases support the extrapolation of this theory to man. Human intrauterine growth retardation may be associated with pre-eclampsia or abnormal placentation, with maternal vascular or connective tissue disorder, with fetal structural or karyotypic abnormalities, or with intrauterine infections. There remains a proportion of cases currently classified as idiopathic, some of which may be associated with uniparental disomy. The Haig-Graham model predicts that human maternal uniparental disomy may cause growth retardation whilst paternal uniparental disomy causes macrosomia. The higher rates of non-disjunction in oocytes6 than in spermatozoa7 suggest that maternal uniparental disomy may be up to ten-fold more frequent than paternal uniparental disomy, which could account for the greater number of cases of unexplained growth retardation than unexplained macrosomia in man.
acrylamide sequencing gel by automated laser fluorescence (Phannacia). For this method, one of the amplification primers had be labelled at the 5’ end with fluorescein. This analysis revealed that in the placenta two chromosomes 16 originated from the mother and the third chromosome 16 from the father (figure). In the child only the two chromosomes 16 of maternal origin are present, indicating loss of the paternal chromosome 16. These results were confirmed by conventional Southern blot analysis of the hypervariable region at the alpha globin locus (3’HVR; D 16S85). Paternity was confirmed by testing HINvI digested genomic DNA with four highly polymorphic single locus probes at D2S44 (YNH24), D5S43 (MS8), D7S21 (MS31), and D12S11 (MS43A). to
This research was funded by Birthright, grant B2/92.
PHILLIP BENNETT JANET VAUGHAN DEBORAH HENDERSON SIOBHAN LOUGHNA GUDRUN MOORE
Action Research Laboratory for the Molecular Biology of Fetal Development, Institute of Obstetrics and Gynaecology, Royal Postgraduate Medical School, Queen Charlotte’s and Chelsea Hospital, London W6 0XG, UK
1. Hall JG. How imprinting is relevant to human disease. Development 1990; suppl: 141-48. 2. Surani MAH, Rashmi K, Allen ND, et al. Genome imprinting and development in mouse. Development 1990; suppl: 89-98. 3. Kalousek DK. Confined placental mosaicism and intrauterine development. Placenta
1990; 10: 69-77. 4. De Chiara TM, Robertson EJ, Efstratiadis A. Parental imprinting of the mouse insulin-like growth factor II gene. Cell 1991; 64: 849-59 5. Haig D, Graham C. Genomic imprinting and the strange case of the insulin-like growth factor II receptor. Cell 1991; 64: 1045-46. 6. Wramsby H, Fredga K, Leidholm P. Chromosome analysis of human oocytes recovered from preovulatory follicles in stimulated cycles. N Engl J Med 1987; 316: 121-24. 7. Martin RH, Rademaker AW, Hildebrande K, Long-Simpson L, Pederson P, Yamamoto J. Variation in the frequency and type of sperm chromosonal abnormalities among normal men. Hum Genet 1987; 77: 108-44.
Uniparental disomy with
normal
phenotype
SIR,-A possible mechanism leading to uniparental disomy (ie, chromosomal loss in an originally trisomic conceptus) was document in two cases of prenatally diagnosed, confined placental 15 and postnatally confirmed uniparental (maternal) disomy resulting in Prader-Willi syndrome.12 We have observed uniparental (maternal) disomy 16 in a phenotypically
mosaic trisomies
normal infant. Chorionic villus sampling at 11weeks’ gestation in a 36-year-old gravida 1, para 1 because of advanced maternal age revealed trisomy 16 in all (n = 15) cells after direct chromosome preparation and chorionic cell cultures. All cells (n 30) from two different amniotic fluid cell cultures as well as cord blood (n 50) had a normal female chromosome count. The pregnancy was complicated by severe intrauterine growth retardation (IUGR). At 35 weeks’ gestation a phenotypically normal girl was born. Apgar scores were 10/10/10. At 3 months, an incarcerated inguinal hernia was corrected surgically. Hypothyroidism, suspected after a screening test, is being investigated. Neurological examination revealed adequate milestones, and physical development was normal at 3 months. To assess the parental origin of the chromosomes 16 in the placenta and in the child, the loci D16S265,3 D16S298, D16S3084 were investigated. These loci represent highly polymorphic (AC)" microsatellites. The lengths of polymerase chain reaction (PCR) amplified DNA fragments were analysed on a denaturing 6% =
=
Locus D16S265.
Major peaks reflect original alleles (All) present in genomic DNA. Additional lower peaks are caused by slippage during PCR resulting in shortened amplification products. Alleles were assigned by visual comparison of PCR peak relative to standard peaks. Mat= maternal,
pat = paternal.
Although the significance of normal clinical findings at 3z limited, this observation provides evidence that uniparental disomy might be compatible with normal development when chromosomes not subject to imprinting are involved. It is not clear whether IUGR results from uniparental disomy or placental malfunction due to the aneuploidy confined to the placenta. The diagnosis of confined placental mosaicism after chorionic villus sampling presumably implies a comparably high risk of uniparental disomy in the fetus because of random chromosome loss.s A closer look at these individuals might help to investigate the frequency and clinical consequences of uniparental disomy to assess whether or when classic chromosome analysis has to be complemented by testing the parental origin of a specific chromosome. months is
We thank Dr Rand, Institute for Rechtsmedizin, for the paternity testing.
Human Genetics Institute, Centre for Paediatrics, Centre for Gynaecology, Westfalische Wilhelms University, 4400 Munster, Germany
University of Miinster, B. DWOKNICZAK B. KOPPERS G. KURLEMANN W. HOLZGREVE J. HORST P. MINY
1. Purvis-Smith SG, Saville T, Manass S, et al. Uniparental disomy 15 resulting from correction of an initial trisomy 15. Am J Hum Genet 1992; 50: 348-50. 2. Cassidy SB, Lai L-W, Erickson RP, et al. Trisomy 15 with loss of the paternal 15 as a cause of Prader-Willi syndrome due to maternal disomy. Am J Hum Genet 1992; 51: 701-08. 3. Weber JL, Kwitek A, May PE. Dinucleotide repeat polymorphisms at the D16S260, D16S261, D16S265, D16S266 and D16S267 loci. Nucleic Acids Res 1990; 18: 4034. 4. Thompson AD, Shen Y, Holman K, Sutherland GR, Callen DR, Richards RI.
Isolation and characterisation of (AC)n microsatellite genetic markers from human chromosome 16. Genomics 1992; 13: 402-08. 5. Kalousek DK, Howard-Peebles PN, Olsen SB, et al. Confirmation of CVS mosaicism in term placentae and high frequency of intrauterine growth retardation association with confined placental mosaicism. Prenat Diagn 1991; 11: 743-50.
LETTERS to the EDITOR
Association between confined
placental trisomy, fetal uniparental disomy, and early intrauterine growth retardation SIR,-As an example of genomic imprinting, deletion of chromosome 15ql 1-13 results in the Prader-Willi syndrome when inherited from the father but in the Angehnan syndrome when inherited from the mother.1 Detection of uniparental disomy, the inheritance of a pair of homologous chromosomes from the same parent, requires analysis of DNA polymorphisms, although cases have been identified after father-to-son transmission of haemophilia A and cystic fibrosis in a baby with only one carrier parent.1 In mice, uniparental disomy may be associated with growth disorder.2 Patients with cystic fibrosis due to uniparental disomy 7 have intrauterine growth retardation although babies with cystic fibrosis are usually of normal birthweight. Isolated placental chromosomal mosaicism is detected in 1-2% of chorionic villus biopsy specimens.3 We postulate that isolated placental mosaicism may be associated with early profound growth disorder when the fetus has uniparental disomy but with relatively normal growth when the fetus is normal. In the first case, a primigravid pregnancy, ultrasonography at 13 and 16 weeks showed no fetal abnormalities and appropriate growth. At 23 weeks, fetal growth had fallen dramatically from the 25th centile to below the 5th centile. No structural abnormalities were detected and umbilical artery waveforms were normal. Chorionic villus biopsy and rapid karyotyping revealed trisomy 16. The parent elected for termination. The structurally normal fetus weighed 350 g below the 3rd centile. Culture of placental trophoblast showed mosaic trisomy 16/normal in the ratio 65/35. Fetal brain, ovary, kidney, liver, and lung were all karyotypically normal. The second case was also a primigravid pregnancy, first seen at 19 weeks when ultrasound measurements were appropriate for gestational age and no fetal abnormalities were detected. Fetal growth continued to be normal at 25,27, and 29 weeks. At 34 weeks, fetal growth velocity had slowed with estimated fetal weight now on the 10th centile. Ultrasound showed placental haematoma at the cord insertion, the umbilical cord itself was oedematous, and there was absence of flow at end-diastole on umbilical artery doppler. A normal 16 kg girl, 10th centile for gestation, was delivered by caesarean section. Umbilical blood gases were normal. No fetal abnormalities were detected but examination of the placenta confirmed the haematoma and oedema of the cord. The body had a normal karyotype but direct examination and long-term culture of the placenta showed trisomy 16 in all cells. DNA analysis was done on parental blood and fetal tissues in the first case and on parental and fetal blood in the second case with two chromosome 16, variable number of tandem repeat (VNTR) probes: p79-2-23 (16q22-24) and pMS625 (16 pter). In the first case the mother was homozygous for p79-2-23 and heterozygous for pMS625, and the father was heterozygous for p79-2-23 and homozygous for pMS625 (figure). The placenta showed two maternal and one paternal allele in each case. All the fetal tissues
showed only maternal alleles, which indicates maternal uniparental disomy for chromosome 16. In the second case both parents were heterozygous at both loci. Fetal blood showed two alleles at each locus, one maternal and one paternal. Therefore this fetus did not have uniparental disomy for chromosome 16. The placenta showed three alleles for p79-2-23, two maternal and one paternal, but only two alleles for pMS625, indicating that there had been a recombination event between these two loci in one of the non-disjoined maternal chromosomes. Growth retardation in our first case was less likely to be due to inadequate placental function but more to underexpression of imprinted growth-related genes on chromosome 16. This phenomenon has been seen in the mouse.4Mouse maternally derived, uniparental disomy for distal chromosome 7 is associated
Case One
Case Two
p79-2-23 (16q22-q24)
pMS625 (16pter) Southern blots of fetal and parental DNA. to
Digestion with restriction enzymes Taq 1 and Mbo 1 and hybridisation chromosome 16 VNTR probes p79-2-23 and pMS625, respectively.
1285
with growth retardation. Paternally derived uniparental disomy for distal chromosome 7 is lethal. The late growth retardation in our second case was associated with structural placental abnormality and significant changes in the umbilical artery blood flow characteristics, which suggests a placental cause. Haig and Graham’s mode}5 predicts the evolution of genomic imprinting for growth-related genes when one female may bear offspring by more than one male and where postnatal care is mostly maternal. Evolution should favour expression of paternal alleles to increase the size of offspring whilst maternal gene expression should reduce size. Our two cases support the extrapolation of this theory to man. Human intrauterine growth retardation may be associated with pre-eclampsia or abnormal placentation, with maternal vascular or connective tissue disorder, with fetal structural or karyotypic abnormalities, or with intrauterine infections. There remains a proportion of cases currently classified as idiopathic, some of which may be associated with uniparental disomy. The Haig-Graham model predicts that human maternal uniparental disomy may cause growth retardation whilst paternal uniparental disomy causes macrosomia. The higher rates of non-disjunction in oocytes6 than in spermatozoa7 suggest that maternal uniparental disomy may be up to ten-fold more frequent than paternal uniparental disomy, which could account for the greater number of cases of unexplained growth retardation than unexplained macrosomia in man.
acrylamide sequencing gel by automated laser fluorescence (Phannacia). For this method, one of the amplification primers had be labelled at the 5’ end with fluorescein. This analysis revealed that in the placenta two chromosomes 16 originated from the mother and the third chromosome 16 from the father (figure). In the child only the two chromosomes 16 of maternal origin are present, indicating loss of the paternal chromosome 16. These results were confirmed by conventional Southern blot analysis of the hypervariable region at the alpha globin locus (3’HVR; D 16S85). Paternity was confirmed by testing HINvI digested genomic DNA with four highly polymorphic single locus probes at D2S44 (YNH24), D5S43 (MS8), D7S21 (MS31), and D12S11 (MS43A). to
This research was funded by Birthright, grant B2/92.
PHILLIP BENNETT JANET VAUGHAN DEBORAH HENDERSON SIOBHAN LOUGHNA GUDRUN MOORE
Action Research Laboratory for the Molecular Biology of Fetal Development, Institute of Obstetrics and Gynaecology, Royal Postgraduate Medical School, Queen Charlotte’s and Chelsea Hospital, London W6 0XG, UK
1. Hall JG. How imprinting is relevant to human disease. Development 1990; suppl: 141-48. 2. Surani MAH, Rashmi K, Allen ND, et al. Genome imprinting and development in mouse. Development 1990; suppl: 89-98. 3. Kalousek DK. Confined placental mosaicism and intrauterine development. Placenta
1990; 10: 69-77. 4. De Chiara TM, Robertson EJ, Efstratiadis A. Parental imprinting of the mouse insulin-like growth factor II gene. Cell 1991; 64: 849-59 5. Haig D, Graham C. Genomic imprinting and the strange case of the insulin-like growth factor II receptor. Cell 1991; 64: 1045-46. 6. Wramsby H, Fredga K, Leidholm P. Chromosome analysis of human oocytes recovered from preovulatory follicles in stimulated cycles. N Engl J Med 1987; 316: 121-24. 7. Martin RH, Rademaker AW, Hildebrande K, Long-Simpson L, Pederson P, Yamamoto J. Variation in the frequency and type of sperm chromosonal abnormalities among normal men. Hum Genet 1987; 77: 108-44.
Uniparental disomy with
normal
phenotype
SIR,-A possible mechanism leading to uniparental disomy (ie, chromosomal loss in an originally trisomic conceptus) was document in two cases of prenatally diagnosed, confined placental 15 and postnatally confirmed uniparental (maternal) disomy resulting in Prader-Willi syndrome.12 We have observed uniparental (maternal) disomy 16 in a phenotypically
mosaic trisomies
normal infant. Chorionic villus sampling at 11weeks’ gestation in a 36-year-old gravida 1, para 1 because of advanced maternal age revealed trisomy 16 in all (n = 15) cells after direct chromosome preparation and chorionic cell cultures. All cells (n 30) from two different amniotic fluid cell cultures as well as cord blood (n 50) had a normal female chromosome count. The pregnancy was complicated by severe intrauterine growth retardation (IUGR). At 35 weeks’ gestation a phenotypically normal girl was born. Apgar scores were 10/10/10. At 3 months, an incarcerated inguinal hernia was corrected surgically. Hypothyroidism, suspected after a screening test, is being investigated. Neurological examination revealed adequate milestones, and physical development was normal at 3 months. To assess the parental origin of the chromosomes 16 in the placenta and in the child, the loci D16S265,3 D16S298, D16S3084 were investigated. These loci represent highly polymorphic (AC)" microsatellites. The lengths of polymerase chain reaction (PCR) amplified DNA fragments were analysed on a denaturing 6% =
=
Locus D16S265.
Major peaks reflect original alleles (All) present in genomic DNA. Additional lower peaks are caused by slippage during PCR resulting in shortened amplification products. Alleles were assigned by visual comparison of PCR peak relative to standard peaks. Mat= maternal,
pat = paternal.
Although the significance of normal clinical findings at 3z limited, this observation provides evidence that uniparental disomy might be compatible with normal development when chromosomes not subject to imprinting are involved. It is not clear whether IUGR results from uniparental disomy or placental malfunction due to the aneuploidy confined to the placenta. The diagnosis of confined placental mosaicism after chorionic villus sampling presumably implies a comparably high risk of uniparental disomy in the fetus because of random chromosome loss.s A closer look at these individuals might help to investigate the frequency and clinical consequences of uniparental disomy to assess whether or when classic chromosome analysis has to be complemented by testing the parental origin of a specific chromosome. months is
We thank Dr Rand, Institute for Rechtsmedizin, for the paternity testing.
Human Genetics Institute, Centre for Paediatrics, Centre for Gynaecology, Westfalische Wilhelms University, 4400 Munster, Germany
University of Miinster, B. DWOKNICZAK B. KOPPERS G. KURLEMANN W. HOLZGREVE J. HORST P. MINY
1. Purvis-Smith SG, Saville T, Manass S, et al. Uniparental disomy 15 resulting from correction of an initial trisomy 15. Am J Hum Genet 1992; 50: 348-50. 2. Cassidy SB, Lai L-W, Erickson RP, et al. Trisomy 15 with loss of the paternal 15 as a cause of Prader-Willi syndrome due to maternal disomy. Am J Hum Genet 1992; 51: 701-08. 3. Weber JL, Kwitek A, May PE. Dinucleotide repeat polymorphisms at the D16S260, D16S261, D16S265, D16S266 and D16S267 loci. Nucleic Acids Res 1990; 18: 4034. 4. Thompson AD, Shen Y, Holman K, Sutherland GR, Callen DR, Richards RI.
Isolation and characterisation of (AC)n microsatellite genetic markers from human chromosome 16. Genomics 1992; 13: 402-08. 5. Kalousek DK, Howard-Peebles PN, Olsen SB, et al. Confirmation of CVS mosaicism in term placentae and high frequency of intrauterine growth retardation association with confined placental mosaicism. Prenat Diagn 1991; 11: 743-50.