Twin-twin transfusion syndrome in a dizygotic monochorionic-diamniotic twin pregnancy.

We present a case of twin-twin transfusion syndrome with discordant gender. Monochorionicity was confirmed by surgical pathology. Cytogenetic analysis showed normal 46,XX and 46,XY karyotypes. Microsatellite analysis using reliable pericentromeric markers was consistent with dispermic fertilization of two separate ova. This suggests that monochorionicity, rather than zygosity, may be responsible for the development of placental vascular anastomoses.

Peripheral neuropathy in the first 2 years of life: diagnostic evaluation including molecular genetics.

Inherited polyneuropathies present in the first 2 years of life are discussed with emphasis on clinical, pathologic, and molecular data. Early-onset polyneuropathies are relatively rare, sometimes life-threatening conditions that demand early recognition by clinical and pathologic examination. Histologic and ultrastructural overlaps among the various conditions are sometimes resolved by molecular genetic analysis. The growth in disease identification by genetic localization allows a more comprehensive understanding of the clinical and morphologic heterogeneity involving rearrangements of the same gene. Molecular mechanisms explaining the acquisition of such gene rearrangements are beginning to be unraveled. Peripheral myelin disorders may be confused with primary axonal disorders, and electrophysiologic examination often helps to distinguish between these two. Furthermore, early-onset central nervous system disorders may present as peripheral polyneuropathies and confound the clinical picture. A tentative diagnosis can often be offered by pathologic examination and confirmed by biochemical enzyme analysis later. The differential clinical diagnostic considerations of early-onset polyneuropathies are offered, to help clinicians sort out these diseases in the most efficient manner.

Three cell line mosaicism involving structural and numerical abnormalities of chromosome 18 in a 3.5-year-old girl: 47,XX,+18/47,XX,+del(18)(q22)/46,XX.

We report on a 3.5-year-old girl with a mosaic karyotype including full trisomy 18, normal cells and a majority of cells with partial trisomy involving an extra chromosome 18 deleted at band q22. She had cardiac and CNS anomalies, dysmorphic facial features failure to thrive and developmental delay. A gastrostomy tube was placed at 2 years of age. The combination of improved nutrition and optimal developmental therapy has led to her sitting supported, attempting to stand and enhancement of her cognitive and non-verbal communication abilities. Molecular investigation of the patient and her parents using microsatellite analysis has led to the conclusion that, as expected, the additional copy of chromosome 18 constituting the full trisomic cell line is maternal meiosis I in origin. The data, however, indicate that in the trisomic cell line containing the deleted chromosome 18q, the structurally abnormal 18 was of paternal origin. We think this case is the first described with both structural and numerical trisomic mosaicism involving chromosome 18 in a liveborn infant. We propose a mechanism of origin and review the literature, comparing the clinical presentation of this case with individuals having full or partial trisomy 18.

Uniparental isodisomy of chromosome 14 in two cases: an abnormal child and a normal adult.

Uniparental disomy (UPD) of a number of different chromosomes has been found in associated with abnormal phenotypes. A growing body of evidence for an imprinting effect involving chromosome 14 has been accumulating. We report on a case of paternal UPD of chromosome 14 studied in late gestation due to polyhydramnios and a ventral wall hernia. A prenatal karyotype documented a balanced Robertsonian 14:14 translocation. The baby was born prematurely with hairy forehead, retrognathia, mild puckering of the lips and finger contractures. Hypotonia has persisted since birth and at age one year, a tracheostomy for laryngomalacia and gastrostomy for feeding remain necessary. Absence of maternal VNTR polymorphisms and homozygosity of paternal polymorphisms using chromosome 14 specific probes at D14S22 and D14S13 loci indicated paternal uniparental isodisomy (pUPID). Parental chromosomes were normal. We also report on a case of maternal UPD in a normal patient with a balanced Robertsonian 14:14 translocation and a history of multiple miscarriages. Five previous reports of chromosome 14 UPD suggest that an adverse developmental effect may be more severe whenever the UPD is paternal in origin. This is the second reported patient with paternal UPD and the fifth reported with maternal UPD, and only few phenotypic similarities are apparent. Examination of these chromosome 14 UPD cases of maternal and paternal origin suggests that there are syndromic imprinting effects.

Fragile X syndrome: discordant levels of CGG repeat mosaicism in two brothers.

Fragile X syndrome is associated with an unstable repeated CGG trinucleotide sequence in the 5' untranslated region of the FMR-1 gene. A significant number of individuals with a mild or atypical presentation are mosaics for the CGG expansion. We report a family with two brothers. The proband had severe mental retardation as well as most of the Fragile X syndrome stigmata, whereas his brother shows only mild learning difficulties. Both inherited a 80 x CGG trinucleotide premutation from the mother. They were negative for the FRAXA fragile site in over 100 metaphases examined. Flanking markers verified that both had inherited the same FMR-1 allele and Xq27-q28 flanking sequences from the mother. The methylation status of the brothers indicated active FMR-1 transcription as determined by using StB12.3/EcoRI + Eagl blots. CGG size or methylation mosaicism was not apparent from Southern blots. Polymerase chain reaction and chemiluminescent detection identified that both brothers had different degrees of mosaicism for the CGG expansion. Large expansions amounting to 70% of the total were visible in the proband, whereas less than 5% of the signal was larger than the premutation in his mildly affected brother. These findings suggest that mosaicism may be responsible for some of the variation in penetrance in this disorder.

Presymptomatic testing for Huntington's disease. Emotional aspects.

Presymptomatic genetic testing for Huntington's disease has been available since 1986. Recent discovery of the distinct gene by the Huntington's Disease Collaborative Research Group will probably simplify testing and encourage more people to pursue it. Unfortunately advances in treatment lag behind genetics diagnosis capabilities. The individual at risk faces pursuing a test for a disease for which there is no cure, only limited treatment. The emotional aspects are reviewed of individuals being tested at the Regional Genetic Program at the University of South Florida from May 1988 until April 1993.

Correction of mucolipidosis III in vitro by gene transfer.

Mucolipidosis II (ML II, I-cell disease) and mucolipidosis III (ML III, pseudo-Hurler polydystrophy) are human autosomal recessive genetic disorders resulting from deficient UDP-N-acetylglucosamine:lysosomal enzyme precursor N-acetylglucosamine-1-phosphotransferase (GNPT) activity. Normally, this enzyme is involved in the processing of most lysosomal enzymes. Cultured fibroblasts from individuals with either disorder are deficient in a broad array of lysosomal enzymes as a result of the diminished GNPT activity. We report the correction of this phenotype by fusing transformed ML III cells generated for this study to lethally irradiated rodent cells. This method of gene transfer does not require selection for the gene of interest, animal models, nor any knowledge of the gene product except a screening method for its presence. It has generated corrected cell hybrids that contain approximately 1% hamster-derived sequences. These cell lines, which contain the hamster analogue to the human phosphotransferase gene, are useful for the molecular cloning of the gene defective in ML II and ML III.

Maternal side effects of prenatal dexamethasone therapy for fetal congenital adrenal hyperplasia.

UNLABELLED: Prenatal treatment of virilizing congenital adrenal hyperplasia in female fetuses via maternal dexamethasone (Dex) therapy (1-1.5 mg/day) from first trimester to birth was associated with excessive weight gain (47-56 pounds at 35-37 weeks gestation), Cushingoid facial features, severe striae resulting in permanent scarring, and hyperglycemic response (8-11.7 nmol/L) to oral glucose administration in all our experience (three cases). Other symptoms included hypertension, gastrointestinal intolerance, or extreme irritability. Previous pregnancies were uncomplicated by these problems. In each case, first or second trimester amniotic fluid 17-hydroxyprogesterone (17OHP, 17-41 nmol/L; normal less than 0.4 nmol/L), androstenedione (22-31 nmol/L, normal less than 5 nmol/L), and testosterone levels (0.54-0.7 nmol/L, normal less than 0.4 nmol/L) during Dex treatment were elevated regardless of the newborn genital outcome. Maternal serum estriol (E3) levels in one mother (normal newborn genitalia) were consistently low (less than 0.2 nmol/L) during the second and third trimester. In two mothers (partially virilized newborn genitalia) initial second trimester E3 levels were unsuppressed (11, 17.4 nmol/L) and suppressed (less than 1.4 nmol/L) following short-term increased dose. CONCLUSION: prenatal Dex treatment of virilizing congenital adrenal hyperplasia at a dose of 1-1.5 mg daily throughout gestation is associated with significant maternal side effects. Parents should be informed of these side effects before initiation of treatment. Careful monitoring for signs of side effects, medical intervention when necessary, and lowering of Dex dose during the second half of gestation would minimize the side effects. Maternal serum E3 levels appear useful for prediction of fetal outcome while amniotic fluid steroid levels may not.

Homozygosity for a dominant negative thyroid hormone receptor gene responsible for generalized resistance to thyroid hormone.

Generalized resistance to thyroid hormones (GRTH) commonly results from mutations in the T3-binding domain of the c-erbA beta thyroid hormone receptor gene. We have reported on a novel deletion mutation in c-erbA beta in a kindred, S, with GRTH. One patient from this kindred was the product of a consanguineous union from two affected members and was homozygous for the beta-receptor defect. This patient at 3.5 weeks of age had unprecedented elevations of TSH, free T4, and free T3 (TSH, 389 mU/L; free T4, 330.8 pmol/L; free T3, 82,719 fmol/L). He displayed a complex mixture of tissue-specific hyperthyroidism and hypothyroidism. He had delayed growth (height age, 1 3/12 yr at chronological age 2 9/12 yr) and skeletal maturation (bone age, 4 months), and developmental delay (developmental age, 8 months), but he was quite tachycardic. The homozygous patient of kindred S is markedly different from a recently reported patient with no c-erbA beta-receptor. This difference indicates that a dominant negative form of c-erbA beta in man can inhibit at least some thyroid hormone action mediated by the c-erbA alpha-receptors.

Lowe oculocerebrorenal syndrome in a female with a balanced X;20 translocation: mapping of the X chromosome breakpoint.

A Hispanic girl with Lowe oculocerebrorenal syndrome (OCRL), an X-linked recessive condition characterized by cataracts, glaucoma, mental retardation, and proteinuria, is reported. A balanced X;20 chromosomal translocation with the X chromosome breakpoint at q26.1 was found with high-resolution trypsin-Giemsa banding. Somatic cell hybridization was used to separate the X chromosome derivative and the chromosome 20 derivative in order to position, with respect to the translocation breakpoint, several DNA loci that are linked to the Lowe syndrome locus (Xq24-q26). DXS10 and DXS53 were found to be distal to the breakpoint, whereas DXS37 and DXS42 were located proximal to it. These studies suggest that the OCRL locus lies in the region between these probes. The translocation chromosome originated from an unaffected male without a visible translocation, indicating that the most likely cause of OCRL in this patient is the de novo translocation that disrupted the OCRL locus.

Cell line segregation in a 45,X/46,XY mosaic child with asymmetric leg growth.

Clinical evaluation of a 13 1/2-year-old male revealed a 4.4-cm leg length discrepancy and a small penis with a normal endocrine evaluation. Cytogenetic analysis of peripheral blood lymphocytes and skin fibroblasts derived from the back showed 45,X/46,XY mosaicism with similar percentages of 45,X cells, 36% and 30% respectively. However, two separate skin fibroblast cultures derived from the thigh and calf of the short (right) leg showed significant lack of Y-bearing cells with 100% and 80% 45,X, respectively. In contrast, skin biopsies of the thigh and calf of the normal (left) leg both showed 100% 46,XY. Similar evidence for differences in the percentages of Y-bearing cells in the left versus right leg fibroblast cultures was obtained using densitometric scanning of dot blots following DNA hybridization with a Y-specific probe at the DYZ4 locus. Asymmetric limb growth in cases of X/XY lymphocyte mosaicism warrants further cytogenetic investigation to substantiate possible genotype-phenotype correlations which may help uncover the fundamental growth deficiency in Turner syndrome.

A homozygous deletion in the c-erbA beta thyroid hormone receptor gene in a patient with generalized thyroid hormone resistance: isolation and characterization of the mutant receptor.

Different point mutations have been identified in the T3-binding domain of the c-erbA beta thyroid hormone receptor gene that are associated with variant phenotypes of generalized thyroid hormone resistance (GTHR). In most cases of GTHR, heterozygotes are affected; a single mutant allele results in the inhibition of the function of normal thyroid hormone receptors. We report here a novel genetic abnormality, a 3-basepair (bp) deletion in the T3-binding domain of the beta-receptor in a kindred, S, with GTHR. One patient, S1, was the product of a consanguineous union of two heterozygotes and was homozygous for this defect. Heterozygotes from kindred S harbored a CAC deletion at nucleotides 1295-1297, which resulted in the deduced loss of amino acid residue threonine at codon 332, and they displayed elevated free T4 levels and inappropriately normal TSH levels characteristic of other kindreds with GTHR. However, patient S1, who had two mutant alleles, had markedly elevated TSH and free T4 levels and displayed profound abnormalities in brain development and linear growth. A fibroblast c-erbA beta cDNA extending from codon 175 to stop codon 457 was cloned from patient S1, sequenced, and used to create a full-length mutant cDNA. The kindred S mutant receptor was synthesized in vitro and did not bind T3. This mutant receptor did bind with similar avidity as the wild-type human beta-receptor to thyroid hormone response elements of the human TSH beta (-12 to 43 bp) and rat GH (-188 to -160 bp) genes. Kindred S showed the effect in man of heterozygous and homozygous expression of a dominant negative form of c-erbA beta.

The human arylsulfatase-C isoenzymes: two distinct genes that escape from X inactivation.

Arylsulfatase C is a microsomal membrane-bound enzyme previously thought to be the same as steroid sulfatase, the only X-linked enzyme known to escape from X inactivation in man. We had shown that arylsulfatase C actually consists of two biochemically distinct isozymes, s and f. Only the s form has steroid sulfatase activity. The f and s forms were thought to be related through posttranslational or posttranscriptional modification of the same gene product. In part consistent with this hypothesis, we now report that in a panel of 28 rodent-human somatic cell hybrids, expression of both s and f was concordant only with the human X chromosome, thus showing that the f form is also X linked. In three separate somatic hybrids containing human X chromosomes in an inactive state, the f form was still expressed. Thus, similar to the s form, the f form also escapes from X inactivation. However, contrary to expectations, the s and f forms were not related by posttranslational modification of the same gene product. A full-length cDNA for the s form failed to hybridize to transcripts produced from an f-expressing cell line, showing that there is little sequence identity between the two. They are also not related by differential splicing of a common primary transcript, since fibroblasts from some patients with steroid sulfatase deficiency due to gene deletion of the s form continue to express the f form. Therefore, although the f and s isozymes of arylsulfatase C are X linked and escape from X inactivation, they are products from separate genes, thus providing a unique isoenzyme system to study possible gene duplication and regulation in the part of the human X chromosome that escapes inactivation.

UDP-N-acetylglucosamine: lysosomal enzyme precursor N-acetylglucosamine-1-phosphate transferase activities in human ovarian tumor tissue and some transformed cell lines.

Uridine diphosphate-N-acetylglucosamine: lysosomal enzyme precursor N-acetyl-glucosamine-1-phosphate transferase, is a key enzyme involved in the intracellular targeting of lysosomal enzymes. This enzyme is elevated fourfold in primary ovarian tumor microsomes with respect to normal ovarian microsomes. This elevation is associated with significant increases in the specific activity of multiple lysosomal hydrolases, including beta-D-hexasaminidase, alpha-L-fucosidase, and beta-D-galactosidase. The activity of the phosphotransferase was also documented in several cell lines derived from human tumors. The possible role of this enzyme in tumor-associated phosphorylation is discussed.

Sialidosis and galactosialidosis: chromosomal assignment of two genes associated with neuraminidase-deficiency disorders.

The inherited human disorders sialidosis and galactosialidosis are the result of deficiencies of glycoprotein-specific alpha-neuraminidase (acylneuraminyl hydrolase, EC 3.2.1.18; sialidase) activity. Two genes were determined to be necessary for expression of neuraminidase by using human-mouse somatic cell hybrids segregating human chromosomes. A panel of mouse RAG-human hybrid cells demonstrated a single-gene requirement for human neuraminidase and allowed assignment of this gene to the (pter----q23) region of chromosome 10. A second panel of mouse thymidine kinase (TK)-deficient LM/TK- -human hybrid cells demonstrated that human neuraminidase activity required both chromosomes 10 and 20 to be present. Analysis of human neuraminidase expression in interspecific hybrid cells or polykaryocytes formed from fusion of mouse RAG (hypoxanthine/guanine phosphoribosyltransferase deficient) or LM/TK- cell lines with human sialidosis or galactosialidosis fibroblasts indicated that the RAG cell line complemented the galactosialidosis defect, but the LM/TK- cell line did not. This eliminates the requirement for this gene in RAG-human hybrid cells and explains the different chromosome requirements of these two hybrid panels. Fusion of LM/TK- cell hybrids lacking chromosome 10 or 20 (phenotype 10+,20- and 10-,20+) and neuraminidase-deficient fibroblasts confirmed by complementation analysis that the sialidosis disorder results from a mutation on chromosome 10, presumably encoding the neuraminidase structural gene. Galactosialidosis is caused by a mutation in a second gene required for neuraminidase expression located on chromosome 20.

Heterogeneity of N-acetylglucosamine 1-phosphotransferase within mucolipidosis III.

The primary defect responsible for mucolipidosis III is a deficiency of UDP-N-acetylglucosamine:lysosomal enzyme N-acetylglucosamine 1-phosphotransferase activity (GlcNAc phosphotransferase). Genetic complementation analysis of cultured fibroblasts derived from 12 patients with mucolipidosis III identified complementation groups A, B, and C (Honey, N. K., Mueller, O. T., Little, L. E., Miller, A. L., and Shows, T. B. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 7420-7424). The GlcNAc phosphotransferase activity present in the cell lines comprising the complementation groups was characterized with respect to endogenous substrates and two exogenous acceptors, alpha-methyl-D-mannoside and high mannose glycopeptides. All group C cell lines and one group A cell line were found to have normal GlcNAc phosphotransferase activity levels at 37 degrees C when screened with these exogenous acceptors. The enzyme activity in group A cell lines was within normal range when assayed at 23 degrees C. Inhibition of the phosphorylation of alpha-methyl-D-mannoside in the presence of increasing amounts of endogenous substrate N-acetyl-beta-D-hexosaminidase B was demonstrated in normal cell lines at 23 and 37 degrees C and in group A cells at 23 degrees C. However, group C cell lines did not show any inhibition at either temperature. This suggests that the alteration of the GlcNAc phosphotransferase from individuals in group C affects the recognition site for the protein portion of lysosomal enzymes, whereas group A individuals have mutations which result in a temperature-sensitive enzyme.

I-cell disease and pseudo-Hurler polydystrophy: heterozygote detection and characteristics of the altered N-acetyl-glucosamine-phosphotransferase in genetic variants.

The human disorders I-cell disease and pseudo-Hurler polydystrophy (also known as mucolipidosis II and III, respectively) are caused by an inherited deficiency of UDP-GlcNAc: lysosomal enzyme precursor GlcNAc-P transferase activity. The most common genetic variants of these diseases (complementation group A) can be identified in homozygotes and heterozygotes using a GlcNAc-P transferase assay with artificial acceptors and commercially available radiochemicals. The kinetic characteristics of the residual GlcNAc-P transferase activity in complementation group A fibroblasts indicates that the low activity is due to a low Vmax. The measured Michaelis-Menten constants for the substrates UDP-GlcNAc and alpha-methyl mannoside are in the normal range. Homozygotes and heterozygotes of another less common variant of pseudo-Hurler polydystrophy (complementation group C) have normal activity and normal kinetic characteristics with this assay using alpha-methyl mannoside as the acceptor substrate. Several PHP variants with unusual characteristics are discussed.