Low incidence of deletion of the esterase D locus in retinoblastoma patients

Esterase D was quantitatively measured in the red blood cells from three patients from three separate kindreds who had abnormalities of chromosome 13. The esterase D activity was proportional to the number of copies of the q14 region of chromosome 13 present. These findings confirm published data localizing the esterase D gene to chromosome band 13q14, a region which is important in the etiology of retinoblastoma. Fifty-one additional retinoblastoma patients not known to have any chromosomal defect also underwent esterase D determination. In none of these patients did the esterase D measurement detect a 13q14 deletion. The normal esterase D levels in this series of 51 retinoblastoma patients suggest that deletion of an esterase D locus is infrequent in retinoblastoma patients. It must be noted that patients who are mosaics, with a 13q14 deletion in only a fraction of all somatic cells, could possibly have normal red blood cell esterase D levels. Further study is necessary to determine if esterase D determination of all retinoblastoma patients is a worthwhile clinical tool.     

Trisomy 13 in the fetus

A significant number of fetuses with trisomy 13 are spontaneously or voluntarily lost before birth; however, very few such fetuses have been systemically autopsied. In the present study, ten trisomy 13 fetuses of 130-305 mm in crown-rump length, estimated gestational age from 108 days to 239 days, were examined following either karyotype or ultrasonographic diagnosis and voluntary termination. Mean maternal age was 35.1 years. The spectrum of anatomical features was similar to that observed in neonates or older infants with trisomy 13, namely, holoprosencephaly, cyclopia, microphthalmia, cleft palate and lip, cardiac defect, polydactyly, and cystic kidney. Kidney weights were significantly increased above normal in eight of nine fetuses. Histologically, the cortex of these kidneys showed increased mitotic activity and blastemic appearance, which extended deep into the medullary areas. The weights and histology of other organs were normal except for slight increases in spleen weight.     

Molecular analysis of the expression of transthyretin in intestine and liver from trisomy 18 fetuses

Human trisomy 18 (Edwards syndrome) provides a model for the role that genes on chromosome 18 play in fetal development. Trisomy 18 occurs in approximately 1 in 3000 live births. Despite its compatibility with life in 5% of cases, prolonged survival is rare. Anomalies involve the urogenital, cardiac, craniofacial and central nervous systems. The abnormalities could be caused by the abnormal expresion of developmentally important genes on chromosome 18. We have investigated the quantity and localisation of the expression of a candidate gene, transthyretin (TTR), on chromosome 18 at the RNA level in intestine and liver tissues from trisomic fetuses and have compared the expression with normal age-matched fetal tissues. The mRNA level of TTR in 10 to 14-week intestine was the same in trisomy 18 and control tissues. However, overexpression was seen for both trisomy 18 liver and intestine at 20–23 weeks. TTR transports both thyroxine and retinol and is therefore important for normal fetal development.     

A null allele of esterase D is a marker for genetic events in retinoblastoma formation

Summary The development of homozygosity or hemizygosity in the 13q14 region by deletion, mitotic recombination, or chromosomal loss has been interpreted as a primary event in retinoblastoma. This finding is consistent with the hypothesis that inactivation of both alleles of a gene located at 13q14.11 is required for tumorigenesis. Observations reported by Benedict and colleagues in one case of bilateral retinoblastoma, LA-RB 69, provided early evidence in favor of this hypothesis. By examining levels of esterase D, an enzyme also mapping to 13q14.11, it was previously inferred that one chromosome 13 in this patient's somatic cells contained a submicroscopic deletion of the Rb and esterase D loci and that this chromosome was retained in her tumor while the normal chromosome 13 was lost. Using a rabbit anti-esterase D antibody and the esterase D cDNA probe, we have found that (1) low but detectable quantities of esterase D protein and enzymatic activity are present in tumor cells from LA-RB 69; (2) fibroblast from this patient contain two copies of the esterase D gene, indicated by heterozygosity at an ApaI polymorphic site within this gene; and (3) tumor cells from the same patient are homozygous at this site, indicating loss and reduplication of the esterase D locus. These results demonstrate that one of the two esterase D alleles in this patient acted as a null or silent allele — that is, was present in the genome with markedly decreased protein expression. This mutant allele acted as a marker for tumor-associated loss of chromosome 13 heterozygosity, in concordance with previous proposals.     

Characterisation of a 19-year-old "long-term survivor" with Edwards syndrome

Trisomy 18 is the second most frequent autosomal aneuploidy affecting about 1 in 8,000 new-borns. Similar to trisomy 13 more than 90% of the patients die within the first year. Main causes of death are failure of vital organ function, in most cases of brain, heart, kidney, and gut, sometimes combined with severe infections. The degree to which essential organs are affected at birth and the clinical course differ considerably. Unknown genetic factors and various environmental effects are most likely involved. A less severe course of Edwards syndrome can be caused by a partial trisomy due to a deletion of the extra chromosome 18 or somatic mosaicism with a trisomic and a normal cell-line in the patient. In this report conventional chromosome analysis, FISH, and QF-PCR have been performed on a 19-year-old female patient with trisomy 18 to investigate a large number of cells including non-mitotic cells from various different tissues. This study supports evidence for an apparently pure form of trisomy 18 in this "long-living" patient with Edwards syndrome.     

Molecular studies of chromosomal mosaicism: relative frequency of chromosome gain or loss and possible role of cell selection.

Studies of uniparental disomy and origin of nonmosaic trisomies indicate that both gain and loss of a chromosome can occur after fertilization. It is therefore of interest to determine both the relative frequency with which gain or loss can contribute to chromosomal mosaicism and whether these frequencies are influenced by selective factors. Thirty-two mosaic cases were examined with molecular markers, to try to determine which was the primary and which was the secondary cell line: 16 cases of disomy/trisomy mosaicism (5 trisomy 8, 2 trisomy 13, 1 trisomy 18, 4 trisomy 21, and 4 involving the X chromosome), 14 cases of 45,X/46,XX, and 2 cases of 45,X/47,XXX. Of the 14 cases of mosaic 45,X/46,XX, chromosome loss from a normal disomic fertilization predominated, supporting the hypothesis that 45,X might be compatible with survival only when the 45,X cell line arises relatively late in development. Most cases of disomy/trisomy mosaicism involving chromosomes 13, 18, 21, and X were also frequently associated with somatic loss of one (or more) chromosome, in these cases from a trisomic fertilization. By contrast, four of the five trisomy 8 cases were consistent with a somatic gain of a chromosome 8 during development from a normal zygote. It is possible that survival of trisomy 8 is also much more likely when the aneuploid cell line arises relatively late in development.     

Mouse trisomy 16 as an animal model of human trisomy 21 (Down syndrome): production of viable trisomy 16 diploid mouse chimeras

We have previously proposed that mice trisomic for chromosome 16 will provide an animal model of human trisomy 21 (Down syndrome). However, the value of this model is limited to some extent because trisomy 16 mouse fetuses do not survive as live-born animals. Therefore, in an effort to produce viable mice with cells trisomic for chromosome 16, we have used an aggregation technique to generate trisomy 16 diploid (Ts 16 2n) chimeras. A total of 79 chimeric mice were produced, 11 of which were Ts 16 2n chimeras. Seven of these Ts 16 2n mice were analyzed as fetuses, just prior to birth, and 4 were analyzed as live-born animals. Unlike nonchimeric Ts 16 mouse fetuses which die shortly before birth with edema, congenital heart disease, and thymic and splenic hypoplasia, all but 1 of the Ts 16 2n animals were viable and phenotypically normal. The oldest of the live-born Ts 16 2n chimeras was 12 months old at the time of necropsy. Ts 16 cells, identified by coat color, enzyme marker, and/or karyotype analyses, comprised 50-60% of the brain, heart, lung, liver, and kidney in the 7 Ts 16 2n chimeric fetuses and 30-40% of these organs in the 4 live-born Ts 16 2n animals. Ts 16 cells comprised an average of 40% of the thymus and 80% of the spleen in the Ts 16 2n chimeras analyzed as fetuses, with no evidence of thymic or splenic hypoplasia. However, we observed a marked deficiency to Ts 16 cells in the blood, spleen, thymus, and bone marrow of live-born Ts 16 2n chimeras as compared to 2n 2n controls. These results demonstrate that although the Ts 16 2n chimeras were, with one exception, viable and phenotypically normal, each animal contained a significant proportion of trisomic cells in a variety of tissues, including the brain. Furthermore, our results suggest that although the abnormal development of Ts 16 thymus and spleen cells observed in Ts 16 fetuses is largely corrected in Ts 16 2n fetuses, Ts 16 erythroid and lymphoid cells have a severe proliferative disadvantage as compared to diploid cells in older live-born Ts 16 2n chimeras. Ts 16 2n chimeric mice will provide a valuable tool for studying the functional consequences of aneuploidy and may provide insight into the mechanisms by which trisomy 21 leads to developmental abnormalities in man.     

Assignment of 48 ESTs to Chromosome 13 Band q14.3 and Expression Pattern for ESTs Located in the Core Region Deleted in B-CLL

An expression map containing 48 ESTs was constructed to identify a tumor-suppressor gene involved in B-cell chronic lymphocytic leukemia (B-CLL), which was previously assigned to chromosome band 13q14.3 close to genetic markers D13S25 and D13S319. Thirty-nine of these 48 ESTs, together with 11 additional ones listed in databases, were initially assigned to chromosome 13q14 between markers D13S168 and D13S176. Nine others have recently been located in the D13S319 region. Our results indicate that 48 of the 59 ESTs analyzed belong to a YAC contig of chromosome 13 band q14, and 22 are contained on YAC 933e9, which encompasses the B-CLL critical region. Ten of these 22 ESTs were accurately assigned on a PAC, BAC, and cosmid contig encompassing the smallest minimal deletion area described so far in B-CLL, and 20 were tested for their expression on 27 normal or tumor tissues. One EST appears to be a likely candidate for the tumor-suppressor gene involved in B-CLL.     

Trisomy 17 in a baboon (Papio hamadryas) with polydactyly, patent foramen ovale and pyelectasis

Trisomy 13 in humans is the third most common autosomal abnormality at birth, after trisomy 21 and trisomy 18. It has a reported incidence of between 1:5,000 and 1:30,000 live births. It is associated with multiple abnormalities, many of which shorten lifespan. We describe here the first reported case of a baboon (Papio hamadryas) with trisomy of chromosome 17, which is homologous to human chromosome 13. The trisomic infant was born to a consanguineous pair of baboons and had morphological characteristics similar to those observed in human trisomy 13, including bilateral polydactyly in the upper limbs, a patent foramen ovale, and pyelectasis. Molecular DNA analysis using human chromosome 13 markers was consistent with the affected infant inheriting two copies of chromosome 17 derived from the same parental chromosome. This trisomy was, therefore, due to either an error in meiosis II or the result of postzygotic nondisjunction. The parental origin, however, could not be determined.     

Trisomy 7, trisomy 10, and loss of the Y chromosome in short-term cultures of normal kidney tissue

Numerical chromosome aberrations are common in several types of malignant tumors. Recently, trisomy 7 and loss of the Y chromosome were described in cultures from nonneoplastic tissue, making the significance of these aberrations as cancer-associated changes doubtful. We herein report the mosaic occurrence of trisomy 7 in four consecutive short-term cultures initiated from normal kidney tissue. Smaller clones with trisomy 10 were present in three cases, and the only culture established from a male also showed mosaic loss of the Y chromosome.     

Human CRYL1, a novel enzyme-crystallin overexpressed in liver and kidney and downregulated in 58% of liver cancer tissues from 60 Chinese patients,and four new homologs from other mammalians

Lambda-crystallin is a composition of lens in rabbit and hare. It contains the putative NAD- or FAD-binding domain, which is named as HCDH domain in 3-hydroxyacyl-CoA dehydrogenase. In our attempt to search for genes differentially expressed between liver cancer tissues and normal tissues, human CRYL1 (crystallin, lambda 1) was identified. It was downregulated in 58% of 60 Chinese HCC tissue samples. The putative protein encoded by CRYL1 shares 83% identity with rabbit lambda-crystallin and contains two HCDH domains. Interestingly, CRYL1 mRNA level is remarkably high in liver and kidney, while it is extremely low in peripheral blood leukocyte and thymus. The CRYL1mRNA levels in liver and kidney are about 1.6 and 1.2 times the total amount of that in other 14 tissues, respectively. Both the special expression pattern and the putative HCDH structure of CRYL1 suggested that the protein may be of the similar function of 3-hydroxyacyl-CoA dehydrogenase. To further understand the lambda-crystallin protein family, we cloned four novel mammalian homologs from mouse, rat, bovine and pig. The unrooted phylogenetic tree of this protein family including human and other 26 species was drawn to analyse their evolutionary relationship. In addition, human CRYL1 was mapped to chromosome 13q12.11 and mouse Cryl1 to chromosome 14 between marker D14Mit83 and D14Mit260.     

Alkaline phosphatase and phosphotyrosine phosphatase activities of cultured amniotic cells with trisomy 18.

In cultured amniotic cells from fetuses with Edward's syndrome (trisomy 18), the activities of two protein phosphatases, alkaline phosphatase and phosphotyrosine phosphatase, were measured. Comparison with normal fetal cells showed a different behavior for each enzyme. Alkaline phosphatase was significantly lowered while phosphotyrosine phosphatase remained at normal levels. The interest of these enzyme assays in the screening procedure of this severe chromosome defect is discussed.     

Nondisjunction of human acrocentric chromosomes: studies of 432 trisomic fetuses and liveborns

The present report summarizes molecular studies on the parent and meiotic stage of origin of the additional chromosome in 432 fetuses or liveborns with an additional chromosome 13, 14, 15, 21, or 22. Our studies suggest that there is little variation in the origin of nondisjunction among the five acrocentric trisomies and that there is no association between the origin of nondisjunction and the likelihood of survival to term of the trisomic conceptus. The proportion of cases of paternal origin was similar among the five trisomies: 12% for trisomy 13, 17% for trisomy 14, 12% for trisomy 15, 9% for trisomy 21, and 11% for trisomy 22. The stage of nondisjunction was also similar among the five trisomies, with the majority of cases of maternal origin being due to nondisjunction at meiosis I, whereas for paternally derived cases, nondisjuction occurred primarily at meiosis II.     

Noninvasive prenatal diagnosis of fetal trisomy 18 and trisomy 13 by maternal plasma DNA sequencing

Massively parallel sequencing of DNA molecules in the plasma of pregnant women has been shown to allow accurate and noninvasive prenatal detection of fetal trisomy 21. However, whether the sequencing approach is as accurate for the noninvasive prenatal diagnosis of trisomy 13 and 18 is unclear due to the lack of data from a large sample set. We studied 392 pregnancies, among which 25 involved a trisomy 13 fetus and 37 involved a trisomy 18 fetus, by massively parallel sequencing. By using our previously reported standard z-score approach, we demonstrated that this approach could identify 36.0% and 73.0% of trisomy 13 and 18 at specificities of 92.4% and 97.2%, respectively. We aimed to improve the detection of trisomy 13 and 18 by using a non-repeat-masked reference human genome instead of a repeat-masked one to increase the number of aligned sequence reads for each sample. We then applied a bioinformatics approach to correct GC content bias in the sequencing data. With these measures, we detected all (25 out of 25) trisomy 13 fetuses at a specificity of 98.9% (261 out of 264 non-trisomy 13 cases), and 91.9% (34 out of 37) of the trisomy 18 fetuses at 98.0% specificity (247 out of 252 non-trisomy 18 cases). These data indicate that with appropriate bioinformatics analysis, noninvasive prenatal diagnosis of trisomy 13 and trisomy 18 by maternal plasma DNA sequencing is achievable.     

Maternal uniparental disomy for human chromosome 14, due to loss of a chromosome 14 from somatic cells with t(13;14) trisomy 14.

Uniparental disomy (UPD) for particular chromosomes is increasingly recognized as a cause of abnormal phenotypes in humans. We recently studied a 9-year-old female with a de novo Robertsonian translocation t(13;14), short stature, mild developmental delay, scoliosis, hyperextensible joints, hydrocephalus that resolved spontaneously during the first year of life, and hypercholesterolemia. To determine the parental origin of chromosomes 13 and 14 in the proband, we have studied the genotypes of DNA polymorphic markers due to (GT)n repeats in the patient and her parents' blood DNA. The genotypes of markers D14S43, D14S45, D14S49, and D14S54 indicated maternal UPD for chromosome 14. There was isodisomy for proximal markers and heterodisomy for distal markers, suggesting a recombination event on maternal chromosomes 14. In addition, DNA analysis first revealed--and subsequent cytogenetic analysis confirmed--that there was mosaic trisomy 14 in 5% of blood lymphocytes. There was normal (biparental) inheritance for chromosome 13, and there was no evidence of false paternity in genotypes of 11 highly polymorphic markers on human chromosome 21. Two cases of maternal UPD for chromosome 14 have previously been reported, one with a familial rob t(13;14) and the other with a t(14;14). There are several similarities among these patients, and a "maternal UPD chromosome 14 syndrome" is emerging; however, the contribution of the mosaic trisomy 14 to the phenotype cannot be evaluated. The study of de novo Robertsonian translocations of the type reported here should reveal both the extent of UPD in these events and the contribution of particular chromosomes involved in certain phenotypes.     

Deletion of chromosome region 13q14 is transmissible and does not always predispose to retinoblastoma

Summary During routine screening of retinoblastoma patients for esterase D activity in red blood cell lysates a patient was identified with only 50% of normal enzyme activity. Chromosome analysis showed that this patient had a small deletion within chromosome region 13q14. Parental studies showed that, whereas the father had normal enzyme levels, the mother had esterase D levels which were also 50% of normal and a similar small 13q14 deletion. Ophthalmological examination failed to demonstrate any retinal abnormality in either parent. Thus wer present the first case not only of the direct transmission of a 13q14 deletion within a family but also of an individual in whom the deletion has not predisposed to tumour formation.     

Separation of retinoblastoma and esterase D loci in a patient with sporadic retinoblastoma and del(13)(q14.1q22.3)

Summary A chromosome 13 deletion in a patient with sporadic retinoblastoma appears to have separated the loci for retinoblastoma and esterase D. This study indicates that: (1) the retinoblastoma locus is distinct from the esterase D locus; and (2) the linear order of these genes is centromere-esterase D-retinoblastoma.