Application of multicolor fluorescent in situ hybridization for enhanced characterization of chromosomal abnormalities in congenital disorders.

OBJECTIVE: To determine the efficacy of multicolor fluorescent in situ hybridization (M-FISH), which paints each chromosome in a unique color, for identification of congenital derivative and marker chromosomes. MATERIAL, METHODS AND CASES: Commercially available M-FISH probes were used to label each chromosome in a specific fluorescent color. Six representative cases involving derivative chromosomes, markers, and subtle anomalies were analyzed by M-FISH. RESULTS: Three familial, rather subtle derivative chromosomes were identified by M-FISH with relative ease. A small ring that was unidentifiable by banded-chromosome analysis was identified by M-FISH. A case of a subtle telomeric anomaly could not be resolved without the use of telomeric-specific probes. The M-FISH results were confirmed by individual chromosome-specific painting probes. CONCLUSION: M-FISH was helpful for identifying a wide range of congenital chromosomal anomalies. However, for subtle chromosomal abnormalities, use of locus-specific probes may be necessary.

Ventricular noncompaction and distal chromosome 5q deletion.

We describe a 7 1/2-year-old girl with mildly unusual phenotype and complex heart disease including ventricular myocardial noncompaction. She was found to have a distal 5q deletion, del(5)(q35.1q35.3). Fluorescent in situ hybridization showed that this deletion included the locus for the cardiac specific homeobox gene, CSX. This suggests that some instances of ventricular myocardial noncompaction may be caused by haploinsufficiency of CSX.

Detection of mosaicism in amniotic fluid cultures: a CYTO2000 collaborative study.

PURPOSE: To evaluate the assumptions on which the American College of Medical Genetics (ACMG) Standards and Guidelines for detecting mosaicism in amniotic fluid cultures are based. METHODS: Data from 653 cases of amniotic fluid mosaicism were collected from 26 laboratories. A chi-square goodness-of-fit test was used to compare the observed number of mosaic cases with the expected number based on binomial distribution theory. RESULTS: Comparison of observed data from the in situ colony cases with the expected distribution of cases detected based on the binomial distribution did not reveal a significant difference (P = 0.525). CONCLUSIONS: The empirical data fit the binomial distribution. Therefore, binomial theory can be used as an initial discussion point for determining whether ACMG Standards and Guidelines are adequate for detecting mosaicism.

Characterization of neo-centromeres in marker chromosomes lacking detectable alpha-satellite DNA.

Recent studies have implicated alpha-satellite DNA as an integral part of the centromere, important for the normal segregation of human chromosomes. To explore the relationship between the normal functioning centromere and alpha-satellite DNA, we have studied eight accessory marker chromosomes in which fluorescence in-situ hybridization could detect neither pancentromeric nor chromosome-specific alpha-satellite DNA. These accessory marker chromosomes were present in the majority of or all cells analyzed and appeared mitotically stable, thereby indicating the presence of a functional centromere. FISH analysis with both chromosome-specific libraries and single-copy YACs, together with microsatellite DNA studies, allowed unequivocal identification of both the origin and structure of these chromosomes. All but one of the marker chromosomes were linear mirror image duplications, and they were present along with either two additional normal chromosomes or with one normal and one deleted chromosome. Indirect immunofluorescence analysis revealed that the centromere protein CENP-B was not present on these markers; however, both CENP-C and CENP-E were present at a position defining a 'neo-centromere'. These studies provide insight into a newly defined class of marker chromosomes that lack detectable alpha-satellite DNA. At least for such marker chromosomes, alpha-satellite DNA at levels detectable by FISH appears unnecessary for chromosome segregation or for the association of CENP-C and CENP-E at a functional centromere.

Novel allele-specific, post-translational reduction in HLA class I surface expression in a mutant human B cell line.

Human B cell line .220 has a novel defect in HLA class I cell surface expression. Mutant .220 was derived from .184TGr, from which both copies of HLA-A and -B are deleted, and has less surface HLA-C than .184TGr. Transfer of class I genes into .220 revealed allele-specific reductions in surface expression: HLA-A1 and -B8 were 1 to 21% of normal; HLA-A11, -A24, and -B5 were moderately reduced; and HLA-A2, -A3, and -B7 were reduced little, if at all. Class I mRNA in .220(A1) and .220(B8) transferents is normal in size and at least normal in quantity. Surface expression of class I molecules was restored by fusing .220 transferents with mutant .174, which lacks the TAP-1 and -2 genes needed for transport of class I-binding peptides. Fusion of .220(A1) cells with beta 2-microglobulin-deficient Daudi cells also fully restored surface expression of class I molecules encoded by both parental cells, indicating beta 2-microglobulin is functional in .220. Pulse-chase experiments showed transgene-encoded HLA-A1 and -B8 alpha-chains are made in apparently normal amounts and associate with beta 2-microglobulin in .220. However, post-translational processing of the HLA-A1 and -B8 molecules is retarded in or before the Golgi apparatus, and immunoprecipitable HLA-A1 molecules disappear after their synthesis. The effects of these abnormalities on surface expression of class I molecules were reversed by incubating .220(A1) and .220(B8) cells at 21 degrees C, which greatly increased the amounts of cell surface HLA-A1 and -B8.

De novo translocation involving chromosomes 1 and 4 resulting in partial duplication of 4q and partial deletion of 1p.

We describe an infant boy with a unique de novo translocation involving chromosomes 1 and 4, resulting in dup(4q) and del(1p). His karyotype was 46,XY,-1,+der(1)t(1;4) (p36.2;q31.2). He had minor anomalies, congenital heart defect, respiratory distress, seizures, and central nervous system abnormalities. He died at age 11 weeks. The patient had manifestations of dup(4q) del(1p), and he was more seriously affected than patients having only one of these. No other patient with an identical chromosomal finding has been reported.

Monosomy 18q12.1----21.1: a recognizable aneuploidy syndrome? Report of a patient and review of the literature.

This report of a patient with an interstitial deletion 18q and review of previously described cases suggest a clinically recognizable syndrome. The phenotype appears to result from a microdeletion of part of 18q12.2 or q12.3, or a deletion of parts of both bands.

Absence of predictable phenotypic expression in proximal 15q duplications.

We describe ten individuals with an insertional duplication 15q12----q13. Phenotypic analysis of these individuals and 15 previously reported cases of proximal 15q duplications fails to show any consistent clinical manifestations. It appears that a duplication of this region is phenotypically silent.

Homozygous tyrosinase gene mutation in an American black with tyrosinase-negative (type IA) oculocutaneous albinism.

We have identified a tyrosinase gene mutation in an American black with classic, tyrosinase-negative oculocutaneous albinism. This mutation results in an amino acid substitution (Cys----Arg) at codon 89 of the tyrosinase polypeptide. The proband is homozygous for the substitution, suggesting that this mutation may be frequently associated with tyrosinase-negative oculocutaneous albinism in blacks.

Simulation of steady cardiovascular flow in the presence of stenosis using a finite element method.

Stenosis could affect one or more segments of the human cardiovascular system. It is a problem capable of causing grave effects. In the present study, the finite element method has been utilised to construct a computer simulation model for the human cardiovascular system in which one or more blood carrying elastic segments are affected by stenosis. Computational effects on the effects of stenosis in aorta arch, carotid, and coronary arteries on parameters of steady flow through the system are presented. It is found that when the total flow rate through the heart is maintained constant, the most notable effect is a very marked increase in pressure drop occurring over the length of the vessel affected with stenosis. Pressure drop in many other segments also increases but by a much smaller extent. On the other hand, when the pressure at the inlet of the ascending aorta and the outlet of the vena cava are maintained constant, the most marked effect is a decrease of flow rate through the stenosed vessel. Stenosis not only causes a pressure drop in the affected segments but it also changes pressures at points distal from the site of stenosis. It also causes a redistribution of flow through the cardiovascular system.

Gene complementing xeroderma pigmentosum group A cells maps to distal human chromosome 9q.

Phenotypic complementation of xeroderma pigmentosum group A (XP-A) cells by microcell-mediated transfer of a single rearranged neo-tagged human chromosome from a human-mouse somatic cell hybrid designated K3SUB1A9-3 was reported previously. Extended growth of this human-mouse hybrid in culture led to deletion of the small arm of the human chromosome, with concomitant loss of complementing ability when introduced into XP-A cells by microcell-mediated chromosome transfer. Cytogenetic analysis of both hybrids suggests that the complementing locus is on chromosome 9q22.2-q34.3, and Southern blot analysis confirms the presence of distal chromosome 9q sequences.

Steady flow of a viscous fluid through a network of tubes with applications to the human arterial system.

The paper presents a finite-element model for the analysis of steady flow of a viscous fluid through a connected system of elastic tubes with the aim of simulating the conditions of blood flow through the human arterial system. The governing equations of the model are non-linear in character and are solved through an iterative computational procedure. This model is capable of incorporating the effects of stenosis on flow and pressure. Typical results are presented and discussed. Quantitative results have been obtained on blood flow through a model of the human arterial system corresponding to the sets of prescribed conditions at the terminations. Also computational results on the effect of stenosis in typical arteries of the system are presented.

Blood flow through the human arterial system in the presence of a steady magnetic field.

The effects of the interaction between a magnetic field and the haemodynamics of the arterial system have been studied. An analysis of a supine subject has been carried out in the presence of an externally applied magnetic field. A finite-element technique has been used to solve the magnetohydrodynamics of the fluid flow problem in a network of rigid tubes. The analysis is carried out by considering arteries as rigid tubes, i.e. the arterial expansion is neglected. In real situations, the arteries are elastic. The method requires the derivation of an expression of the conductance of a single artery in the presence of a transverse magnetic field. Computational results corresponding to two different sets of boundary conditions have been obtained. The quantitative effects of intensity and orientation of the applied magnetic field in the presence and absence of stenosis in the aortic arch are presented and discussed.

Mapping of prolactin and tumor necrosis factor-beta genes on human chromosome 6p using lymphoblastoid cell deletion mutants.

A collection of human B lymphoblastoid cell lines (LCLs) was used to map two genetic sequences for which polymorphism had not been identified: human prolactin (PRL) and tumor necrosis factor-beta (TNFB). The LCLs have overlapping deletions on chromosome 6p produced by gamma-irradiation of LCL 721. After using two chromosome 6p sequences for which LCL 721 is heterozygous to validate our scanning densitometry (SD) method for inferring gene copy number, SD was used to map TNFB and PRL. TNFB maps to the interval between the C4 complement and HLA-B loci within the MHC on chromosome 6p. PRL lies within the 6p21.3-6p22.2 interval distal to HLA-C. We found that LCL 721 is heterozygous for PRL DNA fragment lengths generated by HpaII but not MspI digestion, indicating that the two copies of PRL in LCL 721 are differentially methylated. This novel methylation RFLP was used to corroborate the region PRL assignment.