Vitamin A, differentiation and cancer.

Retinoids, which are derivatives of vitamin A, have a variety of effects on normal cellular differentiation and on the process of carcinogenesis. A number of novel endogenous retinol metabolites have been identified recently. The response of many cell types to retinoid treatment is mediated by retinoid receptors, and involves changes in gene expression, cell growth and cell differentiation. The gene encoding one of the retinoic acid receptors is disrupted by the chromosome translocations associated with acute promyelocytic leukemia, and the expression of another is altered in epithelial tumors; both of these findings have important implications for the use of retinoids as anti-carcinogenic agents. It has been demonstrated recently that certain homeobox genes are regulated by retinoids; these genes may also prove to be useful agents for anti-carcinogenic therapies.

Chromosome microarray analysis in a clinical environment: new perspective and new challenge.

The analysis of the human genome has largely been undertaken in a research environment, but recent developments in technology and associated workflow have allowed diagnostic laboratories to interrogate DNA at significantly improved levels of resolution. Principally, whole genome-based analysis of copy number changes using microarrays has led to this method replacing conventional karyotyping as a routine diagnostic workhorse. The resolution offered by microarrays is an improvement of at least an order of magnitude compared to karyotyping, but it comes at a cost in terms of the time spent in data interpretation. Overall, however, the die has been cast and cytogeneticists need to become familiar with the tools use by molecular geneticists and bioinformaticists. The following review provides a brief background to array technology, but uses a series of case studies to illustrate the usefulness and challenges of interpreting array data.

Molecular characterisation of a der(Y)t(Xp;Yp) with Xp functional disomy and sex reversal.

Sex reversal due to duplication of the Xp21 dosage-sensitive sex reversal locus results in XY females with gonadal dysgenesis. Pure Xp disomy (without a concurrent loss of genetic material) can occur by translocation or interstitial duplication. The case reported here is the rare form with a t(Xp;Yp). The combination of conventional clinical cytogenetic techniques, microsatellite analysis and high-density microarrays identified the X-chromosome breakpoint as centromeric of the NR0B1 gene and its control elements. Cytogenetics and array technology complemented each other in characterizing the translocation event and the extent of the dosage-sensitive sex reversal critical region on the derivative Y-chromosome. The implications of this analysis also lie in genetic counseling that highlight the likely de novo nature of a paternal meiotic event.

Crosses of NOD mice with the related NON strain. A polygenic model for IDDM.

Chromosome locations of non-major histocompatibility complex (MHC) genes contributing to insulin-dependent diabetes mellitus (IDDM) in mice have been determined by outcrossing NOD mice to other inbred strains congenic for the NOD MHC haplotype (H2g7). At least nine non-MHC IDDM susceptibility genes (Idd) were previously identified at first backcross (BC1) after outcross of NOD to C57BL/10.H2g7 congenic mice (B10.H2g7). We investigated whether the same set of Idd loci segregated with IDDM susceptibility after outcross of NOD to NON.H2g7 congenic mice. Since the outcrosses to NON.H2g7 and B10.H2g7 were performed in the same vivarium, direct comparisons were made of the chromosomal locations and relative strengths of Idd alleles in diabetic progeny from the two different outcrosses. In comparison with the NOD x B10.H2g7 outcross, the NOD x NON.H2g7 outcross produced significantly higher IDDM frequencies in F1, F2, and BC1 generations. The high F2 diabetes frequency allowed evaluation of the effects of homozygous expression of both the susceptibility and the resistance allele at Idd loci. This analysis demonstrated that no single non-MHC Idd locus was essential for the onset of diabetes in this cross. After outcross to NON.H2g7, Idd4 (chromosome [Chr] 11), Idd5 (Chr 1), and Idd8 (Chr 14) did not segregate with IDDM in either the BC1 or the F2 generation. Diabetogenic NOD-derived alleles at Idd2 (Chr 9), Idd3 (Chr 3), and Idd10 (Chr 3) were segregating in the BC1. An NON-derived allele contributing to susceptibility on Chr 7 (Idd7) was also detected. Dominant traits, detectable only in the F2 cross, were encoded by Chr 4 (Idd9) and two newly mapped loci on Chr 13 (Idd14) and 5 (Idd15). A third dominant trait was encoded by Chr 6 (possibly Idd6), but here, in contrast to Idd9, Idd14, and Idd15, the NON allele was diabetogenic. Stepwise logistic regression analysis of the BC1 and F2 data confirmed that the ability to identify certainty of the non-MHC Idd loci was contingent on the extent of homozygosity for NOD background genes. This study shows that the diabetogenic phenotype can be achieved through the actions of variable combinations of MHC-unlinked genes and a diabetogenic MHC haplotype.

The effect of hemodialysis and hemoperfusion on serum valproic acid concentration.

We report a patient with dialysis-induced encephalopathy who was taking divalproex sodium for a seizure disorder. Her serum valproic acid concentration appeared to be in the low therapeutic range at 54 mg/l yet she continued to have seizure activity. The elimination half-life and apparent clearance of valproic acid were the same for both a dialysis and nondialysis day, indicating that hemodialysis/hemoperfusion has little effect on the overall removal of valproic acid from the body.

Developmental changes in California Card Sorting Test performance.

The purpose of the present study was to determine 1) if performance on the California Card Sorting Test (CCST) follows the same developmental gradient as other measures of concept formation and 2) whether the components of concept formation tapped by the CCST are developmentally dissociable. Participants were 68 children and young adults in four age-based groups: 7 to 9 years (n = 13); 10 to 12 years (n = 16); 17 to 19 (n = 20); and, 20 to 22 years (n = 19). All were of average or higher measured intelligence and screened for neurological, psychiatric, reading and attentional disorder. The findings of the present study suggest that like many other concept formation tasks, CCST performance approximates adult levels by age 10. Further, the different components of concept formation measured by the CCST are dissociable in the youngest children with the development of concept recognition preceding sorting ability.

Percutaneous aortic valvuloplasty procedure and implications for nursing.

Percutaneous transluminal aortic balloon valvuloplasty (PTABV) is a procedure performed in the cardiac catheterization laboratory for some patients with severe calcific aortic stenosis. A review of the literature indicated that the procedure was beneficial to elderly patients or those who were poor candidates for surgery. The common complications of PTABV include bleeding at the catheterization site and vagal reactions during groin compression. Nursing implications in the cardiac catheterization laboratory involve ensuring patient comfort, managing the effects of cardiac instrumentation, and monitoring vital signs and vascular integrity. Post-PTABV nursing diagnoses include: (1) potential for injury or hemorrhage, (2) potential for alteration in cardiac output, (3) potential for alteration in tissue perfusion, (4) alteration in comfort, (5) potential for infection, and (6) knowledge deficits.

Pair-rule gene expression in the somitic stage chick embryo: association with somite segmentation and border formation.

In vertebrates, metameric organization is high-lighted by the formation of somites from mesenchymal cells of the segmental plate which then differentiate into dermamyotomal and sclerotomal tissues. The resegmentation of the sclerotome into rostral and caudal halves follows, coincident with the production of specific extracellular matrix molecules at the abutment of these two cell types. Ultimately, cells from the caudal sclerotome migrate ventrally and contribute to the chondrogenic prevertebrae. The objective of this work is to investigate the molecular steps regulating these events. Our study is focused on the paired-box containing genes, which have been implicated in delineating boundaries early in development. A chick embryo system, which is readily accessible to manipulation and observation during early development, is used in this study. We have identified the existence of the paired-box motif in the chicken genome by polymerase chain reaction and hybridization with the mouse Pax 1 paired-box sequence. Expression of paired-box genes occurs early in development as shown by Northern analysis, and is localized by in situ hybridization to the edge of each somite, a patch at the central core of each somite, and the periphery of the neural tube. This specific spatial pattern of expression is consistent with the hypothesis that the pair-rule genes function as effecters of border formation in the early embryo. Moreover, the patch of positive cells at the center of a resegmenting somite appear to migrate ventrally, and may contribute to structures of the prevertebrae. These findings are relevant to our understanding of the mechanism of somite resegmentation and implicate the involvement of pair-rule genes in the process.

The molecular mechanism of phase variation of H. influenzae lipopolysaccharide.

Multiple carbohydrate structures on the outer-membrane lipopolysaccharide (LPS) of the gram-negative pathogen H. influenzae undergo high frequency, reversible loss, indicative of phase variation. Characterization of a genetic locus, lic-1, responsible for expression of two LPS epitopes displaying phase variation, showed it to comprise four genes. The first gene mediates phase variation. At its 5' end, within the open reading frame, are a variable number of tandem repeats of the tetramer CAAT. By shifting upstream initiation codons in or out of frame, these 4 bp units create a translational switch. The phenotype of organisms corresponds to the number of 4 bp units. Phase variation between three levels of expression ( +, +, and -) of lic-1-derived epitopes is caused by differences in the three phases of translation of the 5' terminus of this gene. Phase variation also allows for selection of organisms displaying certain LPS epitopes in vivo.

Programs that mitigate the effects of poverty on children.

This article reviews six federally funded in-kind public assistance programs that are intended to mitigate the effects of poverty on low-income children by providing access to basic human necessities such as food, housing, education, and health care. The evidence suggests that, while each program can be improved, these programs do achieve their basic objectives. In general, food stamps, the Special Supplemental Food Program for Women, Infants, and Children (WIC), and school nutrition programs are successful at providing food assistance to low-income children, starting with the prenatal period and continuing through the school years. The Food Stamp Program provides food assistance nationwide to all households solely on the basis of financial need and is central to the food assistance safety net for low-income children. The WIC program has helped reduce the prevalence of iron-deficiency anemia in infants and children and has increased intakes of certain targeted nutrients for program participants. The school nutrition programs provide free or low-cost meals that satisfy the dietary goals of lunches and breakfasts to most school-age children. The Medicaid program has extended health insurance coverage to millions of low-income children. However, many children remain uninsured, and children enrolled in Medicaid do not have the same access to medical care as privately insured children. Relatively little is known about the effects of Medicaid on children's health status. For Head Start, empirical evidence suggests that participating children show enhanced cognitive, social, and physical development in the short term. Studies of the longer-term impacts of Head Start are inconclusive. Although housing assistance improves housing quality and reduces housing costs for recipients, there is a large unmet need for acceptable, affordable housing among poor families. Important gaps remain in our knowledge of the effects of these programs on the well-being of children. Questions regarding a program's effects over time on health and developmental outcomes particularly need more study.

In vitro characterization of chondrogenic cells isolated from chick embryonic muscle using peanut agglutinin affinity chromatography.

Specific binding to the lectin, peanut agglutinin (PNA), has been reported in embryonic precartilage tissues, including the condensing limb bud blastema and the caudal half of the developing somite. The present study aimed to test the hypothesis that PNA-binding may be a surface characteristic of chondroprogenitor cells residing within noncartilage tissues, such as muscle, which have the potential of being induced to form cartilage, e.g., in the presence of bone matrix-derived factors. Day-14 chick embryonic pectoral muscle, which contained histochemically detectable PNA-binding cells, was dissociated into single cells (TM cells) and fractionated by PNA affinity chromatography into PNA-binding (PNA+) and nonbinding (PNA-) cells by PNA-Sepharose 6 MB affinity chromatography. The differentiation potential of the PNA-affinity fractionated cells in vitro was analyzed as a function of culture plating cell density. Immunohistochemistry of a number of cell-type-specific differentiation markers, including sarcomeric actin, collagen type II, and aggrecan core protein, demonstrated that PNA+ cells, when cultured as a micromass at high density (20 x 10(6) cells/ml), exhibited a chondrocyte-like phenotype, whereas the PNA-cells remained myogenic; however, both PNA+ and PNA- monolayer cultures (4 x 10(4) cells/ml) behaved as myoblastic cells. The expression of collagen type II mRNA was also confirmed by coupled reverse transcription/polymerase chain reaction analysis. These observations suggest that PNA binding, i.e., the presence of specific galactose-containing cell surface moieties, is likely to be one of the characteristics of chondrogenic cells residing in mesenchymally derived embryonic tissues.

Evaluation of monoclonal antibodies for subtyping of currently circulating human type A influenza viruses.

The hemagglutinin subtype specificities of six monoclonal antibodies (MAbs) to influenza type A viruses were evaluated in a rapid culture assay by immunoperoxidase staining. Confluent monolayers of MDCK cells in multiwell plates were inoculated with (i) 23 reference viruses, (ii) 200 isolates collected during the influenza season 1995 to 1996, and (iii) 28 clinical specimens previously found to be influenza virus positive. After overnight incubation, the cells were fixed and stained with MAbs IVA1/B10, IIF4/D3, 12L/5, 13L/6, 18L/1, or 18L/4. Type-specific MAbs were included as controls. All antibodies gave intensive cytoplasmic staining with infected cells in the absence of any reaction with uninfected cells. MAbs 12L/5, 13L/6, 18L/1, and 18L/4 exclusively reacted with viruses of the subtype H1, and the antibodies IVA1/B10 and IIF4/D3 exclusively reacted with viruses of the subtype H3. None of these MAbs reacted with viruses of the H2 subtype or with influenza type B viruses. Of the 200 recent isolates, 63 were identified as influenza virus type A, subtype H1, 95 were identified as type A, subtype H3, and 41 were identified as type B. One isolate contained a mixture of a type A (H3) and a type B influenza virus. Of the 28 previously positive clinical specimens, 15 contained an influenza virus A, subtype H3, 1 contained an influenza virus A, subtype H1, and 9 contained an influenza B virus. The subtype of a very weakly positive specimen could not be determined, and two specimens remained negative. The MAbs described here allow for a rapid typing and subtyping of influenza virus isolates and for the type- and subtype-specific detection of influenza viruses in clinical specimens.

Antigenic proteins of Mycobacterium leprae. Complete sequence of the gene for the 18-kDa protein.

Recombinant clones expressing antigenic determinants of the 18-kDa protein antigen from Mycobacterium leprae recognized by the L5 monoclonal antibody were isolated from a lambda gt11 expression library and their nucleotide sequences determined. All clones expressed the M. leprae-specific determinant as part of a large fusion protein with Escherichia coli beta-galactosidase. The deduced amino acid sequence of the coding region indicated that all the lambda gt11 recombinant clones contained an incomplete M. leprae gene sequence representing the carboxy-terminal two-thirds (111 amino acids) of the 18-kDa gene and coding for a peptide of m.w. 12,432. Subsequent isolation and sequencing of a 3.2kb BamHI-PstI DNA fragment from a genomic M. leprae cosmid library permitted the deduction of the complete 148 amino acid sequence with a predicted m.w. of 16,607. A second open reading frame 560 bases downstream from the 18-kDa coding sequence was found to code for a putative protein of 137 amino acids (m.w. = 15,196). Neither this nor the 18-kDa amino acid sequence displayed any significant homologies with any proteins in the GENBANK, EMBL, or NBRF data bases. Crude lysates from recombinant lambda gt11 clones expressing part of the 18-kDa protein have been reported to stimulate the proliferation of some M. leprae-specific helper T cell clones. Thus, it is significant that the complete 18-kDa sequence contains five short peptides predicted to be possible helper T cell antigenic epitopes based on their propensity to form amphipathic helices. Although three of these occur within the 111 amino acid carboxy-terminal peptide expressed by lambda gt11 clones, the most highly amphipathic peptide is found in the amino-terminal region not present in the lambda gt11 recombinants.

Towards construction of a high resolution map of the mouse genome using PCR-analysed microsatellites.

Fifty sequences from the mouse genome database containing simple sequence repeats or microsatellites have been analysed for size variation using the polymerase chain reaction and gel electrophoresis. 88% of the sequences, most of which contain the dinucleotide repeat, CA/GT, showed size variations between different inbred strains of mice and the wild mouse, Mus spretus. 62% of sequences had 3 or more alleles. GA/CT and AT/TA-containing sequences were also variable. About half of these size variants were detectable by agarose gel electrophoresis. This simple approach is extremely useful in linkage and genome mapping studies and will facilitate construction of high resolution maps of both the mouse and human genomes.

Carbamazepine clearance in hemodialysis and hemoperfusion.

A 47-year-old woman with endstage renal disease and dialysis-induced encephalopathy was being treated with carbamazepine for myoclonus. Her carbamazepine serum concentration appeared to be therapeutic at 5.1 micrograms/ml. She experienced a seizure while on hemodialysis/hemoperfusion that was possibly related to the removal of carbamazepine during dialysis. The elimination of carbamazepine on a dialysis day was compared with elimination on a nondialysis day. The half-life and apparent clearance were the same for each day, indicating that hemodialysis/hemoperfusion had little effect on the overall removal of carbamazepine from the body. The possible reasons for this lack of effect are discussed.

Additional microsatellite markers for mouse genome mapping.

Mouse sequence information from the EMBL and GenBank databases, published sequences and genomic clones have been analyzed for simple repetitive elements or microsatellites. Each microsatellite has been amplified by the polymerase chain reaction (PCR) as a single locus marker. PCR primers were designed from unique sequence flanking each repeat. Size variation of PCR products less than 750 base pairs (bp) between mouse strains has been determined using ethidium bromide-stained acrylamide or agarose gels. A further 74 newly characterized microsatellites are presented in this paper, bringing to 185 the total we have analyzed. Of these, 157/185 (85%) have more than one allele, 143/178 (80%) vary in length between C57BL/6J and Mus spretus, and 82/168 (49%) vary between DBA/2J and C57BL/6J. Microsatellites provide informative single locus probes for linkage analysis in the construction of a genetic map of the mouse genome.

Genetic analysis of autoimmune type 1 diabetes mellitus in mice.

Two genes, Idd-3 and Idd-4, that influence the onset of autoimmune type 1 diabetes in the nonobese diabetic mouse have been located on chromosomes 3 and 11, outside the chromosome 17 major histocompatibility complex. A genetic map of the mouse genome, analysed using the polymerase chain reaction, has been assembled specifically for the study. On the basis of comparative maps of the mouse and human genomes, the homologue of Idd-3 may reside on human chromosomes 1 or 4 and Idd-4 on chromosome 17.