Investigation of an N-Terminal Prohormone of Brain Natriuretic Peptide Point-of-Care ELISA in Clinically Normal Cats and Cats With Cardiac Disease.

BACKGROUND: N-terminal prohormone of brain natriuretic peptide (NT-proBNP) concentrations may be increased in cats with various cardiac disorders. The point-of-care (POC) ELISA assay uses the same biologic reagents as the quantitative NT-proBNP ELISA. Previous studies have evaluated the sensitivity and specificity of the POC ELISA in cats with cardiac disease. OBJECTIVES: To prospectively evaluate the diagnostic utility of the POC ELISA in a select population of cats. ANIMALS: Thirty-eight client-owned cats presented to the University of Florida Cardiology Service for cardiac evaluation. Fifteen apparently healthy cats recruited as part of another study. METHODS: Physical examination and echocardiography were performed in all cats. The POC ELISA was assessed visually as either positive or negative by a reader blinded to the echocardiographic findings, and results were analyzed relative to quantitative assay results. RESULTS: Twenty-six cats were diagnosed with underlying cardiac disease, and 27 cats were considered free of cardiac disease. Cats with cardiac disease included: 21 with hypertrophic cardiomyopathy, 2 with unclassified cardiomyopathy, 2 with restrictive cardiomyopathy, and 1 with 3rd degree atrioventricular (AV) block. The POC ELISA differentiated cats with cardiac disease with a sensitivity of 65.4% and specificity of 100%. CONCLUSIONS AND CLINICAL IMPORTANCE: The POC NT-proBNP ELISA performed moderately well in a selected population of cats. A negative test result cannot exclude the presence of underlying cardiac disease, and a positive test result indicates that cardiac disease likely is present, but further diagnostic investigation would be indicated for a definitive diagnosis.

Temperature-sensitive transformation by an Abelson virus mutant encoding an altered SH2 domain.

Abelson murine leukemia virus (Ab-MLV) encodes the v-Abl protein tyrosine kinase and induces transformation of immortalized fibroblast lines and pre-B cells. Temperature-sensitive mutations affecting the kinase domain of the protein have demonstrated that the kinase activity is absolutely required for transformation. Despite this requirement, mutations affecting other regions of v-Abl modulate transformation activity. The SH2 domain and the highly conserved FLVRES motif within it form a phosphotyrosine-binding pocket that is required for interactions between the kinase and cellular substrates. To understand the impact of SH2 alterations on Ab-MLV-mediated transformation, we studied the Ab-MLV mutant P120/R273K. This mutant encodes a v-Abl protein in which the beta B5 arginine at the base of the phosphotyrosine-binding pocket has been replaced by a lysine. Unexpectedly, infection of NIH 3T3 or pre-B cells with P120/R273K revealed a temperature-dependent transformation phenotype. At 34 degrees C, P120/R273K transformed about 10-fold fewer cells than wild-type virus of equivalent titer; at 39.5 degrees C, 300-fold fewer NIH 3T3 cells were transformed and pre-B cells were refractory to transformation. Temperature-dependent transformation was accompanied by decreased phosphorylation of Shc, a protein that interacts with the v-Abl SH2 and links the protein to Ras, and decreased induction of c-Myc expression. These data suggest that alteration of the FLVRES pocket affects the ability of v-Abl to interact with at least some of its substrates in a temperature-dependent fashion and identify a novel type of temperature-sensitive Abelson virus.

Deletional mapping of fifteen mouse VH gene families reveals a common organization for three Igh haplotypes.

In addition to the content of germ-line variable gene segments, the organization of V genes has been implicated in the development of the Ab repertoire. We have searched the expressed VH genes of BALB/c mice for additional VH gene families and utilized deletion mapping to explore the extent of VH gene family interspersion. We have identified and characterized one new VH gene family (VH15) and extended our previous studies of the Igha and Ighb haplotypes to include a third haplotype (Ighj) using a newly developed panel of pre-B cell lines (CXCB cell lines). We conclude that the Igha, Ighb, and Ighj haplotypes have a similar Igh-V locus structure. A refined deletional map for 15 VH gene families and an individual member of the VHSM7 family (H10) has been constructed based on the deletion profiles of 72 rearranged heavy chain loci. These results demonstrate previously unrecognized examples of interspersion among members of the VHS107, VH10, and VHSM7 families.

The utilization of individual VH exons in the primary repertoire of adult BALB/c mice.

The utilization of VH gene families is severely biased during fetal development, such that D-region proximal VH genes are overrepresented. After birth the VH repertoire becomes more equally representative of the total inherited set of functional VH genes. To investigate the extent of this normalization process, we have determined the relative utilization of individual VH exons in the pre-immune repertoire of adult BALB/c spleen cells. Large samples of IgM heavy chain transcripts from polyclonally activated B cells were captured in a cDNA phage library and screened by hybridization using highly specific oligonucleotide probes. These studies revealed that the utilization of particular VH exons can differ by an order of magnitude. Significantly, the VH genes most under-represented in the pre-immune repertoire are located in the region of the Igh locus most distal to the DjhCh region. We suggest that the chromosomal position of a particular VH gene may influence its utilization and that the normalization of the adult repertoire is incomplete.

Simultaneous determination of disopyramide and its mono-N-dealkyl metabolite in plasma and urine by high-performance liquid chromatography.

A high-performance liquid chromatographic procedure is described for the determination of disopyramide and its mono-N-dealkyl metabolite which offers simplicity of extraction with excellent selectivity, sensitivity and reproducibility. The drug and metabolite, following basic diethyl ether extraction and back-extraction with acetic acid, are injected into a reversed-phase high-performance liquid chromatographic column and the absorbance of the eluate measured at 254 nm. Detectability limits of 0.05 micrograms/ml were obtained with both compounds, and studies of the reproducibility, precision, recovery, stability during storage and effect of time in separating plasma from erythrocytes are described. Applications of this high-performance liquid chromatographic procedure to plasma samples from patients on disopyramide therapy and to plasma and urine from a healthy dog administered single doses are reported.

Drug permeation through membranes V: interaction of diazepam with common excipients.

The effects of common tablet excipients on the permeation of diazepam through polydimethylsiloxane membranes and on the turnover time of goldfish were studied. The permeability coefficient decreased and the turnover time increased in the presence of talc, polysorbate 80, and, possibly, fumed silicon dioxide, but these parameters were unaffected by lactose, microcrystalline cellulose, and starch.