Immunological and biophysical properties of hepatitis B antigen labeled by the chloramine-T and by the lactoperoxidase methods.

Optimal conditions were sought for the radiolabeling of microgram quantities of hepatitis B surface antigen (HBs Ag) employing the chloramine-T or lactoperoxidase iodination procedures. Preparations of HBsAg labeled by these procedures are referred to as chloramine-T preparations and lactoperoxidase preparations, respectively. Labeled HBsAg having specific activities between 10-20 muCi/mug were found to display the greatest degree of sensitivity for unlabeled HBsAg and for anti-HBs using a double-antibody radioimmunoassay (RIA-DA). Increasing the specific activity above this level redulted in a decreased affinity of labeled 1251-HBs Ag for anti-HBs, indicating that soluble antigenic alterations had developed. At equivalent specific activities, chloramine-T preparations competed less effectively for unlabeled HBs Ag than lactoperoxidase preparations, and anti-HBs endpoint titers were slightly reduced, especially among preparations of high specific activity (greater than or equal to 65 muCi/mug). Chloramine-T preparations of HBs Ag (sp. act. 15--30 muCi/mug) showed essentially no antigenic deterioration over a 2-month period at minus 196 degrees C or minus 70 degrees C. Utilization of optimally labeled 1251-HBs Ag has increased the sensitivity of the RIA-DA for unlabeled HBs Ag 30-fold to a level below 1 ng/ml and enhanced antiamine-T method revealed that only the most acidic population was labeled (pH 3.75+/-0.5). In contrast, six antigenic components with distinct pI values ranging from 3.7 to 5.2 were detected by RIA-DA in both unlabeled HBs ag and in the chloramine-T preparation. This indicated that the chloramine-T method did not radically change the relative number or charge of each of the pI populations present in purified preparations of HBs Ag. Analysis of HBs Ag iodinated by the lactoperoxidase procedure revealed the presence of three of four populations of particles with pI values ranging from 3.9 to 4.5, suggesting that this procedure labels HBs Ag more uniformly.     

The mutational spectrum of brachydactyly type C

Growth/differentiation factor-5 (GDF5), also known as cartilage-derived morphogenetic protein-1 (CDMP-1), is a secreted signaling molecule that participates in skeletal morphogenesis. Heterozygous mutations in GDF5, which maps to human chromosome 20, occur in individuals with autosomal dominant brachydactyly type C (BDC). Here we show that BDC is locus homogeneous by reporting a GDF5 frameshift mutation segregating with the phenotype in a family whose trait was initially thought to map to human chromosome 12. We also describe heterozygous mutations in nine additional probands/families with BDC and show nonpenetrance in a mutation carrier. Finally, we show that mutant GDF5 polypeptides containing missense mutations in their active domains do not efficiently form disulfide-linked dimers when expressed in vitro. These data support the hypothesis that BDC results from functional haploinsufficiency for GDF5.     

The protective effect of antibody in immunity to murine chlamydial genital tract reinfection is independent of immunoglobulin A

The resolution of primary and secondary chlamydial genital infection in immunoglobulin A (IgA)-deficient (IgA(-/-)) mice was not different from that in IgA(+/+) mice. Furthermore, depletion of either CD4(+) or CD8(+) T cells prior to reinfection of IgA(+/+) or (-/-) mice had limited impact on immunity to reinfection. Thus, although antibody contributes importantly to immunity to chlamydial genital tract reinfection, IgA antibodies are not an absolute requirement of that protective response.     

Comparison of colorimetric,fluorometric, and visual methods for determining anti-influenza (H1N1 and H3N2) virus activities and toxicities of compounds

Methods have been developed previously for rapid evaluation of compounds for antiviral activity in 96-well microplates, which include visual quantitation of antiviral activity based upon inhibition of virus-induced cytopathic effect (CPE) or by less subjective colorimetric or fluorometric means. In the present studies we compared a number of colorimetric (crystal violet, MTT [3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide], and neutral red) and fluorometric (Alamar Blue, bisbenzimide [Hoechst 33258], fluorescein diacetate, and rhodamine 6G) methods to visual scoring of antiviral activity in influenza A virus infections in Madin Darby canine kidney (MDCK) cells. Toxicity determinations using these same methods were also made for anti-influenza inhibitors and other compounds known to inhibit cell proliferation. Against influenza A/Texas/36/91 (H1N1) and A/Sydney/05/97 (H3N2) viruses, visual scoring and dye or stain methods produced results that were not significantly different from each other in deriving 50% virus-inhibitory concentrations (EC(50) values) for six anti-influenza compounds (amantadine, rimantadine, ribavirin, RWJ-270201 [BCX-1812], oseltamivir carboxylate, and zanamivir), with the exception of Alamar Blue which quantified lower EC(50) values than expected. In uninfected replicating cells, the visual and Alamar Blue methods underestimated the 50% cytotoxic concentrations (IC(50) values) of ribavirin, 1-beta-D-arabinofuranosylcytosine, and 6-azauridine, but more accurately assessed the toxicities of amantadine, rimantadine, and cycloheximide. Visual scoring, coupled with the use of one of these dyes or stains except Alamar Blue, can be used to accurately and rapidly quantify the anti-influenza virus activities and toxicities of potential new influenza virus inhibitors. These methods should also be applicable to evaluating antiviral effects against other lytic virus infections.     

Colonic biopsy in verotoxin-induced hemorrhagic colitis and thrombotic thrombocytopenic purpura (TTP)

Sporadic cases and occasional outbreaks of hemorrhagic colitis recently have been associated with the rare Escherichia coli serotype O157:H7, which is now recognized as an important identifiable cause of bloody diarrhea in patients in whom more common gut pathogens cannot be detected. The authors report such a case in a 49-year-old woman who developed thrombotic thrombocytopenic purpura (TTP) and hemorrhagic transverse and descending colitis with a lesion having many of the features of pseudomembranous colitis. While pseudomembrane formation has been described in the hemolytic uremic syndrome (HUS), these features have not, to the authors' knowledge, been described in a patient with hemorrhagic colitis and TTP secondary to a verotoxin-producing serotype of E. coli.     

Isolation of embryonic chick myosatellite and pluripotent stem cells

Summary The current study outlines the isolation and culture of two populations of cells derived from Day 11 embryonic chick leg muscle and associated connective tissues. The two populations consisted of myogenic lineage-committed stem cells (myosatellite stem cells) and lineage-uncommitted stem cells (pluripotent stem cells). After long-term culture the lineage-uncommitted stem cell population displayed differentiated phenotypes suggestive of the following adult tissues, fibroblasts, muscle, fat, cartilage, and bone.     

Body composition measures from underwater weighing and dual energy X-ray absorptiometry in black and white girls: A comparative study

The purpose of this study was to compare estimates of body composition in two ethnic groups, 31 black and 38 white girls 10 through 16 years of age, to establish accurate and precise laboratory standards for field measures of body composition in the NHLBI Growth and Health Study HC 55025. The dual energy X-ray absorptiometry (DXA) measures of fat free mass (FFM) and % body fat (%BF) were made using Hologic QDR-1000/W. Corresponding values of FFM and %BF from underwater weighing (UWW) were determined using the two-component model of Siri, and these were corrected using the model of Lohman for white girls only. In the comparison of the different models and methods, the two-component model overestimated FFM compared to estimates from DXA for black girls, as did the corrected Lohman model for white girls. The two-component model significantly overestimated %BF in both white and black girls compared to corresponding estimates from DXA. The ratio of bone mineral content (BMC)/FFM affected the degree of %BF differences in black girls but not in white girls. Also, as the density of FFM increased or approached adult status in black girls (BMC/FFM increased), differences between the two-component model and estimates from DXA decreased. In both groups of girls, the relationship of %BF from UWW and DXA are a function of the level of body fatness. DXA values of %BF are greater than those from UWW under about 24% body fat, but the converse occurs above 25% body fat. The inability of UWW using the two-component model to account for the body composition in these girls can be corrected in part by measuring the variables for a multicomponent model or more easily by using DXA estimates of body composition. © 1994 Wiley-Liss, Inc.     

Ciliary process microvasculature of the primate eye

Using methylmethacrylate microvascular luminal castings, we studied the three-dimensional angioarchitecture of the primate ciliary process with the scanning electron microscope. We found that the ciliary processes are served by vessels that radiate anteriorly and posteriorly from the "major arterial circle" of the iris. The anterior arterioles possess focal constrictions and supply the anterior and marginal aspects of the major ciliary processes as well as interprocess networks that connect contiguous processes. The posterior arterioles lack focal constrictions and supply the minor ciliary processes via posterior interprocess networks. Major and minor ciliary process capillaries are irregularly dilated and pass posteriorly to drain into the choroidal veins. Finally, venous arcades exist which directly connect anterior and posterior interprocess networks with the choroidal veins and thus bypass the ciliary processes entirely. The presence of focal constrictions in the anterior arterioles suggests a site for possible autonomic or neurohumoral control of blood flow into the major ciliary processes.     

Measuring physician practice site characteristics: A comparison of data from SK&A and a practice site survey

ObjectiveTo evaluate the comparability of commercially available practice site data from SK&A with survey data to understand the implications of using SK&A data for health services research.Data sourcesResponses to the Comprehensive Primary Care Plus (CPC+) Practice Survey and SK&A data.Study designComparison of CPC + Practice Survey responses to SK&A information for 2698 primary care practice sites.Data collectionCPC + Practice Survey data collected through a web‐only survey from April through September 2017, and SK&A data purchased in November 2016.Principal findingsInformation was similar across data sources, although some discrepancies were common. For example, 56% of practice sites had differences in the reported number of practitioners, and larger sites tended to have larger differences. Among practice sites with 1 practitioner in the survey, only 1.3% had a difference of 3 or more practitioners between the data sources, whereas 63% of practice sites with 11 or more practitioners had a difference of 3 or more practitioners.ConclusionsDiscrepancies between data sources could reflect differences of interpretation when defining practice site characteristics, changes over time in those characteristics, or data errors in either SK&A or the survey. Researchers using SK&A data should consider possible ramifications for their studies.