Diagnostic role of an immunoassay-detected polymorphism of factor IX for potential carriers of hemophilia B

In hemophilia B, assays based on a monoclonal antifactor IX specific for the Thr-148 variant of an exonic polymorphism have diagnosed carriers in selected families by either establishing linkage or by indicating the presence or absence of a given normal factor IX. The sensitivity of the immunoassays for detecting heterozygous women was explored by comparing results from immunoassays with solid-phase polyclonal v the monoclonal antifactor IXs. Factor IX with the normal Ala-148 variant gave a flat dilution curve, qualitatively distinct from factor IX with the Thr-148 variant in the monoclonal assay. The two were indistinguishable in the polyclonal assay. Mixtures of equal amounts of the two types gave an intermediate result, about half as reactive in the monoclonal as compared with the polyclonal assay system. Whereas mixtures with 10% Ala-148 and 90% Thr-148 factor IXs could not readily be distinguished from Thr-148 factor IX plasma, as little as 1% of the Thr-148 protein was detected in Ala-148 factor IX plasma. The frequency of the Ala-148 variant varied in individuals with different ethnic backgrounds; it was found in 29% of white, 12% of black, and none of Asian blood donors' factor IX genes in Seattle. Only 4% of samples from South African black men were nonreactive (ie, Ala- 148). The Thr/Ala-148 dimorphism is in strong linkage disequilibrium with Taql restriction fragment length polymorphisms (RFLPs). Three recombinations were noted in normal white genes and one in a normal black factor IX gene (less than 2% of those examined). In 34 white families with at least one woman being a possible carrier, genetically, the immunoassay results were informative in 18. RFLP analyses were informative in eight of the 15 families tested. In five families each, assignment of carrier status was made to a woman by only DNA or only immunoassay results, whereas the other approach was noninformative. The immunoassays provide a rapid, inexpensive screening test and complement DNA analysis in white women who are potential carriers of hemophilia B.     

Expression of ICAM-R (ICAM-3), a novel counter-receptor for LFA-1, in rheumatoid and nonrheumatoid synovium

Objective. To study the distribution of intercellular adhesion molecule receptor (ICAM-R, or ICAM-3), a novel ligand for the leukointegrin lymphocyte function-associated antigen 1 (LFA-1), in normal and rheumatoid synovial membranes and to compare this with the distribution of ICAM-1, ICAM-2, vascular cell adhesion molecule 1 (VCAM-1), and endothelial leukocyte adhesion molecule 1 (ELAM-1). Methods. We performed immunohistochemical analyses of frozen sections of normal and rheumatoid synovial tissue using monoclonal antibodies to the molecules examined. Results. ICAM-1 staining was detectable on the vascular endothelium and the synovial lining cells of both normal and rheumatoid synovial membranes. A variable proportion of lymphocytes infiltrating rheumatoid tissues expressed ICAM-1. ICAM-2 staining was demonstrable in the vascular endothelium of both normal and inflamed tissues, the latter demonstrating a significantly higher proportion of positive vessels. ELAM-1 staining was not detectable in normal synovial membranes but was seen on the endothelium of a limited number of rheumatoid synovial vessels, usually close to the synovial lining cell layer. VCAM-1 staining was intense in both normal and rheumatoid synovial lining cells, but vascular staining was weak in both. In contrast, ICAM-R staining was not detected in association with any synovial blood vessels, but was widely expressed by lymphocytes and macrophages. Cells of the lining layer did not stain for ICAM-R. Conclusion. Although ICAM-R is a ligand for LFA-1 and shares considerable sequence homology with ICAM-1 and ICAM-2, it does not appear to be expressed by the endothelium of normal or inflamed synovial vessels. Intense expression of ICAM-R by rheumatoid synovial lymphocytes and macrophages suggests that it may play a role in processes requiring cell-cell contact, such as antigen presentation and homotypic aggregation.     

Designing a mixed methods study in pediatric oncology nursing research

Despite the appeal of discovering the different strengths of various research methods, mixed methods research remains elusive in pediatric oncology nursing research. If pediatric oncology nurses are to succeed in mixing quantitative and qualitative methods, they need practical guidelines for managing the complex data and analyses of mixed methods research. This article discusses mixed methods terminology, designs, and key design features. Specific areas addressed include the myths about mixed methods research, types of mixed method research designs, steps involved in developing a mixed method research study, and the benefits and challenges of using mixed methods designs in pediatric oncology research. Examples of recent research studies that have combined quantitative and qualitative research methods are provided. The term mixed methods research is used throughout this article to reflect the use of both quantitative and qualitative methods within one study rather than the use of these methods in separate studies concerning the same research problem.     

EHR-based cohort assessment for multicenter RCTs: a fast and flexible model for identifying potential study sites

ObjectiveThe Recruitment Innovation Center (RIC), partnering with the Trial Innovation Network and institutions in the National Institutes of Health-sponsored Clinical and Translational Science Awards (CTSA) Program, aimed to develop a service line to retrieve study population estimates from electronic health record (EHR) systems for use in selecting enrollment sites for multicenter clinical trials. Our goal was to create and field-test a low burden, low tech, and high-yield method.Materials and MethodsIn building this service line, the RIC strove to complement, rather than replace, CTSA hubs’ existing cohort assessment tools. For each new EHR cohort request, we work with the investigator to develop a computable phenotype algorithm that targets the desired population. CTSA hubs run the phenotype query and return results using a standardized survey. We provide a comprehensive report to the investigator to assist in study site selection.ResultsFrom 2017 to 2020, the RIC developed and socialized 36 phenotype-dependent cohort requests on behalf of investigators. The average response rate to these requests was 73%.DiscussionAchieving enrollment goals in a multicenter clinical trial requires that researchers identify study sites that will provide sufficient enrollment. The fast and flexible method the RIC has developed, with CTSA feedback, allows hubs to query their EHR using a generalizable, vetted phenotype algorithm to produce reliable counts of potentially eligible study participants.ConclusionThe RIC’s EHR cohort assessment process for evaluating sites for multicenter trials has been shown to be efficient and helpful. The model may be replicated for use by other programs.     

Iron overload in multiply transfused patients who are HIV seropositive.

AIMS--To assess histologically the amount of iron deposited in liver biopsy specimens from HIV positive patients; and to perform estimations of liver iron on tissue from patients with an increase in parenchymal stainable iron. To correlate the amount of blood transfused and the degree of iron overload. METHODS--Liver biopsy specimens (n = 120) from 109 HIV positive patients, 74 of whom had AIDS, were examined retrospectively and the amount of iron, as visualised with Perls's stain, was graded. Fibrosis was assessed using connective tissue stains. Estimations of liver iron were performed on tissue retrieved from paraffin wax blocks in cases with histological grade 3 or 4 iron overload. The amount of blood transfused before liver biopsy was determined from the notes for each patient. RESULTS--Fifteen of the 120 liver biopsy specimens had significantly increased amounts of iron in their hepatocytes, as assessed histologically, and this was confirmed in seven cases by measurement of liver iron. There was a close correlation between the amount of blood transfused and the degree of iron overload. In the initial biopsy specimens only one case showed portal tract expansion. Three of the five patients who had repeat biopsies, however, showed progressive fibrosis. CONCLUSION--Multiply transfused HIV positive patients may develop clinically important iron overload and are at risk of developing progressive fibrosis. Superimposed liver disease, especially viral hepatitis, in these high risk patients may exacerbate the effects of the iron overload.