Two cases of blastomycosis from a common source: use of DNA restriction analysis to identify strains.

Two friends, one with AIDS, developed severe pulmonary blastomycosis but differed markedly in clinical course. The human immunodeficiency virus-negative patient responded completely to ketoconazole; the patient with AIDS died of progressive disseminated infection despite treatment with fluconazole and amphotericin B. Epidemiologic investigation suggested a common source of infection, but serologic evaluation and environmental cultures were unrevealing. EcoRI digestion of the Blastomyces dermatitidis isolates showed identical restriction fragment patterns that differed from patterns obtained from other clinical isolates. Analysis using a Histoplasma capsulatum ribosomal DNA probe that cross-hybridizes with B. dermatitidis showed that the isolates from the two patients were identical and different from others. Thus, the patients were probably infected with the same strain, possibly from a common source. These data indicate the critical role of cellular immunity in patients with blastomycosis, show that there are multiple genotypes of B. dermatitidis, and suggest that DNA restriction analysis is a useful epidemiologic tool.     

Role of upstream DNase I hypersensitive sites in the regulation of human alpha globin gene expression

Erythroid-specific DNase 1 hypersensitive sites have been identified at the promoters of the human alpha-like genes and within the region from 4 to 40 kb upstream of the gene cluster. One of these sites, HS-40, has been shown previously to be the major regulator of tissue-specific alpha-globin gene expression. We have now examined the function of other hypersensitive sites by studying the expression in mouse erythroleukemia (MEL) cells of various fragments containing these sites attached to HS-40 and an alpha-globin gene. High level expression of the alpha gene was observed in all cases. When clones of MEL cells bearing a single copy of the alpha-globin gene fragments were examined, expression levels were similar to those of the endogenous mouse alpha genes and similar to MEL cells bearing beta gene constructs under the control of the beta-globin locus control region. However, there was no evidence that the additional hypersensitive sites increased the level of expression or conferred copy number dependence on the expression of a linked alpha gene in MEL cells.     

Calcium-dependent cysteine protease activity in the sera of patients with thrombotic thrombocytopenic purpura

Plasma and serum from patients with thrombotic thrombocytopenic purpura (TTP) can cause activation and aggregation of normal human platelets in vitro. It is possible that this platelet-activating factor contributes to the disease. In this report we describe studies designed to identify the platelet-activating factor in TTP. Platelet activation by sera from 15 patients with TTP was inhibited by leupeptin, iodoacetamide, and antipain but not by phenylmethylsulphonylfluoride, epsilon-aminocaproic acid, soybean trypsin inhibitor, aprotinin, and D-phenylanyl-1-prolyl-1- arginine chloromethyl ketone. These studies suggested that the platelet- activating factor in TTP serum was a cysteine protease. We confirmed that a calcium-dependent cysteine protease (CDP) was present in the sera of each of the 15 patients when we used an assay based on the ability of CDP to proteolyse platelet membrane glycoprotein 1b (GP1b) and hence to abolish the ability of CDP-treated normal platelets to agglutinate in the presence of ristocetin and von Willebrand factor. This proteolytic activity was inhibited by EDTA, leupeptin, antipain, iodoacetamide, and by N-ethyl-maleamide (NEM) but not by the serine protease inhibitors. Activity was detected in 15 of 15 patients with TTP tested before therapy was begun. In contrast, no activity was detected in the serum of any of five of the TTP patients tested in remission or in any of the sera from 36 patients with thrombocytopenia and 423 nonthrombocytopenic controls. To look for in vivo CDP activity in patients with TTP, we studied platelets from two patients with acute TTP (drawn into acid-citrate-dextrose, NEM, and leupeptin). These platelets showed a loss of GP1b from the platelet surface. Both patients were also studied in remission: GP1b on the platelet surface had returned to normal. These studies provide evidence that CDP is present in the sera of patients with TTP, that it is specific to this disease, and that is is active in vivo as well as in vitro. We postulate that a disorder of CDP homeostasis plays a major role in the pathophysiology of TTP.     

Central nervous system high-resolution magnetic resonance imaging: effect of increasing spatial resolution on resolving power

Resolving power is a useful measure of the magnetic resonance (MR) imager-determined ability to discriminate subtle disease. Optimizing the resolving power produces the best MR images. The resolving power improves with higher spatial resolution, signal-to-noise ratio, and object contrast. Resolving power degrades with increased patient motion, which can be associated with prolonged imaging times. High and low object contrast 0.35-T MR images of the central nervous system are compared at different levels of spatial resolution and signal-to-noise ratios. In systems that have a marginal signal-to-noise ratio, the resolving power can actually decrease when higher spatial resolution is used due to further lowering of signal-to-noise ratio and to increased motion resulting from longer imaging times. This decreases the conspicuity of small, low contrast lesions.