Clonal relationship between lymphocytic predominance Hodgkin's disease and concurrent or subsequent large-cell lymphoma of B lineage

The occurrence of a large-cell lymphoma (LCL) concurrent with or subsequent to lymphocytic predominance Hodgkin's disease (LPHD) is well documented. Given the well-characterized B-cell nature of the Reed- Sternberg cell variants in LPHD, there may be a clonal relationship between the LPHD and the associated B-cell LCL. In this study, we adapted a highly sensitive, clonospecific assay to test whether the clone comprising the LCL exists in the corresponding LPHD tumor. Nine cases meeting the histologic criteria of nodular LPHD and B-cell LCL were identified, reviewed, and studied. Initially, clonality of both lesions was assessed using consensus primers to conserved regions in the IgH variable (frame-work III) and joining region genes in a polymerase chain reaction (PCR) assay. The PCR assay detected a clonal B-cell population in five of the LCLs, whereas analysis of eight cases of LPHD did not detect a dominant clone. Clonal products from the LCL were then sequenced, and clonospecific oligonucleotides were designed from the unique nucleotide sequence encoding the complementarity- determining region-III. These were then used as primers and/or probes in sensitive PCR-based assays on the corresponding LPHD tumors. In two cases, the clonospecific assay showed that the LPHD and LCL shared a common clone that was further confirmed by sequence analysis. This finding provides genotypic evidence that, at least in some cases, the LCL represents a clonal progression of LPHD.     

Amebic liver abscess: diagnosis and treatment evaluation with MR imaging

Magnetic resonance images were obtained before and after treatment in 17 patients with 29 amebic liver abscesses. Pretreatment T1-weighted images showed a sharply circumscribed, heterogeneous, low-signal-intensity mass, devoid of normal hepatic tissue and corresponding to the abscess cavity as measured sonographically. T2-weighted images showed the abscess cavity as a hyperintense region and also showed a larger region of hyperintensity extending from the cavity margins to the liver surface, corresponding to edematous but morphologically normal liver tissue. After treatment, the abscess cavity became homogeneously hypointense on T1-weighted images, corresponding to liquefaction of the abscess center. With successful treatment, concentric rings corresponding to (a) an inner margin of inflamed granulation tissue, (b) bands of type I collagen, and (c) the outer margin of atrophic and/or mildly inflamed liver tissue became prominent on T1- and T2-weighted images. T2-weighted images showed rapid resolution of the perifocal hepatic edema.     

MRI of female genital and pelvic organs during sexual arousal

We utilized contrast enhanced magnetic resonance imaging (MRI) to delineate the anatomy of the female genital and pelvic organs during sexual arousal. Eleven healthy pre-menopausal women and eight healthy post-menopausal women underwent MRI of the pelvis while watching an erotic video. A 1.5 Tesla MR system was used to produce T1-weighted images following administration of MS-325, a gadolinium-based blood pool contrast agent. Selected structural dimensions and enhancement were measured prior to and during sexual arousal. In both pre- and post-menopausal subjects, vestibular bulb and labia minora width increased with arousal. Enhancement measurements increased in the bulb, labia minora and clitoris in both pre- and post-menopausal subjects, and in the vagina in pre-menopausal subjects. There were no marked changes in size or enhancement of the labia majora, urethra, cervix, or rectum during sexual arousal in pre- or post-menopausal subjects. Using MRI, we observed specific changes in the female genitalia and pelvic organs with sexual arousal, in both pre- and post-menopausal women. MRI can potentially provide detailed anatomical information in the assessment of female sexual function, particularly with regard to changes in blood flow.     

CD63 is a component of Weibel-Palade bodies of human endothelial cells

Weibel-Palade bodies are secretory granules of vascular endothelial cells specialized in the storage of von Willebrand factor (vWF) and P- selectin, two adhesion proteins that can be rapidly mobilized to the cell surface by exocytosis in response to thrombin or other agonists. In this study, we attempted to identify additional components of Weibel- Palade bodies by raising monoclonal antibodies to these granules, purified by cell fractionation. One antibody, 2C6, was found to be specific for CD63, a membrane glycoprotein previously described in the lysosomes of platelets and other cell types. The immunopurified 2C6 antigen was recognized by an anti-CD63 reference antibody, 2.28, by Western blotting. Also, the biosynthetic profile of the 2C6 antigen in endothelial cells showed a nascent molecular mass and a glycosylation pattern identical to that of CD63. Immunofluorescence staining with 2C6 showed the lysosomes, and also elongated structures identified as Weibel-Palade bodies by their shape, distribution, and positive staining with anti-vWF antibodies, CD63 was also found by Western blotting of subcellular fractions highly enriched in Weibel-Palade bodies. Our results indicate that CD63 colocalizes with vWF and P- selectin in the Weibel-Palade bodies of endothelial cells, and together with these adhesion proteins it could be rapidly expressed on the cell surface in areas of vascular injury and inflammation.     

Protein A adsorption of acute myelogenous leukemia serum induces in vitro blast lysis

We studied cytotoxic activity of acute myelogenous leukemia (AML) sera for AML blasts before and after immunoadsorption with Staphylococcus aureus, Cowan I (SAC), which contains protein A. We found in vitro that incubation with treated AML sera reduced viability to 42.7% of control (p less than 0.01) for autologous and 21.0% of control (p less than 0.01) for allogeneic blasts. Normal peripheral blood cells were not killed by treated AML sera. Wood 46 strain of Staphylococcus aureus, which does not contain protein A, did not significantly reduce AML blast viability (94.8%, p greater than 0.4), while Sepharose-bound protein A reduced viability to 63.8% (p less than 0.01). Cytotoxicity does not appear to be complement-mediated, byt cytotoxic activity is trypsin-sensitive and is contained in the immunoglobulin fraction. This model for study of the tumoricidal action of protein A adsorption should be useful for predicting utility of plasma adsorption as a therapeutic adjunct in the future.     

Biochemical, immunological, and in vivo functional characterization of B-domain-deleted factor VIII

Coagulation factor VIII (FVIII) is a cofactor in the intrinsic pathway of blood coagulation for which deficiency results in the bleeding disorder hemophilia A. FVIII contains a domain structure of A1-A2-B-A3- C1-C2 of which the B domain is dispensable for procoagulant activity in vitro. In this report, we compare the properties of B-domain-deleted FVIII (residues 760 through 1639, designated LA-VIII) to wildtype recombinant FVIII. In transfected Chinese hamster ovary (CHO) cells, LA- VIII was expressed at a 10- to 20-fold greater level compared with wildtype FVIII. The specific activity of purified LA-VIII was indistinguishable from wild-type recombinant FVIII and both exhibited similar thrombin activation coefficients. Wildtype recombinant-derived FVIII and LA-VIII also displayed similar timecourses of thrombin activation and heavy chain cleavage. However, compared with wildtype recombinant-derived FVIII, the light chain of LA-VIII was cleaved fivefold more rapidly by thrombin. Addition of purified von Willebrand factor (vWF) did not alter the kinetics of thrombin cleavage or activation of either wildtype recombinant-derived FVIII or LA-VIII. The immunogenicity of LA-VIII was compared with wildtype FVIII in a novel model of neonatal tolerance induction in mice. The results did not detect any immunologic differences between wildtype FVIII and LA-VIII, suggesting that LA-VIII does not contain significant new epitopes that are absent in wildtype FVIII. LA-VIII was tolerated well on infusion into FVIII-deficient dogs and was able to correct the cuticle bleeding time similar to wildtype recombinant factor VIII. In vivo, LA-VIII was bound to canine vWF and exhibited a half-life similar to wildtype recombinant FVIII. These studies support that B-domain-deleted FVIII may be efficacious in treatment of hemophilia A in humans.     

Production of granulocyte-macrophage colony-stimulating factor by primary cultures of unstimulated rat microvascular endothelial cells

Small vessel (microvascular) endothelial cells are in close contact with hematopoietic progenitor cells in the bone marrow and therefore may have an important role in hematopoiesis. Although other studies have shown that endothelial cells produce various colony-stimulating factors (CSFs), these studies examined large vessel endothelial cells, which are different in many respects from microvascular endothelial cells and which do not contact cells in the bone marrow. We show in this study that primary cultures of unstimulated rat fat capillary endothelial cells grown in serum-free medium produce a substantial amount of granulocyte-macrophage CSF (GM-CSF). The medium conditioned by these cells stimulated proliferation of two different lines of GM- CSF-responsive cells--PT-18 mast cells and FDC-P1 cells--and supported the growth of cells of the granulocyte and macrophage lines in cultures of rat bone marrow cells. The factor responsible for this activity had physical properties consistent with those of GM-CSF, namely, a similar apparent mol wt by gel filtration, resistance to repeated freeze-thaws, resistance to boiling for ten minutes but not for 30 minutes, and resistance to heating to 56 degrees C for one hour. The factor causing target cell stimulation was not B cell-stimulating factor-1 (BSF-1, or IL 4), since it failed to stimulate a BSF-1-responsive cell line HT2- JH, and target cells (PT-18) did not respond appreciably to recombinant BSF-1. Northern blot analysis of mRNA from rat fat capillary endothelial cells showed high levels of expression of GM-CSF, confirming that this factor is produced by microvascular endothelial cells. This is the first report of CSF production by unstimulated microvascular endothelial cells, demonstrating that these ubiquitous cells are capable of producing sizable amounts of at least one growth factor for hematopoietic progenitor cells.     

Irradiation induces release of von Willebrand protein from endothelial cells in culture

Human umbilical vein endothelial cells in tissue culture were irradiated with doses between 0 and 40 Gy, and the released von Willebrand (vW) protein and that which remained associated with the cells was quantitated. Doses of 20 Gy and higher produced a statistically significant increase in amount of vW protein secreted. This release was present whether the cells were labeled continuously throughout the experiment or just prelabeled before irradiation. An increase in fibronectin secretion was not observed. The release response to radiation was slow, reaching significance close to 24 hours after irradiation. The release of vW protein was not due to cell lysis, because the secreted vW protein contained very little of the large 260- kilodalton vW precursor subunit present in cell lysates and the cells appeared intact by immunofluorescence staining.