Clear cells in acral melanoma.

Acral melanoma may present clinically and histologically with atypical features causing a delay in proper diagnosis. The aim of the present study was to assess the frequency of a histological variant with clear cell changes. Clinical information, hematoxylin & eosin stained paraffin sections and immunohistochemical staining profiles were reviewed in 49 cases of acral melanoma. Twenty-one (43%) specimens contained tumor cells with clear cell changes in focal areas, whereas in 7 (14%) specimens clear cells were the major tumor constituting cells. The tumor thickness ranged from melanoma in situ to 14 mm. Immunohistochemistry demonstrated weak staining for S100 and HMB45 as well as strong positivity for Melan A and NK1C3. Recognition of clear cell features is important since differential diagnosis includes a variety of other clear cell malignancies, among them metastasis from renal cell carcinoma, clear cell sarcoma and hidradenocarcinoma.

The influence of subinhibitory concentrations of conventional and experimental antifungal drugs on the expression of the iC3b binding protein in Candida albicans strains during filamentation.

The influence of six antifungal agents on the expression of the fungal iC3b binding protein was studied in germ-tubes and the mycelial form of several Candida albicans strains. All antifungal agents inhibited not only the yeast-mycelial transformation, but also the formation of rosettes consisting of complement-coated sheep erythrocytes (EAiC3b) bound to the mycelial form of C. albicans. Immunofluorescence as well as ELISA, employing the monoclonal antibody OKM-1 which recognizes the alpha chain of human CR3 and which cross-reacts with the fungal iC3b binding protein, revealed that subinhibitory concentrations of 0.1 mg l(-1) (which did not affect the growth of either germ-tubes or the mycelial form of C. albicans) inhibited the expression of the iC3b binding protein, while lower concentrations (0.01 mg l(-1)) allowed a comparable and sometimes even slightly higher expression of this protein, in comparison with the untreated control. However, treatment with antifungal agents apparently did not lead to a major cleavage of the protein. The dependence of the amount of the iC3b binding protein expressed on the concentration of added antifungal drugs and on the morphological forms of individual C. albicans isolates suggests a drug dependent influence on the expression of this protein and a possible association with the changing virulence of C. albicans strains during antifungal therapy.

Immunologic changes after transfusion of autologous or allogeneic buffy coat-poor versus white cell-reduced blood to patients undergoing arthroplasty. I. Proliferative T-cell responses and the balance of helper and suppressor T cells.

BACKGROUND: Donor white cells (WBCs) contained in red cell (RBC) transfusions are thought to provoke down-regulation of T-cell-mediated immunity. This study investigated this topic in otherwise healthy patients receiving buffy coat-depleted or WBC-filtered RBCs and undergoing standardized perioperative management. STUDY DESIGN AND METHODS: Patients undergoing elective orthopedic surgery (primary hip and knee replacement surgery) were enrolled in a prospective study. Perioperative changes in T-cell proliferation (stimulation with phytohemagglutinin and mixed lymphocyte culture) and T-cell balance (T-lymphocytes, helper T cells, and suppressor T cells) were compared after random assignment to allogeneic buffy coat-depleted (Group 2, n = 8) or WBC-reduced RBC (Group 3, n = 11) transfusion regimens. Recipients of autologous buffy coat-depleted RBC transfusions (n = 15) served as controls (Group 1). RESULTS: Compared to that in autologous transfusion recipients, alloantigen-induced T-cell proliferation was significantly reduced in recipients of allogeneic WBC-reduced RBCs (Day 3, p = 0.0274). After the transfusion of allogeneic buffy coat-depleted RBCs, a weak trend toward decreased T-cell proliferation was observed (p = 0.0933) and the numbers of CD4+ T cells were also significantly lower (Day 7, p = 0.0389). On Day 10, alloantigen-induced T-cell proliferation remained significantly below baseline after transfusion of WBC-reduced RBCs (p = 0.05), the numbers of CD3+ cells decreased in allogeneic RBC recipients (Group 2, p = 0.078; Group 3, p = 0.05), and those of CD8+ cells decreased significantly after the transfusion of allogeneic buffy coat-depleted RBCs (p = 0.0234) concomitant with an increased CD4:CD8 ratio (p = 0.0391). CONCLUSION: Results of the present study confirm the hypothesis of impaired T-cell-mediated immunity after allogeneic transfusion.

Formation of multinucleated giant cells in vitro is dependent on the stage of monocyte to macrophage maturation.

Multinucleated giant cells (MGC) are a common feature of granulomas that develop during various inflammatory reactions. MGC originate from fusion of monocytes or macrophages, but the exact mechanism of their generation is still unclear. In the present study, we investigated the influence of monocyte to macrophage maturation on the ability of human monocytes/macrophages to fuse with each other. MGC were generated in vitro by stimulation of human peripheral blood monocytes with cytokine containing supernatants. With freshly isolated monocytes, fusion rates of up to 90% were obtained. When monocyte to macrophage maturation was induced by culturing the cells in human serum, fusion rates gradually decreased with advancing time of the preceding culture (corresponding to the stage of differentiation) and almost no MGC formation could be obtained with 8-day-old macrophages. In contrast, fusion rates did not decrease when monocytes had been cultured under serum free conditions before stimulation. When freshly isolated monocytes were added to 1-week cultured macrophages, which had been membrane-labeled with a fluorochrome, fusion between the two populations could be induced. Because the ability for intracellular killing of certain pathogens is reduced in macrophages, fusion with monocytes (newly arriving at the site of inflammation) may represent an attempt to restore this capacity.

Temperature-dependent surface expression of the beta-2-integrin analogue of Candida albicans and its role in adhesion to the human endothelium.

Candida albicans has become one of the most important pathogens in intensive care units. Adherence of C. albicans to the vascular endothelium is believed to represent a critical step in the pathogenesis of disseminated candidiasis and may involve molecules analogous to human beta 2-integrins such as the complement receptor 3 (CR3) analogue of C. albicans (C.a.-CR3). Its expression was detected by a sensitive rosetting assay when Candida was present in its hyphal form but not in its yeast form, the latter being generally considered to be less pathogenic. However, the presence of hyphae alone was not sufficient: C.a.-CR3 expression was found to be temperature-dependent for 4 (out of 10) clinical isolates. Two rosetted better after growth at 30 degrees C, the other 2 after growth at 37 degrees C. This temperature dependence was most pronounced for 1 laboratory strain: C.a.-CR3 expression was best at 30 degrees C and markedly decreased with increasing temperatures. At 37 degrees C no rosettes were detected at all. Modifications of the culture conditions (e.g. agitation, pH) exerted a marked influence on the morphology of this strain but always allowed rosette formation once hyphae were formed at 30 degrees C. However, none of these modifications was able to induce rosettes at 37 degrees C. Adhesion of C. albicans isolates to an endothelial cell line was also temperature-dependent but not strongly correlated with C.a.-CR3 expression. Most strains exhibited a better adherence when grown at 30 degrees C. This finding may be of importance for exogenous infections, with Candida spp. invading the body from the outside, where the temperature is usually lower than the physiological body temperature.

Cell membrane labeling with fluorescent dyes for the demonstration of cytokine-induced fusion between monocytes and tumor cells.

Fusion between monocytes and tumor cells has been suggested as a cause for tumor metastasis. The aim of the present study was to establish an in vitro fusion model representing the in vivo situation as close as possible. For this purpose fusion between cells was induced by cytokine containing conditioned medium. In order to prove that hybrid formation between tumor cells and monocytes occurs, a two-color-fusion-assay based on membrane labeling with the fluorochromes PKH 2 (green) and PKH 26 (red) was established. These fusion experiments were analyzed by microscopy and, in addition, by flow cytometry. The attempt to induce fusion between monocytes and several tumor cell lines of hematopoietic origin revealed quite diverse results. The most extensive hybrid formations were seen with TALL, a T-lymphocytic tumor line. The monocytic tumor line HL60 and the B-lymphocytic tumor line BL41 also clearly yielded hybrids with monocytes but in smaller numbers. With some other hematopoietic tumor lines no evidence for hybrid formation was detected. These studies indicate that fusion of normal monocytes with certain tumor cells may be induced under conditions that may occur in comparable manner in vivo.

Normal granulocyte function but impaired monocyte function in a patient with aplastic anemia.

Granulocytopenia is an invariable finding in aplastic anemia (AA) and bacterial infections are a frequent complication and major cause of death in patients suffering from this bone marrow disorder. Using well-established assays, we determined the chemotaxis of granulocytes and monocytes as well as the phagocytic capacity of monocytes and oxidative metabolism of granulocytes in a patient with aplastic anemia of unknown etiology. Our results indicate a normal granulocyte oxidative metabolism and granulocyte chemotaxis towards formyl-leucyl-methionyl-phenylalanine (fMLP), whereas monocyte chemotaxis and phagocytic capacity were markedly reduced in this patient.

Ca ions stabilize the binding of complement factor iC3b to the pseudohyphal form of Candida albicans.

The pseudohyphal form of Candida albicans is able to bind iC3b. This may play an important role in the pathogenesis of disseminated candidiasis and, in particular, in adherence to endothelium, protection against complement action, and iron acquisition from erythrocytes. Here we report that Ca2+ ions are required to maintain stable binding of iC3b to C. albicans pseudohyphae.