Individual cell analysis of the cytokine repertoire in human immunodeficiency virus-1-infected monocytes/macrophages by a combination of immunocytochemistry and in situ hybridization

The expression of many cytokines is dysregulated in individuals infected with the human immunodeficiency virus-1 (HIV-1). To determine the effects of HIV-1 infection on cytokine expression in individual cells (at the single cell level), we investigated the intracellular levels of proinflammatory cytokines (tumor necrosis factor [TNF]-alpha, interleukin [IL]-1beta, IL-6, and IL-8) and hematopoietic growth factors (granulocyte colony-stimulating factor [G-CSF], granulocyte-macrophage colony-stimulating factor [GM-CSF]) in monocyte-derived macrophages, mock-infected, or infected with HIV-1 by immunocytochemical staining for cytokine protein and compared this with secreted cytokine levels as determined by specific enzyme-linked immunosorbent assay (ELISA). No difference in the frequency or intensity of cell-associated immunocytochemical cytokine staining could be observed between HIV-1 and mock-infected cells even though the level of secreted proinflammatory cytokines increased and the hematopoietic growth factors decreased in HIV-1-infected cultures. Furthermore, equal expression of cytokine mRNA was observed in all cells in the culture regardless of whether the cells were productively infected with HIV-1 as determined by double-labelling immunocytochemical staining for HIV-1 p24 antigen and in situ hybridization for cytokine mRNA expression. These results indicate that HIV-1 infection results in dysregulation of intracellular cytokine mRNA expression and cytokine secretion not only in HIV-1-infected cells, but also through an indirect way(s) affecting cells not producing virus.     

Association of glutathione S-transferase GSTM1 and GSTT1 null genotypes with clinical outcome in epithelial ovarian cancer

Epithelial ovarian cancer is generally associated with a poor outcome, although the mechanisms that determine survival and progression-free interval (PFI) are unclear. Data from ovarian tumors showing associations between (a) null genotypes at the glutathione S-transferase GSTM1 and GSTT1 loci and expression of p53 protein and (b) outcome and expression of p53 suggest that polymorphism at these loci is a factor determining outcome. Accordingly, we have studied the association between the GSTM1 null and GSTT1 null genotypes and survival and PFI in 148 women with epithelial ovarian cancer. Although we did not find an association between individual genotypes and outcome, women with both GSTM1 null and GSTT1 null genotypes demonstrated poorer survival (P = 0.001) and reduced PFI (P = 0.003). Thus, no cases with both these genotypes survived past 42 months postdiagnosis. In contrast, 43% of the women without this combination survived beyond this time. Because response to chemotherapy is a major factor determining outcome in ovarian cancer, we also examined the data for associations between the glutathione S-transferase genotypes and response to such treatment. Thus, in 78 patients treated with chemotherapy, the combination of GSTM1 null and GSTT1 null was associated with unresponsiveness to primary chemotherapy (P = 0.004); none of the eight patients with both these genotypes responded, compared with 38 of 70 (54%) of patients with other genotype combinations. The effect of the combination of genotypes on survival and PFI was lost in a multivariate model that included response to chemotherapy as a confounding factor. This suggests that the combination of GSTM1 null/GSTT1 null is associated with outcome because of its influence on response to chemotherapy. These preliminary findings may provide a basis for the selection of patients for treatment with chemotherapeutic agents.     

Treatment with blood components and coagulation factor concentrates--modern concepts of hemotherapy

Transfusion of whole blood is the most common and successful organ transplantation world wide. It is in use longer than any other transplantation procedure. With the exception of uses in situations of crisis, treatment with whole blood is nowadays obsolete. This is due to enormous progress in preparation technologies. As a result blood from volunteer donors is separated before therapeutic use into its cellular and plasmatic components. The latter may be further processed into subfractions or secondary products. The advantages of this strategy are evident: a) the crude pharmaceutic material blood is used optimally, b) the isolated components may be stored over periods appropriate for their stability be it in a cryopreserved state or after biochemical, immunologic or biological modification, c) the patient receives only the blood components he needs, d) the transfusion of unnecessary fractions of blood is avoided, e) the rate of transfusion-associated adverse events is reduced, f) the physician has a basis for customized hematotherapy. Important implications concern logistics, economy, differentiated therapy and drug safety consideration. This contribution gives an overview on progress made in therapy of hematologic and oncologic patients with blood products and concentrates of coagulation factors. It demonstrates selected aspects of actual interest for optimal hemotherapy.     

Balance retraining in a hemiparetic patient using center of gravity biofeedback: a single-case study

A single-subject experimental design was used to evaluate the effects of center of gravity biofeedback on retraining balance control in a 72-yr.-old male hemiparetic patient. For both assessment and training the subject stood on dual force-plates enclosed on three sides by a visual surround. A 4-wk. training period consisted of subject-initiated shifts in body weight to control movement of a real-time computer display of the subject's center of gravity. A Sensory Organization Test and evaluation of weight distribution indicated that biofeedback retraining positively affected balance control during dynamic conditions and when the ground support moved forward.     

Growth factors and insulin stimulate tyrosine phosphorylation of the 51C/SHIP2 protein

Antibodies raised against the 51C/SHIP2 inositol polyphosphate 5'-phosphatase were used to examine the effects of growth factors and insulin on the metabolism of this protein. Immunoblot analysis revealed that the 51C/SHIP2 protein was widely expressed in fibroblast and nonhematopoietic tumor cell lines, unlike the SHIP protein, which was found only in cell lines of hematopoietic origin. The 51C/SHIP2 antiserum precipitated a protein of approximately 145 kDa along with an activity which hydrolyzed phosphatidylinositol 3,4, 5-trisphosphate to phosphatidylinositol 3,4-bisphosphate. Tyrosine phosphorylation of the 51C/SHIP2 protein occurred in response to treatment of cells with epidermal growth (EGF), platelet-derived growth factor (PDGF), nerve growth factor (NGF), insulin-like growth factor-1 (IGF-1), or insulin. EGF and PDGF induced transient tyrosine phosphorylation of 51C/SHIP2, with maximal tyrosine phosphorylation occurring at 5-10 min following treatment and returning to near basal levels within 20 min. In contrast, treatment of cells with NGF, IGF-1, or insulin resulted in prolonged tyrosine phosphorylation of 51C/SHIP2 protein, with 40-80% maximal phosphorylation sustained for up to 2 h following agonist treatment. The kinetics of activation of the Akt/PKB protein kinase by the various factors correlated well with the kinetics of tyrosine phosphorylation of 51C/SHIP2. EGF, NGF, and PDGF stimulated the association of 51C/SHIP2 protein with the Shc adapter protein; however, no Shc could be detected in 51C/SHIP2-immune precipitates from cells treated with IGF-1 or insulin. The data suggest that 51C/SHIP2 may play a significant role in regulation of phosphatidylinositol 3'-kinase signaling by growth factors and insulin.     

Mammalian and Drosophila dachshund genes are related to the Ski proto-oncogene and are expressed in eye and limb

The human myeloid leukemias are a diverse group of disorders characterized by massive clonal expansion of myeloid cells showing variable degrees of differentiation block. Leukemic dendritic cells were generated in culture from chronic myelogenous leukemia (CML). These were used to stimulate autologous T cells to develop leukemia-specific cytotoxicity. Available data suggest that the cells responsible for the cytolytic activity are at least in part CD8+ and HLA restricted in their function. Additional data suggest that some anti-CML cellular activity may be Fas mediated. T-cell receptor studies provide evidence for an oligoclonal response implying a recognition of a limited number of antigens. We have used culture techniques similar to those used for CML to study the ability of AML cells to differentiate toward dendritic cells. Four of five patients have shown acute leukemia-derived dendritic cells. This work offers an avenue for the development of novel strategies for the control of human myeloid leukemias.     

The Canadian Cardiovascular Society--the first and the next 50 years

BACKGROUND: Although beta2-adrenoceptor agonists are widely used in the treatment of asthma, a number of studies have suggested that their long-term use may exacerbate the condition. One possible mechanism for this stems from the in vitro observation that beta2-agonists increase IgE synthesis by human blood mononuclear cells. OBJECTIVE: We sought to examine the effect of regular beta2-agonist therapy on IgE production in vivo in human volunteers. METHODS: Placebo or salbutamol (8 mg BD) tablets were given in a double-blind, randomized fashion to 25 volunteers allergic to grass pollen throughout a period encompassing the UK grass pollen season (April through September). Levels of serum IgE were measured monthly, and nasal IgE was measured at the height and end of the season. Efficacy was assessed through monthly recordings of symptoms of blocked nose (vascular) and other symptoms of rhinitis (nonvascular). RESULTS: For the whole group the geometric mean of serum IgE levels rose from a baseline of 58.7 IU/mL (range, 0 to 1027 IU/mL) to 140 IU/mL (range, 12 to 878 IU/mL) at the height of the pollen season (P =.0001). There was no significant difference between the magnitude of the rise in IgE between the groups with a ratio of increase for salbutamol/placebo of 1.17 (confidence interval = 0.78 to 1.75). There was no change in nasal IgE levels. Total and nonvascular symptom scores were reduced by salbutamol, reaching statistical significance at the height of the pollen season (P <.05). CONCLUSION: An oral dose of the beta2-agonist salbutamol, sufficient to maintain therapeutic levels and provide clinical benefit, does not accentuate the seasonal increase of IgE in human atopic volunteers.