Alexander Dvilansky

Novel role of 1,25(OH)2D3 in induction of erythroid progenitor cell proliferation

OBJECTIVE Burst-forming unit erythroid and colony-forming unit erythroid growth in vitro is lower in studies of continuous ambulatory peritoneal dialysis patients than healthy controls. Burst-forming unit erythroid growth was potentiated by addition of 1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] and normalized by erythropoietin (Epo) therapy, suggesting an interaction between Epo and 1,25(OH)(2)D(3) at the stem cell level. The objective of this study was to determine the mechanism by which 1,25(OH)(2)D(3) enhances the stimulatory effect of Epo on the growth of erythroid precursor cells. MATERIALS AND METHODS We examined the effect of 1,25(OH)(2)D(3) and Epo on stem cell proliferation. Proliferation of TF1 cells of erythroid origin was measured by the XTT method, 3[H] thymidine incorporation, and cell counting by trypan blue exclusion; cord blood (CB) stem cells were counted. Epo receptor (EpoR) quantitation was evaluated by 125I-Epo binding and Scatchard analysis, immunoprecipitation, and Western blotting. Expression of EpoR mRNA was measured by reverse transcriptase polymerase chain reaction. RESULTS The stem cell factor-dependent CB stem cells and the TF1 cells responded to Epo and 1,25(OH)(2)D(3) by increased proliferation, while their simultaneous addition potentiated cell proliferation in a synergistic manner (25.67% +/- 4.8% of Epo proliferation at day 10 for CB cells; p < 0.005). 1,25(OH)(2)D(3) produced an up-regulation of EpoR number in TF1 cells and increased the expression of EpoR mRNA (p < 0.01). CONCLUSIONS The increase in EpoR expression induced by 1,25(OH)(2)D(3) might explain the synergistic interaction between Epo and 1,25(OH)(2)D(3) in stem cells.

EFFECT OF RADIATION ON RED CELL MEMBRANE AND INTRACELLULAR OXIDATIVE DEFENSE SYSTEMS

Ionizing radiation is currently used for prevention of transfusion associated graft versus host disease (TAGVHD). As radiation damage is associated with the production of activated oxygen species, the aim of this study was to observe the immediate effect of ionizing radiation on red cell membrane and intracellular oxidative defense systems. Neonatal and iron deficiency (IDA) cells, known for their increased sensitivity to oxidative stress, were chosen and compared with normal cells. Irradiation was performed in doses of 1500 cGy, 3000 cGy and 5000 cGy. GSH and methemoglobin levels and the activity of different antioxidant enzymes, measured under optimal in vitro conditions, were preserved in all cells after irradiation. Only radiation at the highest does of 5000 cGy, caused significant potassium leakage in neonatal cells and insignificant increase in IDA cells. Thus, cells with increased sensitivity to oxidative stress are more susceptible to damage by ionizing radiation than normal cells.

Release of platelet 5-hydroxytryptamine by plasma taken from patients during and between migraine attacks

&NA; Experiments were carried out in order to further delineate the pathophysiology of the fall of plasma 5‐hydroxytryptamine (5‐HT) during a migraine attack. Platelets from normal subjects were incubated with 14 C‐labelled 5‐HT, and the release of 5‐HT was measured following exposure of these platelets to plasma taken from migraine patients during an attack or at headache‐free intervals. Plasma taken during attacks released significantly more 5‐HT. It is concluded that factor (s) exist in the serum during migraine attacks, which can cause 5‐HT release from normal platelets. The identification of this factor may be important.

Iron Deficiency Anemia: Recovery from in vitro Oxidative Stress

Red blood cells in iron deficiency anemia (IDA) have a decreased activity of essential antioxidant enzymes. The present study examined the effect of in vitro exposure to oxidative agents in IDA cells and their recovery capacity. Red cells of 26 IDA patients and 10 healthy subjects were examined. Cells of IDA patients had higher levels of reduced glutathione (GSH), and normal methemoglobin and malonyldialdehyde (MDA) levels. Exposure to butyl hydroperoxide revealed a dose-dependent sensitivity in IDA cells, with extensive GSH depletion and increased MDA levels. These changes were partially reversible by incubation with dithiothreitol. Exposure to phenazine methosulfate, to produce intracellular superoxide ions, resulted in moderate GSH depletion and methemoglobin production. IDA cells were more sensitive than control cells to high concentrations of this substance. This effect was further augmented by preincubation with a superoxide dismutase inhibitor. Our data demonstrate that IDA cells are more susceptible to oxidation but have good capacity for recovery.

Membrane dynamic alterations associated with activation of human platelets by thrombin.

Two fluorescent probes, N-carboxymethylisatoic anhydride, which binds to membrane proteins, and 1,6-diphenyl-1,3,5-hexatriene, a lipophilic label, have been used to follow membrane microenvironmental changes. Activation of human platelets by thrombin resulted in a simultaneous increase in values of fluorescence polarization (P) of both probes during the stages of shape change and secretion, which further increased during platelet aggregation. The similar pattern of changes in P for both probes indicates the interdependence of lipids and proteins in the activated platelet membrane.

Radiation Damage to the Erythrocyte Membrane: The Effects of Medium and Cell Concentrations

Human erythrocytes suspended in plasma, or in phosphate buffered saline (PBS), were exposed to ionizing radiation. Potassium leakage from irradiated erythrocytes is significantly higher in PBS than in plasma. The potassium leakage decreases when PBS is gradually replaced by plasma. These findings suggest that some of the plasma constituents have radioprotective properties. The potassium leakage per cell is independent of the hematocrit, Hct. The potassium leakage is attributed to the formation of radiation defects in the membrane. Analysis of the effect of radiation dose, plasma and cell concentrations on the product of the number and surface area of the radiation defects indicates that the radiation damage is mainly due to the direct formation of free radicals in the cell membrane. The radioprotective effect of plasma is attributed to surface reactions of these free radicals with plasma constituents adsorbed on the membrane.

Chronic granulocytic leukemia, neutrophilic type, with paraproteinemia (IgA type K).

A patient with chronic granulocytic leukemia, neutrophilic type, was followed for 28 months. A paraproteinemia, IgA type K, and Bence Jones proteinuria (K) appeared without prior chemotherapy with alkylating agents.

DISTAMYCIN-A DERIVATIVES POTENTIATE TUMOR-NECROSIS-FACTOR ACTIVITY VIA THE MODULATION OF TYROSINE PHOSPHORYLATION

The cytotoxic activities of 2 novel distamycin‐A derivatives, FCE 24517 and FCE 25450A, alone and in combination with tumor‐necrosis factor‐α (TNF), were studied. Both drugs, especially FCE 25450A, analyzed extensively here, inhibited the growth of HL60 promyelocytic cells, and human SV80 and murine L929 transformed fibroblasts in a dose‐dependent manner. The growth‐inhibitory potential of sequential exposure to the distamycin‐A analogs and TNF was determined. A 4‐hr treatment of L929 fibroblasts with 100–1,000 ng/ml FCE 25450A, followed by 2 ng/ml TNF, resulted in a synergistic anti‐proliferative effect. The synergism of FCE 24517 with TNF was less profound. Experiments to elucidate the mechanism underlying the cooperation revealed that FCE 25450A pre‐treatment almost completely abolished the elevated tyrosine phosphorylation of a 137‐kDa and other membranal proteins and prevented the de‐phosphorylation of another protein band observed in L929 cells in the presence of TNF. FCE 25450A alone induced no changes in the phosphotyrosine profile of the cells. The effect of FCE 25450A was counteracted by the tyrosine‐phosphatase inhibitor orthovanadate. In parallel, the inhibitor also diminished the anti‐proliferative action of the FCE 25450A/TNF combination. These findings suggest that, beyond their cytotoxic effects as single agents, the distamycin derivatives increase the sensitivity of cells to TNF. This effect is governed via the inhibition of TNF‐induced tyrosine phosphorylation of specific proteins which are probably involved in the development of TNF resistance. Thus, protein de‐phosphorylation might provide an additional mechanism of action of these novel distamycin‐A‐derived drugs. Int. J. Cancer 72:810–814, 1997.

Thrombocytopenic purpura as the sole manifestation of recurrence in Hodgkin's disease.

A patient with Hodgkin’s disease is described, in whom thrombocytopenic purpura was the presenting manifestation of recurrence 15 years after the initial manifestation of the disease. Diverse factors

A NOVEL AGAROSE ACROBEADS PROTEIN A COLUMN FOR SELECTIVE IMMUNOADSORBANCE OF WHOLE BLOOD: PERFORMANCE, SPECIFICITY AND SAFETY

The present study describes the performance and safety parameters of a novel column composed of protein A linked to acrobeads. Hemoperfusion of anticoagulated whole human blood was carried out in a closed system. Specific adsorption of IgG was documented in whole blood, plasma and IgG diluted in saline. Specific activity of the column was 3.52 mg/ml beads. These studies revealed slight changes in platelet counts and minor hemolysis. The relatively short period needed for maximal hemoperfusion effect and safety may enable the use of this novel therapeutic approach in clinical practice.