The Effect of Ibuprofen on Cytokine Production by Mononuclear Cells from Schizophrenic Patients.

The existence of a restrained inflammatory state in schizophrenic individuals posed the question whether anti-inflammatory drugs may exert antipsychotic effects. Therefore, the effect of ibuprofen (IB) on cytokine production by human peripheral blood mononuclear cells (PBMC) from schizophrenic patients was examined and compared to that of healthy subjects. PBMC from 25 schizophrenic patients and 24 healthy volunteers were incubated for 24 h with lipopolysaccharide (LPS) in the absence or presence of various concentrations of IB. The levels of IL-1ß, IL-6, TNF-α, IL-10 and IL-1ra in the supernatants were tested applying ELISA kits. The secretion of TNF-α by cells from schizophrenic patients was significantly lower compared with controls. IB caused stimulation of TNF-α and IL-6 production by cells of the two groups and enhanced IL-1ß secretion by cells from schizophrenic patients. IB inhibited IL-1ra and IL-10 generation by cells from the two groups. Without IB, IL-1ra secretion was negatively correlated with the disease severity, while 200 µg/ml of IB positively correlated with the PANSS total score. IL-10 production was positively correlated with the PANSS positive subscale score both in the absence or presence of IB. The findings suggest that the effect of IB on the production of inflammatory cytokines may benefit the health of schizophrenic patients.

Vitamin B6 Modifies the Immune Cross-Talk between Mononuclear and Colon Carcinoma Cells.

The role of vitamin B6 as a key component in a number of biological events has been well established. Based on the relationship between chronic inflammation and carcinogenesis on the one hand, and the interaction between immune and cancer cells expressed by modulated cytokine production on the other hand, the aim of the present work was to examine the possibility that vitamin B6 affects cancer development by an interference in the cross-talk between human peripheral blood mononuclear cells (PBMC) and those from two colon carcinoma cell lines. Both non-stimulated PBMC and mononuclear cells induced for cytokine production by HT-29 and RKO cells from human colon carcinoma lines were incubated without and with 4, 20 and 100 µg/ml of pyridoxal hydrochloride (vitamin B6) and secretion of TNF-α, IL-1ß, IL-6, IFN-γ, IL-10, and IL-1ra was examined. Vit B6 caused a dose-dependent decrease in production of all cytokines examined, except for that of IL-1ra. The results indicate that vitamin B6 exerts an immunomodulatory effect on human PBMC. The finding that production of inflammatory cytokines is more pronounced when PBMC are in contact with malignant cells and markedly inhibited by the vitamin suggests an additional way by which vitamin B6 may exert its carcinopreventive effect.

Difference in inflammatory cytokine production by mononuclear cells from obese and non-obese schizophrenic patients.

OBJECTIVE: Schizophrenic patients have an increased risk for obesity compared with the general population. Evidence suggests the existence of an inflammatory process in the etiology of both obesity and schizophrenia. Our study compares in vitro secretion of inflammatory cytokines by peripheral blood mononuclear cells (PBMC) obtained from obese and non-obese schizophrenic patients. METHOD: Mononuclear cells were isolated from 20 obese (BMI >27) and 20 non-obese (BMI <24) schizophrenic in-patients. The levels of TNF-α, IL-1ß, IL-6, IL-1ra, IL-10 or IL-2 and IFN-γ in the supernatants of stimulated PBMC, as well as leptin and adiponectin serum values were evaluated. RESULTS: Peripheral blood mononuclear cells from patients in the obese group showed a significantly increased TNF-α and IL-1ß production, whereas the release of IL-1ra was decreased as compared with the non-obese group. In the obese group, the serum concentration of leptin was significantly higher and that of adiponectin was significantly lower. The results of the remaining cytokines did not differ between the two groups. CONCLUSION: Our study indicates the existence of a difference between obese and non-obese schizophrenic subjects as for inflammatory cytokine production and serum leptin and adiponectin levels, suggesting a 'subclinical inflammatory state' in obese schizophrenic patients that may contribute to a predisposition to inflammation and infections.

Correlation between movement of concanavalin A membrane receptors and cytolysis. A scanning electron microscopy study.

The present study was undertaken to test whether cytolysis induced by Concanavalin A (Con A) requires lateral mobility of membranal lectin receptor sites into caps. Treatment of interphase murine mastocytoma cells with 10(-4) M colchicine promoted cap formation by Con A in about 30% of the cells, followed by cytolysis. Pretreatment of the cells with NaN3, low temperature, or glutaraldehyde decreased the degree of capping and, to the same extent, the degree of cytolysis. The addition of antibodies to cells bound with Con A increased the appearance of capping and cytolysis. A linear relationship with a high correlation coefficient exists between the degree of capping and cytolysis, suggesting that lateral mobility of membrane Con A receptors is required for cytolysis by the lectin. The process of cap formation by Con A up to the stage of cytolysis was followed by scanning electron microscopy.

Erythropoietin effects on fetal mouse erythroid cells. I. Cell population and hemoglobin synthesis.

The effect of the hormone, erythropoietin, on cultures of erythroblasts derived from the livers of fetal C57BL/6J mice was examined. An increase both in the content and in the rate of synthesis of normal adult mouse globin chains was detected in hormone-treated cultures. The rate of protein synthesis by individual erythroblasts does not increase in response to the hormone, whereas the absolute number of hemoglobin-synthesizing cells does increase and accounts for the observed stimulation of hemoglobin synthesis. The principal effect of erythropoietin appears to be upon the population of immature erythroid precursor cells which persists in the presence of the hormone, the cells maintaining their ability to replicate, and their capacity to differentiate into hemoglobinizing erythroblasts. In the absence of hormone, already committed erythroblasts continue their development, but erythropoiesis is not sustained.

Immune response to influenza and pneumococcal vaccines in mice.

The immune response of mice injected with influenza vaccine (FluV) or pneumococcal vaccine (PV) given separately or simultaneously was evaluated. Balb/c mice were divided into six groups. Group I served as control, the mice in group II were injected intraperitoneally with PV, in group III intramuscularly with FluV two weeks after the onset of the study. The mice from group IV received PV and 2 weeks later were injected with FluV, mice in group V were given FluV, whereas group VI received both FluV and PV simultaneously. The results showed that the proliferative response of peripheral blood mononuclear cells (PBMC) significantly increased in animals from groups II, V and VI, whereas the proliferation of splenocytes increased in mice from groups II, III, IV, and VI. These observations indicate a comparable effect of both vaccines, at least when the proliferative response of PBMC and splenocytes were considered.

Induction of primary antiphospholipid syndrome in mice by immunization with a human monoclonal anticardiolipin antibody (H-3).

Antiphospholipid syndrome (APLS) is characterized by thrombocytopenia, thromboembolic phenomena, and recurrent fetal loss, associated with anticardiolipin antibodies (ACA) and/or lupus anticoagulant. The syndrome may be primary or may be associated with other conditions such as systemic lupus erythematosus. We have previously shown the ability to induce APLS in naive mice following passive transfer of serum and monoclonal ACAs. Similarly we generated the secondary APLS in BALB/c mice following immunization with a pathogenic anti-DNA antibody. In the current study we report on the induction of primary APLS following immunization of BALB/c mice with a human monoclonal ACA (H-3). The mice developed high persistent titers of ACA. The APLS was characterized by prolonged activated partial thromboplastin time, low fecundity rate (21% vs. 48% of control immunized mice), high resorption index of fetuses (25% vs. 3%), and low weights of embryos and placentae. Our study points to the ability of inducing primary APLS in naive mice. The induction of various presentations of APLS by different ACA may explain the diversity of clinical manifestations seen in patients with APLS.

Prevention of fetal loss in experimental antiphospholipid syndrome by in vivo administration of recombinant interleukin-3.

Antiphospholipid antibodies are strongly associated with arterial and venous thrombosis and with fetal loss. Recently an experimental model for antiphospholipid syndrome (APLS) was established in our laboratory. In this model, mice are immunized passively or actively with anticardiolipin antibodies and acquire the syndrome, which is characterized by prolonged activated partial thromboplastin time (APTT), thrombocytopenia, low fecundity rate, and fetal loss. In a normal process of pregnancy, lymphokines affect fetal implantation and development. Cytokines from the colony stimulating factor family, like GM-CSF and IL-3, were shown to be positive signals for implantation and to promote placental development and fetal growth. Given our preliminary findings of low IL-3 in mice with APLS and the efficacy of IL-3 in preventing fetal loss in a strain of mice prone to fetal resorption, our aim in the present study was to examine the effect of murine recombinant IL-3 (mrIL-3) on pregnant mice induced with experimental APLS. Mice were passively transfused to the tail vein, 24 h following mating, with anticardiolipin antibodies. The mice were divided into two groups: one group was injected intraperitoneally with mrIL-3 on days 6.5, 8.5, and 10.5 after mating, while the control group was injected with PBS. When the mice were killed on day 15 of pregnancy a 32% +/- 4.2 resorption rate was observed in the anti-cardiolipin-immunized group, which was reduced to 4% +/- 0.3 following treatment with mrIL-3. The thrombocytopenia associated with the experimental APLS was also corrected following lymphokine administration. IL-3 may be effective in prevention of recurrent fetal loss in APLS.

Lycopene affects proliferation and apoptosis of four malignant cell lines.

The beneficial effect of lycopene from tomatoes on a variety of chronic diseases and particularly its association with decreased incidence of prostate and breast cancer seems to be well established. The aim of the study was to examine its anti-proliferative and apoptotic effect on other malignant cell lines. Cells of the following lines were incubated with 1.0, 2.0, and 4.0microM of lycopene: human colon carcinoma (HuCC), B chronic lymphocytic leukemia (EHEB), human erythroleukemia (K562) and Raji, a prototype of Burkitt lymphoma cell line. The results showed that lycopene exerted a significant dose-dependent effect on the proliferation capacity of K562, Raji and HuCC lines, whereas this effect was observed in EHEB cells only with the highest dose used in the study. Increased apoptotic rate was found after incubation of HuCC cells with 2.0 and 4.0microM of lycopene and in Raji cells following incubation with 2.0microM. The findings point out that the anti-proliferative effect of lycopene on tumor cells and its effect on the apoptotic rate depends on its dosage and on the type of the malignant cells.

In vitro DNA and RNA synthesis by human platelets.

In vitro incorporation of [Me-3H] thymidine and [5-3H] uridine into human platelets was demonstrated. Thymidine incorporation was inhibited by three specific inhibitors of DNA synthesis: hydroxyurea, cytosine arabinoside and daunomycin. The effect was dose-dependent. Uridine uptake by platelets was found to be inhibited by specific inhibitors of RNA synthesis such as actinomycin D, rifampicin and vincristine, the effect of actinomycin D being dose dependent. The drug also led to a time-dependent inhibition of protein synthesis when preincubated with platelets. The platelet RNA profile on polyacrylamide gel was demonstrated to be similar to that of embryonic mouse erythroblast RNA. Synthesis of all three fractions, 28 S, 18 S and 4 S, was inhibited by actinomycin D. These findings show that human platelets are capable of DNA and RNA synthesis, and that these activities play a role in controlling protein synthesis in these cells. Detectable amounts of DNA have been found in whole human platelets, and in isolated mitochondria derived from these cells. Isolated platelet mitochondria incorporated [3H] thymidine and [3H] uridine into their macromolecules. These activities were inhibited by daunomycin and by both rifampicin and actinomycin D, respectively. These results support the assumption that DNA and RNA synthesis found in intact cell preparations takes place most probably in platelet mitochondria.

Effect of pentoxifylline on the phagocytic activity, cAMP levels, and superoxide anion production by monocytes and polymorphonuclear cells.

The effect of pentoxifylline (Trental) on the phagocytic capacity, cAMP levels, and superoxide anion production of human peripheral blood monocytes and polymorphonuclears (PMNs) was studied. The drug inhibited the phagocytosis of latex particles by both monocytes and PMNs in a dose-dependent manner. In addition, superoxide anion production during the phagocytic process was also reduced following incubation of the cells with pentoxifylline. It is suggested that this inhibitory effect is due to the increased intracellular levels of cAMP induced by the drug.

Increased salivary calcium levels as an indicator of digoxin intoxication.

One hundred ten individuals were divided into patients with digoxin intoxication; patients treated with digoxin; patients treated with digoxin and diuretics; patients treated with diuretics; and control subjects. Measurement of salivary potassium and calcium levels showed that 81% of the patients with digoxin intoxication had noticeable elevation of the salivary calcium level. In 22%, elevation of the salivary calcium level preceded clinical manifestations of intoxication. The high calcium level in the saliva was not accompanied by changes in serum or urinary calcium levels. The elevation of salivary calcium levels can be used not only as an additional indicator of digoxin intoxication but also for detecting impending intoxication in patients treated with this drug.

Phagocytotic activity of monocytes from diabetic patients.

The phagocytotic activity of monocytes from diabetic patients and healthy controls was studied. It was found that the number of phagocytizing cells from diabetic patients was significantly reduced in comparison with that from control individuals. However, the number of bacteria phagocytized per cell was similar in both groups. Plasma from healthy controls added to diabetic monocytes did not cause any significant change in their phagocytotic capacity. Addition of insulin to the plasma of diabetic patients failed to alter the number of phagocytizing diabetic monocytes. Similarly, addition of glucose to control plasma did not affect the number of control monocytes capable of phagocytosis. Protein synthesis was increased during phagocytosis in both control and diabetic cells. The importance of monocytes in the defense mechanism of the organism is discussed.

Cytotoxic effect of hemin and protoporphyrin on chronic lymphocytic leukemia lymphocytes.

Hemin and protoporphyrin exerted a cytotoxic effect on lymphocytes from 24 patients with chronic lymphocytic leukemia (CLL). The porphyrins inhibited protein and RNA synthesis in a dose dependent pattern. Exposure of leukemic cells to 15 microM hemin for 10 min reduced leucine incorporation to less than 20%, and 78% of the cells were freely permeable to trypan blue. Hemin at 0.17 microM had to be bound to 10(6) cells in the dark to produce this killing effect. On the other hand, the control lymphocytes from 20 healthy subjects were relatively resistant to the hemin effect, and cell damage was reversible. Fetal calf serum (FCS) protected most leukemic lymphocytes from prophyrin toxicity. The combination of human hemoglobin with free protoporphyrin showed a synergistic toxicity, and this combination was the most potent inhibitor of protein synthesis in the presence of serum. 55Fe-hemin was preferentially bound to leukemic lymphocytes, and the binding site of protoporphyrin was observed to be on the cell surface membrane. The hemin quenching effect of diphenyl hexatriene (DPH) stained cell fluorescence indicated that hemin was apparently bound to the lipid layer of the outer membrane. Scanning electron microscopy (SEM) of hemin treated cells revealed an initial development of membranal blebs, followed by total destruction of the cell surface membrane. Transmission electron microscopy (TEM) of the treated cells showed a profound damage in the cytoplasma and nucleus. Control lymphocytes treated by hemin appeared relatively undamaged and free of membranal blebs.

Lectin-like effect of Echis coloratus Venom on human and mouse lymphocytes.

Echis coloratus venom (ECV) treated human and mouse lymphocytes were examined for synthesizing activities and morphologic alterations. RNA, DNA and protein synthesis were markedly inhibited. Human cells were less affected than mouse lymphocytes. No mitogenic activity was observed. Transmission (TEM) and scanning electron microscopy (SEM) revealed membranal damage in all types of lymphocytes and a considerable degree of agglutination. Human lymphocytes showed concentration of the microvilli at one pole of the cell. This phenomenon was considered as a capping effect. The action of ECV may be compared with the effect of the nonmitogenic lectin RCAII, known to penetrate the cells by endocytosis and to inhibit their protein synthesis.

The role of hemin in the regulation of heme synthesis by fetal mouse liver erythroblasts in culture.

The regulatory role of exogenous hemin on the heme synthetic pathway was studied in fetal mouse liver erythroblasts in culture. Hemin added to culture medium of 13th day embryo liver cells inhibited, dose dependently, the incorporation of the porphyrin precursors, 59Fe, 14C-2-glycine and 14C-5-aminolevulinic acid (ALA) by 85%, 70% and 45%, respectively. This suggests a multiple effect of hemin on the porphyrin biosynthetic enzymes. Exogenous ALA competed with 14 C-2-glycine as a porphyrin precursor, but the rate of heme synthesis, measured by 59Fe incorporation, remained unaltered. Protoporphyrin mimicked the hemin effect on the inhibition of glycine incorporation into heme, but reduced iron incorporation by only 20%. Erythroblasts, with an inhibited porphyrin biosynthesis, utilized exogenous 59Fe-hemin for hemoglobin assembly and maintained an undecreased level of hemoglobin synthesis. The results indicate that hemin inhibits the porphyrin biosynthesis in fetal mouse liver erythroblasts mainly at the iron incorporation stage.

Inhibition of leukemic cell proliferation by one or more factors released from splenic BCL1 cells.

Irradiated peripheral blood lymphocytes from patients with chronic B-lymphocytic leukemia (B-CLL) have been shown to secrete a factor or factors that caused inhibition of malignant cell proliferation. In the present study, we used the murine B cell leukemia (BCL1) model system to examine the possible secretion of inhibitory factors from irradiated leukemic spleen cells. It was found that under culture conditions, irradiated spleen cells obtained from leukemic mice produce factors capable of suppressing BCL1 cell proliferation in vitro. The release of an inhibitory factor from nonirradiated cells was also observed, albeit to a lower extent. Supernatants collected from cultured nonirradiated and irradiated cells derived from animals at the first week after BCL1 cell inoculation did not affect the proliferation capacity of cells at any stage of the disease, whereas supernatants obtained at more advanced stages exhibited an inhibitory effect on the proliferation of cells derived from the same stages. The suppressive effect of supernatants of irradiated cells was more pronounced when those of 7 days' culture were used, compared to those of nonirradiated cells. The difference in suppression observed between supernatants of irradiated and nonirradiated BCL1 cells may be attributed either to the same factor produced in a larger amount following irradiation or to two (or more) factors--one produced spontaneously, the other induced by irradiation.

Effect of a low dose of vincristine on platelet production in mice.

The effect of a single low dose (0.1 mg/kg) of vincristine (VCR) on platelet production was investigated in C57B1 mice. A parallel increase of circulating platelets and (75Se)-selenomethionine (Se-Met) uptake was observed. The total megakaryocyte count decreased insignificantly 6 hours after VCR injection, followed by an increase after 18 hours. Plasma taken from mice 24 hours after VCR injection was tested for thrombopoietic activity. The post-VCR plasma caused a significant thrombocytosis, increased Se-Met uptake and increased protein synthesis of the platelets, indicating an active overproduction of platelets. These results suggest that the thrombocytosis induced by a low dose of VCR is mediated by a thrombopoietin-like substance.

Erythroid precursor fusion induced by Sendai virus.

The ultrastructural events of the interaction of Sendai virus (SV) with fetal mouse erythroid precursors, and SV-induced fusion of erythroid precursors at different maturation stages are described. SV was shown to affect the erythroid nuclei causing interruption of the nuclear membrane, enlargement of the nucleolus and nuclear fusion. SV induced fusion also between dividing and non-dividing cells.

SEM observations on Sendai virus-induced fusion of embryonic mouse erythroblasts.

The events of fusion of 11- and 16-day embryonic mouse liver erythroblasts induced by Sendai virus (SV) were followed with the scanning electron microscope (SEM). Erythroid precursors incubated with the virus showed numerous 'pores' on the cell membrane. The cell fusion began with the appearance of a fine 'meshwork' structure between the adjacent cells, followed by a gradual formation of a common cell membrane and terminated with the appearance of polykaryons, in which the nuclei were easily recognized when located in the vicinity of the polykaryon membrane. There was no difference in the process of fusion between erythroblasts at identical and those at different stages of maturation.