Immunotoxicity of stainless-steel nanoparticles obtained after 3D printing.

This study aims to investigate the in vitro effects of nanoparticles (NPs) produced during the selective laser melting (SLM) of 316 L stainless steel metal powder on the immune response in a human blood model. Experimental data did not reveal effect on viability of 316 L NPs for the tested doses. Functional immune assays showed a significant immunosuppressive effect of NPs. There was moderate stimulation (117%) of monocyte phagocytic activity without significant changes in phagocytic activity and respiratory burst of granulocytes. A significant dose-dependent increase in the levels of the pro-inflammatory cytokine TNF-a was found in blood cultures treated with NPs. On the contrary, IL-8 chemokine levels were significantly suppressed. The levels of the pro-inflammatory cytokine IL-6 were reduced by only a single concentration of NPs. These new findings can minimise potential health risks and indicate the need for more research in this area.

Association of Inflammatory and Oxidative Status Markers with Metabolic Syndrome and Its Components in 40-to-45-Year-Old Females: A Cross-Sectional Study.

Oxidative stress and sterile inflammation play roles in the induction and maintenance of metabolic syndrome (MetS). This study cohort included 170 females aged 40 to 45 years who were categorized according to the presentation of MetS components (e.g., central obesity, insulin resistance, atherogenic dyslipidemia, and elevated systolic blood pressure) as controls not presenting a single component (n = 43), those with pre-MetS displaying one to two components (n = 70), and females manifesting MetS, e.g., ≥3 components (n = 53). We analyzed the trends of seventeen oxidative and nine inflammatory status markers across three clinical categories. A multivariate regression of selected oxidative status and inflammatory markers on the components of MetS was performed. Markers of oxidative damage (malondialdehyde and advanced-glycation-end-products-associated fluorescence of plasma) were similar across the groups. Healthy controls displayed lower uricemia and higher bilirubinemia than females with MetS; and lower leukocyte counts, concentrations of C-reactive protein, interleukine-6, and higher levels of carotenoids/lipids and soluble receptors for advanced glycation end-products than those with pre-MetS and MetS. In multivariate regression models, levels of C-reactive protein, uric acid, and interleukine-6 were consistently associated with MetS components, although the impacts of single markers differed. Our data suggest that a proinflammatory imbalance precedes the manifestation of MetS, while an imbalance of oxidative status accompanies overt MetS. Further studies are needed to elucidate whether determining markers beyond traditional ones could help improve the prognosis of subjects at an early stage of MetS.

Titanium Dioxide Nanoparticles Modulate Systemic Immune Response and Increase Levels of Reduced Glutathione in Mice after Seven-Week Inhalation.

Titanium dioxide nanoparticles (TiO2 NPs) are used in a wide range of applications. Although inhalation of NPs is one of the most important toxicologically relevant routes, experimental studies on potential harmful effects of TiO2 NPs using a whole-body inhalation chamber model are rare. In this study, the profile of lymphocyte markers, functional immunoassays, and antioxidant defense markers were analyzed to evaluate the potential adverse effects of seven-week inhalation exposure to two different concentrations of TiO2 NPs (0.00167 and 0.1308 mg TiO2/m3) in mice. A dose-dependent effect of TiO2 NPs on innate immunity was evident in the form of stimulated phagocytic activity of monocytes in low-dose mice and suppressed secretory function of monocytes (IL-18) in high-dose animals. The effect of TiO2 NPs on adaptive immunity, manifested in the spleen by a decrease in the percentage of T-cells, a reduction in T-helper cells, and a dose-dependent decrease in lymphocyte cytokine production, may indicate immunosuppression in exposed mice. The dose-dependent increase in GSH concentration and GSH/GSSG ratio in whole blood demonstrated stimulated antioxidant defense against oxidative stress induced by TiO2 NP exposure.

Copper Oxide Nanoparticles Stimulate the Immune Response and Decrease Antioxidant Defense in Mice After Six-Week Inhalation.

Copper oxide nanoparticles (CuO NPs) are increasingly used in various industry sectors. Moreover, medical application of CuO NPs as antimicrobials also contributes to human exposure. Their toxicity, including toxicity to the immune system and blood, raises concerns, while information on their immunotoxicity is still very limited. The aim of our work was to evaluate the effects of CuO NPs (number concentration 1.40×106 particles/cm3, geometric mean diameter 20.4 nm) on immune/inflammatory response and antioxidant defense in mice exposed to 32.5 µg CuO/m3 continuously for 6 weeks. After six weeks of CuO NP inhalation, the content of copper in lungs and liver was significantly increased, while in kidneys, spleen, brain, and blood it was similar in exposed and control mice. Inhalation of CuO NPs caused a significant increase in proliferative response of T-lymphocytes after mitogenic stimulation and basal proliferative activity of splenocytes. CuO NPs significantly induced the production of IL-12p70, Th1-cytokine IFN-γ and Th2-cytokines IL-4, IL-5. Levels of TNF-α and IL-6 remained unchanged. Immune assays showed significantly suppressed phagocytic activity of granulocytes and slightly decreased respiratory burst. No significant differences in phagocytosis of monocytes were recorded. The percentage of CD3+, CD3+CD4+, CD3+CD8+, and CD3-CD19+ cell subsets in spleen, thymus, and lymph nodes did not differ between exposed and control animals. No changes in hematological parameters were found between the CuO NP exposed and control groups. The overall antioxidant protection status of the organism was expressed by evaluation of GSH and GSSG concentrations in blood samples. The experimental group exposed to CuO NPs showed a significant decrease in GSH concentration in comparison to the control group. In summary, our results indicate that sub-chronic inhalation of CuO NPs can cause undesired modulation of the immune response. Stimulation of adaptive immunity was indicated by activation of proliferation and secretion functions of lymphocytes. CuO NPs elicited pro-activation state of Th1 and Th2 lymphocytes in exposed mice. Innate immunity was affected by impaired phagocytic activity of granulocytes. Reduced glutathione was significantly decreased in mice exposed to CuO NPs.

Six-week inhalation of CdO nanoparticles in mice: The effects on immune response, oxidative stress, antioxidative defense, fibrotic response, and bones.

Due to the growing number of applications of cadmium oxide nanoparticles (CdO NPs), there is a concern about their potential deleterious effects. The objective of our study was to investigate the effect of CdO NPs on the immune response, renal and intestine oxidative stress, blood antioxidant defence, renal fibrotic response, bone density and mineral content. Six-week-old female ICR mice were exposed to CdO NPs for 6 weeks by inhalation (particle size: 9.82 nm, mass concentration: 31.7 µg CdO/m3, total deposited dose: 0.195 µg CdO/g body weight). CdO NPs increased percentage of thymus CD3e+CD8a+ cells and moderately enhanced splenocyte proliferation and production of cytokines and chemokines. CdO NPs elevated pro-fibrotic factors (TGF-ß2, α-SMA and collagen I) in the kidney, and concentrations of AGEs in the intestine. The ratio of GSH and GSSG in blood was slightly reduced. Exposure to CdO NPs resulted in 10-fold higher Cd concentration in tibia bones. No differences were found in bone mass density, mineral content, bone area values, bone concentrations of Ca, P, Mg and Ca/P ratio. Our findings indicate stimulation of immune/inflammatory response, oxidative stress in the intestine, starting fibrotic response in kidneys and accumulation of CdO NPs in bones of mice.

Toxicity evaluation of monodisperse PEGylated magnetic nanoparticles for nanomedicine.

Innovative nanotechnology aims to develop particles that are small, monodisperse, smart, and do not cause unintentional side effects. Uniform magnetic Fe3O4 nanoparticles (12 nm in size) were prepared by thermal decomposition of iron(III) oleate. To make them colloidally stable and dispersible in water and cell culture medium, they were modified with phosphonic acid- (PA) and hydroxamic acid (HA)-terminated poly(ethylene glycol) yielding PA-PEG@Fe3O4 and HA-PEG@Fe3O4 nanoparticles; conventional γ-Fe2O3 particles were prepared as a control. Advanced techniques were used to evaluate the properties and safety of the particles. Completeness of the nanoparticle coating was tested by real-time polymerase chain reaction. Interaction of the particles with primary human peripheral blood cells, cellular uptake, cytotoxicity, and immunotoxicity were also investigated. Amount of internalized iron in peripheral blood mononuclear cells was 72, 38, and 25 pg Fe/cell for HA-PEG@Fe3O4, γ-Fe2O3, and PA-PEG@Fe3O4, respectively. Nanoparticles were localized within the cytoplasm and in the extracellular space. No cytotoxic effect of both PEGylated nanoparticles was observed (0.12-75 µg/cm2) after 24 and 72-h incubation. Moreover, no suppressive effect was found on the proliferative activity of T-lymphocytes and T-dependent B-cell response, phagocytic activity of monocytes and granulocytes, and respiratory burst of phagocytes. Similarly, no cytotoxic effect of γ-Fe2O3 particles was observed. However, they suppressed the proliferative activity of T-lymphocytes (75 µg/cm2, 72 h) and also decreased the phagocytic activity of monocytes (15 µg/cm2, 24 h; 3-75 µg/cm2, 72 h). We thus show that newly developed particles have great potential especially in cancer diagnostics and therapy.

Humoral and cellular immune response in Wistar Han RCC rats fed two genetically modified maize MON810 varieties for 90 days (EU 7th Framework Programme project GRACE).

The genetically modified maize event MON810 expresses a Bacillus thuringiensis-derived gene, which encodes the insecticidal protein Cry1Ab to control some lepidopteran insect pests such as the European corn borer. It has been claimed that the immune system may be affected following the oral/intragastric administration of the MON810 maize in various different animal species. In the frame of the EU-funded project GRACE, two 90-day feeding trials, the so-called studies D and E, were performed to analyze the humoral and cellular immune responses of male and female Wistar Han RCC rats fed the MON810 maize. A MON810 maize variety of Monsanto was used in the study D and a MON810 maize variety of Pioneer Hi-Bred was used in the study E. The total as well as the maize protein- and Cry1Ab-serum-specific IgG, IgM, IgA and IgE levels, the proliferative activity of the lymphocytes, the phagocytic activity of the granulocytes and monocytes, the respiratory burst of the phagocytes, a phenotypic analysis of spleen, thymus and lymph node cells as well as the in vitro production of cytokines by spleen cells were analyzed. No specific Cry1Ab immune response was observed in MON810 rats, and anti-maize protein antibody responses were similar in MON810 and control rats. Single parameters were sporadically altered in rats fed the MON810 maize when compared to control rats, but these alterations are considered to be of no immunotoxicological significance.

The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status.

The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The significant changes of cytotoxic, naive, and memory T-lymphocytes, plasmacytoid dendritic cells (DCs) were in postmenopausal women versus fertile women. Body mass index (BMI) affected markedly the cell surface expression of CD265/RANK. Osteoporosis is linked to reduced percentage of plasmacytoid DCs, and elevated natural Treg cells (p < 0.05). The confounding factors such as women age, BMI, bone mineral density (BMD), waist size and tissue fat affect the expression of RANK on myeloid DCs and CD40L on T-lymphocytes that might be the immunophenotypic modulators after menopause.

The effect of core and lanthanide ion dopants in sodium fluoride-based nanocrystals on phagocytic activity of human blood leukocytes.

Sodium fluoride-based ß-NaLnF4 nanoparticles (NPs) doped with lanthanide ions are promising materials for application as luminescent markers in bio-imaging. In this work, the effect of NPs doped with yttrium (Y), gadolinium (Gd), europium (Eu), thulium (Tm), ytterbium (Yb) and terbium (Tb) ions on phagocytic activity of monocytes and granulocytes and the respiratory burst was examined. The surface functionalization of <10-nm NPs was performed according to our variation of patent pending ligand exchange method that resulted in meso-2,3-dimercaptosuccinic acid (DMSA) molecules on their surface. Y-core-based NCs were doped with Eu ions, which enabled them to be excited with UV light wavelengths. Cultures of human peripheral blood (n = 8) were in vitro treated with five different concentrations of eight NPs for 24 h. In summary, neither type of nanoparticles is found toxic with respect to conducted test; however, some cause toxic effects (they have statistically significant deviations compared to reference) in some selected doses tested. Both core types of NPs (Y-core and Gd-core) impaired the phagocytic activity of monocytes the strongest, having minimal or none whatsoever influence on granulocytes and respiratory burst of phagocytic cells. The lowest toxicity was observed in Gd-core, Yb, Tm dopants and near-infrared nanoparticles. Clear dose-dependent effect of NPs on phagocytic activity of leukocytes and respiratory burst of cells was observed for limited number of samples.

Functionalized porous silica&maghemite core-shell nanoparticles for applications in medicine: design, synthesis, and immunotoxicity.

AIM: To determine cytotoxicity and effect of silica-coated magnetic nanoparticles (MNPs) on immune response, in particular lymphocyte proliferative activity, phagocytic activity, and leukocyte respiratory burst and in vitro production of interleukin-6 (IL-6) and 8 (IL-8), interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), and granulocyte macrophage colony stimulating factor (GM-CSF). METHODS: Maghemite was prepared by coprecipitation of iron salts with ammonia, oxidation with NaOCl and modified by tetramethyl orthosilicate and aminosilanes. Particles were characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS), Fourier-transform infrared (FTIR), and X-ray photoelectron spectroscopy (XPS). Cytotoxicity and lymphocyte proliferative activity were assessed using [3H]-thymidine incorporation into DNA of proliferating human peripheral blood cells. Phagocytic activity and leukocyte respiratory burst were measured by flow cytometry; cytokine levels in cell supernatants were determined by ELISA. RESULTS: γ-Fe2O3&SiO2-NH2 MNPs were 13 nm in size. According to TEM, they were localized in the cell cytoplasm and extracellular space. Neither cytotoxic effect nor significant differences in T-lymphocyte and T-dependent B-cell proliferative response were found at particle concentrations 0.12-75 µg/cm2 after 24, 48, and 72 h incubation. Significantly increased production of IL-6 and 8, and GM-CSF cytokines was observed in the cells treated with 3, 15, and 75 µg of particles/cm2 for 48 h and stimulated with pokeweed mitogen (PHA). No significant changes in TNF-α and IFN-γ production were observed. MNPs did not affect phagocytic activity of monocytes and granulocytes when added to cells for 24 and 48 h. Phagocytic respiratory burst was significantly enhanced in the cultures exposed to 75 µg MNPs/cm2 for 48 h. CONCLUSIONS: The cytotoxicity and in vitro immunotoxicity were found to be minimal in the newly developed porous core-shell γ-Fe2O3&SiO2-NH2 magnetic nanoparticles.

Immunotoxicity and genotoxicity testing of PLGA-PEO nanoparticles in human blood cell model.

A human blood cell model for immunotoxicity and genotoxicity testing was used to measure the response to polylactic-co-glycolic acid (PLGA-PEO) nanoparticle (NP) (0.12, 3, 15 and 75 µg/cm(2) exposure in fresh peripheral whole blood cultures/isolated peripheral blood mononuclear cell cultures from human volunteers (n = 9-13). PLGA-PEO NPs were not toxic up to dose 3 µg/cm(2); dose of 75 µg/cm(2) displays significant decrease in [(3)H]-thymidine incorporation into DNA of proliferating cells after 4 h (70% of control) and 48 h (84%) exposure to NPs. In non-cytotoxic concentrations, in vitro assessment of the immunotoxic effects displayed moderate but significant suppression of proliferative activity of T-lymphocytes and T-dependent B-cell response in cultures stimulated with PWM > CON A, and no changes in PHA cultures. Decrease in proliferative function was the most significant in T-cells stimulated with CD3 antigen (up to 84%). Cytotoxicity of natural killer cells was suppressed moderately (92%) but significantly in middle-dosed cultures (4 h exposure). On the other hand, in low PLGA-PEO NPs dosed cultures, significant stimulation of phagocytic activity of granulocytes (119%) > monocytes (117%) and respiratory burst of phagocytes (122%) was recorded. Genotoxicity assessment revealed no increase in the number of micronucleated binucleated cells and no induction of SBs or oxidised DNA bases in PLGA-PEO-treated cells. To conclude on immuno- and genotoxicity of PLGA-PEO NPs, more experiments with various particle size, charge and composition need to be done.

Hydrophobic sodium fluoride-based nanocrystals doped with lanthanide ions: assessment of in vitro toxicity to human blood lymphocytes and phagocytes.

In vitro immunotoxicity of hydrophobic sodium fluoride-based nanocrystals (NCs) doped with lanthanide ions was examined in this study. Although there is already a significant amount of optical and structural data on NaYF4 NCs, data on safety assessment are missing. Therefore, peripheral whole blood from human volunteers was used to evaluate the effect of 25 and 30 nm hydrophobic NaYF4 NCs dissolved in cyclohexane (CH) on lymphocytes, and of 10 nm NaYF4 NCs on phagocytes. In the concentration range 0.12-75 µg cm(-2) (0.17-106 µg ml(-1) ), both 25 and 30nm NaYF4 NCs did not induce cytotoxicity when measured as incorporation of [(3) H]-thymidine into DNA. Assessment of lymphocyte function showed significant suppression of the proliferative activity of T-lymphocytes and T-dependent B-cell response in peripheral blood cultures (n = 7) stimulated in vitro with mitogens phytohemagglutinin (PHA) and pokeweed (PWM) (PHA > PWM). No clear dose-response effect was observed. Phagocytic activity and respiratory burst of leukocytes (n = 5-8) were generally less affected. A dose-dependent suppression of phagocytic activity of granulocytes in cultures treated with 25 nm NCs was observed (vs. medium control). A decrease in phagocytic activity of monocytes was found in cells exposed to higher doses of 10 and 30 nm NCs. The respiratory burst of phagocytes was significantly decreased by exposure to the middle dose of 30 nm NCs only. In conclusion, our results demonstrate immunotoxic effects of hydrophobic NaYF4 NCs doped with lanthanide ions to lymphocytes and to lesser extent to phagocytes. Further research needs to be done, particularly faze transfer of hydrophobic NCs to hydrophilic ones, to eliminate the solvent effect.

Association between the human immune response and body mass index.

The aim of this study was to determine the strength of the association between the human immune response and body mass index (BMI) and whether differences exist in the effects of obesity on selected immune parameters between men and women. Two hundred ninety participants were divided into groups according to sex and BMI. Parameters CD3, CD4, CD8, CD16+56, CD19, HLADR, CD11b, CD11c, and CD54 were quantified. Leukocyte and differential counts were performed. We observed elevation with regard to the normal weight group in the parameters of white blood cells, neutrophils, monocytes, CD3, CD4, CD19, and CD11b for the whole study group. A decrease was observed in the expression of CD16+56. The effect of BMI on the immune system was much more apparent in women. BMI was correlated with the majority of the measured parameters, reflecting a strong association between BMI and the human immune system.

Changes in immunologic parameters of humoral immunity and adipocytokines in obese persons are gender dependent.

The aim of this study was to investigate several immunologic parameters using of immunonephelometry and adipocytokines by the enzyme immunoassay and their changes in different states of obesity. Obesity is considered to involve a state of chronic low-grade inflammation, with links between adipose cells and the immune system. We found significantly higher complement C3 levels in all obese subjects. Levels of the complement C4 were significantly higher in obese women, but not in men, when compared with the corresponding group of normal weight subjects. The increase in C-reactive protein concentrations was significant in both obese and morbidly obese women, but only in morbidly obese men. No significant differences in tumor necrosis factor-α, interleukin-6, interleukin-10, and soluble intercellular adhesion molecule-1 were found. sE-selectin levels were higher in both overweight and obese women but only in morbidly obese men. We found decreased adiponectin concentrations in obese and morbidly obese women. Concentrations of leptin were significantly higher only in obese men (p < 0.05), whereas in women the increase in leptin levels was significant in overweight, obese, and morbidly obese subjects. In conclusion, our results demonstrate elevated levels of C3, C-reactive protein, sE-selectin, and leptin in obese women and men. In obese women, we also observed increased concentrations of C4 and decreased levels of adiponectin.

Effect of berberine on proliferation, biosynthesis of macromolecules, cell cycle and induction of intercalation with DNA, dsDNA damage and apoptosis in Ehrlich ascites carcinoma cells.

Our primary aim was to study berberine, a potential anti-cancer drug, for its cytotoxic and antiproliferative activity in-vitro using Ehrlich ascites carcinoma (EAC) cells. Cytotoxicity was measured by the growth inhibition assay. We investigated the effect of berberine on the biosynthesis of macro-molecules (DNA, RNA, proteins), cell cycle effects and induction of dsDNA damage and apoptosis in berberine-treated EAC cells. Our results showed that berberine acts cytotoxically on EAC cells. The cytotoxicity was directly concentration and time dependent. The highest cytotoxic concentrations (100 and 50 microg mL(-1)) induced intercalation of berberine with DNA, formation of dsDNA breaks, inhibition of DNA synthesis and death of EAC cells. A concentration of 10 mug mL(-1) induced clear apoptotic cell death, which was followed by inhibition of protein synthesis.

Immunological monitoring in workers occupationally exposed to asbestos.

Occupational exposure to asbestos is strongly associated with pulmonary diseases, cancer and immunotoxic effects. Both systemic and local immunity may play an important role in the pathogenesis of these events. Immune cells appear to be influenced by asbestos exposure, either through direct effects or as a result of the host's protective response to exposure. In this study several immune system parameters were assessed in workers (n = 61) with at least 5 years' exposure to asbestos at an industrial plant. Workers exposed to asbestos fibres had significantly increased levels of immunoglobulin E and concentrations of interleukin-6 and -8 in comparison with two sets of controls (in-plant and town control groups). The levels of soluble adhesion molecule ICAM-1 were higher in the exposed group compared to the town control group. Significantly increased levels of IgA were found in asbestos-exposed group in comparison to the town control. Evaluation of the expression of adhesion molecules on lymphocytes, monocytes and granulocytes by flow cytometry showed significant increases in the class of selectins CD62L on monocytes and granulocytes. Moreover, significantly increased expression of markers CD69 and CD66b on eosinophils was found among workers exposed to asbestos. In conclusion, exposure to asbestos fibres was found to have several effects on immune system. Alterations of these immune parameters may indicate hypersensitivity (increased levels of IgE, increased expression of activation markers CD66b and CD69 on eosinophils) and an elevated inflammatory status (increased levels of interleukins--IL-6, IL-8) in exposed workers.

Immunomodulatory effects of mineral fibres in occupationally exposed workers.

In the context of a large-scale molecular epidemiology study, the possible immunomodulatory effects of mineral fibres, in workers occupationally exposed to asbestos, rockwool and glass fibres, were examined. In each plant, 61, 98 and 80 exposed workers and 21, 43 or 36 control clerical subjects, respectively, were recruited. In the case of the asbestos-exposed subjects, an additional town-control group of 49 people was included. Evidence of pulmonary fibrosis was found in 42% of the asbestos-exposed workers, while evidence of pleural fibrosis was found in 24%. The asbestos-exposed cohort had significantly decreased forced vital capacity of lungs as well as forced expiratory volume per first second. Our findings indicate that exposure to all three types of fibres examined modulates to different degrees the immune response. Suppression of T-cell immunity and to a lesser extent, B-cell immunity was found in the case of workers from a former asbestos cement plant, while stimulation of T-cell response was observed in rockwool workers, and stimulation of T- and B-cell response was seen in glass fibre workers. Depression of the percentage of lymphocyte subpopulation of CD 16+56 (natural killer cells) in peripheral blood was found in glass fibre workers. Statistical analysis showed increased levels of proinflammatory cytokines (IL-6 asbestos; IL-8 all three fibres), expression of adhesion molecule L-selectin on granulocytes and monocytes (asbestos), levels of soluble adhesion molecules (SAMs) in sera (ICAM-1 all three fibres; E-selectin glass fibres), increased levels of immunoglobulin E (asbestos and rockwool) and elevated expression of activation markers on eosinophils (CD66b asbestos, glass fibres; CD69 asbestos). Significant correlations were observed between lymphocyte proliferation and markers of DNA damage and repair. Increased levels of proinflammatory cytokines, SAMs, immunoglobulin E and elevated expression of activation markers on eosinophils was found in people with symptoms of hypersensitivity and an elevated inflammatory status.

Immunotoxic and cancerostatic effects of ethyl-4-isothiocyanatobutanoate in female Lewis rats with implanted fibrosarcoma.

Isothiocyanates (ITCs) have been isolated from plants. Naturally occurring and synthetic ITCs are known as effective chemopreventive agents. Ethyl 4-isothiocyanatobutanoate (E-41B) is a derivative of gamma-aminobutyric acid. Immunotoxic and canocerostatic effects of E-41B in female inbred Lewis rats implanted with experimental fibrosarcoma BP6-TU2 was evaluated in this study. On day 5 after subcutaneous application of tumor cells, animals started to be treated intraperitoneally three times a week with two different doses of E-41B: 28 and 35 mg/kg/day during 28 days. High dose of E-41B was close to maximum tolerated dose (MTD). Control groups of rats with or without tumors injected intraperitoneally only saline or 70% dimethylsulphoxide were added. Administrating of E-41B resulted in suppression of thymus, popliteal lymph node, spleen weight and spleen cellularity. Hematologic evaluation displayed decreased erythrocyte (ERY) count and level of hemoglobin (HB) in rats treated withE-41B. Immune assays--the phagocytic activity of polymorphonuclear leukocytes (PMN) and monocytes, primary antibody response and in vitro proliferative activity of spleen lymphocytes (LY) to mitogens were not significantly affected by E-41B treatment E-41B moderately decreased tumor weights, but this decrease was not statistically significant in comparison with DMSO-exposed rats with tumors. The fibrosarcoma implantation itself increased significantly spleen weight and changed hematological parameters (decreased HB, increased mean cell volume of ERY, increased leukocyte count, increased % PMN, decreased % LY, decreased % EO). Moreover, moderate decreased percentage of CD161+ positive cells (NK cells) were found in peripheral blood. Immune assays showed decline in proliferation of lymphocytes and phagocytic activity of leukocytes. Our findings indicate that administration of E-41B displayed hematoxic effect in rats implanted with fibrosarcoma. Immunotoxic effect was shown as decreased lymphoid organ weight and spleen cytotoxicity although function of immune cells was not impaired.