Krzysztof Krzystyniak

Trichothecene mycotoxins in the dust of ventilation systems in office buildings

SummaryAnalysis of trichothecene mycotoxins in dust samples from ventilation systems of office buildings was applied as a rapid and inexpensive method for the detection of mycotoxins. Dust samples from three different office spaces of the Montreal urban area, reportedly affected by the “sick buildings syndrome”, were analysed by thin-layer chromatography (TLC). Positive colour reaction on TLC plates with 4-(p-nitrobenzyl) pyridine, specific for the 12,13-epoxy group in the trichothecene nucleus, was obtained for the extracts of 0.5- to 50-g dust samples. The dust samples contained at least four trichothecenes: T-2 toxin, diacetoxyscirpenol, roridine A and T-2 tetraol. The results were confirmed by high-performance liquid chromatography analysis. Screening of dust samples from air ventilation systems of reportedly affected buildings provided direct evidence of trichothecene mycotoxins, with the detection limit estimated as 0.4-4 ng/mg dust. Thus, the dust sample analysis is suggested as a rapid technique for detecting the presence of mycotoxins in the dust of ventilation systems.

Heavy metal-specific inhibition of phagocytosis and different in vitro sensitivity of heterogeneous coelomocytes from Lumbricus terrestris (Oligochaeta)

Cell viability and phagocytic activity of coelomocytes from the gastrointestinal tract of Lumbricus terrestris were examined by flow cytometry after in vitro exposure to heavy metals. Control coelomocytes were incubated for 18 h at 15 degrees C, 5% CO2, in Ca(++)-containing LBSS medium with 10(-4)-10(-9) M mercury chloride, methylmercury, cadmium chloride, zinc chloride, lead chloride or lead acetate. Heterogeneity of coelomocyte population was demonstrated by forward scatter (FSC) analysis and cytometric profile showing two different populations of type I/small (60%) and type-II/large (40%) cells. Exposure to either form of Hg, Cd and Zn was relatively highly toxic and affected both cell viability and phagocytosis, whereas Pb was relatively well tolerated by the coelomocytes. A fraction of cells within large coelomocyte population was exceptionally sensitive to the Hg-induced cytotoxicity, which did not affect, however, the relative phagocytic activity of the remaining cells. Overall, at least three different patterns of metal-specific toxicity, affecting both viability and phagocytic functions of earthworm coelomocytes, were confirmed in our in vitro studies. Further characterisation of both the target cells from heterogeneous coelomocyte population and the specific interaction of target cell-xenobiotic can possibly reduce biomonitoring problems in earthworm toxicology and immunotoxicology.

Limited immunotoxic potential of technical formulation of the herbicide atrazine (AAtrex) in mice

Immunotoxicity of the technical atrazine formulation, AAtrex, was examined in C57Bl/6 female mice following a sublethal exposure to equivalent 1/2-1.64 LD50 doses of the herbicide. Animal weight was not affected by the herbicide exposure. No dose-related changes could be concluded for fluctuations in organ weight, changes in the spleen cell number and cell viability. Furthermore, cytofluorometric studies showed no significant changes in the frequency of L3T4-positive and Lyt-2-positive T-cells. Functional in vitro assays of mitogen activation showed no marked effects of AAtrex exposure on lymphocyte stimulation by lipopolysaccharide (LPS), phytohemagglutinin (PHA) and concanavalin A (Con-A). In addition, sublethal exposure to AAtrex did not affect interleukin-2 (IL-2) production by splenic cells. Furthermore, no dose-related effect could be concluded from a transient suppression of a primary humoral IgM response to sheep erythrocytes (SRBC) as well as from a transient inhibition of a specific T-cell response to alloantigens in mixed lymphocyte reaction (MLR). Exposure to equiv. 1/2-1/16 LD50 doses augmented phagocytic activity of peritoneal macrophages, without any visible AAtrex dose-related effect. Normal humoral and cellular responses were restored at 14-40 days after the herbicide exposure. Overall, transient and reversible immunosuppression of humoral-mediated and cell-mediated responses and activated macrophage phagocytic activity could not be attributed to the direct chemical-related effect of sublethal exposure to AAtrex.

Combined effects of selected insecticides on humoral immune response in mice

Biological effects data with single insecticides are far more abundant than with mixtures. These data cannot be used directly to predict the effects of insecticide mixtures. Three insecticides of different chemical classes: organochlorine; dieldrin, organophosphate; malathion, and carbamate; carbofuran, previously evaluated for their immunotoxic potential, were selected for studies of combined acute exposure in C57B1/6 inbred mice. The humoral response to sheep red blood cells (SRBC) and the functional activities of peritoneal macrophages, such as phagocytosis of fluorescent beads and presentation of a single protein antigen, avidin, were examined after in vivo exposure of mice to different combinations of the selected pesticides and compared with the vehicle controls. Regarding exposure to single substances, the data confirmed the immunosuppressive potential of dieldrin and carbofuran and the immunopotentiating effect of malathion. Following the acute concomitant exposure to dieldrin/carbofuran mixture, however, values for the parameters of antigen presentation, primary IgM antibody response to SRBC antigen, and macrophage phagocytosis, returned to control or above-control values, indicating a lack of any synergistic or additive effects of the chemicals on the immune response. Thus, it was concluded the dieldrin/carbofuran mixture had an antagonistic effect on the humoral response to SRBC and the macrophage phagocytic activity, in comparison with the action of administration of each of the insecticides alone.

Protein quality and microbiological changes in aerobically- or vacuum-packaged, irradiated fresh pork loins

The effect of gamma-irradiation on the physicochemical, organoleptic and microbiological properties of pork was studied, during 43 days of storage at 4±1°C. Irradiation treatments were carried out under air or vacuum packaging on fresh pork loins at a dose of 6 kGy, at two different dose rates: 2 kGy/h and 20 kGy/h. The loins were evaluated for protein sulphydryl content and emulsifying capacity, surface hydrophobicity of proteins and sensorial evaluation. Regardless of the type of packaging and dose rate of irradiation, all irradiated pork samples were effectively prevented from bacterial spoilage for at least 43 days. Meat redness and texture of irradiated loins were relatively well preserved during the storage period, especially when samples were stored under vacuum. Overall, the physicochemical and organoleptic changes in pork loins appeared to be relatively little affected by the 6 kGy dose. No marked changes in emulsifying capacity and protein sulphydryl content of proteins were noted throughout the storage period. However, the hydrophobicity was reduced (P⩽0.05) by the faster dose rate of irradiation and by longer storage.

Prevention of lipid radiolysis by natural antioxidants from rosemary (Rosmarinus officinalis L.) and thyme (Thymus vulgaris L.)

Abstract Radiolysis of unsaturated linoleic fatty acid and subsequent generation of volatile alkane and alkene hydrocarbons was studied following relatively low, 3–9 kilo-Gray (kGy) gamma-irradiation, i.e. within the accepted 10 kGy limit for commercial food pasteurization. The low-dose irradiation generated relatively low quantities of hydrocarbons, totalling 83 ± 1–140 ± 4ng/mg for saturated myristic and stearic fatty acids. As expected, unsaturated fatty acids (UFA) were more susceptible to the gamma-irradiation; at 9 kGy dose, the total amount of C10–C19 volatile hydrocarbons were, respectively, 602 ± 18 ng/mg for arachidonic acid and 751 ± 60 ngmg−1 for linoleic acid. Addition of powdered (unextracted) rosemary or thyme, at 1/0.1 g lipid/plant ratio, decreased by 52.5–80.5% the radiolytic generation of C10–C19 hydrocarbons. In conclusion, rosemary and thyme, both known to contain natural antioxidants, markedly reduced the gamma-radiolysis of linoleic acid, when irradiated within the accepted 10 kGy limit for commercial food processing.

Suppression of humoral immunity in inbred mice by dieldrin.

The effect of single, sublethal i.p. injection of dieldrin on the primary antibody response to thymodependent (sheep red blood cells, SRBC) and T-cell-independent (lipopolysaccharide, LPS) antigens were investigated in inbred C57Bl/6 mice. Time-course studies showed significant suppression of the anti-SRBC IgM and anti-LPS IgM response at 7-24 days and at 4-14 days, respectively, after exposure to 0.6 LD50 of dieldrin. The anti-SRBC IgG response was also suppressed by dieldrin exposure, however, maximal suppressory effect was found at 48 days after the pesticide exposure. Similar patterns of the dieldrin-induced suppression of the primary IgM response to the thymodependent and T-cell-independent antigens, in addition to the overall control of cytotoxicity of lymphoid cell populations, suggest rather dysfunction of cellular cooperation during the inductory phase of the immune humoral response.

Evaluation of the immunomodulatory potential of diethyl dithiocarbamate derivatives

The cytotoxicity, immunotoxicity and immunomodulatory potential of four dithiocarbamate derivatives were assessed and compared with the effects of Immuthiol (diethyl dithiocarbamate, DE-DTC) in mice. Cellular stimulation and cell viability were examined after in vitro exposure of spleen lymphocytes to selected DTC analogues: N-methyl-D-glucamine dithiocarbamate (NMG-DTC), dimethyl dithiocarbamate (DM-DTC), dibuthyl dithiocarbamate (DB-DTC) and diisobuthyl dithiocarbamate (DIB-DTC). Lymphocyte activation by plant and bacterial mitogens: concanavalin A (Con A), phytohaemagglutinin (PHA), lipopolysaccharide (LPS) and allogeneic stimulation of cells in mixed lymphocyte reaction (MLR) were examined in vitro in the presence of 10(-4)-10(-9) g/ml DE-DTC and other selected DTC derivatives. No direct in vitro lymphoproliferative activity of DTC derivatives was observed, although a relatively stronger cytotoxicity with DE-DTC and DM-DTC was noted. In addition, the in vivo effects of DTC derivatives were examined by cytofluorometric profile of splenic and bone marrow cells as well as in mitogenic and allogenic responses, after i.v. exposures of animals to two subsequent (25 mg/kg b.w.) doses of the chemical. Less cytotoxic DIB-DTC, NMG-DTC and DB-DTC expressed weak in vivo immunostimulatory potential when compared with the effect of DE-DTC, whereas the effects of DM-DTC on alloantigenic and mitogenic lymphocyte stimulation were comparable with the known effects of DE-DTC. Cytofluorometric studies showed that the number of cytotoxic/suppressor T-cells (Ts) and helper T-cells (Th) in the cell was increased by DE-DTC and NMG-DTC only. In addition, DM-DTC appeared to affect the Ts/Th ratio. DE-DTC did not affect the B-cell subpopulation, whereas other derivatives induced marked modifications of the pre-B-cell subpopulations in bone marrow. Our data suggest that in vivo the immunostimulatory effect of DM-DTC could be accompanied with major changes in bone marrow B-cell frequency and alteration of spleen Ts/Th ratio.

Suppression of avidin processing and presentation by mouse macrophages after sublethal exposure to dieldrin

The molecular events in macrophage antigen processing and presentation were examined to determine the possible site(s) of cell-xenobiotic interaction. Antigenic processing by mouse peritoneal macrophages of a single protein antigen, avidin, was significantly suppressed following sublethal exposure of animals to an organochlorine pesticide, dieldrin. Exposure of C57B1/6 female mice to dieldrin affected the in vitro uptake of [methyl-14C]avidin by peritoneal macrophages and markedly decreased phagocytosis of fluorescein-labelled microspheres and Salmonella typhimurium. Release of the processed avidin, determined by immunochemical quantification of immunogenic avidin and by bioassay of immunogenicity of the released antigen, was also markedly affected. Dieldrin markedly affected presentation of avidin on the macrophage surface, observed by cytoimmunochemical staining of the antigen with fluorescent antibody and flow cytometry. Inhibition of the release of processed avidin was dieldrin dose- and time-dependent, following single sublethal intraperitoneal (ip) exposure to the pesticide. The antigenic properties of processed avidin, determined by biological assay using lymphocyte cultures of normal C57B1/6 mice primed with avidin, were proportional to the antigen concentration in supernatants of macrophage cultures, for both vehicle controls and dieldrin-exposed animals. This observation and analysis of the kinetics of release of processed avidin by macrophages from control and dieldrin-exposed animals suggested that the release of processed avidin, but not the immunogenicity of the antigen itself, was affected by the pesticide exposure. Generally, impairment of avidin processing and presentation appeared to be more dramatic than other pesticide-related injuries to macrophages, such as the uptake of the antigen. In conclusion, antigen processing could be a sensitive target for dieldrin-related injury of macrophage functional activities, which, in consequence, could produce suppression of the humoral immune response.

Immunotoxicity of aminocarb. III. Exposure route-dependent immunomodulation by aminocarb in mice

Aminocarb, a phenylsubstituted methylcarbamate pesticide (4-dimethylamino-3-methyl-N-carbamate; matacil), previously suspected of a relatively low immunotoxic potential, was administered by four different exposure routes to C57BL/6 mice. A single sublethal exposure by oral and dermal routes stimulated humoral immune response at a relatively low dose; 1/256 LD50 of aminocarb. Intraperitoneal (i.p.) injection decreased the humoral PFC response, whereas inhalation of aminocarb had no marked effect on peripheral immune status in exposed animals. Thus, i.p. exposure resulted in higher immunotoxicity over oral administration of aminocarb. Similarly, marked route-related exposure differences in immunomodulatory effects of aminocarb were noted for mitogenic stimulation of spleen lymphocytes and mixed lymphocyte response. Other indices, such as delayed type hypersensitivity (DTH) and production of interleukin-2 (IL-2) were unchanged. Interestingly, expression of major histocompatibility complex (MHC) class II by purified, lipopolysaccharide (LPS)-stimulated B cells increased equally after i.p. and oral exposures to aminocarb. Overall, a weak immunosuppressive potential of aminocarb was concluded, which was possibly due to indirect interaction of the pesticide with the immune system. However, aminocarb may represent an autoimmunity-inducing toxic.