Oral exposure to the herbicide simazine induces mouse spleen immunotoxicity and immune cell apoptosis.

The authors investigated the toxic effects of simazine on mice spleen immune cells and the underlying mechanisms. Mice were given simazine at 0, 90, 200, or 400 mg/kg by gastric gavage for 3 weeks. The authors then measured immune cell proliferation and the expressions of apoptosis-related proteins (Bcl-2, Bax, Fas, and caspase-3), spleen cell intracellular [Ca(2+)], cellular oxidative stress level, and immune functions. After 3 weeks, mice exposed to simazine had reduced proliferation of both spleen T and B cells. The number of spleen CD4(+) T lymphocytes decreased with simazine exposure, while CD8(+) T cells remained unchanged. Exposure to simazine resulted in reduced immune function, higher intracellular [Ca(2+)], and oxidative stress. Finally, simazine induced spleen immune cells apoptosis by reducing Bcl-2, while increasing Fas and Caspase-3 level. Overall, the immunotoxicity of simazine may involve the induction of immune cell apoptosis and alterations in the immune and physiological functions of spleen cells.

Effects of subchronic simazine exposure on some biometric, biochemical, hematological and histopathological parameters of common carp (Cyprinus carpio L.).

OBJECTIVES: This study assessed the subchronic effects of a triazine compound, simazine, on common carp (Cyprinus carpio L.) though, via by means of biometric, biochemical, hematological, and histological examination. DESIGN: One-year-old fish were exposed to simazine at four concentrations, 0.06, (reported concentration in Czech rivers), 4, 20, and 50 microg L-1 for 28 days and compared to carp in a non-treated control group. RESULTS: Exposure of fish to simazine at 0.06 microg L-1 had no effect on measured parameters. However, exposure to simazine at the concentrations of 4, 20, 50 microg L-1 showed significant differences in biochemical, hematological, and histopathological profiles of fish compared to controls. CONCLUSION: Our data suggest that simazine in the recorded environmental concentration 0.06 microg L-1 had no effect on common carp. Subchronic exposure to 4, 20, and 50 microg L-1 of simazine was associated with alterations in biochemical and hematological indices and in fish organ tissues.

Effect of a subchronic exposure to simazine on energetic metabolism of common carp (Cyprinus carpio).

The energetic parameters, such as glycogen, glucose, proteins, lactate and adenosine triphosphate (ATP) content and lactate deshydrogenase (LDH) activity in tissues and blood of carps from simazine (2-chloro-4,6-bis(ethylamino)-s-triazine) contaminated waters were investigated. In order to confirm the field results, a laboratory experiment was designed in which carps were exposed to simazine at the concentration level of 45 micro g. L(- 1) (10-fold of the amount found in natural waters) for 90 days. Fish from a contaminated reservoir showed low glycogen concentrations in hepatopancreas and muscle, while fish in another contaminated reservoir showed high LDH activity together with an increase in lactate content in muscle. Laboratory findings did not confirm field results, and fish exposed to simazine did not show alterations in the parameters studied. The results suggest that carps were not stressed by the presence of the simazine at the concentration levels found in both studies and the mechanisms of defense covered the energetic demand.

Novel s-triazine-degrading bacteria isolated from agricultural soils of central Chile for herbicide bioremediation

s-Triazine-degrading bacterial strains were isolated from long-term simazine-treated agricultural soils of central Chile. The number of culturable heterotrophic bacteria of these agricultural soils (7 x 10(6) CFU/g of dry soil) was not affected by simazine application on field. The simazine-degrading bacterial strains P51, P52 and C53 were isolated by enrichment in minimal medium using simazine as the sole nitrogen source. Resting cells of strains P51 and P52 degraded >80 percent of simazine within 48 hrs, whereas strain C53 was able to remove >60 percent of the herbicide. The atzA and atzD genes of the s-triazine upper and lower catabolic pathways were detected in strains P51 and C53, while only atzD gene was observed in strain P52. To compare the bacterial 16S rRNA gene sequence structure, ARDRA were performed using the restriction enzymes Msp1 and Hha1. ARDRA indicated that strain P52 was a different ribotype than C53 and P51 strains. For further characterization the novel isolates were identified by 16S rRNA gene sequencing. Strains C53 and P51 belong to the genus Stenotrophomonas and the strain P52 belongs to the genus Arthrobacter . s -Triazine-degrading bacterial strains isolated from contaminated soils could be used as biocatalysts for bioremediation of these herbicides.

Immune alterations in mice exposed to the herbicide simazine.

Simazine, a triazine herbicide, was investigated for its in vivo immunomodulatory properties. Male C57Bl/6 mice were treated with vehicle or 300 or 600 mg/kg body weight (bw) simazine daily orally for 4 wk. The immune system was evaluated by the antibody response to sheep red blood cells (SRBC; plaque assay and serum immunoglobulin G), natural killer (NK) and macro-phage activities, lymphocyte subpopulations in the spleen and thymus, and concanavalin A (Con A)- and lipopolysaccharide (LPS)-stimulated lymphocyte proliferation using splenocytes. Body weight and spleen and thymus weight decreased generally in simazine-treated mice, while the weight of adrenal glands was higher than in the control. Simazine treatment (600 mg/kg) induced an increase in the percentage of CD4(+) cells in spleen and CD8 + in thymus. Simazine inhibited the IgM plaque-forming cell numbers and lowered the level of IgG and the proliferation of mitogen-stimulated B cells and T cells. In addition, splenic NK and peritoneal macrophage activities in exposed mice were significantly decreased. Exposure to simazine also decreased cytokine production by macrophages, such as interleukin-1 (IL-1), IL-6, and tumor necrosis factor-alpha (TNF-alpha). Taken together, data indicate that the immune system was suppressed by oral simazine exposure.

Sister chromatid exchanges and micronuclei analysis in lymphocytes of men exposed to simazine through drinking water.

In some cities of the autonomous community of Extremadura (south-west of Spain), levels of simazine from 10 to 30 ppm were detected in tap water. To analyse the possible effect of this herbicide, two biomarkers, sister chromatid exchanges (SCE) and micronuclei (MN), were used in peripheral blood lymphocytes from males exposed to simazine through drinking water. SCE and MN analysis failed to detect any statistically significant increase in the people exposed to simazine when compared with the controls. With respect to high frequency cells (HFC), a statistically significant difference was detected between exposed and control groups.

Genotoxicity studies of three triazine herbicides: in vivo studies using the alkaline single cell gel (SCG) assay.

Triazine herbicides are prevalent contaminants of groundwater in the agricultural regions of the United States. The literature on the genotoxicity of triazines is rife with conflicting data, though the general tendency is for most studies to report negative results. In order to investigate further the genotoxicity of triazines, we exposed mice to triazines by intraperitoneal injection up to the maximum tolerated doses. About 24h later, blood was removed, and the leukocytes subjected to DNA damage analysis using the alkaline single cell gel electrophoresis assay (SCG), one of the most sensitive DNA damage assays available. Our results indicate that atrazine induced a small dose-related increase in DNA damage. Simazine did not induce any dose-related increase in DNA damage. Cyanazine induced a marginal increase in DNA damage with dose, but no individual dose was significantly increased compared to the control. These results indicate that these triazines, even at extremely high concentrations, have only marginal DNA-damaging activity in vivo in mouse leukocytes.

Cytogenetic studies of three triazine herbicides. II. In vivo micronucleus studies in mouse bone marrow.

Atrazine, simazine, and cyanazine are widely used preemergence and postemergence triazine herbicides that have made their way into the potable water supply of many agricultural communities. Although there are several contradictory genotoxicity studies in the literature, our previous in vitro studies with human lymphocytes showed that atrazine, simazine, and cyanazine did not induce sister chromatid exchanges (SCEs) or chromosome aberrations (CAs) up to the limits of solubility in aqueous medium using 0.5% dimethyl sulfoxide. To expand upon these results and to ensure that our in vitro findings could be replicated in an in vivo system, mice were treated with each triazine by two intraperitoneal injections, 24h apart. The animals were sacrificed and the bone marrow removed for micronucleus (MN) analysis, 24h after the last injection. Two to four independent trials were performed for MN analysis in polychromatic erythrocytes, and in some trials the spleen was removed, cultured, and analyzed for SCEs and CAs. None of the triazines investigated induced MN in the bone marrow, even at doses that caused significant bone marrow suppression and/or death. These results indicate that atrazine, simazine, and cyanazine are not genotoxic as measured by the bone marrow MN assay in mice following high dose exposures.

Short-term effects of chlorotriazines on estrus in female Sprague-Dawley and Fischer 344 rats.

Atrazine or simazine (s-chlorotriazines) was administered by gavage daily for 2 wk to female Sprague-Dawley and Fischer 344 rats at oral doses of 100 or 300 mg/kg to evaluate effects on body, ovary, uterus, and adrenal weights, estrous cycle stages, vaginal cytology, and plasma hormone (estradiol, progesterone, prolactin, and corticosterone) levels. Significant reductions in body weights of both Sprague-Dawley and Fischer 344 female rats at both dose levels were accompanied by a significant reduction in ovarian and uterine weights, and a decrease in circulating estradiol levels. The magnitudes of the effects were less in Fischer 344 rats than in Sprague-Dawley rats, and the effects of simazine were less pronounced than those of atrazine at the same dose. A maximum tolerated dose (MTD: > or = 10% body weight reduction) was estimated to be 100 mg/kg for atrazine and 300 mg/kg for simazine for both stains. The Sprague-Dawley female rats exhibited a treatment-related lengthening of the estrous cycle and an increased number of days characterized by cornified epithelial cells. This resulted in a greater percent of the cycle days spent in estrus and reduction in the percent of the cycle days spent in diestrus. Atrazine-dosed Fischer 344 females also exhibited a significant trend toward cycle lengthening, but this was due to reduction in the percent of cycle spent in estrus and a concomitant increase in diestrual days. These findings suggest that treatment with doses of triazine at or above the MTD may result in prolonged exposure to endogenous estrogen in the Sprague-Dawley but not the Fischer 344 rat. These changes may account for the observed earlier onset and/or increased incidence of mammary tumors in chlorotriazine-treated female Sprague-Dawley rats. This strain of rat is already known to be prone to a substantial development of mammary tumors with advancing age, while the Fischer 344 strain is not as likely to exhibit this response.

Possible antiestrogenic properties of chloro-s-triazines in rat uterus.

Several published reports have indicated that certain chloro-s-triazine herbicides may alter endocrine function in rats, possibly by androgen receptor binding. In direct tests of estrogenic bioactivity, oral doses of up to 300 mg/kg/d of atrazine, simazine, or the common metabolite diaminochlorotriazine (DACT) did not significantly increase uterine weight of ovariectomized Sprague-Dawley female rats. The highest dose, which was approximately 10% of the LD50 for these compounds, did cause body weight loss. When administered concomitantly with sc injections of estradiol (2 micrograms/kg), 300 mg/kg of orally administered chlorotriazines significantly reduced uterine weight in comparison to animals given estrogen alone. Neither atrazine, simazine, nor DACT, at oral doses up to 300 mg/kg/d, stimulated incorporation of [3H]thymidine into uterine DNA of immature Sprague-Dawley female rats. However, oral treatment at doses of 50 mg/kg and higher significantly reduced thymidine incorporation into uterine DNA extracted from immature rats given a single injection of 0.15 microgram estradiol. Oral doses of 300 mg/kg of atrazine, simazine, or DACT significantly reduced expression of progesterone receptor binding in cytosol fractions prepared from uteri of ovariectomized rats injected sc with 1 microgram estradiol; 50 mg/kg triazine was not effective in this case. Uterine progesterone receptor levels were not stimulated in rats given oral doses up to 300 mg/kg of these triazines without estradiol injections. These results suggest that atrazine, simazine, and DACT possess no intrinsic estrogenic activity but that they are capable of weak inhibition of estrogen-stimulated responses in the rat uterus. This inhibition may play a role in the previously observed disruptive actions of chlorotriazines on reproductive endocrine function of female rats.

[Immunomorphological analysis of prolonged intoxication by low doses of herbicide simazine]. / Immunomorfologicheskii analiz dlitel'noi intoksikatsii malymi dozami gerbitsida simazina.

Dynamic investigation of several immunologic data and complex morphologic study of the thymus of rats fed for a long time (6 months) by very low doses of herbicide simazine have been carried out. Chronic simazine treatment resulted in the development of the secondary immunodeficient state with the damage of T lymphocytes. The morphologic signs of this process were the disarray in thymus structure (dystrophic changes and intercellular contact break of nurse cells, sclerosis of microvessel walls and stromal elements), severe decrease of the T lymphocyte number in peripheral blood, inhibition of phagocytosis reaction of neutrophils.

[The immunomorphological characteristics of a focal staphylococcal infection against a background of long-term exposure to low doses of simazine]. / Immunomorfologicheskaia kharakteristika ochagovoi stafilokokkovoi infektsii na fone dlitel'nogo vozdeistviia malykh doz simazina.

State of immunologic and nonspecific resistance of the organism, ultra- and histostructure of the thymus, histopathology of the wall of experimental staph abscess reproduced in animals given low doses of the herbicide simazine for a long time have been studied. It is established that simazine induced the immunodeficiency state underlain by pathologic changes in the thymus. Against this background experimental abscesses developed more rapidly, alterative and exudative processes in their wall proceeding more intensively and proliferative ones--attenuating. This provides prolongation of the abscesses healing phase for an indefinite time and chronization of the process.