In this study, the purpose was to investigate the histopathological, genotoxic effect, oxidative stress and cell death due to Metronidazole (MTZ), which is a 5-nitroimidazole compound used widely for the treatment of anaerobic organism infections in fish and humans on gill and liver tissues of Oncorhynchus mykiss.Trout fishes were exposed to 5, 10, and 20 mg/L of MTZ in the aquariums for 2, 4 and 8 days. Staining technics namely H&E, NOS immunohistochemistry, and TUNEL were performed to determine histopathological changes, oxidative damage and apoptosis. Additionally, smear preparations were also prepared from gill blood for genotoxic evaluations. The organ damage started in the 2nd day with 5 mg/L MTZ application and effects increased per duration and dose-dependent manner. It was observed that the gills had the primary and secondary lamellae lengths, with formation of clavate lamellae, fusion in secondary lamellae, separation of epithelium and aneurysm. Regional necrosis, vacuolization of hepatocytes, pycnotic nucleus, enlarged sinusoids were also determined in the liver. NOS immunoreactivity increased with the inducible immunoreactivity (iNOS) that was more prominent when compared to the endothelial immunoreactivity (eNOS). Apoptotic immunoreactivity was higher in the 10 mg 8th day experimental group at liver and gills, and was lower 20 mg 8th day experimental group. When the gills and liver compared with each other, in all doses, immunoreactivity was lower in gills, compared with liver. Genotoxic examinations showed that both number of micronucleated erythrocytes and nuclei abnormalities were higher in MTZ-treated groups.

Nickel nitrate is a heavy metal known as an environmental contaminant due to its toxicity, long environmental half-lives, and capacity for bioaccumulation.  This study aims to determine chromosomal aberration, nuclear alteration, and cell death in Allium cepa var. aggregatum L. root caused by different nickel concentrations.  Roots of Allium cepa var. aggregatum were induced by soaking bulbs in water, then transferred to a solution containing nickel (Ni) at a concentration of 20 ppm, 30 ppm, and 40 ppm for 72 hours.  Root tip mitotic chromosome preparations were done by the squash method.  The chromosome was stained with aceto-orcein and chromosomal damages were observed under a microscope.  The results showed that the mitotic index decreased from 5.025% at control to 3.144%, 2.467%, and 2.181% at immersion with 20 ppm, 30 ppm 40 ppm nickel nitrate, respectively.  Anaphase and telophase indexes in roots with Ni treatments were lower than in control, suggesting that nickel inhibits cell division.  Nickel nitrate induced chromosomal damages and nuclear abnormalities, such as sticky chromosome, fragmented chromosome, chromosome bridge and chromosome laggard, micronuclei, binucleate and nuclear budding.  The percentage of chromosomal damage increases with a higher concentration of Ni.  In situ cell visualization showed that the higher the nickel concentration, the more coloured the root tips indicating high levels of cell death.

The mosquito Aedes aegypti is a primary vector for major arboviruses, and its control is mainly based on the use of insecticides. Caffeine and spent coffee grounds (CG) are potential agents in controlling Ae. aegypti by reducing survival and blocking larval development. In this study, we analyzed the effects of treatment with common CG (CCG: with caffeine), decaffeinated CG (DCG: with low caffeine), and pure caffeine on the survival, behavior, and morphology of the midgut of Ae. aegypti under laboratory conditions. Third instar larvae (L3) were exposed to different concentrations of CCG, DCG, and caffeine. All compounds significantly affected larval survival, and sublethal concentrations reduced larval locomotor activity, delayed development, and reduced adult life span. Damage to the midgut of treated larvae included changes in epithelial morphology, increased number of peroxidase-positive cells (more abundant in DCG-treated larvae), and caspase 3-positive cells (more abundant in CCG-treated larvae), suggesting that the treatments triggered cell damage, leading to activation of cell death. In addition, the treatments reduced the FMRFamide-positive enteroendocrine cells and dividing cells compared to the control. CG and caffeine have larvicidal effects on Ae. aegypti that warrant field testing for their potential to control mosquitoes.

Cigarette smoke (CS) affects immune functions, leading to severe outcomes in smokers. Robust evidence addresses the immunotoxic effects of combustible tobacco products. As heat-not-burn tobacco products (HNBT) vaporize lower levels of combustible products, we here compared the effects of cigarette smoke (CS) and HNBT vapor on Jurkat T cells. Cells were exposed to air, conventional cigarettes or heatsticks of HNBT for 30 min and were stimulated or not with phorbol myristate acetate (PMA). Cell viability, proliferation, reactive oxygen species (ROS) production, 8-OHdG, MAP-kinases and nuclear factor κB (NFκB) activation and metallothionein expression (MTs) were assessed by flow cytometry; nitric oxide (NO) and cytokine levels were measured by Griess reaction and ELISA, respectively. Levels of metals in the exposure chambers were quantified by inductively coupled plasma mass spectrometry. MT expressions were quantified by immunohistochemistry in the lungs and liver of C57Bl/6 mice exposed to CS, HNBT or air (1 h, twice a day for five days: via inhalation). While both CS and HBNT exposures increased cell death, CS led to a higher number of necrotic cells, increased the production of ROS, NO, inflammatory cytokines and MTs when compared to HNBT-exposed cells, and led to a higher expression of MTs in mice. CS released higher amounts of metals. CS and HNBT exposures decreased PMA-induced interleukin-2 (IL-2) secretion and impaired Jurkat proliferation, effects also seen in cells exposed to nicotine. Although HNBT vapor does not activate T cells as CS does, exposure to both HNBT and CS suppressed proliferation and IL-2 release, a pivotal cytokine involved with T cell proliferation and tolerance, and this effect may be related to nicotine content in both products.

Recently, chemical compounds containing lanthanides were used in various fields of biology and medicine. It has been described that such compounds can be applied in scanning electron microscopy (SEM) to increase the contrast and simplify the sample preparation process due to the process of replacing calcium with lanthanides in cell. However cell death by different mechanisms under influence of lanthanides seems possible. Here, we described that mummification process is a cell death physiologically realized in time: some time after lanthanide contrasting, the cell remains metabolically active and is able to biochemically transform neodymium-containing contrast, oxidize it and form large agglomerates. A distinctive feature of mummification induced by neodymium-containing contrast (NCC) is the formation of a high-rigid oxygen-containing “shield” on the surface of a neutrophil granulocyte.

Sulfitobacter porphyrae ZFX1, isolated from surface seawater of the East China Sea during a Prorocentrum donghaiense bloom recession, exhibits high algicidal activity against P. donghaiense. To evaluate the algicidal effect of ZFX1, the algicidal mode and stability were investigated. The results showed that ZFX1 indirectly attacked algae by secreting algicidal compounds, and the algicidal activity of the ZFX1 supernatant was insensitive to different temperatures, light intensities and pH values (pH 3–12). To explore the algicidal mechanism of the ZFX1 supernatant, its effects on the morphological and ultrastructural alterations, photosynthetic capacity, reactive oxygen species (ROS) and antioxidative system of P. donghaiense were investigated. Scanning and transmission electron microscopy revealed that the ZFX1 supernatant destroyed the algal cell membrane structure and caused intracellular leakage. The decrease in the chlorophyll a content and the marked declines in both the photosynthetic efficiency (Fv/Fm) and the electron transport rate (rETR) indicated that the ZFX1 supernatant could damage the photosynthetic system of P. donghaiense. The excessive production of ROS in algal cells demonstrated the oxidative damage triggered by the ZFX1 supernatant. Although the antioxidant defense system of P. donghaiense was activated to scavenge excessive ROS, lipid oxidation occurred. The fatty acid composition profile indicated that the ZFX1 supernatant markedly increased the contents of two saturated fatty acids and a monounsaturated fatty acid and decreased the proportion of two polyunsaturated fatty acids, which resulted in lipids with a lower degree of unsaturation (DU). The decline in the DU decreased the lipid fluidity and rigidified the membrane system, and these effects destroyed the function of the membrane system and ultimately resulted in algal cell death. Therefore, ZFX1 probably plays a key role in mitigating P. donghaiense bloom by inducing lipid oxidation, decreasing the DU of lipids and ultimately destroying the membrane systems of algal cells.

PM2.5 is becoming a worldwide environmental problem, which profoundly endangers public health, thus progressively capturing public attention this decade. As a fragile target of PM2.5, the underlying mechanisms of endothelial cell damage are still obscure. According to the previous microarray data and signaling pathway analysis, a new form of cell death termed ferroptosis in the current study is proposed following PM2.5 exposure. In order to verify the vital role of ferroptosis in PM2.5-induced endothelial lesion and further understand the potential mechanism involved, intracellular iron content, ROS release and lipid peroxidation, as well as biomarkers of ferroptosis were detected, respectively. As a result, uptake of particles increases cellular iron content and ROS production. Meanwhile, GSH depletion, and the decrease of GSH-Px and NADPH play significant roles in PM2.5-induced endothelial cell ferroptosis. Moreover, significantly changed expression of TFRC, FTL and FTH1 hinted that dysfunction of iron uptake and storage is a major inducer of ferroptosis. Importantly, index monitored above can be partially rescued by lipid peroxidation inhibitor ferrostatin-1 and iron chelator deferoxamine mesylate, which mediated antiferroptosis activity mainly depends on the restoration of antioxidant activity and iron metabolism. In conclusion, our data basically show that PM2.5 enhances ferroptosis sensitivity with increased ferroptotic events in endothelial cells, in which iron overload, lipid peroxidation and redox imbalance act pivotal roles.

Ultra-fine-ZnO showed low toxicity in complex water matrix containing multiple components such as PBS buffer and the toxic mechanism of ultra-fine-ZnO has not been clearly elucidated. In present study, enhanced antibacterial activity of 200 nm diameter ultra-fine-ZnO in PBS buffer against Bacillus cereus and Escherichia coli were observed in the presence of several organic acids in comparison with ultra-fine-ZnO in PBS buffer alone. These findings indicated that the toxic effects of the ultra-fine-ZnO was dependent on the concentration of released Zn2+ which was affected by organic acids. The production of reactive oxygen species (ROS) did not responsible to the toxic mechanism of ultra-fine-ZnO which was tested using the antioxidant N-Acetylcysteine (NAC). Indeed, ultra-fine-ZnO induced bacteria cell membrane leakages and cell morphology damages that eventually led to cell death, which were confirmed using propidium monoazide (PMA) in combination with PCR and scanning electron microscopy (SEM). All data gathered herein suggested that released Zn2+ played a major role in the microbial toxicity of ultra-fine-ZnO.

We aimed to verify whether hydrogen peroxide (H₂O₂) accumulation and cell death are detected early in three bioindicators of ozone (O₃), Nicotiana tabacum ‘Bel-W3’, Ipomoea nil ‘Scarlet O’Hara’ and Psidium guajava ‘Paluma’, and whether environmental factors also affect those microscopic markers. The three species were exposed to chronic levels of O₃ in a subtropical area and a histo-cytochemical technique that combines 3,3′-diaminobenzidine (DAB) with Evans blue staining was used in the assessments. The three species accumulated H₂O₂, but a positive correlation with O₃ concentration was only observed in N. tabacum. A positive correlation between O₃ and cellular death was also observed in N. tabacum. In I. nil and P. guajava, environmental factors were responsible for symptoms at the microscopic level, especially in P. guajava. We conclude that the most appropriate and least appropriate bioindicator plant for O₃ monitoring in the subtropics are N. tabacum ‘Bel-W3’ and P. guajava ‘Paluma’, respectively.

An account of histo-cytological and ultrastructural studies on ozone effect on crop and forest species in Italy is given, with emphasis on induced cell death and the underlying mechanisms. Cell death phenomena possibly due to ambient O3 were recorded in crop and forest species. In contrast, visible O3 effects on Mediterranean vegetation are often unclear. Microscopy is thus suggested as an effective tool to validate and evaluate O3 injury to Mediterranean vegetation. A DAB-Evans blue staining was proposed to validate O3 symptoms at the microscopic level and for a pre-visual diagnosis of O3 injury. The method has been positively tested in some of the most important crop species, such as wheat, tomato, bean and onion and, with some restriction, in forest species, and it also allows one to gain some very useful insights into the mechanisms at the base of O3 sensitivity or tolerance. Ozone-induced cell death is a frequent phenomenon in Mediterranean conditions, not only in the most sensitive crops but also in forest species.