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Object: Nowadays, organophosphates appear to be among the most used pesticides all over the world. Their amounts and amounts of their metabolites in surface water are expected to increase in the near future. In the light of potential organelles impairment, this work aims to investigate the effects of chlorpyrifos and malathion in environmentally relevant concentration on morphological and functional traits of zebrafish mitochondria and lysosomes. Methods: Adult zebrafish were exposed to widely used organophosphate pesticides namely malathion (5 and 50 µg L−1) and chlorpyrifos (0.1 and 3 µg L−1) for 14 days. The comprehensive set of biochemical markers that reflect oxidative/carbonyl/nitrosative stress and cytotoxicity (mitochondria swelling, lysosomal membrane stability, and cell damage) was determined in fish liver. Results: Chlorpyrifos and malathion in both concentrations were found to affect mitochondria and lysosome membranes: They caused a substantial decline in neutral red retention and an increase in mitochondria swelling. Also, the concentration-dependent enhancement of lactate dehydrogenase in blood was depicted. Even low pesticide concentrations were able to promote apoptosis and stimulated overproduction of methylglyoxal as reactive carbonyl species. We also revealed that reactive nitrogen species were sparked by organophosphates. This effect occurs only in animals treated with higher insecticides concentration. Moreover, the pesticides induced oxidative stress response measured by increasing of reactive oxygen species level (only after chlorpyrifos), as well as by variation in total antioxidant capacity with signs of deeper injury after chlorpyrifos exposure. The integrative biomarker response (IBR) index calculation supports the hypothesis that chlorpyrifos was significantly more toxic to zebrafish than malathion. Conclusion: Our study suggests that zebrafish markers can serve as a sensitive transferential model for evaluating mitochondrial and lysosomal toxicity of pesticides and extrapolating results to mammalian and human.

Flow cytometry (FCM) is routinely used in medical and veterinary diagnostics although it is also widely applied in environmental studies, including phytoplankton investigations. Cyanobacteria are wide-spread photosynthetic microorganisms that attract attention due to their ecology and potential toxicity. Therefore, novel research tools are being applied in their investigation. This paper characterizes FCM as a technique that enables photopigments (chlorophylls and phycocyanin) expressed by cyanobacteria to be excited and their emission to be subsequently detected. This feature not only allows cells to be counted in a rapid manner but also enables a wide range of potential applications in ecological and biochemical studies. The main advantages of FCM, such as rapid, automatic and precise measurements requiring small sample volumes, are also discussed in this paper along with challenges including analyses of filamentous cyanobacteria and signal overlapping. It is expected that FCM will continue to be used in some fields of cyanobacterial studies.

ABSTRACTHepatocellular carcinoma (HCC) is the most common primary malignancy of the liver with a very poor prognosis and constantly growing incidence. Among other primary risks of HCC, metabolic disorders and obesity have been extensively investigated over recent decades. The latter can promote nonalcoholic fatty liver disease (NAFLD) leading to the inflammatory form of nonalcoholic steatohepatitis (NASH), that, in turn, promotes HCC. Molecular determinants of this pathogenic progression, however, remain largely undefined. In this study, we have focussed on the investigation of α-dicarbonyl compounds (α-dC), highly reactive and tightly associated with overweight-induced metabolic disorders, and studied their potential role in NAFLD and progression toward HCC using murine models. NAFLD was induced using high-fat diet (HFD). Autochthonous HCC was induced using transposon-based stable intrahepatic overexpression of oncogenic NRASG12Vin mice lacking p19Arftumor suppressor. Our study demonstrates that the HFD regimen and HCC resulted in strong upregulation of α-dC in the liver, heart, and muscles. In addition, an increase in α-dC was confirmed in sera of NAFLD and NASH patients. Furthermore, higher expression of the receptor for advanced glycation products (RAGE) was detected exclusively on immune cells and not on stroma cells in livers of mice with liver cancer progression. Our work confirms astable interplay of liver inflammation, carbonyl stress mediated by α-dC, and upregulated RAGE expression on CD8+Tand natural killer (NK) cells in situin NAFLD and HCC, as key factors/determinants in liver disease progression. The obtained findings underline the role of α-dC and RAGE+CD8+Tand RAGE+NK cells as biomarkers and candidates for a local therapeutic intervention in NAFLD and malignant liver disease.

Hypoxia is a major stressor in estuarine and coastal habitats, leading to adverse effects in aquatic organisms. Estuarine bivalves such as blue mussels (Mytilus edulis) and Pacific oysters (Crassostrea gigas) can survive periodic oxygen deficiency but the molecular mechanisms that underlie cellular injury during hypoxia-reoxygenation are not well understood. We examined the molecular markers of autophagy, apoptosis and inflammation during short-term (1 day) and long-term (6 days) hypoxia and post-hypoxic recovery (1 h) in mussels and oysters by measuring the lysosomal membrane stability, activity of a key autophagic enzyme (cathepsin D) and mRNA expression of the genes involved in the cellular survival and inflammation, including caspase 2, 3 and 8, Bcl-2, BAX, TGF-β-activated kinase 1 (TAK1), nuclear factor kappa B1 (NF-κB) and NF-κB activating kinases IKKα and TBK1. Crassostrea gigasexhibited higher hypoxia tolerance, as well as blunted or delayed inflammatory and apoptotic response to hypoxia and reoxygenation as shown by the later onset and/or the lack of transcriptional activation of caspases, BAX and the inflammatory effector NF-κB, compared with M. edulis. Long-term hypoxia resulted in upregulation of Bcl-2 in the oysters and mussels, implying activation of anti-apoptotic mechanisms. Our findings indicate the potential importance of the cell survival pathways in hypoxia tolerance of marine bivalves, and demonstrate the utility of the molecular markers of apoptosis and autophagy for the assessment of sublethal hypoxic stress in bivalve populations.