Cyanobacterial harmful blooms (CyanoHABs) pose a global ecological problem, and their lipopolysaccharides (LPS) are among the bioactive compounds they release. Previous studies on CyanoHAB-LPS from single cyanobacterial species have shown varying bioactivities in different in vitro cell models. In this study, we isolated LPS from 19 CyanoHAB samples collected at 18 water bodies in the Czech Republic over two consecutive seasons. The proportions of cyanobacteria, Gram-negative bacteria (G-), and other bacteria in the biomass were determined by qPCR, while the cyanobacterial genera were identified using light microscopy. In vitro models of keratinocytes (HaCaT), the intestinal epithelium (co-culture of differentiated Caco-2 cells and peripheral blood mononuclear cells - PBMC), and PBMC alone were treated with isolated LPS at concentrations of 50, 100, and 1 µg/ml, respectively. The endotoxin activities of these concentrations were within the range measured in the aquatic environment. Approximately 85-90% of the samples displayed biological activity. However, the potency of individual LPS effects and response patterns varied across the different in vitro models. Furthermore, the observed activities did not exhibit a clear correlation with the taxonomic composition of the phytoplankton community, the relative share of microbial groups in the biomass, endotoxin activity of the LPS, or LPS migration and staining pattern in SDS-PAGE. These findings suggest that the effects of CyanoHAB-LPS depend on the specific composition and abundance of various LPS structures within the complex environmental sample and their interactions with cellular receptors.

• Keywords
• Cyanobacterial harmful blooms, Enterocytes, Immune cells, Inflammation, Keratinocytes, Lipopolysaccharide,
• MeSH
• Biomass MeSH
• Caco-2 Cells MeSH
• Leukocytes, Mononuclear MeSH
• Humans MeSH
• Lipopolysaccharides * toxicity   MeSH
• Cyanobacteria * MeSH
• Harmful Algal Bloom MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Lipopolysaccharides * MeSH

Freshwater cyanobacterial harmful blooms (CyanoHABs) produce a variety of toxic and bioactive compounds including lipopolysaccharides (LPSs). The gastrointestinal tract can be exposed to them via contaminated water even during recreational activities. However, there is no evidence of an effect of CyanoHAB LPSs on intestinal cells. We isolated LPSs of four CyanoHABs dominated by different cyanobacterial species and LPSs of four laboratory cultures representing the respective dominant cyanobacterial genera. Two intestinal and one macrophage cell lines were used to detect in vitro pro-inflammatory activity of the LPS. All LPSs isolated from CyanoHABs and laboratory cultures induced cytokines production in at least one in vitro model, except for LPSs from the Microcystis PCC7806 culture. LPSs isolated from cyanobacteria showed unique migration patterns in SDS-PAGE that were qualitatively distinct from those of endotoxins from Gram-negative bacteria. There was no clear relationship between the biological activity of the LPS and the share of genomic DNA of Gram-negative bacteria in the respective biomass. Thus, the total share of Gram-negative bacteria, or the presence of Escherichia coli-like LPSs, did not explain the observed pro-inflammatory activities. The pro-inflammatory properties of environmental mixtures of LPSs from CyanoHABs indicate their human health hazards, and further attention should be given to their assessment and monitoring.

• Keywords
• cyanobacteria, cyanobacterial harmful bloom, inflammation, intestine, lipopolysaccharide, macrophage,
• MeSH
• Endotoxins metabolism   MeSH
• Humans MeSH
• Lipopolysaccharides pharmacology   MeSH
• Microcystis * MeSH
• Cyanobacteria * metabolism   MeSH
• Harmful Algal Bloom MeSH
• Fresh Water microbiology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Endotoxins MeSH
• Lipopolysaccharides MeSH

Massive toxic blooms of cyanobacteria represent a major threat to water supplies worldwide. Here, the biological activities of lipopolysaccharide (LPS) isolated from Microcystis aeruginosa, the most prominent cyanobacteria in water bloom, were studied. LPS was isolated from complex environmental water bloom samples dominated by M. aeruginosa, and from laboratory cultures of non-axenic as well as axenic M. aeruginosa strains PCC7806 and HAMBI/UHCC130. Employing human blood-based in vitro tests, the LPS isolated from complex water bloom revealed the priming of both major blood phagocyte population monocytes and polymorphonuclear leukocytes documented by the increased surface expression of CD11b and CD66b. This was accompanied by a water bloom LPS-mediated dose-dependent induction of tumor necrosis factor α, interleukin-1β, and interleukin-6 production. In accordance with its priming effects, water bloom LPS induced significant activation of p38 and ERK1/2 kinases, as well as NF-κB phosphorylation, in isolated polymorphonuclear leukocytes. Interestingly, the pro-inflammatory potential of LPS from the axenic strain of M. aeruginosa was not lower compared to that of LPS isolated from non-axenic strains. In contrast to the biological activity, water bloom LPS revealed almost twice higher pyrogenicity levels compared to Escherichia coli LPS, as analyzed by the PyroGene test. Moreover, LPS from the non-axenic culture exhibited higher endotoxin activity in comparison to LPS from axenic strains. Taking the above findings together, M. aeruginosa LPS can contribute to the health risks associated with contamination by complex water bloom mass.

• Keywords
• cyanobacteria, endotoxin, inflammation, leukocytes, lipopolysaccharide, water bloom,
• MeSH
• CD11b Antigen metabolism   MeSH
• Antigens, CD metabolism   MeSH
• Cytokines blood   MeSH
• Eutrophication MeSH
• GPI-Linked Proteins metabolism   MeSH
• Cells, Cultured MeSH
• Laboratories MeSH
• Leukocytes, Mononuclear drug effects   metabolism   MeSH
• Humans MeSH
• Lipopolysaccharides toxicity   MeSH
• Microcystis * MeSH
• Cell Adhesion Molecules metabolism   MeSH
• Immunity, Innate drug effects   MeSH
• Pyrogens toxicity   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• CD11b Antigen MeSH
• Antigens, CD MeSH
• CEACAM8 protein, human MeSH Browser
• Cytokines MeSH
• GPI-Linked Proteins MeSH
• ITGAM protein, human MeSH Browser
• Lipopolysaccharides MeSH
• Cell Adhesion Molecules MeSH
• Pyrogens MeSH

RATIONALE: Pulmonary hypertension (PH) represents a serious health complication accompanied with hypoxic conditions, elevated levels of asymmetric dimethylarginine (ADMA), and overall dysfunction of pulmonary vascular endothelium. Since the prevention strategies for treatment of PH remain largely unknown, our study aimed to explore the effect of nitro-oleic acid (OA-NO2), an exemplary nitro-fatty acid (NO2-FA), in human pulmonary artery endothelial cells (HPAEC) under the influence of hypoxia or ADMA. METHODS: HPAEC were treated with OA-NO2 in the absence or presence of hypoxia and ADMA. The production of nitric oxide (NO) and interleukin-6 (IL-6) was monitored using the Griess method and ELISA, respectively. The expression or activation of different proteins (signal transducer and activator of transcription 3, STAT3; hypoxia inducible factor 1α, HIF-1α; endothelial nitric oxide synthase, eNOS; intercellular adhesion molecule-1, ICAM-1) was assessed by the Western blot technique. RESULTS: We discovered that OA-NO2 prevents development of endothelial dysfunction induced by either hypoxia or ADMA. OA-NO2 preserves normal cellular functions in HPAEC by increasing NO production and eNOS expression. Additionally, OA-NO2 inhibits IL-6 production as well as ICAM-1 expression, elevated by hypoxia and ADMA. Importantly, the effect of OA-NO2 is accompanied by prevention of STAT3 activation and HIF-1α stabilization. CONCLUSION: In summary, OA-NO2 eliminates the manifestation of hypoxia- and ADMA-mediated endothelial dysfunction in HPAEC via the STAT3/HIF-1α cascade. Importantly, our study is bringing a new perspective on molecular mechanisms of NO2-FAs action in pulmonary endothelial dysfunction, which represents a causal link in progression of PH. Graphical Abstract ᅟ.

• Keywords
• Asymmetric dimethylarginine, Human pulmonary artery endothelial cell, Hypoxia, Nitro-oleic acid, Pulmonary hypertension,
• MeSH
• Arginine analogs & derivatives   pharmacology   MeSH
• Pulmonary Artery cytology   MeSH
• Cell Adhesion drug effects   MeSH
• Endothelial Cells drug effects   metabolism   physiology   MeSH
• Hypoxia-Inducible Factor 1, alpha Subunit metabolism   MeSH
• Cell Hypoxia drug effects   MeSH
• Interleukin-6 metabolism   MeSH
• Cells, Cultured MeSH
• Oleic Acids pharmacology   MeSH
• Humans MeSH
• Intercellular Adhesion Molecule-1 metabolism   MeSH
• Nitric Oxide metabolism   MeSH
• Cell Movement drug effects   MeSH
• Nitric Oxide Synthase Type III metabolism   MeSH
• STAT3 Transcription Factor antagonists & inhibitors   metabolism   MeSH
• Cell Survival drug effects   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Arginine MeSH
• Hypoxia-Inducible Factor 1, alpha Subunit MeSH
• HIF1A protein, human MeSH Browser
• IL6 protein, human MeSH Browser
• Interleukin-6 MeSH
• Oleic Acids MeSH
• Intercellular Adhesion Molecule-1 MeSH
• N,N-dimethylarginine MeSH Browser
• NOS3 protein, human MeSH Browser
• Nitric Oxide MeSH
• STAT3 protein, human MeSH Browser
• Nitric Oxide Synthase Type III MeSH
• STAT3 Transcription Factor MeSH