Toxic dinoflagellate Karenia mikimotoi induces apoptosis in Neuro-2a cells through an oxidative stress-mediated mitochondrial pathway.

The dinoflagellate Karenia mikimotoi is a toxic bloom-forming species that threatens aquaculture and public health worldwide. Previous studies showed that K. mikimotoi induces neurotoxicity; however, the underlying mechanism is poorly understood. In this study, three neural cell lines were used to investigate the potential neurotoxicity of K. mikimotoi. The tested cells were exposed to a ruptured cell solution (RCS) of K. mikimotoi at different concentrations (0.5 × 105, 1.0 × 105, 2.0 × 105, 4.0 × 105, and 6 × 105 cells mL-1) for 24 h, and the RCS decreased cell viabilities and promoted Neuro-2a (N2A) cell apoptosis in a dose-dependent manner. The underlying mechanism was further investigated in N2A cells. At the biochemical level, the RCS stimulated reactive oxygen species (ROS) and malondialdehyde (MDA) formation, decreased SOD activity, and reduced mitochondrial membrane potential (MMP). At the gene level, the moderate RCS treatment (2.0 × 105 cells mL-1) upregulated antioxidant response genes (e.g., nrf-2, HO-1, NQO-1, and cat) to alleviate RCS-induced oxidative stress, while the high RCS treatment (4.0 × 105 cells mL-1) downregulated these genes, thereby aggravating oxidative stress. Meanwhile, apoptosis-related genes (e.g., p53, caspase 3, and bax2) were significantly upregulated and the anti-apoptotic gene bcl2 was suppressed after RCS treatment. Western blotting results for Caspase 3, Bax2 and Bcl2 were consistent with the mRNA trends. These results revealed that K. mikimotoi RCS can induce neural cell apoptosis via the oxidative stress-mediated mitochondrial pathway, providing novel insights into the neurotoxicity of K. mikimotoi.

Cadmium chloride exposure impairs the growth and behavior of Drosophila via ferroptosis.

Cadmium (Cd) is a widely distributed toxic heavy metal that enters the environment via anthropogenic mobilization and accumulates in plants and animals, causing metabolic abnormalities even mortality. Although the toxic effects and stress damage of cadmium have been investigated extensively over the past few decades, research on its ability to trigger ferroptosis, growth retardation, and behavioral abnormalities is insufficient. As a result, the effects of CdCl2 exposure on growth and development, activity and sleep, and ferroptosis in this study were examined in fruit fly (Drosophila melanogaster). When exposed to 0.5 mM CdCl2, the entire growth period from larvae to adults was prolonged, and the rates of pupation and eclosion were decreased. Additionally, CdCl2 exposure resulted in a decrease in body weight and individual size of fruit fly and high lethality rate. Moreover, CdCl2 exposure altered fruit fly behavior, including decreased activity and increased sleep duration, particularly in females. Ferrostatin-1 (Fer-1) is a potent selective ferroptosis inhibitor that effectively slows lipid hydroperoxide accumulation to rescue body size reduction and restore activity and sleep in CdCl2-exposed female flies. CdCl2 exposure could induce ferroptosis in fruit fly mechanistically, as evidenced by inhibition of Nrf2 signaling pathway, accumulation of lipid peroxidation, impairment of GPX4 antioxidant system, and upregulation of iron metabolism. Our findings suggest that Cd exposure triggers ferroptosis, which leads to growth retardation and behavioral disorders in fruit fly.

Antagonistic and synergistic effects of warming and microplastics on microalgae: Case study of the red tide species Prorocentrum donghaiense.

Bibliometric network analysis has revealed that the widespread distribution of microplastics (MPs) has detrimental effects on marine organisms; however, the combined effects of MPs and climate change (e.g., warming) is not well understood. In this study, Prorocentrum donghaiense, a typical red tide species in the East China Sea, was exposed to different MP concentrations (0, 1, 5, and 10 mg L-1) and temperatures (16, 22, and 28 °C) for 7 days to investigate the combined effects of MPs and simulated ocean warming by measuring different physiological parameters, such as cell growth, pigment contents (chlorophyll a and carotenoid), relative electron transfer rate (rETR), reactive oxygen species (ROS), superoxide dismutase (SOD), malondialdehyde (MDA), and adenosine triphosphate (ATP). The results demonstrated that MPs significantly decreased cell growth, pigment contents, and rETRmax, but increased the MDA, ROS, and SOD levels for all MP treatments at low temperature (16 °C). However, high temperatures (22 and 28 °C) increased the pigment contents and rETRmax, but decreased the SOD and MDA levels. Positive and negative effects of high temperatures (22 or 28 °C) were observed at low (1 and 5 mg L-1) and high MP (10 mg L-1) concentrations, respectively, indicating the antagonistic and synergistic effects of combined warming and MP pollution. These results imply that the effects of MPs on microalgae will likely not be substantial in future warming scenarios if MP concentrations are controlled at a certain level. These findings expand the current knowledge of microalgae in response to increasing MP pollution in future warming scenarios.

Transcriptome sequencing of a toxic dinoflagellate, Karenia mikimotoi subjected to stress from solar ultraviolet radiation.

Solar ultraviolet radiation (UVR) is a stress factor in aquatic environments and may act directly or indirectly on orgnisms in the upper layers of the water column. However, UVR effects are usually species-specific and difficult to extrapolate. Here we use the HAB-forming, toxic dinoflagellate Karenia mikimotoi (which was found to be relatively resistant in previous studies) to investigate its transcriptional responses to a one-week UVR exposure. For this, batch cultures of K. mikimotoi were grown with and without UVR, and their transcriptomes (generated via RNAseq technology) were compared. RNA-seq generated 45.31 million reads, which were further assembled to 202600 unigenes (>300bp). Among these, ca. 61% were annotated with NCBI, NR, GO, KOG, PFAM, Swiss-Prot, and KEGG database. Transcriptomic analysis revealed 722 differentially expressed unigenes (DEGs, defined as being within a |log2 fold change| ≥ 2 and padj < 0.05) responding to solar UVR, which were only 0.36% of all unigenes. 716 unigenes were down-regulated, and only 6 unigenes were up-regulated in the UVR compared to non-UVR treatment. KEGG pathway further analysis revealed DEGs were involved in the different pathway; genes involved in the ribosome, endocytosis and steroid biosynthesis pathways were highly down-regulated, but this was not the case for those involved in the energy metabolisms (including photosynthesis, oxidative phosphorylation) which may contribute to the sustainable growth observed in UVR treatment. The up-regulated expression of both zinc-finger proteins (ZFPs) and ribosomal protein L11 (RPL11) may be one of the acclimated mechanisms against UVR. In addition, this work identified down-regulated genes involved in fatty acid degradation and the hydrophobic branched chain amino acids (e.g., Valine, leucine, and isoleucine), which act as structural components of cell membranes modulating lipid homeostasis or turnover. In conclusion, the present study suggests that the toxic dinoflagellate K. mikimotoi has limited transcriptomic regulation but confirms that it appears as a tolerant species in response to solar UVR. These findings expand current knowledge of gene expression in HAB-forming species in response to natural environment factors such as solar radiation.

Developmental neurotoxicity of reserpine exposure in zebrafish larvae (Danio rerio).

Reserpine is widely used for treatment of hypertension and schizophrenia. As a specific inhibitor of monoamine transporters, reserpine is known to deplete monoamine neurotransmitters and cause decreased movement symptoms. However, how zebrafish larvae respond to reserpine treatment is not well studied. Here we show that swimming distance and average velocity are significantly reduced after reserpine exposure under various stimulatory conditions. Using liquid chromatograph-mass spectrometer analysis, decreased levels of monoamines (e.g. dopamine, noradrenaline, and serotonin) were detected in reserpine-treated larvae. Moreover, reserpine treatment significantly reduced the number of dopaminergic neurons, which was identified with th (Tyrosine Hydroxylase) in situ hybridization in the preoptic area. Interestingly, dopaminergic neuron development-associated genes, such as otpa, otpb, wnt1, wnt3, wnt5 and manf, were downregulated in reserpine treated larvae. Our data indicates that 2 mg/L reserpine exposure induces dopaminergic neuron damage in the brain, demonstrating a chemical induced depression-like model in zebrafish larvae for future drug development.

Effects of ocean acidification and solar ultraviolet radiation on physiology and toxicity of dinoflagellate Karenia mikimotoi.

A batch culture experiment was conducted to study the interactive effects of ocean acidification (OA) and solar ultraviolet radiation (UVR, 280-400 nm) on the harmful dinoflagellate Karenia mikimotoi. Cells were incubated in 7-days trials under four treatments. Physiological (growth, pigments, UVabc) and toxicity (hemolytic activity and its toxicity to zebrafish embryos) response variables were measured in four treatments, representing two factorial combinations of CO2 (400 and 1000 µatm) and solar irradiance (with or without UVR). Toxic species K. mikimotoi showed sustained growth in all treatments, and there was not statistically significant difference among four treatments. Cell pigment content decreased, but UVabc and hemolytic activity increased in all HC treatments and PAB conditions. The toxicity to zebrafish embryos of K. mikimotoi was not significantly different among four treatments. All HC and UVR conditions and the combinations of HC*UVR (HC-PAB) positively affected the UVabc, hemolytic activity in comparison to the LC*P (LC-P) treatment, and negatively affected the pigments. Ocean acidification (OA) was probably the main factor that affected the chlorophyll-a (Chl-a) and UVabc, but UVR was the main factor that affected the carotenoid (Caro) and hemolytic activity. There were no significant interactive effects of OA*UVR on growth, toxicity to zebrafish embryos. If these results are extrapolated to the natural environment, it can be hypothesized that this strain (DP-C32) of K. mikimotoi cells have the efficient mechanisms to endure the combination of ocean acidification and solar UVR. It is assumed that this toxic strain could form harmful bloom and enlarge the threatening to coastal communities, marine animals, even human health under future conditions.

Light histories influence the impacts of solar ultraviolet radiation on photosynthesis and growth in a marine diatom, Skeletonema costatum.

Phytoplanktonic species acclimated to high light are known to show less photoinhibition. However, little has been documented on how cells grown under indoor conditions for decades without exposure to UV radiation (UVR, 280-400 nm) would respond differently to solar UVR compared to those in situ grown under natural solar radiation. Here, we have shown the comparative photosynthetic and growth responses to solar UVR in an indoor- (IS) and a naturally grown (WS) Skeletonema costatum type. In short-term experiment (<1 day), Phi(PSII) and photosynthetic carbon fixation rate were more inhibited by UVR in the IS than in the WS cells. The rate of UVR-induced damages of PSII was faster and their repair was significantly slower in IS than in WS. Even under changing solar radiation simulated for vertical mixing, solar UVR-induced higher inhibition of photosynthetic rate in IS than in WS cells. During long-term (10 days) exposures to solar radiation, the specific growth rate was much lower in IS than WS at the beginning, then increased 3 days later to reach an equivalent level as that of WS. UVR-induced inhibition of photosynthetic carbon fixation in the IS was identical with that of WS at the end of the long-term exposure. The photosynthetic acclimation was not accompanied with increased contents of UV-absorbing compounds, indicating that repair processes for UVR-induced damages must have been accelerated or upgraded.

Effects of solar ultraviolet radiation on photosynthesis of the marine red tide alga Heterosigma akashiwo (Raphidophyceae).

In order to assess the short- and long-term impacts of UV radiation (UVR, 280-400nm) on the red tide alga, Heterosigma akashiwo, we exposed the cells to three different solar radiation treatments (PAB: 280-700nm, PA: 320-700nm, P: 400-700nm) under both solar and artificial radiation. A significant decrease in the effective quantum yield (Y) during high irradiance periods (i.e., local noon) was observed, but the cells partially recovered during the evening hours. Exposure to high irradiances for 15, 30, and 60min under a solar simulator followed by the recovery (8h) under dark, 9 and 100micromolphotonsm(-2)s(-1) of PAR, highlighted the importance of the irradiance level during the recovery period. Regardless the radiation treatments, the highest recovery (both in rate and total Y) was found at a PAR irradiance of 9micromolphotonsm(-2)s(-1), while the lowest was observed at 100micromolphotonsm(-2)s(-1). In all experiments, PAR was responsible for most of the observed inhibition; nevertheless, the cells exposed only to PAR had the highest recovery in any condition, as compared to the other radiation treatments. In long-term experiments (10 days) using semi-continuous cultures, there was a significant increase of UV-absorbing compounds (UV(abc)) per cell from 1.2 to >4x10(-6)microgUV(abc)cell(-1) during the first 3-5 days of exposure to solar radiation. The highest concentration of UV(abc) was found in samples exposed in the PAB as compared to PA and P treatments. Growth rates (mu) mimic the behavior of UV-absorbing compounds, and during the first 5 days mu increased from <0.2 to ca. 0.8, and stayed relatively constant at this value during the rest of the experiment. The inhibition of the Y decreased with increasing acclimation of cells. All our data indicates that H. akashiwo is a sensitive species, but was able acclimate relatively fast (3-5 days) synthesizing UV-absorbing compounds and thus reducing any impact either on photosystem II or on growth.