Significance of zinc re-absorption in Zn dynamic regulation in marine fish revealed by pharmacokinetic model.

Zinc (Zn) is an essential but toxic trace element and is widely available in the natural environment. In the present study, we developed a re-absorption physiologically based pharmacokinetic (PBPK) model based on long-term dietary exposure to gain insights into the physiological mechanisms of uptake, tissue distribution, storage, and excretion of Zn in marine juvenile gilt-head breams Sparus aurata (with stomach). The PBPK model incorporated the kinetic processes of Zn transfer from fish liver to gastrointestinal system and used the Markov Monte Carlo algorithm to estimate the distribution of model parameters. The model fit indicated that the stomach and intestine of fish were key organs in regulating the concentration of Zn entering the internal environment, with excess exogenous Zn (120 mg/kg) being excreted in feces (rate constant of 5.23 d-1). Modeling results also indicated that liver (3.00 d-1), spleen (1.41 d-1) and kidney (0.51 d-1) were the main tissues responding to blood Zn flux by accumulation and detoxification. Fish kidneys exposed to 60 mg/kg and 120 mg/kg Zn had different regenerative capacities, resulting in different detoxification functions. A higher dietary Zn (120 mg/kg) disrupted the intestinal reabsorption process in marine fish. This study showed that exogenous Zn was directly accumulated in organs through the gastrointestinal-hepatic system, which is an important pathways for regulating metal homeostasis in marine fish. The results provided important understanding of the mechanisms of metal regulation and transport in marine fish.

Cu(II)-Dependent Spine Development Injury in Zebrafish (Danio rerio) with Organ Heterogeneous Cu Imbalance.

Growing evidence suggests that the imbalance of Cu leads to multiorgan diseases or other adverse effects, but the underlying mechanisms remain largely unknown. Herein, we used zebrafish to uncover the mystery of organ heterogeneous responses to Cu stress and Cu(II)-dependent spine developmental injury in the early organogenesis stage. We first demonstrated that Cu(I) was distributed in the entire body, but high contents of Cu(II) were accumulated in the yolk sac and eye in normal zebrafish larvae. Cu exposure from birth to 144 hpf caused no obvious damage to Cu-metabolizing organs (liver and intestine), despite the elevated Cu(I) and Cu(II) levels. However, the spine was more sensitive to the Cu exposure. In the spine region, the Cu(I) level remained stable, whereas the level of Cu(II) significantly increased, which was highly associated with spine development injury. A significant negative correlation between Cu(II) and the spine-related parameters was identified. Moreover, cuproptosis caused spine development deformation during the early embryogenesis stage. Spine-related pathways such as somitegenesis significantly changed in the early embryogenesis period, and 5 spine-related pathways were significantly altered in the larval stage at 96 hpf. Our study suggested that Cu stress induced organ heterogeneous Cu imbalance and Cu(II)-dependent spine development injury in zebrafish.

In vivo bioaccumulation and responses of hemocytes of mussels Perna viridis to microplastics and nanoplastics exposure.

Growing micro- and nano-plastic (MNPs) pollution in the environment poses a threat to marine animals. Due to their excellent filtration capacity, bivalves can easily ingest MNPs, which could be translocated to open circulation system with potential risks. In the present study, the accumulation and elimination of MNPs (200 nm and 1 µm) in the mussel hemolymph serum and hemocytes were firstly quantified, and the differential sensitiveresponses of two subpopulations of hemocytes were then explored by in vivo exposure under environmentally relevant concentration of MNPs (200 µg/L). We demonstrated that MNPs were readily translocated into hemolymph serum, but were immediately followed by efficient internalization by hemocytes. Remarkably, concentrations of MNPs in hemolymph were only 0.63 and 0.39 times lower than the ambient exposure concentration. Granulocytes displayed a much higher potential of accumulating MNPs than the agranulocytes. MPs were more readily internalized by granulocytes, with their estimated maximum bioaccumulation factor (BCF) of 0.29 L/g. Due to the primary function of phagocytic encapsulation of MNPs by granulocytes, lysosome features especially the decline of subsequent lysosome membrane potential could be a potential sensitive biomarker in response to MNPs exposure. Our results provided insights on the bioaccumulation of MNPs at the cellular levels in marine bivalves.

Reduced copper uptake and efflux by the mussel Mytilus coruscus after Cu exposure: Implication for biomonitoring.

The hard-shell mussels Mytilus coruscus have been extensively employed in pollution biomonitoring. Earlier studies indicated that metal concentrations in Mytilus coruscus may not accurately reflect the true metal contamination levels in the sampling areas, possibly due to their modified metal uptake and efflux. Given the likelihood of mussels in the field being exposed to intermittent metal contaminants, this study investigated whether different Cu pre-exposures significantly affected its uptake and efflux upon Cu exposure. We found significant reduction in Cu uptake rate constant (ku) and efflux rate constant (ke) in the mussels with varying Cu pre-exposure regimes. Specifically, the ku decreased from 1.55 ± 0.37 L g-1 d-1 in the control group to 0.65 ± 0.19 after 5 days and 0.53 ± 0.28 after 15 days of exposure to 20 µg L-1 Cu, respectively, and then was further reduced to as low as 0.096 ± 0.046 L g-1 d-1 following a 5-day exposure at 50 µg L-1 Cu. Similarly, the ke decreased from 0.18 ± 0.020 to 0.15 ± 0.015 d-1 following 5-15 days of exposure to 20 µg L-1 Cu, and further decreased to 0.081 ± 0.023 d-1 after a 5-day exposure at 50 µg L-1 Cu. Our subcellular distribution analysis underscored the critical role of the metallothionein-like protein (MTLP) fraction in modifying both Cu ku and ke during the rapid-depuration phase (ke1), whereas the metal-rich granule (MRG) fraction influenced the ke during the second depuration phase (ke2). This study demonstrated that environmental assessments utilizing biomonitoring species should consider the exposure of these organisms to ensure accurate interpretations of metal contamination in marine ecosystems and enhance the effectiveness of these species in environmental monitoring. This crucial factor is often overlooked, potentially skewing data and leading to misinterpretations of environmental health and pollution levels.

Modeling the differential functional responses and selectivity of a marine copepod to nano/microplastics in mixture.

Nano- and microplastics (NMPs) pollution is widespread in the oceans, posing potential risks to marine species. This study examined the accumulation capacity and selectivity potentials of NMPs by a marine copepod Parvocalanus crassirostris under different food mixtures by modeling the combined biokinetic and functional response. We investigated two sizes of NMPs (200 nm and 5 µm) across a concentration gradient (0 - 5000 µg/L) and varying diatom abundances (0, 104, 105 cells/mL). Fluorescence imaging and quantification revealed that P. crassirostris actively ingested NMPs at low concentration. Accumulation increased with NMPs concentration but eventually saturated due to gut capacity limits, following a Holling type II functional response (i.e., hyperbolic curve). Our novel functional response model estimated the key parameters and demonstrated that the maximum accumulation reached 5.3 % of dry weight with averaged half-saturation constants of 229 µg/L. The size of NMPs did not significantly affect the total accumulation or satiety levels. The presence of diatoms influenced the feeding selectivity and decreased the microplastic accumulation by 73 % at 105 cells/mL, while facilitating nanoplastic accumulation by 81 % at 104 cells/mL. This study enhanced our understanding of NMPs bioavailability and environmental fate in marine ecosystems.

Unveiling osmoregulation and immunological adaptations in Eleutheronema tetradactylum gills through high-throughput single-cell transcriptome sequencing.

The fourfinger threadfin fish (Eleutheronema tetradactylum) is an economically significant species renowned for its ability to adapt to varying salinity environments, with gills serving as their primary organs for osmoregulation and immune defense. Previous studies focused on tissue and morphological levels, whereas ignored the cellular heterogeneity and the crucial gene information related to core cell subsets within E. tetradactylum gills. In this study, we utilized high-throughput single-cell RNA sequencing (scRNA-seq) to analyze the gills of E. tetradactylum, characterizing 16 distinct cell types and identifying unique gene markers and enriched functions associated within each cell type. Additionally, we subdivided ionocyte cells into four distinct subpopulations for the first time in E. tetradactylum gills. By employing weighted gene co-expression network analysis (WGCNA), we further investigated the cellular heterogeneity and specific response mechanisms to salinity fluctuant. Our findings revealed the intricate osmoregulation and immune functions of gill cells, highlighting their crucial roles in maintaining homeostasis and adapting to fluctuating salinity levels. This comprehensive cell-type atlas provides valuable insights into the species adaptive strategies, contributing to the conservation and management of this commercially significant fish as well as other euryhaline species.

Copper Toxicity in Acidic Phytoplankton: Impacts of Labile Cu Trafficking and Causes of Mitochondria Dysfunction.

Most studies on Cu toxicity relied on indirect physicochemical parameters to predict Cu toxicity resulting from adverse impacts. This study presents a systematic and intuitive picture of Cu toxicity induced by exogenous acidification in phytoplankton Chlamydomonas reinhardtii. We first showed that acidification reduced the algal resistance to environmental Cu stress with a decreased growth rate and increased Cu bioaccumulation. To further investigate this phenomenon, we employed specific fluorescent probes to visualize the intracellular labile Cu pools in different algal cells. Our findings indicated that acidification disrupted the intracellular labile Cu trafficking, leading to a significant increase in labile Cu(I) pools. At the molecular level, Cu toxicity resulted in the inhibition of the Cu(I) import system and activation of the Cu(I) export system in acidic algal cells, likely a response to the imbalance in intracellular labile Cu trafficking. Subcellular analysis revealed that Cu toxicity induced extensive mitochondrial dysfunction and impacted the biogenesis and assembly of the respiratory chain complex in acidic algal cells. Concurrently, we proposed that the activation of polyP synthesis could potentially regulate disrupted intracellular labile Cu trafficking. Our study offers an intuitive, multilevel perspective on the origins and impacts of Cu toxicity in living organisms, providing valuable insights on metal toxicity.

Contrasting the distribution kinetics of microplastics and nanoplastics in medaka following exposure and depuration.

Direct ingestion of micro/nanoplastics (MNPs) results in significant accumulation in gastrointestinal (GI) tract of fish. The breathing process of fish makes MNPs easily retained in their gills. However, the uptake of MNPs in other fish organs remains largely unknown, let alone their kinetic processes. Herein, microplastics (MPs) and nanoplastics (NPs) in vivo imaging and precise quantification in various tissues (GI tract, gill, liver, brain, eye, and skin) of seawater (SW)- and freshwater (FW)- acclimated medaka Oryzias melastigma were achieved at an environmentally relevant concentration. Subsequently, the distribution kinetics of MNPs was investigated over a 96-h uptake and 48-h depuration period. MNPs were quickly and mostly captured in GI tract and gill of O. melastigma, and then transferred to liver and brain likely via blood circulation. Such transport was more efficient for NPs as compared to MPs, as evidenced by the consistently higher bioconcentration factors in both SW and FW conditions. The detection of MNPs in eye and skin of O. melastigma was more of an adsorption process, although the specific mechanisms of adsorption and absorption process can hardly be clearly differentiated. This study presented distribution kinetics of MNPs in O. melastigma and highlighted their possible transportation among tissues.

Visible Combined Near-Infrared in Situ Imaging Revealed Dynamic Effects of Microplastic Fibers and Beads in Zebrafish.

Microplastics (MPs) as emerging contaminants are widely present in the environment and are ubiquitously ingested and accumulated by aquatic organisms. MPs may be quickly eliminated after a brief retention in aquatic animals (such as the digestive tract); thus, understanding the damage caused by MPs during this process and whether the damage can be recovered is important. Here, we proposed the use of visible light imaging to track MPs combined with near-infrared (NIR) imaging to reveal the in situ impacts of MPs. The combination of these two techniques allows for the simultaneous investigation of the localization and functionality of MPs in vivo. We investigated the effects of two types of MPs on zebrafish, microplastic fibers (MFs) and microplastic beads (MBs). The results showed that MPs larger than 10 µm primarily accumulated in the intestines of zebrafish. Both MFs and MBs disrupted the redox balance of the intestine, and the location of the damage was consistent with the heterogeneous accumulation of MPs. MFs caused greater and more difficult-to-recover damage compared to MBs, which was closely related to the slower elimination rate of MFs. Our study highlights the importance of capturing the dynamic toxicological effects of MPs on organisms. Fibrous MPs and spherical MPs clearly had distinct effects on their toxicokinetics and toxicodynamics in fish.

Cadmium impacts on calcium mineralization of zebrafish skeletal development and behavioral impairment.

Cadmium (Cd) poses significant risks to aquatic organisms due to its toxicity and ability to disrupt the cellular processes. Given the similar atomic radius of Cd and calcium (Ca), Cd may potentially affect the Ca homeostasis, which can lead to impaired mineralization of skeletal structures and behavioral abnormalities. The formation of the spinal skeleton involves Ca transport and mineralization. In this study, we conducted an in-depth investigation on the effects of Cd at environmental concentrations on zebrafish (Danio rerio) skeletal development and the underlying molecular mechanisms. As the concentration of Cd increased, the accumulation of Cd in zebrafish larvae also rose, while the Ca content decreased significantly by 3.0 %-57.3 %, and vertebral deformities were observed. Transcriptomics analysis revealed that sixteen genes involved in metal absorption were affected. Exposure to 2 µg/L Cd significantly upregulated the expression of these genes, whereas exposure to 10 µg/L resulted in their downregulation. Consequently, exposure of zebrafish larvae to 10 µg/L of Cd inhibited the body segmentation growth and skeletal mineralization development by 29.1 %-56.7 %. This inhibition was evidenced by the downregulation of mineral absorption genes and decreased Ca accumulation. The findings of this study suggested that the inhibition of skeletal mineralization was likely attributed to the disruption of mineral absorption, thus providing novel insights into the mechanisms by which metal pollutants inhibit the skeletal development of fish.

Metabolome analysis revealed the critical role of betaine for arsenobetaine biosynthesis in the marine medaka (Oryzias melastigma).

Arsenobetaine (AsB), a non-toxic arsenic (As) compound found in marine fish, structurally resembles betaine (GB), a common methyl donor in organisms. This study investigates the potential role of GB in AsB synthesis in marine medaka (Oryzias melastigma) using metabolomic analysis. Dietary exposure to arsenate (As(V)) and varying GB concentrations (0.05% and 0.1% in diets) increased total As and AsB bioaccumulation, particularly in marine medaka muscle. Metabolomic analysis revealed that GB played a crucial role in promoting up-regulation in methylthioadenosine (MTA) by modulating the methionine cycle and down-regulation in glutathione (GSH) by modulating the glutathione cycle. Methionine metabolism and GSH, potentially binding again to exogenous GB, could synchronously produce more non-toxic AsB. Combining verification experiments of differential metabolites of Escherichia coli in vitro, GB, GSH, S-adenosylmethionine (SAM), and arsenocholine (AsC) entered methionine and glutathione metabolism pathways to generate more AsB. These findings underscore the GB's crucial regulatory role in modulating the synthesis of AsB. This study provides vital insights into the interplay between the structural analogs GB and AsB, offering specific strategies to enhance the detoxification mechanisms of marine fish in As-contaminated environments.

Microplastics Biofragmentation and Degradation Kinetics in the Plastivore Insect Tenebrio molitor.

The insect Tenebrio molitor possesses an exceptional capacity for ultrafast plastic biodegradation within 1 day of gut retention, but the kinetics remains unknown. Herein, we investigated the biofragmentation and degradation kinetics of different microplastics (MPs), i.e., polyethylene (PE), poly(vinyl chloride) (PVC), and poly(lactic acid) (PLA), in T. molitor larvae. The intestinal reactions contributing to the in vivo MPs biodegradation were concurrently examined by utilizing aggregated-induced emission (AIE) probes. Our findings revealed that the intestinal biofragmentation rates essentially followed the order of PLA > PE > PVC. Notably, all MPs displayed retention effects in the intestine, with PVC requiring the longest duration for complete removal/digestion. The dynamic rate constant of degradable MPs (0.2108 h-1 for PLA) was significantly higher than that of persistent MPs (0.0675 and 0.0501 h-1 for PE and PVC, respectively) during the digestive gut retention. Surprisingly,T. molitor larvae instinctively modulated their internal digestive environment in response to in vivo biodegradation of various MP polymers. Esterase activity and intestinal acidification both significantly increased following MPs ingestion. The highest esterase and acidification levels were observed in the PLA-fed and PVC-fed larvae, respectively. High digestive esterase activity and relatively low acidification levels inT. molitor larvae may, to some extent, contribute to more efficient MPs removal within the plastic-degrading insect. This work provided important understanding of MPs biofragmentation and intestinal responses to in vivo MPs biodegradation in plastic-degrading insects.

Differentiation of cellular responses to particulate and soluble constituents in sunscreen products.

Despite the growing awareness of potential human and environmental risks associated with sunscreens, identifying the specific constituents responsible for their potential toxicity is challenging. In this study, we applied three different types of sunscreens with contrasting compositions and compared the effects of their particulate and soluble fractions based on 15 cellular biomarkers of HaCaT cells. Multilinear regression analysis revealed that the internalized soluble fractions played a primary role in the overall cytotoxicity of sunscreen mixtures, which was primarily attributed to their biotransformation, generating metabolites with higher toxicity. The presence of plastic microspheres in sunscreens either inhibited the internalization of soluble fractions or led to their redistribution toward lysosomes. Conversely, subcellular toxicity resulting from the sunscreen mixture was predominantly influenced by particulates. Bio-transformable particulates such as ZnO dissolved in the organelles and induced higher subcellular toxicity compared to bioinert particulates such as microplastics. Subcellular biomarkers including lysosomal count, lysosomal size, mitochondrial count and mitochondrial shape emerged as the potential predictors of sunscreen presence. Our study provides important understanding of sunscreen toxicity by elucidating the differential impacts of particulate and soluble fractions in mixture contaminants.

A novel route of intercellular copper transport and detoxification in oyster hemocytes.

Bivalve hemocytes are oyster immune cells composed of several cellular subtypes with different functions. Hemocytes accumulate high concentrations of copper (Cu) and exert critical roles in metal sequestration and detoxification in oysters, however the specific biochemical mechanisms that govern this have yet to be fully uncovered. Herein, we demonstrate that Cu(I) is predominately sequestered in lysosomes via the Cu transporter ATP7A in hemocytes to reduce the toxic effects of intracellular Cu(I). We also found that Cu(I) is translocated along tunneling nanotubes (TNTs) relocating from high Cu(I) cells to low Cu(I) cells, effectively reducing the burden caused by overloaded Cu(I), and that ATP7A facilitates the efflux of intracellular Cu(I) in both TNTs and hemocyte subtypes. We identify that elevated glutathione (GSH) contents and heat-shock protein (Hsp) levels, as well as the activation of the cell cycle were critical in maintaining the cellular homeostasis and function of hemocytes exposed to Cu. Cu exposure also increased the expression of membrane proteins (MYOF, RalA, RalBP1, and cadherins) and lipid transporter activity which can induce TNT formation, and activated the lysosomal signaling pathway, promoting intercellular lysosomal trafficking dependent on increased hydrolase activity and ATP-dependent activity. This study explores the intracellular and intercellular transport and detoxification of Cu in oyster hemocytes, which may help in understanding the potential toxicity and fate of metals in marine animals.

Copper redox state in cells and aquatic organisms: Implication for toxicity.

Copper (Cu) redox state has been an important issue in biology and toxicology research, but many research gaps remain to be explored due to the limitations in the detecting techniques. Herein, the regulation of Cu homeostasis, including absorption, translocation, utilization, storage, and elimination behavior is discussed. Cuproptosis, a newly identified type of cell death caused by excessive Cu accumulation, which results in the aggregation of DLAT protein or the loss of Fe-S cluster and finally proteotoxic stress, is reviewed. Several longstanding mysteries of diseases such as Wilson disease and toxic effects, may be attributed to cuproptosis. Furthermore, we review the advanced detection methods and application of Cu(I) and Cu(II), especially the in-situ imaging techniques such as XANES, and chemosensors. Most of the existing studies using these detection techniques focus on the bioaccumulation and toxicity of Cu(I) and Cu(II) in cells and aquatic organisms. Finally, it will be important to identify the roles of Cu(I) and Cu(II) in the growth, development, and diseases of organisms, as well as the relationship between bioaccumulation and toxicity of Cu(I) and Cu(II) in cellular and aquatic toxicology.

Tracking the Cellular Degradation of Silver Nanoparticles: Development of a Generic Kinetic Model.

Understanding the degradation of nanoparticles (NPs) after crossing the cell plasma membrane is crucial in drug delivery designs and cytotoxicity assessment. However, the key factors controlling the degradable kinetics remain unclear due to the absence of a quantification model. In this study, subcellular imaging of silver nanoparticles (AgNPs) was used to determine the intracellular transfer of AgNPs, and single particle ICP-MS was utilized to track the degradation process. A cellular kinetic model was subsequently developed to describe the uptake, transfer, and degradation behaviors of AgNPs. Our model demonstrated that the intracellular degradation efficiency of AgNPs was much higher than that determined by mimicking testing, and the degradation of NPs was highly influenced by cellular factors. Specifically, deficiencies in Ca or Zn primarily decreased the kinetic dissolution of NPs, while a Ca deficiency also resulted in the retardation of NP transfer. The biological significance of these kinetic parameters was strongly revealed. Our model indicated that the majority of internalized AgNPs dissolved, with the resulting ions being rapidly depurated. The release of Ag ions was largely dependent on the microvesicle-mediated route. By changing the coating and size of AgNPs, the model results suggested that size influenced the transfer of NPs into the degradation process, whereas coating affected the degradation kinetics. Overall, our developed model provides a valuable tool for understanding and predicting the impacts of the physicochemical properties of NPs and the ambient environment on nanotoxicity and therapeutic efficacy.

Combined simultaneous transsphenoidal and transcranial regimen improves surgical outcomes in complex giant pituitary adenomas: a longitudinal retrospective cohort study.

BACKGROUND: Surgical treatment of complex giant pituitary adenomas (GPAs) presents significant challenges. The efficacy and safety of combining transsphenoidal and transcranial approaches for these tumors remain controversial. In this largest cohort of patients with complex GPAs, we compared the surgical outcomes between those undergoing a combined regimen and a non-combined regimen. We also examined the differences in risks of complications, costs, and logistics between the two groups, which might offer valuable information for the appropriate management of these patients. PATIENTS AND METHODS: This was a multicenter retrospective cohort study conducted at 13 neurosurgical centers. Consecutive patients who received a combined or non-combined regimen for complex GPAs were enrolled. The primary outcome was gross total resection, while secondary outcomes included complications, surgical duration, and relapse. A propensity score-based weighting method was used to account for differences between the groups. RESULTS: Out of 647 patients [298 (46.1%) women, mean age: 48.5 ± 14.0 years] with complex GPAs, 91 were in the combined group and 556 were in the noncombined group. Compared with the noncombined regimen, the combined regimen was associated with a higher probability of gross total resection [50.5% vs. 40.6%, odds ratio (OR): 2.18, 95% confidence interval (CI): 1.30-3.63, P = 0.003]. The proportion of patients with life-threatening complications was lower in the combined group than in the non-combined group (4.4% vs. 11.2%, OR: 0.25, 95% CI: 0.08-0.78, P = 0.017). No marked differences were found between the groups in terms of other surgical or endocrine-related complications. However, the combined regimen exhibited a longer average surgery duration of 1.3 h ( P < 0.001) and higher surgical costs of 22,000 CNY (~ 3,000 USD, P = 0.022) compared with the noncombined approach. CONCLUSIONS: The combined regimen offered increased rates of total resection and decreased incidence of life-threatening complications, which might be recommended as the first-line choice for these patients.

Single-Cell RNA Sequencing Profiling Cellular Heterogeneity and Specific Responses of Fish Gills to Microplastics and Nanoplastics.

Fish gills are highly sensitive organs for microplastic (MP) and nanoplastic (NP) invasions, but the cellular heterogeneity of fish gills to MPs and NPs remains largely unknown. We employed single-cell RNA sequencing to investigate the responses of individual cell populations in tilapia Oreochromis niloticus gills to MP and NP exposure at an environmentally relevant concentration. Based on the detected differentially expressed gene (DEG) numbers, the most affected immune cells by MP exposure were macrophages, while the stimulus of NPs primarily targeted T cells. In response to MPs and NPs, H+-ATPase-rich cells exhibited distinct changes as compared with Na+/K+-ATPase-rich cells and pavement cells. Fibroblasts were identified as a potential sensitive cell-type biomarker for MP interaction with O. niloticus gills, as evidenced by the largely reduced cell counts and the mostly detected DEGs among the 12 identified cell populations. The most MP-sensitive fibroblast subpopulation in O. niloticus gills was lipofibroblasts. Cell-cell communications between fibroblasts and H+-ATPase-rich cells, neurons, macrophages, neuroepithelial cells, and Na+/K+-ATPase-rich cells in O. niloticus gills were significantly inhibited by MP exposure. Collectively, our study demonstrated the cellular heterogeneity of O. niloticus gills to MPs and NPs and provided sensitive markers for their toxicological mechanisms at single-cell resolution.

Dynamic Regulation of Intracellular Labile Cu(I)/Cu(II) Cycle in Microalgae Chlamydomonas reinhardtii: Disrupting the Balance by Cu Stress.

The labile metal pool involved in intracellular trafficking and homeostasis is the portion susceptible to environmental stress. Herein, we visualized the different intracellular distributions of labile Cu(I) and Cu(II) pools in the alga Chlamydomonas reinhardtii. We first demonstrated that labile Cu(I) predominantly accumulated in the granules within the cytoplasmic matrix, whereas the labile Cu(II) pool primarily localized in the pyrenoid and chloroplast. The cell cycle played an integral role in balancing the labile Cu(I)/Cu(II) pools. Specifically, the labile Cu(II) pool primarily accumulated during the SM phase following cell division, while the labile Cu(I) pool dynamically changed during the G phase as cell size increased. Notably, the labile Cu(II) pool in algae at the SM stage exhibited heightened sensitivity to environmental Cu stress. Exogenous Cu stress disrupted the intracellular labile Cu(I)/Cu(II) cycle and balance, causing a shift toward the labile Cu(II) pool. Our proteomic analysis further identified a putative cupric reductase, potentially capable of reducing Cu(II) to Cu(I), and four putative multicopper oxidases, potentially capable of oxidizing Cu(I) to Cu(II), which may be involved in the conversion between the labile Cu(I) pool and labile Cu(II) pool. Our study elucidated a dynamic cycle of the intracellular labile Cu(I)/Cu(II) pools, which were accessible and responsive to environmental changes.