Levels In Zebrafish Research Articles

A novel near-infrared fluorescent probe for butyrylcholinesterase: Research for screening of natural anti-AD inhibitors

Elevated levels of butyrylcholinesterase (BuChE) have the potential to be predictive in the timely detection and diagnosis of Alzheimer's disease (AD). By inhibiting of BuChE activity can raise acetylcholine levels and intervene AD processes. Therefore, BuChE as an important factor in treatment AD has been given more and more attention in clinical studies. Given the facts above, in this study, for precise monitoring of BuChE level changes and screening for possible butyrylcholinesterase inhibitor (BuChEI) for AD diagnosis and therapy, a near-infrared (NIR) fluorescence probe (NFP-BuChE) was created. The probe exhibits excellent sensitivity and selectivity for BuChE. NFP-BuChE has been successfully applied to the detection of endogenous BuChE levels in live cells, and we successfully constructed a screening system for BuChEI on cells and a novel natural efficient BuChEI (matrine) was discovered and identified, which significantly reduced BuChE activity and thus alleviated AD symptoms. Most importantly, for the first time, we measured the changes of BuChE levels in zebrafish (0–4 days) after fertilization, various organs of zebrafish, and AD zebrafish modeled by different concentrations of AlCl3 by NFP-BuChE, and at the same time, we also validated the inhibitory effect of matrine on BuChE by NFP-BuChE in zebrafish. In addition, NFP-BuChE has also been successfully used to measure the changes of BuChE levels in the brains of AD mice. These findings imply that NFP-BuChE is a potentially useful molecular tool for screening possible natural BuChEI quickly and for monitoring changes in BuChE activity, and it is expected that more value will be explored in mice. In addition, matrine and its derivatives are promising options for future Alzheimer's disease treatments.

Dendrobium Officinale Kimura & Migo Attenuates High Cholesterol Diet-Induced Atherosclerosis

Objectives: This study aimed to investigate the potential of Dendrobium officinale Kimura & Migo (DOKM) in ameliorating high cholesterol-induced atherosclerosis (AS) in zebrafish, focusing on the development of safe and effective drugs with low toxicity for AS treatment. Methods: A zebrafish AS model was established by feeding zebrafish a high cholesterol diet (HCD). Treatment was administered using 1 mg/L and 10 mg/L DOKM water extract. Fluorescence microscopy was utilised to observe the effects of DOKM on HCD-induced cholesterol accumulation, neutrophil accumulation, and macrophage infiltration in the vasculature. The impact of DOKM on HCD-induced disorders of lipid metabolism was assessed using Nile red staining. Total cholesterol (TC), triglycerides (TG), and low-density lipoprotein (LDL-C) levels in zebrafish were measured using kits. The expression of pcsk9, srebf2, and hmgcr genes in zebrafish was determined using RT-PCR. HE staining and Elvitegravir (EVG) staining were employed to assess the effect of DOKM on plaque formation in the blood vessels of AS zebrafish. Results: A total of 145 compounds were collected from Dendrobium officinale, along with 680 corresponding targets. Additionally, 1583 atherosclerosis-related targets were identified, with 202 representing interaction targets between Dendrobium officinale and atherosclerosis-related targets. Key compounds identified included Gigantol, Chrysotobibenzyl, Naringenin Chalcone, Hesperetin, and Tangeretin. The core targets of Dendrobium officinale for atherosclerosis treatment were STAT3, PTPN11, JAK2, epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor alpha (PDGFRA), proto-oncogene tyrosine-protein kinase Src (SRC), STAT1, TP53, ESR1, and AKT1. GO functional enrichment analysis revealed a total of 2090 GO biological functional entries. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed 200 AS signalling pathways associated with DOKM treatment for atherosclerosis. DOKM significantly attenuated cholesterol accumulation, neutrophil aggregation, and macrophage infiltration in the blood vessels of AS zebrafish, ultimately leading to reduced plaque formation. AS zebrafish exhibited lipid metabolism disorders, characterised by significant increases in TC, TG, and LDL-C levels, as well as mRNA expression of pcsk9, srebf2, and hmgcr genes. Treatment with DOKM significantly improved these lipid metabolism disorders. Conclusion: DOKM effectively attenuates HCD-induced AS, and this therapeutic effect on AS may be related to its role in improving lipid metabolism dysfunction.

Cross-species evidence for a developmental origin of adult hypersomnia with loss of synaptic adhesion molecules beat-Ia/CADM2.

Idiopathic hypersomnia (IH) is a poorly-understood sleep disorder characterized by excessive daytime sleepiness despite normal nighttime sleep. Combining human genomics with behavioral and mechanistic studies in fish and flies, we uncover a role for beat-Ia/CADM2 , synaptic adhesion molecules of the immunoglobulin superfamily, in excessive sleepiness. Neuronal knockdown of Drosophila beat-Ia results in sleepy flies and loss of the vertebrate ortholog of beat-Ia , CADM2 , results in sleepy fish. We delineate a developmental function for beat-Ia in synaptic elaboration of neuropeptide F (NPF) neurites projecting to the suboesophageal zone (SEZ) of the fly brain. Brain connectome and experimental evidence demonstrate these NPF outputs synapse onto a subpopulation of SEZ GABAergic neurons to stabilize arousal. NPF is the Drosophila homolog of vertebrate neuropeptide Y (NPY), and an NPY receptor agonist restores sleep to normal levels in zebrafish lacking CADM2 . These findings point towards NPY modulation as a treatment target for human hypersomnia.

Multi-faceted potential of sophoridine compound's anti-arrhythmic and antioxidant effects through ROS/CaMKII pathway

Cardiac arrhythmias remain a significant cause of mortality and morbidity, for novel antiarrhythmic therapies. This study states that the first report of sophoridine (SPN), a quinolizidine alkaloid derived from traditional Chinese herbs, shows promise as a potential candidate due to its anti-arrhythmic and antioxidant properties. The study found that cell viability in H9C2 rat cardiomyocytes remained stable even when treated with SPN at a higher dosage of 100 μg/ml. This phenomenon was accompanied by increases in mitochondria-derived reactive oxygen species (ROS) and calcium/calmodulin-dependent protein kinase II (CaMKII) signaling, at 50 and 100 μg/ml. Glucose fluctuations regulate ventricular arrhythmias caused by SPN by activating the ROS/CaMKII pathway. Experimental models using zebrafish provided additional evidence supporting the regulatory effects of SPN on heart rate. In addition, the administration of SPN resulted in substantial deregulation of crucial genes involved in heart development (nppa, nppb, tnnt2a) at the transcriptional level in zebrafish. These findings provide insight into the various pharmacological properties of SPN and this opens up new possibilities for anti-arrhythmic treatment strategies.

Hypolipidemic effect of polysaccharide from Sargassum fusiforme and its ultrasonic degraded polysaccharide on zebrafish fed high-fat diet

Sargassum fusiforme is a brown seaweed that grows abundantly along the rocky coastlines of Asian countries. The polysaccharides derived from Sargassum fusiforme (SFPS) have received much interest due to their various bioactivities, such as hypolipidemic, hypoglycemic, and antioxidant activities. In this study, we extracted and purified SFPS, and obtained the ultrasonic degradation product (SFPSUD). The lipid regulatory effects of SFPS and SFPSUD were investigated in a zebrafish model fed a high-fat diet. The results showed that SFPS significantly decreased the levels of total cholesterol (TC) and triglycerides (TG), and increased the activities of lipoprotein lipase (LPL) and hepatic lipase (HL). SFPSUD was more effective than the SFPS in reducing the TC and TG levels in zebrafish, as well as increasing the LPL and HL activities. Histopathological observations of zebrafish livers showed that SFPSUD significantly improved lipid metabolism disorder in the hepatocytes. The possible lipid-lowering mechanism in zebrafish associated with SFPS and SFPSUD may involve acceleration of the lipid metabolism rate by increasing the activities of LPL and HL. Thus, SFPSUD could be tested as a highly effective hypolipidemic drug. Our results suggest that SFPS and SFPSUD have potential uses as functional foods for the prevention and treatment of hyperlipidemia. Ultrasound can be effectively applied to degrade SFPS to improve its physicochemical properties and bioactivities.

Antibacterial and gut health effects of Amomum tsao-ko in aquatic feed: A sustainable alternative to chemical antibiotics.

Amomum tsao-ko Crevost et Lemarie (Zingiberaceae), an aromatic plant, has been considered to have diverse medicinal values and economic significance. It has been reported to possess antibacterial, antioxidant, and antidiabetic effects. With the increasing risk of diseases in aquaculture, there is a need for alternative solutions to chemical antibiotics. Plant extracts have shown promise as natural feed additives for aquatic animals. In this study, the antibacterial effect of Amomum tsao-ko crude extracts was evaluated using the Oxford cup method. The extracts exhibited significant antimicrobial activity against Salmonella typhimurium and Salmonella enteritidis. Furthermore, the addition of Amomum tsao-ko to fish feed resulted in notable changes in the gut structure of zebrafish and tilapia. The length and morphology of intestinal villi were enhanced, promoting improved digestion. Analysis of the gut microbial community revealed that Amomum tsao-ko supplementation induced key changes in the gut microbial community composition of both zebrafish and tilapia. Notably, a 1% inclusion of Amomum tsao-ko resulted in a marked rise in Proteobacteria levels in zebrafish, which diminished at 10% dosage. The supplement elicited mixed reactions among other bacterial phyla like Actinobacteria and Verrucomicrobiota. Fluctuations were also observed at the genus level, pointing to the concentration of Amomum tsao-ko playing a pivotal role in influencing the structure of intestinal bacteria. The findings of this study suggest that Amomum tsao-ko has antibacterial properties and can positively influence the gut health of fish. The potential use of Amomum tsao-ko as a natural feed additive holds promise for improving aquaculture practices and reducing reliance on chemical antibiotics. Further research is needed to explore the full potential and applications of Amomum tsao-ko in fish feed development.

The RGK protein rrad regulates cardiac calcium levels in zebrafish

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Ministry of Science, Innovation and Universities, Spain Background/Introduction High voltage-activated Ca2+ (CaV) channels are essential for excitation-contraction coupling. RGK proteins (Gem, Rad, Rem, and Rem2) are small Ras-like G proteins that bind CaV β-subunits and profoundly inhibit CaV1/CaV2 channels. A recent study showed that Rad inhibits CaV1.2 channels in cardiomyocytes, thus mediating the enhanced inotropy and chronotropy by β-adrenergic agonists during the fight-or-flight response. The zebrafish has emerged as a useful in vivo animal model in cardiovascular physiology and pathophysiology. The role of zebrafish rrad, the orthologe of the human RAD, in the regulation of cardiac Ca2+ homeostasis in zebrafish has not been investigated. Purpose The aim of this study was to investigate the role of rrad in cardiac Ca2+ regulation in zebrafish. Methods We evaluated the cardiac Ca2+ dynamics, heart rhythm, and hemodynamics in transgenic zebrafish larvae that overexpress rrad in the heart, as well as in heterozygous rrad knockout larvae. Measurements were done in the beating heart of 3 dpf transgenic zebrafish larvae expressing the fluorescent Ca2+ biosensor Twitch-4. Results In some larvae, overexpression of rrad induced a severe enlargement of the atrium with pericardial edema and a high mortality rate. In addition, the ventricle displayed low Ca2+ levels and reduced contractility. The remaining larvae had no altered morphology, but the ventricular Ca2+ levels were reduced. On the other hand, preliminary results showed that down regulation of rrad in zebrafish larvae increased atrial and ventricular Ca2+ levels, like what was found in Rad knockout mice. Conclusion Taken together, these findings suggest that rrad in the zebrafish heart, as in mammals, is an inhibitor of the Ca2+ entry, presumably through CaV1/CaV2 channels, and demonstrate the zebrafish's utility as a model to study the involvement of rrad in cardiac function.

On-Site Quantitative Visualization of Singlet Oxygen in Crops via an Organic Small Molecule-Based Ratiometric Fluorescent Probe and a Mobile Fluorescence Analysis Device.

Singlet oxygen (1O2) plays imperative roles in a variety of biotic or abiotic stresses in crops. The change of its concentration within a crop is closely related to the crop growth and development. Accordingly, there is an urgent need to develop an efficient analytical method for on-site quantitative detection of 1O2 in crops. Here, we judiciously constructed a novel ratiometric fluorescent probe, SX-2, for the detection of 1O2 in crops. Upon treating with 1O2, probe SX-2 displayed highly selective ratiometric fluorescence response, which is favorable for the quantitative detection of 1O2. Concurrently, the fluorescence solution color of probe SX-2 was varied, obviously from blue to yellow, indicating that the probe is beneficial for on-site detection by the naked eye. Sensing reaction mechanism studies showed that the 2,3-diphenyl imidazole group in SX-2 could function as a new selective recognition group for 1O2. Probe SX-2 was utilized for the detection of photoirradiation-induced 1O2 and endogenous 1O2 in living cells. The changes in the 1O2 level in zebrafish were also tracked by fluorescence imaging. In addition, the production of 1O2 in crop leaves under a light source of different wavelengths was studied. The results demonstrated more 1O2 were produced under a light source of 365 nm. Furthermore, to achieve on-site quantitative detection, a mobile fluorescence analysis device has been made. Probe SX-2 and mobile fluorescence analysis device were capable of on-site quantitative detecting of 1O2 in crops. The method developed herein will be convenient for the on-site quantitative measurement of 1O2 in distinct crops.

Threonine dehydrogenase regulates neutrophil homeostasis but not H3K4me3 levels in zebrafish.

l-threonine dehydrogenase (Tdh) is an enzyme that links threonine metabolism to epigenetic modifications and mitochondria biogenesis. In vitro studies show that it is critical for the regulation of trimethylation of histone H3 lysine 4 (H3K4me3) levels and cell fate determination of mouse embryonic stem cells (mESCs). However, whether Tdh regulates a developmental process in vivo and, if it does, whether it also primarily regulates H3K4me3 levels in this process as it does in mESCs, remains elusive. Here, we revealed that, in zebrafish hematopoiesis, tdh is preferentially expressed in neutrophils. Knockout of tdh causes a decrease in neutrophil number and slightly suppresses their acute injury-induced migration, but, unlike the mESCs, the level of H3K4me3 is not evidently reduced in neutrophils sorted from the kidney marrow of adult tdh-null zebrafish. These phenotypes are dependent on the enzymatic activity of Tdh. Importantly, a soluble supplement of nutrients that are able to fuel the acetyl-CoA pool, such as pyruvate, glucose and branched-chain amino acids, is sufficient to rescue the reduction in neutrophils caused by tdh deletion. In summary, our study presents evidence for the functional requirement of Tdh-mediated threonine metabolism in a developmental process in vivo. It also provides an animal model for investigating the nutritional regulation of myelopoiesis and immune response, as well as a useful tool for high-throughput drug/nutrition screening.

Xanthohumol, a prenylated chalcone, regulates lipid metabolism by modulating the LXRα/RXR-ANGPTL3-LPL axis in hepatic cell lines and high-fat diet-fed zebrafish models

Angiopoietin-like 3 (ANGPTL3) acts as an inhibitor of lipoprotein lipase (LPL), impeding the breakdown of triglyceride-rich lipoproteins (TGRLs) in circulation. Targeting ANGPTL3 is considered a novel strategy for improving dyslipidemia and atherosclerotic cardiovascular diseases (ASCVD). Hops (Humulus lupulus L.) contain several bioactive prenylflavonoids, including xanthohumol (Xan), isoxanthohumol (Isoxan), 6-prenylnaringenin (6-PN), and 8-prenylnaringenin (8-PN), with the potential to manage lipid metabolism. The aim of this study was to investigate the lipid-lowering effects of Xan, the effective prenylated chalcone in attenuating ANGPTL3 transcriptional activity, both in vitro using hepatic cells and in vivo using zebrafish models, along with exploring the underlying mechanisms. Xan (10 and 20 μM) significantly reduced ANGPTL3 mRNA and protein expression in HepG2 and Huh7 cells, leading to a marked decrease in secreted ANGPTL3 proteins via hepatic cells. In animal studies, orally administered Xan significantly alleviated plasma triglyceride (TG) and cholesterol levels in zebrafish fed a high-fat diet. Furthermore, it reduced hepatic ANGPTL3 protein levels and increased LPL activity in zebrafish models, indicating its potential to modulate lipid profiles in circulation. Furthermore, molecular docking results predicted that Xan exhibits a higher binding affinity to interact with liver X receptor α (LXRα) and retinoic acid X receptor (RXR) than their respective agonists, T0901317 and 9-Cis-retinoic acid (9-Cis-RA). We observed that Xan suppressed hepatic ANGPTL3 expression by antagonizing the LXRα/RXR-mediated transcription. These findings suggest that Xan ameliorates dyslipidemia by modulating the LXRα/RXR-ANGPTL3-LPL axis. Xan represents a novel potential inhibitor of ANGPTL3 for the prevention or treatment of ASCVD.

Effects of polystyrene nano- and microplastics and of microplastics with sorbed polycyclic aromatic hydrocarbons in adult zebrafish

The presence of nanoplastics (NPs) and microplastics (MPs) in the environment is recognised as a global-scale problem. Due to their hydrophobic nature and large specific surface, NPs and MPs can adsorb other contaminants, as polycyclic aromatic hydrocarbons (PAHs), and modulate their bioavailability and hazard. Adult zebrafish were exposed for 3 and 21 days to: (1) 0.07 mg/L NPs (50 nm), (2) 0.05 mg/L MPs (4.5 μm), (3) MPs with sorbed oil compounds of the water accommodated fraction (WAF) of a naphthenic crude oil (MPs-WAF), (4) MPs with sorbed benzo(a)pyrene (MPs-B(a)P), (5) 5 % WAF and (6) 21 μg/L B(a)P. Electrodense particles resembling NPs were seen in the intestine lumen close to microvilli. MPs were abundantly found in the intestine lumen, but not internalised into the tissues. After 21 days, NPs caused a significant downregulation of cat, and upregulation of gpx1a and sod1, while MPs upregulated cyp1a and increased the prevalence of liver vacuolisation. No histopathological alteration was observed in gills. In this study, contaminated MPs did not increase PAH levels in zebrafish but results highlight the potential differential impact of plastic particles depending on their size, making it necessary to urgently address the ecotoxicological impact of real environmental NPs and MPs.

Test of Antidiabetic Effect of Taro Leaf Extract (Colocasia esculenta L.) on Zebrafish (Danio rerio)

Diabetes mellitus (DM) is a common metabolic disorder defined as chronic hyperglycemia. In addition to the symptoms associated with hyperglycemia itself such as thirst, polyuria and weight loss, it can also cause acute hyperglycemia emergencies that are potentially life threatening. One of the traditional plants that has potential as an antidiabetic drug is the taro plant (Colocasia esculenta L) because it contains chemical compounds such as alkaloids, flavonoids, saponins, tannins and polyphenols which are known to have antidiabetic effects. This study aims to determine the effectiveness and what dose of Taro Leaf Extract gives the best effect of reducing blood glucose levels in zebrafish. This study used zebrafish (Danio rerio) induced with alloxan and glucose to raise blood glucose levels. The 20 test animals used were divided into 6 groups, namely group 1 without treatment (normal), group 2 control (-) alloxan induction 0.1% + glucose 1%, group 3 control (+) metformin, group 4 taro leaf extract 200 mg, group 5 taro leaf extract 300 mg, group 6 taro leaf extract 400 mg. Then glucose levels were measured using a glucometer. Data analysis was carried out statistical tests. The results showed that a dose of 400 mg/2L had the ability to reduce blood glucose levels that were not significantly different from normal zebrafish glucose levels.

Tissue-specific temperature dependence of RNA editing levels in zebrafish

BackgroundRNA editing by adenosine deaminase acting on RNA (ADAR) occurs in all metazoans and fulfils several functions. Here, we examined effects of acclimation temperature (27 °C, 18 °C,13 °C) on editing patterns in six tissues of zebrafish (Danio rerio).ResultsSites and total amounts of editing differed among tissues. Brain showed the highest levels, followed by gill and skin. In these highly edited tissues, decreases in temperatures led to large increases in total amounts of editing and changes in specific edited sites. Gene ontology analysis showed both similarities (e.g., endoplasmic reticulum stress response) and differences in editing among tissues. The majority of edited sites were in transcripts of transposable elements and the 3′UTR regions of protein coding genes. By experimental validation, translation efficiency was directly related to extent of editing of the 3′UTR region of an mRNA.ConclusionsRNA editing increases 3′UTR polymorphism and affects efficiency of translation. Such editing may lead to temperature-adaptive changes in the proteome through altering relative amounts of synthesis of different proteins.

Study on Carbohydrate Metabolism in Adult Zebrafish (Danio rerio).

Excessive carbohydrate intake leads to metabolic disorders in fish. However, few literatures have reported the appropriate carbohydrate level for zebrafish, and the metabolic response to dietary carbohydrate remains largely unknown in zebrafish. This study assessed the responses of zebrafish and zebrafish liver cell line (ZFL) to different carbohydrate levels. In vivo results showed that ≥30% dietary dextrin levels significantly increased the plasma glucose content, activated the expression of hepatic glycolysis-related genes, and inhibited the expression of hepatic gluconeogenesis-related genes in zebrafish. Oil red O staining, triglyceride content, and Hematoxylin-Eosin staining results showed that dietary dextrin levels of ≥30% significantly increased lipid accumulation and liver damage, as well as processes related to glycolipid metabolism and inflammation in zebrafish. In ZFL, the transcription factor sterol regulatory element binding protein-1c signal intensity, 4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene (BODIPY 493/503) signal intensity, and triglyceride content were also significantly increased when incubated in high glucose, along with abnormal glycolipid metabolism and increased inflammation-related genes. In conclusion, we demonstrated that the maximum dietary carbohydrate level in adult zebrafish should be less than 30%. Excess dietary carbohydrates (30%-50%) caused hepatic steatosis and damage to zebrafish, similar to that seen in aquaculture species. Thus, this study assessed responses to different carbohydrate levels in zebrafish and illustrated that zebrafish is an optimal model for investigating glucose metabolism in some aquatic animals.

A near-infrared fluorescent probe for tracking endogenous and exogenous H2O2 in cells and zebrafish

H2O2 is an important signaling molecule in the body, and its levels fluctuate in many pathological sites, therefore, it can be used as a biomarker for early diagnosis of disease. Since the environment in vivo is extremely complex, it is of great significance to develop a probe that can accurately monitor the fluctuation of H2O2 level without interference from other physiological processes. Based on this, we designed and synthesized two new near-infrared H2O2 fluorescent probes, LTA and LTQ, based on the ICT mechanism. Both of them have good responses to H2O2, but LTA has a faster response speed. In addition, the probe LTA has good biocompatibility, good water solubility, and a large Stokes shift (95 nm). The detection limit is 4.525 μM. The probe was successfully used to visually detect H2O2 in living cells and zebrafish and was successfully used to monitor the changes in H2O2 levels in zebrafish due to APAP-induced liver injury.

Comparative analysis of toxicity and metabolomic profiling of rac-glufosinate and L-glufosinate in zebrafish

Glufosinate is a chiral pesticide, with commercial formulations such as racemic glufosinate (rac-glufosinate) and pure L-glufosinate enantiomer (L-glufosinate) on the market. There has been little research on the difference in toxicity to non-target organisms between these two main ingredients. The effects of rac-glufosinate and L-glufosinate on glutathione reductase (GR), catalase (CAT), superoxide dismutase (SOD), and malondialdehyde (MDA) levels in zebrafish were investigated in this study. The effect of two glufosinate agents at low concentrations (0.01 and 0.1 mg/L) on these four oxidative indicators was found to be significantly lower than that of high concentrations (1 and 10 mg/L). L-glufosinate had a stronger enhancing effect on CAT, GR, and MDA content than rac-glufosinate and a stronger inhibitory effect on SOD activity than rac-glufosinate. The researchers used ultra-high-performance liquid chromatography coupled with high-resolution mass spectroscopy metabolomics to compare rac-glufosinate and L-glufosinate for metabolic disorders in adult zebrafish. Stable and obvious metabolic maps of the two agents were obtained using multivariate statistical results, such as principal component analysis and orthogonal partial minimum discriminant analysis. Compared to the control group, the rac-glufosinate and L-glufosinate treatment groups shared 151 differential metabolites, which primarily affected zebrafish energy metabolism, amino acid metabolism, and other metabolic pathways. Caffeine metabolism and biotin metabolism were among the unique pathways disrupted in rac-glufosinate-exposed zebrafish. Contrarily, L-glufosinate treatment primarily affected eight metabolic pathways, including arginine biosynthesis, melanogenesis, and glutathione metabolism. These findings may provide more detailed information on the toxicity of rac-glufosinate and L-glufosinate in zebrafish, as well as some context for assessing the environmental risk of the two glufosinate agents to aquatic organisms.

Toxic effects of triphenyltin on the development of zebrafish (Danio rerio) embryos

Triphenyltin (TPT) is known to be an environmental endocrine disruptor and has adverse effects on aquatic animals. In this study, zebrafish embryos were treated with three different concentrations (12.5, 25, 50 nmol/L) based on the LC50 value at 96 h post fertilization (96 hpf), after TPT exposure. The developmental phenotype and hatchability were observed and recorded. Reactive oxygen species (ROS) levels in zebrafish were detected at 72 hpf and 96 hpf using 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) as a probe. The number of neutrophils after exposure was observed using transgenic zebrafish Tg (lyz: DsRed). RNA-seq analysis was used to compare the gene expression changes in zebrafish embryos at 96 hpf in the control group and 50 nmol/L TPT exposure group. The data revealed that TPT caused a delay in hatching of zebrafish embryos in a time- and dose-dependent manner, as well as causing pericardial edema, spinal curvature and melanin reduction. ROS levels in embryos exposed to TPT increased, and the number of neutrophils increased after TPT exposure to Tg (lyz: DsRed) in transgenic zebrafish. RNA-seq results were also analyzed, and KEGG enrichment analysis showed that significant differential genes were enriched in the PPAR signaling pathway (P < 0.05), and the PPAR signaling pathway mainly affected genes related to lipid metabolism. The RNA-seq results were verified using real-time fluorescence quantitative PCR (RT-qPCR). Oil red O and Nile red staining showed increased lipid accumulation after TPT exposure. These findings suggest that TPT affects the development of zebrafish embryos even at relatively low concentrations.

Impaired retinoic acid signaling in cerebral cavernous malformations

The capillary-venous pathology cerebral cavernous malformation (CCM) is caused by loss of CCM1/Krev interaction trapped protein 1 (KRIT1), CCM2/MGC4607, or CCM3/PDCD10 in some endothelial cells. Mutations of CCM genes within the brain vasculature can lead to recurrent cerebral hemorrhages. Pharmacological treatment options are urgently needed when lesions are located in deeply-seated and in-operable regions of the central nervous system. Previous pharmacological suppression screens in disease models of CCM led to the discovery that treatment with retinoic acid improved CCM phenotypes. This finding raised a need to investigate the involvement of retinoic acid in CCM and test whether it has a curative effect in preclinical mouse models. Here, we show that components of the retinoic acid synthesis and degradation pathway are transcriptionally misregulated across disease models of CCM. We complemented this analysis by pharmacologically modifying retinoic acid levels in zebrafish and human endothelial cell models of CCM, and in acute and chronic mouse models of CCM. Our pharmacological intervention studies in CCM2-depleted human umbilical vein endothelial cells (HUVECs) and krit1 mutant zebrafish showed positive effects when retinoic acid levels were increased. However, therapeutic approaches to prevent the development of vascular lesions in adult chronic murine models of CCM were drug regiment-sensitive, possibly due to adverse developmental effects of this hormone. A treatment with high doses of retinoic acid even worsened CCM lesions in an adult chronic murine model of CCM. This study provides evidence that retinoic acid signaling is impaired in the CCM pathophysiology and suggests that modification of retinoic acid levels can alleviate CCM phenotypes.

UV-aged microplastics induces neurotoxicity by affecting the neurotransmission in larval zebrafish

Microplastics (MPs) are nearly ubiquitous in aquatic ecosystems and may affect aquatic organisms. In this study, virgin and aged polystyrene MPs (PS-MPs) of size 1 μm were selected to analyze their adverse effects on larvae zebrafish. Exposure to PS-MPs significantly reduced the average swimming speed of zebrafish, and the behavioral effects caused by aged PS-MPs on zebrafish were more pronounced. Fluorescence microscopy revealed that 10–100 μg/L of PS-MPs accumulated in tissues of zebrafish. As an endpoint of neurotransmitter concentration, exposure to aged PS-MPs at doses ranging from 0.1 to 100 μg/L significantly increased the dopamine (DA), 5-hydroxytryptamine (5-HT), γ-aminobutyric acid (GABA), and acetylcholine (ACh) levels in zebrafish. Similarly, exposure to aged PS-MPs significantly altered the expression of genes related to these neurotransmitters (e.g., dat, 5ht1aa, and gabral genes). According to Pearson correlation analyses, neurotransmissions was significantly correlated with neurotoxic effects of aged PS-MPs. Thus, aged PS-MPs cause neurotoxicity in zebrafish through their effects on DA, 5-HT, GABA, and ACh neurotransmissions. The results highlight the importance of the neurotoxicity of aged PS-MPs in zebrafish, which has important implications for the risk assessment of aged MPs and the conservation of aquatic ecosystems.

The isl2a transcription factor regulates pituitary development in zebrafish.

ISL LIM homeobox 2, also known as insulin gene enhancer protein ISL-2 (ISL2), is a transcription factor gene that participates in a wide range of developmental events. However, the role of ISL2 in the hypothalamus-pituitary-thyroid axis is largely unknown. In the present study, we characterized the expression patterns of ISL2 and revealed its regulative role during embryogenesis using zebrafish. We used the CRISPR/Cas9 system to successfully establish homozygous ISL2-orthologue (isl2a and isl2b) knockout zebrafish. Moreover, we utilized these knockout zebrafish to analyze the pituitary and thyroid phenotypes in vivo. For further molecular characterization, in situ hybridization and immunofluorescence were performed. The isl2a mutant zebrafish presented with thyroid hypoplasia, reduced whole-body levels of thyroid hormones, increased early mortality, gender imbalance, and morphological retardation during maturity. Additionally, thyrotropes, a pituitary cell type, was notably decreased during development. Importantly, the transcriptional levels of pituitary-thyroid axis hormones-encoding genes, such as tshba, cga, and tg, were significantly decreased in isl2a mutants. Finally, the thyroid dysplasia in isl2a mutant larvae may be attributed to a reduction in proliferation rather than changes in apoptosis. In summary, isl2a regulates the transcriptional levels of marker genes in hypothalamus-pituitary-thyroid axis, and isl2a knockout causing low thyroid hormone levels in zebrafish. Thus, isl2a identified by the present study, is a novel regulator for pituitary cell differentiation in zebrafish, resulting in thyroid gland hypoplasia and phenotypes of hypothyroidism.