Zebrafish Embryos Research Articles

Roxithromycin exposure induces motoneuron malformation and behavioral deficits of zebrafish by interfering with the differentiation of motor neuron progenitor cells

Roxithromycin (ROX), a commonly used macrolide antibiotic, is extensively employed in human medicine and livestock industries. Due to its structural stability and resistance to biological degradation, ROX persists as a resilient environmental contaminant, detectable in aquatic ecosystems and food products. However, our understanding of the potential health risks to humans from continuous ROX exposure remains limited. In this study, we used the zebrafish as a vertebrate model to explore the potential developmental toxicity of early ROX exposure, particularly focusing on its effects on locomotor functionality and CaP motoneuron development. Early exposure to ROX induces marked developmental toxicity in zebrafish embryos, significantly reducing hatching rates (n=100), body lengths (n=100), and increased malformation rates (n=100). The zebrafish embryos treated with a corresponding volume of DMSO (0.1%, v/v) served as vehicle controls (veh). Moreover, ROX exposure adversely affected the locomotive capacity of zebrafish embryos, and observations in transgenic zebrafish Tg(hb9:eGFP) revealed axonal loss in motor neurons, evident through reduced or irregular axonal lengths (n=80). Concurrently, abnormal apoptosis in ROX-exposed zebrafish embryos intensified alongside the upregulation of apoptosis-related genes (bax, bcl2, caspase-3a). Single-cell sequencing further disclosed substantial effects of ROX on genes involved in the differentiation of motor neuron progenitor cells (ngn1, olig2), axon development (cd82a, mbpa, plp1b, sema5a), and neuroimmunity (aplnrb, aplnra) in zebrafish larvae (n=30). Furthermore, the CaP motor neuron defects and behavioral deficits induced by ROX can be rescued by administering ngn1 agonist (n=80). In summary, ROX exposure leads to early-life abnormalities in zebrafish motor neurons and locomotor behavior by hindering the differentiation of motor neuron progenitor cells and inducing abnormal apoptosis.

Shedding a Light on Dark Genes: A Comparative Expression Study of PRR12 Orthologues during Zebrafish Development

Haploinsufficiency of the PRR12 gene is implicated in a human neuro-ocular syndrome. Although identified as a nuclear protein highly expressed in the embryonic mouse brain, PRR12 molecular function remains elusive. This study explores the spatio-temporal expression of zebrafish PRR12 co-orthologs, prr12a and prr12b, as a first step to elucidate their function. In silico analysis reveals high evolutionary conservation in the DNA-interacting domains for both orthologs, with significant syntenic conservation observed for the prr12b locus. In situ hybridization and RT-qPCR analyses on zebrafish embryos and larvae reveal distinct expression patterns: prr12a is expressed early in zygotic development, mainly in the central nervous system, while prr12b expression initiates during gastrulation, localizing later to dopaminergic telencephalic and diencephalic cell clusters. Both transcripts are enriched in the ganglion cell and inner neural layers of the 72 hpf retina, with prr12b widely distributed in the ciliary marginal zone. In the adult brain, prr12a and prr12b are found in the cerebellum, amygdala and ventral telencephalon, which represent the main areas affected in autistic patients. Overall, this study suggests PRR12’s potential involvement in eye and brain development, laying the groundwork for further investigations into PRR12-related neurobehavioral disorders.

Nephroprotective effects of coriander (Coriandrum sativum) leaves aqueous extracts in aristolochic acid-intoxicated zebrafish embryos.

Coriander is a notable medicinal plant known for its diverse properties, including anti-inflammatory, antioxidant, anticancer, analgesic, and anti-diabetic effects. Despite its recognized health benefits, research on its nephroprotective properties is limited. This study aimed to investigate the potential nephroprotective properties of an aqueous extract derived from coriander leaves using an aristolochic acid-intoxicated zebrafish model. To assess kidney abnormalities induced by aristolochic acid (AA), we utilized the transgenic line Tg(wt1b:egfp), which expresses green fluorescent protein (GFP) in the kidney. Our previous report indicated that AA exposure leads to acute renal failure in zebrafish characterized by kidney malformation and impaired renal function. However, pretreatment of coriander extract (CE) can mitigate kidney malformations induced by AA. In addition, CE pretreatment reduces the accumulation of red blood cells in the glomerular region. To verify the nephroprotective effects of CE, we analyzed renal function by measuring the glomerular filtration rate in zebrafish embryos. Results indicate that CE partially mitigates renal function impairment caused by AA exposure, suggesting its potential to attenuate AA-induced renal failure. Mechanistically, pretreatment with CE reduces the expression of proinflammatory and proapoptotic genes induced by AA. This suggests that CE likely alleviates acute renal failure by reducing inflammation and apoptosis. As a result, we regard zebrafish as a valuable model for screening natural compounds that have the potential to alleviate AA-induced nephrotoxicity.

Aspartame Causes Developmental Defects and Teratogenicity in Zebra Fish Embryo: Role of Impaired SIRT1/FOXO3a Axis in Neuron Cells

Aspartame, a widely used artificial sweetener, is present in many food products and beverages worldwide. It has been linked to potential neurotoxicity and developmental defects. However, its teratogenic effect on embryonic development and the underlying potential mechanisms need to be elucidated. We investigated the concentration- and time-dependent effects of aspartame on zebrafish development and teratogenicity. We focused on the role of sirtuin 1 (SIRT1) and Forkhead-box transcription factor (FOXO), two proteins that play key roles in neurodevelopment. It was found that aspartame exposure reduced the formation of larvae and the development of cartilage in zebrafish. It also delayed post-fertilization development by altering the head length and locomotor behavior of zebrafish. RNA-sequencing-based DEG analysis showed that SIRT1 and FOXO3a are involved in neurodevelopment. In silico and in vitro analyses showed that aspartame could target and reduce the expression of SIRT1 and FOXO3a proteins in neuron cells. Additionally, aspartame triggered the reduction of autophagy flux by inhibiting the nuclear translocation of SIRT1 in neuronal cells. The findings suggest that aspartame can cause developmental defects and teratogenicity in zebrafish embryos and reduce autophagy by impairing the SIRT1/FOXO3a axis in neuron cells.

Teratogenic Effects of Di(2-Ethylhexyl) Phthalate (DEHP) on Zebrafish Embryos

Developmental toxicity studies help to understand the impact of a pollutant on this crucial phase of the living organisms which can affect their population dynamics. Zebrafish has become an ideal model for studying environmental and embryo toxicity. The present study was carried out using zebrafish embryos for assessing the environmental toxicity of Di(2-ethylhexyl) phthalate (DEHP) which is universally considered to be an omnipresent environmental contaminant as it is the most widely used plasticizer. The embryos were exposed to DEHP for a range of five concentrations of 0.2, 20, 80, 140 and 200 μg/L for the duration of 96 hours. The treatment resulted in increased mortality and decreased hatch rate, hatchability and heartrate. It also induced teratogenic endpoints like yolk sac edema, pericardial edema and spinal deformity in the embryos which increased in dose and time dependent manner.

Polydopamine-Mediated Metal-Organic Frameworks Modification for Improved Biocompatibility.

Engineered nanomaterials are promising in biomedical application. However, insufficient understanding of their biocompatibility at the cellular and organic levels prevents their widely biomedical applications. Metal-organic frameworks (MOFs) have attracted increasing attention in recent years. In this work, zeolitic imidazolate framework-8 (ZIF-8) and polydopamine (PDA)-modified ZIF-8 are chosen as model nanomaterials due to its emergent role in nanomedicine. In vitro, the results demonstrate that the PDA coating greatly alleviates the cytotoxicity of ZIF-8 to RAW264.7, LO2, and HST6, which represent three different cell types in liver organs. Mechanistically, ZIF-8 entering into the cells can greatly induce the reactive oxygen species generation, which subsequently induces cell cycle delay and autophagy, ultimately leads to enhanced cytotoxicity. Further, human umbilical vein endothelial cells model and zebrafish embryos assay also confirm that PDA can compromise the ZIF-8 toxicity significantly. This study reveals that PDA-coated MOFs nanomaterials show great potential in nano-based drug delivery systems .

Cell-type-specific mRNA transcription and degradation kinetics in zebrafish embryogenesis from metabolically labeled single-cell RNA-seq

During embryonic development, pluripotent cells assume specialized identities by adopting particular gene expression profiles. However, systematically dissecting the relative contributions of mRNA transcription and degradation to shaping those profiles remains challenging, especially within embryos with diverse cellular identities. Here, we combine single-cell RNA-Seq and metabolic labeling to capture temporal cellular transcriptomes of zebrafish embryos where newly-transcribed (zygotic) and pre-existing (maternal) mRNA can be distinguished. We introduce kinetic models to quantify mRNA transcription and degradation rates within individual cell types during their specification. These models reveal highly varied regulatory rates across thousands of genes, coordinated transcription and destruction rates for many transcripts, and link differences in degradation to specific sequence elements. They also identify cell-type-specific differences in degradation, namely selective retention of maternal transcripts within primordial germ cells and enveloping layer cells, two of the earliest specified cell types. Our study provides a quantitative approach to study mRNA regulation during a dynamic spatio-temporal response.

A Review on Unlocking Toxicity: How Zebrafish embryos Help Ensure Safe Chemicals

Aquatic ecotoxicity testing has benefited greatly from zebrafish embryo toxicity testing (ZFET) because of its high throughput, low cost, sensitivity to a variety of toxicants, and ethical issues. This study highlights the benefits of ZFET over conventional fish toxicity testing, such as its capacity to detect teratogenic defects and possible long-term impacts. The limitations of ZFET are also addressed, notably its emphasis on acute dosages and difficulties with interspecies extrapolation. In addition to the generation of in vitro alternatives utilizing zebrafish cell lines, recent developments in ZFET technology are also highlighted, including the use of transgenic zebrafish lines and high-throughput screening techniques. The paper's conclusion includes a discussion of potential prospects for ZFET research. These include improving interspecies extrapolation methods, optimizing chronic toxicity assessment procedures, and integrating ZFET with other ecotoxicity testing strategies. ZFET is a great instrument for environmental protection and a crucial component in avoiding chemical contamination of aquatic ecosystems, so long as it is acknowledged for what it is and is open to future improvement

Impacts of Differentially Shaped Silver Nanoparticles with Increasingly Complex Hydrophobic Thiol Surface Coatings in Small-Scale Laboratory Microcosms

We investigated the impacts of spherical and triangular-plate-shaped lipid-coated silver nanoparticles (AgNPs) designed to prevent surface oxidation and silver ion (Ag+) dissolution in a small-scale microcosm to examine the role of shape and surface functionalization on biological interactions. Exposures were conducted in microcosms consisting of algae, bacteria, crustaceans, and fish embryos. Each microcosm was exposed to one of five surface chemistries within each shape profile (at 0, 0.1, or 0.5 mg Ag/L) to investigate the role of shape and surface composition on organismal uptake and toxicity. The hybrid lipid-coated AgNPs did not result in any significant release of Ag+ and had the most significant toxicity to D. magna, the most sensitive species, although the bacterial population growth rate was reduced in all exposures. Despite AgNPs resulting in increasing algal growth over the experiment, we found no correlation between algal growth and the survival of D. magna, suggesting that the impacts of the AgNPs on bacterial survival influenced algal growth rates. No significant impacts on zebrafish embryos were noted in any exposure. Our results demonstrate that the size, shape, and surface chemistry of AgNPs can be engineered to achieve specific goals while mitigating nanoparticle risks.

Transcription bodies regulate gene expression by sequestering CDK9.

The localization of transcriptional activity in specialized transcription bodies is a hallmark of gene expression in eukaryotic cells. It remains unclear, however, if and how transcription bodies affect gene expression. Here we disrupted the formation of two prominent endogenous transcription bodies that mark the onset of zygotic transcription in zebrafish embryos and analysed the effect on gene expression using enriched SLAM-seq and live-cell imaging. We find that the disruption of transcription bodies results in the misregulation of hundreds of genes. Here we focus on genes that are upregulated. These genes have accessible chromatin and are poised to be transcribed in the presence of the two transcription bodies, but they do not go into elongation. Live-cell imaging shows that disruption of the two large transcription bodies enables these poised genes to be transcribed in ectopic transcription bodies, suggesting that the large transcription bodies sequester a pause release factor. Supporting this hypothesis, we find that CDK9-the kinase that releases paused polymerase II-is highly enriched in the two large transcription bodies. Overexpression of CDK9 in wild-type embryos results in the formation of ectopic transcription bodies and thus phenocopies the removal of the two large transcription bodies. Taken together, our results show that transcription bodies regulate transcription by sequestering machinery, thereby preventing genes elsewhere in the nucleus from being transcribed.

High NaCl Concentrations in Water Are Associated with Developmental Abnormalities and Altered Gene Expression in Zebrafish.

Salt is frequently introduced in ecosystems, where it acts as a pollutant. This study examined how changes in salinity affect the survival and development of zebrafish from the two-cell to the blastocyst stage and from the blastocyst to the larval stage. Control zebrafish embryos were cultured in E3 medium containing 5 mM Sodium Chloride (NaCl), 0.17 mM Potassium Chloride (KCL), 0.33 mM Calcium Chloride (CaCl2), and 0.33 mM Magnesium Sulfade (MgSO4). Experiments were conducted using increasing concentrations of each individual salt at 5×, 10×, 50×, and 100× the concentration found in E3 medium. KCL, CaCl2, and MgSO4 did not result in lethal abnormalities and did not affect early embryo growth at any of the concentrations tested. Concentrations of 50× and 100× NaCl caused embryonic death in both stages of development. Concentrations of 5× and 10× NaCl resulted in uninflated swim bladders in 12% and 65% of larvae, compared to 4.2% of controls, and caused 1654 and 2628 genes to be differentially expressed in blastocysts, respectively. The ATM signaling pathway was affected, and the Sonic Hedgehog pathway genes Shh and Ptc1 implicated in swim bladder development were downregulated. Our findings suggest that increased NaCl concentrations may alter gene expression and cause developmental abnormalities in animals found in affected ecosystems.

Isobavachalcone induces hepatotoxicity in zebrafish embryos and HepG2 cells via the System Xc--GSH-GPX4 signaling pathway in ferroptosis response.

Isobavachalcone (IBC) is a flavonoid component of the traditional Chinese medicine Psoraleae Fructus, with a range of pharmacological properties. However, IBC causes some hepatotoxicity, and the mechanism of toxicity is unclear. The purpose of this paper was to investigate the possible mechanism of toxicity of IBC on HepG2 cells and zebrafish embryos. The results showed that exposure to IBC increased zebrafish embryo mortality and decreased hatchability. Meanwhile, IBC induced liver injury and increased expression of ALT and AST activity. Further studies showed that IBC caused the increase of ROS and MDA the decrease of CAT, GSH, and GSH-Px; the increase of Fe2+ content; and the changes of ferroptosis related genes (acsl4, gpx4, and xct) and iron storage related genes (tf, fth, and fpn) in zebrafish embryos. Through in vitro verification, it was found that IBC also caused oxidative stress and increased Fe2+ content in HepG2 cells. IBC caused depolarization of mitochondrial membrane potential (MMP) and reduction of mitochondrial ATP, as well as altered expression of ACSl4, SLC7A11, GPX4, and FTH1 proteins. Treatment of HepG2 cells with ferrostatin-1 could reverse the effect of IBC. Targeting the System Xc--GSH-GPX4 pathway of ferroptosis and preventing oxidative stress damage might offer a theoretical foundation for practical therapy and prevention of IBC-induced hepatotoxicity.

Single-Step Low-Temperature Synthesis of Carbon Dots for Advanced Multiparametric Bioimaging Probe Applications.

Carbon dots (CDs) have recently emerged in biomedical and agricultural domains, mainly for their probe applications in developing efficient sensors. However, the existing high-temperature approaches limit the industrial level scaling up to further translate them into different products by mass-scale fabrication of CDs. To address this, we have attempted to lower the synthesis temperature to 140 °C and synthesized different CDs using different organic acids and their combinations in a one-step approach (quantum yield 3.6% to 16.5%; average size 3 to 5 nm). Further, sensing applications of CDs have been explored in three different biological models, mainly Danio rerio (zebrafish) embryos, bacterial strains, and the Lactuca sativa (lettuce) plant. The 72 h exposure of D. rerio embryos to 0.5 and 1 mg/mL concentrations of CDs exhibited significant uptake without mortality, a 100% hatching rate, and nonsignificant alterations in heart rate. Bacterial bioimaging experiments revealed CD compatibility with Gram-positive (Bacillus subtilis) and Gram-negative (Serratia marcescens) strains without bactericidal effects. Furthermore, CDs demonstrated effective conduction and fluorescence within the vascular system of lettuce plants, indicating their potential as in vivo probes for plant tissues. The single-step low-temperature CD synthesis approach with efficient structural and optical properties enables the process as industrially viable to up-scale the technology readiness level. The bioimaging of CDs in different biological models indicates the possibility of developing a CD probe for diverse biosensing roles in diseases, metabolism, microbial contamination sensing, and more.

Col1a2+ fibroblasts/muscle progenitors finetune xanthophore countershading by differentially expressing csf1a/1b in embryonic zebrafish.

Animals evolve diverse pigment patterns to adapt to the natural environment. Countershading, characterized by a dark-colored dorsum and a light-colored ventrum, is one of the most prevalent pigment patterns observed in vertebrates. In this study, we reveal a mechanism regulating xanthophore countershading in zebrafish embryos. We found that csf1a and csf1b mutants altered xanthophore countershading differently: csf1a mutants lack ventral xanthophores, while csf1b mutants have reduced dorsal xanthophores. Further study revealed that csf1a is expressed throughout the trunk, whereas csf1b is expressed dorsally. Ectopic expression of csf1a or csf1b in neurons attracted xanthophores into the spinal cord. Blocking csf1 signaling by csf1ra mutants disrupts spinal cord distribution and normal xanthophores countershading. Single-cell RNA sequencing identified two col1a2+ populations: csf1ahighcsf1bhigh muscle progenitors and csf1ahighcsf1blow fibroblast progenitors. Ablation of col1a2+ fibroblast and muscle progenitors abolished xanthophore patterns. Our study suggests that fibroblast and muscle progenitors differentially express csf1a and csf1b to modulate xanthophore patterning, providing insights into the mechanism of countershading.

Black soldier fly bioconversion to cultivated meat media components using blue catfish gut microbiome

This study aimed to develop low-cost media for cell-cultivated seafood using gut-microbial community-assisted fermentation. Black soldier fly larvae (Hermetia illucens) were used as the substrate and exposed to gut microbial communities isolated from blue catfish (Ictalurus furcatus). Fermentation led to a decrease in species richness, with the loss of important chitinase and protease-producing genera such as Pseudomonas and Clostridiaceae. However, there was an increase in Paraclostridium and members of the Enterobacteriaceae. In addition, the effect of fermented hydrolysates from BSFL on the proliferation of zebrafish embryo fibroblasts was tested in comparison to fetal bovine serum (FBS) in in vitro cell cultivation. Lower concentrations of FBS resulted in decreased cell density and altered cell morphology. The supplementation of hydrolysate B at high peptide concentrations had cytotoxic effects on the cells, while at lower peptide concentrations, it improved cell proliferation only in cultures with 2.5 % FBS.

Bifenox induces hepatotoxicity and vascular toxicity in zebrafish embryos via ROS production and alterations in signaling pathways

Existing evidence shows that currently used pesticides pose toxicological risks to exposed wildlife. Chemically, bifenox belongs to diphenyl ethers, a well-known group of herbicides. Its mechanism of action primarily involves inducing lipid peroxidation and blocking protoporphyrinogen oxidases. Toxicity of diphenyl ether herbicides has been elucidated in animal cells; however, in vivo toxicological evaluations of bifenox are required to determine its unexpected effects. This study aimed to determine the negative effects of bifenox, and its effects on higher eukaryotes. We found that early stages of zebrafish embryo exposed to bifenox demonstrated increased mortality and physiological defects, based on the LC50 value. Bifenox severely inhibited blood vessel growth by reducing key elements of complex connectivity; fluorescently tagged transgenic lines (fli1a:EGFP) showed morphological changes. Additionally, transgenic lines that selectively identified hepatocytes (fabp10a:DsRed) showed reduced fluorescence, indicating that bifenox may inhibit liver development. To evaluate the level of oxidative stress, we used 2′,7′-dichlorofluorescein diacetate (DCFH-DA) probes in zebrafish embryos to identify the underlying mechanisms causing developmental damage. Our findings demonstrate that exposure to bifenox causes abnormalities in the hepatic and cardiovascular systems during zebrafish embryogenesis. Therefore, this study provides new information for the evaluation of toxicological risks of bifenox in vertebrates.

Bringing Real Inquiry-Based Science to Diverse Secondary Educational Environments: A Virtual Zebrafish Laboratory to Investigate Environmental Health.

The goal of the University of Wisconsin-Milwaukee WInSTEP SEPA program is to provide valuable and relevant research experiences to students and instructors in diverse secondary educational settings. Introducing an online experience allows the expansion of a proven instructional research program to a national scale and removes many common barriers. These can include lack of access to zebrafish embryos, laboratory equipment, and modern classroom facilities, which often deny disadvantaged and underrepresented students from urban and rural school districts valuable inquiry-based learning opportunities. An online repository of zebrafish embryo imagery was developed in the Carvan laboratory to assess the effects of environmental chemicals. The WInSTEP SEPA program expanded its use as an accessible online tool, complementing the existing classroom experience of our zebrafish module. This virtual laboratory environment contains images of zebrafish embryos grown in the presence of environmental toxicants (ethanol, caffeine, and nicotine), allowing students to collect data on 19 anatomical endpoints and generate significant amounts of data related to developmental toxicology and environmental health. This virtual laboratory offers students and instructors the choice of data sets that differ in the independent variables of chemical concentration and duration of postfertilization exposure. This enables students considerable flexibility in establishing their own experimental design to match the curriculum needs of each instructor.

Tagging the tjp1a Gene in Zebrafish with Monomeric Red Fluorescent Protein Using Biotin Homology Arms.

Tjp1a and other tight junction and adherens proteins play important roles in cell-cell adhesion, scaffolding, and forming seals between cells in epithelial and endothelial tissues. In this study, we labeled Tjp1a of zebrafish with the monomeric red fluorescent protein (mRFP) using CRISPR/Cas9-mediated targeted integration of biotin-labeled polymerase chain reaction (PCR) generated templates. Labeling Tjp1a with RFP allowed us to follow membrane and junctional dynamics of epithelial and endothelial cells throughout zebrafish embryo development. For targeted integration, we used short 35 bp homology arms on each side of the Cas9 genomic target site at the C-terminal of the coding sequence in tjp1a. Through PCR using 5' biotinylated primers containing the homology arms, we generated a double-stranded template for homology directed repair containing a flexible linker followed by RFP. Cas9 protein was complexed with the tjp1a gRNA before mixing with the repair template and microinjected into one-cell zebrafish embryos. We confirmed and recovered a precise integration allele at the desired site at the tjp1a C-terminus. Examination of fluorescence reveals RFP cell-cell junctional labeling using confocal imaging. We are currently using this stable tjp1a-mRFPis86 line to examine the behavior and interactions between cells during vascular formation in zebrafish.

Exposure to acifluorfen induces developmental toxicity in the early life stage of zebrafish

Acifluorfen, a selective herbicide from the diphenyl ether family, targets broad leaf weeds. Diphenyl ether inhibits chlorophyll production in green plants by inhibiting protoporphyrinogen oxidase (PPO), causing cellular damage. Despite its known impacts on plants, the influence of acifluorfen on zebrafish embryo development remains unclear. In this study, we explored the LC50 of acifluorfen in early-stage wild-type zebrafish, determining it to be 54.99 mg/L. Subsequent examinations revealed morphological changes in zebrafish, including reduced body length. Using the cmlc2:dsRED transgenic model, we observed heart dysfunction in acifluorfen-exposed zebrafish, marked by an enlarged heart area, edema, and decreased heart rate. In response to dose-dependent acifluorfen exposure, the inhibition of angiogenesis in the brain was observed in transgenic zebrafish models (fli1a:eGFP). Organ malformations, specifically in the liver and pancreas, were noted, in lfabp:dsRED;elastase:eGFP transgenic models, indicating reduced organ size in acifluorfen-exposed zebrafish. Furthermore, acifluorfen heightened the expression of apoptosis-related genes (casp8, casp9, and tp53) in zebrafish embryos. We then determined whether acifluorfen affected the viability of zebrafish liver (ZFL) cells based on its effects on liver development in vivo. The results indicated that the proliferation of ZFL cells decreased significantly in a dose-dependent manner. Additionally, acifluorfen-treated ZFL cells exhibited a slight increase in apoptotic cells stained with annexin V and propidium iodide. In summary, these findings establish a baseline concentration for acifluorfen's effects on aquatic ecosystems and non-target organisms.

The combined effects of preservative chemicals in consumer products: an analysis using embryonic and adult zebrafish

Benzisothiazolinone (BIT) and propyl paraben (PP) are preservatives in cleaning products; however, their toxicities are not well understood. In this study, zebrafish embryos were exposed to BIT, PP, and mixtures of both for 96 h to investigate the effects on growth hormone (GH), insulin-like growth factor-1 (IGF-1), and the transcription of 19 genes related to the GH/IGFs axis. Concentrations of BIT and PP were measured in the whole body of larvae. Zebrafish pairs were also exposed to BIT, PP, and mixtures for 21 d to evaluate the effects on sex hormones, histology in gonad, and transcription of 22 genes related to the hypothalamus-pituitary-gonad axis and vitellogenin. The mixtures had potentiation effects on development, reproduction, hormones, and gene transcripts than individual exposure. Larvae exposed to 229 μg L-1 BIT, 64.5 μg L-1 PP, and mixtures showed reduced growth. Decreased GH and IGF-1 levels were supported by gene regulation associated with the GH/IGFs axis. In larvae, reactive oxygen species, superoxide dismutase, catalase, and glutathione peroxidase levels were increased under all exposures. The gonadosomatic index in males and number of eggs decreased after mixture exposure. In females exposed to mixtures, the percentage of atretic follicle in ovary was significantly increased. The significant decrease in testosterone in males and significant decrease in 17β-estradiol in females exposed to mixtures suggest anti-estrogenic and anti-androgenic potential. Thus, preservative mixtures in consumer products may be more toxic than the individual substances, which is important for managing the risks of mixing preservatives.