Single-cell profiling of intrahepatic immune cells reveals an expansion of tissue-resident cytotoxic CD4+ T lymphocyte subset associated with pathogenesis of alcoholic-associated liver diseases.

BACKGROUND & AIMS: The immunological mechanisms underpinning the pathogenesis of alcoholic-associated liver disease (ALD) remain incompletely elucidated. This study aims to explore the transcriptomic profiles of hepatic immune cells in ALD compared to healthy individuals and those with metabolic dysfunction-associated steatotic liver disease (MASLD). METHODS: We utilized single-cell RNA sequencing to analyze liver samples from healthy subjects, MASLD, and ALD patients, focusing on the immune cell landscapes within the liver. Key alterations in immune cell subsets were further validated using liver biopsy samples from additional patient cohorts. RESULTS: We observed a significant accumulation of CD4+ T cells in ALD patients' livers, surpassing the prevalence of CD8+ T cells, in contrast to MASLD and healthy counterparts, while natural killer (NK) cells and γδT cells exhibited reduced intrahepatic infiltration. In-depth transcriptional and developmental trajectory analyses unveiled that a distinct CD4+ subset characterized by granzyme K (GZMK) expression, displaying a tissue-resident signature and terminal effector state, prominently enriched among CD4+ T cells infiltrating the livers of ALD patient. Subsequent examination of an independent ALD patient cohort corroborated the substantial enrichment of GZMK+CD4+ T lymphocytes, primarily within liver fibrotic zones, suggesting their potential involvement in disease progression. Additionally, we noted shifts in myeloid populations, with expanded APOE+ macrophage and FCGR3B+ monocyte subsets in ALD samples relative to MASLD and healthy tissues. CONCLUSIONS: In summary, this study unravels the intricate cellular diversity within hepatic immune cell populations, highlighting the pivotal immune pathogenic role of the GZMK+CD4+ T lymphocyte subset in ALD pathogenesis.

Spatial Transcriptomic Study Reveals Heterogeneous Metabolic Adaptation and a Role of Pericentral PPARα/CAR/Ces2a Axis During Fasting in Mouse Liver.

Spatial heterogeneity and plasticity of the mammalian liver are critical for systemic metabolic homeostasis in response to fluctuating nutritional conditions. Here, a spatially resolved transcriptomic landscape of mouse livers across fed, fasted and refed states using spatial transcriptomics is generated. This approach elucidated dynamic temporal-spatial gene cascades and how liver zonation-both expression levels and patterns-adapts to shifts in nutritional status. Importantly, the pericentral nuclear receptor Nr1i3 (CAR) as a pivotal regulator of triglyceride metabolism is pinpointed. It is showed that the activation of CAR in the pericentral region is transcriptionally governed by Pparα. During fasting, CAR activation enhances lipolysis by upregulating carboxylesterase 2a, playing a crucial role in maintaining triglyceride homeostasis. These findings lay the foundation for future mechanistic studies of liver metabolic heterogeneity and plasticity in response to nutritional status changes, offering insights into the zonated pathology that emerge during liver disease progression linked to nutritional imbalances.

Fluorene-9-bisphenol acts on the gut-brain axis by regulating oxytocin signaling to disturb social behaviors in zebrafish.

Previous studies have identified the exposure to ubiquitous environmental endocrine disruptors may be a risk factor of neurological disorders. However, the effects of fluorene-9-bisphenol (BHPF) in environmental exposure concentrations associated with these disorders are poorly understood. In this study, classic light-dark and social behavior tests were performed on zebrafish larvae and adults exposed BHPF exposure to evaluate social behavioral disorders and the microbiota-gut-brain axis was assessed to reveal the potential mechanisms underlying the behavioral abnormalities observed. Our results demonstrated that zebrafish larvae exposed to an environmentally relevant concentration (0.1 nM) of BHPF for 7 days showed a diminished response to external environmental factors (light or dark). Zebrafish larvae exposed to BHPF for 7 days or adults exposed to BHPF for 30 days at 1 µM displayed significant behavioral inhibition and altered social behaviors, including social recognition, social preference, and social fear contagion, indicating autism-like behaviors were induced by the exposure. BHPF exposure reduced the distribution of Nissl bodies in midbrain neurons and significantly reduced 5-hydroxytryptamine signaling. Oxytocin (OXT) levels and expression of its receptor oxtra in the gut and brain were down-regulated by BHPF exposure. In addition, the expression levels of genes related to the excitation-inhibitory balance of synaptic transmission changed. Microbiomics revealed increased community diversity and altered abundance of some microflora, such as an elevation in Bacillota and Bacteroidota and a decline in Mycoplasmatota in zebrafish guts, which might contribute to the abnormal neural circuits and autism-like behaviors induced by BHPF. Finally, the rescue effect of exogenous OXT on social behavioral defects induced by BHPF exposure was verified in zebrafish, highlighting the crucial role of OXT signaling through gut-brain axis in the regulatory mechanisms of social behaviors affected by BHPF. This study contributes to understanding the effects of environmental BHPF exposure on neuropsychiatric disorders and attracts public attention to the health risks posed by chemicals in aquatic organisms. The potential mental disorders should be considered in the safety assessments of environmental pollutants.

Spatial transcriptomics reveals heterogeneity of macrophages in the tumor microenvironment of granulomatous slack skin.

Granulomatous slack skin (GSS) is an extremely rare subtype of cutaneous T-cell lymphoma accompanied by an abundant number of macrophages and is clinically characterized by the development of pendulous skin folds. However, the characteristics of these macrophages in GSS remain unclear. Here, we conducted a spatial transcriptomic study on one frozen GSS sample and drew transcriptomic maps of GSS for the first time. Gene expression analysis revealed the enrichment of three clusters with macrophage transcripts, each exhibiting distinct characteristics suggesting that their primary composition consists of different subpopulations of macrophages. The CD163+ /CD206+ cluster showed a tumor-associated macrophage (TAM) M2-like phenotype and highly expressed ZFP36, CCL2, TNFAIP6, and KLF2, which are known to be involved in T-cell interaction and tumor progression. The APOC1+ /APOE+ cluster presented a non-M1 or -M2 phenotype and may be related to lipid metabolism. The CD11c+ /LYZ+ cluster exhibited an M1-like phenotype. Notably, these cells strongly expressed MMP9, MMP12, CHI3L1, CHIT1, COL1A1, TIMP1, and SPP1, which are responsible for extracellular matrix (ECM) degradation and tissue remodeling. This may partially explain the symptoms of cutaneous relaxation in GSS. Further immunohistochemistry on four GSS cases demonstrated that CD11c predominantly marked granulomas and multinucleated giant cells, whereas CD163 was mainly expressed on scattered macrophages, appearing as a mutually exclusive pattern. The expression pattern of MMP9 overlapped with that of CD11c, implying that CD11c+ macrophages may be a source of MMP9. Our data shed light on the characteristics of macrophages in the GSS microenvironment and provide a theoretical basis for the application of MMP9 inhibitors to prevent cutaneous relaxation of GSS. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

m6A modification-tuned sphingolipid metabolism regulates postnatal liver development in male mice.

Different organs undergo distinct transcriptional, epigenetic and physiological alterations that guarantee their functional maturation after birth. However, the roles of epitranscriptomic machineries in these processes have remained elusive. Here we demonstrate that expression of RNA methyltransferase enzymes Mettl3 and Mettl14 gradually declines during postnatal liver development in male mice. Liver-specific Mettl3 deficiency causes hepatocyte hypertrophy, liver injury and growth retardation. Transcriptomic and N6-methyl-adenosine (m6A) profiling identify the neutral sphingomyelinase, Smpd3, as a target of Mettl3. Decreased decay of Smpd3 transcripts due to Mettl3 deficiency results in sphingolipid metabolism rewiring, characterized by toxic ceramide accumulation and leading to mitochondrial damage and elevated endoplasmic reticulum stress. Pharmacological Smpd3 inhibition, Smpd3 knockdown or Sgms1 overexpression that counteracts Smpd3 can ameliorate the abnormality of Mettl3-deficent liver. Our findings demonstrate that Mettl3-N6-methyl-adenosine fine-tunes sphingolipid metabolism, highlighting the pivotal role of an epitranscriptomic machinery in coordination of organ growth and the timing of functional maturation during postnatal liver development.

AhR-mediated lipid peroxidation contributes to TCDD-induced cardiac defects in zebrafish.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a persistent environmental contaminant that activates the aryl hydrocarbon receptor (AhR) pathway, has been reported to cause cardiac damage. However, the mechanism underlying AhR-induced cardiac defects in response to TCDD exposure remains unclear. In this study, we characterized the impacts of TCDD exposure on heart morphology and cardiac function in zebrafish. TCDD exposure in the early developmental stage of zebrafish embryos led to morphological heart malformation and pericardial edema, concomitant with reduced cardiac function. These cardiac defects were attenuated by inhibiting AhR activity with CH223191. Transcriptome profiling showed that, along with an upregulation of the AhR signaling pathway by TCDD treatment, the expression of pro-ferroptotic genes was upregulated, while that of genes implicated in glutathione metabolism were downregulated. Moreover, lipid peroxidation, as indicated by malonaldehyde (MDA) production, was increased in TCDD-exposed cardiac tissue. Accordingly, inhibiting lipid peroxidation with liproxstatin-1 reversed the adverse cardiac effects induced by TCDD treatment. Taken together, our findings demonstrate that AhR-mediated lipid peroxidation contributes to cardiac defects in the early developmental stage in zebrafish embryos exposed to TCDD.

Proteomics of Salt Gland-Secreted Sap Indicates a Pivotal Role for Vesicle Transport and Energy Metabolism in Plant Salt Secretion.

Soil salinization is one of the major factors restricting crop growth and agricultural production worldwide. Recretohalophytes have developed unique epidermal structures in their aboveground tissues, such as salt glands or salt bladders, to secrete excess salt out of the plant body as a protective mechanism from ion damage. Three hypotheses were proposed to explain how salt glands secrete salts: the osmotic hypothesis, a hypothesis similar to animal fluid transport, and vesicle-mediated exocytosis. However, there is no direct evidence to show whether the salt gland-secreted liquid contains landmark proteins or peptides which would elucidate the salt secretion mechanism. In this study, we collected the secreted liquid of salt glands from Limonium bicolor, followed by extraction and identification of its constituent proteins and peptides by SDS-PAGE and mass spectrometry. We detected 214 proteins and 440 polypeptides in the salt gland-secreted droplets of plants grown under control conditions. Unexpectedly, the proportion of energy metabolism-related proteins increased significantly though only 16 proteins and 35 polypeptides in the droplets of salt-treated plants were detected. In addition, vesicle transport proteins such as the Golgi marker enzyme glycosyltransferase were present in the secreted sap of salt glands from both control and salt-treated plants. These results suggest that trans-Golgi network-mediated vesicular transport and energy production contributes to salt secretion in salt glands.

Metabolic Rewiring of Kynurenine Pathway during Hepatic Ischemia-Reperfusion Injury Exacerbates Liver Damage by Impairing NAD Homeostasis.

Hepatic ischemia-reperfusion (IR) injury remains a common issue lacking effective strategy and validated pharmacological targets. Here, using an unbiased metabolomics screen, this study finds that following murine hepatic IR, liver 3-hydroxyanthranilic acid (3-HAA) and quinolinic acid (QA) decline while kynurenine and kynurenic acid (KYNA) increase. Kynurenine aminotransferases 2, functioning at the key branching point of the kynurenine pathway (KP), is markedly upregulated in hepatocytes during ischemia, shifting the kynurenine metabolic route from 3-HAA and QA to KYNA synthesis. Defects in QA synthesis impair de novo nicotinamide adenine dinucleotide (NAD) biosynthesis, rendering the hepatocytes relying on the salvage pathway for maintenance of NAD and cellular antioxidant defense. Blocking the salvage pathway following IR by the nicotinamide phosphoribosyltransferase inhibitor FK866 exacerbates liver oxidative damage and enhanced IR susceptibility, which can be rescued by the lipid peroxidation inhibitor Liproxstatin-1. Notably, nicotinamide mononucleotide administration once following IR effectively boosts NAD and attenuated IR-induced oxidative stress, inflammation, and cell death in the murine model. Collectively, the findings reveal that metabolic rewiring of the KP partitions it away from NAD synthesis in hepatic IR pathophysiology, and provide proof of concept that NAD augmentation is a promising therapeutic measure for IR-induced liver injury.

Salt-tolerance screening in Limonium sinuatum varieties with different flower colors.

Limonium sinuatum, a member of Plumbaginaceae commonly known as sea lavender, is widely used as dried flower. Five L. sinuatum varieties with different flower colors (White, Blue, Pink, Yellow, and Purple) are found in saline regions and are widely cultivated in gardens. In the current study, we evaluated the salt tolerance of these varieties under 250 mmol/L NaCl (salt-tolerance threshold) treatment to identify the optimal variety suitable for planting in saline lands. After the measurement of the fresh weight (FW), dry weight (DW), contents of Na+, K+, Ca2+, Cl-, malondialdehyde (MDA), proline, soluble sugars, hydrogen peroxide (H2O2), relative water content, chlorophyll contents, net photosynthetic rate, and osmotic potential of whole plants, the salt-tolerance ability from strongest to weakest is identified as Pink, Yellow, Purple, White, and Blue. Photosynthetic rate was the most reliable and positive indicator of salt tolerance. The density of salt glands showed the greatest increase in Pink under NaCl treatment, indicating that Pink adapts to high-salt levels by enhancing salt gland formation. These results provide a theoretical basis for the large-scale planting of L. sinuatum in saline soils in the future.

Dynamic imaging of zebrafish heart with multi-planar light sheet microscopy.

Light sheet fluorescence microscopy has become a research hotspot in biomedicine because of low phototoxicity, high speed, and high resolution. However, the conventional methods to acquire three-dimensional spatial information are mainly based on scanning, which inevitably increases photodamage and is not real-time. Here, we propose a method to generate controllable multi-planar illumination with a dielectric isosceles triangular array and a design of multi-planar light sheet fluorescence microscopy system. We carry out experiments of three-dimensional illumination beam measurement, volumetric imaging of fluorescent microspheres, and dynamic in vivo imaging of zebrafish heart to evaluate the performance of this system. In addition, we apply this system to study the effects of bisphenol fluorene on the heart shape and heart-beating rate of zebrafish. Our experiment results indicate that the multi-planar light sheet microscopy system provides a novel and feasible method for three-dimensional selected plane imaging and low-phototoxicity in vivo imaging.

Environmental level of the antidepressant venlafaxine induces behavioral disorders through cortisol in zebrafish larvae (Danio rerio).

Psychoactive drugs discharged into the environment have different effects on the behavior of vertebrates. The objective of this study was to evaluate the effect of venlafaxine on the behavior of zebrafish, and whether melatonin could reverse the induction of venlafaxine. In this study, a series of venlafaxine concentrations (1 µg/L, 10 µg/L, 100 µg/L) was used to treat zebrafish embryos from 2 hours post-fertilization (hpf) to 5dpf. We found that venlafaxine (1 µg/L) can stimulate the growth of the head area, eye area, and body length of zebrafish. The light-dark test showed that venlafaxine (1 µg/L) could increase the activity of zebrafish larvae. What's more, venlafaxine (1 µg/L) upregulated the expression of steroid regulatory factors including steroidogenic acute regulatory protein (star), cytochrome P450 family member 11A1 (cyp11a1) and 11 ß hydroxylase (cyp11b1) by cAMP-pCREB pathway, affecting the function of the steroidogenic cells, which might be involved in the increased cortisol levels in zebrafish larvae. Whereas, melatonin (230 µg/L) restored the altered locomotion behavior induced by venlafaxine and recovered the altered gene expression. Our results demonstrate that venlafaxine at levels detected in the aquatic environment impacts behavior and may compromise the adaptive responses to the environment in zebrafish larvae.

Acute fluorene-9-bisphenol exposure damages early development and induces cardiotoxicity in zebrafish (Danio rerio).

Fluorene-9-bisphenol (BHPF) is a substitute for bisphenol A (BPA), which is widely used to manufacture plastic products. Previous studies indicate that BHPF has an anti-estrogenic effect and induces cytotoxicity in mice oocytes. However, the effects of acute BHPF exposure on the aquatic organism obtain little attention. In this study, a series of BHPF concentrations (1 µM, 2 µM, 5 µM, 10 µM, 20 µM) was used to exposed zebrafish embryos from 2 h post-fertilization (hpf). The results showed the LC50 at 96hpf was 2.88 µM (1.01 mg/L). Acute exposure induced malformation in morphology, and retarded epiboly rate at 10hpf, increased apoptosis. Moreover, acute BHPF exposure led cardiotoxicity, by impeding cardiac looping, decreasing cardiac contractility (reducing the stroke volume and cardiac output, decreasing fractional shortening of ventricle). Besides that, BHPF exposure altered the expression of cardiac transcriptional regulators and development related genes. In conclusion, acute BHPF exposure induced developmental abnormality, retarded cardiac morphogenesis and injured the cardiac contractility. This study indicated BHPF would be an unneglected threat for the safety of aquatic organisms.

Melatonin attenuates 17ß-trenbolone induced insomnia-like phenotype and movement deficiency in zebrafish.

17ß-trenbolone (17ß-TBOH) is one of the dominant metabolites of trenbolone acetate, which is widely applied in beef cattle operations around the globe. The effects of environmental concentrations of 17ß-trenbolone on the early development of zebrafish embryos have received very little attention. Melatonin could regulate sleep-wake cycle and plays a protective role in various adverse conditions. Here, environmentally realistic concentrations of 17ß-trenbolone (1 ng/L, 10 ng/L, 50 ng/L) has been exposure to zebrafish embryos at 2 h postfertilization (hpf). The results showed that 10 ng/L and 50 ng/L 17ß-trenbolone disturbed the distribution of caudal primary motoneurons and downregulated expression of motoneuron development related genes along with locomotion decreasing. While melatonin could recover the detrimental effects caused by 17ß-trenbolone. Interestingly, 17ß-trenbolone exposure increased waking activity and decreased rest even in a low dose (1 ng/L). Moreover, it upregulated hypocretin/orexin (Hcrt) signaling which promotes wakefulness. Melatonin restored the insomnia-like alternation induced by 17ß-trenbolone exposure. Collectively, we conclude that 17ß-trenbolone disturbed motoneuron development and altered sleep/wake behavior, while melatonin could alleviate the deleterious influence on motoneuron development and recover the circadian rhythm.

Combined treatment of melatonin and sodium tanshinone IIA sulfonate reduced the neurological and cardiovascular toxicity induced by deltamethrin in zebrafish.

The pyrethroid insecticide deltamethrin has been reported to have an effect on vertebrate development and cardiovascular disease. Sodium tanshinone IIA sulfonate (STS) is considered to have cardioprotective effects and melatonin is known to regulate sleep-waking cycles. In this experiment, we used transgenic zebrafish Tg (kdrl:mCherry) and Tg (myl7:GFP) to investigate whether STS and melatonin could reverse the cardiovascular toxicity and neurotoxicity induced by deltamethrin. Zebrafish embryos were exposed to 25 µg/L deltamethrin at 10 hpf and treated with 100 mmol/L STS and 1 µmol/L melatonin showed that deltamethrin treatment affected normal cardiovascular development. In situ hybridization and qRT-PCR results showed that deltamethrin could interfere with the normal expression of cardiovascular development-related genes vegfr2, shh, gata4, nkx2.5, causing functional defects in the cardiovascular system. In addition, deltamethrin could affect the sleep-waking behavior of larvae, increasing the activity of larvae, decreasing the rest behavior and the expression of hcrt, hcrtr, aanat2 were down-regulated. The addition of melatonin and STS can significantly alleviate cardiovascular toxicity and sleep-waking induced by deltamethrin, while restoring the expression of related genes to normal levels. Our study demonstrates the role of STS and melatonin in protecting cardiovascular and sleep-waking behavior caused by deltamethrin.

Venlafaxine plus melatonin ameliorate reserpine-induced depression-like behavior in zebrafish.

Venlafaxine (VEN) is one of the first clinical drugs for the treatment of depression. Long-term use may cause a potentially life-threatening serotonin syndrome. Melatonin (MT) could ameliorate depression behavior. Therefore, the aim of this study was to investigate the antidepressant effects of venlafaxine in combination with melatonin on zebrafish. Reserpine was used to induce depression-like behavioral zebrafish. To explore the effects of combined use of venlafaxine and melatonin on depression-like zebrafish induced by reserpine. We tested the depressive behavior of adult zebrafish through a novel tank test, and evaluated the levels of serotonin (5-HT), dopamine (DA) and noradrenaline (NA) in zebrafish brain using enzyme-linked immunosorbent assay (ELISA), besides that the gene expression of serotonin transporters a (serta), dopamine transporters (dat) and norepinephrine transporters (net), vesicular monoamine transporter2 (vmat2) and monoamine oxidase (mao) were evaluated by qRT-PCR. The results showed that, compared with reserpine-only group, venlafaxine (VEN, 0.025 mg/L) and melatonin (MT, 1 µM) increased the parameters of exploration in the top of the tank and decreased freezing behavior significantly. Compared with reserpine-only group, the use of VEN combined with MT increased serotonin and norepinephrine levels significantly, while there was no obvious difference in dopamine content. The results of qRT-PCR showed that the use of VEN combined with MT significantly reduced the expression of serta and promoted the expression of vmat2, but had no significant effect on the expression of net, dat and mao. The results indicated that venlafaxine combined with melatonin showed more effective role to remedy the depressive symptoms in zebrafish, providing a reference for the clinical application of antidepressants.

The effects of fluorene-9-bisphenol on female zebrafish (Danio rerio) reproductive and exploratory behaviors.

Endocrine disruptor chemicals induce adverse effects to animals' development, reproduction and behavior in environment. We investigated the effects of fluorene-9-bisphenol (BHPF), one substitute of bisphenol A, on courtship behavior and exploratory behavior of adult zebrafish. Customized apparatus was used to evaluate courtship behavior. The result showed that the male spent less time with BHPF and anti-oestrogenic fulvestrant (FULV) treated female in region of approaching (ROA). Courtship index between BHPF-exposed female and male decreased. The body orientation of BHPF- and FULV-exposed female to male decreased. Furthermore, BHPF exposure downregulated the expression of genes related to estrogen receptor, steroidogenesis and upregulated oxidative stress related genes. It indicated that BHPF exposure interfered the preference of male and female in courtship, and induced detrimental effects on reproduction. BHPF treatment decreased locomotor activity and time spent in top, increased freezing bouts, and induced anxiety/depression-like behavior. The tyrosine hydroxylase in brain decreased under BHPF exposure. Here we showed the potential adverse effects of BHPF on reproduction and exploratory behaviors.

Melatonin protects embryonic development and maintains sleep/wake behaviors from the deleterious effects of fluorene-9-bisphenol in zebrafish (Danio rerio).

Environmental endocrine chemicals have various adverse effects on the development of vertebrates. Fluorene-9-bisphenol (BHPF), a substitute of bisphenol A (BPA), is widely used in commercial production. The effects of BHPF on development and behavior are unclear. Melatonin plays a protective role under many unfavorable conditions. In this study, we investigated the effects of BHPF on the development and behaviors of zebrafish and whether melatonin reverses effects induced by BHPF. Zebrafish embryos were exposed to 0.1, 10, or 1000 nmol/L BHPF with or without 1 µmol/L melatonin from 2 hours postfertilization to 6 days postfertilization. The results showed that 0.1 and 10 nmol/L BHPF had little effect on development. High-dose BHPF (1000 nmol/L) delayed the development, increased mortality and surface tension of embryonic chorions, caused aberrant expression of the key genes (ntl, shh, krox20, pax2, cmlc2) in early development detected by in situ hybridization, and damaged the CaP motor neurons, which were associated with locomotion ability detected by immunofluorescence. Melatonin addition reversed or weakened these adverse effects of BHPF on development, and melatonin alone increased surface tension as the effects of high-dose BHPF. However, all groups of BHPF exposure triggered insomnia-like behaviors, with increased waking activity and decreased rest behaviors. BHPF acted on the hypocretin (hcrt) system and upregulated the expression of sleep/wake regulators such as hcrt, hcrt receptor (hcrtr), arylalkylamine N-acetyltransferase-2 (aanat2). Melatonin recovered the alternation of sleep/wake behaviors induced by BHPF and restored abnormal gene expression to normal levels. This study showed that high-dose BHPF had adverse effects on early development and induced behavioral alternations. However, melatonin prevented BHPF-induced aberrant development and sleep/wake behaviors.

Developmental toxicity and neurotoxicity of synthetic organic insecticides in zebrafish (Danio rerio): A comparative study of deltamethrin, acephate, and thiamethoxam.

Synthetic organic insecticides, including pyrethroids, organophosphates, neonicotinoids and other types, have the potential to alter the ecosystems and many are harmful to humans. This study examines the developmental toxicity and neurotoxicity of three synthetic organic insecticides, including deltamethrin (DM), acephate (AP), and thiamethoxam (TM), using embryo-larval stages of zebrafish (Danio rerio). Results showed that DM exposure led to embryo development delay and a significant increase in embryo mortality at 24 and 48 h post-fertilization (hpf). DM and AP decreased embryo chorion surface tension at 24 hpf, along with the increase in hatching rate at 72 hpf. Moreover, DM caused ntl, shh, and krox20 misexpression in a dose-dependent manner with morphological deformities of shorter body length, smaller eyes, and larger head-body angles at 10 µg/L. TM did not show significant developmental toxicity. Furthermore, results of larval rest/wake assay indicated that DM (>0.1 µg/L) and AP (0.1 mg/L) increased activity behavior with different patterns. Interestingly, as an insect-specific pesticide, TM still could alter locomotor activity in zebrafish larvae at concentrations as low as 0.1 mg/L. Our results indicate that different types of synthetic organic insecticides could create different toxicity outcomes in zebrafish embryos and larvae.

Effects of sodium bicarbonate concentration on growth, photosynthesis, and carbonic anhydrase activity of macroalgae Gracilariopsis lemaneiformis, Gracilaria vermiculophylla, and Gracilaria chouae (Gracilariales, Rhodophyta).

There is potential for bicarbonate to improve crop yields and economic efficiency of marine algae. However, few studies have focused on the effect of bicarbonate on the growth, photosynthesis, and enzyme activity associated with carbon utilization, especially in commercial macroalgae. Here, the addition of bicarbonate (up to 420 mg L(-1)) to macroalgal cultures has been evaluated for Gracilariopsis lemaneiformis, Gracilaria vermiculophylla, and Gracilaria chouae with respect to growth rate, photosynthetic activity, carbonic anhydrase activity, and biochemical composition. The results showed that the effects of NaHCO3 on growth, chlorophyll a, phycoerythrin, photosynthetic oxygen evolution, photochemical parameters of PSI and PSII, carbonic anhydrase activity, and nitrogen content were significant (P < 0.05) and followed the same pattern in the three species. The parameter values were promoted in lower NaHCO3 concentrations (up to 252 or 336 mg L(-1)) and inhibited in higher NaHCO3 concentrations (>336 mg L(-1) for Gp. lemaneiformis and >420 mg L(-1) for the other two species). Moreover, species-specific differences induced by supplementation with bicarbonate were discovered during culture. Optimal concentrations of NaHCO3 used in this study were 252 mg L(-1) for Gp. lemaneiformis and 336 mg L(-1) for G. vermiculophylla and G. chouae. These results suggest that an adequate supplementation of sodium bicarbonate is a viable strategy for promoting growth and photosynthetic activity in some macroalgae as well as for improving biochemical composition. The study will help to accelerate the growth rate of algae and improve the quality of thalli, and will also be useful for enhancing the understanding of carbon utilization in macroalgae.