Foxp2 deficiency impairs reproduction by modulating the hypothalamic-pituitary-gonadal axis in zebrafish†.

FOXP2 was initially characterized as a transcription factor linked to speech and language disorders. Single-cell RNA sequencing reveals that Foxp2 is enriched in the gonadotrope cluster of the pituitary gland and colocalized with the hormones LHB and FSHB in chickens and mice, implying that FOXP2 might be associated with reproduction in vertebrates. Herein, we investigated the roles of foxp2 in reproduction in a Foxp2-deficient zebrafish model. The results indicated that the loss of Foxp2 inhibits courtship behavior in adult male zebrafish. Notably, Foxp2 deficiency disrupts gonad development, leading to retardation of follicle development and a decrease in oocytes in females at the full-growth stage, among other phenotypes. The transcriptome analysis (RNA-seq) also revealed that differentially expressed genes clustered into the estrogen signaling and ovarian steroidogenesis-related signaling pathways. In addition, we found that Foxp2 deficiency could modulate the hypothalamic-pituitary-gonadal axis, especially the regulation of lhb and fshb expression, in zebrafish. In contrast, the injection of human chorionic gonadotropin, a specific LH agonist, partially rescues Foxp2-impaired reproduction in zebrafish, suggesting that Foxp2 plays an important role in the regulation of reproduction via the hypothalamic-pituitary-gonadal axis in zebrafish. Thus, our findings reveal a new role for Foxp2 in the regulation of reproduction in vertebrates.

Docosahexaenoic acid-mediated milk protein treated by ultrasound-assisted pH shifting for enhanced astaxanthin delivery and processed cheese application.

Whey protein isolate (WPI)-based nanodelivery systems have recently attracted an increasing amount of attention. Despite this, research focusing on milk protein concentrate (MPC) and micellar casein (MCC) as carriers loaded in hydrophobic compounds is lacking. This study investigated the mediated effect of docosahexaenoic acid (DHA) in 3 different milk proteins for the embedding of astaxanthin (ASTA) after ultrasound-assisted pH-shifting treatment. We then evaluated the application of milk protein carriers in cheese processing by comparing MPC, MCC, and WPI. The particle size, polydispersity index, and zeta potential results of the milk protein-DHA complex suggested that the addition of 0.36 µmol/mL DHA optimized the delivery of milk protein to ASTA. All 3 DHA-mediated milk proteins induced an improvement in encapsulation efficiency and antioxidant properties of ASTA. Furthermore, the DHA-mediated MPC and MCC played a stronger role in improving the bioaccessibility and thermal and storage stability of ASTA than those without DHA. Tests conducted to examine the application in cheese production indicated that MCC carrier had a positive effect on the texture of cheeses. However, the delivery effect was dependent on the milk protein variety, and MCC exhibited the best protection ability of ASTA, followed by MPC and WPI. The simulated digestion and storage stability results of cheese further confirmed that the protein encapsulation mediated by DHA was more conducive to ASTA absorption. These findings suggested that the DHA-mediated milk protein complexes studied here may be suitable hydrophilic delivery carriers for the hydrophobic nutrient ASTA, potentially playing different roles in improving its storage stability and bioaccessibility.

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.

Role of Impurities in Kaolinite Intercalation and Subsequent Formation of Nanoscrolls.

Kaolinite (Kaol)-methanol (MeOH) compounds (Kaol-Me) are widely used as the starting materials for further intercalation. The conventional approach to prepare Kaol-Me compounds is to wash dimethyl sulfoxide (DMSO)-intercalated Kaol (Kaol-DMSO) for 16 days, and MeOH must be refreshed every day. Herein, we report a new and much more efficient method to prepare Kaol-Me from Kaol-DMSO by the promotion of AlCl3 under mild conditions, and the corresponding mechanism is investigated. The X-ray diffraction (XRD), Fourier transform infrared spectroscopy, and X-ray fluorescence characterization results reveal that the electric double layer resulting from the impurities absorbed on the kaolinite surface prevents weakly polar molecules from entering the kaolinite interlayers, which is probably the key reason that MeOH must be refreshed daily in the preparation of Kaol-Me compounds. After being treated with HCl to remove the impurities, Kaol-Me-HCl was successfully intercalated by cetyltrimethyl ammonium bromide and subsequently predominantly curled into nanoscrolls.

Stepwise crosstalk between aberrant Nf1, Tp53 and Rb signalling pathways induces gliomagenesis in zebrafish.

The molecular pathogenesis of glioblastoma indicates that RTK/Ras/PI3K, RB and TP53 pathways are critical for human gliomagenesis. Here, several transgenic zebrafish lines with single or multiple deletions of nf1, tp53 and rb1 in astrocytes, were established to genetically induce gliomagenesis in zebrafish. In the mutant with a single deletion, we found only the nf1 mutation low-efficiently induced tumour incidence, suggesting that the Nf1 pathway is critical for the initiation of gliomagenesis in zebrafish. Combination of mutations, nf1;tp53 and rb1;tp53 combined knockout fish, showed much higher tumour incidences, high-grade histology, increased invasiveness, and shortened survival time. Further bioinformatics analyses demonstrated the alterations in RTK/Ras/PI3K, cell cycle, and focal adhesion pathways, induced by abrogated nf1, tp53, or rb1, were probably the critical stepwise biological events for the initiation and development of gliomagenesis in zebrafish. Gene expression profiling and histological analyses showed the tumours derived from zebrafish have significant similarities to the subgroups of human gliomas. Furthermore, temozolomide treatment effectively suppressed gliomagenesis in these glioma zebrafish models, and the histological responses in temozolomide-treated zebrafish were similar to those observed in clinically treated glioma patients. Thus, our findings will offer a potential tool for genetically investigating gliomagenesis and screening potential targeted anti-tumour compounds for glioma treatment.

Current understanding of gliomagenesis: from model to mechanism.

Glioma, a kind of central nervous system (CNS) tumor, is hard to cure and accounts for 32% of all CNS tumors. Establishing a stable glioma model is critically important to investigate the underlying molecular mechanisms involved in tumorigenesis and tumor progression. Various core signaling pathways have been identified in gliomagenesis, such as RTK/RAS/PI3K, TP53, and RB1. Traditional methods of establishing glioma animal models have included chemical induction, xenotransplantation, and genetic modifications (RCAS/t-va system, Cre-loxP, and TALENs). Recently, CRISPR/Cas9 has emerged as an efficient gene editing tool with high germline transmission and has extended the scope of stable and efficient glioma models that can be generated. Therefore, this review will highlight the documented evidence about the molecular characteristics, critical genetic markers, and signaling pathways responsible for gliomagenesis and progression. Moreover, methods of establishing glioma models using gene editing techniques and therapeutic aspects will be discussed. Finally, the prospect of applying gene editing in glioma by using CRISPR/Cas9 strategy and future research directions to establish a stable glioma model are also included in this review. In-depth knowledge of glioma signaling pathways and use of CRISPR/Cas9 can greatly assist in the development of a stable, efficient, and spontaneous glioma model, which can ultimately improve the effectiveness of therapeutic responses and cure glioma patients.

Silver nanoparticles induce developmental toxicity via oxidative stress and mitochondrial dysfunction in zebrafish (Danio rerio).

Sliver nanoparticles (AgNPs) are widely used in industry, agriculture, and medicine, potentially resulting in adverse effects on human health and aquatic environments. Here, we investigated the developmental toxicity of zebrafish embryos with acute exposure to AgNPs. Our results demonstrated developmental defects in 4 hpf zebrafish embryos after exposure to different concentrations of AgNPs for 72 h. In addition, RNA-seq profiling of zebrafish embryos after AgNPs treatment. Further Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that the differentially expressed genes (DEGs) were enriched in DNA replication initiation, oxidoreductase activity, DNA replication, cellular senescence, and oxidative phosphorylation signaling pathways in the AgNPs-treated group. Notably, we also found that AgNPs exposure could result in the accumulation of reactive oxygen species (ROS) and malondialdehyde (MDA), the inhibition of superoxide dismutase (SOD), catalase (CAT), and mitochondrial complex I-V activities, and the downregulated expression of SOD, CAT, and mitochondrial complex I-IV chain-related genes. Moreover, the expression of mitochondrion-mediated apoptosis signaling pathway-related genes, such as bax, bcl2, caspase-3, and caspase-9, was significantly regulated after AgNPs exposure in zebrafish. Therefore, these findings demonstrated that AgNPs exposure could cause oxidative stress, induce mitochondrial dysfunction, and ultimately lead to developmental toxicity.

Par6 regulates cell cycle progression through enhancement of Akt/PI3K/GSK-3ß signaling pathway activation in glioma.

As the key factor of the polarity protein complex, Par6 not only regulates polarization processes, but also plays important roles in tumor metastasis and progression in many epithelium malignancy tumors. Here, we showed that Par6 is an essential component in glioma tumorigenesis. Our results indicated the aberrant expression of Par6 in malignant glioma tissues and cell lines. We found that the regulation of Par6 expression induces cell proliferation and tumor growth in vivo and in vitro. Additionally, RNA-seq revealed the effects of Par6 were associated with cyclin D1-regulated cell cycle progression in glioma cells. Moreover, our results demonstrated that the regulation of Par6 can enhance the activation of Akt/PI3K signaling pathway, and subsequently upregulate the expression level of GSK-3ß protein, which then regulate cyclin D1-mediated cell cycle regulation. Furthermore, we found that TGF-ß-induced the upregulation of Par6 expression may be involved in this process. The pathological analysis confirmed the correlation between Par6 expression and the prognosis in human glioma tissues, suggesting the regulation of Par6 expression regulates glioma tumorigenesis and progression. Thus, our findings showed that Par6 might be a potential biomarker for the diagnosis and providing a therapeutic strategy for the treatment of malignant glioma.

The oxidative stress responses caused by phthalate acid esters increases mRNA abundance of base excision repair (BER) genes in vivo and in vitro.

The base excision repair (BER) pathway is an important defense response to oxidative DNA damage. It is known that exposures to phthalate esters (PAEs), including Dibutyl phthalate (DBP), Mono-(2-ethylhexyl) phthalate (MEHP), and Di-(2-ethylhexyl) phthalate (DEHP), cause reactive oxygen species-induced DNA damage and oxidative stress. Here, we determined the mRNA levels of BER pathway-related genes (ogg1, nthl1, apex1, parp1, xrcc1, lig3, ung, pcna, polb, pold, fen1, and lig1), pro-apoptotic gene (bax), and apoptotic suppressor gene (bcl2) in different PAEs-exposed zebrafish larvae and HEK293T cells. Further investigations were performed to examine reactive oxygen species (ROS) accumulation, superoxide dismutase (SOD) activity, developmental toxicity, and cell viability after PAEs exposure in vivo and in vitro. The results showed that PAEs exposure can induce developmental abnormalities in zebrafish larvae, and inhibit cell viability in HEK293T cells. Additionally, we found that PAEs exposure results in the accumulation of ROS and the inhibition of SOD activation in vivo and in vitro. Notably, the mRNA levels of BER pathway-related genes (OGG1, NTHL1, APEX1, XRCC1, UNG, POLB, POLD, FEN1) were significantly upregulated after DBP or MEHP exposure, whereas the mRNA levels of NTHL1, UNG, POLB, POLD, and FEN1 were significantly altered in DEHP-treated HEK293T cells. In zebrafish, the mRNA levels of ogg1, pcna, fen1 and lig1 genes were increased after DBP or DEHP exposure, whereas the mRNA levels of nthl1, apex1, parp1, lig3, pcna and polb were decreased after MEHP exposure, respectively. Thus, our findings indicated that PAEs exposure can induce developmental toxicity, cytotoxicity, and oxidative stress, as well as activate BER pathway in vivo and in vitro, suggesting that BER pathway might play critical roles in PAEs-induced oxidative stress through repairing oxidative DNA damage.

Circular RNA hsa_circ_0008285 inhibits colorectal cancer cell proliferation and migration via the miR-382-5p/PTEN axis.

Abundant evidence has showed that circular RNA (circRNA) plays an important role in cancer. Nonetheless, little is known about the roles and mechanisms of specific circRNAs in different cancer types. Hsa_circ_0008285 (circ_0008285), derived from the coding gene chromodomain y-like protein (CDYL), is upregulated in hepatocellular carcinoma and mantle cell lymphoma. However, we previously found, by analyzing two independent high-throughput sequencing datasets, that it was reduced in colon cancer. In this study, we explored the function and mechanism of circ_0008285 in the progression of colorectal cancer (CRC). First, the downregulated expression of circ_0008285 in CRC tissues and cell lines was confirmed using RT-qPCR analysis. In addition, the expression level of circ_0008285 was inversely correlated with tumor size, lymphatic metastasis, and tumor-node-metastasis (TNM) stage through clinicopathological parameter analysis. Functionally, knockdown of circ_0008285 promoted CRC cell proliferation and migration in vitro. Mechanistically, by using RNA-sequencing, bioinformatics analysis, dual-luciferase reporter assay, and western blotting, we determined that circ_0008285 suppressed the PI3K/AKT pathway via the miR-382-5p/PTEN axis. In conclusion, our data demonstrate a tumor suppressor role for circ_0008285 in CRC and suggest circ_0008285 as a potential target for CRC treatment.

Heparanase promotes glioma progression via enhancing CD24 expression.

Heparanase is an endo-ß-d-glucuronidase that cleaves heparan sulfate (HS) side chains of heparan sulfate proteoglycans. Compelling evidence tie heparanase levels with all steps of tumor formation including tumor initiation, growth, metastasis and chemo-resistance, likely involving augmentation of signaling pathways and gene transcription. In order to reveal the molecular mechanism(s) underlying the protumorigenic properties of heparanase, we established an inducible (Tet-on) system in U87 human glioma cells and applied gene array methodology in order to identify genes associated with heparanase induction. We found that CD24, a mucin-like cell adhesion protein, is consistently upregulated by heparanase and by heparanase splice variant devoid of enzymatic activity, whereas heparanase gene silencing was associated with decreased CD24 expression. This finding was further substantiated by a similar pattern of heparanase and CD24 immunostaining in glioma patients (Pearson's correlation; R = 0.66, p = 0.00001). Noteworthy, overexpression of CD24 stimulated glioma cell migration, invasion, colony formation in soft agar and tumor growth in mice suggesting that CD24 functions promote tumor growth. Likewise, anti-CD24 neutralizing monoclonal antibody attenuated glioma tumor growth, and a similar inhibition was observed in mice treated with a neutralizing mAb directed against L1 cell adhesion molecule (L1CAM), a ligand for CD24. Importantly, significant shorter patient survival was found in heparanase-high/CD24-high tumors vs. heparanase-high/CD24-low tumors for both high-grade and low-grade glioma (p = 0.02). Our results thus uncover a novel heparanase-CD24-L1CAM axis that plays a significant role in glioma tumorigenesis.

Hypersensitive assessment of aryl hydrocarbon receptor transcriptional activity using a novel truncated cyp1a promoter in zebrafish.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent organic pollutant (POP), an unintentional byproduct of various industrial processes, and a human carcinogen. The expression of the cytochrome P450 1A (cyp1a) gene is upregulated in the presence of TCDD through activating the aryl hydrocarbon receptor pathway in a dose-dependent manner. Several essential response elements, including the 8 potential xenobiotic response elements in the cyp1a promoter region, have been identified to be the main functional parts for the response to TCDD. Thus, we aimed to develop a convenient and sensitive biomonitoring tool to examine the level of POPs in the environment and evaluate its potential human health risks by TCDD. Here, we established a transgenic zebrafish model with a red fluorescent reporter gene ( mCherry) using the truncated cyp1a promoter. Under exposure to TCDD, the expression pattern of mCherry in the reporter zebrafish mirrored that of endogenous cyp1a mRNA, and the primary target tissues for TCDD were the brain vessels, liver, gut, cloaca, and skin. Our results indicated that exposure of the embryos to TCDD at concentrations as low as 0.005 nM for 48 h, which did not elicit morphologic abnormalities in the embryos, markedly increased mCherry expression. In addition, the reporter embryos responded to other POPs, and primary liver cell culture of zebrafish revealed that Cyp1a protein was mainly expressed in the cytoplasm of liver cells. Furthermore, our transgenic fish embryos demonstrated that TCDD exposure can regulate the expression levels of several tumor-related factors, including epidermal growth factor, TNF-α, C-myc, proliferating cell nuclear antigen, TGF-ß, serine/threonine kinase (Akt), and phosphorylated Akt, suggesting that our transgenic fish can be used as a sensitive model to evaluate the carcinogenicity induced by TCDD exposure.-Luo, J.-J., Su, D.-S., Xie, S.-L., Liu, Y., Liu, P., Yang, X.-J., Pei D.-S. Hypersensitive assessment of aryl hydrocarbon receptor transcriptional activity using a novel truncated cyp1a promoter in zebrafish.

CRISPR/Cas9-based genome engineering of zebrafish using a seamless integration strategy.

Numerous feasible methods for inserting large fragments of exogenous DNA sequences into the zebrafish genome have been developed, as has genome editing technology using programmable nucleases. However, the coding sequences of targeted endogenous genes are disrupted, and the expression patterns of inserted exogenous genes cannot completely recapitulate those of endogenous genes. Here we describe the establishment of a novel strategy for endogenous promoter-driven and microhomology-mediated end-joining-dependent integration of a donor vector using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) 9. We successfully integrated mCherry into the final coding sequence of targeted genes to generate seamless transgenic zebrafish lines with high efficiency. This novel seamless transgenesis technique not only maintained the integrity of the endogenous gene but also did not disrupt the function of targeted gene. Therefore, our microhomology-mediated end-joining-mediated transgenesis strategy may have broader applications in gene therapy. Moreover, this novel seamless gene-editing strategy in zebrafish provides a valuable new transgenesis technique, which was driven by endogenous promoters and in vivo animal reporter modes for translational medicine. It is expected to be a standard gene-editing technique in the field of zebrafish, leading to some important breakthroughs for studies in early embryogenesis.-Luo, J.-J., Bian, W.-P., Liu, Y., Huang, H.-Y., Yin, Q., Yang, X.-J., Pei, D.-S. CRISPR/Cas9-based genome engineering of zebrafish using a seamless integration strategy.

Association of composite dietary antioxidant index with depression and all-cause mortality in middle-aged and elderly population.

Current research has shown an increasing acceptance of interventions for depression through dietary modifications. However, whether composite dietary antioxidant index (CDAI) is associated with depression and all-cause mortality in middle-aged and elderly population remains unknown. This study aimed to explore those associations in American middle-aged and elderly population. Weighted logistic regression models and weighted Cox proportional hazard regression models were used to assess the association of CDAI, covariates, depression, and all-cause mortality, respectively. The stability of the results was also determined by a linear trend test based on CDAI quintiles. Restricted cubic spline curves were employed to test for non-linear relationships. In the model adjusted for all covariates, significant associations were found with the ORs (95% CI) for CDAI and depression [0.77 (0.67, 0.89)] and the HRs (95% CI) for CDAI with all-cause mortality[0.91 (0.83, 1.00)]. Upon conducting restricted cubic spline curves, we found that the association between CDAI and depression was linear, whereas the association between CDAI and all-cause mortality was non-linear with an inflection point of -0.19. Statistical significance was only found before the inflection point. In this study of middle-aged and elderly Americans, CDAI was linearly negatively associated with depression and non-linearly negatively associated with all-cause mortality.

Conjugation of chitopentaose with ß-lactoglobulin using Maillard reaction, and its effect on the allergic desensitization in vivo.

The conjugation of chitopentaose (CHP) on ß-lactoglobulin (ßLg) via Maillard reaction was used to desensitize ßLg. The stable ßLg-CHP conjugate (ßC-4) was formed at 4 h incubation, which contains 5 CHP attached molecules and a conjugated degree of 42 %. The conjugation promoted the thermal stability and emulsifying properties of ßLg, and inhibited the immunoglobulin E (IgE) combining capacity by decreasing the content of ß-sheet in ßLg. Moreover, ßLg-CHP conjugates were imparted with anti-oxidant properties and anti-inflammatory activities. Further, the combined action of inhibited IgE combining capacity and anti-inflammatory activities improved the allergy desensitization in ßLg sensitized mice. The results showed that overexpressed IgE and inflammatory factors, unbalanced Th1-/Th2- immune cytokines were significantly attenuated after ßLg was conjugated with CHP, avoiding the inflammatory lesions in spleen and colon. Additionally, the adverse changes in gut microbiota were alleviated in ßC-4 group with a decrease of Bacteroidetes and increase of Firmicutes at phylum level and the probiotic bacteria of Lactobacillaceae was significantly improved at the family level. Thus, the conjugation of CHP can desensitize allergic reaction caused by ßLg.

Nanoplastic Exposure Mediates Neurodevelopmental Toxicity by Activating the Oxidative Stress Response in Zebrafish (Danio rerio).

The global accumulation and adverse effects of nanoplastics (NPs) are a growing concern for the environment and human health. In recent years, more and more studies have begun to focus on the toxicity of plastic particles for early animal development. Different particle sizes of plastic particles have different toxicities to biological development. Nevertheless, the potential toxicological effects of 20 nm NPs, especially on neurodevelopment, have not been well investigated. In this paper, we used fluorescence microscopy to determine neurotoxicity in zebrafish at different concentrations of NPs. Moreover, the behavioral analysis demonstrated that NPs induced abnormal behavior of zebrafish. The results revealed developmental defects in zebrafish embryos after exposure to different concentrations (0, 0.3, 3, and 9 mg/L) of NPs. The morphological deformities, including abnormal body length and the rates of heart, survival, and hatching, were induced after NP exposure in zebrafish embryos. In addition, the development of primary motor neurons was observed the inhibitory effects of NPs on the length, occurrence, and development of primary motor neurons in Tg(hb9:GFP). Quantitative polymerase chain reaction analysis suggested that exposure to NPs significantly affects the expression of the genes involved in the occurrence and differentiation of primary motor neurons in zebrafish. Furthermore, the indicators associated with oxidative stress and apoptosis were found to be modified in zebrafish embryos at 24 and 48 h following exposure to NPs. Our findings demonstrated that NPs could cause toxicity in primary motor neurons by activating the oxidative stress response and inducing apoptosis, consequently impairing motor performance.

An Image Classification Method Based on Adaptive Attention Mechanism and Feature Extraction Network.

The convolution neural network (CNN) not only has high fault tolerance but also has high computing capacity. The image classification performance of CNN has an important relationship with its network depth. The network depth is deeper, and the fitting ability of CNN is stronger. However, a further increase in the depth of CNN will not improve the accuracy of the network but will produce higher training errors, which will reduce the image classification performance of CNN. In order to solve the above problems, this paper proposes a feature extraction network, AA-ResNet with an adaptive attention mechanism. The residual module of the adaptive attention mechanism is embedded for image classification. It consists of a feature extraction network guided by the pattern, a generator trained in advance, and a complementary network. The feature extraction network guided by the pattern is used to extract different levels of features to describe different aspects of an image. The design of the model effectively uses the image information of the whole level and the local level, and the feature representation ability is enhanced. The whole model is trained as a loss function, which is about a multitask problem and has a specially designed classification, which helps to reduce overfitting and make the model focus on easily confused categories. The experimental results show that the method in this paper performs well in image classification for the relatively simple Cifar-10 dataset, the moderately difficult Caltech-101 dataset, and the Caltech-256 dataset with large differences in object size and location. The fitting speed and accuracy are high.

Protocol for generating mutant zebrafish using CRISPR-Cas9 followed by quantitative evaluation of vascular formation.

The use of vascular-specific transgenic zebrafish provides advantages for identifying new mutations affecting angiogenesis and vascular development. Here, we present a protocol for establishing, screening, and phenotyping CRISPR-Cas9-based mutagenesis in fluorescently labeled transgenic zebrafish. We describe steps for designing single-guide RNA (sgRNA) oligos, synthesizing sgRNA and Cas9 mRNA, and microinjection and generation of mutant lines. We then detail procedures for visualizing dynamic vasculature and quantitatively evaluating vascular formation in transgenic zebrafish. For complete details on the use and execution of this protocol, please refer to Luo et al.1.

Photoredox-Promoted Co-Production of Dihydroisoquinoline and H2 O2 over Defective Zn3 In2 S6.

One of the most sustainable and promising approaches for hydrogen peroxide (H2 O2 ) production in a low-cost and environment-friendly way is photosynthesis, which, however, suffers from poor carrier utilization and low H2 O2 productivity. The addition of proton donors such as isopropanol or ethanol can increase H2 O2 production, which, unfortunately, will inevitably elevate the entire cost while wasting the oxidizing power of holes (h+ ). Herein, the tetrahydroisoquinolines (THIQs) is employed as a distinctive proton donor for the thermodynamically feasible and selective semi-dehydrogenation reaction to highly valuable dihydroisoquinolines (DHIQs), and meanwhile, to couple with and promote H2 O2 generation in one photoredox reaction under the photocatalysis by dual-functional Zn3 In2 S6 photocatalyst. Surprisingly, the suitably defective Zn3 In2 S6 offers an excellent and near-stoichiometric co-production performance of H2 O2 and DHIQs at unprecedentedly high rates of 66.4 and 62.1 mmol h-1 g-1 under visible light (λ ≥ 400 nm), respectively, which outperforms all the previously available reports even though sacrificial agents were employed in those reports. Additionally, photocatalytic redox reaction mechanism demonstrates that H2 O2 can be generated through multiple pathways, highlighting the synergistic effect among ROS (·O2 - and 1 O2 ), h+ and proton donor, which has been ignored in previous studies.

Nanoplastics exposure induces vascular malformation by interfering with the VEGFA/VEGFR pathway in zebrafish (Danio rerio).

The widespread accumulation and adverse effects of nanoplastics (NPs) are a growing concern for environmental and human health. However, the potential toxicological effects of nanoplastics, especially on vascular development, have not been well studied. In this study, the zebrafish model was utilized to systematically study the developmental toxicity of nanoplastics exposure at different concentrations with morphological, histological, and molecular levels. The results revealed developmental defects in zebrafish embryos after exposure to different concentrations of nanoplastics. Specifically, the morphological deformities, including pericardial oedema and spine curvature, as well as the abnormal body length and the rates of survival and hatching were induced after nanoplastics exposure in zebrafish embryos. In addition, we found that nanoplastics exposure could induce vascular malformation, including the ectopic sprouting of intersegmental vessels (ISVs), malformation of superficial ocular vessels (SOVs), and overgrowth of the common cardinal vein (CCV), as well as the disorganized vasculature of the subintestinal venous plexus (SIVP). Moreover, further study indicated that SU5416, a specific vascular endothelial growth factor receptor (VEGFR) inhibitor, partially rescued the nanoplastics exposure-impaired vasculature, suggesting that the VEGFA/VEGFR pathway might be associated with nanoplastics-induced vascular malformation in zebrafish embryos. Further quantitative polymerase chain reaction assays revealed that the mRNA levels of VEGFA/VEGFR pathway-related genes, including vegfa, nrp1, klf6a, flt1, fih-1, flk1, cldn5a, and rspo3, were altered in different groups, indicating that nanoplastics exposure interferes with the VEGFA/VEGFR pathway, thereby inducing vascular malformation during the early developmental stage in zebrafish embryos. Therefore, our findings illustrated that nanoplastics might induce vascular malformation by regulating VEGFA/VEGFR pathway-related genes at the early developmental stage in zebrafish.