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Background: Qilong capsule (QLC) is a traditional Chinese medicine commonly used to treat ischemic stroke (IS). But the pharmacodynamic mechanism of QLC is not clear. Method: we used an existing zebrafish model to explore the protective mechanism of QLC on IS. We treated normally-developing zebrafish larvae with QLC and ponatinib 2 days post fertilization, and used the area of cerebral vascular thrombosis, red blood cell staining intensity, and brain cell apoptosis to quantitate QLC efficacy against IS. Real-time quantitative polymerase chain reaction (qPCR) was used to detect changes in the expression of genes involved in coagulation, inflammation, vascular endothelium, and apoptosis. Result: We found that QLC reduced the area affected by ponatinib-induced cerebral vascular embolism, erythrocyte staining intensity, and the number of apoptotic brain cells. qPCR showed that QLC inhibited the expression of genes related to coagulation, inflammation, and apoptosis. Conclusion: QLC had a significant protective efficacy in ponatinib-induced IS.

Objective. Single-walled carbon nanotubes (SWCNTs) are able to cross the blood-brain barrier, penetrate through the cell membrane, and accumulate in the cell nucleus, which purposefully allows their use in the health sciences as imaging probes and drug carriers in the cancer therapy. The aim of this study was to investigate the effect of low doses of SWCNTs on the expression of microRNAs associated with the cell proliferation and the brain development in zebrafish (Danio rerio) embryos. Methods. The zebrafish embryos (72 h post fertilization) were exposed to low doses of SWCNTs (2 and 8 ng/ml of medium) for 24 or 72 h. The microRNAs (miR-19, miR-21, miR-96, miR-143, miR-145, miR-182, and miR-206) expression levels were measured by quantitative polymerase chain reaction analysis. Results. It was found that low doses of SWCNTs elicited dysregulation in the expression of numerous cell proliferation and brain development-related microRNAs (miR-19, miR-21, miR-96, miR-143, miR-145, miR-182, and miR-206) in dose- (2 and 8 ng/ml of medium) as well as malformations in the zebrafish embryos brain development in a time-dependent (24 and 72 h) manner. Conclusion. Taken together, the present data indicate that the low doses of SWCNTs disturbed the genome functions and reduced the miR-19, miR-21, miR-96, miR-143, miR-145, miR-182, and miR-206 expression levels in dose- and time-dependent manners and interrupted the brain development in the zebrafish embryos indicating for both the genotoxic and the neurotoxic interventions.

Threatened abortion (TA) is the most common complication in early pregnancy and is caused by anxiety and depressive symptoms. The Zishen Yutai Pill (ZYP) is a traditional herbal formula that is commonly used to treat TA. However, the pharmacological mechanisms underlying the effect of ZYP have yet to be elucidated. To disclose the mechanism of ZYP in the treatment of TA, first, we identified the chemical constituents of ZYP from multiple databases and then predicted the potential targets of TA by applying the GeneCards database. A protein-protein interaction (PPI) network was then constructed to allow the screening of hub targets. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGGs) enrichment analyses were also performed and the Database for Annotation Visualization and Integrated Discovery server was used to identify critical biological processes and signaling pathways. Cytoscape software was used to construct a Compound-Target-Pathway. Furthermore, we analyzed ZYP by UPLC-Q-TOF/MS, then the RU486-induced TA rat model was established, and the reliability of the network pharmacology prediction results was verified. Finally, the mechanisms responsible for the action of ZYP on TA were revealed by qRT-PCR and molecular docking. Database screening identified a total of 161 active compounds in ZYP and 324 TA-related targets. And, we identified 42 compounds from ZYP by UPLC-Q-TOF/MS. Inflammation and apoptosis were identified as the main biological processes. GO and KEGG analyses identified that the MAPK and PI3K/Akt pathways were the key functional pathways that respond to ZYP. The results showed that ZYP treatment significantly increased maternal weight, significantly increased the levels of estradiol and progesterone, and attenuated histopathological changes in a rat model of TA. Data indicated that ZYP treatment improved pregnancy outcomes in the rat model of TA. QRT-PCR data showed that ZYP reduced inflammation and apoptosis by regulating the MAPK and PI3K/Akt pathways. In addition, molecule docking results identified a range of key compounds, including Pik3a, Mapk14, Mapk1, Mapk3, Mapk8 Tnf, Il6, and Cas8. In summary, we performed network pharmacological analysis and experimental validation and identified that ZYP exerts an effect on TA by regulating the MAPK and PI3K/Akt pathways and by inhibiting the expression levels of proinflammatory cytokines and genes related to apoptosis.

LIBS coupled with chemometric methods reveals that Ca and Mg levels in the PTC tissues are significantly higher than those in normal ones, and it may serve as a potential technique for detection of papillary thyroid cancers.

Metal nanoclusters (NCs) have emerged as novel antibacterial agents featuring broad-spectrum antibacterial activity without drug resistance for bacteria, but suffer from fast antibacterial invalidation due to their consumption by bacteria. Herein we report the design of a visible-light-driven photodynamic antibacterial agent based on conjugating aggregation-induced emission (AIE)-featured AuAg NCs with highly luminescent carbon dots (CDs). The conjugation of CDs with AuAg NCs could not only enhance the visible-light harvest, but also promote charge carrier generation/separation

Sanguinarine, derived from the root of Sanguinaria canadensis, have multiple biological activities, such as antimicrobial, insecticidal, antitumor, anti-inflammatory and anti-angiogenesis effect, but little is known about its toxicity on normal embryonic development. Here, we study the developmental toxicity using zebrafish model. Notably, sanguinarine caused a significant increase of the malformation rate and decrease of hatching rates and body length of zebrafish embryos. Sanguinarine also impaired the normal development of heart, liver and nerve system of zebrafish embryos. Further, the ROS level and MDA concentrations were remarkably increased, while the activity of T-SOD was decreased. In addition, obvious increase of apoptosis were observed by AO staining or TUNEL assay. Further studies showed that the oxidative stress-, apoptosis-related genes were changed, while genes of nrf2 and wnt pathways were inhibited by sangunarine. To sum up, our study will be helpful to understand the adverse effect of sanguinarine on embryonic development and the underlying molecular mechanism.

Hepatitis E virus (HEV) infection is the most common cause of acute viral hepatitis worldwide. However, host-HEV interactions have yet to be fully understood. Zinc-finger antiviral protein (ZAP) is a novel interferon (IFN)-stimulated gene product that inhibits a variety of viruses in synergy with IFN-β. To evaluate the role of ZAP in HEV infection, its expressions in HEV-infected patients and in cell cultures were measured. We report a significant inhibition of ZAP expression in patients with HEV genotype four acute infection. The expression of ZAP in the HEV life cycle was monitored in cultures of HEV-infected cells. Results indicated that the ZAP level decreased significantly after HEV infection. ZAP over-expression inhibited HEV replication, whereas its knockdown by RNA interference significantly increased HEV RNA. These suggest that ZAP serves as an antiviral in HEV infection. Moreover, silencing ZAP decreased IFN regulatory factor 3 (IRF3) phosphorylation in HEV-infected cells treated with poly(I:C), indicating that ZAP synergizes with IFN-β. In conclusion, ZAP is an important anti-HEV host factor and in synergy with IFN-β, inhibits HEV replication.

Young Prunus persica fruits (YPF) contain substances that are distinct from those found in the mature fruits. Response surface methodology was used to explore the influences of extraction conditions including ultrasonic time (X1), ethanol proportion (X2), liquid-to-solid ratio (X3) and temperature (X4) on UV-absorbing components from YPF. To purify the extract, the adsorption/desorption properties of 280 nm-absorbing components on AB-8 resin were investigated. A total of 11 metabolites (amino acids, glycosylated amino acids and phenolics) were identified in the UV-absorbing fraction of YPF (YPF-F) based on LC-MS/MS assays. In a study of in vivo anti-inflammatory activity, YPF-F significantly decreased the number of inflammatory cells that migrated to the lateral line location in CuSO4-induced transgenic fluorescent zebrafish. YPF should be utilized as a high value resource of functional foods.

The clinical use of clozapine (CLZ), an atypical antipsychotic drug, was affected by side effects, such as cardiotoxicity. We selected normally developing zebrafish embryos to explore the antagonism of salvianolic acid A (SAA) against clozapine-induced cardiotoxicity. Embryos were treated with CLZ and SAA, and zebrafish phenotypes were observed at 24 h, 48 h, 72 h, and 96 h after treatment. The observed phenotypes included heart shape, heart rate, and venous sinus-arterial bulb (SV-BA) interval. Real-time quantitative PCR was used to detect changes in the expression of genes involved in heart inflammation, oxidative stress, and apoptosis. The results showed that SAA relieved pericardial edema, increased heart rate, and reduced the SV-BA interval. The PCR results also showed that when the zebrafish embryos were incubated with SAA and CLZ for 96 h, the expression of il-1b and nfkb2 were significantly downregulated, the expression of sod1 and cat were significantly upregulated, and the expressions of mcl1a and mcl1b were significantly downregulated. In summary, SAA can antagonize clozapine-induced cardiotoxicity.

Glioma is a tumor in the brain and spinal cord originating in the glial cells that surround the nerve cells. Among several microRNAs reported, miRNA-363 is associated with human glioma. Based on miRNA-363 levels, the development and progression of glioma can be monitored. The current study used an interdigitated electrode sensor to monitor microRNA-363 levels, which indeed reflects the severity of glioma. The interdigitated electrode was generated using a photolithography technique followed by surface chemical modification carried out to insert miRNA-363 complementary oligo as the probe complexed with gold nanoparticles. The proposed sensor works based on the dipole moment between two electrodes, and when molecular immobilization or interaction occurs, the response by the signal output changes. The changes in the target microRNA-363 sequence were standardized to identify glioma. The limit of detection of miRNA-363 was 10 fM with an R

Clozapine is one of the antipsychotic drugs for treating schizophrenia, but its cardiotoxicity was the primary obstacle for its clinical use, due to the unknown mechanism of clozapine-induced cardiotoxicity. In this study, we studied the cardiotoxicity of clozapine by employing zebrafish embryos. Acute clozapine exposure showed dose-dependent mortality with the LC50 at 59.36 μmol L−1 and 49.60 μmol L−1 when determined at 48 and 72 h post exposure, respectively. Morphological abnormalities like pericardial edema, incompletely heart looping, and bradycardia were detected after clozapine exposure in a time- and dose-dependent manner. Clozapine treatment also resulted in a slower heart rate and disturbed rhythm in zebrafish embryos. Also, oxidative stress was observed after clozapine exposure by measurement of ROS (reactive oxygen species), MDA (a lipid peroxidation marker), antioxidant enzyme activities, and oxidative stress-related gene expression. The elevation of inflammation coincided with oxidative stress by the assay of inflammation-related genes expression accompanied by clozapine incubation. Collectively, the data indicate that clozapine might achieve cardiotoxic effect in zebrafish larva through increasing oxidative stress, attenuation in antioxidant defense, and up-regulation of inflammatory cytokines. The data could provide experimental explanations for myocarditis and pericarditis induced by clozapine in clinics, and help find an effective solution to reduce its cardiotoxicity.

The SKU5 similar (SKS) genes encode a family of multi-copper-oxidase-like proteins with cupredoxin domains similar to those in laccase and ascorbate oxidase. Although SKS proteins are known to function in root growth and cotyledon vascular patterning in Arabidopsis, their role in plant reproductive processes is poorly understood. Here, we identified a seed mutant of maize (Zea mays), generated by ethyl methane sulfonate (EMS) mutagenesis, that we designated defective kernel-zk1 (dek-zk1). The mutant produced small, shriveled kernels with an aberrant basal endosperm transfer layer (BETL) and placento-chalazal (PC) layer and irregular starch granules. Map-based cloning revealed that Dek-zk1 encodes an SKU5 similar 13 (GenBank: ONM36900.1), so it was named ZmSKS13. ZmSKS13 comprises a paralogous pair with Zm00001d012524, but the transcript abundance of ZmSKS13 in developing kernels is 15 times higher than that of Zm00001d012524, resulting in dek-zk1 mutation conveying a distinct kernel phenotype. ZmSKS13 loss of function led to overaccumulation of reactive oxygen species (ROS) and severe DNA damage in the nucellus and BETL and PC layer cells, and exogenous antioxidants significantly alleviated the defects of the mutant kernels. Our results thus demonstrate that ZmSKS13 is a novel regulator that plays a crucial role in kernel development in maize through the modulation of ROS homeostasis.

Introduction Macleaya cordata (Willd) R. Br. (Papaveraceae family) is a well-known traditional Chinese medicine used to treat muscle pain, inflamed wounds, and bee bites. Benzo[c]phenanthridine alkaloids are the main active ingredients in M. cordata. In this work, sanguinarine and chelerythrine were efficiently extracted and purified by ultrahigh-pressure extraction (UHPE) technique and pH-zone-refining counter-current chromatography (PZRCCC) from M. cordata. Objective To develop an efficient UHPE method followed by an efficient separation technique using PZRCCC for benzo[c]phenanthridine alkaloids from the study plant species, and to evaluate the study samples for anti-breast cancer activity. Methodology The optimal extraction conditions were optimised as extraction pressure 200 MPa, extraction solvent 95% ethanol, solid-liquid ratio 1:30 (g/mL) and extraction time 2 min. A two-phase n-hexane/ethyl acetate/i-propanol/water (1:3:1.5:4.5, v/v) solvent system was optimised with 10 mmol triethylamine in the upper phase and 10 mmol trifluoroacetic acid in lower phase in PZRCCC. The sample loading was optimised as 2.50 g. Moreover, the samples were evaluated for anti-breast cancer activity later on. Results The 2.50 g sample loading yielded 0.45 g of sanguinarine and 0.59 g chelerythrine in one-step separation using PZRCCC. The anti-breast cancer activities of sanguinarine and chelerythrine were found stronger than positive control (vincristine 5.04 μg/mL) with half-maximal inhibitory concentration values of 0.96 and 3.00 μg/mL, respectively. Conclusion This study showed that the established methods were efficient in extraction (UHPE) and separation (PZRCCC) of the sanguinarine and chelerythrine from M. cordata.

Objective. The aim of the present investigation was to study the expression of genes encoding homeobox proteins ZEB2 (zinc finger E-box binding homeobox 2), TGIF1 (TGFB induced factor homeobox 1), SPAG4 (sperm associated antigen 4), LHX1 (LIM homeobox 1), LHX2, LHX6, NKX3-1 (NK3 homeobox 1), and PRRX1 (paired related homeobox 1) in U87 glioma cells in response to glucose deprivation in control glioma cells and cells with knockdown of ERN1 (endoplasmic reticulum to nucleus signaling 1), the major pathway of the endoplasmic reticulum stress signaling, for evaluation of it possible significance in the control of glioma growth through ERN1 signaling and chemoresistance. Methods. The expression level of homeobox family genes was studied in control (transfected by vector) and ERN1 knockdown U87 glioma cells under glucose deprivation condition by real-time quantitative polymerase chain reaction. Results. It was shown that the expression level of ZEB2, TGIF1, PRRX1, and LHX6 genes was up-regulated in control glioma cells treated by glucose deprivation. At the same time, the expression level of three other genes (NKX3-1, LHX1, and LHX2) was down-regulated. Furthermore, ERN1 knockdown of glioma cells significantly modified the effect glucose deprivation condition on the expression almost all studied genes. Thus, treatment of glioma cells without ERN1 enzymatic activity by glucose deprivation condition lead to down-regulation of the expression level of ZEB2 and SPAG4 as well as to more significant up-regulation of PRRX1 and TGIF1 genes. Moreover, the expression of LHX6 and NKX3-1 genes lost their sensitivity to glucose deprivation but LHX1 and LHX2 genes did not change it significantly. Conclusions. The results of this investigation demonstrate that ERN1 knockdown significantly modifies the sensitivity of most studied homeobox gene expressions to glucose deprivation condition and that these changes are a result of complex interaction of variable endoplasmic reticulum stress related and unrelated regulatory factors and contributed to glioma cell growth and possibly to their chemoresistance.

Background: Zishen Guchong Pill (ZGP), a traditional oriental herbal medicine, has been applied broadly in the treatment of threatened or recurrent miscarriages in clinics, but little is known about its developmental toxicity. Aim of the study: This study aimed to investigate the potential toxicity of ZGP and its mechanisms. methods: We investigated the developmental toxicity of ZGP using a zebrafish model. Zebrafish embryos at 4 hours post-fertilization (hpf) were exposed to 25, 50, 100, and 200 µg/mL ZGP until 120 hpf. The zebrafish morphology, organ development, reactive oxygen species (ROS) content, oxidative stress-related enzyme activity, and mRNA levels of oxidative stress and apoptosis-related genes were measured. Results: Our results demonstrated that ZGP had no toxicity at 50 µg/mL ZGP but induced phenotypic defects and decreased the hatching rate and body length at more than 50 µg/mL ZGP (high dose). High doses of ZGP caused severe heart failure, including a significant increase in the sinus venosus (SV) and bulbus arteriosus (BA) distance and pericardial area, and reduced heart rate and numbers of red blood cells. In addition, high doses of ZGP caused liver atrophy and decreased the length of dopamine ganglia and neurovascular. High doses of ZGP elicited the generation of ROS and elevated malondialdehyde levels, but reduced the activity of superoxide dismutase (SOD) and catalase (CAT). Real-time PCR data showed the downregulation of oxidative stress-related genes (sod and cat) and the activation of Nrf2 (nrf2, keap1, ho-1, nqo1, gclc, and gclm) and P53 (p53, bcl-2, and bax) signal pathway genes after high ZGP exposure. Conclusions: Taken together, our results indicate that ZGP has no developmental toxicity under 50 µg/mL ZGP but induced developmental toxicity in zebrafish embryos at more than 50 µg/mL ZGP, and oxidative stress contributed to the toxic response. • ZGP is safe and reliable in clinical use. • ZGP (high does) caused an increase in ROS, which induced oxidative stress, leading to apoptosis and developmental toxicity. • The Nrf2 signaling pathway is impaired during the early stages of ZGP-induced developmental toxicity.

Activator of heat shock protein 90 (hsp90) ATPase (Aha1) is a Hsp90 co-chaperone required for Hsp90 ATPase activation. Aha1 is essential for yeast survival and muscle development in C. elegans under elevated temperature and hsp90-deficeiency induced stress conditions. The roles of Aha1 in vertebrates are poorly understood. Here, we characterized the expression and function of Aha1 in zebrafish. We showed that zebrafish genome contains two aha1 genes, aha1a and aha1b, that show distinct patterns of expression during development. Under the normal physiological conditions, aha1a is primarily expressed in skeletal muscle cells of zebrafish embryos, while aha1b is strongly expressed in the head region. aha1a and aha1b expression increased dramatically in response to heat shock induced stress. In addition, Aha1a-GFP fusion protein exhibited a dynamic translocation in muscle cells in response to heat shock. Moreover, upregulation of aha1 expression was also observed in hsp90a1 knockdown embryos that showed a muscle defect. Genetic studies demonstrated that knockout of aha1a, aha1b or both had no detectable effect on embryonic development, survival, and growth in zebrafish. The aha1a and aha1b mutant embryos showed normal muscle development and stress response in response to heat shock. Single or double aha1a and aha1b mutants could grow into normal reproductive adults with normal skeletal muscle structure and morphology compared with wild type control. Together, data from these studies indicate that Aha1a and Aha1b are involved in stress response. However, they are dispensable in zebrafish embryonic development, growth, and survival.

Hepatitis E virus (HEV) is the major pathogen of viral hepatitis. However, the understanding of the HEV life cycle is limited. In the present study, cells were separately infected with non-enveloped HEV (derived from feces or bile) or quasi-enveloped HEV (derived from the cell culture after serial passages, eHEV) and observed by confocal fluorescence microscopy to investigate the life cycle of HEV. HEV finished its binding and entry into host cells at first 6 h post-inoculation (hpi). Cells inoculated with eHEV showed less infectivity than cells inoculated with non-enveloped HEV. Newly synthesized progeny virions were released into the supernatant of cell cultures from 48 hpi. qRT-PCR and Western blotting results showed that the supernatant's progeny viruses were infectious even after 5 serial passages. These results show the significant difference between non-enveloped HEV and eHEV, which will provide novel insights into the HEV replication cycle. The efficient cell culture of HEV will promote the development of anti-HEV drugs and vaccines. This article is protected by copyright. All rights reserved.

The Traditional Chinese Medicine formula Fufang Kushen Injection (FKI) has demonstrated potential to enhance the efficacy and reduce the toxicity of the chemotherapeutic drug cisplatin. However, there is insufficient evidence to determine whether the combination of matrine and cisplatin were linked to the angiogenesis pathway. In this study, we selected two zebrafish lines, AB and Tg (vegfr2: GFP), as in vivo models to rapidly assess the anti-angiogenesis effects. KFI and cisplatin had no obvious effects when used individually, but combined KFI (5 and 10 μL/mL) and cisplatin (50μg/mL) significantly inhibited the zebrafish intersegmental vessel (ISV) formation and growth. Matrine at 50 μg/mL also showed synergetic anti-angiogenesis activity with cisplatin (50μg/mL) in 48hpf zebrafish larvae. This study has shown the potential of FKI to enhance cisplatin efficacy and reduce its toxicity by inhibiting angiogenesis. These results contribute to the scientific evidence supporting the use of KFI in combination with cisplatin to treat cancer in the clinic.

In this study, we investigated the metabonomic profiles of Apostichopus japonicus using an LC-MS-based method in conjunction with multivariate data analysis. Based on the PLS-DA model, 85 differential metabolites (VIP value >1.0) were obtained from viscera and body wall samples. The MS/MS and NMR experiments were used for the qualitative identification of the characteristic peaks. Sphingoid-based nucleoside analogues were the main components in Chinese A. japonicus viscera. Our findings demonstrate that A. japonicus viscera contain a large number of compounds that may have applications as nutraceuticals or pharmaceuticals.

Seizures are sustained neuronal hyperexcitability in brain that result in loss of consciousness and injury. Understanding how the brain responds to seizures is critical to help developing new therapeutic strategies for epilepsy, a neurological disorder characterized by recurrent and unprovoked seizures. However, the mechanisms underlying seizure-dependent alterations of biological properties are poorly understood. In this study, we analyzed gene expression profiles of the zebrafish heads that were undergoing seizures and identified 1776 differentially expressed genes. Gene-regulatory network analysis revealed that BDNF-TrkB signaling pathway positively regulated brain inflammation in zebrafish during seizures. Using K252a, a TrkB inhibitor to block BDNF-TrkB signaling pathway, attenuated pentylenetetrazole (PTZ)-induced seizures, which also confirmed BDNF-TrkB mediated inflammatory responses including regulation of il1β and nfκb, and neutrophil and macrophage infiltration of brain. Our results have provided novel insights into seizure-induced brain inflammation in zebrafish and anti-inflammatory related therapy for epilepsy.

Marine phospholipids (MPLs) are valuable components that can be applied within diverse areas like nutrition, pharmacy, and medicine as well as basic scientific research, particularly due to their high concentration of long chain polyunsaturated fatty acids. In this work, lipidomics approach by UPLC-Q-Exactive Orbitrap/MS was used for the identification, quantification, comparison, and characterization of phospholipids (PLs) from three different marine resources (shrimp head, codfish roe, and squid gonad). In total, 310 PL molecular species containing 34 different structures of fatty acid chains were identified simultaneously by Lipidsearch software. Significant differences between three groups in the PL classes, compositions and contents were revealed. A list of characteristic PL species that represent significant differences among groups was determined by lipidomics analysis. Until now, the information about the deep comprehensive description of PL from marine sources is limited. Thus, this study will give a very potential starting point to develop MPL products and establish the quality standards for different marine raw materials.

Wenlong Sheng,1 Shijun Weng,2 Fei Li,1 Yun Zhang,1 Qiuxia He,1 Wenxiang Sheng,1 Ying Fu,3 Haiyue Yan,4 Kechun Liu1 1Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China; 2State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Department of Ophthalmology, Zhongshan Hospital, Fudan University, Shanghai, People’s Republic of China; 3Shandong Science and Technology Exchange Center, Jinan, People’s Republic of China; 4Shandong Institute of Scientific and Technical Information, Jinan, People’s Republic of ChinaCorrespondence: Wenlong Sheng; Kechun Liu Email shengwenlong1121@163.com; liukechun2000@163.comBackground: Melatonin (N-acetyl-5-methoxytryptamine), a significant indoleamine neuromodulator implicated in circadian rhythms and sleep patterns, regulates diverse rhythmic functions via activating its high-affinity G-protein-coupled receptors. However, the detailed cellular expression of the Mel1a receptor in the retina is still a research gap.Methods: The expression of the Mel1a receptor in pigeon retina was assessed using Western blot analysis and immunofluorescent staining. The cellular localization of the Mel1a receptor was studied using double immunofluorescent staining and laser-scanning confocal microscopy.Results: Our data suggested that the Mel1a receptor was extensively expressed in the outer segment of Rho4D2-labeled rod and L/M-opsin-labeled red/green cone and in the somata of the CB-labeled horizontal cell, TH-labeled dopaminergic amacrine cell, ChAT-labeled cholinergic amacrine cell, PV-labeled AII amacrine cell, Brn3a-labeled conventional ganglion cell, melanopsin-containing ganglion celland CRALBP-labeled Müller glial cell. In addition, the Mel1a receptor was diffusely distributed throughout the full thickness of the inner plexiform layer. However, the outer segment of S-opsin-labeled blue cone, the somata of ChX-10-labeled bipolar cell and outer plexiform layer seemed to lack immunoreactivity of the Mel1a receptor.Conclusion: The finding that multiple types of retinal cells express the Mel1a receptor provides a new neurobiological basis for the participation of melatonin in the regulation of retinal functions through activating the Mel1a receptor.Keywords: circadian rhythm, retinal Mel1a receptor, cellular localization, diurnal animal

Nano-titanium nitride (Nano-TiN) has strong resistance to wear and corrosion, good biocompatibility, and an attractive metallic luster. Nano-TiN is widely used in medical devices, such as orthopedic implants, syringe needles, coronary stents, and long-term dental implants, and also in imitation gold jewelry. Despite its widespread use, there are few reports describing safety evaluations of Nano-TiN. Here, we exposed healthy zebrafish embryos to different concentrations of Nano-TiN solution for five days, starting at about four hours post fertilization, and found that Nano-TiN caused dose- and time-dependent developmental toxicity. With increasing Nano-TiN concentration and length of exposure, mortality, and deformities gradually increased; body length shortened and hatching rate and motility were significantly reduced. We also found that exposure to Nano-TiN affected development of the heart, liver, nerves, and other organs, and led to elevated levels of reactive oxygen species and reduced antioxidant capacity. Exposure to Nano-TiN resulted in downregulation of expression of antioxidant genes, such as

Macleaya cordata (Willd) R. Br. (Papaveraceae family) is a well-known traditional Chinese medicine used to treat muscle pain, inflamed wounds, and bee bites. Benzo[c]phenanthridine alkaloids are the main active ingredients in M. cordata. In this work, sanguinarine and chelerythrine were efficiently extracted and purified by ultrahigh-pressure extraction (UHPE) technique and pH-zone-refining counter-current chromatography (PZRCCC) from M. cordata.To develop an efficient UHPE method followed by an efficient separation technique using PZRCCC for benzo[c]phenanthridine alkaloids from the study plant species, and to evaluate the study samples for anti-breast cancer activity.The optimal extraction conditions were optimised as extraction pressure 200 MPa, extraction solvent 95% ethanol, solid-liquid ratio 1:30 (g/mL) and extraction time 2 min. A two-phase n-hexane/ethyl acetate/i-propanol/water (1:3:1.5:4.5, v/v) solvent system was optimised with 10 mmol triethylamine in the upper phase and 10 mmol trifluoroacetic acid in lower phase in PZRCCC. The sample loading was optimised as 2.50 g. Moreover, the samples were evaluated for anti-breast cancer activity later on.The 2.50 g sample loading yielded 0.45 g of sanguinarine and 0.59 g chelerythrine in one-step separation using PZRCCC. The anti-breast cancer activities of sanguinarine and chelerythrine were found stronger than positive control (vincristine 5.04 μg/mL) with half-maximal inhibitory concentration values of 0.96 and 3.00 μg/mL, respectively.This study showed that the established methods were efficient in extraction (UHPE) and separation (PZRCCC) of the sanguinarine and chelerythrine from M. cordata.

The small 295-bp ZmPht1; 5 promoter is sufficient to drive high-intensity expression of target genes, especially under phosphate deprivation conditions, and is therefore useful for crop improvement via transgenic techniques. Phosphate (Pi) deficiency has become a major challenge and limiting factor in world agricultural production. Manipulating the gene expression using appropriate promoters to improve the Pi absorption and utilization efficiency of crops could reduce the requirement for Pi fertilizers. In the study, a 295-bp strong promoter (M2P-7) of maize high-affinity phosphate transporter ZmPht1; 5 was isolated and functionally validated in transgenic Nicotiana benthamiana and maize by analyzing the ZmPht1; 5 promoter (M2P-1) and its 5' truncated variants (M2P-2 ~ M2P-8) in different sizes under normal and Pi-deprivation conditions. The M2P-7 displayed the highest promoter activities among 5' truncated fragments in all tested tissues of transgenic Nicotiana benthamiana at different development stages, which was 1.5 and 3 times higher than the well-used CaMV35S promoter under normal and Pi-deprivation conditions, respectively. In maize, the M2P-7 promoter activity was comparable to the maize ubiquitin1 promoter widely used in monocots under normal condition, which was about 1.3 times that of the ubiquitin1 promoter under Pi-deprivation environments. Moreover, the M2P-7 fragment is only 295 bp in length, thus reducing the construct size, and is therefore beneficial for genetic transformation. Thus, the small promoter M2P-7 of plant origin could be of great use for monocotyledonous and dicotyledonous crop improvement via transgenic techniques based on its promoter activities, expression patterns and small size.

Sea cucumbers, soft-bodied wormlike echinoderms, are traditionally consumed in China. Sea cucumber viscera are often discarded during production. In this study, we characterized saponins present in the viscera of the Chinese sea cucumber Apostichopus japonicus. Sea cucumber saponins were obtained by a two-stage process consisting of foam fractionation and macroporous resin separation. Furthermore, the saponins were characterized by HPLC-Q-TOF-MS. Three novel saponins, i.e., apostichoposide A–C, were detected and characterized based on their unique fragmentation patterns in the negative ion mode tandem MS spectra as well as isolation and NMR studies. The abundance of saponins in sea cucumber viscera holds promise for industrial applications.

Isopsoralen is the main component of the Chinese medicine psoralen, which has antitumour activity and can be used for the treatment of osteoporosis. However, the mechanism behind its hepatotoxicity has not yet been elucidated. In this study, the hepatotoxicity of isopsoralen was investigated using zebrafish. Isopsoralen treatment groups of 25, 50 and 100 μM were established. The mortality, liver morphology changes, levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), liver histopathology and mRNA levels of liver injury–related genes in zebrafish larvae were measured. The results showed that isopsoralen resulted in the development of malformed zebrafish, dose-dependent increases in ALT and AST, decreased liver fluorescence and weakened fluorescence intensity. Histopathological examination showed that high-dose isopsoralen caused a large number of vacuolated structures in the larvae liver. The polymerase chain reaction results showed a significant decrease in the mRNA levels of genes related to antioxidant capacity ( lfabp, gstp2 and sod1) and drug transport ( mdr1, mrp1 and mrp2), indicating that isopsoralen significantly inhibited liver antioxidant capacity and drug efflux capacity in zebrafish larvae. Isopsoralen is hepatotoxic to zebrafish larvae via inhibition of drug transporter expression resulting in the accumulation of isopsoralen in the body and decreased antioxidant capacity, leading to liver injury.

The sensitive and accurate detection of intracellular gasotransmitter carbon monoxide (CO) is of great significance for unraveling its diverse cellular functions. Herein, we report a simple highly specific and sensitive 4-hydroxynaphthalimide-based ratiometric fluorescent probe Ratio-CO for monitoring CO levels in living cells. Probe Ratio-CO could quantitatively detect CO in the range of 0–50 μM by the ratiometric fluorescence spectroscopy method and the detection limit was measured to be about 17.9 nM. Additionally, probe Ratio-CO has been proven to possess preeminent selectivity towards CO over other bioactive species. Most importantly, the excellent response properties of probe Ratio-CO enable its applications in the monitoring of the fluctuations of CO levels in living cells by the ratiometric fluorescence imaging technique, and we thus anticipate that this probe would be a novel tool for further elucidating the biological functions of CO in living systems.