Thrombosis inhibited by Corydalis decumbens through regulating PI3K-Akt pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Corydalis decumbens (Thunb.) Pers. was used as stasis-eliminating medicine traditionally to treat cardiovascular disease potentially attributed to its antithrombotic effect, but lack of pharmacological research on it. AIM OF THE STUDY: To investigate the antithrombotic effect of C. decumbens and its preliminary mechanism. MATERIALS AND METHODS: A carrageenan-induced mouse thrombus model and adenosine diphosphate stimulated platelet aggregation of rabbits were used to confirm the inhibitory effect of C. decumbens extract and compounds on thrombosis in vivo. Then, H2O2-induced human umbilical vein endothelial cells (HUVECs) injury model was further adopted to verify the effects of bioactive compounds in vitro. Moreover, in silico network pharmacology analyses and molecular docking were performed to predict the underlying mechanisms, targets, and pathways, and which were further confirmed through western blotting assay. RESULTS: The administration of total extract (TE), total alkaloids (TA) and tetrahydropalmatine (TET) resulted in a significant reduction in black tail thrombus and congestion, along with a decreasing in platelet aggregation of rabbits. A superior antithrombotic effect indicated the bioactive fraction, and then the isolated bioactive compounds, TET and protopine (PRO) increased cell survival, and decreased reactive oxygen species (ROS) and lactate dehydrogenase (LDH) release in H2O2-induced HUVECs injury model. Moreover, the two alkaloids targeted 33 major proteins and influenced 153 pathways in network pharmacology prediction. Among these, HSP90AA1, COX-2, NF-κB/p65, MMP1 and HIF-1α were the key proteins and PI3K-Akt emerged as the major signaling pathway. Further western blotting results supported that five key proteins were downregulated by the two bioactive compounds in H2O2-stimulated HUVECs model. CONCLUSION: C. decumbens exerted protective effect on thrombosis through inhibiting PI3K-Akt pathway and related key proteins, which supported the traditional use and presented potential antithrombotic alkaloids for further investigation.

Co-delivery of Paclitaxel/Atovaquone/Quercetin to regulate energy metabolism to reverse multidrug resistance in ovarian cancer by PLGA-PEG nanoparticles.

Ovarian cancer is a malignant tumor that seriously endangers the lives of women, with chemotherapy being the primary clinical treatment. However, chemotherapy encounters the problem of generating multidrug resistance (MDR), mainly due to drug efflux induced by P-glycoprotein (P-gp), which decreases intracellular accumulation of chemotherapeutic drugs. The drugs efflux mediated by P-gp requires adenosine triphosphate (ATP) hydrolysis to provide energy. Therefore, modulating energy metabolism pathways and inhibiting ATP production may be a potential strategy to reverse MDR. Herein, we developed a PTX-ATO-QUE nanoparticle (PAQNPs) based on a PLGA-PEG nanoplatform capable of loading the mitochondrial oxidative phosphorylation (OXPHOS) inhibitor atovaquone (ATO), the glycolysis inhibitor quercetin (QUE), and the chemotherapeutic drug paclitaxel (PTX) to reverse MDR by inhibiting energy metabolism through multiple pathways. Mechanistically, PAQNPs could effectively inhibit the OXPHOS and glycolytic pathways of A2780/Taxol cells by suppressing the activities of mitochondrial complex III and hexokinase II (HK II), respectively, ultimately decreasing intracellular ATP levels in tumor cells. Energy depletion can effectively inhibit cell proliferation and reduce P-gp activity, increasing the chemotherapeutic drug PTX accumulation in the cells. Moreover, intracellular reactive oxygen species (ROS) is increased with PTX accumulation and leads to chemotherapy-resistant cell apoptosis. Furthermore, PAQNPs significantly inhibited tumor growth in the A2780/Taxol tumor-bearing NCG mice model. Immunohistochemical (IHC) analysis of tumor tissues revealed that P-gp expression was suppressed, demonstrating that PAQNPs are effective in reversing MDR in tumors by inducing energy depletion. In addition, the safety study results, including blood biochemical indices, major organ weights, and H&E staining images, showed that PAQNPs have a favorable in vivo safety profile. In summary, the results suggest that the combined inhibition of the two energy pathways, OXPHOS and glycolysis, can enhance chemotherapy efficacy and reverse MDR in ovarian cancer.

An immunochromatographic test for serological diagnosis of scrub typhus.

A fluorescent immunochromatographic test (FM-ICT) was developed for rapid detection of anti-Orientia tsutsugamushi antibodies in serum samples. The FM-ICT was constructed based on the dual-antigen sandwich method. Truncated 56 kDa outer membrane protein of O. tsutsugamushi strain SJ, was expressed in E. coli and mixed with those of Ptan and Gillam strains. A thin line of the protein mixture was precisely sprayed across a nitrocellulose membrane making this the "Test" line. Polyclonal antibodies (pAbs) to O.tsutsugamushi were sprayed in another line across the membrane making this the "Control" line. Fluorescent microspheres conjugated 56 kDa proteins reacting with sample serum will be captured on the "Test" line if the sample contains antibodies to O.tsutsugamushi. Several experimental parameters were optimized. After optimizing the reaction procedure, the results are visible, within 6 min, with the naked eye under ultraviolet light. The limit of detection (LOD) was determined to be 7.63 ng/mL with prepared polyclonal antibodies. No cross-reaction was observed with sera samples from other febrile diseases. In clinical evaluations, the strips showed 94.92% sensitivity (106/112) and 93.75% specificity (56/60). The FM-ICT we developed will provide a new tool for on-site diagnosis of scrub typhus.

The whitening effect of cuscutin responsible for traditional use of Bergenia purpurascens.

ETHNOPHARMACOLOGICAL RELEVANCE: The roots and rhizomes of Bergenia purpurascens (Hook. f. et Thomson) Engl., was used as a sunscreen to protect against ultraviolet rays in Tibet of China historically, but its skin whitening constituents and pharmacological effects of this plant remained unknown. AIM OF THE STUDY: To investigate the anti-melanogenesis effect of B. purpurascens in vitro and in vivo, and then explore the preliminary mechanism. MATERIALS AND METHODS: An ultraviolet B (UVB)-induced skin injury model of mice was used to verify the ameliorative effect of B. purpurascens extract (BPE) on ultraviolet damage. Then, alpha-melanocyte stimulating hormone (α-MSH)-induced murine melanoma cell line (B16F10) melanin generation model was further adopted to approval the effects of BPE and its bioactive compound, cuscutin, in vitro. Moreover, α-MSH stimulated melanogenesis model in zebrafish was employed to confirm the anti-pigmentation effect of cuscutin. Then, proteins expressions associated with melanin production were observed using western blotting assay to explore preliminary mechanism. RESULTS: BPE inhibited UVB-induced mice injury and restored skin barrier function observably in vivo. BPE and cuscutin suppressed the overproduction of melanin in α-MSH induced B16F10 significantly, in which cuscutin exhibited better effect than well-known whitening agent α-arbutin at same 10 µg/mL concentration. Moreover, the pigmentation of zebrafish embryo was decreased by cuscutin. Finally, cuscutin showed significant downregulation of expressions of tyrosinase (TYR) and tyrosinase related protein-1 (TRP-1), TRP-2 and microphthalmia-associated transcription factor (MITF) in the melanogenic signaling pathway. CONCLUSION: B. purpurascens extract and its major bioactive constituent, cuscutin, showed potent anti-melanogenesis and skin-whitening effect by targeting TYR and TRP-2 proteins for the first time, which supported its traditional use.

A rapid colloidal gold immunochromatographic assay based on polyclonal antibodies against HtpsC protein for the detection of Streptococcus suis.

An efficient and rapid immunochromatographic assay (ICA) has been engineered for the detection of Streptococcus suis (S. suis). The underpinning principle of this ICA test lies in the use of polyclonal antibodies (pAbs) decorated with colloidal gold, which are specific to S. suis. These pAbs were derived from rabbits immunized with type II histidine triad protein (HtpsC) and HtpsC-N of S. suis. The sensitivity of the ICA was noteworthy, identifying S. suis at bacterial concentrations as diminutive as 1.0 × 103 CFU/mL. Moreover, the assay demonstrated respectable specificity and did not indicate false positives for other bacterial species (Escherichia coli, Salmonella, Staphylococcus aureus, Listeria monocytogenes, Streptococcus pyogenes, Streptococcus lactis, or Enterococcus faecalis). The assay was also capable of detecting multiple S. suis serotypes containing the htpsC gene, including serotypes 1-9, 12, 14, 16 and 23. Nonetheless, the detection of S. suis that lacks the htpsC gene remained beyond the capabilities of this assay. A simultaneous analysis of 16 samples utilizing PCR substantiated the reliability of the ICA test. The assay's results can be procured within a 15-min window, making it a suitable option for field application. Broadly, this study underscores the potential of the HtpsC protein as a target antigen for the detection of S. suis, and proposes that the HtpsC protein be evaluated further in other detection assays specific for S. suis.

The promoting effects of pyriproxyfen on autophagy and apoptosis in silk glands of non-target insect silkworm, Bombyx mori.

Pyriproxyfen is a juvenile hormone analogue. The physiological effects of its low-concentration drift during the process of controlling agricultural and forestry pests on non-target organisms in the ecological environment are unpredictable, especially the effects on organs that play a key role in biological function are worthy of attention. The silk gland is an important organ for silk-secreting insects. Herein, we studied the effects of trace pyriproxyfen on autophagy and apoptosis of the silk gland in the lepidopteran model insect, Bombyx mori (silkworm). After treating fifth instar silkworm larvae with pyriproxyfen for 24 h, we found significant shrinkage, vacuolization, and fragmentation in the posterior silk gland (PSG). In addition, the results of autophagy-related genes of ATG8 and TUNEL assay also demonstrated that autophagy and apoptosis in the PSG of the silkworm was induced by pyriproxyfen. RNA-Seq results showed that pyriproxyfen treatment resulted in the activation of juvenile hormone signaling pathway genes and inhibition of 20-hydroxyecdysone (20E) signaling pathway genes. Among the 1808 significantly differentially expressed genes, 796 were upregulated and 1012 were downregulated. Among them, 30 genes were identified for autophagy-related signaling pathways, such as NOD-like receptor signaling pathway and mTOR signaling pathway, and 30 genes were identified for apoptosis-related signaling pathways, such as P53 signaling pathway and TNF signaling pathway. Further qRT-PCR and in vitro gland culture studies showed that the autophagy-related genes Atg5, Atg6, Atg12, Atg16 and the apoptosis-related genes Aif, Dronc, Dredd, and Caspase1 were responsive to the treatment of pyriproxyfen, with transcription levels up-regulated from 24 to 72 h. In addition, ATG5, ATG6, and Dronc genes had a more direct response to pyriproxyfen treatment. These results suggested that pyriproxyfen treatment could disrupt the hormone regulation in silkworms, promoting autophagy and apoptosis in the PSG. This study provides more evidence for the research on the damage of juvenile hormone analogues to non-target organisms or organs in the environment, and provides reference information for the scientific and rational use of juvenile hormone pesticides.

New Monoterpenoid Indole Hybrids from Gelsemium elegans with Anti-Inflammatory and Osteoclast Inhibitory Activities.

Gelsegansymines A (1) and B (2), two new indole alkaloids along with six known analogues (3-8) were isolated from the aerial parts of Gelsemium elegans. Their structures were elucidated by means of spectroscopic techniques. Structurally, compounds 1 and 2 possessed the rare cage-like gelsedine skeleton hybrid with bicyclic monoterpenoid. The anti-inflammatory activities of isolated compounds (1-3) were tested on LPS induced RAW264.7 cells. Under the treated concentration without toxicity for cells, the cytokines levels of nitric oxide (NO), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were evaluated by Griess method and enzyme-linked immunosorbent assay (ELISA). The results showed that compounds 1-3 exhibited anti-inflammatory activities with dose-dependent manner range from 12.5 to 50 µmol/L. Furthermore, the inhibitory activities of compounds 1 and 2 on receptor activator of NF-κB ligand (RANKL) induced osteoclast formation were tested in vitro. Compounds 1 and 2 at 5 µmol/L exhibited the significant inhibitory effect on the osteoclastogenesis induced by RANKL. This work reported the anti-inflammatory and osteoclast inhibitory activities of new monoterpenoid indole hybrids, which may inspire the further light on the related traditional application research of G. elegans.

Molecular characterization of MyD88 as a potential biomarker for pesticide-induced stress in Bombyx mori.

The widespread use of pesticides hampers the immune system of non-target organisms, however, there is a lack of common biomarkers to detect such effects. Myeloid differentiation primary response factor 88 (MyD88) is a crucial junction protein in the Toll-like receptor signaling pathway, which plays an important role in the inflammatory response. In this study, we investigated MyD88 as a potential biomarker for pesticide-induced stress. Phylogenetic analysis revealed that MyD88 was a conserved protein in the evolution of vertebrates and invertebrates. MyD88s usually have death domain (DD) and Toll/interleukin-1 receptor (TIR) domain. Bombyx mori (B. mori) is an important economic insect that is sensitive to toxic substances. We found microbial pesticides enhanced the expression level of MyD88 in B. mori. Transcriptome analysis demonstrated that MyD88 expression level was increased in the fatbody after dinotefuran exposure, a third-generation neonicotinoid pesticide. Moreover, the expression of MyD88 was upregulated in fatbody and midgut by imidacloprid, a first-generation neonicotinoid pesticide. Additionally, insect growth regulator (IGR) pesticides, such as methoprene and fenoxycarb, could induce MyD88 expression in the fatbody of B. mori. These results indicated that MyD88 is a potential biomarker for pesticide-induced stress in B. mori. This study provides novel insights into screening common biomarkers for multiple pesticide stresses and important implications for the development of more sustainable pest management strategies.

Dinotefuran induces oxidative stress and autophagy on Bombyx mori silk gland: Toxic effects and implications for nontarget organisms.

Dinotefuran, a third-generation neonicotinoid insecticide, is widely utilized in agriculture for pest control; however, its environmental consequences and risks to non-target organisms remain largely unknown. Bombyx mori is an economically important insect and a good toxic detector for environmental assessments. In this study, ultrastructure analysis showed that dinotefuran exposure caused an increase in autophagic vesicles in the silk gland. Dinotefuran exposure triggered elevated levels of oxidative stress in silk glands. Reactive oxygen species, oxidized glutathione disulfide, glutathione peroxidase, the activities of UDP glucuronosyl-transferase and carboxylesterase were induced in the middle silk gland, while malondialdehyde, reactive oxygen species, superoxide dismutase ， oxidized glutathione disulfide were increased in the posterior silk gland. Global transcription patterns revealed the physiological responses were induced by dinotefuran. Dinotefuran exposure substantially induced the expression levels of many genes involved in the mTOR and PI3K - Akt signaling pathways in the middle silk gland, whereas many differentially expressed genes involved in fatty acid and pyrimidine metabolism were found in the posterior silk gland. Additionally, functional, ultrastructural, and transcriptomic analysis indicate that dinotefuran exposure induced an increase of autophagy in the silk gland. This study illuminates the toxicity effects of dinotefuran exposure on silkworms and provides new insights into the underlying molecular toxicity mechanisms of dinotefuran to nontarget organisms.

Cobra venom P-III class metalloproteinase atrase A induces inflammatory response and cell apoptosis in endothelial cells via its metalloproteinase domain.

Snake venom metalloproteinases (SVMPs), which are a critical component of viperid and crotalid venoms, play various important roles in the pathogenesis of snakebite envenomation. The SVMPs from elapid venoms are not well elucidated, as compared with those from viperid and crotalid venoms. Atrase A is a nonhemorrhagic P-III SVMP purified from Naja atra venom that possesses only weak fibrinogenolytic activity. In our prior study, we found that atrase A detached adherent cells from the substrate. In this work, we investigated further the effect and mechanism of atrase A on endothelial cells. Oxidative damage, inflammatory mediators, apoptosis, and activation of the NF-κB and MAPK signaling pathways were measured after HMEC-1 cells were exposed to atrase A. The results showed that HMEC-1 cells released inflammatory mediators, exihibited oxidative damage and apoptosis after exposure to atrase A. The Western blot analysis results revealed that atrase A increased Bax/Bcl-2 and caspase-3 levels and activated the NF-κB and MAPK signaling pathways in endothelial cells. The effects on endothelial cells were nearly completely abolished after atrase A was treated with ethylenediamine tetraacetic acid. These results showed that atrase A led to an inflammatory response, cellular injury and apoptosis in endothelial cells, and this effect was due to its metalloproteinase domain. The study contributes to a better understanding of the structures and functions of cobra venom P-III class metalloproteinases.

Dinotefuran exposure induces autophagy and apoptosis through oxidative stress in Bombyx mori.

As a third-generation neonicotinoid insecticide, dinotefuran is extensively used in agriculture, and its residue in the environment has potential effects on nontarget organisms. However, the toxic effects of dinotefuran exposure on nontarget organism remain largely unknown. This study explored the toxic effects of sublethal dose of dinotefuran on Bombyx mori. Dinotefuran upregulated reactive oxygen species (ROS) and malondialdehyde (MDA) levels in the midgut and fat body of B. mori. Transcriptional analysis revealed that the expression levels of many autophagy and apoptosis-associated genes were significantly altered after dinotefuran exposure, consistent with ultrastructural changes. Moreover, the expression levels of autophagy-related proteins (ATG8-PE and ATG6) and apoptosis-related proteins (BmDredd and BmICE) were increased, whereas the expression level of an autophagic key protein (sequestosome 1) was decreased in the dinotefuran-exposed group. These results indicate that dinotefuran exposure leads to oxidative stress, autophagy, and apoptosis in B. mori. In addition, its effect on the fat body was apparently greater than that on the midgut. In contrast, pretreatment with an autophagy inhibitor effectively downregulated the expression levels of ATG6 and BmDredd, but induced the expression of sequestosome 1, suggesting that dinotefuran-induced autophagy may promote apoptosis. This study reveals that ROS generation regulates the impact of dinotefuran on the crosstalk between autophagy and apoptosis, laying the foundation for studying cell death processes such as autophagy and apoptosis induced by pesticides. Furthermore, this study provides a comprehensive insight into the toxicity of dinotefuran on silkworm and contributes to the ecological risk assessment of dinotefuran in nontarget organisms.

Combined analysis of transcriptomics and metabolomics on the cumulative effect of nano-titanium dioxide on mulberry seedlings.

Introduction: Titanium dioxide nanoparticles (TiO2 NPs) are among the most widely used inorganic nanomaterials in industry, medicine and food additives. There are increasing concerns regarding their potential risks to plants and the environment. Mulberry trees are widely grown in China due to their high survival rate and ability to aid ecological recovery. Methods: Herein, the effects of TiO2 NPs with different concentrations (100, 200, 400 and 800 mg/L) on the growth and physiology of the mulberry tree were systematically evaluated in aspects of physiology, transcriptomics and metabolomics. Results: Results showed that TiO2 NPs could be absorbed by the mulberry sapling root system and be transferred to the plant shoot. This results in the destruction of mulberry sapling root and leaf tissue. Furthermore, the number of chloroplasts and their pigment contents were reduced and the homeostasis of metal ions was disrupted. The toxic effects of TiO2 NPs attenuated the mulberry sapling's stress resistance, the contents of malondialdehyde in 100 mg/L, 200 mg/L 400 mg/L and 800 mg/L treatment groups increased by 87.70%, 91.36%, 96.57% and 192.19% respectively compared with the control group. The transcriptomic data showed that TiO2 NPs treatment mainly affected the expression of genes related to energy synthesis and transport, protein metabolism, and response to stress. Meanwhile, the results of metabolomics showed that 42 metabolites produced significant differences in mulberry, of which 26 differential metabolites were up-regulated in expression and 16 differential metabolites were down-regulated, mainly including metabolic pathways such as secondary metabolite biosynthesis, citric acid cycle, and tricarboxylic acid cycle, and was not conducive to the seed germination and or growth of the mulberry sapling. Discussion: This study enriches the understanding of the effects of TiO2 NPs on plants and provides a reference for the comprehensive scientific assessment of the potential risks of nanomaterials on plants.

The influence of extrovert-introvert personality on children's cortical activation with attention training systems.

Extrovert-introvert personality can take an active role in affecting people's attitudes, tastes, and behaviors in education. However, little research has been conducted to study whether and how extrovert-introvert personality may influence children's interaction with the attention training system. In this manuscript, we present the results of a user study that not only measured the influence of children's extrovert-introvert personality on their preference for two typical types of attention training systems (i.e., cognitive-based and neurofeedback-based) but also employed functional near-infrared spectroscopy (fNIRS) to investigate how the personality may influence cortical activation in children. Our results show that, for extroverted children, the neurofeedback attention training system elicited significantly greater activation in the prefrontal cortex and posterior parietal cortex, and was more likely to be preferred. The findings could be useful for developing more effective attention training systems based on user personality.

Toxicological effects of trace amounts of pyriproxyfen on the midgut of non-target insect silkworm.

Pyriproxyfen is an insect growth regulator that is widely used in public health and pest control in agriculture. Our previous studies have shown that trace amounts of pyriproxyfen in the environment can cause serious toxic effects in the non-target insect silkworm, including failing to pupate, metamorphose and spin cocoons. However, it is unknown why pyriproxyfen not only has no lethal effects on fifth instar larvae but also tend to increase their body weight. The midgut is the main digestive organs of the silkworm, our results showed that the residual of pyriproxyfen in the silkworm at 24 h after 1 × 10-4 mg/L pyriproxyfen treatment caused severe damage to the midgut microvilli, goblet cells, and nuclei of the silkworm, but body weight and digestibility of the larval were both increased. In addition, pyriproxyfen significantly (p < 0.05) increased the activities of digestive enzymes (α-amylase, trehalase, trypsin and lipase) in the midgut of silkworm. However, it caused down-regulation of ecdysone synthesis-related genes at the end of the fifth instar silkworm, decreased ecdysone titer, and prolonged larval instar. At the same time, pyriproxyfen also activated transcription of detoxification enzymes-related genes such as the cytochrome P450 enzyme genes Cyp9a22 and Cyp15C1, the carboxylesterase genes CarE-8 and CarE-11, and the glutathione S-transferase gene GSTo2. This study elucidated a novel toxicological effect of pyriproxyfen to insects, which not only expands the understanding of the effects of juvenile hormone pesticides on lepidopteran insects but also provides a reference for exploring the ecological security of non-target organisms.

Salvianolic acid A alleviates lipopolysaccharide-induced disseminated intravascular coagulation by inhibiting complement activation.

INTRODUCTION: Disseminated intravascular coagulation (DIC) is a syndrome characterized by coagulopathy, microthrombus, and multiple organ failure. The complement system in DIC is overactivated, and the functions of complement and coagulation pathways are closely related. Our previous screening revealed that salvianolic acid A (SAA) has anti-complement activity. The hyper-activated complement system was involved in the lipopolysaccharide (LPS) induced DIC in rats. The effects of SAA anti-complement action on LPS-induced DIC in rats were investigated. METHODS: The complement activity of the classical pathway and alternative pathway was detected through an in vitro hemolysis assay. The binding sites of SAA and complement C3b were predicted by molecular docking. LPS-induced disseminated coagulation experiments were performed on male Wistar rats to assess coagulation function, complement activity, inflammation, biochemistry, blood routine, fibrinolysis, and survival. RESULTS: SAA had an anti-complement activity in vivo and in vitro and inhibited the complement activation in the classical and alternative pathway of complement. The infusion of LPS into the rats impaired the coagulation function, increased the plasma inflammatory cytokine level, complemented activation, reduced the clotting factor levels, fibrinogen, and platelets, damaged renal, liver, and lung functions, and led to a high mortality rate (85%). SAA treatment of rats inhibited complement activation and attenuated the significant increase in D-dimer, interleukin-6, alanine aminotransferase, and creatinine. It ameliorated the decrease in plasma levels of fibrinogen and platelets and reversed the decline in activity of protein C and antithrombin III. The treatment reduced kidney, liver, and lung damage, and significantly improved the survival rate of rats (46.2 and 78.6% for the low- and high-dose groups, respectively). CONCLUSION: SAA reduced LPS-induced DIC by inhibiting complement activation. It has considerable potential in DIC treatment.

Pipiserpin, a Culex factor Xa inhibitor, affects female reproductive capacity and serves as a potential target for mosquito control.

BACKGROUND: Mosquito control is still the main prevention and control measure for numerous mosquito-borne diseases causing millions of deaths each year. New strategies for mosquito control are in demand. Proteases play an important role in mosquito physiology, therefore this study explored the inhibition of a serpin (serine protease inhibitor) in mosquitoes and its effect on reproductive capacity. RESULTS: A factor Xa inhibitor homolog (named Pipiserpin) was amplified and identified in Culex pipiens pallens mosquitoes. We expressed a recombinant Pipiserpin protein in vitro against which a mouse antiserum was generated. We found that female mosquitoes expressed more Pipiserpin protein than male mosquitoes. After mating, female mosquitoes were fed with blood mixed with different amounts of antisera and results showed that consumption of Pipiserpin impeded ovary development and decreased eggs hatching rates compared to that of the pre-immune serum group. CONCLUSION: We identified a Culex mosquito factor Xa inhibitor, Pipiserpin, which affects female reproductive potential. Our results suggest that Pipiserpin may be a novel target for mosquito population control. The conclusions from our study on Cx. pipiens pallens might serve as a reference for the development of control measures for other mosquitoes as well. © 2022 Society of Chemical Industry.

The JH-Met2-Kr-h1 pathway is involved in pyriproxyfen-induced defects of metamorphosis and silk protein synthesis in silkworms, Bombyx mori.

Environmental residues of pryriproxyfen, a juvenile hormone analogue (JHA) type pesticide, may have on unintended consequences on non-target insects. However, the mechanism of pyriproxyfen action and silk protein synthesis in silkworms has not been reported. In the present study, we treated the silkworms with trace pyriproxyfen (1 × 10-4 mg/L) and found that the silkworm larvae showed no obvious poisoning symptoms, while the development of silk glands and cocoon-forming function were both seriously damaged due to the accumulation of pyriproxyfen in posterior silk gland (PSG). The titer of the juvenile hormone (JH) was increased, whereas the content of 20-hydroxyecdysone (20E) was reduced in pyriproxyfen-exposed hemolymph. Met2 is a component of the JH receptor complex and JH can promote its phosphorylation. We found Met2 and SRC were up-regulated in the larval stage after pyriproxyfen exposure, the JH-Met2/SRC complex led to the up-regulation of downstream genes Kr-h1, and Dimm, and then specifically inhibited the transcription of Fib-H. Meanwhile, the transcription of ecdysone inducible transcription factor Br-C Z4 was also inhibited by pyriproxyfen and resulted in the defects of metamorphosis. In conclusion, the trace pyriproxyfen could affect the metamorphosis and silk protein synthesis through the Met2-mediated pathway. Our study provided new evidence that Met2 might be a potential target gene of JHA in Lepidoptera.

Purification, structural analysis, and stability of antioxidant peptides from purple wheat bran.

Protein derived from purple wheat bran was hydrolyzed sequentially using alcalase proteases for the production of antioxidant peptides. Purple wheat bran protein (PWBP) hydrolysates were fractionated using size-exclusion (G-25) and ion-exchange chromatography methods to identify the structure of antioxidant peptides. The free radical scavenging activity of peptides purified from PWBP hydrolysates was evaluated using superoxide anion radical-scavenging activity and determination assays of Trolox equivalent antioxidant capacity (TEAC). Results demonstrated that purple wheat bran peptide F4-4 exhibited the highest antioxidant activity among other hydrolysates. F4-4 was further identified as Cys-Gly-Phe-Pro-Gly-His-Cys, Gln-Ala-Cys, Arg-Asn-Phe, Ser-Ser-Cys, and Trp-Phe by high performance liquid chromatography (HPLC) spectrometer coupled with Orbitrap Elite™ mass spectrometer (LC-MS/MS). Antioxidant peptides 2 and 4 showed improved stability when the temperature was lower than 80 °C. These peptides also demonstrated good digestive stability in vitro system by simulating gastrointestinal digestion.

TP53, PIK3CA, FBXW7 and KRAS Mutations in Esophageal Cancer Identified by Targeted Sequencing.

BACKGROUND/AIM: Esophageal cancer (EC) is a common malignancy with significant morbidity and mortality. As individual cancers exhibit unique mutation patterns, identifying and characterizing gene mutations in EC that may serve as biomarkers might help predict patient outcome and guide treatment. Traditionally, personalized cancer DNA sequencing was impractical and expensive. Recent technological advancements have made targeted DNA sequencing more cost- and time-effective with reliable results. This technology may be useful for clinicians to direct patient treatment. MATERIALS AND METHODS: The Ion PGM and AmpliSeq Cancer Panel was used to identify mutations at 737 hotspot loci of 45 cancer-related genes in 64 EC samples from Chinese patients. RESULTS: Frequent mutations were found in TP53 and less frequent mutations in PIK3CA, FBXW7 and KRAS. CONCLUSION: These results demonstrate that targeted sequencing can reliably identify mutations in individual tumors that make this technology a possibility for clinical use.

Structural insights into the coordination of iron by Thermus thermophilus HB8 ferric binding protein A.

The ferric binding protein belongs to the substrate-binding protein super-family and transports ferric ions across the periplasmic space in gram negative bacteria. This process involves the binding and release of ferric ions through conformational changes of the ferric binding protein, and the assistance of a synergistic anion. Here we report the crystal structure of Thermus thermophilus HB8's (TtFbpA) ferric binding protein A in four different forms, which represent the apo state (apo-TtFbpA), the carbonate-bound state (TtFbpACO3),and the iron- and carbonate-bound state (TtFbpAFeCO3). The ferric ion in TtFbpAFeCO3 is bound by three tyrosine residues from TtFbpA and one synergistic carbonate ion. Structural comparisons among the three different states reveal the molecular mechanisms of iron-binding by TtFbpA. Our results, together with previous studies on other bacterial periplasmic ferric binding proteins, provide a complete understanding of the structural basis for iron binding and release in the periplasm of gram-negative bacteria.