Search

Bovine serum albumin (BSA) was induced to form cold-set gel by using glucono-δ-lactone (GDL) and sodium chloride (NaCl), and characteristics of the gel and its embedding potential to bioactive molecules were investigated. The results from texture analyzer indicated that hardness and springiness of BSA cold-set gel were improved by increasing GDL or NaCl, but the gel with lower concentration of GDL (1.0%) and NaCl (0.2 M) had better water holding capacity (WHC). GDL had a significant influence on storage modulus (G’) and loss modulus (G″) but it did not appear to be applicable to NaCl. Light transmittance at 500 nm and average particle size were used to characterize formation of BSA cold-set gel as pH values decreased, and the results suggested that BSA molecules had aggregated at the pH value of the gel point. Microstructure and fractal analysis from confocal laser scanning microscope (CLSM) further suggested that electrostatic and hydrophobic interaction played important roles in the formation of the cold-set gel and it could be deduced that structure and properties of the gel were closely related to the forces. BSA cold-set gel showed an excellent embedding performance to encapsulate bioactive molecules (used quercetin as representative), and achieved effective slow-release action, especially in simulated intestinal juice of pH 7.5. BSA cold-set gel may have important potential application in encapsulation, protection, delivery and controlled-release of some fragile bioactive components, so the research provides a theoretical basis for its application.

Rationale: Colorectal cancer (CRC) is one of the most common cancers worldwide. Ciclopirox olamine (CPX) has recently been identified to be a promising anticancer candidate; however, novel activities and detailed mechanisms remain to be uncovered. Methods: The cytotoxic potential of CPX towards CRC cells was examined in vitro and in vivo. The global gene expression pattern, ROS levels, mitochondrial function, autophagy, apoptosis, etc. were determined between control and CPX-treated CRC cells. Results: We found that CPX inhibited CRC growth by inhibiting proliferation and inducing apoptosis both in vitro and in vivo. The anti-cancer effects of CPX involved the downregulation of DJ-1, and overexpression of DJ-1 could reverse the cytotoxic effect of CPX on CRC cells. The loss of DJ-1 resulted in mitochondrial dysfunction and ROS accumulation, thus leading to CRC growth inhibition. The cytoprotective autophagy was provoked simultaneously, and blocking autophagy pharmacologically or genetically could further enhance the anti-cancer efficacy of CPX. Conclusion: Our study demonstrates that DJ-1 loss-induced ROS accumulation plays a pivotal role in CPX-mediated CRC inhibition, providing a further understanding for CRC treatment via modulating compensatory protective autophagy.

GBM (glioblastoma multiforme) is the most common and aggressive brain tumor with no curative options available. Therefore, it is imperative to develop novel potent therapeutic drugs for GBM treatment. Here, we show that regorafenib, an oral multi-kinase inhibitor, exhibits superior therapeutic efficacy over temozolomide, the first-line chemotherapeutic agent for GBM treatment both in vitro and in vivo. Mechanistically, regorafenib directly stabilizes PSAT1 (phosphoserine aminotransferase 1), a critical enzyme for serine synthesis, to trigger PRKAA-dependent autophagy initiation and inhibit RAB11A-mediated autophagosome-lysosome fusion, resulting in lethal autophagy arrest in GBM cells. Maintenance of PSAT1 at a high level is essential for regorafenib-induced GBM suppression. Together, our data provide novel mechanistic insights of regorafenib-induced autophagy arrest and suggest a new paradigm for effective treatment of GBM. Abbreviations: 3-MA: 3-methyladenine; ACACA: acetyl coenzyme A carboxylase alpha; ACTB/β-actin: actin, beta; AMPK: adenosine monophosphate-activated protein kinase; ATG5: autophagy related 5; CTSD: cathepsin D; DN-: dominant-negative; GBM: glioblastoma multiforme; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PRKAA/AMPKα: protein kinase AMP-activated catalytic subunit alpha; PSAT1: phosphoserine aminotransferase 1; SQSTM1/p62: sequestosome 1; TKIs: tyrosine kinase inhibitors.

Checkpoint blockade therapy targeting the programmed-death ligand 1 (PD-L1) and its receptor programmed cell death 1 promotes T-cell-mediated immunosurveillance against tumours, and has been associated with marked clinical benefit in cancer patients. Antibodies against PD-L1 function by blocking PD-L1 on the cell surface, but intracellular storage of PD-L1 and its active redistribution to the cell membrane can minimize the therapeutic benefits, which highlights the importance of targeting PD-L1 throughout the whole cell. Here, we show that PD-L1 is palmitoylated in its cytoplasmic domain, and that this lipid modification stabilizes PD-L1 by blocking its ubiquitination, consequently suppressing PD-L1 degradation by lysosomes. We identified palmitoyltransferase ZDHHC3 (DHHC3) as the main acetyltransferase required for the palmitoylation of PD-L1, and show that the inhibition of PD-L1 palmitoylation via 2-bromopalmitate, or the silencing of DHHC3, activates antitumour immunity in vitro and in mice bearing MC38 tumour cells. We also designed a competitive inhibitor of PD-L1 palmitoylation that decreases PD-L1 expression in tumour cells to enhance T-cell immunity against the tumours. These findings suggest new strategies for overcoming PD-L1-mediated immune evasion in cancer.

Metabolic disorders of the circulatory system of animals (e.g., hyperglycemia and hyperlipidemia) can significantly affect immune function, however, since there is currently no reliable animal model for hyperproteinemia, its effects on immunity remain unclear. In this study, we established an animal model for hyperproteinemia in an invertebrate silkworm model, with a controllable plasma protein concentration (PPC) and no primary disease effects. We evaluated the influence of hyperproteinemia on innate immunity. The results showed that high PPC enhanced hemolymph phagocytosis via inducing a rapid increase in granulocytes. Moreover, while oenocytoids increased, the plasmacytes quickly dwindled. High PPC inhibited hemolymph melanization due to decreased phenoloxidase (PO) activity in the hemolymph via inhibiting the expression of the prophenoloxidase-encoding genes, PPO1 and PPO2. High PPC upregulated the gene expression of antimicrobial peptides via differential activation of the Toll and Imd signaling pathways associated with NF-κB signaling, followed by an induction of inconsistent antibacterial activity towards Gram-positive and Gram-negative bacteria in an animal model of high PPC. Therefore, high PPC has multiple significant effects on the innate immune function of the silkworm circulatory system.

Activation of PI3K/AKT pathway is one of the most recurrent resistant mechanisms for BRAF-targeted therapy, and the combination of MAPK and PI3K/AKT inhibitors becomes one of the most promising regimens for BRAF-targeted relapsed melanoma patients. Although the potent drug efficacy was observed in preclinical experiments and early clinical trials, the dual-drug resistance is inevitable observed. In this study, we systematically explored the mechanisms of dual-drug resistance to MAPKi and PI3K/mTORi in melanoma. With transcriptomic dissection of dual-drug resistant models, we identified that the drug tolerance was mediated by ECM-integrins α3β1 and α11β1 signaling. Upon binding ECM, the integrins activated downstream kinase Src rather than FAK, WNT, or TGFβ. Knockdown of integrins α3, α11, and β1 significantly inhibited the proliferation of dual-drug resistant sublines while with trivial effects on parental cells. Although Src inhibition suppressed the phosphorylation of AKT, c-JUN, and p38, none of inhibitors targeting these kinases reversed the dual-drug resistance in model cells. Notably, Src inhibitor promoted the phosphorylations of LATS1 and YAP1, subsequently, re-localized YAP1 from nucleus to cytosol facilitating further degradation. Both small molecule inhibitors and shRNAs targeting YAP1 or Src overcame the MAPKi and PI3K/mTORi dual-drug resistance. In conclusion, our data not only illuminated an integrin-Src-YAP1 pathway mediated MAPKi and PI3K/mTORi dual-drug resistant mechanism but also provided a potential combinatorial regimen for the drug-relapsed melanoma patients.

Programmed cell death protein 1 (PD-1) is a crucial anticancer target, but the relatively low response rate and acquired resistance to existing antibody drugs highlight an urgent need to develop alternative targeting strategies. Here, we report the palmitoylation of PD-1, discover the main DHHC enzyme for this modification, reveal the mechanism of its effect on PD-1 protein stability, and rationally develop a peptide for targeting PD-1 expression. Palmitoylation promoted the trafficking of PD-1 to the recycling endosome, thus preventing its lysosome-dependent degradation. Palmitoylation of PD-1, but not of PD-L1, promoted mTOR signaling and tumor cell proliferation, and targeting palmitoylation displayed significant anti-tumor effects in a three-dimensional culture system. A peptide was designed to competitively inhibit PD-1 palmitoylation and expression, opening a new route for developing PD-1 inhibitors and combinatorial cancer immunotherapy., We show for the first time that PD-1 is palmitoylated, identify DHHC9 as the predominant enzyme for its palmitoylation, and reveal the molecular mechanisms underlying its effects on PD-1 stability and functions. Importantly, we also designed PD1-PALM, a competitive inhibitor of PD-1 palmitoylation, and this first-in-class molecule may inspire the development of new checkpoint inhibitors.

Tumor necrosis factor-α (TNF-α) is a pluripotent mediator of pro-inflammatory and antimicrobial defense mechanisms and a regulator of lymphoid organ development. Although two types of TNF-α have been identified in several teleost species, their functions in pathogen infection remain largely unexplored, especially in pathogen clearance. Herein, we cloned and characterized two types of TNF-α, termed shTNF-α1 and shTNF-α2, and their receptors, shTNFR1 and shTNFR2, from snakehead (Channa argus). These genes were constitutively expressed in all tested tissues, and were induced by Aeromonas schubertii and Nocardia seriolae in head kidney and spleen in vivo, and by lipoteichoic acid (LTA), lipopolysaccharides (LPS), and Polyinosinic-polycytidylic acid [Poly (I:C)] in head kidney leukocytes (HKLs) in vitro. Moreover, recombinant shTNF-α1 and shTNF-α2 upregulated the expression of endogenous shTNF-α1, shTNF-α2, shTNFR1, and shTNFR2, and enhanced intracellular bactericidal activity, with shTNF-α1 having a greater effect than shTNF-α2. These findings suggest important roles of fish TNFα1, TNFα2, and their receptors in bacterial infection and pathogen clearance, and provide a new insight into their function in antibacterial innate immunity.

As a central pro-inflammatory cytokine, interleukin-1β (IL-1β) plays critical roles in the inflammatory response, pathogen infection, and immunological challenges in mammals. Although fish IL-1β has been confirmed to participate in inflammatory response to pathogen infection, few studies have been performed to characterize the antibacterial and bactericidal functions of fish IL-1β. In this study, snakehead (Channa argus) IL-1β (shIL-1β) and its receptors, shIL-1R1 and shIL-1R2, were cloned and functionally characterized. ShIL-1β contained the IL-1 family signature domain, and a potential cutting site at Asp96 that presented in all vertebrate IL-1β sequences. ShIL-1R1 had three extracellular IG-like domains and one intracellular signal TIR domain, while shIL-1R2 had three extracellular IG-like domain but lacked the intracellular signal TIR domain. ShIL-1β, shIL-1R1, and shIL-1R2 were constitutively expressed in all tested tissues, and their expressions could be induced by Aeromonas schubertii and Nocardia seriolae in the head kidney and spleen in vivo, and by LTA, LPS, and Poly (I:C) in head kidney leukocytes (HKLs) in vitro. Moreover, recombinant shIL-1β upregulated the expression of endogenous shIL-1β, shIL-R1, and shIL-R2 in snakehead HKLs, and enhanced intracellular bactericidal activity. Taken together, this study found that, like IL-1β and its receptors in mammals, shIL-1β and its receptors play crucial roles in antibacterial innate immunity. This provides new insight into the evolution of IL-1β function in vertebrates.

Immune checkpoint blockades (ICBs), as a major breakthrough in cancer immunotherapy, target CTLA-4 and the PD-1/PD-L1 axis and reinvigorate anti-tumor activities by disrupting co-inhibitory T-cell signaling. With unprecedented performance in clinical trials, ICBs have been approved by FDA for the treatment of malignancies such as melanoma, non-small-cell lung cancer, colorectal cancer, and hepatocellular carcinoma. However, while ICBs are revolutionizing therapeutic algorithms for cancers, the frequently observed innate, adaptive or acquired drug resistance remains an inevitable obstacle to a durable antitumor activity, thus leading to non-response or tumor relapse. Researches have shown that resistance could occur at each stage of the tumor’s immune responses. From the current understanding, the molecular mechanisms for the resistance of ICB can be categorized into the following aspects: 1. Tumor-derived mechanism, 2. T cell-based mechanism, and 3. Tumor microenvironment-determined resistance. In order to overcome resistance, potential therapeutic strategies include enhancing antigen procession and presentation, reinforcing the activity and infiltration of T cells, and destroying immunosuppression microenvironment. In future, determining the driving factors behind ICB resistance by tools of precision medicine may maximize clinical benefits from ICBs. Moreover, efforts in individualized dosing, intermittent administration and/or combinatory regimens have opened new directions for overcoming ICB resistance.

Encapsulation and protection of hydrophobic bioactive compounds using food-based sustainable materials are challenging issues in recent decade. Herein, we provided a feasible strategy to fabricate bovine serum albumin (BSA) cold-set emulsion gel by using microbial transglutaminase (MTGase) and glucono-δ-lactone (GDL) as substrate precursors. Its embedding potential towards quercetin and vitamin E (VE) were thereby evaluated. The results showed that hardness and springiness of the gels decreased along with increasing of GDL (≥0.3%, w/v) in the presence of 0.1% (w/v) MTGase. Storage modulus (G′) and loss modulus (G'') reached the maximum at 0.3% (w/v) GDL, while the water-holding capacity (WHC) of gels (≥85% (w/v)) was positively correlated to GDL addition. The pattern of confocal laser scanning microscope (CLSM) further confirmed the occurrence of smaller oil droplets in fabricated gels with relatively high abundance of GDL. Consequently, the embedding efficiencies for quercetin and VE reached up to 94.80% (w/v) and 97.94% (w/v) respectively. The harder gels had much better controlled-release performance than the softer ones. The work shed light on the rational design of food protein gels to protect, stabilize and delivery of small hydrophobic bioactive molecules.

The effects of meal size on the postprandial metabolic response and of digestion on the post-exercise metabolic recovery process were investigated in juvenile black carp (Mylopharyngodon piceus) . ...

Bleomycin is a clinically potent anticancer drug used for the treatment of germ-cell tumors, lymphomas and squamous-cell carcinomas. Unfortunately, the therapeutic efficacy of bleomycin is severely hampered by the development of pulmonary fibrosis. However, the mechanisms underlying bleomycin-induced pulmonary fibrosis, particularly the molecular target of bleomycin, remains unknown. Here, using a chemical proteomics approach, we identify ANXA2 (annexin A2) as a direct binding target of bleomycin. The interaction of bleomycin with ANXA2 was corroborated both in vitro and in vivo. Genetic depletion of anxa2 in mice mitigates bleomycin-induced pulmonary fibrosis. We further demonstrate that Glu139 (E139) of ANXA2 is required for bleomycin binding in lung epithelial cells. A CRISPR-Cas9-engineered ANXA2E139A mutation in lung epithelial cells ablates bleomycin binding and activates TFEB (transcription factor EB), a master regulator of macroautophagy/autophagy, resulting in substantial acceleration of autophagic flux. Pharmacological activation of TFEB elevates bleomycin-initiated autophagic flux, inhibits apoptosis and proliferation of epithelial cells, and ameliorates pulmonary fibrosis in bleomycin-treated mice. Notably, we observe lowered TFEB and LC3B levels in human pulmonary fibrosis tissues compared to normal controls, suggesting a critical role of TFEB-mediated autophagy in pulmonary fibrosis. Collectively, our data demonstrate that ANXA2 is a specific bleomycin target, and bleomycin binding with ANXA2 impedes TFEB-induced autophagic flux, leading to induction of pulmonary fibrosis. Our findings provide insight into the mechanisms of bleomycin-induced fibrosis and may facilitate development of optimized bleomycin therapeutics devoid of lung toxicity.

Gut microbiota is closely related to the tumorigenesis and progression of colorectal cancer (CRC), especially Fusobacterium nucleatum (Fn) could promote colorectal tumor growth and cause chemoresistance. Here, we developed a tumor-triggered release drug delivery gel to inhibit CRC cell growth in situ by intestinal perfusion. Antibiotic metronidazole (MTZ) and 5-fluorouracil (5-FU), the first-line chemotherapy drug of CRC, are individually incorporated into metal polyphenol network-coated mesoporous silica nanoparticle (MSN), and then blended with carboxymethyl cellulose (CMC) to obtain an anti-CRC gel, AB-Gel. In vivo studies showed that AB-Gel could efficiently inhibit the growth and metastasis of CRC in an orthotopic mouse model attributing to the MTZ-induced modulation in the CRC-related microbiota. Chemotherapy efficacy against orthotopic CRC was significantly enhanced by the elimination of Fn. This strategy based on the combination of gut microbiota modulation and chemotherapy would be considered as an enhanced strategy for CRC treatment with potential use in the clinic.

Memory enables access to past experiences to guide future behavior. Humans can determine which memories to trust (high confidence) and which to doubt (low confidence). How memory retrieval, memory confidence, and memory-guided decisions are related, however, is not understood. In particular, how confidence in memories is used in decision making is unknown. We developed a spatial memory task in which rats were incentivized to gamble their time: betting more following a correct choice yielded greater reward. Rat behavior reflected memory confidence, with higher temporal bets following correct choices. We applied machine learning to identify a memory decision variable and built a generative model of memories evolving over time that accurately predicted both choices and confidence reports. Our results reveal in rats an ability thought to exist exclusively in primates and introduce a unified model of memory dynamics, retrieval, choice, and confidence.

Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3) is a serine/threonine protein kinase that acts as a key regulator and is widely involved in various innate and acquired immune signaling pathways. In this study, we first cloned the complete open reading frame (ORF) of the MEKK3 gene (named CcMEKK3) in a hybrid snakehead (Channa maculate ♀ × Channa argus ♂). The full-length ORF of CcMEKK3 is 1851 bp, and encodes a putative protein of 616 amino acids containing a serine/threonine kinase catalytic (S-TKc) domain and a Phox and Bem1p (PB1) domain. A sequence alignment and phylogenetic tree analysis showed that CcMEKK3 is highly conserved relative to the MEKK3 proteins of other teleost species. CcMEKK3 was constitutively expressed in all the healthy hybrid snakehead tissues tested, with greatest expression in the immune tissues, such as the head kidney and spleen. The expression of CcMEKK3 was usually upregulated in the head kidney, spleen, and liver at different time points after infection with Nocardia seriolae or Aeromonas schubertii. Similarly, the dynamic expression levels of CcMEKK3 in head kidney leukocytes after stimulation revealed that CcMEKK3 was induced by LTA, LPS, and poly(I:C). In the subcellular localization analysis, CcMEKK3 was evenly distributed in the cytoplasm of HEK293T cells, and its overexpression significantly promoted the activities of NF-κB and AP-1. These results suggest that CcMEKK3 is involved in the immune defense against these two pathogens, and plays a crucial role in activating the NF-κB and MAPK signaling pathways.

Diapause is a developmental transition in insects based on seasonal adaptation to adversity, it is regulated by a circadian clock system and the endocrine system. However, the molecular node and its mechanism underlying the effects of these systems are still unclear. Here, a mutant of Bombyx mori with the circadian clock gene Period (Per) knocked out was constructed, which dramatically changed the classic diapause-destined pathway. Per-knockout silkworms powerfully attenuated, but could not completely block, the predetermined effects of temperature and photoperiod on diapause determination, and this effect depended on the diapause hormone (DH) pathway. The impaired transcription-translation feedback loop of the circadian clock system lacking the Per gene caused direct up-regulation of the expression of GRD, a receptor of γ-aminobutyric acid (GABA), by changing expression level of Cycle. The synthesis of GABA in the tissue complex of brain-suboesophageal ganglion then increased and restricted the decomposition, which continuously promoted the GABAergic signal to play a role, and finally inhibiting (delaying) the release of DH to the hemolymph, and reducing the diapause-inducing effect of DH. The results provided an example to explain the regulatory mechanism of the circadian clock on endocrine hormones in the silkworm.

Extracellular adenosine 5'-triphosphate (ATP), secreted by living cancer cells or released by necrotic tumor cells, plays an important role in tumor invasion and metastasis. Our previous study demonstrated that ATP treatment in vitro could promote invasion in human prostate cancer cells via P2Y2, a preferred receptor for ATP, by enhancing EMT process. However, the pro-invasion mechanisms of ATP and P2Y2 are still poorly studied in breast cancer. In this study, we found that P2Y2 was highly expressed in breast cancer cells and associated with human breast cancer metastasis. ATP could promote the in vitro invasion of breast cancer cells and enhance the expression of β-catenin as well as its downstream target genes CD44, c-Myc and cyclin D1, while P2Y2 knockdown attenuated above ATP-driven events in vitro and in vivo. Furthermore, iCRT14, a β-catenin/TCF complex inhibitor, could also suppress ATP-driven migration and invasion in vitro. These results suggest that ATP promoted breast cancer cell invasion via P2Y2-β-catenin axis. Thus blockade of the ATP-P2Y2-β-catenin axis could suppress the invasive and metastatic potential of breast cancer cells and may serve as potential targets for therapeutic interventions of breast cancer.

Pure bacterial cultures were isolated from different tissues of moribund Megalobrama terminalis from a high mortality event that occurred at a farm in Foshan, China. Two isolates (F2 and F3) were identified as Streptococcus dysgalactiae subsp. dysgalactiae based on morphological and biochemical detection as well as molecular analysis. In brain heart infusion broth, the best growth conditions of isolate F3 were 35oC, salinity 5‰ and pH 7. Furthermore, infection with isolate F3 (1.2 × 106 CFU/fish) led to the death of M. terminalis and zebrafish (Danio rerio). However, isolate F3 had no obvious pathogenicity to tilapia (GIFT, Oreochromis niloticus). When the water temperature was 29oC, the corresponding mortality rates for zebrafish infected by isolate F3 were higher than those at 23oC. Culture for 24 and 72 hr with isolate F3 resulted in the same mortality rates for zebrafish. The antimicrobial susceptibility assay revealed that isolate F3 was susceptible to ampicillin, florfenicol and several other antibiotics but resistant to nalidixic acid, streptomycin, sulfamethoxazole/trimethoprim, neomycin and amikacin. To our knowledge, this is the first report that S. dysgalactiae infected the subtropical freshwater fish M. terminalis, which indicates that this bacterium is a potential threat to subtropical freshwater fish.

Breast cancer is the most common cancer among women worldwide, yet successful treatment remains a clinical challenge. Ivermectin, a broad-spectrum antiparasitic drug, has recently been characterized as a potential anticancer agent due to observed antitumor effects. However, the molecular mechanisms involved remain poorly understood. Here, we report a role for ivermectin in breast cancer suppression by activating cytostatic autophagy both in vitro and in vivo. Mechanistically, ivermectin-induced autophagy in breast cancer cells is associated with decreased P21-activated kinase 1 (PAK1) expression via the ubiquitination-mediated degradation pathway. The inhibition of PAK1 decreases the phosphorylation level of Akt, resulting in the blockade of the Akt/mTOR signaling pathway. In breast cancer xenografts, the ivermectin-induced cytostatic autophagy leads to suppression of tumor growth. Together, our results provide a molecular basis for the use of ivermectin to inhibit the proliferation of breast cancer cells and indicate that ivermectin is a potential option for the treatment of breast cancer. Cancer Res; 76(15); 4457–69. ©2016 AACR.

Adenosine monophosphate-activated protein kinase (AMPK) is a crucial energy sensor that maintains cellular energy homeostasis. AMPK plays a critical role in macroautophagy/autophagy, and autophagy facilitates hepatitis B virus (HBV) replication. To date, the intrinsic link among AMPK, autophagy and HBV production remains to be elucidated. Here, we demonstrate that PRKAA (a catalytic subunit of AMPK) is activated in response to HBV-induced oxidative stress, which in turn decreases the production of HBV. Mechanistic studies reveal that the autophagy machinery is associated with the inhibitory effect of PRKAA/AMPK on HBV production. Activation of PRKAA/AMPK promotes autolysosome-dependent degradation through stimulation of cellular ATP levels, which then leads to the depletion of autophagic vacuoles. Taken together, our data suggest that the activation of AMPK might be a stress response of host cells to restrict virus production through promotion of autophagic degradation. These findings therefore indicate that AMPK could provide a potential therapeutic target for HBV infection.

Expression of programmed cell death 1 (PD-1) ligand 1 (PD-L1) protects tumor cells from T cell-mediated immune surveillance, and immune checkpoint blockade (ICB) therapies targeting PD-1 and PD-L1 have exhibited significant clinical benefits. However, the relatively low response rate and observed ICB resistance highlight the need to understand the molecular regulation of PD-L1. Here we show that HIP1R targets PD-L1 to lysosomal degradation to alter T cell-mediated cytotoxicity. HIP1R physically interacts with PD-L1 and delivers PD-L1 to the lysosome through a lysosomal targeting signal. Depletion of HIP1R in tumor cells caused PD-L1 accumulation and suppressed T cell-mediated cytotoxicity. A rationally designed peptide (PD-LYSO) incorporating the lysosome-sorting signal and the PD-L1-binding sequence of HIP1R successfully depleted PD-L1 expression in tumor cells. Our results identify the molecular machineries governing the lysosomal degradation of PD-L1 and exemplify the development of a chimeric peptide for targeted degradation of PD-L1 as a crucial anticancer target.

The objectives of this study were to examine whether sustained exercise training at four water velocities, i.e. nearly still water (control), 1 body length (BL) s−1, 2 BL s−1 and 4 BL s−1, has effects on swimming performance and digestive metabolism in juvenile black carp (Mylopharyngodon piceus). The results demonstrated that fish subjected to sustained training at 2 and 4 BL s−1 showed significantly higher critical swimming speed (Ucrit) and maximum metabolic rate (MMR) over the control group. Fish subjected to sustained training at 1 and 2 BL s−1 showed a significantly (30 and 54%) prolonged duration, 14 and 17% higher postprandial ṀO2 increment (i.e. ṀO2peak), and 62 and 92% more energy expended on specific dynamic action (SDA), respectively, after consuming a similar meal over fish kept in nearly still water. These results suggest that (1) sustained exercise training at a higher speed (2 or 4 BL s−1) had a positive influence on the aerobic swimming performance of juvenile M. piceus, which may be associated with improved aerobic metabolism; and (2) sustained exercise training at a lower speed (1 or 2 BL s−1) resulted in elevated postprandial metabolic responses in juvenile M. piceus.

Objective: This study aims to understand the distribution of pathogenic bacteria in the region of HIV/AIDS patients with opportunistic infection. Methods: To count the number of the bacterial culture of HIV/AIDS patients in our hospital from October 2011 to December 2014, and observe the distribution of all kinds of pathogenic bacteria. Results: From the 4269 cases of HIV/AIDS patients’ bacteria, 5045 cases were cultured whose main flora distribution was Candida albicans, 1759 cases. The second one was penicillium, 982 cases. The third one was mycobacteria, 557 cases. And then there are 213 cases of Cryptococcus neoformans, 212 cases of Klebsiella pneumonia, 209 cases of E. coli, 157 cases of coagulase-negative staphylococci, 112 cases of Candida tropicalis, 90 cases of glabrata, 81 cases of Staphylococcus aureus, 75 cases of Pseudomonas aeruginosa, 60 cases of Salmonella, 48 cases of Acinetobacter and the distribution of the rest of cultured bacterial was less than 40 cases. Conclusion: There are many kinds of types of Pathogenic bacteria in HIV/AIDS patients with the opportunity to infectious. And the majorities are Candida albicans, Penicillium marneffei, Penicillium, Mycobacterium, Cryptococcus neoformans and so on. The infection sites are widely distributed; respiratory and circulatory are the main infected system. Improving the detection rate and reducing the contamination rate can truly reflect the distribution of pathogenic bacteria, and the distribution can guide the infection work in hospital. At the same time, it’s good to predict and prevent opportunistic infection. Thus, the patients can get immediate treatment.

Dysregulation of microRNAs (miRNAs) has long been implicated in tumorigenesis, whereas the underlying mechanisms remain largely unknown. Oxidative stress is a hallmark of cancer that involved in multiple pathophysiological processes, including the aberrant regulation of miRNAs. Compelling evidences have implied complicated interplay between reactive oxygen species (ROS) and miRNAs. Indeed, ROS induces carcinogenesis through either reducing or increasing the miRNA level, leading to the activation of oncogenes or silence of tumor suppressors, respectively. In turn, miRNAs target ROS productive genes or antioxidant responsive elements to affect cellular redox balance, which contributes to establishing a microenvironment favoring cancer cell growth and metastasis. Both miRNAs and ROS have been identified as potential biomarkers and therapeutic targets in human malignancies, and comprehensive understanding of the molecular events herein will facilitate the development of novel cancer therapeutic strategies.

The purpose of this study was to investigate the chemical composition and biological activity of the volatile oils (VOs) from the flowers of three buckwheat species, Fagopyrum esculentum, Fagopyrum tataricum and Fagopyrum cymosum. The VOs were obtained from the fresh buckwheat flowers by hydrodistillation, and were analyzed for their chemical composition by gas chromatography-mass spectrometry (GC-MS). Nonanoic acid (7.58%), (E)-3-hexen-1-ol (6.52%), and benzothiazole (5.08%) were the major constituents among the 28 identified components which accounted for 92.89% of the total oil of F. esculentum. 2-Pentadecanone (18.61%), eugenol (17.18%), 1,2-benzenedicarboxylic acid, bis(2-methylpropyl) ester (13.19%), and (E,E)-farnesylacetone (7.15%) were the major compounds among the 14 identified components which accounted for 88.48% of the total oil of F. tataricum. Eugenol (12.22%), (E)-3-hexen-1-yl acetate (8.03%), linalool oxide (7.47%), 1-hexanol (7.07%), and benzothiazole (6.72%) were the main compounds of the 20 identified components which accounted for 90.23% of the total oil of F. cymosum. The three VOs were screened to have broad spectrum antibacterial activity with minimum inhibitory concentration (MIC) values ranged from 100.0 μg/mL to 800.0 μg/mL against the tested bacteria, and their median inhibitory concentration (IC50) values were from 68.32 μg/mL to 452.32 μg/mL. Xanthomonas vesicatoria was the most sensitive bacterium. Moreover, the flower VOs of F. esculentum, F. tataricum and F. cymosum also exhibited noteworthy antioxidant capacity with the IC50 value of 354.15 μg/mL, 210.63 μg/mL, and 264.92 μg/mL for the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assay, and the value of 242.06 μg/mL, 184.13 μg/mL, and 206.11 μg/mL respectively for the β-carotene-linoleic bleaching test. These results suggested the volatile oils of buckwheat flowers could be potential resource of natural antimicrobial and antioxidant agents.

Perfluorooctane sulfonate (PFOS) is a ubiquitous environmental contaminant that has been found to pose various risks to fish health and the safety of aquatic ecosystem. Swimming performance is an integrated index of fitness in fish. However, little research has sought on the effects of PFOS on swimming performances of fish. Experiments were carried out to clarify the impacts of acute exposure to PFOS on behavior, swimming ability and metabolic rate in topmouth gudgeon (Pseudorasbora parva), to understand the underlying ecotoxicological effects of waterborne PFOS exposure on fish physiology and behavior. Fish were exposed to PFOS (0, 0.5, 2, 8 or 32 mg/L) for 96 h. Afterwards, the routine metabolic rate (RMR), spontaneous swimming behavior (SSB), fast-start swimming performance (FSP) and critical swimming speed (Ucrit) of the topmouth gudgeon were examined. The results show reduced behavioral performance and increased physiological stress with increasing PFOS concentration. Both RMR, SSB and Ucrit were significantly affected by PFOS exposure (p 0.05). The results indicate that the anaerobic swimming capacity was conserved, but the metabolic level, SSB and aerobic swimming performance in topmouth gudgeon were susceptible to PFOS contamination, and hence might be useful as considerable potential biomarkers of pollution.

In the version of this Article originally published, 'palmitoyltransferase ZDHHC3 (DHHC3)' was incorrectly referred to as an 'acetyltransferase' rather than an as an 'acyltransferase'; this has now been corrected in five instances. In Fig. 3a, the label for the bottom row of the blots was mistakenly written as 'GAPHD'; it should have read 'GAPDH'. In the two right-most panels of Fig. 4j, the antibody labels 'α-PD-L1' for the reciprocal co-immunoprecipitation of DHHC3 were incorrect; they should have been 'α-DHHC3'. These errors have been corrected in all versions of the Article.

The basal forebrain (BF) plays key roles in multiple brain functions, including sleep-wake regulation, attention, and learning/memory, but the long-range connections mediating these functions remain poorly characterized. Here we performed whole-brain mapping of both inputs and outputs of four BF cell types – cholinergic, glutamatergic, and parvalbumin-positive (PV+) and somatostatin-positive (SOM+) GABAergic neurons – in the mouse brain. Using rabies virus -mediated monosynaptic retrograde tracing to label the inputs and adeno-associated virus to trace axonal projections, we identified numerous brain areas connected to the BF. The inputs to different cell types were qualitatively similar, but the output projections showed marked differences. The connections to glutamatergic and SOM+ neurons were strongly reciprocal, while those to cholinergic and PV+ neurons were more unidirectional. These results reveal the long-range wiring diagram of the BF circuit with highly convergent inputs and divergent outputs and point to both functional commonality and specialization of different BF cell types. DOI: http://dx.doi.org/10.7554/eLife.13214.001

Aeromonas is regarded as an important pathogen of freshwater animals but little is known about the genetics of its antimicrobial resistance in Chinese aquaculture. The aim of this study was to investigate the presence of integrons and characterize multidrug resistant Aeromonas spp. isolated from diseased farmed freshwater animals. These animal samples included fish, ornamental fish, shrimp, turtles, and amphibians which were collected from 64 farms in Guangdong province of South China. One hundred and twelve Aeromonas spp. isolates were examined for antimicrobial resistance phenotypes and the presence of class 1 integron sequences. Twenty-two (19.6%) of these isolates carried a class 1 integron comprising six different gene insertion cassettes including drfA12-orfF-aadA2, drfA12-orfF, aac(6')-II-bla OXA-21 -cat3, catB3, arr-3, and dfrA17. Among these, drfA12-orfF-aadA2 was the dominant gene cassette array (63.6%, 14/22) and this is the first report of aac(6')-II-bla OXA-21 -cat3 in an Aeromonas hydrophila isolate from a Chinese giant salamander (Andrias davidianus). All the integron-positive strains were resistant to more than five agents and 22 contained other resistance genes including bla CTX-M-3, bla TEM-1, aac(6')-Ib-cr, and tetA. All integron-positive isolates also contained mutations in the quinolone resistance determining regions (QRDR). Our investigation demonstrates that freshwater animals can serve as a reservoir for pathogenic Aeromonas strains containing multiple drug-resistance integrons. This data suggests that surveillance for antimicrobial resistance of animal origin and a prudent and responsible use of antimicrobials in aquaculture is necessary in these farms.

To investigate the effects of hypoxia acclimation, exercise training and fasting on the swimming performance of juvenile qingbo (Spinibarbus sinensis), we measured the critical swimming speed (U crit), resting and excess post-exercise oxygen consumption (EPOC) of control, hypoxia-acclimated, exercise-acclimated and fasting fish at 25°C. The muscle and plasma metabolites before and after a bout of exhaustive exercise (produced by chasing) were also measured. The fish were acclimated to hypoxia (48 h at 1.0 mg L−1, 12.5% air saturation), exercise training (2 weeks at 60% of U crit, 6 h daily) or fasting (2 weeks). All treatments resulted in significantly lower resting oxygen consumption ( $$ {\dot{\text{M}}} $$ O2rest) but had no effect on the magnitude of EPOC. Hypoxia acclimation had no effect on U crit or peak post-exercise oxygen consumption ( $$ {\dot{\text{M}}} $$ O2peak) but produced a higher depletion of muscle [glycogen] post-chasing (P

The objective of this study was to determine whether acclimation to hypoxia and sustained exercise would increase hypoxia tolerance (as indicated by a decrease in critical oxygen tension, P crit) and swimming performance in goldfish ( Carassius auratus ), and to investigate the relationship between changes in performance and gill remodelling and tissue metabolic capacity. Goldfish were acclimated to either hypoxia (48 h at 0.3 mg O2 l–1) or sustained exercise (48 h at 70% of critical swimming speed, U crit) and then P crit and U crit were determined in normoxia (10 mg O2 l–1) and hypoxia (1 mg O2 l–1) and compared with values from control fish. Acclimation to both hypoxia and sustained exercise improved hypoxia tolerance ( P crit was reduced by 49% and 39%, respectively), which was associated with an increase in lamellar surface area (71% and 43%, respectively) and an increase in blood \[Hb\] (26% in both groups). Exercise acclimation also resulted in a decrease in routine ![Graphic][1] (![Graphic][2] ). Acclimation to both hypoxia and sustained exercise resulted in a significant increase in U crit in hypoxia (18% and 17%, respectively), which was associated with an increase in maximal O2 consumption rate at U crit (![Graphic][3] ; 35% and 39%, respectively). While hypoxia acclimation resulted in an increase in U crit in normoxia, acclimation to sustained exercise did not improve subsequent swimming performance in normoxia. This lack of improvement was possibly due to depleted oxidizable substrates during exercise acclimation. [1]: /embed/inline-graphic-1.gif [2]: /embed/inline-graphic-2.gif [3]: /embed/inline-graphic-3.gif

To investigate the effect of fasting on maintenance metabolism, feeding metabolism and aerobic swimming performance as well as their metabolic interactions in juvenile southern catfish, we measured the following: (1) the postprandial oxygen consumption ( M ˙ O 2 ) response (16% body mass meal size) after 0 (control), 1, 2 and 4 weeks of fasting and (2) the swimming performance of non-digesting and digesting fish after either 0, 1, 2 or 4 weeks of fasting. The fasting groups displayed with lower resting M ˙ O 2 ( M ˙ O 2 rest), lower peak postprandial M ˙ O 2 ( M ˙ O 2 peak), larger energy expenditures and longer digestive processes than those of the control groups. The critical swimming speed (Ucrit), the active M ˙ O 2 ( M ˙ O 2 active) and the metabolic scope ( M ˙ O 2 active - M ˙ O 2 rest, MS) of both non-digesting and digesting fish all decreased progressively after 1, 2 and 4 weeks of fasting, with those of non-digesting fish decreased more acutely than digesting fish (P M ˙ O 2 active than that of non-digesting fish in the 0-, 1-, 2- and 4-week fasting groups. Digestion only caused a significantly lower Ucrit and MS in both the 0- and 1-week fasting groups (P M ˙ O 2 increased greatly with the swimming speed, and digestion caused a higher M ˙ O 2 when compared to that of the fasting fish in any groups. The M ˙ O 2 of fish in the 4-week fasting group was significantly lower than that of other groups when the swimming speed was the same. In conclusion, both digestive and locomotive functions were down-regulated during fasting. In the 0- and 1-week fasting groups, the decreased MS for swimming during digestion caused a lower Ucrit (i.e., a digestion priority model). However, because the M ˙ O 2 active of digesting fish decreased much more slowly than that of fasting fish, the MS of the 2- and 4-week fasting groups did not change during digestion, and the fish could handle both physiological activities independently (i.e., an additive model).

To examine the effects of exercise training on the metabolic interaction between digestion and locomotion in juvenile darkbarbel catfish (Peltebagrus vachelli) (5.58+/-0.04 g), the postprandial metabolic response, critical swimming speeds (U(crit)) and oxygen consumption rates (VO(2)) during swimming were measured on fish held at a constant temperature (25 degrees C). Fish were fed a diet of cutlets of freshly killed loach. Fish in the trained group were forced to swim at 60% U(crit) for 50 min followed by an exhaustive 10-min chase once daily for 21 days. Exercise training did not produce significant differences in resting VO(2) (VO(2rest)) and postprandial peak VO(2) (VO(2peak)) compared to the non-trained groups. However, exercise training elicited a significant decrease in both the duration and energy expenditure of digestion when fed with similar food (P

To test whether the effects of feeding on swimming performance vary with acclimation temperature in juvenile southern catfish (Silurus meridionalis), we investigated the specific dynamic action (SDA) and swimming performance of fasting and feeding fish at acclimation temperatures of 15, 21, 27, and 33 degrees C. Feeding had no effect on the critical swimming speeding (U(crit)) of fish acclimated at 15 degrees C (p=0.66), whereas it elicited a 12.04, 18.70, and 20.98% decrease in U(crit) for fish acclimated at 21, 27 and 33 degrees C, respectively (p

The critical swimming speed (U(crit), aerobic swimming performance) and endurance (anaerobic swimming performance) of juvenile southern catfish Silurus meridionalis Chen (9.8+/-0.1 cm body length and 8.09+/-0.17 g body mass, n=226) were investigated at 10, 15, 20, 25, and 30 degrees C. Both absolute U(crit) (cm s(-1)) and relative critical swimming speed (U(r), BL s(-1)) of juvenile southern catfish increased in the temperature zone from 10 to 25 degrees C (p

To investigate the effects of exhaustive chasing training and detraining on the swimming performance of juvenile darkbarbel catfish (Peltebagrus vachelli Richardson), we performed exhaustive chasing training daily for 14 days and subsequently detrained fish for 7 days. Chasing training resulted in significant increases in critical swimming speed (U (crit)), post-chasing peak oxygen consumption rate (VO(2 peak)), and heart and gill indexes compared with non-trained controls. Both resting oxygen consumption (VO(2 rest)) and excess post-chasing VO(2) (EPOC) were unaffected by exhaustive chasing training. Fish that underwent chasing training had lower levels of whole-body lipid content and reduced food intake and growth compared with non-trained control fish; however, condition factor was not affected by chasing training. Seven days of detraining reversed the effects of exhaustive chasing training. Overall, these data suggested that short-term exhaustive chasing training improves aerobic swimming capacity in darkbarbel catfish, but the training effects are reversible over a short period of time.

Effects of feeding on pre-exercise VO(2) and excess post-exercise oxygen consumption (EPOC) after exhaustive exercise were investigated in sedentary southern catfish, active herbivorous grass carp, omnivorous crucian carp, and sluggish omnivorous darkbarbel catfish to test whether feeding had different effects on EPOC and to compare EPOC in fishes with different ecological habits. For fasting fish, the pre-exercise and peak post-exercise VO(2) were higher and recovery rates were faster in crucian carp and grass carp compared to those of darkbarbel catfish and southern catfish. EPOC magnitudes of grass carp and southern catfish were significantly larger than those of crucian carp and darkbarbel catfish. Feeding had no significant effect on peak post-exercise VO(2), recovery rate, and EPOC magnitude in grass carp. Both the pre-exercise and peak post-exercise VO(2) increased with meal size, while the EPOC magnitude and duration decreased significantly in the larger meal size groups of crucian carp and southern catfish. In darkbarbel catfish, both the pre-exercise and peak post-exercise VO(2) increased with meal size, but the VO(2) increment elicited by exercise was larger in feeding groups compared with the fasting group. These results suggest that (1) the characteristics of the post-exercise VO(2) profile, such as peak post-exercise VO(2) and recovery rate, were closely related to the activity of fishes, whereas the EPOC magnitude was not and (2) the effects of feeding on EPOC were more closely related to the postprandial increase in VO(2).

Variations of 12 morphological characters and 78 isozymic bands among 78 isolates of five Fusarium spp. from Dongtan wetland were described and analysed with cladistic parsimony and phenetic UPGMA methods. Hierarchical cluster analysis of 12 morphological characters grouped 78 strains into five defined species with a high overlap between isolates. Hierarchical cluster analysis of isozyme patterns showed a higher degree of relationship among five Fusarium spp., in which Fusarium nivale, Fusarium semitectum and Fusarium oxysporum clustered as one group, and F. semitectum was closer to F. nivale than to F. oxysporum; Fusarium graminearum and Fusarium moniliforme formed one group and showed clearly distinct from the first group. Groups of individual isolates indicated by a plot of principal component analysis were consistent with these findings. The comparison of two different data sets revealed that isozyme patterns showed higher variations between species and among individual isolates than morphological characters. Parsimony analysis of morphological characters yielded unresolved cladograms. Parsimony analysis of isozymes as presence/absence characters revealed the same five species in general as the results indicated by phenetic analysis, differing in the relative position of species in subclusters.

A rapid spectroscopic approach for whole-organism fingerprinting of Fourier transform infrared (FT-IR) spectroscopy was used to analyse 16 isolates from five closely related species of Fusarium: F. graminearum, F. moniliforme, F. nivale, F. semitectum and F. oxysporum. Principal components analysis and hierarchical cluster analysis were used to study the clusters in the data. On visual inspection of the clusters from both methods, the spectra were not differentiated into five separate clusters corresponding to species and these unsupervised methods failed to identify these fungal strains. When the data were trained by back propagation algorithm of artificial neural networks (ANNs) with principal components scores of spectra used as input modes, the strains were accurately predicted and recognized. The results in this study show that FT-IR spectroscopy in combination with principal component artificial neural networks (PC-ANNs) is well suited for identifying Fusarium spp. It would be advantageous to establish a comprehensive database of taxonomically well-defined Fusarium species to aid the identification of unknown strains.

The effect of purified macronutrients on specific dynamic action (SDA) was assessed in juvenile southern catfish (39.81 ± 1.35 g) at 27.5 � C by stuffing iso-energetic (5 kJ) casein, corn oil, corn starch in sections of grass carp intestine (0.25 g). A control (without filler) and mixed diets were also made (each macronutrient provide one-third energy). The postprandial metabolic response, i.e. the oxygen consumption curve, was markedly different among fish fed different food. The peak postprandial metabolic rate of fish fed mixed diet (76.72 mgO2 h )1 ) was significantly higher than that of protein group (66.08 mgO2 h )1 ), while the latter was significantly higher than those of other groups (56.11 mgO2 h )1 for lipid, 54.00 mgO2 h )1 for carbohydrate and 56.09 mgO2 h )1 for control) (P < 0.05). The SDA durations were 32, 32, 30, 14 and 10 h for mixed, casein, corn oil, corn starch and control groups, respectively. The SDA coefficient (energy expended on SDA as per cent of ingested energy) of casein (9.41% 2 ) was significantly higher than that of mixed diet (7.43%) (P < 0.05), while the latter was significantly higher than those of corn oil (3.97%) and corn starch (0.84%) (P < 0.05). The results of this study suggested that the method of using casing on investigation of SDA in carnivorous fish was feasible. The SDA response of carnivorous southern catfish fed protein was higher than lipid while latter was higher than carbohydrate (nearly to zero). The result of this study may give interesting information for understanding the physiological mechanism under SDA.

The effect of relative meal size (0.5-24% body mass) on specific dynamic action (SDA) was assessed in Chinese catfish (Silurus asotus Linnaeus) (30.90+/-1.30 g) at 25.0 degrees C; the cutlets of freshly killed loach without viscera, head and tail were used as a test meal. There was no significant difference in either SDA duration or peak oxygen consumption (VO2) among low meal size ranges. But both increased linearly as meal size increased from 2 to 24% without reaching a plateau. Factorial metabolic scope was 5.92 in fish fed with 24% body mass, the highest documented feeding metabolic scope value in fish till now. The Peak VO2 of satiated meal size groups (175.85+/-10.55 mg O2 h(-1)) was above 80% of maximum metabolic rate during locomotion recovery process (215.48+/-7.07 mg O2 h(-1)). The relationship between energy expended on SDA (E) and energy ingested (I) was described as: E=0.0000432I(2)+0.140I+2.12. The lowest value of SDA coefficient appeared at 2% body mass group.

Glioblastoma is one of the most aggressive human cancers with poor prognosis, and therefore a critical need exists for novel therapeutic strategies for management of glioblastoma patients. Itraconazole, a traditional antifungal drug, has been identified as a novel potential anticancer agent due to its inhibitory effects on cell proliferation and tumor angiogenesis; however, the molecular mechanisms involved are still unclear. Here, we show that itraconazole inhibits the proliferation of glioblastoma cells both in vitro and in vivo. Notably, we demonstrate that treatment with itraconazole induces autophagic progression in glioblastoma cells, while blockage of autophagy markedly reverses the antiproliferative activities of itraconazole, suggesting an antitumor effect of autophagy in response to itraconazole treatment. Functional studies revealed that itraconazole retarded the trafficking of cholesterol from late endosomes and lysosomes to the plasma membrane by reducing the levels of SCP2, resulting in repression of AKT1-MTOR signaling, induction of autophagy, and finally inhibition of cell proliferation. Together, our studies provide new insights into the molecular mechanisms regarding the antitumor activities of itraconazole, and may further assist both the pharmacological investigation and rational use of itraconazole in potential clinical applications.