An insulin-regulated arrestin domain protein controls hepatic glucagon action.

Glucagon signaling is essential for maintaining normoglycemia in mammals. The arrestin fold superfamily of proteins controls the trafficking, turnover, and signaling of transmembrane receptors as well as other intracellular signaling functions. Further investigation is needed to understand the in vivo functions of the arrestin domain-containing 4 (ARRDC4) protein family member and whether it is involved in mammalian glucose metabolism. Here, we show that mice with a global deletion of the ARRDC4 protein have impaired glucagon responses and gluconeogenesis at a systemic and molecular level. Mice lacking ARRDC4 exhibited lower glucose levels after fasting and could not suppress gluconeogenesis at the refed state. We also show that ARRDC4 coimmunoprecipitates with the glucagon receptor, and ARRDC4 expression is suppressed by insulin. These results define ARRDC4 as a critical regulator of glucagon signaling and glucose homeostasis and reveal a novel intersection of insulin and glucagon pathways in the liver.

Complete genome analysis of a novel narnavirus in sweet viburnum (Viburnum odoratissimum).

Trees and shrubs provide important ecological services. However, few studies have surveyed the virome in trees and shrubs. In this study, we discovered a new positive-sense RNA virus originating from Viburnum odoratissimum, which we named "Vo narna-like virus". The complete genome of Vo narna-like virus is 3,451 nt in length with an open reading frame (ORF) encoding the RNA-dependent RNA polymerase (RdRP) protein. Phylogenetic analysis placed this virus within the betanarnavirus clade, sharing 53.63% amino acid sequence identity with its closest relative, Qingdao RNA virus 2. The complete sequence of the virus was confirmed by rapid amplification of cDNA ends (RACE) and Sanger sequencing. Small interfering RNA (siRNA) analysis indicated that this virus interacts with the RNA interference (RNAi) pathway of V. odoratissimum. This is the first report of a narnavirus in V. odoratissimum.

The Sapria himalayana genome provides new insights into the lifestyle of endoparasitic plants.

BACKGROUND: Sapria himalayana (Rafflesiaceae) is an endoparasitic plant characterized by a greatly reduced vegetative body and giant flowers; however, the mechanisms underlying its special lifestyle and greatly altered plant form remain unknown. To illustrate the evolution and adaptation of S. himalayasna, we report its de novo assembled genome and key insights into the molecular basis of its floral development, flowering time, fatty acid biosynthesis, and defense responses. RESULTS: The genome of S. himalayana is ~ 1.92 Gb with 13,670 protein-coding genes, indicating remarkable gene loss (~ 54%), especially genes involved in photosynthesis, plant body, nutrients, and defense response. Genes specifying floral organ identity and controlling organ size were identified in S. himalayana and Rafflesia cantleyi, and showed analogous spatiotemporal expression patterns in both plant species. Although the plastid genome had been lost, plastids likely biosynthesize essential fatty acids and amino acids (aromatic amino acids and lysine). A set of credible and functional horizontal gene transfer (HGT) events (involving genes and mRNAs) were identified in the nuclear and mitochondrial genomes of S. himalayana, most of which were under purifying selection. Convergent HGTs in Cuscuta, Orobanchaceae, and S. himalayana were mainly expressed at the parasite-host interface. Together, these results suggest that HGTs act as a bridge between the parasite and host, assisting the parasite in acquiring nutrients from the host. CONCLUSIONS: Our results provide new insights into the flower development process and endoparasitic lifestyle of Rafflesiaceae plants. The amount of gene loss in S. himalayana is consistent with the degree of reduction in its body plan. HGT events are common among endoparasites and play an important role in their lifestyle adaptation.

Chromosome-level assembly of the Neolamarckia cadamba genome provides insights into the evolution of cadambine biosynthesis.

Neolamarckia cadamba (Roxb.), a close relative of Coffea canephora and Ophiorrhiza pumila, is an important traditional medicine in Southeast Asia. Three major glycosidic monoterpenoid indole alkaloids (MIAs), cadambine and its derivatives 3ß-isodihydrocadambine and 3ß-dihydrocadambine, accumulate in the bark and leaves, and exhibit antimalarial, antiproliferative, antioxidant, anticancer and anti-inflammatory activities. Here, we report a chromosome-scale N. cadamba genome, with 744.5 Mb assembled into 22 pseudochromosomes with contig N50 and scaffold N50 of 824.14 Kb and 29.20 Mb, respectively. Comparative genomic analysis of N. cadamba with Co. canephora revealed that N. cadamba underwent a relatively recent whole-genome duplication (WGD) event after diverging from Co. canephora, which contributed to the evolution of the MIA biosynthetic pathway. We determined the key intermediates of the cadambine biosynthetic pathway and further showed that NcSTR1 catalyzed the synthesis of strictosidine in N. cadamba. A new component, epoxystrictosidine (C27H34N2O10, m/z 547.2285), was identified in the cadambine biosynthetic pathway. Combining genome-wide association study (GWAS), population analysis, multi-omics analysis and metabolic gene cluster prediction, this study will shed light on the evolution of MIA biosynthetic pathway genes. This N. cadamba reference sequence will accelerate the understanding of the evolutionary history of specific metabolic pathways and facilitate the development of tools for enhancing bioactive productivity by metabolic engineering in microbes or by molecular breeding in plants.

Decoupling subgenomes within hybrid lavandin provide new insights into speciation and monoterpenoid diversification of Lavandula.

Polyploidization and transposon elements contribute to shape plant genome diversity and secondary metabolic variation in some edible crops. However, the specific contribution of these variations to the chemo-diversity of Lamiaceae, particularly in economic shrubs, is still poorly documented. The rich essential oils (EOs) of Lavandula plants are distinguished by monoterpenoids among the main EO-producing species, L. angustifolia (LA), L. × intermedia (LX) and L. latifolia (LL). Herein, the first allele-aware chromosome-level genome was assembled using a lavandin cultivar 'Super' and its hybrid origin was verified by two complete subgenomes (LX-LA and LX-LL). Genome-wide phylogenetics confirmed that LL, like LA, underwent two lineage-specific WGDs after the γ triplication event, and their speciation occurred after the last WGD. Chloroplast phylogenetic analysis indicated LA was the maternal source of 'Super', which produced premium EO (higher linalyl/lavandulyl acetate and lower 1,8-cineole and camphor) close to LA. Gene expression, especially the monoterpenoid biosynthetic genes, showed bias to LX-LA alleles. Asymmetric transposon insertions in two decoupling 'Super' subgenomes were responsible for speciation and monoterpenoid divergence of the progenitors. Both hybrid and parental evolutionary analysis revealed that LTR (long terminal repeat) retrotransposon associated with AAT gene loss cause no linalyl/lavandulyl acetate production in LL, and multi-BDH copies retained by tandem duplication and DNA transposon resulted in higher camphor accumulation of LL. Advances in allelic variations of monoterpenoids have the potential to revolutionize future lavandin breeding and EO production.

The Gastrodia menghaiensis (Orchidaceae) genome provides new insights of orchid mycorrhizal interactions.

BACKGROUND: To illustrate the molecular mechanism of mycoheterotrophic interactions between orchids and fungi, we assembled chromosome-level reference genome of Gastrodia menghaiensis (Orchidaceae) and analyzed the genomes of two species of Gastrodia. RESULTS: Our analyses indicated that the genomes of Gastrodia are globally diminished in comparison to autotrophic orchids, even compared to Cuscuta (a plant parasite). Genes involved in arbuscular mycorrhizae colonization were found in genomes of Gastrodia, and many of the genes involved biological interaction between Gatrodia and symbiotic microbionts are more numerous than in photosynthetic orchids. The highly expressed genes for fatty acid and ammonium root transporters suggest that fungi receive material from orchids, although most raw materials flow from the fungi. Many nuclear genes (e.g. biosynthesis of aromatic amino acid L-tryptophan) supporting plastid functions are expanded compared to photosynthetic orchids, an indication of the importance of plastids even in totally mycoheterotrophic species. CONCLUSION: Gastrodia menghaiensis has the smallest proteome thus far among angiosperms. Many of the genes involved biological interaction between Gatrodia and symbiotic microbionts are more numerous than in photosynthetic orchids.

An all-in-one magnetic SERS nanosensor for ratiometric detection of Escherichia coli in foods.

An all-in-one nanosensor was developed for the magnetic enrichment and ratiometric surface-enhanced Raman scattering (SERS) detection of Escherichia coli (E. coli). The all-in-one nanosensor was constructed through the chemical integration of four components into a single nanoparticle, which include a manganese ferrite nanoparticle serving as the magnetic core, a thin silver shell as the SERS substrate, a self-assembled layer of 4-mercaptobenzoic acid (MBA) molecules as the SERS internal standard, and a MBA-conjugated layer of aptamer sequences as the capture probe of E. coli. In the detection of E. coli in food, the target cells were first captured by the nanosensors and magnetically enriched in a short time of 15 min, and then the ratiometric SERS was performed through the Raman intensity ratio between two specific SERS peaks produced by the captured E. coli and the internal MBA. The pre-concentration and ratiometry enabled the nanosensor to detect E. coli with a detection limit down to 10 CFU/mL. The all-in-one nanosensor was successfully applied for the detection of E. coli in various liquid foods including milk, juice, tea, and coffee, with recoveries ranging from 89 to 110% and relative standard deviation lower than 1.7%. In comparison with the previous sandwich strategy adopted by most SERS sensors, this nanosensor endowed with an easier use and a lower cost is more sensitive and reproducible, leading to a great potential in practical applications.

Wnt/ß-catenin signaling pathway participates in the effect of miR-626 on oral squamous cell carcinoma by targeting RASSF4.

BACKGROUND: The role of miR-626 in oral squamous cell carcinoma (OSCC) was investigated by targeting RASSF4. METHODS: The miR-626 and RASSF4 expression was detected in normal oral mucosa or OSCC tissues and OSCC or normal cells. The methylation status of RASSF4 was analyzed using methylation-specific polymerase chain reaction (PCR). The cytoplasmic/nuclear ratios (C/N ratios) targeted by miR-626 were examined using microarray, followed by a dual-luciferase reporter assay. The subcellular localization of RASSF4 and miR-626 in OSCC cells was determined using RNA fluorescence in situ hybridization (FISH) and immunocytochemistry (ICC), respectively. Ca9-22 and HSC2 cells were divided into mock, inhibitor NC, miR-626 inhibitor, scramble, RASSF4 and miR-626 mimic + RASSF4 groups, and then CCK-8, Annexin V-FITC/PI, wound healing, Transwell, qRT-PCR and western blotting assays were performed. RESULTS: OSCC tissues and cells had increased miR-626 expression and decreased RASSF4 expression. Patients with RASSF4 methylation had lower RASSF4 expression than those without methylation. In addition, a negative correlation between miR-626 and RASSF4 was found in OSCC tissues, both of which were correlated with the pathological grade, pathological stage, lymph node metastasis and patient prognosis. MiR-626 targeted RASSF4 in OSCC cells. Overexpressed RASSF4 inhibited the proliferation, invasion, migration and epithelial-mesenchymal transition (EMT) of OSCC cells, promoted cell apoptosis, and blocked the Wnt/ß-Catenin pathway, which was reversed by miR-626 overexpression. CONCLUSIONS: Inhibiting miR-626 can regulate the biological characteristics of OSCC cells, including proliferation, invasion, migration, EMT and apoptosis, by targeting RASSF4, which may be related to the Wnt/ß-Catenin pathway.

Hepatocyte-Specific Deficiency of BAP31 Amplified Acetaminophen-Induced Hepatotoxicity via Attenuating Nrf2 Signaling Activation in Mice.

Liver-specific deficiency of B-cell receptor-associated protein 31 knockout mice (BAP31-LKO) and the littermates were injected with acetaminophen (APAP), markers of liver injury, and the potential molecular mechanisms were determined. In response to APAP overdose, serum aspartate aminotransferase and alanine aminotransferase levels were increased in BAP31-LKO mice than in wild-type controls, accompanied by enhanced liver necrosis. APAP-induced apoptosis and mortality were increased. Hepatic glutathione was decreased (1.60 ± 0.31 µmol/g tissue in WT mice vs. 0.85 ± 0.14 µmol/g tissue in BAP31-LKO mice at 6 h, p < 0.05), along with reduced glutathione reductase activity and superoxide dismutase; while malondialdehyde was significantly induced (0.41 ± 0.03 nmol/mg tissue in WT mice vs. 0.50 ± 0.05 nmol/mg tissue in BAP31-LKO mice for 6 h, p < 0.05). JNK signaling activation and APAP-induced hepatic inflammation were increased in BAP31-LKO mice. The mechanism research revealed that BAP31-deficiency decreased Nrf2 mRNA stability (half-life of Nrf2 mRNA decreased from ~1.3 h to ~40 min) and miR-223 expression, led to reduced nuclear factor erythroid 2-related factor 2 (Nrf2) signaling activation and antioxidant genes induction. BAP31-deficiency decreased mitochondrial membrane potentials, reduced mitochondria-related genes expression, and resulted in mitochondrial dysfunction in the liver. Conclusions: BAP31-deficiency reduced the antioxidant response and Nrf2 signaling activation via reducing Nrf2 mRNA stabilization, enhanced JNK signaling activation, hepatic inflammation, and apoptosis, amplified APAP-induced hepatotoxicity in mice.

CRISPR-delivery particles targeting nuclear receptor-interacting protein 1 (Nrip1) in adipose cells to enhance energy expenditure.

RNA-guided, engineered nucleases derived from the prokaryotic adaptive immune system CRISPR-Cas represent a powerful platform for gene deletion and editing. When used as a therapeutic approach, direct delivery of Cas9 protein and single-guide RNA (sgRNA) could circumvent the safety issues associated with plasmid delivery and therefore represents an attractive tool for precision genome engineering. Gene deletion or editing in adipose tissue to enhance its energy expenditure, fatty acid oxidation, and secretion of bioactive factors through a "browning" process presents a potential therapeutic strategy to alleviate metabolic disease. Here, we developed "CRISPR-delivery particles," denoted CriPs, composed of nano-size complexes of Cas9 protein and sgRNA that are coated with an amphipathic peptide called Endo-Porter that mediates entry into cells. Efficient CRISPR-Cas9-mediated gene deletion of ectopically expressed GFP by CriPs was achieved in multiple cell types, including a macrophage cell line, primary macrophages, and primary pre-adipocytes. Significant GFP loss was also observed in peritoneal exudate cells with minimum systemic toxicity in GFP-expressing mice following intraperitoneal injection of CriPs containing Gfp-targeting sgRNA. Furthermore, disruption of a nuclear co-repressor of catabolism, the Nrip1 gene, in white adipocytes by CriPs enhanced adipocyte browning with a marked increase of uncoupling protein 1 (UCP1) expression. Of note, the CriP-mediated Nrip1 deletion did not produce detectable off-target effects. We conclude that CriPs offer an effective Cas9 and sgRNA delivery system for ablating targeted gene products in cultured cells and in vivo, providing a potential therapeutic strategy for metabolic disease.

Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.

Obesity-mediated inflammation is a major cause of insulin resistance, and macrophages play an important role in this process. The 78-kDa glucose-regulated protein (GRP78) is a major endoplasmic reticulum chaperone that modulates unfolded protein response (UPR), and mice with GRP78 heterozygosity were resistant to diet-induced obesity. Here, we show that mice with macrophage-selective ablation of GRP78 (Lyz- GRP78-/-) are protected from skeletal muscle insulin resistance without changes in obesity compared with wild-type mice after 9 wk of high-fat diet. GRP78-deficient macrophages demonstrated adapted UPR with up-regulation of activating transcription factor (ATF)-4 and M2-polarization markers. Diet-induced adipose tissue inflammation was reduced, and bone marrow-derived macrophages from Lyz- GRP78-/- mice demonstrated a selective increase in IL-6 expression. Serum IL-13 levels were elevated by >4-fold in Lyz- GRP78-/- mice, and IL-6 stimulated the myocyte expression of IL-13 and IL-13 receptor. Lastly, recombinant IL-13 acutely increased glucose metabolism in Lyz- GRP78-/- mice. Taken together, our data indicate that GRP78 deficiency activates UPR by increasing ATF-4, and promotes M2-polarization of macrophages with a selective increase in IL-6 secretion. Macrophage-derived IL-6 stimulates the myocyte expression of IL-13 and regulates muscle glucose metabolism in a paracrine manner. Thus, our findings identify a novel crosstalk between macrophages and skeletal muscle in the modulation of obesity-mediated insulin resistance.-Kim, J. H., Lee, E., Friedline, R. H., Suk, S., Jung, D. Y., Dagdeviren, S., Hu, X., Inashima, K., Noh, H. L., Kwon, J. Y., Nambu, A., Huh, J. R., Han, M. S., Davis, R. J., Lee, A. S., Lee, K. W., Kim, J. K. Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.

De novo transcriptome assembly of the eight major organs of Sacha Inchi (Plukenetia volubilis) and the identification of genes involved in α-linolenic acid metabolism.

BACKGROUND: Sacha Inchi (Plukenetia volubilis L.), which belongs to the Euphorbiaceae, has been considered a new potential oil crop because of its high content of polyunsaturated fatty acids in its seed oil. The seed oil especially contains high amounts of α-linolenic acid (ALA), which is useful for the prevention of various diseases. However, little is known about the genetic information and genome sequence of Sacha Inchi, which has largely hindered functional genomics and molecular breeding studies. RESULTS: In this study, a de novo transcriptome assembly based on transcripts sequenced in eight major organs, including roots, stems, shoot apexes, mature leaves, male flowers, female flowers, fruits, and seeds of Sacha Inchi was performed, resulting in a set of 124,750 non-redundant putative transcripts having an average length of 851 bp and an N50 value of 1909 bp. Organ-specific unigenes analysis revealed that the most organ-specific transcripts are found in female flowers (2244 unigenes), whereas a relatively small amount of unigenes are detected to be expressed specifically in other organs with the least in stems (24 unigenes). A total of 42,987 simple sequence repeats (SSRs) were detected, which will contribute to the marker assisted selection breeding of Sacha Inchi. We analyzed expression of genes related to the α-linolenic acid metabolism based on the de novo assembly and annotation transcriptome in Sacha Inchi. It appears that Sacha Inchi accumulates high level of ALA in seeds by strong expression of biosynthesis-related genes and weak expression of degradation-related genes. In particular, the up-regulation of FAD3 and FAD7 is consistent with high level of ALA in seeds of Sacha Inchi compared with in other organs. Meanwhile, several transcription factors (ABI3, LEC1 and FUS3) may regulate key genes involved in oil accumulation in seeds of Sacha Inchi. CONCLUSIONS: The transcriptome of major organs of Sacha Inchi has been sequenced and de novo assembled, which will expand the genetic information for functional genomic studies of Sacha Inchi. In addition, the identification of candidate genes involved in ALA metabolism will provide useful resources for the genetic improvement of Sacha Inchi and the metabolic engineering of ALA biosynthesis in other plants.

IL-10 prevents aging-associated inflammation and insulin resistance in skeletal muscle.

Altered energy balance and insulin resistance are important characteristics of aging. Skeletal muscle is a major site of glucose disposal, and the role of aging-associated inflammation in skeletal muscle insulin resistance remains unclear. To investigate, we examined glucose metabolism in 18-mo-old transgenic mice with muscle-specific overexpression of IL-10 (MIL10) and in wild-type mice during hyperinsulinemic-euglycemic clamping. Despite similar fat mass and energy balance, MIL10 mice were protected from aging-associated insulin resistance with significant increases in glucose infusion rates, whole-body glucose turnover, and skeletal muscle glucose uptake (∼60%; P < 0.05), as compared to age-matched WT mice. This protective effect was associated with decreased muscle inflammation, but no changes in adipose tissue inflammation in aging MIL10 mice. These results demonstrate the importance of skeletal muscle inflammation in aging-mediated insulin resistance, and our findings further implicate a potential therapeutic role of anti-inflammatory cytokine in the treatment of aging-mediated insulin resistance.-Dagdeviren, S., Jung, D. Y., Friedline, R. H., Noh, H. L., Kim, J. H., Patel, P. R., Tsitsilianos, N., Inashima, K., Tran, D. A., Hu, X., Loubato, M. M., Craige, S. M., Kwon, J. Y., Lee, K. W., Kim, J. K. IL-10 prevents aging-associated inflammation and insulin resistance in skeletal muscle.

Genetic ablation of lymphocytes and cytokine signaling in nonobese diabetic mice prevents diet-induced obesity and insulin resistance.

Obesity is characterized by a dysregulated immune system, which may causally associate with insulin resistance and type 2 diabetes. Despite widespread use of nonobese diabetic (NOD) mice, NOD with severe combined immunodeficiency (scid) mutation (SCID) mice, and SCID bearing a null mutation in the IL-2 common γ chain receptor (NSG) mice as animal models of human diseases including type 1 diabetes, the underlying metabolic effects of a genetically altered immune system are poorly understood. For this, we performed a comprehensive metabolic characterization of these mice fed chow or after 6 wk of a high-fat diet. We found that NOD mice had ∼50% less fat mass and were 2-fold more insulin sensitive, as measured by hyperinsulinemic-euglycemic clamp, than C57BL/6 wild-type mice. SCID mice were also more insulin sensitive with increased muscle glucose metabolism and resistant to diet-induced obesity due to increased energy expenditure (∼10%) and physical activity (∼40%) as measured by metabolic cages. NSG mice were completely protected from diet-induced obesity and insulin resistance with significant increases in glucose metabolism in peripheral organs. Our findings demonstrate an important role of genetic background, lymphocytes, and cytokine signaling in diet-induced obesity and insulin resistance.

Transient receptor potential vanilloid type-1 channel regulates diet-induced obesity, insulin resistance, and leptin resistance.

Insulin resistance is a major characteristic of obesity and type 2 diabetes, but the underlying mechanism is unclear. Recent studies have shown a metabolic role of capsaicin that may be mediated via the transient receptor potential vanilloid type-1 (TRPV1) channel. In this study, TRPV1 knockout (KO) and wild-type (WT) mice (as controls) were fed a high-fat diet (HFD), and metabolic studies were performed to measure insulin and leptin action. The TRPV1 KO mice became more obese than the WT mice after HFD, partly attributed to altered energy balance and leptin resistance in the KO mice. The hyperinsulinemic-euglycemic clamp experiment showed that the TRPV1 KO mice were more insulin resistant after HFD because of the ∼40% reduction in glucose metabolism in the white and brown adipose tissue, compared with that in the WT mice. Leptin treatment failed to suppress food intake, and leptin-mediated hypothalamic signal transducer and activator of transcription (STAT)-3 activity was blunted in the TRPV1 KO mice. We also found that the TRPV1 KO mice were more obese and insulin resistant than the WT mice at 9 mo of age. Taken together, these results indicate that lacking TRPV1 exacerbates the obesity and insulin resistance associated with an HFD and aging, and our findings further suggest that TRPV1 has a major role in regulating glucose metabolism and hypothalamic leptin's effects in obesity.

Preparation and Performance Evaluation of Castor Oil-Based Asphalt Regeneration Agent.

Regeneration agents play a critical role in modifying the mechanical properties and durability of RAP asphalt mixtures. This paper aimed to develop a castor oil-based asphalt regeneration agent. The effects of this regeneration agent on the pavement performance of laboratory-aged asphalt and an RAP asphalt mixture were comparatively studied by a series of laboratory tests. For the developed castor oil-based asphalt regeneration agent, the weight ratio of the castor oil to dibutyl phthalate was determined as 1:4. Moreover, the regeneration effectiveness of the castor oil-based regeneration agent was tested on three laboratory-aged asphalt binders and an RAP asphalt binder; the penetration, softening point and ductility of the RAP asphalt binder recovered to 83 dmm, 50.3 °C, and more than 100 cm, respectively. The optimum content of the regeneration agent was 5% by the weight of the aged asphalt binder. Furthermore, the castor oil-based regeneration agent could effectively restore the pavement performance of an RAP asphalt mixture. In this study, the RAP percentage can reach up to 60% by the weight of the HMA mixture using the castor oil-based asphalt regeneration agent according to the Chinese specification.

The Effect of Water during the Compaction Process on Surface Characteristics of HMA Pavement.

During the compaction process of HMA pavement, it is common to spray cold water on the wheel of a road roller to prevent the mixture from sticking to the wheel, which might deteriorate the bonding strength between the asphalt binder and aggregate, and consequently lead to surface polishing of the pavement. This paper aims to demonstrate whether the water used during the compaction process affects the surface performance of HMA pavement. In this study, the black pixel ratio and mass loss ratio were used to evaluate the water effect on the surface performance of asphalt pavement, considering the water consumption, molding temperature and long-term ageing process. The test results indicated that the water used during the compaction process would increase the risk of surface polishing of HMA pavement. This adverse effect became more significant if the HMA samples were prepared using greater water consumption, a greater molding temperature and a long-term ageing process. Moreover, there exists a certain correlation between the black pixel ratio and mass loss ratio, and their relationships were demonstrated by the experimental results in this study. It is recommended that further research concentrates on the influencing mechanism and the treatment strategy for the adverse effect caused by the water used during the compaction process. The use of more types of asphalt binders, aggregate and methodologies is also recommended in further studies.

Effect of dietary aspirin eugenol ester on the growth performance, antioxidant capacity, intestinal inflammation, and cecal microbiota of broilers under high stocking density.

This study was designed to examine the impact of aspirin eugenol ester (AEE) on the growth performance, serum antioxidant capacity, jejunal barrier function, and cecal microbiota of broilers raised under stressful high density (HD) stocking conditions compared with normal density broilers (ND). A total of 432 one-day-old AA+ male broilers were randomly divided into 4 groups: normal density (ND, 14 broilers /m2), high density (HD, 22 broilers /m2), ND + AEE, and HD + AEE. The results of the study revealed a significant decrease in the growth performance of broiler chickens as a result of HD stress (P < 0.05). The total antioxidant capacity (T-AOC) in serum demonstrated a significant decrease (P < 0.05) at both 28 and 35 d. Conversely, the serum level of malondialdehyde (MDA) exhibited a significant increase (P < 0.05). Dietary supplementation of AEE resulted in a significant elevation (P < 0.05) of serum GSH-PX, SOD and T-AOC activity at both 28 and 35 d. Moreover, exposure to HD stress resulted in a considerable reduction in the height of intestinal villi and mRNA expression of tight junction proteins in the jejunum, along with, a significant elevation in the mRNA expression of inflammatory cytokines (P < 0.05). However, the administration of AEE reversed the adverse effects of HD-induced stress on villus height and suppressed the mRNA expression of the pro-inflammatory genes, COX-2 and mPGES-1. Additionally, the exposure to HD stress resulted in a substantial reduction in the α-diversity of cecal microbiota and disruption in the equilibrium of intestinal microbial composition, with a notable decrease in the relative abundance of Bacteroides and Faecalibacterium (P < 0.05). In contrast, the addition of AEE to the feed resulted in a notable increase in the relative abundance of Phascolarctobacterium and enhanced microbial diversity (P < 0.05). The inclusion of AEE in the diet has been demonstrated to enhance intestinal integrity and growth performance of broilers by effectively mitigating disruptions in gut microbiota induced by HD stress.

GprC of the nematode-trapping fungus Arthrobotrys flagrans activates mitochondria and reprograms fungal cells for nematode hunting.

Initiation of development requires differential gene expression and metabolic adaptations. Here we show in the nematode-trapping fungus, Arthrobotrys flagrans, that both are achieved through a dual-function G-protein-coupled receptor (GPCR). A. flagrans develops adhesive traps and recognizes its prey, Caenorhabditis elegans, through nematode-specific pheromones (ascarosides). Gene-expression analyses revealed that ascarosides activate the fungal GPCR, GprC, at the plasma membrane and together with the G-protein alpha subunit GasA, reprograms the cell. However, GprC and GasA also reside in mitochondria and boost respiration. This dual localization of GprC in A. flagrans resembles the localization of the cannabinoid receptor CB1 in humans. The C. elegans ascaroside-sensing GPCR, SRBC66 and GPCRs of many fungi are also predicted for dual localization, suggesting broad evolutionary conservation. An SRBC64/66-GprC chimaeric protein was functional in A. flagrans, and C. elegans SRBC64/66 and DAF38 share ascaroside-binding sites with the fungal GprC receptor, suggesting 400-million-year convergent evolution.