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Purpose Glaucoma is a leading cause of global irreversible blindness, and characterized by the progressive loss of retinal ganglion cells (RGCs). Ligustrazine (TMP) is a natural product that has shown beneficial effects on various diseases. This study aimed to determine whether ligustrazine produces a therapeutic effect on glaucoma and to investigate its underlying mechanisms. Methods A rat chronic hypertensive glaucoma model was induced by episcleral vein cauterization (EVC). Adult Sprague-Dawley (SD) rats were intraperitoneally administered TMP at a dose of 80 mg/kg once a day, from two days before EVC to one month after EVC. To elucidate the role of the mammalian target of rapamycin (mTOR) and phosphoinositide 3-kinase (PI3K), TMP-treated experimental rats were co-treated with the mTOR inhibitor rapamycin (5 mg/kg) or the PI3K inhibitor Ly294002 (10 mg/kg). The intraocular pressure (IOP) of the experimental and control rats was measured every six days. Retinal cells were examined by hematoxylin-eosin and terminal deoxynucleotidyltransferase-mediated biotinylated UTP nick end labeling (TUNEL) staining, as well as transmission electron microscopy. Immunohistochemistry and western blot analysis were performed to measure proteins involved in apoptosis and autophagy. Results Ligustrazine protected retinal cells from death in experimental glaucoma rats, which was not due to the lowering of IOP, but could be attributable to direct suppression of retinal cell apoptosis. In glaucoma rats, autophagy was markedly activated in retina cells, as evidenced by increased numbers of autophagosomes and the expression of autophagy-related proteins (ATG5 and LC3-II/I). Notably, such alterations in glaucoma rats were almost completely reversed by ligustrazine. The suppressive effects of ligustrazine on apoptosis and autophagy of retina cells were markedly attenuated by the mTOR inhibitor rapamycin or the PI3K inhibitor Ly294002. Additionally, ligustrazine significantly increased the protein levels of phosphorylated PI3K (p-PI3K), protein kinase B (p-Akt), and mTOR (p-mTOR) in glaucoma rats, whereas such increases were attenuated by rapamycin or Ly294002. Conclusions These results demonstrate that ligustrazine is protective in experimental glaucoma by inhibiting autophagy via the activation of the PI3K-Akt/mTOR pathway, providing compelling evidence that ligustrazine is potentially therapeutic for patients with glaucoma.

This research aims to investigate the effect of gemcitabine (GEM) on various activities and functions of macrophages. Phagocytosis, cell autophagy and reactive oxygen species (ROS) were analysed by laser scanning confocal microscope. The cell cycle status and major histocompatibility complex II (MHC-II) expression were examined by flow cytometry. Inflammatory cytokine secretion such as tumour necrosis factor α (TNF-α) and interleukin 6 (IL-6) was detected by Elisa assay. The expression of proteins was analysed by western blot method. The results revealed that GEM-induced immune inhibition of M1-type RAW264.7 macrophages activated by interferon-γ (IFN-γ) and lipopolysaccharide (LPS). We also found that GEM inhibited autophagy, as evidenced by the reduced formation of autophagosome-like vacuoles and autophagosomes. Further study showed that incubation of activated macrophages with the autophagy inhibitor 3-MA induced immune suppression. In contrast, treatment with the autophagy inducer trehalose (Tre) restored phagocytosis, TNF-α and IL-6 secretion, and MHC-II expression in GEM-induced immune-inhibited macrophages. GEM reduced immune effect of M1-type RAW264.7 macrophages via inhibiting TNF-α, IL-6 and MHC-II expression. Furthermore, activation of autophagy by Tre reversed GEM-induced immune inhibition of RAW264.7 macrophages. Supplementary Information The online version contains supplementary material available at 10.1007/s12026-021-09210-7.

In tropical and subtropical countries, dengue virus (DENV) infections have been increasing; however, we still lack effective therapy. In the present study, we aimed to engineer a bispecific antibody (subsequently named LUZ-8F2-6B1), based on monoclonal antibody 6B1, which has anti DENV-1, 2, and 3 activity, and 8F2, which has anti DENV-4 activity. LUZ-8F2-6B1 displayed potent neutralization activity against four serotypes of DENV by binding to the envelop protein. In vivo, we demonstrated that LUZ-8F2-6B1 could provide protection against infection by four serotypes of DENV in a mouse model. In addition, the deletion of nine amino acids in the Fc region (LUZ-8F2-6B1-9del) completely abolished the antibody-dependent enhancement observed at lower doses of the antibody. Thus, LUZ-8F2-6B1 is a promising, safe, and effective agent for the prophylaxis and treatment of DENV infection.

Bacillus thuringiensis Cry proteins have been widely used over the past decades for many different insect pests, which are safe for users and the environment. The coleopteran-specific Cry3Aa toxin from B. thuringiensis exhibits toxicity to the larvae of Monochamus alternatus. Receptors play a key role in the mechanisms underlying the toxic action of Cry. However, the binding receptor for Cry3Aa has yet to be identified in the midgut of M. alternatus larvae. Therefore, the aim of this study was to identify the receptor for Cry3Aa toxin in the brush border membrane vesicles (BBMVs) of M. alternatus larvae. Our results indicate that the Cry3Aa toxin binds to the BBMVs (Kd = 247 nM) of M. alternatus via a 107 kDa aminopeptidase N (APN) (Kd = 57 nM). In silico analysis of the APN protein predicted that an 18 amino acid sequence in the N-terminal acted as a signal peptide, and that the Asn residue, located at position 918 in the C-terminus is an anchored site for glycosyl phosphatidyl inositol. Further analysis showed that M. alternatus APN exhibits 75% homology to the APN from Anoplophora glabripenis. Our work, therefore, confirmed that APN, which is localized in the BBMVs in the midgut of M. alternatus larvae, acts as a binding protein for Cry3Aa toxins.

Kai Jia,* Yan Wang,* Xin Tong, Rong Wang Department of Nutrition, Beijing Chao-Yang Hospital, Capital Medical University, Beijing, People’s Republic of China*These authors contributed equally to this workCorrespondence: Kai JiaDepartment of Nutrition, Beijing Chao-Yang Hospital, No. 8 Gongti South Road, Chaoyang District, Beijing 100020, People’s Republic of ChinaTel +86-85231275Fax +86-10-83911276Email kaijiamyemail@sina.comIntroduction: KGF-modified MSCs can promote the repair of spinal cord injury and pulmonary fibrosis injury in rats. However, the effect of KGF-modified MSCs on UC rats is unclear. We aimed to explore the therapeutic effect and possible mechanism of KGF gene-modified MSCs on trinitrobenzene sulfonic acid (TNBS)-induced UC rats.Methods: The lentivirus-mediated KGF gene was introduced into bone marrow MSCs of male rats. Female SD rats were induced to establish a UC model by TNBS. Untreated MSCs, MSCs carrying empty vectors (MSCs-vec) or MSCs carrying KGF gene (MSCs-KGF) were transplanted into UC rats by tail vein injection.Results: Significantly high expression of KGF was observed in the intestinal tissues of the MSCs-KGF group. Compared with the challenged control group, the DAI score, CMDI score and TDI score of the MSCs group, MSCs-vec group and MSCs-KGF group were markedly lower. Treatment with MSCs obviously promoted the expression of claudin-1 and PCNA in intestinal tissues of UC rats. Simultaneously, compared with the challenged control group, the levels of TNF-α, IL-6 and IL-8 in the intestinal tissues of the MSCs groups were significantly decreased, while the levels of IL-10 were significantly increased. Most importantly, we found that MSCs-KGF significantly improved colonic morphology and tissue damage and inflammation in UC rats compared with MSCs and MSCs-vec. Further analysis showed that MSCs-KGF clearly promoted phosphorylation of PI3K and Akt and inhibited nuclear translocation of NF-κB in intestinal tissues of UC rats.Discussion: MSCs, especially KGF-modified MSCs, can improve colonic tissue damage in UC rats by promoting intestinal epithelial cell proliferation and reducing colonic inflammatory response, which may be related to activation of PI3K/Akt pathway and inhibition of NF-κB activation.Keywords: ulcerative colitis, bone marrow mesenchymal stem cells, keratinocyte growth factor, gene modification, PI3K/Akt pathway, NF-κB

Background Monochamus alternatus Hope is one of the insect vectors of pinewood nematode (Bursaphelenchus xylophilus), which causes the destructive pine wilt disease. The microorganisms within the ecosystem, comprising plants, their environment, and insect vectors, form complex networks. This study presents a systematic analysis of the bacterial microbiota in the M. alternatus midgut and its habitat niche. Methods Total DNA was extracted from 20 types of samples (with three replicates each) from M. alternatus and various tissues of healthy and infected P. massoniana (pines). 16S rDNA amplicon sequencing was conducted to determine the composition and diversity of the bacterial microbiota in each sample. Moreover, the relative abundances of bacteria in the midgut of M. alternatus larvae were verified by counting the colony-forming units. Results Pinewood nematode infection increased the microbial diversity in pines. Bradyrhizobium, Burkholderia, Dyella, Mycobacterium, and Mucilaginibacter were the dominant bacterial genera in the soil and infected pines. These results indicate that the bacterial community in infected pines may be associated with the soil microbiota. Interestingly, the abundance of the genus Gryllotalpicola was highest in the bark of infected pines. The genus Cellulomonas was not found in the midgut of M. alternatus, but it peaked in the phloem of infected pines, followed by the phloem of heathy pines. Moreover, the genus Serratia was not only present in the habitat niche, but it was also enriched in the M. alternatus midgut. The colony-forming unit assays showed that the relative abundance of Serratia sp. peaked in the midgut of instar II larvae (81%). Conclusions Overall, the results indicate that the bacterial microbiota in the soil and in infected pines are correlated. The Gryllotalpicola sp. and Cellulomonas sp. are potential microbial markers of pine wilt disease. Additionally, Serratia sp. could be an ideal agent for expressing insecticidal protein in the insect midgut by genetic engineering, which represents a new use of microbes to control M. alternatus.

B1-type human adenoviruses (HAdVs) HAdV-3, HAdV-7, and HAdV-55 have caused epidemics in North America, Asia, and Europe. However, to date, no adenovirus vaccines or antiviral drugs have been approved for general use. In the present work, a scFv-phage immune library was constructed and mouse monoclonal antibody (MMAb) 10G12 was obtained through selection. 10G12 is specific for HAdV-7 and binds the hexon loop1 and loop2 (LP12), resulting in strong neutralization activity against HAdV-7. Additionally, it is stable in serum and buffer at various pH values. The findings provide insight into adenovirus and antibody responses and may facilitate the design and development of adenovirus vaccines and antiviral drugs.

Tanshinol, a water-soluble component isolated from Salvia miltiorrhiza Bunge, has a variety of biological activities involving anti-fibrotic effect. However, the exact role and the underlying mechanisms remain largely unclear. This study mainly focused on the anti-hepatic fibrotic activities and mechanisms of tanshinol on carbon tetrachloride (CCl4)-induced liver fibrosis in rats via anti-oxidative and anti-inflammation pathways. The rats were divided into 4 groups as follows: control, model, tanshinol 20 mg/kg, and tanshinol 40 mg/kg. Except for the control group, CCl4 was used to induce liver fibrosis processing for 8 weeks, meanwhile rats in tanshinol groups were intraperitoneally injected with additional tanshinol. Control group simultaneously received the same volumes of olive oil and saline. The potentially protective effect and mechanisms of tanshinol on liver fibrosis in rats were evaluated. The serum levels of alanine aminotransferase, aspartate aminotransferase, and total bilirubin were obviously lower in the tanshinol treatment groups related to model group. Compared with the model group, the levels of hyaluronic acid, type IV collagen, Laminin (LN), and procollagen III peptide (PIIIP) in serum were significantly decreased after tanshinol treatment. Furthermore, tanshinol could regulate Nrf2/HO-1 signaling pathway and increase the level of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and also decrease the level of malondialdehyde (MDA) to against damage induced by oxidative stress. Simultaneously tanshinol could regulate nuclear factor kappa B signaling pathway to inhibit expression of inflammation factors, including transforming growth factor-β, tumor necrosis factor-α, Cox-2, interleukin-1β, and interleukin-6. In summary, our research demonstrated that tanshinol has protective effect on CCl4-induced liver fibrosis via inhibiting oxidative stress and inflammation, which may be associated with the regulation of nuclear factor erythroid2-related factor 2/hemeoxygenase-a and nuclear factor kappa B/inhibitor of kappa B alpha signaling pathways.

Human adenovirus serotype 7 (HAdV-7), belonging to species B, has caused severe lower respiratory tract diseases and even deaths recently. However, no adenovirus vaccine or therapeutic is available thus far. In this study, a HAdV-7-specific human monoclonal antibody (HMAb), 3-3E, isolated from single plasma cells obtained from the peripheral blood mononuclear cells of HAdV-7-infected patients showed potent HAdV-7 neutralization activity. The results showed HMAb 3-3E only binds to the hexon protein of intact HAdV-7 or the recombinant hexon protein and it does not bind to other intact virion particles. This could mean the antibody recognizes a conformational epitope of the hexon protein. Further, HMAb 3-3E potently neutralized HAdV-7 in vitro at low concentrations. In vivo studies showed HMAb 3-3E protected from HAdV-7 infection in a murine model. Therefore, HMAb 3-3E is promising as a safe and effective prophylactic and therapeutic treatment for HAdV-7 infection.

Anaerobic digesters have been widely used to treat wastewaters in livestock farms. With the increasing risk of antibiotic resistance originated from livestock husbandry, removal of antibiotics and antibiotic resistance genes (ARGs) via anaerobic digesters deserved more attention. Here we investigated the removal of antibiotics and ARGs in swine wastewater by three on-farm full-scale anaerobic digesters, including buried biogas digester (BBD), up-flow anaerobic sludge blanket (UASB) and high density polyethylene covered biogas digester (HDPE-BD). Variations of antibiotic resistome in swine wastewater were further revealed by metagenomic sequencing. Results showed the removal efficiencies for antibiotics, ARGs and mobile genetic elements (MGEs) varied in the three digesters, ranging from 65.1% to 98.1%, 3.5%-71.0% and 26.9%-77.2%, respectively. In general, UASB and HDPE-BD showed better removal efficiencies than BBD. However, enrichment of metal resistance genes (MRGs) was noted in UASB. Pathogens could not be effectively removed by all of the three digesters. What’s more, accumulation of pathogens was found in UASB (removal efficiencies: −8.5%-13.6%). Bacterial community succession, horizontal genetic transfer and biocide and metal resistance genes (BMRGs) profiles jointly structured the variation of antibiotic resistome during anaerobic digestion. A total of 334 high-quality bins were identified from swine wastewater, 96 of which belonged to phylum of Firmicutes, Bacteroidetes and Proteobacteria carried ARGs. Proteobacteria was the dominant multi-drug resistant flora. Meanwhile, ARG-carrying pathogens (Bacteroides and Mycolicibacter) were found in the swine wastewater, suggesting a potential threat to human and animal health. The findings from this study showed that HDPE-BD is the most eco-friendly and effective anaerobic digester in controlling risks from antibiotic resistance determinants in swine wastewater.

Higher and prolonged viral replication is critical for the increased pathogenesis of the highly pathogenic avian influenza (HPAI) subtype of H5N1 influenza A virus (IAV) over the lowly pathogenic H1N1 IAV strain. Recent studies highlighted the considerable roles of cellular miRNAs in host defence against viral infection. In this report, using a 3′UTR reporter system, we identified several putative miRNA target sites buried in the H5N1 virus genome. We found two miRNAs, miR‐584‐5p and miR‐1249, that matched with the PB2 binding sequence. Moreover, we showed that these miRNAs dramatically down‐regulated PB2 expression, and inhibited replication of H5N1 and H1N1 IAVs in A549 cells. Intriguingly, these miRNAs expression was differently regulated in A549 cells infected with the H5N1 and H1N1 viruses. Furthermore, transfection of miR‐1249 inhibitor enhanced the PB2 expression and promoted the replication of H5N1 and H1N1 IAVs. These results suggest that H5N1 virus may have evolved a mechanism to escape host‐mediated inhibition of viral replication through down‐regulation of cellular miRNAs, which target its viral genome.

Phenolic endocrine disrupting chemicals (EDCs) may pose a great hazard to wildlife and humans, owing to their ubiquitous presence in the environment and potential bioaccumulation ability. We investigated the bioaccumulation, metabolism, and human health risks of six phenolic EDCs, including bisphenol A (BPA), 4-tert-octylphenol (4-t-OP), 4-nonylphenol (4-NP), estrone (E1), 17β-estradiol (E2), and 17α-ethinylestradiol (EE2), in wild fish from the Pearl River system, South China. Except EE2, the other five EDCs were detected in at least one of the four fish tissues (bile, liver, plasma, and muscle). The concentrations of BPA and 4-NP were greater than those of 4-t-OP, E1, and E2 in all tissues. The median values of log bioaccumulation factors for EDCs at the range of 3.86-4.52 in bile, 2.06-3.16 in liver, 2.69-3.87 in plasma, and 1.34-2.30 in muscle, indicating a higher bioaccumulation potential in fish bile than in other tissues. Greater levels of glucuronide/sulfate conjugated EDCs were found in fish bile and liver than in the plasma and muscle, suggesting that the liver and bile played an important role in the metabolism and excretion of phenolic EDCs in fish. The calculated hazard quotient values were below 1 for each compound, implying low risk to human health by intake of edible fish.

Purpose To examine the cognition, spontaneous epilepsy, and electroencephalography (EEG) characteristics of rats with malformations of cortical development (MCD) and their use as an animal model for investigating the pathogenesis of intractable epilepsy and screening novel antiepileptic drugs. Methods An epileptic rat model of MCD was established with the F1 generation of pregnant rats after X-irradiation with 175 cGy (Group L), 195 cGy (Group M), or 215 cGy (Group H). Long-term video-EEG monitoring was used to record the seizures in the rats with MCD. Cognition was assessed with the Morris water maze. The EEGs were recorded and analyzed in the frontal and parietal lobes and hippocampi of adult rats. Finally, the brain tissues were processed for Nissl staining. Results The model groups exhibited markedly prolonged escape latencies and distinct decrements in the percent distance traveled in the target quadrant and platform-crossing frequency. These findings were dose-dependent. Frequent interictal epileptiform discharges were observed in the frontal and parietal lobes and hippocampi of adult rats, and their incidences were markedly higher in the model groups compared with that in the normal controls, with Group M having the highest incidence. Spontaneous seizures were observed in the model groups (mean incidence, 46.7%). The daily mean frequency of seizures and the incidence of spontaneous seizures were highest in Group M. Nissl staining revealed a dose-dependent pattern of hippocampal abnormalities, cortical and subcortical nodular heterotopia, and callosal agenesis in the model groups. Conclusion The 195 cGy dose was most appropriate for establishing an epileptic model of MCD with X-irradiation.

Human-induced temperature changes influence coastal regions, both via thermal pollution and ocean warming, which exerts profound effects on the chemistry of metals and the physiology of organisms. However, it remains unknown whether the increased temperature of discharged water or ocean warming, as a result of climate change, lead to an increase of human health risks associated with the consumption of sea foods. In this study, the influence of temperature on metal accumulation by oysters was studied in individuals collected from a coastal area affected by the thermal water discharge of the Houshi Power Plant, China. The bioaccumulation factor (BAF) and oral bioavailability (OBA) of metals in oysters was determined. Elevated temperatures led to an increase in BAF for Cu, Zn, Hg, and Cd (p 0.05), but no change was observed for As and Pb (p 0.05). The OBA for Cd, As, and Pb correlated positively to elevated temperatures (p 0.05). However, for Cu and Zn, OBA was negatively correlated with increasing temperature (p 0.05). As, Pb, and Cd in the trophically available metal (defined as a sum of heat-stable proteins, heat-denaturable proteins, and organelles) was significantly elevated at the highest temperature seawater site (site A) compared to the lowest seawater site (site B). Thus, the irregular variation of OBA for each metal may be the result of variations in the subcellular distribution of metals and the protein quality influenced by the increased temperature. Moreover, the increased temperature and increased the hazard quotient values of As and Cd (p 0.05 for As, n = 6, p 0.05 for Cd, n = 6), which provided an indication of the potential risks of the consumption of oysters or other seafood to future warming under climate change scenarios.

The casuarina moth, Lymantria xylina Swinhoe (Lepidoptera: Erebidae), is an important pest in the Australian pine tree, Casuarina equisetifolia, forest in the coastal area of South China. At the same time, as a closely related species of Lymantria dispar L. (Lepidoptera: Erebidae), it is also a potential quarantine pest. In the present study, specific primers were designed for identification of L. xylina based on the COI barcoding sequence between L. xylina and four other common forest pests. A 569-bp fragment was successfully amplified from 40 L. xylina from five geographical populations in four Chinese provinces. In addition, even through the analysis came from five highly diverse populations of L. xylina, the genetic distances ranged from 0.001 to 0.031. The neighbor-joining tree showed that the species from Hubei and Chongqing were clustered within a distinct group.

The pine aphid Cinara pinitabulaeformis Zhang et Zhang is the main pine pest in China, it causes pine needles to produce dense dew (honeydew) which can lead to sooty mold (black filamentous saprophytic ascomycetes). Although common chemical and physical strategies are used to prevent the disease caused by C. pinitabulaeformis Zhang et Zhang, new strategies based on biological and/or genetic approaches are promising to control and eradicate the disease. However, there is no information about genomics, proteomics or transcriptomics to allow the design of new control strategies for this pine aphid. We used next generation sequencing technology to sequence the transcriptome of C. pinitabulaeformis Zhang et Zhang and built a transcriptome database. We identified 80,259 unigenes assigned for Gene Ontology (GO) terms and information for a total of 11,609 classified unigenes was obtained in the Clusters of Orthologous Groups (COGs). A total of 10,806 annotated unigenes were analyzed to identify the represented biological pathways, among them 8,845 unigenes matched with 228 KEGG pathways. In addition, our data describe propagative viruses, nutrition-related genes, detoxification related molecules, olfactory related receptors, stressed-related protein, putative insecticide resistance genes and possible insecticide targets. Moreover, this study provides valuable information about putative insecticide resistance related genes and for the design of new genetic/biological based strategies to manage and control C. pinitabulaeformis Zhang et Zhang populations.

A sensitive and selective quick, easy, cheap, effective, rugged, and safe (QuEChERS) extraction combined with dispersive solid-phase extraction (d-SPE) cleanup method was developed to simultaneously extract a wide range of personal care products (16 biocides, 4 synthetic musks, and 4 benzotriazoles) in fish muscle and liver tissues. In order to get satisfactory recoveries, different extraction parameters were optimized, including extraction salts and d-SPE materials, extraction solvents and acetic acid contents in organic phase, and the ratios of solvent and water. Ultra pressure liquid chromatography-tandem mass spectrometry and gas chromatography-mass spectrometry were used to analyze the target compounds in the extracts. Among the 24 personal care products, the recoveries in the range of 70-120 % were obtained for 20, 19, and 12 analytes in fish muscle at the spiking concentrations of 10, 5, and 1 ng/g ww, respectively, and for 13, 12, and 11 analytes in liver at the spiking concentrations of 40, 20, and 4 ng/g ww, respectively. Method quantification limits (MQLs) of all analytes were 0.02-2.12 ng/g ww for fish muscle and 0.22-12.2 ng/g ww for fish liver tissues. The method was successfully applied to wild fish samples collected from Dongjiang River, south China. Twenty-one and 17 of the analytes were found in fish muscle and liver samples, respectively, in at least one site of the river with the concentrations between below MQLs and 119 ng/g ww, respectively. Graphical abstract Achieved satisfactory recoveries, high precision, and low method quantification limits (MQLs) for PCPs in wild fish tissues by QuEChERS procedure optimization combined with UPLC-MS/MS and GC-MS analyses.

Global environmental change presents a clear need for improved leading indicators of critical transitions, especially those that can be generated from compositional data and that work in empirical cases. Ecological theory of community dynamics under environmental forcing predicts an early replacement of slowly replicating and weakly competitive “canary” species by slowly replicating but strongly competitive “keystone” species. Further forcing leads to the eventual collapse of the keystone species as they are replaced by weakly competitive but fast‐replicating “weedy” species in a critical transition to a significantly different state. We identify a diagnostic signal of these changes in the coefficients of a correlation between compositional disorder and biodiversity. Compositional disorder measures unpredictability in the composition of a community, while biodiversity measures the amount of species in the community. In a stochastic simulation, sequential correlations over time switch from positive to negative as keystones prevail over canaries, and back to positive with domination of weedy species. The model finds support in empirical tests on multi‐decadal time series of fossil diatom and chironomid communities from lakes in China. The characteristic switch from positive to negative correlation coefficients occurs for both communities up to three decades preceding a critical transition to a sustained alternate state. This signal is robust to unequal time increments that beset the identification of early‐warning signals from other metrics.

Invading pathogens trigger specific host responses, an understanding of which might identify genes that function in pathogen recognition and elimination. In this study, we performed trans-species expression quantitative trait locus (ts-eQTL) analysis using genotypes of the Plasmodium yoelii malaria parasite and phenotypes of mouse gene expression. We significantly linked 1,054 host genes to parasite genetic loci (LOD score ≥ 3.0). Using LOD score patterns, which produced results that differed from direct expression-level clustering, we grouped host genes that function in related pathways, allowing functional prediction of unknown genes. As a proof of principle, 14 of 15 randomly selected genes predicted to function in type I interferon (IFN-I) responses were experimentally validated using overexpression, small hairpin RNA knockdown, viral infection, and/or infection of knockout mice. This study demonstrates an effective strategy for studying gene function, establishes a functional gene database, and identifies regulators in IFN-I pathways.

BACKGROUND: Alzheimer's disease (AD) is an age‐related chronic degenerative disease that damages the nervous system. A noninvasive and simple method for early detection of AD is extremely important for the diagnosis and prognosis of AD. Thus, we aimed to develop an enzyme‐linked immunosorbent assay (ELISA) kit to detect urine Alzheimer‐associated neuronal thread protein (AD7C‐NTP), and to evaluate its clinical value for the diagnosis of AD. METHODS: Immunogenic AD7C‐NTP peptide fragments were synthesized by the solid‐phase method and used for immunizing mice or rabbits to generate anti‐AD7C‐NTP antibodies. The urine AD7C‐NTP ELISA kit was then established; the generated mouse anti‐AD7C‐NTP antibody was used as a capture antibody, the biotin‐labeled rabbit anti‐AD7C‐NTP antibody was used as a detection antibody, and avidin labeled by horseradish peroxidase was used as a substrate. The first morning urine specimens of 121 AD patients and 118 age‐matched controls were collected, and the urine AD7C‐NTP levels were detected by the above ELISA kit. RESULTS: Mouse and rabbit anti‐AD7C‐NTP antibody ELISA titer was found to be 1:8,000 and 1:32,000, respectively. A single band with a relative molecular mass of 41 kDa was found in human brain specimens by Western blot assay, which was identified as AD7C‐NTP antibody. The urine AD7C‐NTP concentration of the AD patients was higher than that of the age‐matched controls, the sensitivity was 89.3% and the specificity was 84.7%. CONCLUSIONS: Our study demonstrated that our newly developed urine AD7C‐NTP ELISA kit has suggested potential for diagnosing AD in a Chinese population, suggesting it may be a useful diagnostic kit for detecting early AD.

In the present study, we investigated whether CVVH can reduce HMGB1, TLR4, NF-κB and other serum cytokine levels, preventing organ injury in a dog sepsis model. A total of 10 dogs were injected with LPS and treated with either CVVH group (n = 5) or nothing (Control, n = 5) for 24 h. EILSA was used for examining the concentration of TNF-α, IL-6, HMGB 1 and TLR4. The histological change of lung, liver and kidney tissues was determined. The mRNA expression of HMGB1, TLR4 and NF-κB was examined by RT-PCR. The protein of HMGB1 and phosphated NF-κB was examined by Western-blot. The levels of serum HMGB1 came to the peak at 8 h, 16 h and then declined. The LPS-induced increase in HMGB1 level was suppressed by CVVH compared with Control. Likewise, serum TNF-α and IL-6 levels decreased with CVVH along with a significant improvement in the function of main organs. Histologic examination revealed significant reduction in inflammation in lung; liver and kidney tissues harvested 24 h after CVVH compared with Control. The mRNA of HMGB1, TLR4 and NF-κB in the kidney was expressed at high level after LPS administration, which was significantly decreased by CVVH. The increased protein expression of HMGB1 and phosphated NF-κB was reduced after CVVH compared with control. CVVH by reducing the level of HMGB1, TLR4, NF-κB and other cytokines could weaken the cascade of cytokines and restore the immune system, and reduce the damage of important organs in sepsis.

We investigated the bioaccumulation of antibiotics in bile, plasma, liver and muscle tissues of wild fish from four rivers in the Pearl River Delta region. In total, 12 antibiotics were present in at least one type of fish tissues from nine wild fish species in the four rivers. The mean values of log bioaccumulation factors (log BAFs) for the detected antibiotics in fish bile, plasma, liver, and muscle tissues were at the range of 2.06–4.08, 1.85–3.47, 1.41–3.51, and 0.48–2.70, respectively. As the digestion tissues, fish bile, plasma, and liver showed strong bioaccumulation ability for some antibiotics, indicating a different bioaccumulation pattern from hydrophobic organic contaminants. Human health risk assessment based on potential fish consumption indicates that these antibiotics do not appear to pose an appreciable risk to human health. To the best of our knowledge, this is first report of bioaccumulation patterns of antibiotics in wild fish bile and plasma.

Background Arterial medial calcification (AMC) is frequent prevalence in patients with end stage renal disease. Evidence about hyperphosphatemia induced anabolic crosstalk between osteoblast and osteoclast in AMC of uremia is rare. Lanthanum carbonate as an orally administered phosphate-binding agent to reduce phosphate load and ameliorate AMC, but direct evidence is missing. Methods Detailed time-course studies were conducted of Sprague–Dawley rats fed with adenine and high phosphate diet to imitate the onset and progression of AMC of uremia. Calcification in great arteries was evaluated by VonKossa’s and Masson's trichrome staining. Osteoblast (Runx2, Osteocalcin) and osteoclast (RANKL, Cathepsin K, TRAP) related genes were analyzed by Immunohistochemistry and qRT-PCR. Serum PTH, RANKL and OPG levels were detected by ELISA kit. Results Serum phosphate was markedly increased in CRF group (6.94 ± 0.97 mmol/L) and 2%La group (5.12 ± 0.84 mmol/L) at week 4, while the latter group diminished significantly (2.92 ± 0.73 mmol/L vs CRF Group 3.48 ± 0.69, p

Although somatic cell reprogramming to generate inducible pluripotent stem cells (iPSCs) is associated with profound epigenetic changes, the roles and mechanisms of epigenetic factors in this process remain poorly understood. Here we identify Jmjd3 as a potent negative regulator of reprogramming. Jmjd3-deficient MEFs produced significantly more iPSC colonies than did wild-type cells, while ectopic expression of Jmjd3 markedly inhibited reprogramming. We show that the inhibitory effects of Jmjd3 are produced through both histone demethylase-dependent and -independent pathways. The latter pathway is entirely novel and involves Jmjd3 targeting of PHF20 for ubiquitination and degradation via recruitment of an E3 ligase, Trim26. Importantly, PHF20-deficient MEFs could not be converted to fully reprogrammed iPSCs, even with knockdown of Jmjd3, Ink4a or p21, indicating that this protein exerts predominant effects on reprogramming. Our findings demonstrate a previously unrecognized role of Jmjd3 in cellular reprogramming and provide molecular insight into the mechanisms by which the Jmjd3-PHF20 axis controls this process.

To investigate the role of liver-specific expression of glucokinase (GCK) in the pathogenesis of hyperglycemia and to identify candidate genes involved in mechanisms of the onset and progression of maturity onset diabetes of the young, type 2 (MODY-2), we examined changes in biochemical parameters and gene expression in GCK knockout (gck(w/-)) and wild-type (gck(w/w)) mice as they aged. Fasting blood glucose levels were found to be significantly higher in the gck(w/-) mice, compared to age-matched gck(w/w) mice, at all ages (P0.05), except at 2 weeks. GCK activity of gck(w/-) mice was about 50% of that of wild type (gck(w/w)) mice (P0.05). Glycogen content at 4 and 40 weeks of age was lower in gck(w/-) mice compared to gck(w/w) mice. Differentially expressed genes in the livers of 2 and 26 week-old liver-specific GCK knockout (gck(w/-)) mice were identified by suppression subtractive hybridization (SSH), which resulted in the identification of phosphoenolpyruvatecarboxykinase (PEPCK, also called PCK1) and Sterol O-acyltransferase 2 (SOAT2) as candidate genes involved in pathogenesis. The expressions of PEPCK and SOAT2 along with glycogen phosphorylase (GP) and glycogen synthase (GS) were then examined in GCK knockout (gck(w/-)) and wild-type (gck(w/w)) mice at different ages. Changes in PEPCK mRNA levels were confirmed by real-time RT-PCR, while no differences in the levels of expression of SOAT2 or GS were observed in age-matched GCK knockout (gck(w/-)) and wild-type (gck(w/w)) mice. GP mRNA levels were decreased in 40-week old gck(w/-) mice compared to age-matched gck(w/w) mice. Changes in gluconeogenesis, delayed development of GCK and impaired hepatic glycogen synthesis in the liver potentially lead to the onset and progression of MODY2.

High-flow through abnormal blood vessels underlies many life-threatening diseases. The ability to safely and non-invasively normalize these vessels by molecular intervention holds promise to treat these devastating conditions. Here we studied high-flow AV shunts caused by upregulation of Notch signaling via endothelial expression of constitutively-active Notch4 (Notch4*). Using 4D, two-photon imaging with cellular resolution in live mice, we found that normalizing Notch signaling by turning off Notch4* promptly converted large caliber, high-flow AV shunts to capillary-like vessels. The process was initiated by vessel narrowing without the loss of endothelial cells. Restoration of venous receptor EphB4 is an underlying mechanism, as EphB4 expression was recovered upon Notch normalization and required for the vessel regression. The structural regression of the high-flow AV shunts returned shunting flow to perfusing vessels, reversing tissue hypoxia and dysfunction. Our data provide direct, in vivo evidence that a single genetic manipulation in Notch pathway can exert dominant effects over hemodynamics leading to safe degeneration of the high-flow AV shunts at the core of AV Malformations.

Background During the process that AIV infect hosts, the NS1 protein can act on hosts, change corresponding signal pathways, promote the translation of virus proteins and result in virus replication. Results In our study, we found that PARP domain and Glu-rich region of PARP10 interacted with NS1, and the presence of NS1 could induce PARP10 migrate from cytoplasm to nucleus. NS1 high expression could reduce the endogenous PARP10 expression. Cell cycle analysis showed that with inhibited PARP10 expression, NS1 could induce cell arrest in G2-M stage, and the percentage of cells in G2-M stage rise from the previous 10%-45%, consistent with the cell proliferation result. Plague forming unit measurement showed that inhibited PARP10 expression could help virus replication. Conclusions In a word, our results showed that NS1 acts on host cells and PARP10 plays a regulating role in virus replication.

Stringent control of the NF-kappaB and type I interferon signaling pathways is critical to effective host immune responses, yet the molecular mechanisms that negatively regulate these pathways are poorly understood. Here, we show that NLRC5, a member of the highly conserved NOD-like protein family, can inhibit the IKK complex and RIG-I/MDA5 function. NLRC5 inhibited NF-kappaB-dependent responses by interacting with IKKalpha and IKKbeta and blocking their phosphorylation. It also interacted with RIG-I and MDA5, but not with MAVS, to inhibit RLR-mediated type I interferon responses. Consistent with these observations, NLRC5-specific siRNA knockdown not only enhanced the activation of NF-kappaB and its responsive genes, TNF-alpha and IL-6, but also promoted type I interferon signaling and antiviral immunity. Our findings identify NLRC5 as a negative regulator that blocks two central components of the NF-kappaB and type I interferon signaling pathways and suggest an important role for NLRC5 in homeostatic control of innate immunity.

Endogenous retroviral gene products have been found in some human tumors, and therefore, may serve as antigens for immunotherapy approaches. The murine colorectal carcinoma CT26 and melanoma B16 have recently been found to express the endogenous retroviral gene products gp70 and p15E, respectively, that can serve as antigens recognized by T cells. To date, though, there has been no demonstration of tumor treatment using an endogenous retroviral protein. In this study, we demonstrate that mice immunized with recombinant vaccinia encoding the gp70 H2-L(d)-restricted minimal determinant were protected from CT26 tumor challenge. Splenocytes from mice immunized with vaccinia gp70 specifically secreted IFN-gamma in response to gp70 peptide-pulsed stimulators. Although this strategy could protect against subsequent tumor challenge, it was ineffective against established tumors. Therefore, to investigate the treatment of established CT26 or B16 lung metastases, mice were treated with cultured dendritic cells (DCs) pulsed with gp70 or p15E peptide. Significant inhibition of established lung metastases required immunization with peptide-pulsed DCs pretreated with CD40 ligand that has been demonstrated to increase the T-cell stimulatory activity of DCs. The ability to immunize against endogenous retroviral tumor antigens may have relevance in the induction of antitumor immunity for some human cancers.