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Oriented ligand immobilization is one of the most effective strategies used in the design and construction of a high-capacity protein A chromatography. In this work, cysteine was introduced as anchoring sites by substituting a specific residue on Helix I, II, and at C-terminus of antibody binding domain Z from protein A, respectively, to investigate structural evolution and binding behavior of protein A ligands at liquid-solid interfaces. Among the three affinity dextran-coated Fe3O4 magnetic nanoparticles (Fe3O4@Dx MNPs), affinity MNPs with the immobilized ligand via N11C on Helix I (Fe3O4@Dx-Z1 MNPs) had the highest helical content, and MNPs with the immobilized ligand via G29C on Helix II (Fe3O4@Dx-Z2 MNPs) had the lowest helical content at the same pHs. It was attributed to less electrostatic attraction of ligand to negatively charged surface on Fe3O4@Dx-Z1 MNPs because of less positive charged residues on Helix I (K6) than Helix II (R27/K35). Among the three affinity MNPs, moreover, the highest affinity to immunoglobulin G (IgG) binding was observed on Fe3O4@Dx-Z1 MNPs in isothermal titration calorimetry measurement, further validating greater structural integrity of the ligand on Fe3O4@Dx-Z1 MNPs. Finally, the study of IgG binding on MNPs and 96-well plates showed that anchoring sites for ligand immobilization had distinct influences on IgG binding and IgG-mediated antigen binding. This work illustrated that anchoring sites of the ligands had a striking significance for the molecular structure of the ligand at liquid-solid interfaces and raised an important implication for the design and optimization of protein A chromatography and protein A-based immunoassay analysis.

Panax notoginseng saponins (PNS), the main bioactive constituents of a traditional Chinese herb Panax notoginseng, were commonly used for ischemic stroke in China. However, the associated cellular and molecular mechanisms of PNS have not been well examined. This study aimed to decipher the underlying molecular target of PNS in the treatment of cerebral ischemia. The oxygen-glucose-deprived (OGD) model of rat brain microvascular endothelial cells (BMECs) was used in this study. The alteration of gene expression in rat BMECs after PNS treatment was measured by microarray and indicated that there were 38 signaling pathways regulated by PNS. Among them, RIG-I receptor and related signaling molecules TNF receptor-associated factor 2 (Traf2) and nuclear factor-kappa B (NF-κB) were significantly suppressed by PNS, which was verified again in OGD-induced BMECs measured by FQ-PCR and western blotting and in middle cerebral artery occlusion (MCAO) rats measured by immunohistochemistry. The levels of TNF-α, IL-8, and the downstream cytokines regulated by RIG-I receptor pathway were also decreased by PNS. Meanwhile, the neurological evaluation, hematoxylin and eosin (HE) staining, and Evans blue staining were conducted to evaluate the effect of PNS in MCAO rats. Results showed PNS significantly improved functional outcome and cerebral vascular leakage. Flow cytometry showed the number of the inflammatory cells infiltrated in brain tissue was decreased in PNS treatment. Our results identified that RIG-I signaling pathway mediated anti-inflammatory properties of PNS in cerebral ischemia, which provided the novel insights of PNS application in clinics.

The formation of a stable spatial arrangement of protein A ligands is a great challenge for the development of high‐capacity polymer‐grafted protein A adsorbents due to the complexity in interplay between coupled ligands and polymer chain. In this work, carboxymethyl dextrans (CMDs) with different molecular weight were introduced to provide stable spatial ligand arrangement in CMD‐grafted protein A gels to improve IgG adsorption. The result showed that coupling of protein A ligand in CMD‐grafted layer had no marked influence on pore size and dextran layers coupled with the ligands were stable in experimental range of salt concentrations. The result of IgG adsorption revealed that carboxymethyl dextran T10, a short CMD, was more suitable as a scaffold for the synthesis of high‐capacity protein A gels. Moreover, the maximal adsorption capacity for IgG was obtained to be 96.4 mg/g gel at ionic capacities of 300–350 mmol/L and a ligand density of 15.2 mg/g gel. Dynamic binding capacity for IgG exhibited a higher capacity utilization in CMD‐grafted protein A gels than non‐grafted protein A gel. The research presented a tactics to establish a stable dextran layer coupled with protein A ligands and demonstrated its importance to improve binding capacity for IgG.

Background Corticotropin-releasing hormone (CRH) is an important neuromodulator that is widely distributed in the brain and plays a key role in mediating stress responses and autonomic functions. While the distribution pattern of fluorescently labeled CRH-expressing neurons has been studied in different transgenic mouse lines, a full appreciation of the broad diversity of this population and local neural connectivity can only come from integration of single-cell morphological information as a defining feature. However, the morphologies of single CRH neurons and the local circuits formed by these neurons have not been acquired at brain-wide and dendritic-scale levels. Results We screened the EYFP-expressing CRH-IRES-Cre;Ai32 mouse line to reveal the morphologies of individual CRH neurons throughout the whole mouse brain by using a fluorescence micro-optical sectioning tomography (fMOST) system. Diverse dendritic morphologies and projection fibers of CRH neurons were found in various brain regions. Follow-up reconstructions showed that hypothalamic CRH neurons had the smallest somatic volumes and simplest dendritic branches and that CRH neurons in several brain regions shared a common bipolar morphology. Further investigations of local CRH neurons in the medial prefrontal cortex unveiled somatic depth-dependent morphologies of CRH neurons that exhibited three types of mutual connections: basal dendrites (upper layer) with apical dendrites (layer 3); dendritic-somatic connections (in layer 2/3); and dendritic-dendritic connections (in layer 4). Moreover, hypothalamic CRH neurons were classified into two types according to their somatic locations and characteristics of dendritic varicosities. Rostral-projecting CRH neurons in the anterior parvicellular area had fewer and smaller dendritic varicosities, whereas CRH neurons in the periventricular area had more and larger varicosities that were present within dendrites projecting to the third ventricle. Arborization-dependent dendritic spines of CRH neurons were detected, among which the most sophisticated types were found in the amygdala and the simplest types were found in the hypothalamus. Conclusions By using the CRH-IRES-Cre;Ai32 mouse line and fMOST imaging, we obtained region-specific morphological distributions of CRH neurons at the dendrite level in the whole mouse brain. Taken together, our findings provide comprehensive brain-wide morphological information of stress-related CRH neurons and may facilitate further studies of the CRH neuronal system.

Protein A chromatography is a key technology in the industrial production of antibodies, and a variety of commercial protein A adsorbents are available in shelf. High stability and binding capacity of a protein A adsorbent are two key issues for successful practice of protein A chromatography. Earlier versions of protein A adsorbents ever exhibited serious fragility to typical cleaning-in-place protocols (e.g. washing with sodium hydroxide solution), and suffered from low binding capacity, harsh elution, ligand leakage and other problems involved in industrial applications. During the last three decades, various techniques and approaches have been applied in the improvement of chemical stability and enhancement of binding capacity of protein A-based ligands and adsorbents for antibody purifications. This mini-review focuses on the technical explorations in protein A-based affinity adsorbents, especially protein A-based ligands, including the efforts to increase the chemical stability by site-directed mutations and to improve the binding capacity by ligand polymerization and site-directed immobilization. Moreover, the efforts to develop short peptide ligands based on the structure of protein A, including the biomimetic design strategies and the synthesis of peptide-mixed mode hybrid ligands are discussed. These peptide and peptide-based hybrid ligands exhibit high affinity and selectivity to antibodies, but noteworthy differences in the binding mechanism of antibody from protein A. As a result, bound antibody to the ligands could be effectively eluted under mild conditions. Perspectives for the development of the protein A-based peptide ligands have been extensively discussed, suggesting that the ligands represent a direction for technological development of antibody purification.

STUDY QUESTION Is there an association between the human testis-specific gene, testis developmental related gene 1 (TDRG1) and human sperm motility? SUMMARY ANSWER TDRG1 is associated with asthenozoospermia and involved in regulating human sperm motility. WHAT IS KNOWN ALREADY Many testis-specific proteins potentially regulate spermatogenesis and sperm motility. We have identified a novel human testis-specific gene, TDRG1, which encodes a 100-amino-acid protein localized in the human sperm tail, yet little is known about its role in human spermatozoa. STUDY DESIGN, SIZE, DURATION Sperm samples were obtained from normozoospermic men and asthenozoospermic men who visited the reproductive medical center at Jiangxi Maternal and Child Health Hospital, Nanchang, Jiangxi, China between February 2018 and January 2019. In total, 27 normozoospermic men and 25 asthenozoospermic men were recruited to participate in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS The level of TDRG1 in sperm of normozoospermic and asthenozoospermic men was examined by immunoblotting and immunofluorescence assays. Progressive motility was examined by computer-aided sperm analysis. The correlation between the TDRG1 protein level and progressive motility was analyzed by linear regression. TDRG1 was imported into the sperm of normozoospermic and asthenozoospermic men using a cell-penetrating peptide (CPP)-fused TDRG1 recombinant protein (CPP-TDRG1), and the progressive motility was examined. Also, the altered proteins associated with TDRG1 in asthenozoospermic sperm were detected using label-free quantification method-based quantitative proteomic technology. TDRG1-interacting proteins were identified by co-immunoprecipitation coupled with tandem mass spectrometry analysis. MAIN RESULTS AND THE ROLE OF CHANCE The mean level of TDRG1 was significantly decreased in sperm of asthenozoospermic men compared with normozoospermic men (P LARGE SCALE DATA N/A. LIMITATIONS, REASONS FOR CAUTION The sample size was limited and larger cohorts are needed for verifying the positive effect of CPP-TDRG1 on human sperm motility. Furthermore, the caution should be paid that a comprehensive safety examination would be performed to evaluate whether CPP-TDRG1 is a possible treatment approach for asthenozoospermia. WIDER IMPLICATIONS OF THE FINDINGS Our results provide new insights into the mechanisms of sperm motility which may contribute to the diagnosis and treatment for asthenozoospermia. STUDY FUNDING/COMPETING INTEREST(S) National Natural Science Foundation of China (81501317 and 81871207 to H.C.; 81771644 to T.L.; 31671204 to X.Z.; 81571432 to Y.T.). The authors have no conflicts of interest to declare.

Carbaryl is a widely used carbamate pesticide in agriculture. The strain Rhizobium sp. X9 possesses the typical carbaryl degradation pathway in which carbaryl is mineralized via 1-naphthol, salicylate, and gentisate. In this study, we cloned a carbaryl hydrolase gene cehA and a novel two-component 1-naphthol hydroxylase gene cehC1C2. CehA mediates carbaryl hydrolysis to 1-naphthol and CehC1, an FMNH2 or FADH2-dependent monooxygenase belonging to the HpaB superfamily, and hydroxylates 1-naphthol in the presence of reduced nicotinamide-adenine dinucleotide (FMN)/flavin adenine dinucleotide (FAD), and the reductase CehC2. CehC1 has the highest amino acid similarity (58%) with the oxygenase component of a two-component 4-nitrophenol 2-monooxygenase, while CehC2 has the highest amino acid similarity (46%) with its reductase component. CehC1C2 could utilize both FAD and FMN as the cofactor during the hydroxylation, although higher catalytic activity was observed with FAD as the cofactor. The optimal molar ratio of CehC1 to CehC2 was 2:1. The Km and Kcat/Km values of CehC1 for 1-naphthol were 74.71 ± 16.07 μM and (8.29 ± 2.44) × 10-4 s-1·μM-1, respectively. Moreover, the enzyme activities and substrate spectrum between CehC1C2 and previously reported 1-naphthol hydroxylase McbC were compared. The results suggested that McbC had a higher 1-naphthol hydroxylation activity, while CehC1C2 had a broader substrate spectrum.

Degradation of the fungicide iprodione by the Paenarthrobacter sp. strain YJN-5 is initiated via hydrolysis of its N1 amide bond to form N-(3,5-dichlorophenyl)-2,4-dioxoimidazolidine. In this study, another iprodione-degrading strain, Paenarthrobacter sp. YJN-D, which harbours the same metabolic pathway as strain YJN-5 was isolated and characterized. The genes that encode the conserved iprodione catabolic pathway were identified based on comparative analysis of the genomes of the two iprodione-degrading Paenarthrobacter sp. and subsequent experimental validation. These genes include an amidase gene, ipaH (previously reported in AEM e01150-18); a deacetylase gene, ddaH, which is responsible for hydantoin ring cleavage of N-(3,5-dichlorophenyl)-2,4-dioxoimidazolidine, and a hydrolase gene, duaH, which is responsible for cleavage of the urea side chain of (3,5-dichlorophenylurea)acetic acid, thus yielding 3,5-dichloroaniline as the end product. These iprodione-catabolic genes are distributed on three plasmids in strain YJN-5 and are highly conserved between the two iprodione-degrading Paenarthrobacter strains. However, only the ipaH gene is flanked by a mobile genetic element. Two iprodione degradation cassettes bearing ipaH-ddaH-duaH were constructed and expressed in strains Pseudomonas putida KT2440 and Bacillus subtilis SCK6 respectively. Our findings enhance the current understanding of the microbial degradation mechanism of iprodione.

Three undescribed prenylated benzoic acid derivatives berberisides A-C (1-3) and a new clerodane glycoside berberiside D (4) were isolated from Berberis tsarica Aherndt. Their structures were elucidated on the basis of extensive NMR and HR-ESI-MS analysis. The in vitro cytotoxic activities of all isolates were studied against lung carcinoma A549, hepatocellular carcinoma HepG2 and breast carcinoma MDA-MB-231 cell lines. Among them, compounds 1 and 4 exhibited anti-proliferative effects against three tumor cell lines with IC50 ranging from 28.97 ± 2.18 to 35.83 ± 0.72 μM.

Most tumors of the digestive system, including esophageal, gastric, liver and colorectal cancer, are malignant tumors that are associated with rates of high morbidity and mortality. The lack of effective methods for early diagnosis is an important cause of poor prognosis for these malignancies. Circular RNAs (circRNAs) belong to a family of endogenous, covalently closed non‑coding RNAs that are characterized as having no 5' cap structures or 3' poly‑A tails. Shortly following discovery, circRNAs were considered to be a product of mis‑splicing and have no significant biological function. However, in recent years, accumulating evidence is demonstrating that they serve key roles in tumorigenesis and have the potential to serve as diagnostic markers. The present article summarizes the biogenesis and function of circRNAs and reviews their role in seven common types of tumor of the digestive system whilst exploring their potential as tumor markers and the significant roles they can serve in the digestive system, in addition to providing a referencing point for future studies of digestive system malignancies.

A Gram-stain-positive, aerobic, non-motile and coccoid-shaped bacterium, designated XNB-1T, was isolated from farmland soil in Taian, Shandong province, China. Strain XNB-1T contained iso-C15 : 0 and iso-C16 : 0 as the predominant fatty acids. The diagnostic diamino acid of the peptidoglycan was ornithine, and the interpeptide bridge was l-Orn←Gly(1, 2)←d-Glu. The polar lipid profile of strain XNB-1T consisted of diphosphatidylglycerol, phosphatidylglycerol, an unidentified phosphoglycolipid and three unidentified phospholipids. The predominant menaquinone of strain XNB-1T was MK-8(H4) and the DNA G+C content was 70.1 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain XNB-1T belonged to the genus Ornithinicoccus , and shared the highest similarity with Ornithinicoccus hortensis HKI 0125T (96.0 %), followed by Ornithinicoccus halotolerans EGI 80423T (95.5 %). Genome-based analysis of average nucleotide identity of strain XNB-1T with O. hortensis HKI 0125T and O. halotolerans EGI 80423T yielded values of 73.1 and 73.3 %, respectively, while the digital DNA–DNA hybridization values were 19.5 and 19.9 %, respectively. On the basis of phenotypic, chemotaxonomic and phylogenetic data, strain XNB-1T is considered to represent a novel species of the genus Ornithinicoccus , for which the name Ornithinicoccus soli sp. nov. is proposed. The type strain is XNB-1T (=CCTCC AB 2019099T=KCTC 49259T).

A Gram-stain-positive, strictly aerobic, non-motile, non-spore-forming and rod-shaped bacterium, designated as strain G-1T, was isolated from farmland soil sampled in in Fuyang, Anhui Province, PR China. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain G-1T was closely related to Cumulibacter manganitolerans 2-36T (97.7 % similarity). Strain G-1T contained iso-C16 : 0, C17 : 1ω6c, iso-C15 : 0 and iso-C14 : 0 as the predominant fatty acids. The polar lipids of strain G-1T were diphosphatidylglycerol, phosphatidylethanolamine, an unidentified phospholipid, an unidentified lipid and two unidentified glycolipids. The predominant respiratory quinone of strain G-1T was MK-9(H4). The cell wall contained meso-diaminopimelic acid as the diagnostic diamino acid. The G+C content of the genomic DNA based on genome calculations was 64.2 mol%. Average nucleotide identity and the digital DNA–DNA hybridization values for the draft genomes between strain G-1T and strain 2-36T were 75.7 and 20.2 %, respectively. On the basis of phenotypic and phylogenetic data, strain G-1T is considered to represent a novel species of the genus Cumulibacter , for which the name Cumulibacter soli sp. nov. is proposed. The type strain is G-1T (=CCTCC AB2019021T=KCTC 49258T).

Schizophrenia is a complex mental illness with genetic heterogeneity, which is often accompanied by alterations in brain structure and function. The neurobiological mechanism of schizophrenia associated with heredity remains unknown. Recently, the development of trans-scale and multi-omics methods that integrate gene and imaging information sheds new light on the nature of schizophrenia. In this article, we summarized the results of brain structural and functional changes related to the specific single-nucleotide polymorphisms (SNPs) in the past decade, and the SNPs were divided into non-coding regions and coding regions, respectively. It is hoped that the relationship between SNPs and cerebral alterations can be displayed more clearly and intuitively, so as to provide fresh approaches for the discovery of potential biomarkers and the development of clinical accurate individualized treatment decision-making.

Since the discovery of ketamine anti-depressant effects in last decade, it has effectively revitalized interest in investigating excitatory synapses hypothesis in the pathogenesis of depression. In the present study, we aimed to reveal the excitatory synaptic regulation of corticotropin-releasing hormone (CRH) neuron in the hypothalamus, which is the driving force in hypothalamic-pituitary-adrenal (HPA) axis regulation. This study constitutes the first observation of an increased density of PSD-93-CRH co-localized neurons in the hypothalamic paraventricular nucleus (PVN) of patients with major depression. PSD-93 overexpression in CRH neurons in the PVN induced depression-like behaviors in mice, accompanied by increased serum corticosterone level. PSD-93 knockdown relieved the depression-like phenotypes in a lipopolysaccharide (LPS)-induced depression model. Electrophysiological data showed that PSD-93 overexpression increased CRH neurons synaptic activity, while PSD-93 knockdown decreased CRH neurons synaptic activity. Furthermore, we found that LPS induced increased the release of glutamate from microglia to CRH neurons resulted in depression-like behaviors using fiber photometry recordings. Together, these results show that PSD-93 is involved in the pathogenesis of depression via increasing the synaptic activity of CRH neurons in the PVN, leading to the hyperactivity of the HPA axis that underlies depression-like behaviors.

Cotinine is a stable toxic contaminant, produced as a by-product of smoking. It is of emerging concern due to its global distribution in aquatic environments. Microorganisms have the potential to degrade cotinine; however, the genetic mechanisms of this process are unknown. Nocardioides sp. strain JQ2195 is a pure-culture strain that has been reported to degrade cotinine at micropollutant concentrations. This strain utilizes cotinine as its sole carbon and nitrogen source. In this study, a 50-kb gene cluster (designated cot), involved in cotinine degradation, was predicted based on genomic and transcriptomic analyses. A novel three-component cotinine hydroxylase gene (designated cotA1A2A3), which initiated cotinine catabolism, was identified and characterized. CotA from Shinella sp. strain HZN7 was heterologously expressed and purified and was shown to convert cotinine into 6-hydroxycotinine. H218O-labeling and electrospray ionization-mass spectrometry (ESI-MS) analysis confirmed that the hydroxyl group incorporated into 6-hydroxycotinine was derived from water. This study provides new molecular insights into the microbial metabolism of heterocyclic chemical pollutants. IMPORTANCE In the human body, cotinine is the major metabolite of nicotine, and 10 to 15% of generated cotinine is excreted in urine. Cotinine is a structural analogue of nicotine and is much more stable than nicotine. Increased tobacco consumption has led to high environmental concentrations of cotinine, which may have detrimental effects on aquatic ecosystems and human health. Nocardioides sp. strain JQ2195 is a unique cotinine-degrading bacterium. However, the underlying genetic and biochemical foundations of cotinine degradation are still unknown. In this study, a 50-kb gene cluster (designated cot) was identified by genomic and transcriptomic analyses as being involved in the degradation of cotinine. A novel three-component cotinine hydroxylase gene (designated cotA1A2A3) catalyzed cotinine to 6-hydroxy-cotinine. This study provides new molecular insights into the microbial degradation and enzymatic transformation of cotinine.

Alzheimer’s disease (AD) diagnoses are greatly increasing in frequency as the global population ages, highlighting an urgent need for new anti-AD strategies. With the aim to search for human acetylcholinesterase (hAChE) inhibitors from the species of Myrtaceae family, ten acylphloroglucinol trimers (APTs), including eight new APTs, callistemontrimers A–H (1a, 1b, 2a, 2b, 3a, 3b, 4b, and 5b), and two naturally occurring ones (4a and 5a), along with one reported triketone–acylphloroglucinol–monoterpene adduct (6), were obtained and structurally characterized from the hAChE inhibitory acetone extract of Callistemon salignus seeds. The structures and their absolute configurations for new APTs were unequivocally established via the detailed interpretation of extensive spectroscopic data (HRESIMS and NMR), ECD calculations, and single crystal X-ray diffraction, whereas the absolute configurations of known APTs were determined by further chiral separation, and calculated ECD calculations. The results of hAChE inhibitory assay revealed that an enantiomeric mixture of 2a/2b, 2a, and 2b are good hAChE inhibitors with IC50 values of 1.22 ± 0.23, 2.28 ± 0.19, and 4.96 ± 0.39 μM, respectively. Molecular docking was used to uncover the modes of interactions for bioactive compounds with the active site of hAChE. In addition, 2 and 6 displayed moderate neurite outgrowth-promoting effects with differentiation rates of 6.16% and 6.19% at a concentration of 1.0 μM, respectively.

BACKGROUND In 2017 Tuta absoluta was identified as an invasive species in China. Due to its rapid geographic expansion and the severe crop damage it causes, T. absoluta poses a serious threat to China's tomato production industry. To determine its geographic distribution and host range, intensive surveys and routine monitoring were conducted across the Chinese mainland between 2018 and 2019. The population colonization coefficient (PCC; ratio of colonized sites and prefectures) and population occurrence index (POI; ratio of infested host species and PCCs) were calculated. RESULTS In northwestern China, T. absoluta populations established in Xinjiang exhibited a medium PCC value (~0.03). In southwestern China, populations in Yunnan and its five neighboring provinces exhibited high (~0.50 in Yunnan and Guizhou), or low (

Salicylate is a typical aromatic compound widely distributed in nature. Microbial degradation of salicylate has been well studied and salicylate hydroxylases play essential roles in linking the peripheral and ring-cleavage catabolic pathways. The direct hydroxylation of salicylate catalyzed by salicylate-1-hydroxylase or salicylate-5-hydroxylase has been well studied. However, the CoA mediated salicylate 5-hydroxylation pathway has not been characterized in detail. Here, we elucidate the molecular mechanism of the reaction in the conversion of salicylate to gentisate in the carbaryl-degrading strain Rhizobium sp. X9. Three enzymes (salicylyl-CoA ligase CehG, salicylyl-CoA hydroxylase CehH and gentisyl-CoA thioesterase CehI) catalyzed the conversion of salicylate to gentisate via a route, including CoA thioester formation, hydroxylation and thioester hydrolysis. Further analysis indicated that genes cehGHI are also distributed in other bacteria from terrestrial environment and marine sediments. These genomic evidences highlight the role of this salicylate degradation pathway in the carbon cycle of soil organic compounds and marine sediments. Our findings of this three-step strategy enhanced the current understanding of CoA mediated degradation of salicylate.

Strobilurin fungicides are widely used in agricultural production due to their broad-spectrum and fungal mitochondrial inhibitory activities. However, their massive application has restrained the growth of eukaryotic algae and increased collateral damage in freshwater systems, notably harmful cyanobacterial blooms (HCBs). In this study, a strobilurin fungicide-degrading strain, Hyphomicrobium sp. strain DY-1, was isolated and characterized successfully. Moreover, a novel esterase gene, strH, responsible for the de-esterification of strobilurin fungicides, was cloned, and the enzymatic properties of StrH were studied. For trifloxystrobin, StrH displayed maximum activity at 50°C and pH 7.0. The catalytic efficiencies (k(cat)/K(m)) of StrH for different strobilurin fungicides were 196.32 ± 2.30 μM(−1) · s(−1) (trifloxystrobin), 4.64 ± 0.05 μM(−1) · s(−1) (picoxystrobin), 2.94 ± 0.02 μM(−1) · s(−1) (pyraclostrobin), and (2.41 ± 0.19)×10(−2 )μM(−1) · s(−1) (azoxystrobin). StrH catalyzed the de-esterification of a variety of strobilurin fungicides, generating the corresponding parent acid to achieve the detoxification of strobilurin fungicides and relieve strobilurin fungicide growth inhibition of Chlorella. This research will provide insight into the microbial remediation of strobilurin fungicide-contaminated environments. IMPORTANCE Strobilurin fungicides have been widely acknowledged as an essential group of pesticides worldwide. So far, their residues and toxic effects on aquatic organisms have been reported in different parts of the world. Microbial degradation can eliminate xenobiotics from the environment. Therefore, the degradation of strobilurin fungicides by microorganisms has also been reported. However, little is known about the involvement of enzymes or genes in strobilurin fungicide degradation. In this study, a novel esterase gene responsible for the detoxification of strobilurin fungicides, strH, was cloned in the newly isolated strain Hyphomicrobium sp. DY-1. This degradation process detoxifies the strobilurin fungicides and relieves their growth inhibition of Chlorella.

Ligand immobilization is indeed key fundamental for the development of high-capacity protein A chromatographic material. In this work, mono- and tetra-mers of Z domain from protein A were immobilized onto magnetic nanoparticle (NP) coated with carboxymethyl dextran (CMD), Fe3O4@CMD NP, via oriented coupling. Spectral results of Fe3O4@CMD-oZ NP with oriented Z domain showed that the molecular structure of the ligand was dominated by ligand – surface interaction. At pH 4.5, the ligand was disrupted by electrostatic attraction between NP surface and ligand. As buffer pH was higher than pI of the ligand, the ligand tended to be native structure with an increase of electrostatic repulsion between NP surface and ligands. At pH 10.0, therefore, the ligand remained intact on Fe3O4@CMD-oZ NP. Furthermore, ligand polymerization promoted the structural stability of the tetrameric ligand by decreasing interaction between NP surface and three domains at amino terminus of the ligand. It was more significant at pH 7.0 and the highest stoichiometric number of human IgG binding was obtained. Moreover, oriented immobilization was more beneficial for structural stability of the tetrameric ligand than random immobilization. Likely, spectral and calorimetric evidences at different ligand densities further demonstrated that the crowding effect induced at high ligand density maintained molecular structure of the tetrameric ligand better.

In this study, we isolated and characterized the bacterial strain Pseudomonas sp. BYT-1, which is capable of degrading pymetrozine and using it as the sole carbon source for growth. Strain BYT-1 could degrade 2.30 mM pymetrozine within 20 h under the optimal conditions of 30 °C and pH 7.0. Investigation of the degradation pathway showed that pymetrozine was oxidatively hydrolyzed to 4-amino-6-methyl-4,5-dihydro-2 H-[1,2,4]triazin-3-one (AMDT) and nicotinic acid (NA). The former accumulates as the end product in the culture, whereas the latter was hydroxylated to 6-hydroxynicotinic acid (6HNA) and subjected to further degradation. The transformation of pymetrozine to AMDT and NA by the cell-free extracts of strain BYT-1 also supported that the oxidative hydrolysis of the C═N double bond in pymetrozine was the initial degradation step. This is the first report on a pure bacterial culture with the ability to degrade pymetrozine. These findings enhance our understanding of the microbial degradation mechanism of pymetrozine.

In this work, a range of investigation were conducted to illustrate the mechanism of hIgG –protein A from Staphylococcal aureus (SpA) binding for the development of high-capacity SpA gels. Results of surface plasmon resonance demonstrated that oriented immobilization at carboxyl terminus of Z domain was more preferred than that at amino terminus and bound hIgG with the oriented domain had a flat-on orientation over chip surface. Furthermore, much negative enthalpy changes as well as lower binding stoichiometry were observed in the binding of tetrameric Z domains (denoted as Z4cys) to hIgG, indicating that each of both ligands could bind 2.1 hIgG molecules. Adsorption equilibria of hIgG adsorption showed that by far the maximum adsorption capacity for hIgG in Sepharose-based protein A gels was determined to be about 120.0 mg/g gel. Its excellent performance was further manifested by the result of dynamic binding capacity. In protein A gel, moreover, there was a critical density around 20 mg/g gel, above which the availability of the ligand decreased rapidly due to serious steric exclusion effect. The research provided insight into the IgG – SpA binding and strategic guidance for the development of high-capacity protein A gels.

Pymetrozine is a synthetic pesticide that can be utilized as the sole carbon source by Pseudomonas sp. strain BYT-1. However, the genes involved in the degradation of pymetrozine remain unknown. We used transposon mutagenesis to create a mutant that unable to hydrolyze pymetrozine. The transposon interrupted the gene pyzH, which was cloned by self-formed adaptor PCR. PyzH hydrolyzed the C=N double bond of pymetrozine to produce 4-amino-6-methyl-4,5-dihydro-2H-[1,2,4]triazin-3-one (AMDT) and nicotinaldehyde; the latter inhibits PyzH activity. PyzH can completely hydrolyze pymetrozine in the presence of dehydrogenase ORF6, which can convert nicotinaldehyde into nicotinic acid and relieve the inhibition. H2 18 O-labeling experiments showed that the oxygen atom of nicotinaldehyde came from water instead of oxygen. PyzH homologous genes were also found in other soil isolates able to degrade pymetrozine.

Methomyl {bis[1-methylthioacetaldehyde-O-(N-methylcarbamoyl)oximino]sulfide} is a highly toxic oxime carbamate insecticide. Several methomyl-degrading microorganisms have been reported so far, but the role of specific enzymes and genes in this process is still unexplored. In this study, a protein annotated as a carbamate C-N hydrolase was identified in the methomyl-degrading strain Aminobacter aminovorans MDW-2, and the encoding gene was termed ameH. A comparative analysis between the mass fingerprints of AmeH and deduced proteins of the strain MDW-2 genome revealed AmeH to be a key enzyme of the detoxification step of methomyl degradation. The results also demonstrated that AmeH was a functional homodimer with a subunit molecular mass of approximately 34 kDa and shared the highest identity (27%) with the putative formamidase from Schizosaccharomyces pombe ATCC 24843. AmeH displayed maximal enzymatic activity at 50°C and pH 8.5. K(m) and k(cat) of AmeH for methomyl were 87.5 μM and 345.2 s(−1), respectively, and catalytic efficiency (k(cat)/K(m)) was 3.9 μM(−1) s(−1). Phylogenetic analysis revealed AmeH to be a member of the FmdA_AmdA superfamily. Additionally, five key amino acid residues (162, 164, 191, 193, and 207) of AmeH were identified by amino acid variations. IMPORTANCE Based on the structural characteristic, carbamate insecticides can be classified into oxime carbamates (methomyl, aldicarb, oxamyl, etc.) and N-methyl carbamates (carbaryl, carbofuran, isoprocarb, etc.). So far, research on the degradation of carbamate pesticides has mainly focused on the detoxification step and hydrolysis of their carbamate bond. Several genes, such as cehA, mcbA, cahA, and mcd, and their encoding enzymes have also been reported to be involved in the detoxification step. However, none of these enzymes can hydrolyze methomyl. In this study, a carbamate C-N hydrolase gene, ameH, responsible for the detoxification step of methomyl in strain MDW-2 was cloned and the key amino acid sites of AmeH were investigated. These findings provide insight into the microbial degradation mechanism of methomyl.

Hepatocellular carcinoma (HCC) is a malignant tumor and is associated with necroinflammation driven by various immune cells, such as dendritic cells, macrophages and natural killer cells. Innate immune cells can directly affect HCC or regulate the T-cell responses that mediate HCC. In addition, innate immune cells and T cells are not isolated, which means the interaction between them is important in the HCC microenvironment. Considering the current unsatisfactory efficacy of immunotherapy in patients with HCC, understanding the relationship between innate immune cells and T cells is necessary. In the present review the roles and clinical value of innate immune cells that have been widely reported to be involved in HCC, including dendritic cells, macrophages (including kupffer cells), neutrophils, eosinophils, basophils and innate lymphoid cells and the crosstalk between the innate and adaptive immune responses in the antitumor process have been discussed. The present review will facilitate researchers in understanding the importance of innate immune cells in HCC and lead to innovative immunotherapy approaches for the treatment of HCC.

Heterotrophic nitrification bacteria play a critical role in nitrogen cycling and pollution removal. However, the underlying nitrification mechanisms are diverse and have rarely been investigated at the genetic level. In this study, the new heterotrophic nitrifier Pseudomonas sp. strain JQ170 was isolated. Strain JQ170 can utilize ammonia (NH4+-N), nitrite (NO2−-N), or nitrate (NO3−-N) as sole nitrogen sources, preferring NH4+-N. A ratio of 96.4% of 1.0 mM NH4+-N was removed in 24 h. The optimum pH, temperature, and carbon source for NH4+-N removal were pH 7.0, 30 °C, and citrate, at a C/N ratio of 9–18, respectively. During the NH4+-N removal process, only NO2−-N but neither hydroxylamine, NO3−-N, nor gaseous nitrogen were detected. A random transposon insertion mutagenesis library of strain JQ170 was constructed. Two NO2−-N-production deficient mutants were screened and transposon insertion sites were located in nap genes (which encode periplasmic NO3−-N reductase Nap). Further gene knockout and complementation of the napA gene confirmed nap as essential for NO2−-N production. The following nitrification processes in strain JQ170 is proposed: NH4+-N to NO3−-N in the cytoplasm; then NO3−-N to NO2−-N in the periplasmic space by Nap; finally, NO2−-N secreted out of cells. Overall, this paper provides new insight towards understanding heterotrophic nitrification at the genetic level.

The residues of bensulfuron-methyl (BSM), a sulfonylurea herbicide, in soil have caused serious damage to the rotation of susceptible crops. Many studies have reported that the removal of BSM in soil was achieved by adding degrading bacteria. However, the mechanisms used by bacteria to degrade BSM in the crop rhizosphere remain unclear. In this study, a BSM-degrading bacterium, Hansschlegelia zhihuaiae S113, was applied to investigate the enhancement of effects mediated by organic acids during the bioremediation of BSM-contaminated maize rhizosphere soil. Organic acids, such as L -malic acid, tartaric acid, and fumaric acid, identified in maize root exudates, significantly stimulated the expression of cheA, which encoded the histidine kinase in strain S113 and contributed to the chemotactic response. This process accelerated the accumulation of strain S113 around the maize roots and promoted the colonization process on maize roots. The growth of strain S113 was significantly increased by L -malic acid but not tartaric acid or fumaric acid. After the S113 suspension was root-irrigated to BSM-contaminated soil, the density of strain S113 colonizing root surfaces and in rhizosphere soil reached 1.1 × 104 cells/g for roots and 4.9 × 104 cells/g in dry soil at 15 d, leading to 80.9% BSM degradation efficiency. The treatment with the addition of a mixture of S113 and L -malic acid completely degraded BSM in rhizosphere soil due to the strong attraction and growth promotion of strain S113 by L -malic acid, with a higher efficiency than that with the extra addition of fumaric acid (89.7%) or tartaric acid (87.0%). This paper revealed the enhancement effects of organic acids identified in root exudates for the in situ bioremediation of BSM-contaminated rhizosphere soil.

The ventral part of the anteromedial thalamic nucleus (AMv) is in a position to convey information to the cortico-hippocampal-amygdalar circuit involved in the processing of fear memory. Corticotropin-releasing-factor (CRF) neurons are closely associated with the regulation of stress and fear. However, few studies have focused on the role of thalamic CRF neurons in fear memory. In the present study, using a conditioned fear paradigm in CRF transgenic mice, we found that the c-Fos protein in the AMv CRF neurons was significantly increased after cued fear expression. Chemogenetic activation of AMv CRF neurons enhanced cued fear expression, whereas inhibition had the opposite effect on the cued fear response. Moreover, chemogenetic manipulation of AMv CRF neurons did not affect fear acquisition or contextual fear expression. In addition, anterograde tracing of projections revealed that AMv CRF neurons project to wide areas of the cerebral cortex and the limbic system. These results uncover a critical role of AMv CRF neurons in the regulation of conditioned fear memory. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12264-020-00592-6) contains supplementary material, which is available to authorized users.

IL-33 is a member of the IL-1 family of cytokines whose role remains controversial in rheumatoid arthritis (RA). The present study was performed to evaluate the correlation of IL-33 with other cytokines and chemokines in serum and the synovia, and to explore the nature of the association. The concentration of IL-33 in samples from 96 patients with RA was analyzed. The response of fibroblast-like synoviocytes (FLSs) to treatment with different concentrations of IL-33 was assessed in vitro. IL-33 was indicated to exhibit an association with multiple cytokines and chemokines in synovial fluid with an inverted-U-shaped trend, including IL-6, IL-1β, IL-8, MIG and IP-10, but not in the serum. Furthermore, in vitro experiments confirmed that IL-33 also exerted a U-type dose-dependent regulatory effect on FLS function. In addition, the data-points do not exactly follow the U-shaped curve fit in most cases, therefore, the applicability of this mathematical model in clinical practice is limited.

Due to their fast growth rate and robustness, some haloalkalitolerant methanotrophs from the genus Methylotuvimicrobium have recently become not only promising biocatalysts for methane conversion but also favorable materials for obtaining fundamental knowledge on methanotrophs. Here, to realize unmarked genome modification in Methylotuvimicrobium bacteria, a counterselectable marker (CSM) was developed based on pheS, which encodes the α-subunit of phenylalanyl-tRNA synthetase. Two-point mutations (T252A and A306G) were introduced into PheS in Methylotuvimicrobium buryatense 5GB1C, generating PheS AG , which can recognize p-chloro-phenylalanine (p-Cl-Phe) as a substrate. Theoretically, the expression of PheS AG in a cell will result in the incorporation of p-Cl-Phe into proteins, leading to cell death. The P tac promoter and the ribosome-binding site region of mmoX were employed to control pheS AG , producing the pheS AG -3 CSM. M. buryatense 5GB1C harboring pheS AG -3 was extremely sensitive to 0.5 mM p-Cl-Phe. Then, a positive and counterselection cassette, PZ (only 1.5 kb in length), was constructed by combining pheS AG -3 and the zeocin resistance gene. A PZ- and PCR-based strategy was used to create the unmarked deletion of glgA1 or the whole smmo operon in M. buryatense 5GB1C and Methylotuvimicrobium alcaliphilum 20Z. The positive rates were over 92%, and the process could be accomplished in as few as eight days.

Zanthoxylum armatum, its peels possessed better special flavour, as well as various bioactivities, such as anti-inflammatory, anti-microbial and anti-tumour. In our chemical investigation on the peels of Z. armatum, two new lignans (1 and 2) and three known lignans (3–5) were isolated by silica gel column chromatography, ODS column and preparative HPLC and their structures were established as zanthlignans A and B (1–2), (-)-asarinin (3), phylligenin (4) and planispine A (5) through various spectroscopic techniques including UV, IR, HR-ESI-MS, NMR and CD methods.

Phthalate esters (PAEs), a class of toxic anthropogenic compounds, have been predominantly used as additives or plasticizers, and great concern and interests have been raised regarding its environmental behavior and degradation mechanism. In the present study, a bacterial consortium consisting of Microbacterium sp. PAE-1 and Pandoraea sp. PAE-2 was isolated by the enrichment method, which could degrade dibutyl phthalate (DBP) completely by biochemical cooperation. DBP was converted to phthalic acid (PA) via monobutyl phthalate (MBP) by two sequential hydrolysis steps in strain PAE-1, and then PA was further degraded by strain PAE-2. Strain PAE-1 could hydrolyze many dialkyl Phthalate esters (PAEs) including dimethyl, diethyl, dibutyl, dipentyl, benzyl butyl, dihexyl, di-(2-ethyhexyl) and their corresponding monoalkyl PAEs. Two esterase genes named dpeH and mpeH, located in the same transcription unit, were cloned from strain PAE-1 by the shotgun method and heterologously expressed in Escherichia. coli (DE3). The Km and kcat values of DpeH for DBP were 9.60 ± 0.97 μM and (2.72 ± 0.06) × 106 s−1, while those of MpeH for MBP were 18.61 ± 2.00 μM and (5.83 ± 1.00) × 105 s−1, respectively. DpeH could only hydrolyze dialkyl PAEs to the corresponding monoalkyl PAEs, which were then hydrolyzed to PA by MpeH. DpeH shares the highest similarity (53%) with an alpha/beta hydrolase from Microbacterium sp. MED-G48 and MpeH shows only 25% identity with a secreted lipase from Trichophyton benhamiae CBS 112371, indicating that DpeH and MpeH are two novel hydrolases against PAEs.

Perioperative neurocognitive disorders (PND) are characterized by deficits in cognitive functions in the elderly following anesthesia and surgery. Effective clinical interventions for preventing this disease are limited. Growing evidence demonstrates that activation of NOD-like receptor protein3 (NLRP3) inflammasome is involved in neurodegenerative diseases. We therefore hypothesized that activation of NLRP3 inflammasome is linked to neuroinflammation and the subsequent cognitive impairments that occurred in an animal model of PND. In this study, 18-month-old C57BL/6 mice were subjected to an exploratory laparotomy under isoflurane anesthesia to mimic clinical human abdominal surgery. For interventional studies, mice received NLRP3 specific inhibitor MCC950 (10 mg/kg) or the vehicle only intraperitoneally. Behavioral studies were performed at 6 and 7 d after surgery using open field and fear conditioning tests, respectively. Interleukin-1β (IL-1β), interleukin-18 (IL-18), tumor necrosis factor-α (TNF-α), ionized calcium-binding adaptor molecule-1 (IBA1) positive cells, glial fibrillary acidic protein (GFAP) positive cells, NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), and cleaved caspase-1 were measured at 3 days post-surgery. Brain-derived neurotrophic factor (BDNF) and postsynaptic density protein 95 (PSD95) were measured at 7 days post-surgery. Our data indicates that surgery-induced cognitive impairments were associated with significant increases in IL-1β, IL-18, TNF-α, NLRP3, ASC, cleaved caspase-1, IBA1-positive cells and GFAP-positive cells, and decreases in BDNF and PSD95 expression in the hippocampus. Notably, administration with MCC950 attenuated inflammatory changes and rescued surgery-induced cognitive impairments. Our study suggests that surgery induces neuroinflammation and cognitive deficits that are partly attributed to the activation of NLRP3 inflammasome in the hippocampus of aged mice.

A Gram-staining-negative, aerobic, non-motile and rod-shaped bacterium that produced yellow viscous colonies, designated FL-8T, was isolated from farmland soil in Chuzhou, Anhui province, PR China. 16S rRNA gene sequence similarities between strain FL-8T and the type strains of species of the genus Terrimonas with validly published names ranged from 94.6 to 96.1 %. Strain FL-8T contained iso-C15 : 1 G, iso-C15 : 0 and iso-C17 : 0 3-OH as the predominant fatty acids. The predominant polar lipid of strain FL-8T was phosphatidylethanolamine. The sole respiratory quinone of strain FL-8T was MK-7 and the DNA G+C content was 44.8 mol%. On the basis of phenotypic, chemotaxonomic and phylogenetic data, strain FL-8T is considered to represent a novel species of the genus Terrimonas , for which the name Terrimonas soli sp. nov. is proposed. The type strain is FL-8T (=CCTCC AB 2017059T=JCM 32095T).

A Gram-stain-negative, aerobic, non-motile, non-spore-forming and rod-shaped bacterium, designated YHM-9T, was isolated from soil in Yangquan, Shanxi Province, PR China. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YHM-9T belonged to the genus Pedobacter and shared the highest similarity (97.4 %) to the type strain Pedobacter lignilitoris W-WS13T. Strain YHM-9T exhibited low DNA–DNA relatedness with P. lignilitoris W-WS13T (21.7±1.3 %). The DNA G+C content was 38.9 mol%. The major fatty acids were iso-C15 : 0, summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and iso-C17 : 0 3-OH. The respiratory quinone was MK-7, the major polyamine was sym-homospermidine and the major polar lipids were phosphatidylethanolamine. Based on the morphological, physiological, biochemical and chemotaxonomic characteristics, strain YHM-9T was recognized as a representative of a novel species within the genus Pedobacter , for which the name Pedobacter agrisoli sp. nov. is proposed. The type strain is YHM-9T (=JCM 32093T=CCTCC AB 2017125T).

Dipicolinic acid (DPA) is a widely distributed pyridine derivative containing two carboxylate groups. However, the catabolism of DPA mediated by microorganisms are rarely reported. Alcaligenes faecalis strain JQ135, previously isolated in our lab, was able to degrade various pyridine derivatives, including nicotinic acid (NA). In this study, it was found that strain JQ135 could utilize DPA as the sole carbon and nitrogen source for growth and degrade 1.8 mM (300 mg/L) DPA to a non-detectable level within 12 h. The catabolism of DPA by strain JQ135 extracted two intermediates, i.e., 3-hydroxy-2,6-dipicolinic acid and 4-imino-pent-2-enedioic acid. Moreover, deletion of the key genes essential for NA catabolism (such as moaCDEBmoeA and maiA) did not affect DPA catabolism, indicating that the maleamate pathway and molybdenum cofactor are not involved in the DPA catabolism by strain JQ135. This study will provide insights into the bacterial catabolism of DPA.

Four new cytochalasans, phychaetoglobins A–D (1–4), together with twelve known cytochalasans (5–16), were isolated from a mangrove-associated fungus Chaetomium globosum kz-19. The new structures were elucidated on the basis of extensive 1D and 2D NMR, HR ESIMS spectroscopic analyses, and electronic circular dichroism (ECD) calculations. The absolute configuration of 2 was established by application of Mosher’s method. Compounds 4–8 exhibited moderate cytotoxicities against A549 and HeLa cell lines with the IC50 values less than 20 μM.

As a widely used fungicide, the environmental fate of carbendazim and its residues in agricultural products have caused great concern. However, its effects on soil microbial communities are largely unknown. Herein, we used high-throughput sequencing to reveal the effects of high and low dose of carbendazim and its degrading strain, Rhodococcus qingshengii strain djl-6, on the composition, diversity, and interrelationship of soil bacterial and fungal communities in short- and medium-term under laboratory conditions. The results showed that carbendazim exhibited an increased negative impact on bacterial communities and reduced the proportion of dominant fungal phylum Ascomycota during a 14-day incubation period. Only the impacts of low-dose carbendazim (2 mg·kg−1 dry soil) on fungal community were weakened. Network analysis showed that carbendazim increased the connectivity and modularity of microbial co-occurrence networks. Strain djl-6 exhibited good potential for bioremediation of carbendazim-contaminated soils. Moreover, it driven the assembly of potential carbendazim-degrading consortia from indigenous microbial communities; and members of the genera Arthrobacter, Bacillus, Brevundimonas, Lysinibacillus, Massilia, Mycobacterium, Paenibacillus, and Pseudarthrobacter might be participated in the degradation of carbendazim. Taken together, our study provides a relatively comprehensive understanding of the effects of carbendazim and its degrading strain djl-6 on soil microbial communities.

Glucagon-like peptide-1 (GLP-1)-based therapy via G protein-coupled receptor (GPCR) GLP-1R, to attenuate hyperglycemia in critical care has attracted great attention. However, the exaggerated inflammation by GLP-1R agonist, Exendin-4, in a mouse model of burn injury was quite unexpected. Recent studies found that GPCR might elicit proinflammatory effects by switching from Gαs to Gαi signaling in the immune system. Thus, we aimed to investigate the possible Gαs to Gαi switch in GLP-1R signaling in monocyte following burn injury. Splenic monocytes from sham and burn mice 24 h following burn injury were treated with consecutive doses of Exendin-4 alone or in combination with an inhibitor of Gαi signaling (pertussis toxin, PTX), or a blocker of protein kinase A (H89). Cell viability was assessed by CCK-8, and the supernatant was collected for cytokine measurement by ELISA. Intracellular cAMP level, phosphorylated PKA activity, and nuclear NF-κB p65 were determined by ELISA, ERK1/2 activation was analyzed by Western blot. The expression of GLP-1R downstream molecules, Gαs, Gαi and G-protein coupled receptor kinase 2 (GRK2) were examined by immunofluorescence staining and Western blot. Exendin-4 could inhibit the viability of monocyte from sham rather than burn mice. Unexpectedly, it could also reduce TNF-α secretion from sham monocyte while increase it from burn monocyte. The increased secretion of TNF-α by Exendin-4 from burn monocyte could be reversed by pretreatment of PTX or H89. Accordingly, Exendin-4 could stimulates cAMP production dose dependently from sham instead of burn monocyte. However, the blunt cAMP production from burn monocyte was further suppressed by pretreatment of PTX or H89 after 6-h incubation. Nevertheless, phosphorylated PKA activity was significantly increased by low dose of Exendin-4 in sham monocyte, by contrast, it was enhanced by high dose of Exendin-4 in burn monocyte after 1-h incubation. Following Exendin-4 treatment for 2 h ex vivo, total nuclear NF-κB and phosphorylated NF-κB activity, as well as cytoplasmic pERK1/2 expressions were reduced in sham monocyte, however, only pERK1/2 was increased by Exendin-4 in burn monocytes. Moreover, reduced expressions of GLP-1R, GRK-2 and Gαs in contrast with increased expression of Gαi were identified in burn monocyte relative to sham monocyte. This study presents an unexpected proinflammatory switch from Gαs to Gαi signaling in burn monocyte, which promotes ERK1/2 and NF-κB activation and the downstream TNF-α secretion. This phenomenon is most probably responsible for proinflammatory response evoked by Gαs agonist Exendin-4 following burn injury.

// Chao Ren 1, 2 , Ya-Lin Tong 3 , Jun-Cong Li 2 , Ning Dong 2 , Ji-Wei Hao 2 , Qing-Hong Zhang 2 and Yong-Ming Yao 1, 2, 4 1 School of Medicine, Nankai University, Tianjin 300071, People’s Republic of China 2 Trauma Research Center, First Hospital Affiliated to The Chinese PLA General Hospital, Beijing 100048, People’s Republic of China 3 Department of Burns and Plastic Surgery, The 181st Hospital of Chinese PLA, Guilin 541002, People’s Republic of China 4 State Key Laboratory of Kidney Disease, The Chinese PLA General Hospital, Beijing 100853, People’s Republic of China Correspondence to: Yong-Ming Yao, email: c_ff@sina.com Keywords: HMGB1, sepsis, lateral ventricles, antagonists, brain injuries Received: July 29, 2016 Accepted: September 04, 2017 Published: October 05, 2017 ABSTRACT Sepsis induced brain injury acts as an acute complication and accounts for deterioration and high mortality rate of septic condition. HMGB1 is a late inflammatory mediator that plays a critical role in brain dysfunction and diseases. However, the role of HMGB1 in sepsis induced brain dysfunction remains intricate. The current study investigated the effect of HMGB1 on brain injury in septic mice model with intracerebroventricular injection of BoxA (a specific antagonist of HMGB1). The expression of HMGB1, morphological changes of brain tissues, apoptosis of brain cells, and alteration of behavior were determined. The expressions of HMGB1 in cortex, hippocampus, and striatum were significantly enhanced in the sepsis group when compared with the sham group. In septic conditions, brain tissues showed significant abnormalities in tissue structure, and increased apoptosis of brain cells which was caspase-3 dependent. Septic mice showed suppression of locomotor activity and impairment of memory and learning. Neutralizing brain HMGB1 significantly improved brain injury and apoptosis of brain cells, and further ameliorated disturbed locomotor activities and damaged memory and learning. However, no significant improvement of survival rate was seen after inhibiting central HMGB1. These results reveal that HMGB1 is a potential target for ameliorating sepsis induced brain injury with early antagonizing.

A novel Gram-stain-positive bacterium, designated djl-8T, was isolated from farmland soil in Nanjing, Jiangsu province, PR China. Cells of strain djl-8T were aerobic, non-motile, non-spore-forming and rod-shaped. The organism grew at 25–37 °C, pH 5.5–8.0 and 0.5–4.0 % NaCl (w/v). The DNA G+C content was 69.3 mol%. The diagnostic diamino acid in the cell-wall peptidoglycan was LL-2, 6-diaminopimelic acid. The major fatty acids (>5 %) were iso-C16 : 0, anteiso-C17 : 0, iso-C15 : 0, 10-Me C17 : 0 and C17 : 1ω8c. The respiratory quinone was MK-8 (H4) and the major polar lipids were phosphatidylglycerol, phosphatidylinositol, diphosphatidylglycerol and unknown phospholipids. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain djl-8T is a member of the genus Nocardioides and shared the highest similarity with Nocardioides ginkgobilobae SYP-A7303T (97.1 %), followed by Nocardioides soli mbc-2T (96.9 %), Nocardioide spyridinolyticus OS4T (96.6 %) and Nocardioides maradonensis RP-B30T (96.6 %). Strain djl-8T exhibited low DNA–DNA relatedness with Nocardioides ginkgobilobae SYP-A7303T (26.9±2.1 %). On the basis of the morphological, physiological, biochemical and chemotaxonomic characteristics presented in this study, strain djl-8T represents a novel species of the genus Nocardioides , for which the name Nocardioides agrisoli sp. nov. is proposed. The type strain is djl-8T (=KCTC 39844T=CCTCC AB 2017058T).

The purpose of the study was to investigate the role of the signal transducer and activator of transcription 3 (STAT3), signal transduction protein in regulating the biological characteristics of stem cells in cervical carcinoma. Overexpressed plasmid of STAT3 was constructed and used to transfect SiHa into cervical carcinoma cells. STAT3-targeted specific siRNA was designed and produced. The effects of STAT3 upregulation (or inhibition) on the expression of NANOG, OCT4 and SOX2 markers of stem cells were measured, using western blot analysis and RT-qPCR. In addition, the tumor sphere experiment was also conducted to detect the formation of tumor spheres after the intervention of expression of STAT3 and the expression of STAT3, NANOG, OCT4 and SOX2 was detected in 35 cases of cervical carcinoma tissues and 31 cases of normal cervical tissues using immunohistochemistry. We determined whether the STAT3 overexpression plasmid was successfully constructed using enzyme digestion, PCR for bacterium solution, western blot analysis and RT-qPCR and found that the plasmid met the requirements of subsequent procedures. Compared with the empty plasmid group and STAT3 low expression group, the mRNA and protein expression of markers of stem cells, OCT4, SOX2 and NANOG were significantly elevated in the STAT3 overexpression group with statistically significant differences (P

A novel carbendazim (methyl-1H-benzimidazol-2-ylcarbamate, or MBC) degrading strain SD-4 was isolated and identified preliminarily as Mycobacterium sp. according to its phenotypic features and phylogenetic analysis. This strain could utilize MBC as the sole carbon and nitrogen sources for growth and degrade 50mgL-1 MBC at the average degradation rate of 0.63mgL-1h-1. Strain SD-4 degraded MBC through the typical pathway, in which MBC was first hydrolyzed by MheI to 2-aminobenzimidazole (2-AB) and then converted to 2-hydroxybenzimidazole (2-HB). The MBC hydrolase encoding gene mheI was cloned from strain SD-4 and successfully expressed in Escherichia coli by codon optimization. The sulfhydryl-blocking assay revealed that the activity of MheI was closely related to cysteine, and the site-directed mutation experiment showed that Cys16 and Cys222 played important roles during the hydrolysis of MBC by MheI. Therefore they affected its activity directly and were defined as the key amino acid sites.

Arctiin is the most abundant bioactive compound contained in the Arctium lappa plant. In our previous study, we isolated one single bacterium capable of bioconverting arctigenin, an aglycone of arctiin, to 3′-desmethylarctigenin (3′-DMAG) solely. However, to date, a specific bacterium capable of producing other arctiin metabolites has not been reported. In this study, we isolated one single bacterium, which we named Eggerthella sp. AUH-JLD49s, capable of bioconverting 3′-DMAG under anaerobic conditions. The metabolite of 3′-DMAG by strain AUH-JLD49s was identified as 3′-desmethyl-4′-dehydroxyarctigenin (DMDH-AG) based on electrospray ionization mass spectrometry (ESI–MS) and 1H and 13C nuclear magnetic resonance spectroscopy. The bioconversion kinetics and bioconversion capacity of strain AUH-JLD49s were investigated. In addition, the metabolite DMDH-AG showed an inhibitory effect on cell growth of human colon cancer cell line HCT116 and human breast cancer cell line MDA-MB-231.

Methomyl (S-methyl N-(methylcarbamoyloxy) thioacetimidate) is a kind of oxime carbamate insecticide. It is considered to be extremely toxic to nontarget organism. To date, no pure culture or consortium has been reported to have the ability to degrade methomyl completely. In this study, a methomyl-degrading enrichment E1 was obtained by using the sludge from the wastewater-treating system of a pesticide manufacturer as the original inoculant. Two bacterial strains named MDW-2 and MDW-3 were isolated from this enrichment, and they were preliminarily identified as Aminobacter sp. and Afipia sp. respectively. Strains MDW-2 and MDW-3 could coexist and degrade 50 mg l-1 methomyl completely within 3 days by the cooperative metabolism. Methomyl was first converted to methomyl oxime and methylcarbamic acid by strain MDW-2, and the latter could be used as the carbon source for the growth of strain MDW-2. But methomyl oxime could not be sequentially degraded by strain MDW-2. However, it could be degraded and used as the carbon source by strain MDW-3. Significance and impact of the study This study presents a bacterial combination of Aminobacter sp. MDW-2 and Afipia sp. MDW-3, which could degrade methomyl completely by biochemical cooperation. This study also proposes the biodegradation pathway of methomyl for the first time and highlights the application potential of a bacterial combination in the remediation of methomyl-contaminated environments.

The present study investigated the efficiency of the use of chitosan nanoparticles containing plasmid‑bone morphogenetic protein 2 (pBMP2) sequences (CNPBs) to induce the differentiation of bone marrow stem cells (BMSCs) into osteoblast‑like cells that may be able to promote ectopic bone formation. pBMP2s were constructed, and chitosan nanoparticles were incubated with 50, 100 or 200 µg/ml pBMP2. BMSCs were collected from the tibiae and femurs of 6‑week old rats, cultured and treated with the CNPBs or 200 µg/ml pBMP2 as a positive control. Transfection efficiency was confirmed using the green fluorescent protein assay. Histological staining methods, including alkaline phosphatase, Wright's and von Kasso staining, were used to identify features of osteoblast‑like cells differentiated from BMSCs. Expression levels of the markers of osteoblasts, such as alkaline phosphatase, osteoprotegerin, osteocalcin and osteopontin, were determined to verify the differentiation of BMSCs into osteoblast‑like cells. Ectopic bone formation was observed following the integration of polyglycolic acid (PGA) scaffolds with CNPBs and BMSCs, which were implanted into the dorsal muscles of Sprague‑Dawley rats. Exposure to CNPBs led to the transfection of BMSCs with BMP2. The transfected BMSCs possessed the characteristic phenotypes of osteoblasts. The expression levels of alkaline phosphatase, osteoprotegerin, osteocalcin and osteopontin were significantly higher in the transfected cells compared with the control group, particularly the CBP200 group. PGA scaffolds integrated with BMSCs and CNPBs induced ectopic bone formation, as changes in the morphology of cells were observed using histological staining. Therefore, CNPBs may be a promising method of promoting the formation of novel bone tissue.

A copper-tolerant phenanthrene (PHE)-degrading bacterium, strain Sphingobium sp. PHE-1, was newly isolated from the activated sludge in a wastewater treatment plant. Two key genes, ahdA1b-1 encoding polycyclic aromatic hydrocarbon ring-hydroxylating dioxygenase (PAH-RHDɑ) and xyLE encoding catechol-2,3-dioxygenase (C23O), involved in the PHE metabolism by strain PHE-1 were identified. The PAH-RHD gene cluster showed 96% identity with the same cluster of Sphingomonas sp. P2. Our results indicated the induced transcription of xylE and ahdA1b-1 genes by PHE, simultaneously promoted by Cu(II). For the first time, high concentration of Cu(II) is found to encourage the expression of PAH-RHDɑ and C23O genes during PHE degradation. Applying Sphingomonas PHE-1 in PHE-contaminated soils for bioaugmentation, the abundance of xylE gene was increased by the planting of ryegrass and the presence of Cu(II), which, in turn, benefited ryegrass growth. The best performance of PHE degradation and the highest abundance of xylE genes occurred in PHE-copper co-contaminated soils planted with ryegrass.

Focally amplified lncRNA on chromosome 1 (FALEC) is novel lncRNA located in a focal amplicon on chromosome 1q21.2, and has been identified as an oncogenic properties in a variety of human cancers. However, there was no report about the expression pattern and biological function of FALEC in endometrial cancer. In our research, FALEC expression was increased in endometrial cancer tissue samples and cell lines compared with corresponding paracancerous normal tissue samples and cell line, respectively. Furthermore, we investigated the clinical significance of FALEC in endometrial cancer patients, and found endometrial cancer patients with advanced clinical stage or large tumor size had higher levels of FALEC expression than those with early clinical stage or small tumor size. The in vitro studies showed silencing of FALEC expression inhibited cell proliferation and arrested cell cycle at G0/G1. In conclusion, FALEC is overexpressed in endometrial cancer tissues and cells, and involved in regulating cell proliferation and cell-cycle.

Picolinic acid (PA) is a natural toxic pyridine derivative. Microorganisms can degrade and utilize PA for growth. However, the full catabolic pathway of PA and its physiological and genetic foundation remain unknown. In this study, we identified a gene cluster, designated picRCEDFB4B3B2B1A1A2A3, responsible for the degradation of PA from Alcaligenes faecalis JQ135. Our results suggest that PA degradation pathway occurs as follows: PA was initially 6-hydroxylated to 6-hydroxypicolinic acid (6HPA) by PicA (a PA dehydrogenase). 6HPA was then 3-hydroxylated by PicB, a four-component 6HPA monooxygenase, to form 3,6-dihydroxypicolinic acid (3,6DHPA), which was then converted into 2,5-dihydroxypyridine (2,5DHP) by the decarboxylase PicC. 2,5DHP was further degraded to fumaric acid through PicD (2,5DHP 5,6-dioxygenase), PicE (N-formylmaleamic acid deformylase), PicF (maleamic acid amidohydrolase), and PicG (maleic acid isomerase). Homologous pic gene clusters with diverse organizations were found to be widely distributed in Alpha-, Beta-, and Gammaproteobacteria. Our findings provide new insights into the microbial catabolism of environmental toxic pyridine derivatives. IMPORTANCE Picolinic acid is a common metabolite of l-tryptophan and some aromatic compounds and is an important intermediate in organic chemical synthesis. Although the microbial degradation/detoxification of picolinic acid has been studied for over 50 years, the underlying molecular mechanisms are still unknown. Here, we show that the pic gene cluster is responsible for the complete degradation of picolinic acid. The pic gene cluster was found to be widespread in other Alpha-, Beta-, and Gammaproteobacteria. These findings provide a new perspective for understanding the catabolic mechanisms of picolinic acid in bacteria.