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X-box protein 1 (Xbp1), an essential transcription factor including an unstable form (Xbp1-u) and a stable form (Xbp1-s), plays an vital role in B cell activation and differentiation to plasma cells. In this study, we cloned and identified Xbp1-u gene from Nile tilapia (Oreochromis niloticus), containing 783 bp of nucleotide sequence encoding 260 amino acids. The deduced protein possesses a basic region leucine zipper domain (bZIP) and 26 ribonucleotides of OnXbp1-u transcript. Transcription analysis revealed OnXbp1-u and OnXbp1-s were widely distributed in all examined tissues, with a high expression in immune-related tissues. When stimulated with Streptococcus agalactiae in vivo, the expressions of OnXbp1-u and OnXbp1-s were significantly up-regulated in liver, spleen, head kidney, blood, skin and intestine. After in vitro challenge upon S.agalactiae, the similar up-regulations of OnXbp1-u and OnXbp1-s were also demonstrated in head kidney leukocytes. Moreover, the OnXbp1-u and OnXbp1-s could get involved in LPS-inducible B cell activation and (r)OnIL6-inducible B cell differentiation. Taken together, the results indicated that OnXbp1-u and OnXbp1-s might not only involved in the immune response against S. agalactiae challenge, but also in the B cell activation and differentiation in Nile tilapia.

Chemokines are a class of small molecular weight cytokines of 6–14 kDa, exerting important roles in the regulation of various inflammatory diseases and immune cell migration. In this study, we have identified the CXCL12 gene from Nile tilapia (Oreochromis niloticus), including CXCL12a (OnCXCL12a) and CXCL12b (OnCXCL12b). The open reading frames of OnCXCL12a and OnCXCL12b are 309 and 297 bp, encoding 102 and 98 amino acids, respectively. Multiple alignment showed that OnCXCL12a and OnCXCL12b have characteristics of CXC chemokines and share high identity with CXCL12 amino acid sequences from the known species. Tissue distribution in the healthy fish indicated that OnCXCL12a and OnCXCL12b expressed in all examined tissues, with the highest expression in muscle and anterior kidney, respectively. After challenged by Streptococcus agalactiae, Poly(I:C) and LPS in vivo and in vitro, OnCXCL12 is transcriptionally up-regulated in immune tissues and cells significantly. The recombinant OnCXCL12 proteins, (r)OnCXCL12a and (r)OnCXCL12b, enhance the release of nitric oxide and increase the expression of inflammatory cytokines (TNF-α, IL-6, and IL-10) in anterior kidney leukocytes, as well as exhibit chemotactic activity for leukocytes from anterior kidney. Summarizing, these results indicate that OnCXCL12 is involved in the immune response of Nile tilapia against pathogen infection and may play an important role in mediating inflammatory response.

The potential synergistic effects of Flavan-3-ols to inhibit the formation of acrylamide have attracted wide attention. This study aims to investigate the synergistic inhibitory effects of the B-type procyanidin and flavan-3-ols monomer on acrylamide production in food matrix. By comparing the inhibitory effects of different compounds in food matrix, procyanidin B2 and catechin were screened out for examining their synergistic inhibitory effects on acrylamide by Isobologram analysis. When the interaction index (γ) was lower than 1, the interaction was synergistic reaction. The results showed the procyanidin B2/catechin ratio of 1:3 (γ = 0.57) had similar synergistic effect to that of 1:9 (γ = 0.53), both of which showed better effects than the ratio of 1:1 (γ = 0.71). The optimal synergistic inhibitory effect (70.11 ± 2.07%) was achieved when the procyanidin B2 and catechin concentrations were 0.6 µg/mL and 5.4 µg/mL, respectively. Compared with single use, the combined use of B2 and catechin could decrease the dosage of B2 to 1/10 and that of catechin to 1/3.

In arable soils co-influenced by mining and farming, soil bacteria significantly affect metal (Cadmium, Cd) bioavailability and accumulation. To reveal the soil microecology response under this co-influence, three intersection areas (cornfield, vegetable field, and paddy field) were investigated. With a similar nutrient condition, the soils showed varied Cd levels (0.31-7.70 mg/kg), which was negatively related to the distance from mining water flow. Different soils showed varied microbial community structures, which were dominated by Chloroflexi (19.64-24.82%), Actinobacteria (15.49-31.96%), Acidobacteriota (9.46-20.31%), and Proteobacteria (11.88-14.57%) phyla. A strong correlation was observed between functional microbial taxon (e. g. Acidobacteriota), soil physicochemical properties, and Cd contents. The relative abundance of tolerant bacteria including Vicinamibacteraceae, Knoellia, Ardenticatenales, Lysobacter, etc. elevated with the increase of Cd, which contributed to the enrichment of heavy metal resistance genes (HRGs) and integration genes (intlI), thus enhancing the resistance to heavy metal pollution. Cd content rather than crop species was identified as the dominant factor that influenced the bacterial community. Nevertheless, the peculiar agrotype of the paddy field contributed to its higher HRGs and intlI abundance. These results provided fundamental information about the crop-specific physiochemical-bacterial interaction, which was helpful to evaluate agricultural environmental risk around the intersection of farmland and pollution sources.

Complement 1q-binding protein (gC1qR), a multi-functional cellular protein, is able to bind the globular head region of C1q molecule (gC1q), playing an important role in regulation of immune response and host defense against bacterial infection. In this study, the full length of a gC1qR ortholog (OngC1qR) was identified and characterized from Nile tilapia (Oreochromis niloticus). The cDNA of OngC1qR ORFs consisted of 870 bp of nucleotide sequence encoding a 289 amino acid residue polypeptide. The deduced amino acid sequence is highly homologous to teleost, containing a MAM33p domain, two potential glycosylation sites, and a putative cell adhesion site EGD (Glu-Gly-Asp). Spatial mRNA expression analysis revealed that the OngC1qR was expressed ubiquitously in all examined tissues, with a high expression in the liver. The OngC1qR expression was significantly up-regulated in liver, spleen and head kidney following in vivo challenges with Streptococcus agalactiae. A high up-regulation was also detected in head kidney leukocytes in vitro stimulation with Streptococcus agalactiae. In addition, immunofluorescence detection illustrated that the OngC1qR was located at the cell surface of leukocytes. Recombinant OngC1qR protein was able to not only bind LPS, LTA, S. agalactiae and Aeromonas hydrophila, but also possess binding capability with the ligand OnC1qs. Taken together, the results indicated that OngC1qR might be involved in host defense against bacterial infection in Nile tilapia.

Polymeric immunoglobulin receptor (pIgR) plays important roles in transport of polymeric immunoglobulins across mucosal epithelial cells and prevention of access of pathogens into the organisms. In this study, a pIgR ortholog (OnpIgR) was cloned and identified from Nile tilapia (Oreochromis niloticus). The open reading frame (ORF) of OnpIgR is 1023 bp, encoding a predicted protein of 341 amino acids. The deduced amino acid sequence of OnpIgR is highly homologous to other teleost's pIgRs, containing two Ig-like domains (ILDs), a transmembrane region and an intracellular region. Expression analysis revealed that OnpIgR was exhibited in all tested tissues including liver, spleen, skin and intestines, and highly expressed in the liver. Immunofluorescence detection indicated that the natural pIgR expressed on the membrane of the epithelial cells. After challenge of bacterial pathogen (Streptococcus agalactiae), OnpIgR expression was significantly up-regulated in intestines and skin, as well as intestinal epithelial cells. In addition, recombinant OnIFN-γ induced the expression of pIgR in the intestinal epithelial cells. Moreover, recombinant OnpIgR protein was able to bind both S. agalactiae and A. hydrophila in vitro, and interact with (r)OnIgM and (r)OnIgT. Taken together, the results indicated that OnpIgR, involving in the polymeric immunoglobulin transport, might play an important role in mucosal immunity against bacterial infection.

Sorghum procyanidins (SPC) tetramers were reported to have inhibitory effects on the adhesion of Streptococcus mutans (S. mutans), which is one of the primary pathogenic bacteria that cause caries. To make clear the mechanism underlying the inhibitory effects of SPC tetramers on the adhesion of S. mutans, it is important to understand the interaction between SPC tetramers and the catalytic region of glucosyltransferases-I (GTF-I/CAT) from S. mutans. In this study, fluorescence quenching, UV-visible (UV-Vis) absorption and circular dichroism (CD) spectroscopies were applied to investigate the interaction between SPC tetramers and the GTF-I/CAT from S. mutans UA159. The fluorescence quenching results demonstrated that SPC tetramers combined with GTF-I/CAT protein on one binding site, and the fluorescence intensity of GTF-I/CAT protein was decreased regularly by static quenching with increasing concentrations of SPC tetramers. The UV-Vis absorption spectra of GTF-I/CAT protein were also changed with a red shift owing to the impact of SPC tetramers. In addition, the proportions of α-helix, β-sheet and random coil in the secondary structure of GTF-I/CAT protein were obviously altered by SPC tetramers. In conclusion, SPC tetramers have a strong affinity for GTF-I/CAT protein by altering its conformation and micro-environment. Our results suggest that SPC tetramers interact with GTF-I/CAT to interfere with the adhesion of S. mutans, which may facilitate the better application of SPC tetramers in the prevention of dental caries.

The red-light running rate by type of road users has not been reported in China so far. We conducted an observation study to report the violation rate in Changsha, China. Portable digital devices were used to record red-light running violations at five selected intersections. The observation was performed for three days (weekday, weekend and holiday), four time periods per day and an hour per time period (peak and off-peak hours in the morning and in the afternoon). Violation rate was calculated as number of violations divided by total number of vehicles/pedestrians × 100%. We used adjusted violation rate ratio (VRR) to quantify the effects of type of day and time period based on Poisson model. Totally, 162,124 vehicles (including motor vehicles, motorcycles and bicycles) and 31,649 pedestrians were recorded. The red-light running rate was 0.14% for motor vehicle drivers, far lowering than those for motorcyclists (18.64%), bicyclists (18.74%) and pedestrians (18.54%). The rate on holiday was 1.89 times that on weekday for drivers. The rate for motorcyclists was high in off-peak hours (adjusted VRR: 1.11), but low on weekend and on holiday (adjusted VRRs: 0.80 and 0.65). The rate for bicyclists was 32% lower on weekend than on weekday. For pedestrians, the rates were high on weekend and holiday and in off-peak hours (adjusted VRR: 1.09, 1.67 and 1.30). The red-light running rate of motor vehicle drivers is far lower than those for motorcyclists, bicyclists and pedestrians. The effects of type of day and time period on violation rate vary with road users, indicating the type of day and time period should be considered when developing and implementing interventions to reduce red-light running of different road users.

It has been reported that B-type procyanidins can alleviate oxidative damage of prostatic cells, but there has been limited information on the similar role of A-type procyanidins. This study investigated the protective effect of procyanidin A-type dimers from peanut skin against H2O2-induced oxidative stress damage in prostate cancer DU145 cells. According to the UPLC-Q-TOF-MS/MS analysis and comparison with standards, the fourth fraction of peanut skin procyanidin (PSP-4) was identified as procyanidin A-type dimers, namely, procyanidin A1 and A2. Results revealed that PSP-4 treatment prior H2O2 exposure increased cell activity and attenuated the cell cycle arrest and apoptosis rate. The H2O2-induced increase in intracellular reactive oxygen species (ROS) was remarkably inhibited by PSP-4. PSP-4 treatment enhanced the activity of catalase (CAT) and total super oxide dismutase (T-SOD) and restored glutathione (GSH) content, compared with the H2O2 treatment. Furthermore, the results indicated that PSP-4 protected DU145 cells by attenuating phosphorylation of the mitogen-activated protein kinases (MAPKs), by increasing the Bcl-2/Bax ratio, and by reducing the activation of caspase-3 and caspase-9 by cascade reactions. This study reveals that procyanidin A-type dimers from peanut skin have the potential function in preventing oxidative stress damage of prostatic cells.

Interleukin 6 (IL-6) is a pleiotropic cytokine that exerts its biological functions through interaction with its receptor system consisting of a ligand-specific IL-6 receptor (IL-6R) and a common signal-transducing receptor (gp130). In this study, OnIL-6R and Ongp130 genes from Nile tilapia (Oreochromis niloticus) were identified, and their roles in bacterial or viral infection and in regulation of inflammatory response involved in IL-6 were investigated. The open reading frames (ORFs) of OnIL-6R and Ongp130 are 2019 bp and 2679 bp, encoding 672 and 892 amino acids, respectively. Domain analysis of the deduced amino acid sequences of OnIL-6R and Ongp130 showed that both of them contained a conserved Ig-like domain, FNIII domains, and a WSXWS motif. The transcripts of OnIL-6R and Ongp130 were widely expressed in all examined tissues. Following in vivo challenges with Streptococcus agalactia, Poly I: C and lipopolysaccharide (LPS), the mRNAs of OnIL-6R and Ongp130 were notably induced in liver, head kidney and spleen. The transcriptional up-regulations of OnIL-6R and Ongp130 were also detected in Nile tilapia monocytes/macrophages and lymphocytes after in vitro stimulations with S. agalactiae, Poly I: C and LPS. Besides, increasing mRNA levels of the inflammation-related cytokines (IL-1β, TNF-α, IL-6, IL-10, and MIF) induced by recombinant OnIL-6 could be further enhanced by co-treatment with recombinant soluble OnIL-6R in lymphocytes. Furthermore, recombinant soluble Ongp130 suppressed the induction of expression of these cytokines in lymphocytes when co-stimulated with (r)OnIL-6 and (r)sOnIL-6R. Taken together, these results indicated that OnIL-6R and Ongp130 were likely involved in the resistance to bacterial or viral infection in Nile tilapia. Moreover, soluble OnIL-6R and soluble Ongp130 have an agonistic effect or antagonistic effect in the inflammation response involved in OnIL-6.

The mechanisms of the electrochemical oxidation of orientin and its interaction with deoxyribonucleic acid have been investigated using glassy carbon electrode. The electrochemical response of orientin is due to oxidation of phenolic hydroxyl groups on orientin. The whole process is controlled by the adsorption step and concerned 4 electrons and 4 protons. Negative shift of potential and decrease of peak current for electrochemical oxidation of orientin can be observed at bare glassy carbon electrode and deoxyribonucleic acid modified electrode in 0.05 M Na2HPO4-KH2PO4 (pH 7.48), confirming the dominant electrostatic interaction between orientin and deoxyribonucleic acid. Moreover, a reliable and sensitive method of reversed-phase high-performance liquid chromatography-electrochemical detection has been developed for simultaneous determination of the isomer pair orientin/isoorientin. Chromatographic separation was carried out on a reversed-phase C18 column and a mobile phase comprised of acetonitrile and acetate (0.5%, pH 2.97) by amperometric detection with a glassy carbon electrode at the working potential of +1.00 V. The method was validated for linearity, accuracy, precision, and limit of detection. Under the optimized conditions, linear regression analysis for the calibration curve showed a good linear relationship between peak area and their concentrations in the linear range of 89.2 nM to 59.5 μM for orientin with detection limit of 14.9 nM and 78.0 nM to 52.0 μM for isoorientin with a detection limit of 13.0 nM, respectively. Compared to the method using ultraviolet detection, the detection limits are greatly lowered. Finally, the proposed method has been successfully applied to the determination of orientin and isoorientin in Lophatherum gracile Brongn.

Complement component 1q (C1q), together with C1r and C1s to form C1, recognize and bind immune complex to initiate the classical complement pathway. In this study, C1q subunit molecules (XlC1qA, XlC1qB, XlC1qC) were cloned and analyzed from Xenopus laevis (X. laevis). The open reading frame (ORF) of XlC1qA is 819 bp of nucleotide sequence encoding 272 amino acids, the ORF of XlC1qB is 711 bp encoding 236 aa, and the XlC1qC is consists of 732 bp encoding 243 aa. The deduced amino acid sequences contain a collagen-like region (CLR), Gly-X-Y repeats in the N-terminus and a C1q family domain at the C-terminus. Phylogenetic analysis revealed that the XlC1qs are clustered with the amphibian clade. Expression analysis indicated that the XlC1qs exhibited constitutive expression in all examined tissues, with the highest expression in liver. Additionally, XlC1q could interact with heat-aggregated mouse IgG and IgM, Xenopus IgM and Nile tilapia IgM, respectively, indicating the functional conservation of XlC1q binding to immunoglobulins. Further, XlC1qs can inhibit C1q-dependent hemolysis of sensitized sheep red blood cells with concentration-dependent manner. These data collectively suggest that the function of C1qs in X. laevis may be conserved in interaction with immunoglobulins, as that of mammals and teleosts.

A bifunctional aptamer-based electrochemical biosensor for label-free detection of thrombin and adenosine has been developed. It was based on the principle of switching structures of aptamers from DNA/DNA duplex to DNA/target complex upon target addition. Thrombin aptamer was firstly immobilized on gold electrode surface by self-assembly, and then it was hybridized with adenosine aptamer. Methylene blue (MB), as an electrochemical indicator, was abundantly adsorbed on the aptamers via specific interaction of MB with guanine base. In the presence of thrombin or adenosine, the aptamer part could bind with thrombin or adenosine instead of MB. The decreased peak current of MB was intimately related to the concentration of thrombin or adenosine. In present work, the assay was linear in the ranges from 6 to 60 nM for thrombin and from 10 to 1000 nM for adenosine. Detection limits were determined to be 3 nM for thrombin and 10 nM for adenosine, respectively. Moreover, bovine plasma albumin (BSA) and lysozyme, or uridine and guanosine, did not influence performance of the biosensor, indicating good selectivity of this sensor for thrombin or adenosine detection. Finally, this sensor was successfully applied to thrombin and adenosine analysis in human plasma samples.