Identification of Isthmin-1 in common carp (Cyprinus carpio L.) and the effects on glucose metabolism in vivo and in vitro.

Isthmin-1 (Ism1) plays roles in glucose uptake in mammals as an adipokine. To investigate its role in the glucose metabolism of common carp (Cyprinus carpio. L), the Ism1 sequence was cloned, and its expression and distribution in tissues were detected. In addition, we prepared and purified the recombinant Ism1 protein using the E. coli expression system and assessed changes in the expression of key genes related to glucose metabolism through both in vivo injection experiments and primary hepatocyte experiments in vitro. The results revealed that the open reading frame of Ism1 was 1377 bp long, encoding 458 amino acids. Similarity analysis indicated that Ism1 exhibited a close evolutionary relationship with goldfish (Carassius auratus), sharing 98.35% amino acid similarity. Ism1 was expressed in all tissues of common carp, with the highest level observed in the heart, followed by the gill, head kidney, and hepatopancreas. Distinct patterns of Ism1 expression were identified during the oral glucose tolerance test and long-term high-carbohydrate and high-fat diet feeding experiments. In vivo studies demonstrated that the serum glucose concentration was reduced on treatment with Ism1, accompanied by a significant upregulation of mRNA levels for gk, hk, and pfk genes in hepatopancreas; conversely pepck and g6pase mRNA levels were significantly downregulated in the hepatopancreas under these conditions as well. Furthermore, our primary hepatocyte experiment confirmed that Ism1 could inhibit pepck and g6pase mRNA expression, while promoting gk, hk, and pfk mRNA expression levels. In conclusion, Ism1, in common carp, could participate in the glucose metabolism, which provides essential information for future studies on the function of Ism1.

Establishment and characterization of Yellow River carp (Cyprinus carpio haematopterus) muscle cell line and its application to fish virology and immunology.

The Yellow River carp (Cyprinus carpio haematopterus) is a vital economically farmed fish of the Cyprinidae family. With the development of intensive aquaculture, carp production has increased dramatically, leading to the frequent occurrence of various diseases. Cell lines are considered the most cost-effective resource for in vitro studies and are widely used for physiological and pathological studies because of accessibility and convenience. This research established a novel immortal cell line CCM (Yellow River carp muscle cells) derived from the carp muscle. CCM has been passed over 71 generations for 1 year. The morphology of CCM and the adhesion and extension processes were captured by light and electron microscopy. CCM were passaged every 3 days with 20% FBS DMEM/F12 at 1:3. The optimum conditions for CCM growth were 28 °C and 20% FBS concentration. DNA sequencing of 16S rRNA and COI showed that CCM was derived from carp. CCM positively reacts to anti-PAX7 and anti-MyoD antibodies of carp. Analysis of chromosomes revealed that the chromosomal pattern number of CCM was 100. Transfection experiment demonstrated that CCM might be utilized to express foreign genes. Furthermore, cytotoxicity testing showed that CCM was susceptible to Aeromonas hydrophila, Aeromonas salmonicida, Aeromonas veronii, and Staphylococcus Aureus. The organophosphate pesticides (chlorpyrifos and glyphosate) or heavy metals (Hg, Cd, and Cu) exhibited dose-dependent cytotoxicity against CCM. After LPS treatment, the MyD88-IRAKs-NFκB pathway stimulates inflammatory-related factor il1ß, il8, il10, and nfκb expression. LPS did not seem to cause oxidative stress in CCM, and the expression of cat and sod was not affected. Poly (I:C) through TLR3-TRIF-MyD88-TRAF6-NFκB and TRIF-TRAF3-TBK1-IRF3 activated the transcription of related factors, increased expression of anti-viral protein, but no changes in apoptosis-related genes. To our knowledge, this is the first muscle cell line in Yellow River carp and the first study on the immune response signal pathways of Yellow River carp based on the muscle cell line. CCM cell line provides a more rapid and efficient experimental material for fish immunology research, and this study preliminarily elucidated its immune response strategy to LPS and poly (I:C).

Dietary Bile Acid Supplementation Could Regulate the Glucose, Lipid Metabolism, and Microbiota of Common Carp (Cyprinus carpio L.) Fed with a High-Lipid Diet.

This study sought to examine the role of bile acids in the regulation of glucose and lipid metabolism, intestinal flora, and growth in high-fat diet-fed common carp (Cyprinus carpio L.). Fish (6.34 ± 0.07 g) were fed for 56 days with three different diets, the control diet (CO, 5.4% lipid), high-fat diet (HF, 11% lipid), and high-fat diet with 60 mg/kg bile acids (BAs, 11% lipid). The results showed that high-fat diets resulted in poor growth performance and increased triglyceride (TG) in serum and the liver. The addition of bile acids significantly alleviated the adverse effects of a high-fat diet. The mRNA expression results indicated that bile acids may improve lipid metabolism through the enhancement of the peroxisome proliferator-activated receptor (PPARa). The expression of gluconeogenesis-related phosphoenolpyruvate carboxykinase (PEPCK) mRNA was inhibited, while fibroblast growth factor 19 (FGF19) was significantly higher. Bile acids reshaped the intestinal microflora community, with the level of Bacteroidetes increasing. The correlation analysis indicated that Patescibacteria, Dependentiae, Myxococcota, and Planctomycetota in the gut are associated with genes involved in glucose and lipid metabolism. These results indicated that bile acids could ameliorate the negative effects of high-fat diets on common carp.

Effects of Genistein on Lipid Metabolism, Antioxidant Activity, and Immunity of Common Carp (Cyprinus carpio L.) Fed with High-Carbohydrate and High-Fat Diets.

A 56-day feeding trial was conducted to investigate the effects of genistein on growth, lipid metabolism, antioxidant capacity, and immunity of common carp fed with high-carbohydrate or high-fat diets. Five diets were used to feed fish: control diet (5% fat; CO), high-fat diet (11% fat; HF), high-carbohydrate diet (45% carbohydrate; HC), and HF or HC diet with 500 mg/kg genistein (FG or CG). Results showed that final body weight (FW) and specific growth rate (SGR) were significantly reduced, but the supplementation with genistein resulted in higher values of FW and SGR than the HF or HC group. Both high carbohydrate and high fat belong to high-energy diets, which may promote lipid deposition. Genistein obviously decreased liver triglyceride (TG) content and alleviated hepatic fat vacuolation in the HF and HC groups. The expression of lipid metabolism genes (cpt-1 and atgl) was markedly higher in the FG group than in the HF group. The lipid synthesis-related genes (fas, acc, and pparγ) were elevated in high-energy diets but recovered to the control level or reduced after genistein treatments. With respect to fatty acid transporter genes, fatp increased in the FG group, and cd36 increased in the CG group. Furthermore, the antioxidant and immune indexes, such as total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-PX), superoxide dismutase (SOD), acid phosphatase (ACP), and lysozyme (LZM) activities, were decreased, while malonate aldehyde (MDA) content, activities of alanine aminotransferase (ALT), and aspartate aminotransferase (AST) were enhanced in the HF and HC groups. The antioxidant and immunity values could be ameliorated by treatment with genistein. Moreover, the transcript levels of antioxidant-related genes (cat, gr, and nrf2) in the liver and anti-inflammatory factors (tgf-ß and il-10) and lyz in the head kidney tissue were promoted, although the expression levels of proinflammatory factors (tnf-α and il-6) declined in the genistein supplementation group, which confirmed the antioxidant and immune-enhancing effects of genistein. Therefore, 500 mg/kg genistein could ameliorate the negative effects of high-energy diets on immunity.

Fabrication of a Cation-Exchange Membrane via the Blending of SPES/N-Phthaloyl Chitosan/MIL-101(Fe) Using Response Surface Methodology for Desalination.

In the present work, a novel mixed matrix cation exchange membrane composed of sulfonated polyether sulfone (SPES), N-phthaloyl chitosan (NPHCs) and MIL-101(Fe) was synthesized using response surface methodology (RSM). The electrochemical and physical properties of the membrane, such as ion exchange capacity, water content, morphology, contact angle, fixed ion concentration and thermal stability were investigated. The RSM based on the Box-Behnken design (BBD) model was employed to simulate and evaluate the influence of preparation conditions on the properties of CEMs. The regression model was validated via the analysis of variance (ANOVA) which exhibited a high reliability and accuracy of the results. Moreover, the experimental data have a good fit and high reproducibility with the predicted results according to the regression analysis. The embedding of MIL-101(Fe) nanoparticles contributed to the improvement of ion selective separation by forming hydrogen bonds with the polymer network in the membrane. The optimum synthesis parameters such as degree of sulfonation (DS), the content of SPES and NPHCs and the content of MIL-101(Fe) were acquired to be 30%, 85:15 and 2%, respectively, and the corresponding desalination rate of the CEMs improved to 136% while the energy consumption reduced to 90%. These results revealed that the RSM was a promising strategy for optimizing the preparation factors of CEMs and other similar multi-response optimization studies.

Vitreoscilla Hemoglobin Improves Sophorolipid Production in Starmerella Bombicola O-13-1 Under Oxygen Limited Conditions.

Sophorolipids (SLs) are homologous microbial secondary metabolites produced by Starmerella bombicola and have been widely applied in many industrial fields. The biosynthesis of SLs is a highly aerobic process and is often limited by low dissolved oxygen (DO) levels. In this study, the Vitreoscilla hemoglobin (VHb) gene was transformed into S. bombicola O-13-1 by homologous recombination to alleviate oxygen limitation. VHb expression improved the intracellular oxygen utilization efficiency under either oxygen-rich or oxygen-limited conditions. In shake flask culture, the production of SLs was higher in the recombinant (VHb+) strain than in the wild-type (VHb-) strain, while the oxygen uptake rate of the recombinant (VHb+) strain was significantly lower than that of the wild-type (VHb-) strain. In a 5 L bioreactor, the production of SLs did not increase significantly, but the DO level in the fermentation broth of the VHb+ strain was 21.8% higher than that of VHb- strain under oxygen-rich conditions. Compared to wide-type strains (VHb-), VHb expression enhanced SLs production by 25.1% in the recombinants (VHb+) under oxygen-limited conditions. In addition, VHb expression raised the transcription levels of key genes involved in the electron transfer chain (NDH, SDH, COX), TCA cycle (CS, ICD, KDG1) and SL synthesis (CYP52M1 and UGTA1) in the recombinant (VHb+) strains. VHb expression in S. bombicola could enhance SLs biosynthesis and intracellular oxygen utilization efficiency by increasing ATP production and cellular respiration. Our findings highlight the potential use of VHb to improve the oxygen utilization efficiency of S. bombicola in the industrial-scale production of SLs using industrial and agricultural by-products like molasses and waste oil as fermentation feedstock.

Molecular identification of glucose transporter 4: The responsiveness to starvation, glucose, insulin and glucagon on glucose transporter 4 in common carp (Cyprinus carpio L.).

Glucose transporter 4 (GLUT4) is comprehensively investigated in mammals, while the comparative research of GLUT4 in common carp is deficient. To investigate the function of GLUT4, carp glut4 was first isolated. The open reading frame of carp glut4 was 1518 bp in length, encoding 505 amino acids. A high-sequence homology was identified in carp and teleost, and the phylogenetic tree displayed that the carp GLUT4 was clustered with the teleost. A high level of glut4 mRNA was analysed in fat, red muscle and white muscle. After fasting treatment, glut4 mRNA expression was increased significantly in muscle. In the oral glucose tolerance test experiment, glut4 mRNA was also significantly elevated in muscle, gut and fat. Furthermore, intraperitoneal injection of insulin resulted in the upregulation of glut4 gene expression significantly in white muscle, gut and fat. On the contrary, the glut4 mRNA level in the white muscle, gut and fat was markedly downregulated after glucagon injection. These results suggest that GLUT4 might play important roles in food intake and could be regulated by nutrient condition, insulin and glucagon in common carp. Our study is the first to report on GLUT4 in common carp. These data provide a basis for further study on fish GLUT4.

Multipronged Regulatory Functions of Serum Albumin in Early Stages of Amyloid-ß Aggregation.

Human serum albumin (HSA) is a major interacting-partner of Alzheimer's amyloid-ß (Aß) peptide in the plasma and has emerged as a promising therapeutic target. HSA inhibits Aß fibrillization, but the underlying molecular mechanism is not well elucidated. In this work, we investigated the role of HSA in the early stages of Aß aggregation by simulating the binding process of multiple Aß monomers and protofibrils to HSA with extensive molecular dynamics simulations. HSA could simultaneously trap multiple Aß monomers and accommodate the formation of nonfibrillar Aß oligomers after binding. In particular, domains I and III show stronger binding capacities and hold preferable interaction sites for oligomers. Consequently, HSA prevents the formation of fibrillar oligomers in water, thus interfering with the nucleation process. On the other aspect, when protofibrils are preformed, HSA tends to block the ß-strand spanning the central hydrophobic core located at the protofibril end, preventing the addition of monomers to protofibrils. Furthermore, Aß protofibril structures are severely disrupted both globally and locally. Thus, further growth of protofibrils to fibrils is impeded by HSA. Our results collectively indicate that HSA performs multipronged regulatory functions in the early stages of Aß aggregation. Our work advances the understanding of the amyloid inhibition of Aß by HSA and provides theoretical guidance for developing rational therapies of Alzheimer's disease.

Caregiver burden and parenting stress among left-behind elderly individuals in rural China: a cross-sectional study.

BACKGROUND: One public health problem that cannot be ignored is the mental health of left-behind elderly individuals in rural areas. However, the burden of care and parenting stress among left-behind elderly individuals has never been analyzed. The purpose of this study was to explore the level of caregiver burden and parenting stress and their relationship among left-behind elderly individuals. METHODS: A total of 261 left-behind elderly people responded to the study. The 22-item Zarit Burden Interview and the 36-item Parenting Stress Index-Short Form were used. RESULTS: We sent out 300 questionnaires in total. The effective rate was 87% (n = 261). Among the left-behind elderly individuals, most respondents were female (n = 171; 65.5%). The results showed that older age (OR:3.04; 95%CI: 1.307-7.048), an annual income of ¥5000-¥9900 (OR:3.25; 95%CI: 1.192-8.852) and higher parenting stress (OR:1.17; 95%CI: 1.103-1.242) were the risk factors related to higher caregiver burden in the left-behind elderly individuals. The influencing factor for lower caregiver burden in the left-behind elderly was gender (being male) (OR:0.08; 95%CI:0.036-0.178). Age (r = - 0.789; P < 0.001) and gender (r = 0.325; P < 0.001) were significant positively correlated with parenting stress, and annual income (r = - 0.717; P < 0.001) was negatively correlated with parenting stress. CONCLUSION: Parenting stress is a risk factor affecting caregiver burden of left-behind elderly individuals. Healthcare professionals should pay close attention to the caregiver burden and parenting stress of left-behind elderly individuals, especially those who are older, female and have lower income.

Relationship between neck circumference and hypertension of primary and middle school students:a Meta-analysis / 中国学校卫生

Objective@#To systematically evaluate the relationship between neck circumference and hypertension of primary and secondary school students.@*Methods@#Web of science, PubMed, Scopus, CNKI and WanFang databases were searched by computer, and the retrieval time was from inception to December 2019. Two researchers independently screened the literature, extracted the data and evaluated the quality, and then performed Meta-analysis using Stata 14.0 software.@*Results@#A total of 8 studies were included, including 20 475 subjects. Meta-analysis results showed that the risk of hypertension increased by 35% in people with a high neck circumference compared with the normal population(OR=1.35, 95%CI=1.20-1.51, P<0.01). The results of subgroup analysis showed that the correlation between neck circumference and hypertension of obese primary and secondary school students was 1.41 times higher than that of normal weight students(OR=1.41, 95%CI=1.23-1.61, P<0.01). The correlation between the neck circumference and the risk of hypertension of primary and secondary school students in Europe and America was more significant than that in Asia(OR=1.31, 95%CI=1.11-1.53, P=0.01). When the mean value of neck circumference was greater than 28.5 cm(OR=1.29, 95%CI=1.02-1.64, P=0.03), it was associated with the incidence of hypertension.@*Conclusion@#The neck circumference of primary and middle school students is related to the risk of hypertension, especially in obese people. Blood pressure monitoring and health education should be strengthened to prevent hypertension.