Goose Hepatic IGFBP2 Is Regulated by Nutritional Status and Participates in Energy Metabolism Mainly through the Cytokine-Cytokine Receptor Pathway.

Changes in the nutritional status of animals significantly affect their health and production performance. However, it is unclear whether insulin-like growth factor-binding protein 2 (IGFBP2) mediates these effects. This study aimed to investigate the impact of changes in nutritional and energy statuses on hepatic IGFBP2 expression and the mechanism through which IGFBP2 plays a mediating role. Therefore, the expression of IGFBP2 was first determined in the livers of fasting/refeeding and overfeeding geese. The data showed that overfeeding inhibited IGFBP2 expression in the liver compared with the control (normal feeding) group, whereas the expression of IGFBP2 in the liver was induced by fasting. Interestingly, the data indicated that insulin inhibited the expression of IGFBP2 in goose primary hepatocytes, suggesting that the changes in IGFBP2 expression in the liver in the abovementioned models may be partially attributed to the blood insulin levels. Furthermore, transcriptome sequencing analysis showed that the overexpression of IGFBP2 in geese primary hepatocytes significantly altered the expression of 337 genes (including 111 up-regulated and 226 down-regulated genes), and these differentially expressed genes were mainly enriched in cytokine-cytokine receptor, immune, and lipid metabolism-related pathways. We selected the most significant pathway, the cytokine-cytokine receptor pathway, and found that the relationship between the expression of these genes and IGFBP2 in goose liver was in line with the findings from the IGFBP2 overexpression assay, i.e., the decreased expression of IGFBP2 was accompanied by the increased expression of LOC106041919, CCL20, LOC106042256, LOC106041041, and IL22RA1 in the overfed versus normally fed geese, and the increased expression of IGFBP2 was accompanied by the decreased expression of these genes in fasting versus normally fed geese, and refeeding prevented or attenuated the effects of fasting. The association between the expression of these genes and IGFBP2 was verified by IGFBP2-siRNA treatment of goose primary hepatocytes, in which IGFBP2 expression was induced by low serum concentrations. In conclusion, this study suggests that IGFBP2 mediates the biological effects induced by changes in nutritional or energy levels, mainly through the cytokine-cytokine receptor pathway.

Study on the Mechanism of MC5R Participating in Energy Metabolism of Goose Liver.

Nutrition and energy levels have an important impact on animal growth, production performance, disease occurrence and health recovery. Previous studies indicate that melanocortin 5 receptor (MC5R) is mainly involved in the regulations of exocrine gland function, lipid metabolism and immune response in animals. However, it is not clear how MC5R participates in the nutrition and energy metabolism of animals. To address this, the widely used animal models, including the overfeeding model and the fasting/refeeding model, could provide an effective tool. In this study, the expression of MC5R in goose liver was first determined in these models. Goose primary hepatocytes were then treated with nutrition/energy metabolism-related factors (glucose, oleic acid and thyroxine), which is followed by determination of MC5R gene expression. Moreover, MC5R was overexpressed in goose primary hepatocytes, followed by identification of differentially expressed genes (DEGs) and pathways subjected to MC5R regulation by transcriptome analysis. At last, some of the genes potentially regulated by MC5R were also identified in the in vivo and in vitro models, and were used to predict possible regulatory networks with PPI (protein-protein interaction networks) program. The data showed that both overfeeding and refeeding inhibited the expression of MC5R in goose liver, while fasting induced the expression of MC5R. Glucose and oleic acid could induce the expression of MC5R in goose primary hepatocytes, whereas thyroxine could inhibit it. The overexpression of MC5R significantly affected the expression of 1381 genes, and the pathways enriched with the DEGs mainly include oxidative phosphorylation, focal adhesion, ECM-receptor interaction, glutathione metabolism and MAPK signaling pathway. Interestingly, some pathways are related to glycolipid metabolism, including oxidative phosphorylation, pyruvate metabolism, citrate cycle, etc. Using the in vivo and in vitro models, it was demonstrated that the expression of some DEGs, including ACSL1, PSPH, HMGCS1, CPT1A, PACSIN2, IGFBP3, NMRK1, GYS2, ECI2, NDRG1, CDK9, FBXO25, SLC25A25, USP25 and AHCY, was associated with the expression of MC5R, suggesting these genes may mediate the biological role of MC5R in these models. In addition, PPI analysis suggests that the selected downstream genes, including GYS2, ECI2, PSPH, CPT1A, ACSL1, HMGCS1, USP25 and NDRG1, participate in the protein-protein interaction network regulated by MC5R. In conclusion, MC5R may mediate the biological effects caused by changes in nutrition and energy levels in goose hepatocytes through multiple pathways, including glycolipid-metabolism-related pathways.

Glucose-induced enhanced anti-oxidant activity inhibits apoptosis in goose fatty liver.

The development of mammalian nonalcoholic fatty liver disease is associated with oxidative stress, reduced mitochondrial function, and increased apoptosis in hepatocytes; however, the expressions of mitochondria-related genes are elevated in goose fatty liver, suggesting that there may be a unique protective mechanism in goose fatty liver. The aim of the study was to investigate this protective mechanism in terms of anti-oxidant capacity. Our data showed no substantial differences in the mRNA expression levels of the apoptosis-related genes including B-cell lymphoma-2 (Bcl-2), BCL2-associated X (Bax), cysteinyl aspartate-specific proteinase-3 (Caspase-3), and cysteinyl aspartate-specific proteinase-9 (Caspase-9) in the livers of the control and overfeeding Lander geese groups. The protein expression levels of Caspase-3 and cleaved Caspase-9 were not markedly different between the groups. Compared with the control group, malondialdehyde content was significantly lower (P < 0.01), glutathione peroxidase (GSH-Px) activity, glutathione (GSH) content, and mitochondrial membrane potential levels were higher (P < 0.01) in the overfeeding group. The mRNA expression levels of the anti-oxidant genes superoxide dismutase 1 (SOD1), glutathione peroxidase 1 (GPX1), and glutathione peroxidase 2 (GPX2) were increased in goose primary hepatocytes after 40 mM and 60 mM glucose treatment. Reactive oxygen species (ROS) levels were significantly reduced (P < 0.01), whereas the mitochondrial membrane potential was maintained at normal levels. The mRNA expression levels of the apoptosis-related genes Bcl-2, Bax, and Caspase-3 were not substantial. There were no significant differences in the expression levels of Caspase-3 and cleaved Caspase-9 proteins. In conclusion, glucose-induced enhanced anti-oxidant capacity may help protect the function of mitochondria and inhibit the occurrence of apoptosis in goose fatty liver.

No significant pathological symptoms were observed in the liver of goose after overfeeding, suggesting that a specific protection mechanism exists in goose liver. Previous studies have shown that mitochondria may participate in the formation of goose fatty liver by improving its energy metabolism and the production of precursor metabolites. To further understand the role of mitochondria in the formation of goose fatty liver, the present study investigated the changes of mitochondrial function, anti-oxidant capacity, and apoptosis in goose fatty liver. There were found that the level of mitochondrial membrane potential was increased, no apoptosis was observed and anti-oxidant capacity was improved in goose fatty liver, no apoptosis was observed and anti-oxidant genes expressions were increased in goose primary hepatocytes after 40 mM glucose treatment. Our findings imply that apoptosis is inhibited by glucose-induced enhanced anti-oxidant activity in goose fatty liver. Our study not only contributes to revealing the protective mechanism in goose fatty liver but also providing new references for the study of nonalcoholic fatty liver in mammals.

A compact mesoband microwave radiation system.

A compact mesoband radiation system is designed and tested. The system is composed of a charging power supply, a Marx generator, a switched quarter-wavelength oscillator, and a circularly polarized patch antenna. The Marx generator outputs a fast high-voltage pulse with a rise time of 10 ns and an amplitude of 100 kV, which is used to charge the oscillator. The oscillator consists of a quarter-wavelength coaxial transmission line, a ring ground switch, and a coupler. It generates a mesoband oscillation pulse with a center frequency of 350 MHz and a percentage bandwidth of 10%. The oscillation pulse is radiated by the circularly polarized patch antenna. The measured radiation factor is greater than 50 kV, and the radiation waveform is consistent with the simulation waveform.

Synthesis of Azetidine Nitrones and Exomethylene Oxazolines through a Copper(I)-Catalyzed 2,3-Rearrangement and 4π-Electrocyclization Cascade Strategy.

A variety of azetidine nitrones are prepared in moderate to good yields through copper(I) combined with 2-aminopyridine to catalyze skeletal rearrangement of O-propargylic oximes. Mechanistic studies reveal that the reaction undergoes a copper(I)-catalyzed tandem [2,3]-rearrangement, 4π-electrocyclization, ring opening, and recyclization over four steps in one pot. Substituents at the terminus of alkyne and oxime moieties have a significant impact on the formation of azetidine nitrones and exomethylene oxazolines, respectively. Furthermore, the obtained azetidine nitrone could easily participate in [3 + 2] cycloaddition with alkynoates, and a [2.2]-paracyclophane-derived azetidine nitrone is synthesized in 45% yield over five steps from bromo[2.2]-paracyclophane.

Avenanthramide C induces cellular senescence in colorectal cancer cells via suppressing ß-catenin-mediated the transcription of miR-183/96/182 cluster.

Cellular senescence is representing a potential anticancer therapeutic arsenal. Avenanthramide C (AVN C), as a signature compound of oats, exhibits antioxidant, anti-inflammatory, anti-atherosclerotic, and anti-tumor activities. However, the relationship between AVN C and cellular senescence in tumors remains largely unclear. Here, we elucidated that AVN C treatment predisposed colorectal cancer cells to senescent phenotype confirmed by flattened and enlarged shape characteristics, elevated senescence-associated ß-galactosidase (SA-ß-Gal) activity, and G1 phase arrest. Furthermore, AVN C triggered cellular senescence via transcriptionally repressing miR-183/96/182 cluster and subsequently reduced the levels of mature miR-183, -96, and -182. Mechanistically, AVN C exerted its senescence induction by attenuating ß-catenin-mediated transactivation of miR-183/96/182 cluster to unleash its common target FOXO1 and two other targets, FOXO3 and SMAD4, which subsequently foster the p21 and p16 expression. In addition, AVN C is also noted to facilitate p53-mediated p21 transactivation via suppressing ß-catenin. Collectively, we identified a novel mechanism of ß-catenin/miR-183/96/182 cluster/FOXO1 mediated-CRC cellular senescence that entails that AVN C serves as an auxiliary agent for CRC treatment.

Identification and characterization of a central replication origin of the mega-plasmid pSCATT of Streptomyces cattleya.

Streptomyces linear plasmids often contain an internal replication origin. In this study, a new replication origin was identified and confirmed in the 1.8-Mb plasmid pSCATT of Streptomyces cattleya DSM46488. The real-time qPCR results indicated that the copy number of pSCATT was one copy per chromosome. The identified replication origin oriC1-II was found to locate in the central region of pSCATT and was 2 kb in size. This replication origin consists of a protein-coding gene SCATT_p08010 with an unknown function and the upstream non-coding sequence. Deletion or disruption analysis of SCATT_p08010 or the upstream non-coding sequence revealed that both SCATT_p08010 and the non-coding sequence were essential for replication. However, the identified replication origin was shown to endow the plasmid with the ability to replicate in a circular model but not in a linear model in S. lividans. Interestingly, the knockout of the replication origin did not result in the curing of pSCATT, indicating that there might be other replication origins present in the mega-plasmid. The experimental validation of the central replication origin oriC1-II might be helpful for the investigation of the replication mechanism of the mega-plasmid and the genome evolution of Streptomyces.

Thermogravimetric analysis of the pyrolysis and combustion kinetics of surface dead combustibles in the Daxing'an Mountains.

In boreal regions, the frequency of forest fires is increasing. In this study, thermogravimetric analysis was used to analyze the pyrolysis kinetics of dead surface combustibles in different forest types within the Daxing'an Mountains, China. The results show that the combustible material load of forest types, the Larix forest (LG) is relatively high. Base on the E of kinetic parameters, the LG, and Quercus forest (QM) forest types had relatively high combustibility values and comprehensive combustibility values for 1-, 10-, and 100-h time lags. According to the obtained P values, the pyrolysis of dead surface fuels with 1-, 10-, and 100-h time lags is relatively difficult in the Larix / Betula mixed forest (L-B) and QM forest types. Therefore, mixed forests of the LG, L-B, and QM tree species can be established as fire-resistant forests to establish a fire barrier, reduce the combustibility of forest stands, and reduce the possibility of forest fires.

Nestin Promotes Peritoneal Fibrosis by Protecting HIF1-α From Proteasomal Degradation.

BACKGROUND: Peritoneal dialysis (PD) is a treatment for end stage renal disease patients, but it can also cause peritoneal fibrosis. Nestin is known as a neural stem cell marker and it has many functions. The hypoxia induced factor (HIF) signaling pathway can be activated under hypoxia conditions, leading to the overexpression of some angiogenesis related genes. The aim of our study is to demonstrate Nestin's role in the development of peritoneal fibrosis (PF), and to provide a new target (Nestin) to treat PF. METHODS: PD mice models were constructed by an intraperitoneal administration of PDS at 10 ml/100g/d for 4 weeks. Nestin-positive cells were isolated from peritonea of Nestin-GFP mice by flow cytometry. The relationship of Nestin and HIF1-α-VEGFA pathway was detected by Nestin knockdown, Co-immunoprecipitation and immunofluorescence. Also, proteasomal activity was demonstrated by CHX and MG132 application, followed by Western blotting and Co-immunoprecipitation. RESULTS: In our experiments, we found that Nestin expression resulted in PF. Also, HIF1-α/VEGFA pathway was activated in PF. Nestin knockdown reduced the level of HIF1-α. Nestin directly bound to HIF1-α and protected HIF1-α from proteasomal degradation. Overexpression of HIF1-α reverts the fibrosis levels in Nestin-knockdown cells. In brief, Nestin inhibited the degradation of HIF1-α by mitigating its ubiquitination level, leading to the activation of HIF1-α signaling pathway, and eventually promoted PF. CONCLUSION: We found a novel mechanism of PF that Nestin promotes by protecting HIF1-α from proteasomal degradation. Taken together, our key findings highlight a novel mechanism by which the silencing of Nestin hinders HIF1- α -induced PF.

Exploring the pharmacological effects and potential targets of paeoniflorin on the endometriosis of cold coagulation and blood stasis model rats by ultra-performance liquid chromatography tandem mass spectrometry with a pattern recognition approach.

This study was employed to explore the potential biomarkers of endometriosis of cold coagulation and blood stasis (ECB) model rats and the effective mechanism of action of paeoniflorin (PF). The serum metabolomics approach was carried out using the UPLC-MS technique with a pattern recognition approach to prove the possible biomarkers of the ECB model rats and the perturbed pathways. Subsequently, the mechanism of PF treatment of this disease model was elucidated. The results revealed that the serum metabolism profiles in two groups were also separated significantly. Moreover, 8 biomarkers were found in the positive mode, and 5 biomarkers were found in the negative mode. Totally, 13 biomarkers participated in the metabolism of phenylalanine, arachidonic acid, etc. After treatment with PF, 10 biomarkers were regulated. Among the 10 biomarkers, 4 were statistically significant: l-phenylalanine, l-tryptophan, LysoPC (18:4(6Z,9Z,12Z,15Z)), and LysoPC (16:1(9Z)). We initially confirmed that PF could significantly regulate the metabolic expression of multiple metabolic pathways in the ECB model rats. For the first time, this study explored the mechanism of action of PF treatment based on the metabolic pathways of the organism and demonstrated the potential of the metabolomics techniques for the study of drug action mechanisms.

Comparative Analysis of CRISPR Loci Found in Streptomyces Genome Sequences.

The interspaced short palindromic repeats (CRISPR) system is an immune system widely distributed in prokaryotes, resisting the invasion of the foreign mobile genetic elements like phages or plasmids. In this study, we present the comparative analysis of 182 CRISPR loci found in 46 publicly available complete genome sequences of Streptomyces. Overall, nine direct repeats (DRs) groups are identified while all the 2104 spacers are divided into three main groups according to the multiple sequence alignment. Only 11 spacers are identical with parts of 10 plasmid sequences, which indicates a possible origin. The cas gene clusters near the CRISPR arrays are found to mainly belong to the I-E subtype. These CRISPR loci might play an important role in the genome evolution of Streptomyces.

High-throughput metabolomics used to identify potential therapeutic targets of Guizhi Fuling Wan against endometriosis of cold coagulation and blood stasis.

Metabolomics is an emerging and robust discipline and involves the comprehensive evaluation of small molecule endogenous metabolites and enables the exploration of the pathogenesis of diseases. For example, endometriosis - a common disease which mostly occurs in women of childbearing age. A cure for endometriosis of cold coagulation and blood stasis (ECB) is highly sought after. This study was conducted to discover the potential biomarkers of ECB and the effective mechanism undertaken by Guizhi Fuling Wan (GFW) in treating ECB in rats. Urinary metabolomics were performed by using UPLC-Q-TOF-MS with pattern recognition methods to evaluate the changes in metabolic profiles and to identify biomarkers for elucidating the mechanism of the treatment of ECB with GFW. The results showed that urinary metabolism in the two groups were distinctly separated on the 28th day, and a total of 20 differential biomarkers (16 in the positive mode, 4 in the negative mode) were confirmed involving several key metabolic pathways which included phenylalanine, tyrosine and tryptophan biosynthesis, valine, leucine and isoleucine biosynthesis, glyoxylate and dicarboxylate metabolism, tyrosine metabolism and the citrate cycle. Following the oral administration of GFW, certain pathways were affected; these included the following: phenylalanine, tyrosine and tryptophan biosynthesis, valine, leucine and isoleucine biosynthesis, glyoxylate and dicarboxylate metabolism, tyrosine metabolism, citrate cycle, steroid hormone biosynthesis, tryptophan metabolism, phenylalanine metabolism, primary bile acid biosynthesis, and aminoacyl-tRNA biosynthesis. This study also demonstrated that the administration of GFW affected the levels of urine endogenous metabolites, thereby laying a foundation for further study of the pharmacodynamical mechanism of GFW.

RASAL1 influences the proliferation and invasion of gastric cancer cells by regulating the RAS/ERK signaling pathway.

The aim of this study was to investigate the biological characteristics of the RASAL1 gene in a well-differentiated gastric cancer cell line MKN-28 and a poorly differentiated gastric cancer cell line BGC-823 cells, using RNA interference and gene transfection technology, respectively. MKN-28 cells were transfected with the shRNA of RASAL1 and BGC-823 cells were transfected with the pcDNA 3.1 plasmid vector containing RASAL1. RT-PCR and western blotting were then used to detect the expression of RASAL1 mRNA and protein. The activities of RAS and extracellular signal-regulated kinase 1/2 were analyzed by the pull-down method and western blotting. The proliferate capacity, apoptosis rate, invasive and migratory potentials of MKN-28 or BGC-823 cells were also measured by Cell Counting Kit-8 cell proliferation assay, propidium iodide/Annexin V staining coupled with flow cytometry, and transwell chamber assays, respectively. Measurement of RASAL1 mRNA and protein expression in two cells revealed successful transfection of the shRNA of RASAL1 and RASAL1-pcDNA3.1 plasmid into these two cells. Moreover, decreased expression of RASAL1 in MKN-28 cells resulted in increased expression of RAS-GTP and p-ERK1/2. Interestingly, decreased expression of RASAL1 inhibited apoptosis and facilitated cell proliferation, invasion and migration. The increased expression of RASAL1 in BGC-823 cells caused declined expression of RAS-GTP and p-ERK1/2, as well as promoted apoptosis and restrained cell proliferation, invasion and migration. The down-regulation of RASAL1 promoted the proliferation, invasion and migration of gastric cancer MKN-28 cells, and up-regulation of RASAL1 inhibited the proliferation, invasion and migration of BGC-823 gastric cancer cells by regulating the RAS/ERK signaling pathway. Thus, our results suggest that RASAL1 may play an important role as a tumor suppressor gene in gastric cancer.

Detection method and toxicity study of a new disinfection by-product, 2,2,4-trichloro-5-methoxycyclopenta-4-ene-1,3-dione (TCMCD), in chlorinated drinking water.

The detection method of 2,2,4-trichloro-5-methoxycyclopenta-4-ene-1,3-dione (TCMCD), a new disinfection by-product, was established and optimized. Drinking water samples from eight cities in Jiangsu Province, China, were analyzed and TCMCD was detected ranging from 42 to 171 ng/L among the water samples from the three cities. Toxicity of TCMCD was studied using the exposure of zebrafish and the human peripheral blood lymphocyte micronucleus assay. The results showed that TCMCD is fatal to zebrafish embryos and is a potential mutagen to human beings.

The interference of 2-chloro-5-oxo-3-hexene diacyl chloride (COHC) in the detection of strong mutagen MX.

The MX formation potential of some compounds belong to benzoic acids, benzoic aldehydes and phenols families was investigated. Only some structures with aldehyde group can form MX, which is inconsistent with other research. A new compound 2-chloro-5-oxo-3-hexene diacyl chloride (COHC) was found. It will seriously interfere with the detection of MX when the GC/MS instrument is operated in select ion monitoring mode because its retention time is very close to that of methylated MX and it has a set of characteristic ions m/z 199, 201, 203, which is widely used in the detection of MX. The detection of COHC has not been reported previously. However, according to its formation mechanism it will widely occur in chlorinated drinking and humic water. Due to the presence of COHC, some of the former research results on MX should be revised.

[Detection of the strong mutagen MX in drinking water].

MX [3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone] was found to be the most potent mutagen in chlorinated drinking water. MX in some tap waters in Jiangsu Province was absorbed by XAD-8 resin, methylized with saturated BF3-CH3OH and detected using GC/MS. The contents of MX in the tap waters were between 0.58 to 20.0 ng.L-1, except one of those samples. More MX was produced in low-water season.

[Multidrug resistance mechanisms in cell line HL-60/VCR].

BACKGROUND & OBJECTIVE: Drug resistance is a major factor in chemotherapeutic failure of leukemia. Multidrug resistant cell lines are the good models for investigating the mechanisms and reversal of acquired drug resistance. This study was designed to explore the multidrug resistance (MDR) mechanisms in cell line HL-60/VCR. METHODS: Flow cytometry and a panel of antibodies were used to analyze the expression of MDR proteins (P-gp, MRP, LRP, BCRP, GST-pi) and apoptosis-modulating proteins (bcl-2, bcl-x, bax, bad) in MDR cell line HL-60/VCR and drug sensitive cell line HL-60. RESULTS: The expression levels of MDR proteins (P-gp, MRP, BCRP, GST-pi) were (18.62, 1.19, 1.50, 1.32-flod) higher in HL-60/VCR than in HL-60, while the expression of LRP level was similar. The levels of apoptosis-modulating proteins(bcl-2, bcl-x, bad) were (2.48, 1.25, 1.08-fold) higher in HL-60/VCR than in HL-60, while the pro-apoptosis protein bax contrarily decreased in HL-60/VCR. CONCLUSION: Various MDR mechanisms were involved in multi-drug resistance HL-60/VCR cell line, which including increasing expression of drug-resistance protein (P-gp, MRP, BCRP, and GST-pi); the apoptosis-modulating proteins (bcl-2, bcl-x, bax, and bad) might take part in the mechanism of drug resistance.