Elevated circulating GPHB5 levels in women with insulin resistance and polycystic ovary syndrome: A cross-sectional study and multiple intervention studies

ObjectiveGPHB5 has been found to be associated with glucose and lipid metabolism in animal studies. However, the association of GPHB5 with IR and metabolic disorders remains unknown, and there is a lack of research in humans. Our aim in this study was to investigate the relationship between circulating GPHB5 and metabolic disorders in humans.MethodsBioinformatics analysis was performed to understand the relationship between GPHB5 and metabolic disorders. GPHB5 mRNA expression in mice and rats was determined using RT-qPCR. Circulating GPHB5 concentrations were measured with an ELISA kit. EHC and OGTT were performed in humans.ResultsBioinformatics analysis shows that GPHB5 is associated with metabolic disorders and PCOS. GPHB5 mRNA expression levels in the metabolic-related tissues of HFD-fed mice, db/db and ob/ob mice, and PCOS rats were significantly higher than those of WT mice or rats. In human studies, we find that circulating GPHB5 levels were significantly higher in women with IR and PCOS. GPHB5 levels were positively correlated with age, BMI, WHR, BP, FBG, 2 h-BG, FIns, 2 h-Ins, TC, LDL-C, HbA1c, and FFA, but negatively correlated with adiponectin. Furthermore, GPHB5 was positively correlated with DHEAS and FAI, while negatively correlated with SHBG, FSH, SHBG and FSH. The increased GPHB5 concentration was related to IR and PCOS. After the treatment of metformin, GLP-1RA (Lira), and TZDs, circulating GPHB5 levels were decreased.ConclusionsOur results reveal that circulating GPHB5 could be a biomarker and potential therapeutic target for IR and PCOS in women

Transcriptomic analysis of gills provides insights into the molecular basis of molting in Chinese mitten crab (Eriocheir sinensis)

Chinese mitten crab (Eriocheir sinensis) is an economically important freshwater aquaculture species and is a model species for research on the mechanism of molting. This study aimed to identify important candidate genes associated with the molting process and to determine the role of gills in the regulation of molting with the help of transcriptomic analysis. The transcriptomes of crabs at different molting stages—postmolt (PoM), intermolt (InM), premolt (PrM) and ecdysis (E)—were de novo assembled to generate 246,232 unigenes with a mean length of 851 bp. A total of 86,634 unigenes (35.18% of the total unigenes) were annotated against reference databases. Significantly upregulated genes were identified in postmolt compared to intermolt (1,475), intermolt compared to premolt (65), premolt compared to ecdysis (1,352), and ecdysis compared to postmolt (153), and the corresponding numbers of downregulated genes were 1,276, 32, 1,573 and 171, respectively. Chitin synthase, endochitinase, chitinase A, chitinase 3, chitinase 6 and chitin deacetylase 1 were upregulated during the postmolt and ecdysis stages, while phosphoglucomutase 3 (PGM3), glucosamine 6-phosphate deaminase (GNPDA) and glucosamine glycoside hydrolase (nagZ) were upregulated during the intermolt and premolt stages compared to the other stages. The upregulated genes were enriched in several lipid-related metabolic pathways, such as “fatty acid elongation”, “glycerophospholipid metabolism” and “sulfur metabolism”. Meanwhile, three signaling pathways, including the “phosphatidylinositol signaling system”, the “calcium signaling pathway” and the “GnRH signaling pathway” were also enriched. Tetraspanin-18, an important effector gene in the lysosomal pathway involved in cell apoptosis, up-regulate with the beginning of molting (in premolt stage) and reach the top in the ecdysis stage, and barely expressed in the intermolt stage. The expression variations in the tetraspanin-18 gene indicated that it may play an important role in the beginning of molting cycle, which might be regulated by the stress of salinity. This study revealed that the gills could participate in chitin degradation, in reestablishment of the exoskeleton and the signaling process. Based on transcriptomic analysis of the gills, we not only explored novel molecular mechanisms of molting in E. sinensis but also acquired foundational genetic data for E. sinensis

Sequencing and <i>De Novo</i> Analysis of the Hemocytes Transcriptome in <i>Litopenaeus vannamei</i> Response to White Spot Syndrome Virus Infection

<div><p>Background</p><p>White spot syndrome virus (WSSV) is a causative pathogen found in most shrimp farming areas of the world and causes large economic losses to the shrimp aquaculture. The mechanism underlying the molecular pathogenesis of the highly virulent WSSV remains unknown. To better understand the virus-host interactions at the molecular level, the transcriptome profiles in hemocytes of unchallenged and WSSV-challenged shrimp (<i>Litopenaeus vannamei</i>) were compared using a short-read deep sequencing method (Illumina).</p><p>Results</p><p>RNA-seq analysis generated more than 25.81 million clean pair end (PE) reads, which were assembled into 52,073 unigenes (mean size = 520 bp). Based on sequence similarity searches, 23,568 (45.3%) genes were identified, among which 6,562 and 7,822 unigenes were assigned to gene ontology (GO) categories and clusters of orthologous groups (COG), respectively. Searches in the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG) mapped 14,941 (63.4%) unigenes to 240 KEGG pathways. Among all the annotated unigenes, 1,179 were associated with immune-related genes. Digital gene expression (DGE) analysis revealed that the host transcriptome profile was slightly changed in the early infection (5 hours post injection) of the virus, while large transcriptional differences were identified in the late infection (48 hpi) of WSSV. The differentially expressed genes mainly involved in pattern recognition genes and some immune response factors. The results indicated that antiviral immune mechanisms were probably involved in the recognition of pathogen-associated molecular patterns.</p><p>Conclusions</p><p>This study provided a global survey of host gene activities against virus infection in a non-model organism, pacific white shrimp. Results can contribute to the in-depth study of candidate genes in white shrimp, and help to improve the current understanding of host-pathogen interactions.</p></div

Molecular Characterization and Expression Analysis of ATP-Gated P2X7 Receptor Involved in Japanese Flounder (<i>Paralichthys olivaceus</i>) Innate Immune Response

<div><p>ATP-gated P2X7 receptor (P2RX7) channel is a key component for purinergic signaling and plays important roles in the innate immune response in mammals. However, the expression, molecular properties and immune significances of P2RX7 in lower vertebrates are still very limited. Here we identified and characterized a novel bony fish <i>P2RX7</i> homologue cDNA, termed <i>poP2RX7</i>, in Japanese flounder (<i>Paralichthys olivaceus</i>). PoP2RX7 protein shares about 60–88% sequence similarity and 45–78% sequence identity with known vertebrate P2RX7 proteins. Phylogenetic analysis placed poP2RX7 and other P2RX7 proteins within their own cluster apart from other P2RX members. While the functional poP2RX7 channel shares structural features in common with known P2RX7 homologs, electrophysiological studies revealed that BzATP, the more potent agonist for known mammalian and fish P2RX7s, shows similar potency to ATP in poP2RX7 activation. <i>poP2RX7</i> mRNA constitutively expressed in all examined tissues from unstimulated healthy Japanese flounder with dominant expression in hepatopancreas and the lowest expression in head kidney, trunk kidney, spleen and gill. <i>poP2RX7</i> mRNA expression, however, was significantly induced in Japanese flounder head kidney primary cells by Poly(I:C) and bacterial endotoxin LPS stimulations. <i>In vivo</i> experiments further revealed that <i>poP2RX7</i> gene expression was substantially up-regulated by immune challenge with infectious bacteria <i>Edwardsiella tarda</i> and <i>Vibrio anguillarum</i>. Moreover, activation of poP2RX7 results in an increased gene expression of multifunctional cytokines <i>IL-1β</i> and <i>IL-6</i> in the head kidney primary cells. Collectively, we identified and characterized a novel fish P2RX7 homolog which is engaged in Japanese flounder innate immune response probably through modulation of pro-inflammatory cytokines expression.</p></div

A Service Computing Framework for Proteomics Analysis and Collaboration of Pathogenic Mechanism Studies

The booming of proteomics data has positioned multiple disciplines and research areas in a more complicated and challenging place. Moreover, the proteomics data of any defined research interests, such as for pathogenic mechanism studies of infectious diseases, have presented unstructured and heterogeneous characteristics. Thus, a service computing framework for proteomics analysis is desired to bring biologists and computer scientists into this area seamlessly and efficiently. With this regard, this work is dedicated to detail the proteomics analysis and collaboration process of pathogenic mechanism studies. We articulate this framework to serve the requirements and ease the task design by broadly reviewing the state-of-theart research and development efforts and collectively designing different informative stages. Thus, the framework has a focus of distilling different aspects, including data curation, resources distribution, standard construction and computational tasks identification, into the proteomics analysis. The framework is designed as Proteomics Analysis as a Service to deepen the understanding of the interdisciplinary research

Multiple sequence alignment of poP2X7 receptor with selected P2X7 receptor proteins.

<p>Sequence alignment was carried out by ClustalW program. Representative P2X7 receptor proteins from different species with GenBank accession numbers are hP2X7 (<i>Homo sapiens</i>, NP_002553), rP2X7 (<i>Rattus norvegicus</i>, NP_062129), sP2X7 (<i>Sparus aurata</i>, CAI59608), zP2X7 (<i>Danio rerio</i>, AAI63071) and poP2X7 (<i>Paralichthys olivaceus</i>, KC748421). Highly conserved (:), less conserved (.) and identical (*) amino acid residues identified in all the proteins are indicated. TM: transmembrane domain. The P2X family signature motif (²⁴⁴Gly–²⁷⁰Phe) was boxed in black. The two residues ¹²⁷Lys and ²⁸⁴Asn accounted for species difference of P2X7 receptors in ATP/BzATP agonist sensitivity were boxed in green <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096625#pone.0096625-Young1" target="_blank">[28]</a>. Five important residues for nucleotide binding were boxed in red and the predicted LPS/lipid-binding domain was boxed in yellow.</p

Distribution of unigenes size (>500 bp) in assembled <i>L. vannamei</i> transcriptome.

<p>Distribution of unigenes size (>500 bp) in assembled <i>L. vannamei</i> transcriptome.</p

Characterization of poP2RX7 permeability to NMDG⁺.

<p>A. Representative −100/+40 mV ramp experiment was applied to an oocyte expressing the poP2RX7 and bathed with NMDG⁺ media. For clarity only two ramps are shown, corresponding to the beginning and after 3 min of continuous application of 1 mM ATP. The values of reversal potential are shown by arrows. B. The same protocol was applied to an oocyte expressing the rP2RX7 and bathed with NMDG⁺ media. C. Representative ramp protocol applied to an oocyte expressing the poP2RX7 and bathed with LD media, no change in reversal potential is observed under these conditions. D. Summary of the changes in reversal potential (ΔE_rev) after 3 min of ATP application in oocytes expressing the poP2RX7 or the rP2RX7 bathed in NMDG⁺ or LD media. *<i>p</i><0.05, estimated by Mann-Whitney test. n = 3–7.</p

LPS and Poly(I:C)-induced gene expression of <i>poP2RX7</i> in <i>P. olivaceus</i> head kidney primary cells.

<p><i>P. olivaceus</i> head kidney primary cells were prepared as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096625#s2" target="_blank"><i>Material and methods</i></a> and stimulated with 25 µg/ml (final concentration) LPS (A) or Poly(I:C) (B). Total RNA from different time points (0, 2, 4, 6, 8, 12 and 24 h post stimulation) was extracted and the gene expression changes of <i>poP2RX7</i> were determined by qRT-PCR. <i>β-actin</i> was employed as an internal reference gene. Values labeled with different <i>lowercase</i> letters indicate significant difference (<i>p</i><0.05) among treatments. Asterisks (*) mark the significant up-regulation of <i>poP2RX7</i> mRNA compared with the untreated control group (<i>p</i><0.05). Data in this and following figures are the mean ± standard deviation of triplicate determinations from one representative experiment; similar results were obtained on two other separate experiments.</p

Sequence of primers used in this study.

<p>Sequence of primers used in this study.</p