Abstract-We have previously shown that oxytocin receptors are present in the heart and that perfusion of isolated rat hearts with oxytocin results in decreased cardiac flow rate and bradycardia. The mechanisms involved in the negative inotropic and chronotropic effects of oxytocin were investigated in isolated dog right atria in the absence of central mechanisms. Perfusion of atria through the sinus node artery with 10 Ϫ6 mol/L oxytocin over 5 minutes (8 mL/min) significantly decreased both beating rate (Ϫ14.7Ϯ4.9% of basal levels, nϭ5, PϽ0.004) and force of contraction (Ϫ52.4Ϯ9.1% of basal levels, nϭ5, PϽ0.001). Co-perfusion with 10 Ϫ6 mol/L oxytocin receptor antagonist (nϭ3) completely inhibited the effects of oxytocin on frequency (PϽ0.04) and force of contraction (PϽ0.004), indicating receptor specificity. The effects of oxytocin were also totally inhibited by co-perfusion with 5ϫ10Ϫ8 mol/L tetrodotoxin (PϽ0.02) or 10 Ϫ6 mol/L atropine (PϽ0.03) but not by 10 Ϫ6 mol/L hexamethonium, which implies that these effects are neurally mediated, primarily by intrinsic parasympathetic postganglionic neurons. Co-perfusion with 10 Ϫ6 mol/L NO synthase inhibitor (L-NAME) significantly inhibited oxytocin effects on both beating rate (Ϫ1.85Ϯ1.27% versus Ϫ14.7Ϯ4.9% in oxytocin alone, PϽ0.05) and force of contraction (Ϫ24.9Ϯ4.4% versus Ϫ52.4Ϯ9.1% in oxytocin alone, nϭ4, PϽ0.04). The effect of oxytocin on contractility was further inhibited by L-NAME at 10 Ϫ4 mol/L (Ϫ8.1Ϯ1.8%, PϽ0.01). These studies imply that the negative inotropic and chronotropic effects of oxytocin are mediated by cardiac oxytocin receptors and that intrinsic cardiac cholinergic neurons and NO are involved in these actions. (Hypertension. 2001;38:292-296.) Key Words: receptors Ⅲ neurons Ⅲ heart rate Ⅲ contractility Ⅲ nervous system, parasympathetic Ⅲ nitric oxide O xytocin, 1 of 2 major posterior pituitary hormones (oxytocin and vasopressin), is a neuropeptide synthesized primarily in magnocellular neurons in the paraventricular and supraoptic nuclei of the hypothalamus, which project to posterior pituitary, median eminence, and several brain regions. Oxytocin is also produced in peripheral tissues, 1 including the heart. We have recently shown that the heart is an important source of oxytocin production and synthesis. 2 In addition to its well-known effects on reproductive functions, such as uterine contraction, milk-ejection reflex, and induction of maternal behavior, 3,4 oxytocin was recently shown to be involved in endocrine and neuroendocrine regulation through receptor-mediated actions exerted on the heart, vasculature, and kidneys. [5][6][7][8][9] In humans, intravenous oxytocin injections cause biphasic dose-dependent changes in mean arterial pressure that consist of an initial pressor response accompanied by bradycardia and a decrease in cardiac output, followed by a prolonged fall in mean arterial pressure accompanied by increased cardiac output. 10 Oxytocin administration at the rat upper spinal cord 11 or into acutely decentralized canine stel...

Background Cordyceps militaris, an ascomycete caterpillar fungus, has been used as a traditional Chinese medicine for many years owing to its anticancer and immunomodulatory activities. Currently, artificial culturing of this beneficial fungus has been widely used and can meet the market, but systematic molecular studies on the developmental stages of cultured C. militaris at transcriptional and translational levels have not been determined.Methodology/Principal FindingsWe utilized high-throughput Illumina sequencing to obtain the transcriptomes of C. militaris mycelium and fruiting body. All clean reads were mapped to C. militaris genome and most of the reads showed perfect coverage. Alternative splicing and novel transcripts were predicted to enrich the database. Gene expression analysis revealed that 2,113 genes were up-regulated in mycelium and 599 in fruiting body. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed to analyze the genes with expression differences. Moreover, the putative cordycepin metabolism difference between different developmental stages was studied. In addition, the proteome data of mycelium and fruiting body were obtained by one-dimensional gel electrophoresis (1-DGE) coupled with nano-electrospray ionization liquid chromatography tandem mass spectrometry (nESI-LC-MS/MS). 359 and 214 proteins were detected from mycelium and fruiting body respectively. GO, KEGG and Cluster of Orthologous Groups (COG) analysis were further conducted to better understand their difference. We analyzed the amounts of some noteworthy proteins in these two samples including lectin, superoxide dismutase, glycoside hydrolase and proteins involved in cordycepin metabolism, providing important information for further protein studies.Conclusions/SignificanceThe results reveal the difference in gene expression between the mycelium and fruiting body of artificially cultivated C. militaris by transcriptome and proteome analysis. Our study provides an effective resource for the further developmental and medicinal research of this promising fungus.

Background Agrocybe aegerita, the black poplar mushroom, has been highly valued as a functional food for its medicinal and nutritional benefits. Several bioactive extracts from A. aegerita have been found to exhibit antitumor and antioxidant activities. However, limited genetic resources for A. aegerita have hindered exploration of this species.Methodology/Principal FindingsTo facilitate the research on A. aegerita, we established a deep survey of the transcriptome and proteome of this mushroom. We applied high-throughput sequencing technology (Illumina) to sequence A. aegerita transcriptomes from mycelium and fruiting body. The raw clean reads were de novo assembled into a total of 36,134 expressed sequences tags (ESTs) with an average length of 663 bp. These ESTs were annotated and classified according to Gene Ontology (GO), Clusters of Orthologous Groups (COG), and Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathways. Gene expression profile analysis showed that 18,474 ESTs were differentially expressed, with 10,131 up-regulated in mycelium and 8,343 up-regulated in fruiting body. Putative genes involved in polysaccharide and steroid biosynthesis were identified from A. aegerita transcriptome, and these genes were differentially expressed at the two stages of A. aegerita. Based on one-dimensional gel electrophoresis (1-DGE) coupled with electrospray ionization liquid chromatography tandem MS (LC-ESI-MS/MS), we identified a total of 309 non-redundant proteins. And many metabolic enzymes involved in glycolysis were identified in the protein database.Conclusions/SignificanceThis is the first study on transcriptome and proteome analyses of A. aegerita. The data in this study serve as a resource of A. aegerita transcripts and proteins, and offer clues to the applications of this mushroom in nutrition, pharmacy and industry.

Background Ganoderma lucidum is a basidiomycete white rot fungus and is of medicinal importance in China, Japan and other countries in the Asiatic region. To date, much research has been performed in identifying the medicinal ingredients in Ganoderma lucidum. Despite its important therapeutic effects in disease, little is known about Ganoderma lucidum at the genomic level. In order to gain a molecular understanding of this fungus, we utilized Illumina high-throughput technology to sequence and analyze the transcriptome of Ganoderma lucidum.Methodology/Principal FindingsWe obtained 6,439,690 and 6,416,670 high-quality reads from the mycelium and fruiting body of Ganoderma lucidum, and these were assembled to form 18,892 and 27,408 unigenes, respectively. A similarity search was performed against the NCBI non-redundant nucleotide database and a customized database composed of five fungal genomes. 11,098 and 8, 775 unigenes were matched to the NCBI non-redundant nucleotide database and our customized database, respectively. All unigenes were subjected to annotation by Gene Ontology, Eukaryotic Orthologous Group terms and Kyoto Encyclopedia of Genes and Genomes. Differentially expressed genes from the Ganoderma lucidum mycelium and fruiting body stage were analyzed, resulting in the identification of 13 unigenes which are involved in the terpenoid backbone biosynthesis pathway. Quantitative real-time PCR was used to confirm the expression levels of these unigenes. Ganoderma lucidum was also studied for wood degrading activity and a total of 22 putative FOLymes (fungal oxidative lignin enzymes) and 120 CAZymes (carbohydrate-active enzymes) were predicted from our Ganoderma lucidum transcriptome.ConclusionsOur study provides comprehensive gene expression information on Ganoderma lucidum at the transcriptional level, which will form the foundation for functional genomics studies in this fungus. The use of Illumina sequencing technology has made de novo transcriptome assembly and gene expression analysis possible in species that lack full genome information.

A novel lectin was isolated from the mushroom Agrocybe aegerita (designated AAL-2) by affinity chromatography with GlcNAc (N-acetylglucosamine)-coupled Sepharose 6B after ammonium sulfate precipitation. The AAL-2 coding sequence (1224 bp) was identified by performing a homologous search of the five tryptic peptides identified by MS against the translated transcriptome of A. aegerita. The molecular mass of AAL-2 was calculated to be 43.175 kDa from MS, which was consistent with the data calculated from the amino acid sequence. To analyse the carbohydrate-binding properties of AAL-2, a glycan array composed of 465 glycan candidates was employed, and the result showed that AAL-2 bound with high selectivity to terminal non-reducing GlcNAc residues, and further analysis revealed that AAL-2 bound to terminal non-reducing GlcNAc residues with higher affinity than previously well-known GlcNAc-binding lectins such as WGA (wheatgerm agglutinin) and GSL-II (Griffonia simplicifolia lectin-II). ITC (isothermal titration calorimetry) showed further that GlcNAc bound to AAL-2 in a sequential manner with moderate affinity. In the present study, we also evaluated the anti-tumour activity of AAL-2. The results showed that AAL-2 could bind to the surface of hepatoma cells, leading to induced cell apoptosis in vitro. Furthermore, AAL-2 exerted an anti-hepatoma effect via inhibition of tumour growth and prolongation of survival time of tumour-bearing mice in vivo.

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