Diastereoselective Generation of C2-Azlactonized 2H-Chromenes via Brønsted Acid-Catalyzed Oxo-Cyclization of Propargyl Alcohols.

A new Brønsted acid-catalyzed oxo-cyclization of propargyl alcohols with azlactones to synthesize C2-azlactonized 2H-chromenes has been established that uses 1,1'-binaphthyl-2,2'-diyl hydrogen phosphate (BiNPO4H) as the catalyst and gives excellent diastereoselectivities (≥19:1 dr) in most cases. This protocol has a high compatibility with various substituents of substrates, offering a catalytic and useful entry to the fabrication of the synthetically important C2-functionalized 2H-chromene scaffold.

Succinate induces skeletal muscle fiber remodeling via SUNCR1 signaling.

The conversion of skeletal muscle fiber from fast twitch to slow-twitch is important for sustained and tonic contractile events, maintenance of energy homeostasis, and the alleviation of fatigue. Skeletal muscle remodeling is effectively induced by endurance or aerobic exercise, which also generates several tricarboxylic acid (TCA) cycle intermediates, including succinate. However, whether succinate regulates muscle fiber-type transitions remains unclear. Here, we found that dietary succinate supplementation increased endurance exercise ability, myosin heavy chain I expression, aerobic enzyme activity, oxygen consumption, and mitochondrial biogenesis in mouse skeletal muscle. By contrast, succinate decreased lactate dehydrogenase activity, lactate production, and myosin heavy chain IIb expression. Further, by using pharmacological or genetic loss-of-function models generated by phospholipase Cß antagonists, SUNCR1 global knockout, or SUNCR1 gastrocnemius-specific knockdown, we found that the effects of succinate on skeletal muscle fiber-type remodeling are mediated by SUNCR1 and its downstream calcium/NFAT signaling pathway. In summary, our results demonstrate succinate induces transition of skeletal muscle fiber via SUNCR1 signaling pathway. These findings suggest the potential beneficial use of succinate-based compounds in both athletic and sedentary populations.

Substrate-Controlled Generation of 3-Sulfonylated 1-Indenones and 3-Arylated ( Z)-Indenes via Cu-Catalyzed Radical Cyclization Cascades of o-Alkynylbenzonitriles.

Substrate-controlled generation of 3-sulfonylated 1-indenones and 3-arylated ( Z)-indenes through radical cyclization cascades of o-alkynylbenzonitriles with sulfonyl hydrazides was established by combining copper(II) with tert-butyl hydroperoxide (TBHP) as a catalytic oxidative system. With the installation of the aryl group (R1) into the alkynyl unit, sulfonyl radicals triggered addition-cyclization cascades to access 3-sulfonylated 1-indenones with good yields by using a Cu/TBHP system, whereas a series of new functionalized 3-arylated ( Z)-indenes were obtained in good yields when o-alkynylbenzonitriles bearing the alkyl group in the alkynyl unit were subjected with arylsulfonyl hydrazides with an electron-rich nature under the above conditions.

miR-361-3p regulates FSH by targeting FSHB in a porcine anterior pituitary cell model.

FSH plays an essential role in processes involved in human reproduction, including spermatogenesis and the ovarian cycle. While the transcriptional regulatory mechanisms underlying its synthesis and secretion have been extensively studied, little is known about its posttranscriptional regulation. A bioinformatics analysis from our group indicated that a microRNA (miRNA; miR-361-3p) could regulate FSH secretion by potentially targeting the FSHB subunit. Herein, we sought to confirm these findings by investigating the miR-361-3p-mediated regulation of FSH production in primary pig anterior pituitary cells. Gonadotropin-releasing hormone (GnRH) treatment resulted in an increase in FSHB synthesis at both the mRNA, protein/hormone level, along with a significant decrease in miR-361-3p and its precursor (pre-miR-361) levels in time- and dose-dependent manner. Using the Dual-Luciferase Assay, we confirmed that miR-361-3p directly targets FSHB. Additionally, overexpression of miR-361-3p using mimics significantly decreased the FSHB production at both the mRNA and protein levels, with a reduction in both protein synthesis and secretion. Conversely, both synthesis and secretion were significantly increased following miR-361-3p blockade. To confirm that miR-361-3p targets FSHB, we designed FSH-targeted siRNAs, and co-transfected anterior pituitary cells with both the siRNA and miR-361-3p inhibitors. Our results indicated that the siRNA blocked the miR-361-3p inhibitor-mediated upregulation of FSH, while no significant effect on non-target expression. Taken together, our results demonstrate that miR-361-3p negatively regulates FSH synthesis and secretion by targeting FSHB, which provides more functional evidence that a miRNA is involved in the direct regulation of FSH.

Revelation of mRNAs and proteins in porcine milk exosomes by transcriptomic and proteomic analysis.

BACKGROUND: Milk is a complex liquid that provides nutrition to newborns. Recent reports demonstrated that milk is enriched in maternal-derived exosomes that are involved in fetal physiological and pathological conditions by transmission of exosomal mRNAs, miRNAs and proteins. Until now, there is no such research relevant to exosomal mRNAs and proteins in porcine milk, therefore, we have attempted to investigate porcine milk exosomal mRNAs and proteins using RNA-sequencing and proteomic analysis. RESULTS: A total of 16,304 (13,895 known and 2,409 novel mRNAs) mRNAs and 639 (571 known, 66 candidate and 2 putative proteins) proteins were identified. GO and KEGG annotation indicated that most proteins were located in the cytoplasm and participated in many immunity and disease-related pathways, and some mRNAs were closely related to metabolisms, degradation and signaling pathways. Interestingly, 19 categories of proteins were tissue-specific and detected in placenta, liver, milk, plasma and mammary. COG analysis divided the identified mRNAs and proteins into 6 and 23 categories, respectively, 18 mRNAs and 10 proteins appeared to be involved in cell cycle control, cell division and chromosome partitioning. Additionally, 14 selected mRNAs were identified by qPCR, meanwhile, 10 proteins related to immunity and cell proliferation were detected by Western blot. CONCLUSIONS: These results provide the first insight into porcine milk exosomal mRNA and proteins, and will facilitate further research into the physiological significance of milk exosomes for infants.

miR-146a-5p inhibits TNF-α-induced adipogenesis via targeting insulin receptor in primary porcine adipocytes.

TNF-α is a multifunctional cytokine participating in immune disorders, inflammation, and tumor development with regulatory effects on energy metabolism. Our work focused on the function of TNF-α in adipogenesis of primary porcine adipocytes. TNF-α could suppress the insulin receptor (IR) at the mRNA and protein levels. Microarray analysis of TNF-α-treated porcine adipocytes was used to screen out 29 differentially expressed microRNAs (miRNAs), 13 of which were remarkably upregulated and 16 were intensely downregulated. These 29 differentially expressed miRNAs were predicted to mainly participate in the insulin signaling pathway, adipocytokine signaling pathway, and type 2 diabetes mellitus pathway by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses. miR-146a-5p, reportedly involved in immunity and cancer relevant processes, was one of the most highly differentially expressed miRNAs after TNF-α treatment. Red Oil O staining and TG assay revealed that miR-146a-5p suppressed adipogenesis. A dual-luciferase reporter and siRNA assay verified that miR-146a-5p targeted IR and could inhibit its protein expression. miR-146a-5p was also validated to be involved in the insulin signaling pathway by reducing tyrosine phosphorylation of insulin receptor substrate-1. Our study provides the first evidence of miR-146a-5p targeting IR, which facilitates future studies related to obesity and diabetes using pig models.

DDQ-Mediated Three-Component Dioxygenation of Alkenes.

A new DDQ-mediated three-component dioxygenation of alkenes has been established, providing a direct and metal-free access toward densely functionalized 4,5-dichloro-3-hydroxyphthalonitrile derivatives with generally good to excellent yields under mild conditions. During this process, DDQ plays dual roles as both a dehydrogenation reagent and a coupling partner, enabling oxidative coupling to form two C-O functionalities in a highly atom-economy fashion.

Genome-wide discovery of novel and conserved microRNAs in white shrimp (Litopenaeus vannamei).

Of late years, a large amount of conserved and species-specific microRNAs (miRNAs) have been performed on identification from species which are economically important but lack a full genome sequence. In this study, Solexa deep sequencing and cross-species miRNA microarray were used to detect miRNAs in white shrimp. We identified 239 conserved miRNAs, 14 miRNA* sequences and 20 novel miRNAs by bioinformatics analysis from 7,561,406 high-quality reads representing 325,370 distinct sequences. The all 20 novel miRNAs were species-specific in white shrimp and not homologous in other species. Using the conserved miRNAs from the miRBase database as a query set to search for homologs from shrimp expressed sequence tags (ESTs), 32 conserved computationally predicted miRNAs were discovered in shrimp. In addition, using microarray analysis in the shrimp fed with Panax ginseng polysaccharide complex, 151 conserved miRNAs were identified, 18 of which were significant up-expression, while 49 miRNAs were significant down-expression. In particular, qRT-PCR analysis was also performed for nine miRNAs in three shrimp tissues such as muscle, gill and hepatopancreas. Results showed that these miRNAs expression are tissue specific. Combining results of the three methods, we detected 20 novel and 394 conserved miRNAs. Verification with quantitative reverse transcription (qRT-PCR) and Northern blot showed a high confidentiality of data. The study provides the first comprehensive specific miRNA profile of white shrimp, which includes useful information for future investigations into the function of miRNAs in regulation of shrimp development and immunology.

Exploration of microRNAs in porcine milk exosomes.

BACKGROUND: Breast milk contains complex nutrients and facilitates the maturation of various biological systems in infants. Exosomes, membranous vesicles of endocytic origin found in different body fluids such as milk, can mediate intercellular communication. We hypothesized that microRNAs (miRNAs), a class of non-coding small RNAs of 18-25 nt which are known to be packaged in exosomes of human, bovine and porcine milk, may play important roles in the development of piglets. RESULTS: In this study, exosomes of approximately 100 nm in diameter were isolated from porcine milk through serial centrifugation and ultracentrifugation procedures. Total RNA was extracted from exosomes, and 5S ribosomal RNA was found to be the major RNA component. Solexa sequencing showed a total of 491 miRNAs, including 176 known miRNAs and 315 novel mature miRNAs (representing 366 pre-miRNAs), which were distributed among 30 clusters and 35 families, and two predicted novel miRNAs were verified targeting 3'UTR of IGF-1R by luciferase assay. Interestingly, we observed that three miRNAs (ssc-let-7e, ssc-miR-27a, and ssc-miR-30a) could be generated from miRNA-offset RNAs (moRNAs). The top 10 miRNAs accounted for 74.5% (67,154 counts) of total counts, which were predicted to target 2,333 genes by RNAhybrid software. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses using DAVID bioinformatics resources indicated that the identified miRNAs targeted genes enriched in transcription, immunity and metabolism processes, and 14 of the top 20 miRNAs possibly participate in regulation of the IgA immune network. CONCLUSIONS: Our findings suggest that porcine milk exosomes contain a large number of miRNAs, which potentially play an important role in information transfer from sow milk to piglets. The predicted miRNAs of porcine milk exosomes in this study provide a basis for future biochemical and biophysical function studies.

Differential gene expression pattern in hypothalamus of chickens during fasting-induced metabolic reprogramming: functions of glucose and lipid metabolism in the feed intake of chickens.

Fasting-induced hypothalamic metabolic reprogramming is involved in regulating energy homeostasis and appetite in mammals, but this phenomenon remains unclear in poultry. In this study, the expression patterns of a panel of genes related to neuropeptides, glucose, and lipid metabolism enzymes in the hypothalamus of chickens during fasting and refeeding were characterized by microarray analysis and quantitative PCR. Results showed that 48 h of fasting upregulated (P < 0.05) the mRNA expressions of orexigenic neuropeptide Y and agouti-related protein but downregulated (P < 0.05) that of anorexigenic neuropeptide pro-opiomelanocortin; growth hormone-releasing hormone; islet amyloid polypeptide; thyroid-stimulating hormone, ß; and glycoprotein hormones, α polypeptide. After 48 h of fasting, the mRNA expression of fatty acid ß-oxidation [peroxisome proliferator-activated receptor α (PPARα), carnitine palmitoyltransferase 1A, and forkhead box O1], energy sensor protein [sirtuin 1 (SIRT1) and forkhead box O1], and glycolysis inhibitor (pyruvate dehydrogenase kinase, isozyme 4) were enhanced, but that of fatty acid synthesis and transport associated genes (acetyl-CoA carboxylase α, fatty acid synthase, apolipoprotein A-I, endothelial lipase, and fatty acid binding protein 7) were suppressed. Liver and muscle also demonstrated similar expression patterns of genes related to glucose and lipid metabolism with hypothalamus, except for that of acetyl-CoA carboxylase α, acyl-CoA synthetase long-chain family member 4, and apolipoprotein A-I. The results of intracerebroventricular (ICV) injection experiments confirmed that α-lipoic acid (ALA, pyruvate dehydrogenase kinase, isozyme 4 inhibitor, 0.10 µmol) and NADH (SIRT1 inhibitor, 0.80 µmol) significantly suppressed the appetite of chickens, whereas 2-deoxy-d-glucose (glycolytic inhibitor, 0.12 to 1.20 µmol) and NAD(+) (SIRT1 activator, 0.08 to 0.80 µmol) increased feed intake in chickens. The orexigenic effect of NAD(+) was also blocked by cotreatment with NADH. However, ICV injection of either GW7647 (PPARα agonist) or GW6471 (PPARα antagonist) showed no effects on feed intake. Results suggested that hypothalamic glycolysis (inhibited by ALA and promoted by 2-deoxy-d-glucose) and SIRT1 (inhibited by NADH and promoted by NAD(+)), not PPARα, were probably involved in feed intake regulation in chickens.

Roles of α-linolenic acid on IGF-I secretion and GH/IGF system gene expression in porcine primary hepatocytes.

The main purposes of this study were to investigate the effects of α-linolenic acid (ALA) on the insulin-like growth factor (IGF) system of porcine primary hepatocytes with or without growth hormone (GH) or insulin and the potential role of peroxisome proliferator-activated receptor α and -γ (PPARα/γ) pathway. We found that 1 µM ALA increased IGF-I secretion from hepatocytes at 48 and 72 h. Expression of hepatocytes IGF-I, IGF-II, GH receptor (GHR), insulin receptor (IR), IGF-binding protein 3 (IGFBP3), and IGFBP4 mRNAs was up-regulated by ALA treatment. GH (15 nM) alone or co-treated with ALA increased hepatocytes IGF-I secretion and the expression of GHR and IGFBP1 mRNAs, but down-regulated IGFBP5 mRNA compared with appropriate control across ALA. GH also enhanced the ALA-induced increase in the transcript levels of IGF-II and GHR, but tended to attenuate that of IGFBP4. Insulin (1 µM) alone or co-treated with ALA improved IGF-I secretion and the expression of IGFBP3 mRNA, but decreased IGFBP1 mRNA versus appropriate control across ALA. Insulin also up-regulated the expression of GHR, IR, IGFBP3, and IGFBP4 mRNAs, and tended to prevent the transcript levels of IGF-I and IGFBP4 improved by ALA. Both PPARγ agonist rosiglitazone and its antagonist GW9662 could elevated the IGF-I secretion in dose-dependent manner but they had no interaction with ALA. However, GW7647, a PPARα agonist, increased IGF-I secretion dose-dependently, but the antagonist GW6471 was without effect. Moreover, GW6471 prevented the IGF-I promoting effect of ALA. This suggests that the IGF-I promoting effect of ALA may be mediated by the PPARα pathway.

Identification and comparison of microRNAs from skeletal muscle and adipose tissues from two porcine breeds.

MicroRNAs (miRNAs) are an abundant class of small regulatory RNAs that negatively regulate gene expression at the post-transcriptional level. Although an increasing number of porcine miRNAs recently have been identified, research has yet to identify the full repertoire of miRNAs in pig skeletal and adipose tissues and their differences between breeds. We extracted small RNA from skeletal muscle and adipose tissues of Landrace and Lantang pigs, and the expression of a total of 184 known porcine miRNAs (113 from Solexa sequencing and 171 from miRNA chip hybridization) as well as 521 novel miRNA candidates was detected. Moreover, 20 miRNAs were selected randomly from the 184 miRNAs and analysed by quantitative real-time PCR to confirm the aforementioned results. In the skeletal muscle tissues, 21 miRNAs were up-regulated in Lantang and another 33 were highly expressed in Landrace pigs. In the adipose tissues, 25 miRNAs were down-regulated in Lantang and another 23 were lowly expressed in Landrace pigs. miRNA divergence between tissues was also detected in this study. Ten miRNAs were highly expressed in the skeletal muscle tissue in comparison with adipose tissue, and another 10 miRNAs exhibited the opposite expression profile. To investigate the regulatory mechanism of the miRNAs in muscle and adipose tissues, the 10 miRNAs with the most divergent expression profiles were functionally categorized using the Kyoto Encyclopedia of Genes and Genomes database. Most of the miRNAs strongly corresponded to myogenesis and adipogenesis processes. In addition, 84 of the 521 miRNA candidates were potentially porcine-specific miRNAs. This study adds new valuable information to comparative miRNA profiles of skeletal muscle and adipose tissues in porcine species. The great diversity of miRNA composition and expression levels both between breeds and between tissues suggests that a complex regulatory network exists in porcine subcutaneous fat development.

Osteoarticular Involvement-Associated Biomarkers and Pathways in Psoriasis: The Shared Pathway With Ankylosing Spondylitis.

Psoriatic arthritis (PsA) is a unique immune-mediated disease with cutaneous and osteoarticular involvement. However, only a few studies have explored the susceptibility of osteoarticular involvement in psoriasis (Ps) at the genetic level. This study investigated the biomarkers associated with osteoarticular participation and potential shared molecular mechanisms for PsA and ankylosing spondylitis (AS). Methods: The RNA-seq data of Ps, PsA, and AS in the Gene Expression Omnibus (GEO) database were obtained. First, we used the limma package and the weighted gene co-expression network analysis (WGCNA) to identify the potential genes related to PsA and AS. Then, the shared genes in PsA and AS were performed using the GO, KEGG, and GSEA analyses. We also used machine learning to screen hub genes. The results were validated using external datasets and native cohorts. Finally, we used the CIBERSORT algorithm to estimate the correlation between hub genes and the abundance of immune cells in tissues. Results: An overlap was observed between the PsA and AS-related modules as 9 genes. For differentially expressed genes in AS and PsA, only one overlapping gene was found (COX7B). Gene enrichment analysis showed that the above 9 genes might be related to the mRNA surveillance pathway. The GSEA analyses showed that COX7B was involved in adaptive immune response, cell activation, etc. The PUM1 and ZFP91, identified from the support vector machine, had preferable values as diagnostic markers for osteoarticular involvement in Ps and AS (AUC > 0.7). Finally, CIBERSORT results showed PUM1 and ZFP91 involvement in changes of the immune microenvironment. Conclusion: For the first time, this study showed that the osteoarticular involvement in psoriasis and AS could be mediated by the mRNA surveillance pathway-mediated abnormal immunologic process. The biological processes may represent the cross talk between PsA and AS. Therefore, PUM1 and ZFP91 could be used as potential biomarkers or therapeutic targets for AS and Ps patients.

The effect of dietary Panax ginseng polysaccharide extract on the immune responses in white shrimp, Litopenaeus vannamei.

The immunostimulatory effects of orally administered Panax ginseng root or its polysaccharides (GSP) in white shrimp, Litopenaeus vannamei, were investigated in this study. Shrimp were fed a diet containing 0.4 g kg⁻¹ GSP over a period of 84 days, during which the activities of total superoxide dismutase (T-SOD), catalase (CAT), glutathione peroxidase (GSH-Px), acid phosphatase (ACP), and alkaline phosphatase (AKP), as well as malondialdehyde (MDA) content, and expressions of cytosolic superoxide dismutase (cyt-SOD), CAT, GSH-Px, and peroxiredoxin (Prx) genes were determined in various tissues of the shrimp. Results showed that the shrimp fed the GSP diet had significantly increased ACP and AKP activities in the gills. The GSP-fed shrimp also displayed significantly increased T-SOD and GSH-Px activities in the gills and hepatopancreas of the shrimp; meanwhile there was enhanced CAT activity in the gills, but decreased MDA content in the gills, hepatopancreas and muscle. The mRNA expressions of cyt-SOD, CAT, GSH-Px and Prx were significantly elevated in the gills and hepatopancreas of the shrimp fed the GSP diet for 84 days, compared with that of the control. Therefore, GSP can be used as an immunostimulant for shrimp through dietary administration to increase immune enzyme activity and modify expression of immune genes in shrimp.

2,6-Diphenyl-4-(2-thien-yl)-1,4-dihydro-pyridine-3,5-dicarbonitrile.

The asymmetric unit of the title compound, C(23)H(15)N(3)S, contains two crystallographically independent mol-ecules. The pyridine rings adopt envelope conformations. The thio-phene rings are oriented at dihedral angles of 77.97 (4)/53.53 (4) and 78.44 (4)/57.11 (4)° with respect to the phenyl rings, while the dihedral angles between the phenyl rings are 48.51 (4) and 44.49 (4)°. In the crystal structure, inter-molecular N-H⋯N hydrogen bonds link the mol-ecules into chains along the c axis. The S, C and H atoms of one of the thio-phene rings are disordered over two orientations, with occupancy ratios of 0.314 (15):0.686 (15).

2-Amino-5,7-bis-(4-fluoro-phen-yl)-1',3'-dimethyl-7,8-dihydro-spiro-[pyrido[2,3-d]pyrimidine-6(5H),5'-pyrimidine]-2',4,4',6'(3H,1'H,3'H,5'H)-tetra-one ethanol solvate.

In the mol-ecule of the title compound, C(24)H(20)F(2)N(6)O(4)·C(2)H(5)OH, the pyrimidine ring is oriented at dihedral angles of 42.64 (3) and 62.94 (3)° with respect to the benzene rings, while the dihedral angle between the benzene rings is 74.45 (3)°. The pyridine ring adopts an envelope conformation. In the crystal structure, inter-molecular N-H⋯O and O-H⋯N hydrogen bonds link the mol-ecules into a two-dimensional network, forming R(2) (2)(8) ring motifs. π-π contacts between the pyrimidine and benzene rings [centroid-centroid distances = 3.516 (1) and 3.927 (1) Å] may further stabilize the structure.

Kirenol Inhibits the Function and Inflammation of Fibroblast-like Synoviocytes in Rheumatoid Arthritis in vitro and in vivo.

Kirenol is a diterpenoid extracted from the Chinese herbal medicine Siegesbeckiae. Siegesbeckiae has been used to treat Rheumatoid arthritis (RA) in China for several centuries. RA is characterized by the proliferation of synoviocytes in inflamed synovia, as well as by their expression of inflammatory cytokines. In the present study, we found that Kirenol inhibited the migration, invasion, and proinflammatory of IL-6 secretion of RA-associated synovial fibroblasts (FLS) at a concentration of 100-200 µg/ml in vitro. Proinflammatory cytokines production and synovium hyperplasia and cartilage erosion were also inhibited in a collagen-induced arthritis (CIA) mouse model upon Kirenol treatment. Together, our results thus confirm that Kirenol has potent therapeutic efficacy in RA owing to its ability to suppress negative FLS activities.

Obestatin: its physicochemical characteristics and physiological functions.

Obestatin, a novel 23 amino acid amidated peptide encoded by the same gene with ghrelin, was initially reported to reduce food intake, body weight gain, gastric emptying and suppress intestinal motility through an interaction with the orphan receptor GPR39. However, recently reports have shown that above findings had been questioned by several groups. Further studies explained that obestatin was involved in inhibiting thirst and anxiety, improving memory, regulating sleep, affecting cell proliferation, and increasing the secretion of pancreatic juice enzymes. We also identified that obestatin could stimulate piglet liver and adipose cell proliferation, and inhibit the secretion of IGF-I. According to the controversy over the effects and the cognate ligand of obestatin, here we provide the latest review on the structure, distribution and physiological functions of obestatin.