Application and limitations of the methyl imidate protection strategy of N-acetylglucosamine for glycosylations at O-4: synthesis of Lewis A and Lewis X trisaccharide analogues

We describe here the synthesis of the allyl Le^a trisaccharide antigen as well as that of an analogue of the Le^x trisaccharide antigen, in which the galactose residue has been replaced by a glucose unit. Although successful fucosylations at O-4 of N-acetylglucosamine acceptors have been reported using perbenzylated thioethyl fucosyl donors under MeOTf activation, such conditions led in our case to the conversion of our acceptor to the corresponding alkyl imidates. Indeed, in this synthesis of the Le^a analogue, we demonstrate that the temporary protection of the N-acetyl group as a methyl imidate is advantageous to fucosylate at O-4. In contrast, we report here that glucosylation at O-4 of an N-acetylglucosamine monosaccharide acceptor using the α-trichloroacetimidate of peracetylated glucopyranose as a donor proceeded in better yields under activation with excess BF₃·OEt₂ than that of the corresponding methyl imidate. Therefore, we conclude that activation of thioglycoside donors by MeOTf to glycosylate at O-4 of a glucosamine acceptor is best accomplished following the temporary protection of the N-acetyl group as a methyl imidate, especially when the donors are highly reactive and prone to degradation. In contrast, if donor and acceptor can withstand multiple equivalents of BF₃·OEt₂, glycosylations at O-4 of a glucosamine acceptor with a trichloroacetimidate donor does not benefit from the temporary protection of the N-acetyl group as a methyl imidate.     

Selective separation of microorganisms by lectins: yeast and concanavalin A as a model system

Specific aggregation and separation of microorganisms was investigated using yeasts and concanavalin A as a model system. Cells of Saccharomyces cerevisiae were specifically aggregated and so separated from those of Schizosaccharomyces pombe. Optimum aggregation with over 99% of cells aggregated was achieved by adjustment to pH value and applied agitation. Dimeric lectin structure caused a far higher degree of aggregation than did tetrameric. Degree of aggregation was also strongly influenced by the ratio of lectin/cell concentrations, optimum aggregation occurring in the middle range of ratios. A high ratio of lectin to cells inhibited aggregation, occupation of most of the available receptors preventing intercellular bonding by divalent lectins. Detachment and reuse of concanavalin A was demonstrated using switching from moderate to low pH value. Potential uses for species-specific-separation of microorganisms are discussed. (c) 1992 John Wiley & Sons, Inc.     

Novel Ca2+‐independent carbohydrate recognition of the C‐type lectins,SPL‐1 and SPL‐2, from the bivalve Saxidomus purpuratus

Novel Ca²⁺‐independent C‐type lectins, SPL‐1 and SPL‐2, were purified from the bivalve Saxidomus purpuratus. They are composed of dimers with either identical (SPL‐2 composed of two B‐chains) or distinct (SPL‐1 composed of A‐ and B‐chains) polypeptide chains, and show affinity for N‐acetylglucosamine (GlcNAc)‐ and N‐acetylgalactosamine (GalNAc)‐containing carbohydrates, but not for glucose or galactose. A database search for sequence similarity suggested that they belong to the C‐type lectin family. X‐ray crystallographic analysis revealed definite structural similarities between their subunits and the carbohydrate‐recognition domain (CRD) of the C‐type lectin family. Nevertheless, these lectins (especially SPL‐2) showed Ca²⁺‐independent binding affinity for GlcNAc and GalNAc. The crystal structure of SPL‐2/GalNAc complex revealed that bound GalNAc was mainly recognized via its acetamido group through stacking interactions with Tyr and His residues and hydrogen bonds with Asp and Asn residues, while widely known carbohydrate‐recognition motifs among the C‐type CRD (the QPD [Gln‐Pro‐Asp] and EPN [Glu‐Pro‐Asn] sequences) are not involved in the binding of the carbohydrate. Carbohydrate‐binding specificities of individual A‐ and B‐chains were examined by glycan array analysis using recombinant lectins produced from Escherichia coli cells, where both subunits preferably bound oligosaccharides having terminal GlcNAc or GalNAc with α‐glycosidic linkages with slightly different specificities.     

6-<Emphasis Type="Italic">O</Emphasis>-Sulfo sialylparagloboside and sialyl Lewis X neo-glycolipids containing lactamized neuraminic acid: Synthesis and antigenic reactivity against G159 monoclonal antibody

Synthesis and antigenic reactivity of 6-O-sulfo sialylparagloboside (SPG) and sialyl Lewis X (sLe^X) neo-glycolipids containing lactamized neuraminic acid are described. The suitably protected GlcNAc-β (1 → 3)-Gal-β (1 → 4)-GlcOSE derivative was glycosylated with NeuTFAc-α (2 → 3)-Gal imidate to give NeuTFAc-α (2 → 3)-Galβ (1 → 4)-GlcNAc-β (1 → 3)-Gal-β (1 → 4)-GlcOSE pentasaccharide. The partial N,O-deacylation in the NeuTFAc-α (2→3)-Gal part afforded N-deacetylated SPG derivative which was converted to the desired oligosaccharide containing lactamized neuraminic acid. Similar treatment of the sLe^X hexasaccharide derivative, NeuTFAc-α (2 → 3)-Gal-β (1 → 4) [Fuc-α (1 →3)]-GlcNAc-β (1 → 3)-Gal-β (1 → 4)-GlcOSE, gave the key hexasaccharide intermediate containing lactamized neuraminic acid. These suitably protected SPG and sLe^x oligosaccharides were converted stepwise into the desired neo-glycolipids (GSC-551 and GSC-552) by the coupling with 2-(tetradecyl)hexadecanol, 6-O-sulfation at C-6 of the GlcNAc residure, and complete deprotection.Both lactamized-sialyl 6-O-sulfo SPG (GSC-551) and sLe^x (GSC-552) neo-glycolipids were clearly recognized with G159 monoclonal antibody showing that both the lactamized neuraminic acid and the 6-O-sulfate at C-6 of GlcNAc would be involved in the G159-defined determinant. However, the Fuc residue and the lipophilic (ceramide) part may not be critical for this recognition. Published in 2005Synthetic studies on sialoglycoconjugates, Part 138. For part 136, see Ref [1], and for part 137, see Ref [19].     

The N-glycolyl form of mouse sialyl Lewis X is recognized by selectins but not by HECA-452 and FH6 antibodies that were raised against human cells

E-, P- and L-selectins critically function in lymphocyte recirculation and recruiting leukocytes to inflammatory sites. MECA-79 antibody inhibits L-selectin-mediated lymphocyte adhesion in several species and does not require sialic acid in its epitope. Many other antibodies, however, recognize human selectin ligands expressing N-acetylneuraminic acid but not mouse selectin ligands expressing N-glycolylneuraminic acid, suggesting that difference in sialic acid in sialyl Lewis X leads to differential reactivity. We found that HECA-452 and FH6 monoclonal antibodies bind Chinese hamster ovary (CHO) cells expressing N-acetylneuraminyl Lewis X oligosaccharide but not its N-glycolyl form. Moreover, synthetic N-acetylneuraminyl Lewis X oligosaccharide but not its N-glycolyl oligosaccharide inhibited HECA-452 and FH6 binding. By contrast, E-, P- and L-selectin bound to CHO cells regardless of whether they express N-acetyl or N-glycolyl form of sialyl Lewis X, showing that selectins have a broader recognition capacity than HECA-452 and FH-6 anti-sialyl Lewis x antibodies. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Prediction of antiviral peptides derived from viral fusion proteins potentially active against herpes simplex and influenza A viruses

There are very few antiviral drugs available to fight viral infections and the appearance of viral strains resistant to these antiviralsis not a rare event. Hence, the design of new antiviral drugs is important. We describe the prediction of peptides with antiviralactivity (AVP) derived from the viral glycoproteins involved in the entrance of herpes simplex (HSV) and influenza A viruses intotheir host cells. It is known, that during this event viral glycoproteins suffer several conformational changes due to protein-proteininteractions, which lead to membrane fusion between the viral envelope and the cellular membrane. Our hypothesis is that AVPscan be derived from these viral glycoproteins, specifically from regions highly conserved in amino acid sequences, which at thesame time have the physicochemical properties of being highly exposed (antigenic), hydrophilic, flexible, and charged, since theseproperties are important for protein-protein interactions. For that, we separately analyzed the HSV glycoprotein H and B, andinfluenza A viruses hemagglutinin (HA), using several bioinformatics tools. A set of multiple alignments was carried out, to findthe most conserved regions in the amino acid sequences. Then, the physicochemical properties indicated above were analyzed. Wepredicted several peptides 12-20 amino acid length which by docking analysis were able to interact with the fusion viralglycoproteins and thus may prevent conformational changes in them, blocking the viral infection. Our strategy to design AVPsseems to be very promising since the peptides were synthetized and their antiviral activities have produced very encouragingresults.     

Matrilysin (MMP‐7) cleaves C‐type lectin domain family 3 member A (CLEC3A) on tumor cell surface and modulates its cell adhesion activity

Matrilysin (MMP‐7) plays important roles in tumor progression. Previous studies have suggested that MMP‐7 binds to tumor cell surface and promotes their metastatic potential. In this study, we identified C‐type lectin domain family 3 member A (CLEC3A) as a membrane‐bound substrate of MMP‐7. Although this protein is known to be expressed specifically in cartilage, its message was found in normal breast and breast cancer tissues as well as breast and colon cancer cell lines. Because few studies have been done on CLEC3A, we overexpressed its recombinant protein in human cancer cells. CLEC3A was found in the cell membrane, extracellular matrix (ECM), and culture medium of the CLEC3A‐expressing cells. CLEC3A has a basic sequence in the NH₂‐terminal domain and showed a strong heparin‐binding activity. MMP‐7 cleaved the 20‐kDa CLEC3A protein, dividing it to a 15‐kDa COOH‐terminal fragment and an NH₂‐terminal fragment with the basic sequence. The 15‐kDa fragment no longer had heparin‐binding activity. Treatment of the CLEC3A‐expressing cells with MMP‐7 released the 15‐kDa CLEC3A into the culture supernatant. Furthermore, the 20‐kDa CLEC3A promoted cell adhesion to laminin‐332 and fibronectin substrates, but this activity was abrogated by the cleavage by MMP‐7. These results suggest that CLEC3A binds to heparan sulfate proteoglycans on cell surface, leading to the enhancement of cell adhesion to integrin ligands on ECM. It can be speculated that the cleavage of CLEC3A by MMP‐7 weakens the stable adhesion of tumor cells to the matrix and promotes their migration in tumor microenvironments. J. Cell. Biochem. 106: 693–702, 2009. © 2009 Wiley‐Liss, Inc.     

Disorder and order in unfolded and disordered peptides and proteins: A view derived from tripeptide conformational analysis. I. Tripeptides with long and predominantly hydrophobic side chains

We performed a conformational analysis of the central residues of three tripeptides glycyl‐L ‐isoleucyl‐glycine (GIG), glycyl‐L ‐tyrosyl‐glycine (GYG) and glycyl‐L ‐arginyl‐glycine (GRG) in aqueous solution, based on a global analysis of amide I′ band profiles and NMR J‐coupling constants. The results are compared with recently reported distributions of GVG, GFG and GEG. For GIG and GYG, we found that even though the polyproline II (pPII) fraction is below 0.5, it is still the most populated conformation, whereas GVG and GFG show both a larger β‐strand fraction. For GRG, we observed a clear dominance of pPII over β‐strand, reminiscent of observations for GEG and GKG. This finding indicates that terminal charges on otherwise hydrophobic residue side chains stabilize pPII over β‐strand conformations. For all peptides investigated we found that a variety of compact and turn‐like conformations constitute nearly 20 percent of their conformational distributions. Attempts to analyze our data with a simple two‐state pPII??β model therefore do not yield any satisfactory reproduction of experimental results. A comparison of the obtained GxG ensembles with conformational distributions of GxG segments in truncated coil libraries (helices and sheets omitted) revealed a much larger fraction of type II β_i+2 and type III β like conformations for the latter. Thus, a comparison of conformational distributions of unfolded peptide segments in solution and in coil libraries reveal interesting information on how the interplay between intrinsic propensities of amino acid residues and non‐local interactions in polypeptide chains determine the conformations of loop segments in proteins. Proteins 2013; © 2012 Wiley Periodicals, Inc.     

Antiviral activity of Basidiomycete mycelia against influenza type A(serotype H1N1) and herpes simplex virus type 2 in cell culture

In this study, we investigated the in vitro antiviral activity of the mycelia of higher mushrooms against influenza virus type A(serotype H1N1) and herpes simplex virus type 2(HSV-2), strain BH. All 10 investigated mushroom species inhibited the reproduction of influenza virus strain A/FM/1/47(H1N1) in MDCK cells reducing the infectious titer by 2.0–6.0 lg ID50. Four species, Pleurotus ostreatus, Fomes fomentarius, Auriporia aurea, and Trametes versicolor, were also determined to be effective against HSV-2 strain BH in RK-13 cells, with similar levels of inhibition as for influenza. For some of the investigated mushroom species—Pleurotus eryngii, Lyophyllum shimeji, and Flammulina velutipes—this is the first report of an anti-influenza effect. This study also reports the first data on the medicinal properties of A. aurea, including anti-influenza and antiherpetic activities. T. versicolor 353 mycelium was found to have a high therapeutic index(324.67), and may be a promising material for the pharmaceutical industry as an anti-influenza and antiherpetic agent with low toxicity. Mycelia with antiviral activity were obtained in our investigation by bioconversion of agricultural wastes(amaranth flour after CO2 extraction), which would reduce the cost of the final product and solve some ecological problems.     

A类Ⅰ型清道夫受体调节小鼠腹腔巨噬细胞抗肺炎克雷伯菌和铜绿假单胞菌感染

本研究旨在探讨A类Ⅰ型清道夫受体(scavenger receptor class A type Ⅰ,SR-AI)在呼吸道感染常见病原体肺炎克雷伯菌及铜绿假单胞菌感染过程中的免疫调节功能。以肺炎克雷伯菌和铜绿假单胞菌临床分离株与野生型小鼠和SR-AI~(-/-)小鼠腹腔原代巨噬细胞互作,研究SR-AI在吞噬和炎症反应中的作用。荧光染料染色菌体及检测胞内荧光强度,数据显示SR-AI敲除后巨噬细胞对肺炎克雷伯菌的吞噬能力下降,但对铜绿假单胞菌的吞噬能力升高。采用实时定量荧光聚合酶链反应检测相关炎症因子mRNA水平,发现SRAI敲除后肺炎克雷伯菌和铜绿假单胞菌刺激巨噬细胞引发的炎症反应均增强。结果表明,SR-AI参与巨噬细胞对肺炎克雷伯菌的吞噬,但不参与对铜绿假单胞菌的吞噬,且可能抑制了肺炎克雷伯菌和铜绿假单胞菌引发的炎症反应。     

Cell expression of GDAP1 in the nervous system and pathogenesis of Charcot-Marie-Tooth type 4A disease

Mutations in the mitochondrial protein GDAP1 are the cause of Charcot-Marie-Tooth type 4A disease (CMT4A), a severe form of peripheral neuropathy associated with either demyelinating, axonal or intermediate phenotypes. GDAP1 is located in the outer mitochondrial membrane and it seems that may be related with the mitochondrial network dynamics. We are interested to define cell expression in the nervous system and the effect of mutations in mitochondrial morphology and pathogenesis of the disease. We investigated GDAP1 expression in the nervous system and dorsal root ganglia (DRG) neuron cultures. GDAP1 is expressed in motor and sensory neurons of the spinal cord and other large neurons such as cerebellar Purkinje neurons, hippocampal pyramidal neurons, mitral neurons of the olfactory bulb and cortical pyramidal neurons. The lack of GDAP1 staining in the white matter and nerve roots suggested that glial cells do not express GDAP1. In DRG cultures satellite cells and Schwann cells were GDAP1-negative. Overexpression of GDAP1-induced fragmentation of mitochondria suggesting a role of GDAP1 in the fission pathway of the mitochondrial dynamics. Missense mutations showed two different patterns: most of them induced mitochondrial fragmentation but the T157P mutation showed an aggregation pattern. Whereas null mutations of GDAP1 should be associated with loss of function of the protein, missense mutations may act through different pathogenic mechanisms including a dominant-negative effect, suggesting that different molecular mechanisms may underlay the pathogenesis of CMT4A.     

Antiviral activity of Basidiomycete mycelia against influenza type A (serotype H1N1) and herpes simplex virus type 2 in cell culture

In this study, we investigated the in vitro antiviral activity of the mycelia of higher mushrooms against influenza virus type A(serotype H1N1) and herpes simplex virus type 2(HSV-2), strain BH. All 10 investigated mushroom species inhibited the reproduction of influenza virus strain A/FM/1/47(H1N1) in MDCK cells reducing the infectious titer by 2.0–6.0 lg ID50. Four species, Pleurotus ostreatus, Fomes fomentarius, Auriporia aurea, and Trametes versicolor, were also determined to be effective against HSV-2 strain BH in RK-13 cells, with similar levels of inhibition as for influenza. For some of the investigated mushroom species—Pleurotus eryngii, Lyophyllum shimeji, and Flammulina velutipes—this is the first report of an anti-influenza effect. This study also reports the first data on the medicinal properties of A. aurea, including anti-influenza and antiherpetic activities. T. versicolor 353 mycelium was found to have a high therapeutic index(324.67), and may be a promising material for the pharmaceutical industry as an anti-influenza and antiherpetic agent with low toxicity. Mycelia with antiviral activity were obtained in our investigation by bioconversion of agricultural wastes(amaranth flour after CO2 extraction), which would reduce the cost of the final product and solve some ecological problems.     

Monoclonal antibodies specific for type 3 and type 4 chain-based blood group determinants: Relationship to the A1 and A2 subgroups

Two monoclonal antibodies, specific for A type 3 and A type 4 blood group determinants, are described. These antibodies recognized A1 but not A2 erythrocytes. A third monoclonal antibody showing specificity for A type 3 and A type 4, and also for H type 3 and H type 4, did not discriminate between A1 and A2 erythrocytes. On red cells these three antibodies recognized glycosphingolipids and binding to glycoproteins could not be demonstrated. On paraffin-embedded tissue sections the three antibodies labelled a supranuclear area, characteristic of the Golgi apparatus, of all cells producing A antigens. This labelling occurred irrespective of the A1, A2 status.The results suggest that glycolipids of erythrocytes and possibly of other cell types bear the A type 3/4 determinant specific for the A1 subgroup and that A type 3/4 determinants of glycoproteins might be present in both A1 and A2 subgroups on short oligosaccharide chains which are only detectable at the level of the Golgi apparatus.     

Binding and precipitating activities ofErythrina lectins with complex type carbohydrates and synthetic cluster glycosides. A comparative study of the lectins fromE. corallodendron,E. cristagalli,E. flabelliformis,andE. indica

Erythrina lectins possess similar structural and carbohydrate binding properties. Recently, tri- and tetra-antennary complex type carbohydrates with non-reducing terminal galactose residues have been shown to be precipitated as tri- and tetravalent ligands, respectively, with certainErythrina lectins [Bhattacharyya L, Haraldsson M, Brewer CF (1988) Biochemistry 271034-41]. The present work describes a comparative study of the binding and precipitating activities of fourErythrina lectins,viz. E. corallodendron, E. cristagalli, E. flabelliformis, andE. indica, with multi-antennary complex type carbohydrates and synthetic cluster glycosides. The results show that though their binding affinities are very similar, theErythrina lectins show large differences in their precipitating activities with the carbohydrates. The results also indicate significant dependence of the precipitating activities of the lectins on the core structure of the carbohydrates. These findings provide a new dimension to the structure-activity relationship of the lectins and their interactions with asparagine-linked carbohydrates.Abbreviations EAL, ECorL, ECL, EFL, and EIL represent the lectins from the seeds ofErythrina arborescens, - E. corallodendron, E. cristagalli, E. flabelliformis, andE. indica respectively - AFOS thetri-antennary complex type oligosaccharide from asialofetuin - AFGP the tri-antennary glycopeptide from asialofetuin - MeGal methyl -d-galactopyranoside Unless stated otherwise all sugars are in thed-configuration.     

Disorder and order in unfolded and disordered peptides and proteins: A view derived from tripeptide conformational analysis. II. Tripeptides with short side chains populating asx and β‐type like turn conformations

In the preceding paper, we found that ensembles of tripeptides with long or bulky chains can include up to 20% of various turns. Here, we determine the structural and thermodynamic characteristics of GxG peptides with short polar and/or ionizable central residues (D, N, C), whose conformational distributions exhibit higher than average percentage (>20%) of turn conformations. To probe the side‐chain conformations of these peptides, we determined the ³J(H^α,H^β) coupling constants and derived the population of three rotamers with χ₁‐angles of ?60°, 180° and 60°, which were correlated with residue propensities by DFT‐calculations. For protonated GDG, the rotamer distribution provides additional evidence for asx‐turns. A comparison of vibrational spectra and NMR coupling constants of protonated GDG, ionized GDG, and the protonated aspartic acid dipeptide revealed that side chain protonation increases the pPII content at the expense of turn populations. The charged terminal groups, however, have negligible influence on the conformational properties of the central residue. Like protonated GDG, cationic GCG samples asx‐turns to a significant extent. The temperature dependence of the UVCD spectra and ³J(H^NH^α) constants suggest that the turn populations of GDG and GNG are practically temperature‐independent, indicating enthalpic and entropic stabilization. The temperature‐independent J‐coupling and UVCD spectra of GNG require a three‐state model. Our results indicate that short side chains with hydrogen bonding capability in GxG segments of proteins may serve as hinge regions for establishing compact structures of unfolded proteins and peptides. Proteins 2013. © 2012 Wiley Periodicals, Inc.     

Monoclonal antibody recognizing a determinant on type 2 chain blood group A and B oligosaccharides detects oncodevelopmental changes in azoxymethane-induced rat colon tumors and human colon cancer cell lines

Altered expression of ABH blood group substances is a common feature of human colorectal carcinoma, yet it remains unclear how these structural changes influence the biological properties of tumor cells. Azoxymethane-induced rat colon tumors display many features of the human disease, thereby providing a potentially useful model to study the role of blood group substances in colon cancer progression. We have prepared monoclonal antibodies to a microsomal fraction isolated from an azoxymethane-induced rat colon tumor and selected an antibody that detects cancer-associated changes. Monoclonal antibody (mAb) 3A7 recognizes a determinant on type 2 chain blood group A (GalNAcα1–3[Fucα1–2]Galβ1–4GlcNAc-R) and B (Galα1–3[Fucα1–2]Galβ1–4GlcNAc-R) oligosaccharides. Expression of the epitope detected by this antibody was developmentally regulated in rat colon, with maximal expression from day 4–21 after birth. Immunohistochemical staining and Western blotting analyses of azoxymethane-induced colon tumors revealed increased expression of the epitope in all of the 21 colonic tumors examined, including preneoplastic glands within transitional mucosa. Conventional and signet-ring adenocarcinomas that had invaded through the muscularis propria (Duke's B2) consistently showed the most intense staining with mAb 3A7, including regions depicting angioinvasion. Some of the lymph node metastases (Duke's C2) stained poorly with the antibody. The epitope was also expressed in blood group A positive human colon carcinoma cell lines, including HT29 and SW480 but not by SW620, a cell line derived from a lymph node metastasis isolated in vivo from the SW480 primary tumor, or in the blood group B cell line SW1417. The glycoproteins detected by mAb 3A7 in rat colon tumors and HT29 cells ranged in size between 50 and 200 kd, including a major species of 140 kd. Affinity chromatography of detergent lysates of normal rat colon on the blood group A specific lectin Dolichos biflorus (DBA)-agarose resulted in nearly quantitative binding of glycoprotein species detected by the antibody. By contrast, immunoreactive glycoproteins from rat colon tumors or HT29 cells bound poorly to DBA-agarose but were retained by another blood group A-binding lectin, Helix-pomatia (HPA)-agarose. These results indicate that colon carcinogenesis results in quantitative as well as qualitative changes in oligosaccharides detected by mAb 3A7 and suggest that the combined use of mAb 3A7 and blood group A-specific lectins may provide a useful tool for early detection of colon cancer.     

MicroRNA‐146b‐5p overexpression attenuates premature ovarian failure in mice by inhibiting the Dab2ip/Ask1/p38‐Mapk pathway and γH2A.X phosphorylation

ObjectiveTo examine the role of high‐fat and high‐sugar (HFHS) diet‐induced oxidative stress, which is a risk factor for various diseases, in premature ovarian failure (POF).Materials and methodsOvarian granulosa cells (OGCs) were isolated from mice and cultured in medium supplemented with HFHS and poly (lactic‐co‐glycolic acid) (PLGA)‐cross‐linked miR‐146b‐5p nanoparticles (miR‐146@PLGA). RNA and protein expression levels were examined using quantitative real‐time polymerase chain reaction and Western blotting, respectively. HFHS diet‐induced POF model mice were administered miR‐146@PLGA.ResultsThe ovarian tissue of mice fed a HFHS diet exhibited the typical pathological characteristics of POF. HFHS supplementation induced oxidative stress injury in the mouse OGCs, activation of the Dab2ip/Ask1/p38‐Mapk signalling pathway and phosphorylation of γH2A.X in vitro and in vivo. The results of the luciferase reporter assay revealed that miR‐146 specifically downregulated p38‐Mapk14 expression. Meanwhile, co‐immunoprecipitation and Western blot analyses revealed that HFHS supplementation upregulated nuclear p38‐Mapk14 expression and consequently enhanced γH2A.X (Ser139) phosphorylation. The HFHS diet‐induced POF mouse model treated with miR‐146@PLGA exhibited downregulated p38‐Mapk14 expression in the OGCs, mitigated OGC ageing and alleviated the symptoms of POF.ConclusionsThis study demonstrated that HFHS supplementation activates the Dab2ip/Ask1/p38‐Mapk signalling pathway and promotes γH2A.X phosphorylation by inhibiting the expression of endogenous miR‐146b‐5p, which results in OGC ageing and POF development.     

Angiotensin-converting enzyme insertion/deletion (rs106180) and angiotensin type 1 receptor A1166C (rs106165) genotypes and psoriasis: Correlation with cellular immunity,lipid profile,and oxidative stress markers

Psoriasis is a chronic inflammatory skin condition and angiotensin-converting enzyme (ACE) is a key circulating enzyme converting angiotensin (Ang) I to the vasoactive peptide Ang II. The exact role of ACE insertion (I)/deletion (D) polymorphism (rs106180) in psoriasis is not clear. We aimed to examine whether the ACE I/D and Ang II type 1 receptor (AT1R) A₁₁₆₆C-polymorphisms (rs106165), lipid profile, and stress oxidative are associated with susceptibility to psoriasis. One hundred patients with psoriasis and 100 sex- and age-matched unrelated healthy controls were recruited for this case-control study. ACE I/D and AT1R A₁₁₆₆C polymorphisms were identified by the polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism, respectively, malondialdehyde (MDA) was detected by the high-performance liquid chromatography, serum arylesterase (ARE) activity of paraoxonase and catalase activities were detected by the spectrophotometry, superoxide dismutase (SOD) activity and vascular adhesion protein (VAP)-1 were measured by ELISA. The presence of C allele of AT1R A₁₁₆₆C and I allele of ACE considerably increased the risk of psoriasis by 6.42-fold (P < 0.001). The distribution of II-genotype of ACE was significantly higher in psoriasis patients than in control group and increased the risk of disease by 3.11-times (P = 0.023). The higher levels of MDA in patients and the higher activity of SOD, ARE, and CAT was observed in healthy controls with I/D+I/I-genotype of ACE I/D. This study for the first time demonstrated that the ACE I/D and AT1R A ₁₁₆₆C genes polymorphisms robustly increases the risk of developing psoriasis in population from west of Iran. In addition, these individuals had significantly higher VAP-1 and MDA concentration and lower enzymatic and nonenzymatic antioxidant-status, suggesting that psoriatic patients carrying C allele of AT1R₁₁₆₆ polymorphism may be more susceptible to cardiovascular disease and myocardial infarction compared with A allele.