Effects of capric acid on rumen methanogenesis and biohydrogenation of linoleic and α-linolenic acid

Capric acid (C10:0), a medium chain fatty acid, was evaluated for its anti-methanogenic activity and its potential to modify the rumen biohydrogenation of linoleic (C18:2n-6) and α-linolenic acids (C18:3n-3). A standard dairy concentrate (0.5 g), supplemented with sunflower oil (10 mg) and linseed oil (10 mg) and increasing doses of capric acid (0, 10, 20 and 30 mg), was incubated with mixed rumen contents and buffer (1 : 4 v/v) for 24 h. The methane inhibitory effect of capric acid was more pronounced at the highest (30 mg) dose compared to the medium (20 mg) (-85% v. -34%), whereas the lower dose (10 mg) did not reduce rumen methanogenesis. A 23% decrease in total short-chain fatty acid (SCFA) production was observed, accompanied by shifts towards increased butyrate at 20 mg and increased propionate at 30 mg of capric acid (P < 0.001). Capric acid linearly decreased the extent of biohydrogenation of C18:2n-6 and C18:3n-3, by up to 60% and 86%, respectively. This reduction was partially due to a lower extent of lipolysis when capric acid was supplemented. Capric acid at 20 and 30 mg completely inhibited the production of C18:0 (P < 0.001), resulting in an accumulation of biohydrogenation intermediates, mainly C18:1t10 + t11 and C18:2t11c15. In contrast to effects on rumen fermentation (methane production and proportions of SCFA), 30 mg of capric acid did not induce major changes in rumen biohydrogenation as compared to the medium (20 mg) dose. This study revealed the dual action of capric acid, being inhibitory to both methane production and biohydrogenation of C18:2n-6 and C18:3n-3.     

The effect of α2,6-linked sialic acid on anti-IgM antibody-induced apoptosis in Ramos cells

Apoptosis in B cells is induced through the B cell antigen receptor (BCR) and affects the sialic acid recognition molecules on B cells. We investigated the effects of ₁-acid glycoprotein (AGP), which mainly contains 2,6-linked sialic acid, on anti-IgM antibody (Ab)-induced apoptosis in Ramos cells, which are derived from Burkitt's lymphoma. When Ramos cells were incubated with anti-IgM-Ab in plates coated with AGP, neuraminidase-digested AGP (asAGP) or 2,3-sialylated AGP (2,3AGP), apoptosis was suppressed only in those coated with AGP. We also studied the effects of CD22, which is expressed on the surface of mature B cells and binds to sugar chains containing 2,6-linked sialic acid, with anti-CD22 monoclonal antibody (mAb). Anti-CD22mAb enhanced anti-IgM Ab-induced apoptosis in Ramos cells. These contradictory results suggested that the recognition molecules for 2,6-linked sialic acid on AGP, which inhibits B-cell apoptosis, is distinct from CD22, or that different binding domains of CD22 between 2,6-linked sialic acid and anti-CD22 mAb exert opposite functions of suppression or enhancement to anti-IgM Ab-induced B cells.     

Chemoenzymatic synthesis of C8-modified sialic acids and related α2-3- and α2-6-linked sialosides

Naturally occurring 8-O-methylated sialic acids, including 8-O-methyl-N-acetylneuraminic acid and 8-O-methyl-N-glycolylneuraminic acid, along with 8-O-methyl-2-keto-3-deoxy-d-glycero-d-galacto-nonulosonic acid (Kdn8Me) and 8-deoxy-Kdn were synthesized from corresponding 5-O-modified six-carbon monosaccharides and pyruvate using a sialic acid aldolase cloned from Pasteurella multocida strain P-1059 (PmNanA). In addition, α2-3- and α2-6-linked sialyltrisaccharides containing Neu5Ac8Me and Kdn8Deoxy were also synthesized using a one-pot multienzyme approach. The strategy reported here provides an efficient approach to produce glycans containing various C8-modified sialic acids for biological evaluations.     

Effects of ascorbic acid and ��-carotene on HepG2 human hepatocellular carcinoma cell line

Recent studies have demonstrated that vegetable rich diets have protective effects on the occurrence and prognosis of various cancers. In addition to dietary intakes, ascorbic acid and β-carotene are also taken as supplements. The aim of this study was to assess effects of ascorbic acid, β-carotene and their combinations on human hepatocellular carcinoma cell line HepG2. Ascorbic acid and β-carotene were applied to cells as plasma peak concentrations (70 and 8 μM, respectively) and their half concentrations (35 and 4 μM, respectively) for 24 and 48 h. Genotoxic and cytotoxic effects of ascorbic acid and β-carotene were evaluated by alkali single cell gel electrophoresis (SCGE), acridine orange/ethidium bromide staining patterns of cells (apoptosis and necrosis) and lipid peroxidation (thiobarbituric acid reactive substances, TBARS). Results of the SCGE demonstrated that both ascorbic acid and β-carotene caused DNA damage on HepG2 which were also concordant to increased apoptosis and necrosis of cells. Increased TBARS values also demonstrated increased lipid peroxidation in these cells. Results of the present study demonstrates that when dietary intakes of ascorbic acid and β-carotene and their relevant achievable plasma level concentrations were considered, both ascorbic acid and β-carotene induce genotoxic and cytotoxic damage on HepG2 together with increased oxidative damage in contrast to their protective effect on healthy cells. This may be correlated to oxidative status and balance of ROS in hepatocellular carcinoma cells.     

Studies on type II glycogenosis. Effects of cortisone derivatives on acid α-glucosidase

Cortisone causes a marked increase in the activity of liver acid alpha-glucosidase 2h after injection into male Wistar rats. Studies on rat liver tissue slices, isolated lysosomes and cultured skin fibroblasts have demonstrated similar elevations of acid alpha-glucosidase activity after incubation with cortisone. Cortisone-treated human liver tissue, obtained by needle biopsy, also shows an increase in acid alpha-glucosidase activity. Neutral alpha-glucosidase activity was not stimulated by cortisone in vivo or in liver slices.     

Effects of α-tocopherol analogs on lysosome membranes and fatty acid monolayers

The surface pressures of α-tocopherol analogs, fatty acids, and their mixtures were measured in their spread monolayers at an air—water interface. The surface pressure—area isotherms for the mixed monolayers of α-tocopherol and either stearic acid, oleic acid or linoleic acid deviated positively from those calculated on the basis of the additivity rule, and the magnitude depended on the length of the phytyl side chain in α-tocopherol and on the degree of unsaturation of the fatty acid chains. Lysosome membranes of mouse liver were stabilized by addition of α-tocopherol. A decrease in the length of the phytyl side chain in α-tocopherol reduced its ability to stabilize lysosome membranes. A good correlation was obtained between the extent of stabilizing activity of α-tocopherol analogs on lysosome membranes and the degree of positive deviation of the surface pressure for their mixtures with fatty acids.     

Effects of α-ketoglutarate on neutrophil intracellular amino and α-keto acid profiles and ROS production

The aim of this study was to determine the effects of α-ketoglutarate on neutrophil (PMN), free α-keto and amino-acid profiles as well as important reactive oxygen species (ROS) produced [superoxide anion (O₂ ^?), hydrogen peroxide (H₂O₂)] and released myeloperoxidase (MPO) acitivity. Exogenous α-ketoglutarate significantly increased PMN α-ketoglutarate, pyruvate, asparagine, glutamine, asparatate, glutamate, arginine, citrulline, alanine, glycine and serine in a dose as well as duration of exposure dependent manner. Additionally, in parallel with intracellular α-ketoglutarate changes, increases in O₂ ^– formation, H₂O₂-generation and MPO acitivity have also been observed. We therefore believe that α-ketoglutarate is important for affecting PMN “susceptible free amino- and α-keto acid pools” although important mechanisms and backgrounds are not yet completely explored. Moreover, our results also show very clearly that changes in intragranulocytic α-ketoglutarate levels are relevant metabolic determinants in PMN nutrition considerably influencing and modulating the magnitude and quality of the granulocytic host defense capability as well as production of ROS.     

Effects of gibberellic acid and abscisic acid on α-amylase activity and ultrastructure of aleurone cells ofAvena sativa L.

The effects of GA₃ and/or ABA on the α-amylase activity and the ultrastructure of aleurone cells in halves of seeds without embryos (embryo-less half seeds) of oats (Avena sativa L.) were studied. α-Amylase activity was detected by the starch-agar gel method in the aleurone layers of embryo-less half seeds soaked in 1 μM GA₃ solution or 100 μM GA₃+10 μM ABA solution but not in those of seeds soaked in distilled water, 10 μM ABA solution, or 1 μM GA₃+10 μM ABA solution. Ultrastructural examinations of aleurone cells with α-amylase activity showed a decrease in the number of sphaerosomes, the appearance of flattened saccules pressed to the surface of aleurone grains, and the development and transformations of the rER from a slender form to the one with wide inner spaces. In the aleurone cells in which the enzyme activity was not detected, components of the rER showed only slender profiles. The number of sphaerosomes did not decrease, and no flattened saccules appeared in the aleurone cells treated with 10 μM ABA or 1 μM GA₃+10 μM ABA.     

Effects of non-linearity on cell–ECM interactions

Filamentous biopolymers such as F-actin, vimentin, fibrin and collagen that form networks within the cytoskeleton or the extracellular matrix have unusual rheological properties not present in most synthetic soft materials that are used as cell substrates or scaffolds for tissue engineering. Gels formed by purified filamentous biopolymers are often strain stiffening, with an elastic modulus that can increase an order of magnitude at moderate strains that are relevant to cell and tissue deformation in vivo. This review summarizes some experimental studies of non-linear rheology in biopolymer gels, discusses possible molecular mechanisms that account for strain stiffening, and explores the possible relevance of non-linear rheology to the interactions between cell and extracellular matrices.     

Effects of curvature and cell–cell interaction on cell adhesion in microvessels

It has been found that both circulating blood cells and tumor cells are more easily adherent to curved microvessels than straight ones. This motivated us to investigate numerically the effect of the curvature of the curved vessel on cell adhesion. In this study, the fluid dynamics was carried out by the lattice Boltzmann method (LBM), and the cell dynamics was governed by the Newton’s law of translation and rotation. The adhesive dynamics model involved the effect of receptor-ligand bonds between circulating cells and endothelial cells (ECs). It is found that the curved vessel would increase the simultaneous bond number, and the probability of cell adhesion is increased consequently. The interaction between traveling cells would also affect the cell adhesion significantly. For two-cell case, the simultaneous bond number of the rear cell is increased significantly, and the curvature of microvessel further enhances the probability of cell adhesion.     

α2-3- and α2-6- N-linked sialic acids allow efficient interaction of Newcastle Disease Virus with target cells

Receptor recognition and binding is the first step in the viral cycle. It has been established that Newcastle Disease Virus (NDV) interacts with sialylated molecules such as gangliosides and glycoproteins at the cell surface. Nevertheless, the specific receptor(s) that mediate virus entry are not well known. We have analysed the role of the sialic acid linkage in the early steps of the viral infection cycle. Pretreatment of ELL-0 cells with both α2,3 and α2,6 specific sialidases led to the inhibition of NDV binding, fusion and infectivity, which were restored after α2,3(N)- and α2,6(N)-sialyltransferase incubation. Moreover, α2,6(N)-sialyltransferases also restored NDV activities in α2-6-linked sialic acid deficient cells. Competition with α2-6 sialic acid-binding lectins led to a reduction in the three NDV activities (binding, fusion and infectivity) suggesting a role for α2-6- linked sialic acid in NDV entry. We conclude that both α2-3- and α2-6- linked sialic acid containing glycoconjugates may be used for NDV infection. NDV was able to efficiently bind, fuse and infect the ganglioside-deficient cell line GM95 to a similar extent to that of its parental MEB4, suggesting that gangliosides are not essential for NDV binding, fusion and infectivity. Nevertheless, the fact that the interaction of NDV with cells deficient in N-glycoprotein expression such as Lec1 was less efficient prompted us to conclude that NDV requires N-linked glycoproteins for efficient attachment and entry into the host cell.     

Effects of α-eleostearic acid on asolectin liposomes dynamics: Relevance to its antioxidant activity

In this study, the effect of α-eleostearic acid (α-ESA) on the lipid peroxidation of soybean asolectin (ASO) liposomes was investigated. This effect was correlated to changes caused by the fatty acid in the membrane dynamics. The influence of α-ESA on the dynamic properties of liposomes, such as hydration, mobility and order, were followed by horizontal attenuated total reflection Fourier transform infrared spectroscopy (HATR-FTIR), nuclear magnetic resonance (NMR), differential scanning calorimetry (DSC) and UV–vis techniques. The α-ESA showed an in vitro antioxidant activity against the damage induced by hydroxyl radical (OH) in ASO liposomes. The analysis of HATR-FTIR frequency shifts and bandwidths and ¹H NMR spin–lattice relaxation times, related to specific lipid groups, showed that α-ESA causes an ordering effect on the polar and interfacial regions of ASO liposomes, which may restrict the OH diffusion in the membrane. The DSC enthalpy variation analysis suggested that the fatty acid promoted a disordering effect on lipid hydrophobic regions, which may facilitate interactions between the reactive specie and α-ESA. Turbidity results showed that α-ESA induces a global disordering effect on ASO liposomes, which may be attributed to a change in the lipid geometry and shape. Results of this study may allow a more complete view of α-ESA antioxidant mode of action against OH, considering its influence on the membrane dynamics.     

“Just a spoonful of sugar...”: import of sialic acid across bacterial cell membranes

Eukaryotic cell surfaces are decorated with a complex array of glycoconjugates that are usually capped with sialic acids, a large family of over 50 structurally distinct nine-carbon amino sugars, the most common member of which is N-acetylneuraminic acid. Once made available through the action of neuraminidases, bacterial pathogens and commensals utilise host-derived sialic acid by degrading it for energy or repurposing the sialic acid onto their own cell surface to camouflage the bacterium from the immune system. A functional sialic acid transporter has been shown to be essential for the uptake of sialic acid in a range of human bacterial pathogens and important for host colonisation and persistence. Here, we review the state-of-play in the field with respect to the molecular mechanisms by which these bio-nanomachines transport sialic acids across bacterial cell membranes.     

Effects of the 3-hydroxyanthranilic acid analogue NCR-631 on anoxia-, IL-1β- and LPS-induced hippocampal pyramidal cell lossin vitro

Summary The kynurenine pathway intermediate 3-hydroxyanthranilic acid (3-HANA) is converted by 3-HANA 3,4-dioxygenase (3-HAO) to the putative neuropathogen quinolinic acid (QUIN). In the present study, the neuroprotective effects of the 3-HANA analogue and 3-HAO inhibitor NCR-631 was investigated using organotypic cultures of rat hippocampus. An anoxic lesion was induced by exposing the cultures to 100% N₂ for 150 min, resulting in a pronounced loss of pyramidal neurons, as identified using NMDA-R1 receptor subunit immunohistochemistry. NCR-631 provided a concentration-dependent protective effect against the anoxia. NCR-631 was also found to counteract the loss of pyramidal neurons in two models of neuroinflammatory-related damage; incubation with either LPS (10 ng/ml) or IL-1 (10 IU/ml). The findings suggest that NCR-631 has neuroprotective properties and that it may be a useful tool to study the role of kynurenines in neurodegeneration.Abbreviations EAA excitatory amino acid - 3-HANA 3-hydroxyanthranilic acid - 3-HAO 3-hydroxyanthranilic acid 3,4-dioxygenase - IL-1 interleukin-1 - KYNA kynurenic acid - LPS lipopolysaccaride - NCR-631 4,6-dibromo-3hydroxyanthranilic acid - NMDA N-methyl-d-aspartate - QUIN quinolinic acid     

Effects of decorin on the expression of α-smooth muscle actin in a human myofibroblast cell line

Myofibroblasts are metabolically and morphologically distinctive fibroblasts expressing α-smooth muscle actin (α-SMA), and their activation plays a key role in development of the fibrotic response. In an activated state, myofibroblasts cease to proliferate and start to synthesize large amounts of extracellular component proteins. The expression of α-SMA correlates with the activation of myofibroblasts. Decorin, a member of the small leucine-rich proteoglycan gene family, has been implicated in the negative control of cell proliferation primarily by upregulating the expression of p21, a potent inhibitor of cyclin-dependent kinase. In order to examine the effect of decorin on myofibroblast cell growth, we rendered a human lung myofibroblast cell line, MRC-5, quiescent by either cell–cell contact or serum starvation, and examined the relationship between decorin and α-SMA expression in these cells. The expression of decorin in cells made quiescent by serum starvation was lower than that in cells made quiescent by cell–cell contact. In contrast, the expression of α-SMA in cells made quiescent by cell–cell contact was lower than that in cells made quiescent by serum starvation. Furthermore, forced expression of decorin was accompanied by a suppression of α-SMA expression, whereas knocking down of decorin expression by RNA interference increased the expression of α-SMA.     

Lectin affinity chromatography of articular cartilage fibromodulin: Some molecules have keratan sulphate chains exclusively capped by α(2-3)-linked sialic acid

Fibromodulin from bovine articular cartilage has been subjected to lectin affinity chromatography by Sambucus nigra lectin which binds α(2-6)- linked N-acetylneuraminic acid, and the structure of the keratan sulphate in the binding and non-binding fractions examined by keratanase II digestion and subsequent high pH anion exchange chromatography. It has been confirmed that the keratan sulphate chains attached to fibromodulin isolated from bovine articular cartilage may have the chain terminating N-acetylneuraminic acid residue α(2-3)- or α(2-6)-linked to the adjacent galactose residue. Although the abundance of α(2-6)-linked N-acetylneuraminic acid (ca. 22%) is such that this could cap one of the four chains in almost all fibromodulin molecules, it was found that ca. 34% of the fibromodulin proteoglycan molecules from bovine articular cartilage were capped exclusively with α(2-3)-linked N-acetylneuraminic acid. The remainder of the fibromodulin proteoglycans, which bound to the lectin had a mixture of α(2-3)- and α(2-6)-linked N-acetylneuraminic acid capping structures. The keratan sulphates attached to fibromodulin molecules capped exclusively with α(2-3)- linked N-acetylneuraminic acid were found to have a higher level of galactose sulphation than those from fibromodulin with both α(2-3)- and α(2-6)-linked N-acetylneuraminic acid caps, which bound to the Sambucus nigra lectin. In addition, both pools contained chains of similar length (ca. 8–9 disaccharides). Both also contained α(1-3)-linked fucose, showing that this feature does not co-distribute with α(2-6)-linked N-acetylneuraminic acid, although these two features are present only in mature articular cartilage. These data show that there are discrete populations of fibromodulin within articular cartilage, which may have differing impacts upon tissue processes.     

Effects of α-zearalenol on the metabolome of two breast cancer cell lines by 1H-NMR approach

Introduction

Zearalenone (ZEN) is one of the most widely distributed toxins that contaminates many crops and foods. Its major metabolites are α-Zearalenol (α-zol) and β-Zearalenol. Previous studies showed that ZEN and α-zol have estrogenic properties and are able to induce growth promoting effect in breast tissues.

Objectivies

Considering that tumorigenesis is dependent on the reprogramming of cellular metabolism and that the evaluation of the cellular metabolome is useful to understand the metabolic changes that can occur during the cancer development and progression or after treatments, aim of our work is to study, for the first time, the effects of α-zol on the metabolomic profile of an estrogen positive breast cancer cell line, MCF-7, and of an estrogen negative breast cancer cell lines MDA-MB231.

Methods

Firstly, we tested the effects of α-zol on the cell viability after 24, 48 and 72 h of treatments with 10^?10, 10^?8 and 10^?6 M concentrations on breast cancer MCF-7 and MDA-MB231 cell lines in comparison to human non-cancerous breast MCF10A cell line. Then, we evaluated cell cycle progression, levels of reactive oxygen species (ROS) and the metabolomic profiling by ¹H-NMR approach on MCF-7 and MDA-MB231 before and after 72 h treatments. Principal component analysis was used to compare the obtained spectra.

Results

α-zol is resulted able to induce: (i) an increase of the cell viability on MCF-7 cells mainly after 72 h treatment, (ii) a slight decrease of the cell viability on MDA-MB231 cells, and (iii) an increase of cells in S phase of the cell cycle and of ROS only in MCF-7 cells. Moreover, the evaluation of metabolomics profile evidenced that after treatment with α-zol the levels of some metabolites increased in MCF-7 cells whereas decreased slightly in MDA-MB231 cells.

Conclusions

Our results showed that α-zol was able to increase the protein biosynthesis as well as the lipid metabolism in MCF-7 cells, and, hence, to induce an estrogen positive breast cancer progression.

     

An improved method for the measurement of total lipid-bound sialic acids after cleavage of α2,8 sialic acid linkage withVibrio cholerae sialidase in the presence of cholic acid,SDS and Ca2+

In the measurement of total lipid-bound sialic acids involving periodic acid oxidation, as in the periodate-resorcinol assay, the inner sialic acids of disialoglycolipids (such as GD₃ and GD₂) are not involved because their 2,8 ketosidic linkages are resistant to periodic acid oxidation, even after acid/enzyme hydrolysis or alkali pretreatment. However, the sialic acids from these glycolipids can be recovered completely after cleavage of 2,8 linkages byV. cholerae sialidase in the presence of cholic acid, sodium dodecyl sulphate and calcium. Interestingly, removal of calcium or detergent(s) or both significantly minimizes the sialidase action on the disialyl residues of these gangliosides. Therefore, we recommend sialidase (Vibrio cholerae) pretreatment of the glycolipids in the presence of cholic acid, SDS and Ca²⁺ for complete recovery of sialic acids from di- and polysialogangliosides and for accurate measurement of total lipid-bound sialic acids by periodate-resorcinol assay.Presented at the Second International Glycobiology Symposium which was held in San Francisco, CA, USA (14 February 1994).     

Characterization of subfractions of β2-glycoprotein I: Evidence for sialic acid microheterogeneity

• 1.1. β₂-Glycoprotein I is a sialic acid microheterogeneous protein and contains on the average 11 mol sialic acid/mol.
• 2.2. Linear correlation was found between sialic acid content and pI of isolated subfractions.
• 3.3. Asialo-β₂2-glycoprotein I consists of 2 isoforms. Each of which can originate from the same subfraction.
• 4.4. The isolated subfractions exhibited almost the same amino acid composition.