In vitro cytotoxicity assay of Fucoidan extracted from Turbinaria conoides against cancer cell lines MCF7, A549, and normal cell line L929

Abstract The main aim of the study is to quantify the cytotoxic property of the Fucoidan extracted from the Turbinaria conoides using the MTT assay with the standard fucose. Fucoidan was extracted using the soaked water method and it was determined using the HPLC procedure the obtained Test sample Fucoidan extracted from the Turbinaria conoides and standard fucose was subjected to the cytotoxicity assay against the MCF7 Human breast cancer cell line, A549 lung cancer cell line, and L929 normal mouse fibroblast cell line. From the results it was found that the Test sample showed good IC50 value for MCF7 cell line then A549 with an increasing concentration 24 hours incubation at 37°C The IC50 for MCF7 was 115.21 µg/ml and A549 396.46µg/ml and the Fucoidan extract was checked for its cytotoxicity against the normal mouse fibroblast cell line L929, Fucoidan was found non-lethal to the L929 mouse fibroblast normal cell line. Standard fucose also gave a significant result towards MCF7 and against the L929. This indicates that the Fucoidan extracted from Tubinaria conoides shows better anticancer potential in it. Hence its application can be further extended in the pharmacological fields.

Hidratos de carbono complejos en la leche materna: los oligosacáridos (Parte 1) / Complex carbohydrates in breast milk: oligosaccharides (Part 1)

Los oligosacáridos de la leche materna (HMOs) son unas 200 moléculas distintas sintetizadas y secretadas por la glándula mamaria a partir de lactosa a la que diversos enzimas unen monosacáridos simples (glucosa, galactosa, n-acetil galactosamina, fucosa y ácido siálico). Estas uniones y sus diferentes orientaciones espaciales generan una gran diversidad de estructuras químicas y de funcionalidades. La concentración de los HMOs es mayor en el calostro (± 25 g/L), está relacionada con la duración del embarazo y la lactancia: disminuyen progresivamente hasta la mitad de los niveles iniciales. La genética materna influye en el perfil de algunos HMOs; el gen FUT2, que codifica la síntesis de la fucosiltransferasa 2 (FUT2) condiciona el llamado carácter secretor en 75-85% de las mujeres y hace que los antígenos del grupo ABO(H) sean secretados en los líquidos orgánicos (saliva, lágrimas, semen). La ausencia de actividad del gen FUT2 condiciona el carácter no-secretor (15-25% de las mujeres). La actividad del gen FUT3 condiciona la actividad de la fucosiltransferasa 3 (FUT3) que se asocia con el grupo sanguíneo Lewis+ mientras que su ausencia caracteriza a los portadores como Lewis 0. Los HMOs son absorbidos a nivel del intestino como trazas (1%) pero incluso en esas cantidades ejercerían efectos sistémicos.

Human milk oligosaccharides (HMOs) are a family of some 200 different molecules synthesized by the mammary gland. At the core is a molecule of lactose, which is linked by different enzymes to glucose, galactose, n-acetyl galactosamine, fucose or sialic acid. These linkages and their different spatial orientation generate, besides the possibilities of numerous chemical structures, the potential for different spatial isomers. The concentration of HMOs in human milk depends on pregnancy and breastfeeding duration. They are highest in colostrum (± 25 g/L) and decrease over time to half this initial level. Maternal genetics modifies the concentration and profile of some oligosaccharides. For example, the FUT2 gene codifies the synthesis of fucosyltransferase 2 (FUT2) whose activity generates the secretor status for antigens of the ABO(H) blood group in organic fluids (saliva, milk, tears, semen) among 75-85% of the carriers of the trait. The absence of activity of the FUT2 gene conditions the non-secretor status (15-25% of women). The FUT3 gene regulates the activity of the fucosyltransferase 3 (FUT3) that is associated with the Lewis blood group. Traces of HMOs (1%) are absorbed in the intestinal tract, however, they exert important systemic effects even at low concentrations.

Protective effect of Cardiospermum halicacabum leaf extract on glycoprotein components on STZ-induced hyperglycemic rats

OBJECTIVE@#To investigate the protective role of Cardiospermum halicacabum (C. halicacabum) leaf extract on glycoprotein metabolism in streptozotocin (STZ)-induced diabetic rats.@*METHODS@#Diabetes was induced in male albino Wistar rats by intraperitonial administration of STZ. The C. halicacabum leaf extract (CHE) was administered orally to normal and STZ-diabetic rats for 45 days. The effects of C. halicacabum leaf extract (CHE) on plasma and tissue glycoproteins (hexose, hexosamine, fucose and sialic acid) were determined.@*RESULTS@#The levels of plasma and tissues glycoproteins containing hexose, hexosamine and fucose were significantly increased in STZ-induced diabetic rats. In addition, the level of sialic acid significantly increased in plasma and liver while decreased in kidney of STZ-induced diabetic rats. After administration of CHE to diabetic rats, the metabolic alteration of glycoprotein reverted towards normal levels.@*CONCLUSIONS@#The present study indicates that the CHE possesses a protective effect on abnormal glycoprotein metabolism in addition to its antihyperglycemic activity.

Generation of glyco-engineered BY2 cell lines with decreased expression of plant-specific glycoepitopes

Plants are known to be efficient hosts for the production of mammalian therapeutic proteins. However, plants produce complex N-glycans bearing β1,2-xylose and core α1,3-fucose residues, which are absent in mammals. The immunogenicity and allergenicity of plant-specific Nglycans is a key concern in mammalian therapy. In this study, we amplified the sequences of 2 plant-specific glycosyltransferases from Nicotiana tabacum L. cv Bright Yellow 2 (BY2), which is a well-established cell line widely used for the expression of therapeutic proteins. The expression of the endogenous xylosyltranferase (XylT) and fucosyltransferase (FucT) was downregulated by using RNA interference (RNAi) strategy. The xylosylated and core fucosylated N-glycans were significantly, but not completely, reduced in the glycoengineered lines. However, these RNAi-treated cell lines were stable and viable and did not exhibit any obvious phenotype. Therefore, this study may provide an effective and promising strategy to produce recombinant glycoproteins in BY2 cells with humanized N-glycoforms to avoid potential immunogenicity.

Serum fucose level in malignant diseases.

We review the present knowledge of serum fucose with special attention to its relation with various malignant diseases. We summarize the role of serum fucose as a useful diagnostic and prognostic marker when used singly or in combination. The purpose of this review is to provide an expert opinion on the practical and applied aspect of serum fucose level in clinical practice and research settings. Our review is based on information from published research studies, library books, and electronic searches through Medline and PubMed. The available published data were used as the basis for recommendations. Each of the subsections concludes to provide information to assist the clinicians and the research scientists make informed decisions.

Composition and Partial Structure Characterization of Tremella Polysaccharides

Heteropolysaccharides isolated from liquid cultures of nine Tremella species contained 0.3 to 1.2% protein, 2.7 to 5% ash, 0.9 to 3.4% acetyl groups, 76.5 to 84.2% carbohydrates and trace amounts of starch. The polysaccharides in aqueous solution were slightly acidic (pH 5.1 to 5.6). They consisted of the following monomeric sugars: fucose, ribose, xylose, arabinose, mannose, galactose, glucose and glucuronic acid. The backbones of the polysaccharide structures consisted of alpha-(1-->3)-links while the side chains were beta-linked.

Analysis of monosaccharides in Radix Rehmanniae by GC / 中国中药杂志

<p><b>OBJECTIVE</b>To isolate and purify the polysaccharides from Radix Rehmanniae and analysis the monosaccharides composition.</p><p><b>METHOD</b>The polysaccharides were extracted with hot water and precipitated by alcohol. Proteins in the precipitates were removed by TCA method. The products were further purified with column chromatography on Superdex 200 and Sephadex G100. The SRP I and SRP II were identified as homogeneous polysaccharide by HPLC, respectively, and then analyzed by GC after being hydrolysised.</p><p><b>RESULT</b>Two homogeneous polysaccharides (SRP I and SRP II) were obtained from Radix Rehmanniae.</p><p><b>CONCLUSION</b>SRP I contained rhamnose, arabinose, glucose and galactose in the percentage of 6.11%, 66.46%, 3.93% and 21.50%. SRP I was composed of rhamnose, fucose, mannose, galactose and fructose in the percentage of 21.82%, 24.47%, 10.48%, 29.94% and 13.29%.</p>

Histochemical detection of sugar residues in lizard teeth (Liolaemus gravenhorsti): a lectin-binding study

The structural diversity of the many oligosaccharide chains of surface glycoconjugates renders them likely candidates for modulators of cell-interactions, cellular movements, differentiation, and cellular recognition. A selection of different lectins was used to investigate the appearance of cellular distribution and changes in sugar residues during tooth development in the polyphyodont lizard, Liolaemus gravenhorsti. Lectins from three groups were used: (1) N-acetylgalactosamine specificity: BS-1, PNA, RCA-120; (2) N-acetylglucosamine specificity: ECA; and (3) fucose specificity: UEA 1 and LTA.. Digital images were processed using Scion Image. Grayscale graphics in each image were obtained. The lectins used showed a strong, wide distribution of the L-fucose and N-acetylgalactosamine at the cell surface and in the cytoplasm of multinucleate odontoclast cell, while mononuclear odontoclast cells showed no binding, suggesting some roles that the residues sugar might play in the resorption of dentine or with multinucleation of odontoclast after the attachment to the dentine surface in this polyphyodont species. Further studies must be planned to determine the specific identities of these glycoconjugates,and to elucidate the roles played by these sugar residues in the complex processes related to odontogenesis in polyphyodont species.

The Study on the Recognition of Apoptotic Thymocyte by Macrophage / 대한해부학회지

The present study was carried out to investigate the correlation with the sugar specificity of the apoptotic cells and the phagocytosis of the macrophage. Young rat thymocytes were isolated from the thymus and macrophages were isolated from the peritoneal cavity. Isolated thymocytes were treated with dexamethasone to induce apoptosis. After identified the apoptotic cells by TUNEL method, nine kinds of lectins (PSA, UEA I, GSL I B4, ECL, DBA, SBA, DSL, GSL II or WGA) were used to investigate the sugar specificity of the apoptotic thymocyte. And we carried out the phagocytosis assay using the lectin-inhibitory sugars. By TUNEL method, the most numbers of apoptotic thymocytes were observed in the cultured thymocytes treated with 10(-6)M of dexamethasone for 6 hours. From lectin histochemistry, apoptotic thymocyte showed positive reaction with PSA, UEA I, SBA, DSL and WGA, which indicate apoptotic cells have the sugar residues of alpha-D-mannose, alpha-L-fucose, terminal beta-N-acetylgalactosamine and internal beta-1, 4-N-acetylglucosamine oligomers. From phagocytosis assay using cultured thymocytes and macrophages, apoptotic thymocytes of positive reaction with TUNEL method were phagocytosed by macrophage. When the apoptotic thymocytes treated with the lectin-inhibitory sugars were mixed with the macrophages, it was observed that the phagocytosis of macrophages was reduced by methyl-alpha-mannopyranoside and alpha-L-fucose. Considering overall results, it can be assumed that the mannose and fucose may play an important role in the recognition of apoptotic thymocytes by macrophages.

Characterization of mouse fibroblast (NIH3T3) and fibrosarcoma cell lines (WEHI-164 and MFS 8) using PMRS.

Proton magnetic resonance Spectroscopy (PMRS) has been used to study the differences between immortalized fibroblasts and fibrosarcoma cells of different grade. One and two dimensional purged correlation spectroscopy (PCOSY) have been used to assess intact viable fibroblast and fibrosarcoma cells, and differences in the triglyceride, cellular metabolite, and cell surface fucosylation patterns between the three cell lines have been observed. The clinical implication of this study is the potential use of PMRS as an adjunct to conventional histopathology.

Raddioactive fucose as a tool for studying glycoprotein secretion

The efficiency and reliability of radioactive fucose as a specific label for newly synthesized glycoproteins were investigated. Young adult male rabbits were injected intravitreally with [3H]-fucose, [3H]-galactose, [3H]-mannose, N-acetyl-[3H]-glucosamine or N-acetyl-[3H]-mannosamine, and killed 40 h after injection. In another series of experiments rabbits were injected with either [3H]-fucose or several tritiated amino acids and the specific activity of the vitreous proteins was determined. Vitreous samples were also processed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and histological sections of retina, ciliary body and lens (the eye components around the vitreous body) were processed for radioautography. The specific activity (counts per minute per microgram of protein) of the glycoproteins labeled with [3H]-fucose was always much higher than that of the proteins labeled with any of the other monosaccharides or any of the amino acids. There was a good correlation between the specific activity of the proteins labeled by any of the above precursors and the density of the vitreous protein bands detected by fluorography. This was also true for the silver grain density on the radioautographs of the histological sections of retina, ciliary body and lens. The contribution of radioautography (after[3H]-fucose administration) to the elucidation of the biogenesis of lysosomal and membrane glyco-proteins and to the determination of the intracellular process of protein secretion was reviewed. Radioactive fucose is the precursor of choice for studying glycoprotein secretion because it specific, efficient and practical for this purpose.

The FML (Fucose Mannose Ligand) of Leishmania donovani: a new tool in diagnosis, prognosis, transfusional control and vaccination against human kala-azar

The Fucose-Mannose Ligand (FML) of Leishmania donovani is a complex glycoproteic fraction. Its potential use as a tool for diagnosis of human visceral leishmaniasis was tested with human sera from Natal, Rio Grande do Norte, Brazil. The FML-ELISA test, showed 100 per cent sensitivity and 96 per cent specificity, identifying patients with overt kala-azar (p < 0.001, when compared to normal sera), and subjects with subclinical infection. More than 20 per cent apparently healthy subjects with positive reaction to FML developed overt kala-azar during the following 10 months. In the screening of human blood donnors, a prevalence of 5 per cent of sororeactive subjects was detected, attaining 17 per cent in a single day. The GP36 glycoprotein of FHL is specifically reconized by human kala-azar sera. The immunoprotective effect of FML on experimental L. donovani infection was tested in swiss albino mice. The protection scheemes included three weekly doses of FML, supplemented or not with saponin by the subcutaneous or intraperitoneal routes and challenge with 2 x 10(7) amastigotes of Leishmania donovani. An enhancement of 80.0 per cent in antibody response (p < 0.001) and reduction of 85.5 per cent parasite liver burden (p < 0.001) was detected in animals immunized with FML saponin, unrespectively of the immunization route.

Fucosylated glycoconjugates of the human spermatozoon. Comparison of the domains of these glycoconjugates with the alpha-fucosyl binding sites, and with lactosaminic glycoconjugates and beta-D-galactosyl binding site domains

Fucosylated glycoconjugates play an important role in fertilization as the recognition signal of the zona pellucida. In this work, using "critical" concentrations of either, FITC Lotus tetragonolobus lectin or FITC alpha-L-fucosyl-BSA neoglycoprotein as molecular probes, population densities of fucosylated glycoconjugates and of their "complementary" molecules (carrying fucosyl receptors), were found all over the sperm surface with higher population densities in post acrosomal sheath, neck and midpiece. These results were compared with previously reported data on the population densities of lactosaminic compounds and their "complementary" molecules, obtained on same samples of spermatozoa. Statistical data demonstrate that fucosylated glycoconjugates share the same domains with biantennary N-acetyllactosaminic oligosaccharides carrying outer galactose and bisected N-acetylglucosamine residues. These domains highly differ with those of the lactosaminic glycoproteins carrying tri and tetraantennary N-acetyllactosaminic oligosaccharides. These studies also show that the domains of fucosylated glycoconjugates and their "complementary" molecules (carrying fucosyl receptors) locate in different zones of the spermatozoon than those of the compounds carrying beta-galactosyl receptors. Besides, the results suggest structural differences between fucosylated glycoconjugates of the acrosome, equatorial zone and post acrosomal sheath. This may be relevant to the different biological behavior of these compounds and zones, in fertilization.

Caracterización de fracciones mayoritarias del plasma seminal humano: estudio comparativo entre individuos normospérmicos, azoospérmicos, necrozoospérmicos, oligozoospérmicos y vasectomizados / Caracterization of major human seminal plasma fractions: comparative study with normospermic, azoospermic, necrozoospermic, oligozoospermic and vasectomised donors

El plasma seminal humano (PSH) de individuos normospérmicos y con diferentes patologías fue analizado por cromatografía de afinidad en Concanavalina-A Sepharose. Se obtuvieron dos fracciones mayoritarias, F1 y F2, en proporciones variables y con diferencias en la composición de monosacáridos y aminoácidos de acuerdo con el tipo de patología. Galactosa es el principal miembro, juntamenete con fucosa y proporciones variables de manosa, N-acetilglucosamina y N-acetilgalactosamina. Se encontraron diferencias en la estructura primaria de los glicanos (enlaces N- y O-glicosídicos) presentes en las fracciones de los individuos con diferentes patologías. En F1 todas las cadenas de oligosacáridos son del tipo O-glicosídica correspondiendo a glicoproteínas tipo mucina; en F2 se encontró una relación molar O-/N- es 5,3/1. La fracción de F1 de individuos azoospérmicos y oligozoospérmicos son del tipo O-glicosídico mientras que en vasectomizados la relación molar O-/N- es 1,7/1. En todos los casos, F2 presenta cadenas de oligosacáridos O- y N-glicosídicas en diferentes proporciones, con la excepción del plasma seminal de vasectomizado que pertenece al tipo O-glicosídico. Los resultados obtenidos sugieren que la composición de las fracciones F1 y F2 de donantes con diferentes patologías está caracterizada por diferencias importantes en el contenido de carbohidratos y pépticos

Contenido de glicoproteínas del humor vítreo humano / Glycoproteins content of the human vitreous human

Evaluar el contenido de glicoproteínas en el vítreo y determinar las posibles modificaciones en relación al tiempo de fallecimiento, edad. Se obtuvo humor vítreo (HV) de 120 pacientes (60 hombres y mujeres), en edades comprendidas entre 20-40 años fallecidos recientemente por causas que no afectaban los tejidos oculares. Los resultados fueron sometidos a los análisis estadísticos habituales, calculándose las medias, desviaciones y los correspondientes errores estándares. Se practicó análisis de varianza (ANAVA) de los factores y tres períodos (4, 6 y 8 horas) a un nivel de significación del 0,01. Los valores promedios de fucosa, de PBH y de ácido siálico en los 120 HV fueron de 0,8798 ñ 0,1189; 8.18 ñ 0,62 y 13,09 ñ 0,50 mg/dl, sin diferencias entre los sexos. El ANAVA de dos vías demuestra que ni el sexo, ni el tiempo influyen significativamente sobre el contenido vítreo de fucosa, de PBH y de ácido siálico. La edad tampoco ejerce una influencia significativa sobre la composición glicoproteíca de HV, como lo demostró el análisis de varianza de una vía (f= 0,08 gl= 4/119 no significativo). El contenido de glicoproteínas en el HV humano, con pocas excepciones es muy similar al de la mayoría de las especies animales. La concentración de proteínas en el HV no es influenciada ni por el sexo ni por el tiempo de recolección postmortem. Entre 20 y 40 años no existe una influencia significativa de la edad sobre el contenido de glicoproteína vítrea

Study of serum fucose and serum sialic acid levels in oral squamous cell carcinomia.

Serum fucose and sialic acid levels were determined in 50 oral cancer patients and 25 healthy controls. A statistically significant increase was noted in the study group. The increase in serum fucose level correlated well with the clinical staging in the study group whereas sialic acid did not. These values were independent of age, sex and histopathological grading. The result suggest that the serum fucose level is a better biochemical tumor marker than sialic acid level. However its usefulness may be limited if used judiciously to assess the prognosis of the disease.

The FML vaccine (fucose-mannose ligand) protects hamsters from experimental kala-azar

Visceral leishmaniais or kala-azar, is a chronic and frequently lethal disease, caused by Leishmania donovani. Clinical signs include malaise, hepatosplenomegaly, hypergammaglobulinemia, fever, cachexia and progressive suppresion of the cellular immune response. Only few studies on prophylactic immunization against this disease have been understaken, mostly with crude antigens, and no vaccine against kala-azar is yet available. In previous studies, we have isolated the Fucose-Mannose Ligand (FML) of L. donovani that strongly and specifically inhibits the in vitro infection of macrophages by promastigotes and amastigotes. The FML behaves as a pontent immunogen for rabbits and mice, and is specifically recognized by kala-azar patient sera. The protective pontential of FML on kala-azar was now analyzed in the CB-hamster model. We studied the efect of three intraperitoneal weekly doses of FML (100 mg) in saponin (100 mg), folowed by an intracardiac injection of 107 amastigotes. Saponin- and saline-treated controls were also included. Protection was highly significant regarding the enhancement of anti-FML antibodies titers, the splenocyte proliferative response, and the intradermal delayed hypersensitivity reaction to antigen, as well as the decrease of the parasite burden in spleen and of splenomegaly. Protection to kala-azar was due to specific FML antigenic properties, since the results obtained by saponin alone were significantly different. We conclude that the use of FML and saponin as a vaccine reduced the disease impact and retarded its onset.

Fucosidosis: una enfermedad por almacenamiento lisosomal / Fucosidosis: a lysosomal storage disease

Se describen 3 pacientes: 2 del sexo masculino y 1 del femenino, con enfermedad por almacenamiento lisosomal del tipo fucosidosis. En los 3 casos existió el antecedente de retardo psicomotor y posteriormente se hizo evidente el deterioro progresivo de las capacidades psíquicas motoras adquiridas. Todos tenían facies toscas, retardo mental profundo y alteraciones morfológicas en vértebras lumbares. Desde el punto de vista bioquímico, los 3 pacientes presentaron excreción urinaria de oligisacáridos ricos en fucosa, así como disminución de la actividad de la enzima alfa-L-fucosidasa en leucocitos. Se hace énfasis en la demostración del defecto enzimático como único criterio diagnóstico confirmatorio

Metabolic analysis of galactose toxicity in Escherichia coli with 2-deoxygalactose as the probe.

Biochemical basis of galactose toxicity has been studied in gal T mutants (CGSC 4974) using 2-deoxygalactose, a non-metabolizable analogue of galactose, as the probe. It is found that biochemical features of toxicity in wild type cells either with 2-deoxygalactose or with 2-deoxyglucose are very similar to the picture obtained with gal T mutants and the observed bacteriostasis is probably due to futile phosphorylation and not due to any specific inhibitory effect of phosphorylated galactose.

Serum fucose levels in gynaecological disorders including carcinoma cervix.

Serum protein bound fucose levels and the ratio between serum protein bound fucose and total serum protein were determined in 40 healthy females, 60 patients with benign gynaecological disorders and 50 patients with previously untreated cases of carcinoma cervix. The mean serum fucose level in healthy subjects was 8.96 mg% and the ratio between serum protein bound fucose and total serum protein was 1.29 x 10(-3). Mean serum fucose level in patients with benign gynaecological disorders was 9.42 mg% and the ratio between serum protein bound fucose and total serum protein was 1.42 x 10(-3). In stage I and II carcinoma cervix patients mean serum fucose was 11.92 mg% and the ratio between serum protein bound fucose and total serum protein was 1.68 x 10(-3). In stage III and IV carcinoma cervix patients the mean serum fucose was 17.76 mg% and the ratio between serum protein bound fucose and total serum protein was 2.64 x 10(-3).