Regulation of chondroitin sulfate biosynthesis by specific sulfation: acceptor specificity of serum {beta}-GalNAc transferase revealed by structurally defined oligosaccharides

The relationship between sulfation and polymerization in chondroitinsulfate (CS) biosynthesis has been poorly understood. In thisstudy, we investigated the specificity of bovine serum UDP-GalNAc:CS ß-GalNAc transferase responsible for chain elongationusing structurally defined acceptor substrates. They consistedof tetra- and hexasaccharide-serines that were chemically synthesizedand various regular oligosaccharides with a GlcA residue atthe nonreducing terminus, prepared from chondroitin and CS usingtesticular hyaluronidase. The enzyme preparation was obtainedfrom fetal bovine serum by means of heparin-Sepharose affinitychromatography. The preparation did not contain the     

Immunohistochemical localization and possible physiological roles of tumor necrosis factor-α in mouse cumulus-oocyte complexes

Tumor necrosis factor-α (TNF-α), a cytokine which is produced by activated macrophages, has been shown to participate in the regulation of ovarian functions. In the course of our investigation on the mechanism of maturation, fertilization and degeneration of mouse oocytes, immunoreactivity to TNF-α was found in the cytoplasm of the cells surrounding the maturing oocytes and of granulosa cells facing the antral cavity. Immunoblot analysis with the specific antibody to TNF-α identified the 17 kDa Mr band in the extract of cumulusoocyte complexes. Various concentrations of TNF-α (mouse, recombinant) and anti TNF-α antiserum (polyclonal rabbit anti-mouse recombinant TNF-α) were then used to determine their effect on the germinal vesicle breakdown (GVBD), polar body extrusion, fertilization and fragmentation of mouse oocytes/eggs. TNF-α at concentrations of 10 ng/mL or less and anti-TNF-α antiserum at concentrations of 10% or less, had no effect on the spontaneous GVBD and polar body extrusion of mouse oocytes in culture. Mouse follicular oocytes cultured for more than 72 h in modified Krebs-Ringer solution in vitro undergo spontaneous fragmentation, which is a degenerative change to form 'blastomeres' with or without nuclear fragments or chromatin. Ghost-like blastomeres were also identified in the space among fragmented 'blastomeres'. The spontaneous fragmentation of mouse follicular oocytes was suppressed in the presence of TNF-α at concentrations of 1 ng/mL or greater. Anti-TNF-α antiserum (1%) accelerated the induction of fragmentation of oocytes cultured in vitro . The addition of anti TNF-α antiserum (10%) to the culture medium did not influence the fertilization rates of the eggs surrounded by the expanded cumulus. These results appear to indicate that the process of degeneration of mouse oocytes/eggs is modulated by TNF-α accumulated in the expanded cumulus during oocyte maturation.     

Daily protein and energy intakes of infants fed a commercial infant formula with a reduced protein concentration of 2.2 g/100 kcal: an impact of feeding interval on energy intake

ABSTRACT

We evaluated the protein and energy intakes of infants fed commercial infant Formula A (protein, 2.2 g/100 kcal; energy, 68 kcal/100 mL) and examined whether changes in feeding intervals are involved in constant energy intake. Daily nutritional intake of 378 Formula A-fed infants was assessed using reference values and compared to that of infants fed Formulas B (protein: 2.3 g/100 kcal, energy: 68 kcal/100 mL) and C (protein: 2.4 g/100kcal, energy: 70 kcal/100 mL). From 15 to 149 days of age, the mean formula volume and protein intake were 758–887 mL/day and 11.4–13.3 g/day, respectively, higher than the protein intake of breast-fed infants. Daily energy intake (86–129 kcal/kg/day) was comparable to the estimated energy requirements. Feeding intervals were shorter in infants fed Formulas A and B than in those fed Formula C, whereas energy intake was similar. The protein intake of infants decreased as the protein concentration per energy in infant formula was reduced, and accordingly the protein intake of Formula A-fed infants was significantly lower than that of Formula C-fed infants. In conclusion, the new composition of Formula A is suitable in protein and energy intake of infants, and daily energy intake remains constant by shortening in feeding intervals when the energy concentration in infant formula is reduced.

Clinical Trial Registration: UMIN000023110     

Ultrastructural characterization of microlipophagy induced by the interaction of vacuoles and lipid bodies around generative and sperm cells in Arabidopsis pollen

Protoplasma - During pollen maturation, various organelles change their distribution and function during development as male gametophytes. We analyzed the behavior of lipid bodies and vacuoles...     

Alleviation of High-Fat Diet-Induced Fatty Liver Damage in Group IVA Phospholipase A2-Knockout Mice

Hepatic fat deposition with hepatocellular damage, a feature of non-alcoholic fatty liver disease, is mediated by several putative factors including prostaglandins. In the present study, we examined whether group IVA phospholipase A₂ (IVA-PLA₂), which catalyzes the first step in prostanoid biosynthesis, is involved in the development of fatty liver, using IVA-PLA₂-knockout mice. Male wild-type mice on high-fat diets (20% fat and 1.25% cholesterol) developed hepatocellular vacuolation and liver hypertrophy with an increase in the serum levels of liver damage marker aminotransferases when compared with wild-type mice fed normal diets. These high-fat diet-induced alterations were markedly decreased in IVA-PLA₂-knockout mice. Hepatic triacylglycerol content was lower in IVA-PLA₂-knockout mice than in wild-type mice under normal dietary conditions. Although high-fat diets increased hepatic triacylglycerol content in both genotypes, the degree was lower in IVA-PLA₂-knockout mice than in wild-type mice. Under the high-fat dietary conditions, IVA-PLA₂-knockout mice had lower epididymal fat pad weight and smaller adipocytes than wild-type mice. The serum level of prostaglandin E₂, which has a fat storage effect, was lower in IVA-PLA₂-knockout mice than in wild-type mice, irrespective of the kind of diet. In both genotypes, high-fat diets increased serum leptin levels equally between the two groups, but did not affect the serum levels of adiponectin, resistin, free fatty acid, triacylglycerol, glucose, or insulin. Our findings suggest that a deficiency of IVA-PLA₂ alleviates fatty liver damage caused by high-fat diets, probably because of the lower generation of IVA-PLA₂ metabolites, such as prostaglandin E₂. IVA-PLA₂ could be a promising therapeutic target for obesity-related diseases including non-alcoholic fatty liver disease.     

Paradesulfovibrio onnuriensis gen. nov., sp. nov., a chemolithoautotrophic sulfate-reducing bacterium isolated from the Onnuri vent field of the Indian Ocean and reclassification of Desulfovibrio senegalensis as Paradesulfovibrio senegalensis comb. nov.

An anaerobic, rod-shaped, mesophilic, chemolithoautotrophic, sulfate-reducing bacterial strain IOR2T was isolated from a newly found deep-sea hydrothermal vent (OVF, Onnuri Vent Field) area in the central Indian Ocean ridge (11°24′88″ S 66°25′42″ E, 2021 m water depth). The 16S rRNA gene sequence analysis revealed that the strain IOR2T was most closely related to Desulfovibrio senegalensis BLaC1T (96.7%). However, it showed low similarity with the members of the family Desulfovibrionaceae, such as Desulfovibrio tunisiensis RB22T (94.0%), D. brasiliensis LVform1T (93.9%), D. halophilus DSM 5663T (93.7%), and Pseudodesulfovibrio aespoeensis Aspo-2T (93.2%). The strain IOR2T could grow at 23–42°C (optimum 37°C), pH 5.0–8.0 (optimum pH 7.0) and with 0.5–6.5% (optimum 3.0%) NaCl. The strain could use lactate, pyruvate, H2, and glycerol as electron donors and sulfate, thiosulfate, and sulfite as electron acceptors. The major fatty acids of the strain IOR2T were iso-C15:0, iso-C17:0, ante-iso-C15:0, and summed feature 9 (C16:0 methyl/iso-C17:1ω9c). Both the strains IOR2T and BLaC1T could grow with CO2 and H2 as the sole sources of carbon and energy, respectively. Genomic evidence for the Wood-Ljungdahl pathway in both the strains reflects chemolithoautotrophic growth. The DNA G + C content of the strain IOR2T and BLaC1T was 58.1–60.5 mol%. Based on the results of the phylogenetic and physiologic studies, Paradesulfovibrio onnuriensis gen. nov., sp. nov. with the type strain IOR2T (= KCTC 15845T = MCCC 1K04559T) was proposed to be a member of the family Desulfovibrionaceae. We have also proposed the reclassification of D. senegalensis as Paradesulfovibrio senegalensis comb. nov.     

Fertile Arabidopsis cyp704b1 mutant,defective in sporopollenin biosynthesis,has a normal pollen coat and lipidic organelles in the tapetum

The exine acts as a protectant of the pollen from environmental stresses, and the pollen coat plays an important role in the attachment and recognition of the pollen to the stigma. The pollen coat is made of lipidic organelles in the tapetum. The pollen coat is necessary for fertility, as pollen coat-less mutants, such as those deficient in sterol biosynthesis, show severe male sterility. In contrast, the exine is made of sporopollenin precursors that are biosynthesized in the tapetum. Some mutants involved in sporopollenin biosynthesis lose the exine but show the fertile phenotype. One of these mutants, cyp704b1, was reported to lose not only the exine but also the pollen coat. To identify the cause of the fertile phenotype of the cyp704b1 mutant, the detailed structures of the tapetum tissue and pollen surface in the mutant were analyzed. As a result, the cyp704b1 mutant completely lost the normal exine but had high-electron-density granules localized where the exine should be present. Furthermore, normal lipidic organelles in the tapetum and pollen coat embedded between high-electron-density granules on the pollen surface were observed, unlike in a previous report, and the pollen coat was attached to the stigma. Therefore, the pollen coat is necessary for fertility, and the structure that functions like the exine, such as high-electron-density granules, on the pollen surface may play important roles in retaining the pollen coat in the cyp704b1 mutant.     

Chick embryo chorioallantoic membrane (CAM): an alternative predictive model in acute toxicological studies for anti-cancer drugs

The chick embryo chorioallantoic membrane (CAM) is a preclinical model widely used for vascular and anti-vascular effects of therapeutic agents in vivo. In this study, we examine the suitability of CAM as a predictive model for acute toxicology studies of drugs by comparing it to conventional mouse and rat models for 10 FDA-approved anticancer drugs (paclitaxel, carmustine, camptothecin, cyclophosphamide, vincristine, cisplatin, aloin, mitomycin C, actinomycin-D, melphalan). Suitable formulations for intravenous administration were determined before the average of median lethal dose (LD₅₀) and median survival dose (SD₅₀) in the CAM were measured and calculated for these drugs. The resultant ideal LD₅₀ values were correlated to those reported in the literature using Pearson’s correlation test for both intravenous and intraperitoneal routes of injection in rodents. Our results showed moderate correlations (r²=0.42 − 0.68, P<0.005–0.05) between the ideal LD₅₀ values obtained using the CAM model with LD₅₀ values from mice and rats models for both intravenous and intraperitoneal administrations, suggesting that the chick embryo may be a suitable alternative model for acute drug toxicity screening before embarking on full toxicological investigations in rodents in development of anticancer drugs.     

Generation of monoclonal antibodies to vertebrate albumins for analysis of arthropod blood meals

An immunoassay using monoclonal antibodies (MAbs) that are specific for different vertebrate taxa (from class to species) has been developed that simplifies and facilitates analysis of vertebrate blood meals from arthropod vectors. The MAbs have been prepared against the single protein albumin, the most abundant protein in vertebrate sera. A panel of these antibodies has been generated against albumins from 33 species of vertebrates, representing four classes, 15 orders, and 25 families. Immunoreactivity of albumin in mosquito blood meals can be detected as late as 48 h after feeding. Immunoassays with MAbs can be carried out in the field as well as the laboratory. Used in conjunction with nucleic acid assays or used alone with an appropriate assortment of antibodies, the assay is simple, sensitive, and unambiguous.