Two positional isomers of sialylheptasaccharides isolated from the urine of a patient with sialidosis

We describe the structures of two positional isomers of sialylheptasaccharide isolated from the urine of a patient with sialidosis with partial deficiency of beta-galactosidase. Based on structural studies including compositional sugar analysis, exoglycosidase digestion, chemical ionization mass spectrometry, proton nuclear magnetic resonance spectrometry, and methylation analysis, their structures were deduced to be as follows: AcNeu alpha 2----6Gal beta 1----4GlcNac beta 1----2Man alpha 1----3(Man alpha 1----6)Man beta 1----4GlcNac; AcNeu alpha 2----6Gal beta 1----4GlcNac beta 1----2Man alpha 1----6(Man alpha 1----3)Man beta 1----4GlcNac. Sialyloligosaccharide 1 has previously been found in the urine and liver of patients with mucolipidosis I and II and sialidosis, but sialyloligosaccharide 2 has not been found yet in human urine. These two sialyloligosaccharides could not be completely separated by any chromatographic procedures tested. The analytical techniques, including methylation study and NMR spectroscopy, could not clearly detect the differences between them. However, alpha-mannosidase treatment gave important information for the structural analyses of these sialyloligosaccharides.     

Lysosomal acid lipase deficiency in rats: lipid analyses and lipase activities in liver and spleen

We report the biological characterization of an animal model of a genetic lipid storage disease analogous to human Wolman's disease. Affected rats accumulated cholesteryl esters (13.3-fold), free cholesterol (2.8-fold), and triglycerides (5.4-fold) in the liver, as well as cholesteryl esters (2.5-fold) and free cholesterol (1.33-fold) in the spleen. Triglycerides did not accumulate, and the levels actually decreased in the spleen. Analysis of the fatty acid composition of the cholesteryl esters and triglycerides showed high percentages of linoleic acid (18:2) and arachidonic acid (20:4) in both organs, especially in the liver. No accumulation of phospholipids, neutral glycosphingolipids, or gangliosides was found in the affected rats. Acid lipase activity for [14C]triolein, [14C]cholesteryl oleate, and 4-methyl-umbelliferyl oleate was deficient in both the liver and spleen of affected rats. Lipase activity at neutral pH was normal in both liver and spleen. Heterozygous rats showed intermediate utilization of these substrates in both organs at levels between those for affected rats and those for normal controls, although they did not accumulate any lipids. These data suggest that these rats represent an animal counterpart of Wolman's disease in humans.     

Phylogeography of the temperate grassland plant Tephroseris kirilowii (Asteraceae) inferred from multiplexed inter-simple sequence repeat genotyping by sequencing (MIG-seq) data

Journal of Plant Research - A group of temperate grassland plant species termed the “Mansen elements” occurs in Japan and is widely distributed in the grasslands of continental East...     

Cloning and expression of a porcine UDP-GalNAc: polypeptideN-acetylgalactosaminyl transferase

By employing a bovine UDP-N-acetylgalactosamine: polypeptideN-acetylgalactosaminyl transferase (O-GalNAc transferase) cDNA as a probe, we isolated four overlapping cDNAs from a porcine lung cDNA library. Both the nucleotide sequence of the porcine cDNA and the predicted primary structure of the protein (559 amino acids) proved to be very similar to those of the bovine enzyme (95% and 99% identity, respectively). Transient expression of the clone in COS-7 cells, followed by enzymatic activity assays, demonstrated that this cDNA sequence encodes a porcine O-GalNAc transferase. The intracellular O-GalNAc transferase activity was increased approximately 100-fold by transfecting cells with the porcine cDNA.Abbreviations O-GalNAc transferase UDP-N-acetylgalactosamine: polypeptideN-acetylgalactosaminyltransferase - PCR polymerase chain reaction - SDS sodium dodecyl sulfate - PAGE polyacrylamide gel electrophoresis - GnT-III UDP-N-acetylglucosamine: -mannoside -1,4N-acetylglucosaminyltransferase III     

Estimation of the fertility rates of Japanese wild boars (Sus scrofa leucomystax) using fetuses and corpora albicans

Effective population control of Japanese wild boar (Sus scrofa leucomystax) requires reliable information about population dynamics. Fertility rate is the fundamental component of reproduction to evaluate population dynamics. However, little is known regarding the fertility rate of Japanese wild boar. The traditional hunting practices make it difficult to obtain pregnant females and calculate the fertility rate by checking fetuses as is performed in other countries. Therefore, we focused on the corpora albicans (CA) as the CA remains in the ovaries of postpartum females after pregnancy. This study aimed to evaluate the utility of CA and estimate the fertility rate of Japanese wild boars using CA. Histological analysis of ovaries enabled us to discriminate type 1 CA, which remains for 1 year after breeding. Type 1 CA is a superior indicator compared with lactation in the non-pregnancy season because it allows verification of postpartum females over a long period. The fertility rate was calculated by the combination of pregnant and postpartum females using fetuses and type 1 CA from April to November. The fertility rate of the females captured after the second pregnancy season was 90.3 % during the pregnancy period and 100 % during the non-pregnancy period. The high fertility rate of adult females suggests that intensive adult female harvesting is needed. Our new method to determine fertility rates contributes to developing a monitoring system to adequately control Japanese wild boar population.     

Taurine inhibits apoptosis by preventing formation of the Apaf-1/caspase-9 apoptosome

Cardiomyocyte apoptosis contributes to cell death during myocardial infarction. One of the factors that regulate the degree of apoptosis during ischemia is the amino acid taurine. To study the mechanism underlying the beneficial effect of taurine, we examined the interaction between taurine and mitochondria-mediated apoptosis using a simulated ischemia model with cultured rat neonatal cardiomyocytes sealed in closed flasks. Exposure to medium containing 20 mM taurine reduced the degree of apoptosis following periods of ischemia varying from 24 to 72 h. In the untreated group, simulated ischemia for 24 h led to mitochondrial depolarization accompanied by cytochrome c release. The apoptotic cascade was also activated, as evidenced by the activation of caspase-9 and -3. Taurine treatment had no effect on mitochondrial membrane potential and cytochrome c release; however, it inhibited ischemia-induced cleavage of caspase-9 and -3. Taurine loading also suppressed the formation of the Apaf-1/caspase-9 apoptosome and the interaction of caspase-9 with Apaf-1. These findings demonstrate that taurine effectively prevents myocardial ischemia-induced apoptosis by inhibiting the assembly of the Apaf-1/caspase-9 apoptosome. ischemia; cultured cardiomyocytes     

IRS-1 transgenic mice show increased epididymal fat mass and insulin resistance

Insulin receptor substrate-1 (IRS-1) is the major substrate of both the insulin receptor and the IGF-1 receptor. In this study, we created IRS-1 transgenic (IRS-1-Tg) mice which express human IRS-1 cDNA under control of the mouse IRS-1 gene promoter. In the IRS-1-Tg mice, IRS-1 mRNA expression was significantly increased in almost all tissues, but its protein expression was increased in very limited tissues (epididymal fat and skeletal muscle). IRS-1-Tg mice showed glucose intolerance and significantly enlarged epididymal fat mass, as well as elevated serum TNF-α concentrations. Importantly insulin signaling was significantly attenuated in the liver of IRS-1-Tg mice, which may contribute to the glucose intolerance. Our results suggest that excess IRS-1 expression may not provide a beneficial impact on glucose homeostasis in vivo.     

Plant regeneration via direct and indirect adventitious shoot formation and chromosome-doubled somaclonal variation in Titanotrichum oldhamii (Hemsl.) Solereder

The gesneriaceous perennial plant Titanotrichum oldhamii has beautiful foliage and attractive bright yellow flowers. However, breeding of T. oldhamii by conventional sexual hybridization may be difficult because sexual reproduction of this species is very rare. In the present study, plant regeneration systems via both direct and indirect formation of adventitious shoots from leaf explants were established as the first step toward breeding T. oldhamii by using biotechnological techniques. Adventitious shoots were formed efficiently on medium containing 0.1 mg l⁻¹ benzyladenine. Histological observation showed that shoot formation on this medium occurred directly from leaf epidermal cells without callus formation. On the other hand, leaf explants formed calluses on medium containing 0.1 mg l⁻¹ 2,4-dichlorophenoxyacetic acid. The calluses could be maintained by monthly subculturing to fresh medium of the same composition. When the calluses were transferred to plant growth regulator-free medium, they formed adventitious shoots. Directly and indirectly formed shoots rooted well on medium containing 0.1 mg l⁻¹ indole-3-butyric acid. Plantlets thus obtained were successfully acclimatized and grew vigorously in the greenhouse. Flow cytometry analysis indicated that no variation in the ploidy level was observed in plants regenerated via direct shoot formation, whereas chromosome doubling occurred in several plants regenerated via indirect shoot formation. Regenerated plants with the same ploidy level as the mother plants showed almost the same phenotype as the mother plants, whereas chromosome-doubled plants showed apparent morphological alterations: they had small and crispate flowers, and round and deep green leaves.     

N-cadherin-mediated cell adhesion determines the plasticity for cell alignment in response to mechanical stretch in cultured cardiomyocytes

Mechanical stretch has been implicated as the growth stimuli in the heart. Physiologically, mechanical stretch is reported to contribute to the orientation of cardiomyocytes, though the molecular mechanism remains to be elucidated. This study was designed to make clear functional significances of N-cadherin in plasticity of cell alignment in response to mechanical stretch. Neonatal rat cardiomyocytes, cultured on silicone dishes, were subjected to artificial uniaxial cyclic stretch. Mechanical stretch was started at certain times (3-75h) after seeding and continued for 24h. Stretch stimulation in 3h after cultivation promoted cell orientation running parallel to tension direction. In contrast, cardiac myocytes fail to align when exposed to stretch 24-75h after cultivation. To address the importance of N-cadherin in the responsiveness to stretch, the expression and distribution of N-cadherin were analyzed. Immediately after seeding, N-cadherin showed dispersed distributions. During cultivation, N-cadherin localized to cell-cell contacts accompanied by the upregulation of its protein. Next, to investigate influence of cell-cell adhesion, cardiomyocytes cultured for 72h were replated by trypsin treatment and exposed to stretch 3h after replating. The cardiomyocytes replated by trypsinization were oriented in parallel to tension direction by mechanical stretch. Finally, adenoviral transfection of dominant-negative N-cadherin recovered the ability to exhibit cell orientation in response to stretch. Our results suggested that N-cadherin was involved in the oriented responses of cardiomyocytes induced by mechanical stretch.