Lipid droplets are formed in 2-cell-like cells

Embryonic stem (ES) cells, derived from the inner cell mass of a blastocyst, are believed to pluripotent cells and give rise to embryonic, but not extraembryonic, tissues. In mice, totipotent 2-cell stage embryo-like (2-cell-like) cells, which are identified by reactivation of murine endogenous retrovirus with leucin transfer RNA primer (MuERV-L), arise at a very few frequencies in ES cell cultures. Here, we found that a lipid droplet forms during the transition from ES cells to 2-cell-like cells, and we propose that 2-cell-like cells utilize a unique energy storage and production pathway.     

The effects of medetomidine and xylazine on gastrointestinal motility and gastrin release in the dog

The effect of medetomidine, a potent and highly selective α₂-adrenoceptor agonist, on the motility of the gastric antrum, duodenum, mid-jejunum and ileum was investigated in ten dogs. Its effect on the release of gastrin was also determined. Administration of medetomidine intramuscularly (i.m.) at a dose of 40 μg/kg inhibited the motility of the gastric antrum, duodenum, mid-jejunum and ileum significantly, in comparison to administration of xylazine intramuscularly at a dose of 2.0 mg/kg. The release of gastrin was also significantly decreased in dogs receiving medetomidine. It was found to inhibit the motility in the gastric antrum and duodenum longer than in the mid-jejunum and ileum, presumably by acting specifically on α₂-adrenoceptors, likely at the peripheral level. Medetomidine also inhibited the gastric contraction associated with gastrin secretion.     

Glucocorticoid sensitivity depends on expression levels of glucocorticoid receptors in canine neoplastic mast cells

Glucocorticoid (GC) administration with or without other chemotherapeutic reagents is a commonly used option in the treatment of mast cell malignancies. However, the responsiveness of mast cell tumors to GC treatment varies in individuals, and the regulatory mechanisms determining the GC sensitivity of malignant mast cells remain unclear. Since the expression of the GC receptor (GR) has been reported to be associated with GC sensitivity in human neoplastic lymphocytes, we attempted to investigate the relationship between GR levels and GC sensitivity by using neoplastic mast cells derived from canine mast cell tumors (MCTs). To elucidate the regulatory mechanisms involved in GC responsiveness, we analyzed various canine MCT cell lines and tissue samples from dogs with MCT. While the proliferation of canine MCT cells was suppressed by the addition of GC to the culture, we found that MCT cells derived from humans and rodents, as well as canine lymphoma cells, responded poorly to GC. However, there were also some variations in responsiveness to GC treatment among canine MCT cell lines used in this study. Using real-time polymerase chain reaction and Western blot analysis, we elucidated the relationship between GR expression and responsiveness to GC in canine MCT cells. Furthermore, to assess the involvement of GR expression in GC sensitivity in vivo, clinical investigations were conducted on dogs with cutaneous MCT. Written informed consent was obtained from owners, and the affected dogs were treated with prednisolone (0.5-2.0 mg kg(-1)day(-1), administered orally) 1 or 2 weeks prior to the surgical removal of the tumors. Tumor volume was measured according to WHO criteria both before and after prednisolone treatment, and the GC sensitivity of each MCT was determined on the basis of the reduction in tumor volume. Of the 15 dogs with MCT, 11 responded to treatment with prednisolone completely or partially, whereas 4 dogs showed no response. Examination of clinical samples obtained by surgical removal revealed that GR expression levels were significantly lower in GC-resistant MCT tissues than in GC-sensitive MCT tissues. Thus, these results strongly indicate that GR expression may contribute to GC sensitivity in canine MCT.     

Molecular cloning, nucleotide sequence and characteristics of a xylanase gene (xynA) from Ruminococcus albus 7

A 2.6-kb DNA fragment encoding a xylanase gene ( xyn A) was cloned from the rumen hemicellulolytic bacterium Ruminococcus albus 7. The deduced primary structure of the protein (XynA) was divided into a signal peptide region and 3 domains. Domain A was identified as a family 11 (G) catalytic domain, but one amino acid residue was replaced by another in an active site signature 1 of family 11. Domain B is a stabilizing domain for the catalytic domains of families 10 and 11. Deletion of domain B reduced stability of the xylanase at high temperature and at high and low pH. Domain B may be useful for protein engineering of xylanase. Domain C has sequence similarity to deacetylases and NodB proteins.     

In vitro effects of insulin on glucose and lipid metabolism in rat embryos

Glucose is utilized for oxidation and synthesis of various lipids in cultured rat embryos. The present experiment examined the effect of insulin on the incorporation of glucose into lipid fractions in rat embryos in vitro . Embryos at the 2-cell, 8-cell and blastocyst stages were incubated for 5 h in hamster embryo culture medium (HECM)-1 containing ¹⁴C-glucose and 170 nmol/L insulin, or in HECM-1 containing only ¹⁴C-glucose, and the oxidation of glucose in these embryos was examined. In addition, the total lipids of blastocysts were separated by thin layer chromatography and the radioactivity of the separated lipid fractions was measured. Oxidation of glucose was significantly increased after insulin treatment compared with that without insulin treatment in 8-cell embryos and blastocysts ( P  < 0.05), but not in 2-cell embryos. Incorporation of glucose into lipids in blastocysts was significantly lowered by insulin treatment compared with that without insulin treatment ( P  < 0.05). Most of the radioactivity was recovered from triacylglycerols of blastocysts and the remaining radioactivity was found in other neutral lipids and phospholipids. We conclude that insulin accelerates the utilization for oxidation of glucose and inhibits the storage of triacylglycerols in rat blastocysts.     

Urinary free metanephrines measurement in dogs with adrenal gland diseases using a new simple liquid chromatography tandem mass spectrometry method

Measurement of urinary metanephrines in spot samples is used for the diagnosis of canine pheochromocytoma (PC). We describe a simple analytical method based on liquid chromatography tandem mass spectrometry (LC-MS/MS) for measuring free metanephrine (MN) and normetanephrine (NMN) in spot urine samples. Using the developed method, we evaluated the stability of urinary free-MN and free-NMN at various storing conditions. In addition, we assessed the feasibility of urinary free-MN and -NMN measurement for diagnosing PC. Urine samples were mixed with stable isotope internal standards and thereafter purified by ultrafiltration. The purified samples were analyzed by LC-MS/MS in the multiple reaction monitoring mode after separation on a multimode octa decyl silyl column. The coefficient of variation of free-MN and -NMN measurement was 7.6% and 5.5%, respectively. The linearity range was 0.5–10 µg/l for both analytes. Degradation was less than 10% for both analytes under any of the storage conditions. The median free-NMN ratio to creatinine of 9 PC dogs (595, range 144–47,961) was significantly higher (P<0.05) than that of 13 dogs with hypercortisolism (125, range 52–224) or 15 healthy dogs (85, range 50–117). The developed method is simple and may not require acidification of spot urine. The results of this preliminary retrospective study suggest that the measurement of urinary free metanephrines is a promising tool for diagnosing canine PC.     

Radiocesium interception potential (RIP) of smectite and kaolin reference minerals containing illite (micaceous mineral) as impurity

Cesium-137 (¹³⁷Cs) is strongly adsorbed on clay minerals, especially on illite. The adsorption of Cs⁺ on reference clay minerals, however, has not been fully investigated in relation to the presence of illite. The objective of this study was to clarify the effect of impurities (i.e., illite and vermiculite), present in reference smectite group minerals and kaolin minerals, on the retention of Cs⁺. The clay mineralogy of the reference minerals was characterized by X-ray diffraction (XRD). The radiocesium interception potential (RIP) was measured as an index of the Cs⁺ retention ability of clays. The content of illite in clay was represented by the total potassium (K) content given that illite is a major source of K in the clay fraction. The content of vermiculite in clay was represented by the Cs fixation capacity induced by Cs saturation followed by heating of samples at 110°C. Metabentonite and beidellite gave extremely high RIP values compared with other smectite group minerals, although a peak for illite (at 1.0 nm) was not observed in XRD analysis. The reference smectite and kaolin minerals showed a range of RIP values, even though their RIP values are theoretically zero. The RIP values had a significant positive correlation with the total K content of all the reference clay minerals (r_s = 0.621*). This indicated that the retention ability for ¹³⁷Cs depended more on the content of illite, as impurity, rather than the type of bulk mineral. Hence, the contribution of illite to the magnitude of the RIP was elucidated by the combination of measurement of total K content and XRD analysis.     

A critical role for the innate immune signaling molecule IRAK-4 in T cell activation

IRAK-4 is a protein kinase that is pivotal in mediating signals for innate immune responses. Here, we report that IRAK-4 signaling is also essential for eliciting adaptive immune responses. Thus, in the absence of IRAK-4, in vivo T cell responses were significantly impaired. Upon T cell receptor stimulation, IRAK-4 is recruited to T cell lipid rafts, where it induces downstream signals, including protein kinase C activation through the association with Zap70. This signaling pathway was found to be required for optimal activation of nuclear factor kappaB. Our findings suggest that T cells use this critical regulator of innate immunity for the development of acquired immunity, suggesting that IRAK-4 may be involved in direct signal cross talk between the two systems.