Enzymatic sulfation of galactosyl- and lactosylceramides in cell lines derived from renal tubules.

1. The renal cell lines, JTC-12 and MDCK, not only synthesize galactosylceramide 3-sulfate and lactosylceramide 3'-sulfate in vivo, but also contain enzymes that catalyze the transfer of sulfate to galactosylceramide and lactosylceramide in vitro. 2. Concentration of cations necessary for maximum sulfotransferase activity occurred at 40 mM Ca2+ with galactosylceramide and 15 mM Ca2+ with lactosylceramide as the substrate. Na+ was also found to stimulate the sulfation of galactosylceramide, but was slightly inhibitory for the sulfation of lactosylceramide. 3. The products of the in vitro assay mixture were characterized as galactosylceramide 3-sulfate and lactosylceramide 3'-sulfate by a variety of TLC separations. 4. The apparent Km of JTC-12 cells for galactosylceramide was 17 microM, while that for lactosylceramide was 82 microM. The Km values of MDCK cells were comparable to those of JTC-12 cells. Competition studies suggested that galactosylceramide and lactosylceramide were sulfated by a single enzyme in both cell lines.     

Differential induction of regulatory genes during mesoderm formation in Xenopus laevis embryos

Mesoderm development in Xenopus laevis depends on inductive cell interactions mediated by diffusible molecules. The mesoderm inducer activin is capable of redirecting the development of animal explants both morphologically and biochemically. We have studied the induction of four regulatory genes, Mix. 1, goosecoid (gsc), Xlim-1 and Xbra in such explants by activin, and the influence of other factors on this induction. Activin induction of gsc is strongly enhanced by dorsalization of the embryo by LiCl, while expression of the other genes is only slightly enhanced. The protein synthesis inhibitor cycloheximide (CHX) inhibits the activin-dependent induction of Xbra partially, while induction of Mix. 1 and Xlim- 1 is essentially unaffected. In contrast, gsc shows strong superinduction in the presence of activin and CHX, and can be induced in animal explants by CHX alone. Induction and superinduction by CHX have previously been observed for immediate early genes in a variety of systems, notably for the activation of c-fos expression by serum stimulation, but have not been reported in early amphibian embryos. © 1993Wiley-Liss, Inc.     

Gait analysis using gravitational acceleration measured by wearable sensors

A novel method for measuring human gait posture using wearable sensor units is proposed. The sensor units consist of a tri-axial acceleration sensor and three gyro sensors aligned on three axes. The acceleration and angular velocity during walking were measured with seven sensor units worn on the abdomen and the lower limb segments (both thighs, shanks and feet). The three-dimensional positions of each joint are calculated from each segment length and joint angle. Joint angle can be estimated mechanically from the gravitational acceleration along the anterior axis of the segment. However, the acceleration data during walking includes three major components; translational acceleration, gravitational acceleration and external noise. Therefore, an optimization analysis was represented to separate only the gravitational acceleration from the acceleration data. Because the cyclic patterns of acceleration data can be found during constant walking, a FFT analysis was applied to obtain some characteristic frequencies in it. A pattern of gravitational acceleration was assumed using some parts of these characteristic frequencies. Every joint position was calculated from the pattern under the condition of physiological motion range of each joint. An optimized pattern of the gravitational acceleration was selected as a solution of an inverse problem. Gaits of three healthy volunteers were measured by walking for 20 s on a flat floor. As a result, the acceleration data of every segment was measured simultaneously. The characteristic three-dimensional walking could be shown by the expression using a stick figure model. In addition, the trajectories of the knee joint in the horizontal plane could be checked by visual imaging on a PC. Therefore, this method provides important quantitive information for gait diagnosis.     

Effects of growth on residual stress distribution along the radial depth of cortical cylinders from bovine femurs

Residual stress is defined as the stress that remains in bone tissue without any external forces. This study investigated the effects of growth on residual stress distributions from the surface to deeper regions of cortical cylinders obtained from less-than-one-month-old (Group Y) and two-year-old (Group M) bovine femurs. In these experiments, five diaphysis specimens from each group were used. Residual stress was measured using a high-energy synchrotron white X-ray beam to penetrate X-rays into the deeper region of the bone specimens. The measurements in the cortical cylinders from Groups Y and M were performed at 0.5- and 1-mm intervals, respectively, from the outer surface to the deeper region of the diaphysis specimens at four positions: anterior, posterior, lateral, and medial. The residual stress was calculated on the basis of variation in the interplanar spacing of hydroxyapatite crystals in the bone tissue. According to the results, the diaphysis specimens from Group Y were not subjected to large residual stresses (average −1.2 MPa and 2.4 MPa at the surface region and 1.5 mm depth, respectively). In Group M, the surface region of the diaphysis specimens was subjected to tensile residual stresses (average 6.7 MPa) and the deeper region was subjected to compressive stresses (average −8.2 MPa at 3 mm depth). There was a strong significant difference between both these regions. The value of residual stresses at the surface region of the diaphysis specimens in both the groups had a positive statistical correlation with the cortical thickness at the measured locations.     

Residual stress distribution in rabbit limb bones

The presence of the residual stresses in bone tissue has been noted and the authors have reported that there are residual stresses in bone tissue. The aim of our study is to measure the residual stress distribution in the cortical bone of the extremities of vertebrates and to describe the relationships with the osteon population density. The study used the rabbit limb bones (femur, tibia/fibula, humerus, and radius/ulna) and measured the residual stresses in the bone axial direction at anterior and posterior positions on the cortical surface. The osteons at the sections at the measurement positions were observed by microscopy. As a result, the average stresses at the hindlimb bones and the forelimb bones were 210 and 149 MPa, respectively. In the femur, humerus, and radius/ulna, the residual stresses at the anterior position were larger than those at the posterior position, while in the tibia, the stress at the posterior position was larger than that at the anterior position. Further, in the femur and humerus, the osteon population densities in the anterior positions were larger than those in the posterior positions. In the tibia, the osteon population density in the posterior position was larger than that in the anterior position. Therefore, tensile residual stresses were observed at every measurement position in the rabbit limb bones and the value of residual stress correlated with the osteon population density (r=0.55, P<0.01).     

Identification of MK-1925: A selective,orally active and brain-penetrable opioid receptor-like 1 (ORL1) antagonist

Structure–activity relationship studies directed toward improving the metabolic stability of compound 1 resulted in the identification of 3-[5-(3,5-difluorophenyl)-3-({[(1S,3R)-3-fluorocyclopentyl]amino}methyl)-4-methyl-1H-pyrazol-1-yl]propanenitrile 39 (MK-1925) as a selective, orally available and brain-penetrable opioid receptor-like 1 (ORL1) antagonist. The compound also showed in vivo efficacy after oral dosing. Therefore, compound 39 was selected to undergo further studies as a clinical candidate.     

Normalization of phospholipids concentration of the gastric mucosa was observed in patients with peptic ulcer after eradication of Helicobacter pylori

Background. Phospholipids concentration in the gastric mucosa decreased in patients with Helicobacter pylori infection. The aim of this study is to examine the effects of eradication of H. pylori on decreasing the phospholipids concentration in the gastric mucosa in patients with gastric or duodenal ulcer. Materials and Methods. Phospholipids (phosphatidylcholine, phosphatidylethanolamine, and sphingonomyeline) were measured in biopsy specimens from the antrum and corpus using thin‐layer chromatography. In H. pylori positive patients with gastric ulcer (n = 26) and duodenal ulcer (n = 13), and H. pylori negative controls (n = 20), the biopsy specimens were obtained before and 3 months after eradication. Eradication was performed using lansoprazole, amoxycillin, and clarithromycin. Results. Compared with the H. pylori negative control group, the concentrations of phosphatidylcholine and phosphatidylethanolamine decreased significantly in the gastric ulcer group in both antrum and corpus mucosa, and in the duodenal ulcer group in antrum mucosa. This decrease returned to the control level after eradication. Conclusions. This study demonstrates that the eradication of H. pylori in patients with peptic ulcer normalized the decrease of phosphatidylcholine and phosphatidylethanolamine in the gastric mucosa.     

Selective inhibition of MAO-A in serotonergic synaptosomes by two amphetamine metabolites, p-hydroxyamphetamine and p-hydroxynorephedrine

The present study was carried out mainly to clarify whether the two amphetamine metabolites, p-hydroxyamphetamine (P-OHA) and p-hydroxynorephedrine (p-OHN) are taken up by mouse brain 5-hydroxytryptamine (5-HT) nerve terminals to inhibit type A monoamine oxidase (MAO-A) and then potentiate the abnormal behavior, head-twitch. Of the two metabolites, only intracerebroventricular p-OHA, at 80 μg/mouse, sufficient to cause a head-twitch response (HTR), appreciably inhibited MAO-A activity without affecting MAO-B activity in homogenates of the mouse striatum, hypothalamus and the rest of the forebrain; and p-OHN did not inhibit either type of MAO at the dose tested. Estimation of intra- and extrasynaptosomal MAO-A activity showed that both metabolites significantly inhibited only the intrasynaptosomal deamination of 5-HT by MAO-A with p-OHA being more potent. Taken together with our previous findings, these present results clearly indicate that p-OHA may accumulate in the 5-HT nerve terminals through the uptake system, and concomitantly inhibit MAO-A activity. These actions of p-OHA may increase intraneuronal 5-HT levels and then potentiate 5-HT release to cause interaction with the post-synaptic 5-HT receptors.     

Decreased calcium/calmodulin-dependent protein kinase II and protein kinase C activities mediate impairment of hippocampal long-term potentiation in the olfactory bulbectomized mice

Olfactory bulbectomized (OBX) mice showed significant impairment of learning and memory-related behaviors 14 days after olfactory bulbectomy, as measured by passive avoidance and Y-maze tasks. We here observed a large impairment of hippocampal long-term potentiation (LTP) in the OBX mice. Concomitant with decreased acetylcholinesterase expression, protein kinase C (PKC)alpha autophosphorylation and NR1(Ser-896) phosphorylation significantly decreased in the hippocampal CA1 region of OBX mice. Both PKCalpha and NR1(Ser-896) phosphorylation significantly increased following LTP in the control mice, whereas increases were not observed in OBX mice. Like PKC activities, calcium/calmodulin-dependent protein kinase II (CaMKII) autophosphorylation significantly decreased in the hippocampal CA1 region of OBX mice as compared with that of control mice. In addition, increased CaMKII autophosphorylation following LTP was not observed in OBX mice. Finally, the impairment of CaMKII autophosphorylation was closely associated with reduced pGluR1(Ser-831) phosphorylation, without change in synapsin I (site 3) phosphorylation in the hippocampal CA1 region of OBX mice. Taken together, in OBX mice NMDA receptor hypofunction, possibly through decreased PKCalpha activity, underlies decreased CaMKII activity in the post-synaptic regions, thereby impairing LTP induction in the hippocampal CA1 region. Both decreased PKC and CaMKII activities with concomitant LTP impairment account for the learning disability observed in OBX mice.     

Genetic structure and differentiation of the Japanese extremely long-tailed chicken breed (Onagadori), associated with plumage colour variation: suggestions for its management and conservation

The Onagadori is a distinguished chicken breed that is characterized by an extremely long tail in the male. In this breed, three different plumage colour varieties have been developed (black-breasted white, black-breasted red and white) in which the black-breasted white is believed to be the original colour of the Onagadori, based on historical records. To establish a conservation strategy, 176 birds were genotyped for autosomal microsatellites. Significant genetic distinctness was found between the original (black-breasted white) and two derivative varieties ( F _ST = 0.091 and 0.093). At the same time, a Bayesian model-based clustering revealed that the majority of individuals belonging to the black-breasted red and white varieties had an extremely low proportion of the genome shared with the original type (black-breasted white). This suggests that derivative varieties were created by crossing with other breeds, with low introgression of the original-type genome. We propose that the three plumage colour varieties should be treated as separate genetic units in a conservation programme.