Thiaminepyrophosphatase activity in the plasma membrane of microglia

An intense thiaminepyrophosphatase (TPPase) activity was demonstrated in glial cells and blood vessels in the central nervous system (CNS), when incubation was carried out with thiaminepyrophosphte (TPP, cocarboxylase), using the method of Novikoff and Goldfischer (1961). Glial cells with TPPSase activity were identified as microglia because they were morphologically similar to microglial cells in the sections stained with silver impregnation. TPPase activity was localized in the microglial perikaryon and in the processes, as viewed under a light microscope. Electron microscopically, enzyme activity was localized in the plasma membrane of microglia. We consider this activity to be a true TPPase activity hydrolyzing TPP, and we then went on to examine the substrate specificity, optimum pH, effect of chemical inhibitors and activators, and the effect of glutaraldehyde fixation. Our data are reported herein.     

Cells capable of uptake of horseradish peroxidase in some circumventricular organs of the cat and rat

Summary The structure of mesenchymal cells distributed in some of the hypendymal organs of the circumventricular system in the cat and rat was demonstrated after intravenous injection of high doses of horseradish peroxidase. These cellular elements were observed in the vicinity of blood vessels of the organon vasculosum laminae terminalis, subfornical organ and area postrema. Electron-microscopically, these cells located between the basal laminae of the brain parenchyma and the blood capillaries show long cellular processes encircling fenestrated capillaries. Light and electron-microscopic examination revealed that this cell type is identical with the horseradish peroxidase-uptake cells, previously reported in the vicinity of the hypophysial portal system. Such phagocytic cells may be considered as a cellular component intervening between the brain parenchyma and the blood stream, playing a role in selective barrier functions in the above-mentioned circumventricular organs where a blood-brain barrier in the classical sense of the definition is lacking.This work was supported by grant No. 437002 from the Ministry of Education, Science and Culture, Japan     

Mass fragmentographic determination of lofepramine and its metabolites in human plasma and urine using deuterated internal standards

A sensitive and specific method for the determination of lofepramine and its metabolites, desipramine and 2-hydroxydesipramine, in human plasma and urine is described. Lofepramine, desipramine and 2-hydroxydesipramine were derivatized to ethyl p-chlorobenzoate, the bis(heptafluorobutyryl) derivative and the N,O-bis(trifluoroacetyl) derivative, respectively, and then analysed by gas chromatography—mass fragmentography. Corresponding deuterated compounds were used as internal standards. Determination was possible at levels as low as 2 ng/ml for lofepramine and desipramine and 20 ng/ml for 2-hydroxydesipramine.     

Diversity of the cadherin family: evidence for eight new cadherins in nervous tissue.

To examine the diversity of the cadherin family, we isolated cDNAs from brain and retina cDNA preparations with the aid of polymerase chain reaction. The products obtained included cDNAs for two of three known cadherins as well as eight distinct cDNAs, of which deduced amino acid sequences show significant similarity with the known cadherin sequences. Larger cDNA clones were isolated from human cDNA libraries for six of the eight new molecules. The deduced amino acid sequences show that the overall structure of these molecules is very similar to that of the known cadherins, indicating that these molecules are new members of the cadherin family. We have tentatively designated these cadherins as cadherin-4 through -11. The new molecules, with the exception of cadherin-4, exhibit features that distinguish them as a group from previously cloned cadherins; they may belong to a new subfamily of cadherins. Northern blot analysis showed that most of these cadherins are expressed mainly in brain, although some are expressed in other tissues as well. These findings show that the cadherin family of adhesion molecules is much larger than previously thought, and suggest that the new cadherins may play an important role in cell-cell interactions within the central nervous system.     

Differences in pathological findings and growth hormone responses in patients with growth hormone-producing pituitary adenoma.

Plasma growth hormone (GH) responses to various stimuli were examined in 21 patients with GH-producing pituitary adenomas, classified into three types by the immunohistochemistry of cytokeratin and the glycoprotein hormone alpha-subunit distribution. Seven type 1 adenomas were exclusively composed of cells in which the cytokeratin formed a dot-like pattern; they were chromophobic to hematoxylin and eosin (H&E), occasionally positive for GH, and almost completely negative for the alpha-subunit. Thirteen type 2 adenomas were composed of cells with cytokeratin that had a perinuclear distribution; they were eosinophilic to H&E, and diffusely positive for both GH and the alpha-subunit. One patient had a type 3 adenoma which had a mixed pattern of intracellular cytokeratin distribution and was chromophobic and eosinophilic to H&E. Clinically, type 1 is characterized by earlier onset, larger tumor size, and more frequent aggressive extension. Paradoxical GH responses to TRH and OGTT were seen in 1 of 6 patients (16.7%) of type 1 and 8 of 9 patients (88.9%) of type 2, and 0% of type 1 and 62.5% of type 2, respectively. Type 2 cases showed higher plasma GH response to GH-releasing hormone, and a tendency to greater suppression of plasma GH by bromocriptine compared with type 1. Octreotide acetate administration revealed that the nadir/basal ratio of plasma GH levels was 42.9 +/- 6.6% in type 1 and 13.5 +/- 5.8% in type 2. These results suggest that there is a pathophysiological difference between these two distinct types of GH-producing pituitary adenomas.     

Differential establishment and survival of Hymenolepis diminuta in syngeneic and outbred rat strains.

Experimental Hymenolepis diminuta infection was carried out in inbred strains of rats (F344/N, JAR-2, LOU/M, TM, DA and DA-bg/bg) and outbred Wistar rats. All strains became infected with this cestode, but clear strain-dependent variation in the susceptibility to H. diminuta infection was observed. Marked differences in worm persistence and worm weight were found at 6 weeks post-infection in TM and DA rats. These strains would be useful to clarify the interactions between H. diminuta and its rat host.     

A DBL-homologous region of the yeast CLS4/CDC24 gene product is important for Ca(2+)-modulated bud assembly.

The CLS4/CDC24 is essential for the budding process of the yeast Saccharomyces cerevisiae. Disruption of the CLS4/CDC24 gene is lethal, and expression of the CLS4 product under the control of the GAL1 promoter is sufficient for cellular growth. The CLS4 product is detected in yeast cell lysate with an apparent molecular mass of 93 kD (854 amino acid residues) and shows homology with the human DBL oncogene product. Temperature-sensitive cdc24-1 mutation is located in the N-terminal portion of the protein whereas Ca(2+)-sensitive cls4-1 mutation is present after the DBL-homologous region (amino acid residues 281-518) near the putative Ca(2+)-binding site. Mutations within the DBL-homologous region are responsible for the Ca(2+)-sensitive phenotype. Thus the CLS4 gene product seems to have several functional domains within the molecule essential for bud assembly.     

Isolation and identification of alpha-(beta-alanyl)hypusine from bovine brain

A unique dipeptide was isolated from bovine brain using five steps of ion-exchange chromatography. Its acid hydrolysate contained equimolar amounts of beta-alanine and hypusine. The structure of the peptide was elucidated as alpha-(beta-alanyl)hypusine using dansylation technique. About 1 mumol of the compound was isolated from 1090 g of bovine brain.     

Effect of streptokinase on prostacyclin synthesis and phospholipase activity in cultured pulmonary artery endothelial cells

In this study, we examined the effects of streptokinase on arachidonic acid release and prostacyclin biosynthesis in cultured bovine pulmonary artery endothelial cells. When intact cells were incubated with streptokinase, a significant stimulatory effect on prostacyclin biosynthetic activity in cells was evident without any cellular damage at all concentrations used (1-10,000 units/ml). Streptokinase also caused a marked release of arachidonic acid. It induced rapid phospholipid hydrolysis, resulting in the release of up to 15% of incorporated [3H]arachidonic acid into the medium. After the addition of streptokinase, degradation of phosphatidylcholine and phosphatidylethanolamine was observed and lysophosphatidylcholine and lysophosphatidylethanolamine were produced. We also observed a transient rise in diacylglycerol after the addition of streptokinase. To test for phospholipase C activity, the release of incorporated [3H]choline, [3H]inositol and [3H]ethanolamine into the culture medium was determined. The level of radioactive inositol showed an increase, but the changes in choline and ethanolamine were comparatively small. An increase in inositol was detectable within 1 min after streptokinase addition and peaked after 15 min. Inositol phosphate and inositol trisphosphate were released, and these releases were suppressed by the addition of neomycin (50 microM). These results suggest that streptokinase stimulates phospholipase A2 and C activity, and that prostacyclin biosynthesis is subsequently increased in cultured endothelial cells.