Effects of Hydrostatic Pressure on Carcinogenic Properties of Epithelia

The relationship between chronic inflammation and cancer is well known. The inflammation increases the permeability of blood vessels and consequently elevates pressure in the interstitial tissues. However, there have been only a few reports on the effects of hydrostatic pressure on cultured cells, and the relationship between elevated hydrostatic pressure and cell properties related to malignant tumors is less well understood. Therefore, we investigated the effects of hydrostatic pressure on the cultured epithelial cells seeded on permeable filters. Surprisingly, hydrostatic pressure from basal to apical side induced epithelial stratification in Madin-Darby canine kidney (MDCK) I and Caco-2 cells, and cavities with microvilli and tight junctions around their surfaces were formed within the multi-layered epithelia. The hydrostatic pressure gradient also promoted cell proliferation, suppressed cell apoptosis, and increased transepithelial ion permeability. The inhibition of protein kinase A (PKA) promoted epithelial stratification by the hydrostatic pressure whereas the activation of PKA led to suppressed epithelial stratification. These results indicate the role of the hydrostatic pressure gradient in the regulation of various epithelial cell functions. The findings in this study may provide clues for the development of a novel strategy for the treatment of the carcinoma.     

Fermentative Production of Succinic Acid from n-Paraffin by Candida brumptii IFO 0731

An investigation of succinic acid production from n-paraffin under various culture conditions was carried out with Candida brumptii IFO 0731. Ammonium nitrogen was required for both cell growth and succinic acid production. Favorable culture conditions for succinic acid production were ascertained. The productivity was markedly increased by the additions of CaCO₃ and organic nutrients. Under the best condition, the largest quantity of succinic acid production, 23.6 mg/ml, was obtained in a 67% yield from super heavy n-paraffin after 8 days cultivation.     

Changes in Targeting Efficiencies of Proteins to Plant Microbodies Caused by Amino Acid Substitutions in the Carboxy-terminal Tripeptide

It has been demonstrated that the carboxyl terminus of microbodyenzymes functions as a targeting signal to microbodies in higherplants. We have examined an ability of 24 carboxy-terminal aminoacid sequences to facilitate the transport of a cytosolic passengerprotein, ß-glucuroni-dase, into microbodies in greencotyledonary cells of trans-genic Arabidopsis. Immunoelectronmicroscopic analysis revealed that carboxy-terminal tripeptidesequences of the form [C/A/S/P]-[K/R]-[I/L/M] function as amicrobody-targeting signal, although tripeptides with prolineat the first amino acid position and isoleucine at the carboxylterminus show weak targeting efficiencies. All known micro-bodyenzymes that are synthesized in a form similar in size to themature molecule, except catalase, contain one of these tripeptidesequences at their carboxyl terminus. (Received April 14, 1997; Accepted April 8, 1997)     

Inhibitory effect of norepinephrine on immunoreactive corticotropin-releasing factor release from the rat hypothalamus in vitro

Effects of catecholamines on immunoreactive corticotropin-releasing factor (I-CRF) release from the rat hypothalamus were examined using a rat hypothalamic perifusion system and a rat CRF RIA in vitro. Norepinephrine had a potent inhibitory effect on I-CRF release in a dose-dependent manner at 0.1 nM-1 microM concentrations, but dopamine did not. This inhibitory effect of norepinephrine was completely blocked by propranolol, but only partially blocked by phentolamine. Isoproterenol also had a potent inhibitory effect at 0.01-100 nM concentrations, and a high dose of phenylephrine (10 nM) inhibited I-CRF release. Clonidine did not influence I-CRF release. These results suggest that norepinephrine inhibits I-CRF release mainly through the beta-adrenergic receptor and partially through the alpha 1-receptor.     

Spatial variation in density of stream benthic fishes in northern Hokkaido, Japan: does riparian vegetation affect fish density via food availability?

The densities of two benthic fishes, the Siberian stone loach (Noemacheilus barbatulus) and the wrinklehead sculpin (Cottus nozawae), and the biomass of their food resources (i.e., periphyton and benthic invertebrates) were compared between forest and grassland streams in northern Hokkaido, Japan, to examine whether riparian deforestation had positive effects on the benthic fishes via enhancement of food availability. The comparisons indicated that riparian vegetation had little influence on periphyton, invertebrates, or fishes. Regression analysis indicated that spatial variations in loach and sculpin densities were explained more by substrate heterogeneity, competitor abundance, or both, rather than by food abundance. However, when the two species were combined as benthic insectivores, a strong correlation was found between total benthic fish density and invertebrate biomass. Our results suggest that, although total benthic fish abundance was food limited, riparian vegetation had no positive effects via food availability on the benthic fishes in our streams.     

Isolation and characterization of a cDNA encoding mitochondrial chaperonin 10 from Arabidopsis thaliana by functional complementation of an Escherichia coli groES mutant

Chaperonin (Cpn) is one of the molecular chaperones. Cpn10 is a co-factor of Cpn60, which regulates Cpn60-mediated protein folding. It is known that Cpn10 is located in mitochondria and chloroplasts in plant cells. The Escherichia coli homologue of Cpn10 is called GroES. A cDNA for the Cpn10 homologue was isolated from Arabidopsis thaliana by functional complementation of the E. coli groES mutant. The cDNA was 647 bp long and encoded a polypeptide of 98 amino acids. The deduced amino acid sequence showed approximately 50% identity to mammalian mitochondrial Cpn10s and 30% identity to GroES. A Northern blot analysis revealed that the mRNA for the Cpn10 homologue was expressed uniformly in various organs and was markedly induced by heat-shock treatment. The Cpn10 homologue was constitutively expressed in transgenic tobaccos. Immunogold and immunoblot analyses following the subcellular fractionation of leaves from transgenic tobaccos revealed that the Cpn10 homologue was localized in mitochondria and accumulated at a high level in transgenic tobaccos.     

Differentiation-associated changes in membrane proteins of mouse myeloid leukemia cells

The mouse myeloid leukemia cell line (M1) is known to differentiate in vitro into macrophages and granulocytes upon treatment with various inducer including mouse ascitic fluid. Changes of cell surface proteins during differentiation of M1 cells were analyzed by the lactoperoxidase-catalyzed radioiodination method and SDS-polycrylamide slab gel electrphoresis. Treatment of the cells with ascitic fluid changed the electrophoretic pattern of the iodinated proteins, the prominent change being the appearance of a new protein with a molecular weight of 180 000 (P180). Iodinated P180 was also detected in normal macrophages in granulocytes, which are similar to differentiated M1 cells. This protein was metabolically labeled with l-[¹⁴C]fucose, increasing with the period of the treatment. P180 was not expressed on ascitic fluid-treatment of a resistant clone of M1 cells that could not be induced to differentiate. These results indicate that P180 is a glycoprotein that is exposed on the outer surface of differentiated M1 cells, and that its expression is associated with differentiation of the cells.P180 was solubilized from ¹²⁵I-labeled macrophages with detergents bound to concanavalin A-Sepharose. This suggests that P180 is one of the receptors for concanavalin A. Therefore, P180 may contribute partly to the increases in agglutinability by concanavalin A and in the number of concanavalin A binding sites on the surface of M1 cells, which are known to be associated with differentiation of M1 cells.     

Microbody proliferation and segregation cycle in the single-microbody alga Cyanidioschyzon merolae

The proliferation cycle of the microbody was studied in the primitive red alga Cyanidioschyzon merolae, which contains one microbody per cell. Cells were synchronized with a dark/light cycle, and the morphology of the microbody and its interaction with other organelles were observed three-dimensionally by fluorescence microscopy, transmission electron microscopy, and computer-assisted three-dimensional reconstruction of serial thin sections. The microbody in interphase cells is a sphere of 0.3 μm in diameter without a core. In M-phase, the microbody passes through a series of irregular shapes, in the order rod, worm, branched, H-shaped and dumbbell, and symmetric fission occurs just before cytokinesis. The microbody duplicates its volume in M-phase and three-dimensional quantitative analysis revealed that its surface area increases before its volume does. The microbody touches the mitochondrion and the chloroplast throughout its proliferation cycle, except briefly in interphase cells, winding around the divisional plane of the mitochondrion at one phase. Immunocytochemical labeling of catalase as a marker of matrix proteins of the microbody revealed that the duplication of catalase occurs in tandem with the volume increase. While no specific apparatus was identified in the microbody divisional areas, we identified an electron-dense apparatus about 30–50 nm in diameter between the microbody and the mitochondrion that may play a role in segregating the daughter microbodies. These results are the first characterization to show the morphological changes of one microbody in a one-microbody alga without proliferation-inducing substrates, which have been used in many studies, and clearly show that two daughter microbodies arise by binary fission of the pre-existing microbody. Received: 11 November 1998 / Accepted: 22 December 1998     

New Neplanocin Analogues II. Enzymatic One-Step Synthesis and Antitumor Activity of 6′-(3-sn-Phosphatidyl)Neplanocin a Derivatives

Abstract

A novel series of neplanocin analogues, 6′-(3-sn-phosphatidyl)neplanocin As bearing a variety of fatty acyl or alkyl residues in the glyceride moiety (2b-2h), were synthesized by means of phospholipase D-catalyzed transphosphatidylation. Among them, 2b, 2c, and 2e each exhibited significant antitumor effect against P388 leukemia in mice, which evidently surpassed that of parent compound neplanocin A.     

Re-evaluation of physical interaction between plant peroxisomes and other organelles using live-cell imaging techniques

The dynamic behavior of organelles is essential for plant survival under various environmental conditions. Plant organelles, with various functions,migrate along actin filaments and contact other types of organelles, leading to physical interactions at a specific site called the membrane contact site. Recent studies have revealed the importance of physical interactions in maintaining efficient metabolite flow between organelles.In this review, we first summarize peroxisome function under different environmental conditions and growth stages to understand organelle interactions. We then discuss current knowledge regarding the interactions between peroxisome and other organelles, i.e., the oil bodies, chloroplast, and mitochondria from the perspective of metabolic and physiological regulation, with reference to various organelle interactions and techniques for estimating organelle interactions occurring in plant cells.