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.     

Monodeiodination of thyroxine to 3, 5, 3'-triiodothyronine and to 3, 3', 5'-triiodothyronine in isolated dog renal cortical tubuli

Monodeiodination of T4 to T3 and rT3 in the intact cells of dog renal tubuli and glomeruli was investigated. The tubuli and glomeruli were obtained by a sieve method. T4 (2 micrograms/ml) was incubated in Tris-HCl buffer, pH 7.5, with renal cells (180 micrograms protein/ml) and 5 mM DTT for 1 h at 37 degrees C and the T3 and rT3 generated during incubation were measured by specific radioimmunoassays. In order of decreasing activity, dog renal cortical tubuli, cortical homogenate, glomeruli and medullary tubuli were capable of converting T4 to T3. Net rT3 production from T4 in cortical tubuli was also greater than that in cortical homogenate. The conversion of T4 to T3 and also to rT3 in cortical tubuli was enzymatic in nature, since the reactions showed dependence on time and protein concentration; instability to heating; temperature and pH optimum. The production of T3 and rT3 from T4 was maximum at pH 6.5 and at pH 9.5, respectively, indicating that two different enzymic systems, a 5- and a 5'-monodeiodinase, might be involved in the deiodination of the tyrosyl and the phenolic ring of T4 in dog kidney.     

Effect of changes in thyroid state on metabolism of thyroxine by rat placenta

We studied the effect of the state of the thyroid on T4 monodeiodination in the rat placenta, and it was compared with those in the liver and kidney. The tissues, maternal serum, and amniotic fluid were obtained from pregnant rats. The tissues were homogenized in cold 50 mM Tris-HCl buffer, pH 7.5. The homogenate (1 mg protein) was incubated at 37 degrees C for 60 min with 1 microgram T4 in the presence of 5 mM DTT. The T3 and reverse T3 generated in the reaction mixture were extracted into cold ethanol and measured by RIAs. The conversion of T4 to reverse T3 in rat placenta was not significantly changed in MMI-induced hypothyroidism or T4 induced hyperthyroidism. On the other hand, conversion of T4 to T3 in the liver and kidney were changed in parallel with the thyroid state. The concentration of reverse T3 in the amniotic fluid was increased in accordance with the increase in the maternal serum T4 concentration. These results indicate that the placental T4 inner ring deiodination is not affected by the thyroid state, and that the change in the amniotic fluid reverse T3 concentration in this study is mainly dependent upon the change in maternal thyroid function.     

Nuclear magnetic resonance studies on the spatial relationship of aromatic donor molecules to the heme iron of horseradish peroxidase

The geometry of hydrogen donor molecules bound to horseradish peroxidase was investigated using nuclear magnetic resonance techniques. Between resorcinol and 2-methoxy-4-methylphenol which showed different optical difference spectra, little difference was observed in the orientation of the molecules bound to horseradish peroxidase: the minimal distances between the enzyme iron and the protons of the phenol rings are in the range of 8.4-11.0 A. This situation was not greatly different for the third compound studied in this paper, benzhydroxamic acid, providing evidence against the view that its side chain coordinates to the heme iron. Furthermore, it was found that transferred nuclear Overhauser effect for the signals of these compounds was observable only when the heme peripheral 8-methyl proton signal was irradiated. These results, together with a hypothetical model of the enzyme structure obtained by computer-aided simulation procedures, suggest that the binding of these donor molecules and competitive inhibitors occur in the vicinity of the heme peripheral 8-methyl group, with hydrophobic interactions probably with Tyr-185 and with hydrogen bond with adjacent amino acid residues such as Arg-183.     

On the degradation of dermorphin and D-Arg2-dermorphin analogs by a soluble rat brain extract

Degradation of dermorphin, [D-Arg2]dermorphin and [D-Arg2, Gly3, Phe4]dermorphin in a soluble rat brain extract was examined. The former two heptapeptides were degraded in a similar fashion to produce corresponding N-terminal tetrapeptide as the main degradation product along with the parallel release of Tyr5, Pro6 and Ser7-NH2. Tyr-D-Arg-Phe-Gly showed a good enzymatic stability. When captopril, an angiotensin-converting enzyme inhibitor, was present in the incubation mixture, hydrolysis of the Gly4-Tyr5 bond was markedly suppressed and resulted in release of the corresponding N-terminal hexapeptide as the main degradation product. Combined use of captopril and amastatin, an aminopeptidase inhibitor, markedly suppressed the hydrolysis of these peptides. On the other hand, [D-Arg2, Gly3, Phe4]dermorphin was hydrolyzed easier than the other two heptapeptides and considerable amounts of Tyr1 and Phe4 were released after 20 hr incubation while the N-terminal tetrapeptide, Tyr-D-Arg-Gly-Phe, showed a good enzymatic stability. On the basis of these results, possible degradation pathways of these heptapeptides were discussed.     

Measurement of Substance P Metabolites in Rat CNS

A procedure based on ion-exchange chromatography for chemical separation and radioimmunoassays for quantitation of substance P (SP), the SP(1-7), and C-terminal fragments, respectively, has been developed. The procedure allows the determination of these fragments in the presence of large (i.e., 50- to 100-fold) excess of parent compound. The chemical identity of isolated SP and fragments was studied with preparative electrophoresis on dilute agarose gel and with HPLC. The activity identified as SP(1-7) comigrated with the authentic standard whereas practically all activity isolated as C-terminal fragments comigrated with SP(5-11). The levels of C-terminal fragments in rat brain areas rich in SP and in spinal cord were 1-2% of those of parent compound. The levels of SP(1-7) were always higher, in the spinal cord markedly higher (three to five times). Postmortem storage of samples from brain and spinal cord indicated that SP(1-7) levels fell more rapidly than those of SP or C-terminal fragments.     

Role of Tween 80 and Monoolein in a Lipid-Sterol-Protein Complex Which Enhances Ethanol Tolerance of Sake Yeasts

An exogenous ternary complex composed of Tween 80, ergosterol, and albumin increased the final ethanol concentration of fermentation by sake yeasts from 17.2 to 19.0% (vol/vol) and reduced the fermentation time from 30 to 25 days. Likewise, a complex of monoolein, albumin, and either ergosterol or ergosteryl oleate increased the final ethanol concentration of fermentation to 19.7 or 19.8% (vol/vol), respectively, and reduced the fermentation time to 25 days. Both Tween 80 and monoolein promoted the fermentative activity (Q_CO2) of cells, and the effect was enhanced by the presence of ergosterol.     

Purification and characteristics of cytosolic chitinase from Piromyces communis OTS1

Abstract A chitinase was purified from the cytosolic fraction of the anaerobic rumen fungus Piromyces communis OTS1 by affinity chromatography using regenerated chitin, gel filtration and chromatofocusing. The chitinase was most active at pH 6.2 and at 60 °C in a 20-min assay. The molecular mass of the purified protein was estimated by SDS-PAGE to be 42 kDa and its pI was 4.9. The enzyme activity, which was of the 'endo' type, was inhibited by A⁺, Hg²⁺ and allosamidin. N -Acetyl- β -glucosaminidase and 'exo' type chitinase activity were absent from the purified preparation.     

The types of bioreactors used for shoots and embryos

Plant regenerated organs such as shoots, bulbs, microtubers, corms, embryos, etc. have been successfully proliferated in the bioreactor. The use of a bioreactor leads to the development of technology suitable for large scale plant propagation. The basic construction and characteristics of various types of bioreactor systems are reviewed in relation to shoot and embryo cultures. A pilot scale 500 liter bioreactor system was applied to the production of large scale Stevia rebaudiana shoots.Abbreviations DW dry weight - EC electrical conductivity - FW fresh weight - ORP oxidation-reduction potential     

Pock Formation of Streptomycetes endus with Production of Phage Taillike Particles

Plate (or slope) cultures of endomycin-producing Streptomyces endus (KCC S-0213) showed spontaneously developing pocks which increased in number during subculturing. Neither spore formation nor typical aerial hyphae formation was observed in the pocks, whereas formation of substrate hyphase was not inhibited. Almost all of the hyphae were broken or lysed in the pocks, and many phage tail tiplike particles were observed in the pocks. No self-replication activity was associated with the particles. The particles often formed a hexagonal crystal or a large crystal mass. The production of these particles did not occur in the liquid culture or in young or normal plate cultures having no pocks. These results were similar to those obtained from the plaque-making phenomenon, except for active phage production, in thiostrepton-producing Streptomyces azureus (ATCC 14921), which has been described previously.