Early changes in mitochondrial monoamine oxidase activity of rat liver during 2-acetylaminofluorene feeding

The activities of mitochondrial type A and B monoamine oxidase were determined in the liver of rats fed a diet containing 2-acetylaminofluorene (AAF). Three days after the initiation of AAF-feeding, there was a significant decrease of type B monoamine oxidase activity without affect on type A enzyme. The decreased activity of type B monoamine oxidase, which reached a minimum after three weeks, was sustained for as long as AAF-feeding was continued. Sex-related difference in response to AAF was seen in the rat with respect to the onset and the intensity of the decreased type B monoamine oxidase activity, male rats being more sensitive to the carcinogen than female rats. In contrast to the in vivo effect, AAF showed a potent inhibitory effect on type A monoamine oxidase, rather than on type B enzyme, when added in vitro. The pI₅₀ values were estimated to be 7.5 against type A monoamine oxidase and 4.1 against type B enzyme, respectively. The in vitro inhibition of both types of monoamine oxidase by AAF was competitive. The K_i values for AAF were calculated to be 9.51 · 10^?9 M for type A monoamine oxidase and 1.30 · 10^?5 M for type B enzyme, respectively. In accordance with the potent inhibitory effect of AAF on type A monoamine oxidase in vitro, a single administration of the carcinogen, at a dose of 50 mg/kg, resulted in a marked and temporal decrease of the enzyme activity in the mitochondria of male rat liver. Recovery of the decreased type B monoamine oxidase activity was slow, and the enzyme activity did not return to control levels, even if rats were fed the basal diet for 2 or 4 weeks after the cessation of AAF-feeding.     

Assignment of the human myeloperoxidase gene (MPO) to bands q21.3----q23 of chromosome 17

Using a human myeloperoxidase cDNA, we have mapped the human myeloperoxidase gene to chromosome 17 at q21.3----q23 by in situ hybridization to metaphase chromosomes from human lymphocyte preparations.     

Relationship between general or specific immunoreactivity and prognosis in postoperative patients with hepatocellular carcinoma

Summary The levels of a variety of immunological parameters were examined in 203 preoperative patients with hepatocellular carcinoma (HCC) at various stages (I–IV). The changes in the peripheral blood lymphocyte (PBL) count, the serum level of immunosuppressive acidic protein and the degree of the skin reaction to purified protein derivative were associated significantly with the stage of HCC progression. However, the percentages of lymphocyte subsets, mitogenic responsiveness of PBL and serum immunoglobulin concentration remained at the levels of stage I. Further study demonstrated that in patients undergoing hepatic artery ligation, there were statistically significant correlations between the PBL count, immunosuppressive acidic protein concentration and intensity of the skin reaction to purified protein derivative, assayed 1 month after surgery, and the prognosis. HCC-specific immunity was examined in 34 patients treated by hepatic resection or hepatic artery ligation using in vitro responses of PBL to HCC extracts (ATS test). This test was performed using culture medium containing added arginine. None of the PBL from the patients showed a positive response to allogeneic HCC extracts, but the PBL from 12 patients (9 hepatic resections, 3 hepatic artery ligations) were stimulated significantly (SI 2.5) with autologous HCC extracts. In 7 of 9 hepatic resection patients who were positive in the ATS test, tumor recurrence was identified. Statistical analysis indicated that the ATS test result was significantly correlated with tumor recurrence in hepatic resection patients. Autologous-PBL-stimulating activities were isolated in a fraction at pH 8.3 and in fractions at pH 6.7–7.0 by chromatofocusing of the crude extract. Although identification of the HCC-specific antigen remains to be done, use of the above fractions may simplify the ATS test procedure and improve its sensitivity.     

Mast cell degranulation and its inhibition by an anti-allergic agent tranilast. An electron microscopic study

Degranulation of IgE-sensitized rat mast cells by antigen was studied quantitatively in vitro and in vivo by electron microscopy. The inhibition of this degranulation by an anti-allergic drug, N-(3,4-dimethoxycinnamoyl)anthranilic acid (Tranilast), was also examined both in vitro and in vivo. In the in vitro study using peritoneal mast cells, alteration of the granules, cavity formation by fusion of the perigranular membrane and granule discharge due to fusion of the cavity membrane with the cell membrane were observed and were accompanied by histamine release. Scanning electron microscopy disclosed the extrusion of smooth, round bodies from pores formed on the cell surface. In the in vivo study of passive cutaneous anaphylaxis (PCA), the characteristic features of mast cell degranulation were obvious 5 min after the injection of antigen; leakage of dye increased progressively from 5 to 30 min but was not found at 6 h. From quantitative analysis of the substructure of mast cells, it was demonstrated that degranulation of IgE-sensitized mast cell induced by antigen was achieved by sequential exocytosis both in vitro and in vivo. Tranilast inhibited these changes to a remarkable extent and it was concluded that the inhibition of mast cell degranulation by this drug might play an important role in anti-allergic treatment.     

Type II restriction endonucleases cleave single-stranded DNAs in general.

Restriction endonucleases (13 out of 18 species used for the test) were certified to cleave single-stranded(ss)DNA. Such enzymes as AvaII, HaeII, DdeI, AluI, Sau3AI, AccII,TthHB8I and HapII were newly reported to cleave ssDNA. A model to account for the cleavage of ssDNA by restriction enzymes was proposed with supportive data. The essential part of the model was that restriction enzymes preferentially cleave transiently formed secondary structures (called canonical structures) in ssDNA composed of two recognition sequences with two fold rotational symmetry. This means that a restriction enzyme can cleave ssDNAs in general so far as the DNAs have the sequences of restriction sites for the enzyme, and that the rate of cleavage depends on the stabilities of canonical structures.     

Electron microscopic radioautographic study of the localization of an anti-allergic agent, Tranilast, in rat mast cells.

We have previously reported that Tranilast, an anti-allergic agent, was rapidly taken into the cytoplasm of rat mast cells in vitro by means of light microscopic radioautography. The present study was performed at the electron microscopic level to elucidate the fine localization of this agent in the mast cells. The results revealed that the number of radioautographic silver grains in the cells increased by the incubation with 3H-labelled Tranilast for 0 to 60 min. and that many silver grains were localized on the specific granules, especially on the perigranular membranes. These results suggest that the mode of inhibitory action of mast cell degranulation by Tranilast is related to the specific localization of this agent on the perigranular membranes.     

Selective inhibitors of germination of legume seeds in activated sludge compost

Water extracts of the compost produced from activated sludge and coffee residue were found to be selectively inhibitory to seed germination of some legumes. Germination rate of white clover (Trifolium repens L.), red clover (Trifolium pratense L.) and alfalfa (Medicago sativa L.) seeds were reduced to 2, 29 and 73% of the control, respectively, by water extracts of the compost (20 g l^–1). However, the extracts did not show any inhibition to seed germination of sorghum (Sorghum bicolor Moench), African millet (Eleusine coracana Gaertn.), and Komatsuna (Brassica rapa L.) at the same concentration. The inhibitors in the compost extracts were separated by ion-exchange chromatography and reverse-phase high performance liquid chromatography (HPLC) and the inhibitory activities of seed germination were tested with white clover seeds. Five inhibitors were isolated and identified as 3,4-dichlorophenylacetic acid (3,4-DCP), 3,4-dichlorobenzoic acid (3,4-DCB), 3,4,5-trichlorophenylacetic acid, 3,4,5-trichlorobenzoic acid and mono-2-ethylhexylphthalate by ¹H-, ¹³C-NMR spectroscopy and mass spectrometry. The inhibitory activities of some authentic chemicals of the inhibitors and the related compounds were compared. The results indicated that the main inhibitor in the compost could be 3,4-DCB, which was contained at the concentration of 6.58 mg kg^–1 compost and showed the strongest inhibitory effect on seed germination of white clover among the tested compounds.     

Effects of estradiol and testosterone on the synthesis,expression and degradation of androgen receptor in human uterine endometrial fibroblasts

The mechanism of known receptor-mediated androgen effects on the endometrial stroma was studied in endometrial fibroblasts derived from human uterus. 17-Estradiol (E) induced the expressions of androgen receptor (AR) mRNA, and predominantly increased the level of testosterone-binding sites (TBS) in uterine endometrial fibroblasts. The effect on the level of dihydrotestosterone-binding sites (DHTBS) was similar but smaller. This result suggests that the AR mRNA expressed might encode TBS, but probably not DHTBS. The TBS level increased by estrogen was down-regulated by testosterone (T) + E, but the AR mRNA expression increased by E was not down-regulated by E + T in the fibroblasts. Although the synthesis rate of AR was slightly increased (p<0.05) by E alone or E + T, the degradation rate of AR was significantly accelerated (p<0.05) by E + T in the fibroblasts. This result suggests that T might stimulate the metabolic rate of TBS, but does not inhibit the synthesis rate of AR mRNA to TBS in endometrial fibroblasts.     

Identification and Biological Activity of Germination-Inhibiting Long-Chain Fatty Acids in Animal-Waste Composts

Long-chain fatty acids in germination-inhibiting animal-wastecomposts were identified by gas chromatography-mass spectrometryas myristic, palmitic, stearic, oleic, linoleic, and linolenicacids. These acids were found at concentrations greater than0.25 mg (g dry compost)^–1. The identified acids, togetherwith lauric acid, and five kinds of short- and medium-chainfatty acid, were tested for their effects on the germinationprocess of sorghum seeds. The authentic long-chain fatty acids, which were dissolved ina 1:9 (v/v) mixture of methanol and distilled water at 40 mgliter^–1, significantly reduced the -amylase activity,physiological water uptake, and ATP content of the germinatingseeds during the first 24h of imbibition, as well as the rateof germination of seeds. Among the tested fatty acids, myristicand palmitic acids were the most potent inhibitors of germination.The inhibitory effects of long-chain fatty acids were strongerthan those of the phenolic acids. The short- and medium-chainfatty acids did not have any significant germination-inhibitoryeffects at 40 mg liter^–1. The results indicate that thelong-chain fatty acids are the dominant inhibitors of germinationin animal-waste composts, and that the inhibition of the -amylaseactivity in germinating sorghum seeds is one aspect of the modeof action of these long-chain fatty acids. ¹On leave from the Department of Crop Science, Faculty of Agriculture,University of Peradeniya, Sri Lanka.