L-canavanine inhibits L-arginine uptake by broiler chicken intestinal brush border membrane vesicles.

1. Intestinal brush border membrane vesicles (BBMV) were prepared from 3-week-old broiler chickens. 2. Electron microscopy of the BBMV fraction showed single membrane vesicles of different sizes with no electron dense material inside. No other organelles were observed. The sucrase and maltase activities were enriched by factors of 16 and 18, respectively, in the BBMV fraction in comparison with the homogenate. On the other hand, the Na+/K+-ATPase sensitivity to ouabain was increased by a factor of 0.8. 3. The BBMV showed a maximum L-[14C]-arginine uptake (944.9 +/- 22.9 pmoles/mg protein) at 45 s and thereafter it declined slowly. In the presence of 0.5 mM L-canavanine, the L-[14C]-arginine uptake by BBMV was reduced by 43.6% at 45 s. 4. It is concluded that L-canavanine inhibits L-arginine Na+-dependent transport across the enterocyte apical membrane in a highly purified intestinal BBMV from broiler chickens.

Effects of Bauhinia megalandra aqueous leaf extract on intestinal glucose absorption and uptake by enterocyte brush border membrane vesicles.

Aqueous extract of Bauhinia megalandra leaves was able to inhibit the intestinal glucose absorption in a concentration-dependent way and additive to phlorizine. Moreover, B. megalandra leaf extract drastically reduced the 14C-glucose uptake by enterocyte brush border membrane vesicles. The B. megalandra leaf extract administrated orally, simultaneously with glucose improved the glucose tolerance with a significant reduction of the 30-min peak. The extract did not have an effect on the glucose tolerance when glucose was administrated subcutaneously.

Inhibición de la neoglucogénesis y de la glucosa 6-fosfatasa hepática por flavonas purificadas a partir de hojas de bauhinia megalandra / Inhibition of the neoglucogenic and glucose 6-hepatic phosphatase for flavones purified from bauhinia megalandra leaves

En microsomas intactos, dos flavonas purificadas a partir de hojas de Bauhinia megalandra, inhiben la actividad de la glucosa 6-fosfatasa en una manera dependiente de su concentración. Ambas flavonas incrementan el valor del Km para glucosa-6-fosfato, sin modificar el valor de Vmax. Los resultados permiten concluir que ambas flavonas inhiben competitivamente al transportador de glucosa-6-fosfato (T1), sin afectar ni a la subunidad catalítica ni al transportador de fosfato/pirofosfato (T2) del sistema de la glucosa-6-fosfatasa. Las flavonas inhiben la capacidad neoglucogénica de rebanadas de hígado incubadas en buffer Krebs-Ringer bicarbonato suplementado con lactato y albúmina saturada con oleato. La inhibición que ejercen las flavonas sobre el transportador T1 del sistema de la glucosa-6-fosfatasa microsomal, puede explicar la disminución de la capacidad neoglucogénica de las rebanadas de hígado, lo cual podría reducir la concentración de glucosa en pacientes diabéticos

Biological screening of plants of the Venezuelan Amazons.

A total of 21 extracts derived from 17 different plant species collected in Venezuelan Amazons have been tested for the following biological activities: cardiovascular activity, brine shrimp lethality, and inhibitory effects on the hydrolysis of glucose-6-phosphate in intact and disrupted microsomes. Eight extracts diminished rat blood pressure with or without changes in heart rate. The fruit extract of Swartzia leptopetala and the leaf and twig extract of Connarus lambertii resulted in death of experimental animals. The majority of extracts (17 extracts) showed significant toxicity against Artemia salina. Concerning the hydrolysis of glucose-6-phosphate, better inhibitory effects were observed in intact microsomes than in disrupted ones for all the extracts, suggesting that these extracts intervene with variable potency in glucose-6-phosphate transport through the microsomal membrane.

Characterisation of hepatic microsomal glucose-6-phosphatase from broiler chickens.

1. The hepatic glucose-6-phosphatase (G-6-Pase) kinetic variables from chickens were studied in intact and disrupted microsomes using two substrates: glucose-6-phosphate (G-6-P) and pyrophosphate (PPi). They were studied from embryonic life to 51 d of age. 2. The phosphohydrolase activity studied in the broiler chicken liver microsomes corresponds to a true glucose-6-phosphatase. 3. The enzyme VMAX with both substrates in intact and disrupted microsomes showed 2 maxima: one in 19-d-old embryos and the other in 9-d-old chickens. Pyrophosphatase (PPase) VMAX in intact microsomes was higher than that of the G-6-Pase at all ages studied, except in 12 d embryos and 3-d-old chicks. In disrupted microsomes the VMAX of both enzymatic activities were similar. The G-6-Pase latency was high in the 19-d-old embryos and 51-d-old chickens. 4. The KM for PPi and G-6-Pase decreased when microsomes were disrupted. In intact microsomes the G-6-P KM was low in embryos and 3-d-old chicks and later increased. On the other hand, the PPi KM in intact microsomes showed little change during the animal's life and was lower than that of G-6-P. In disrupted microsomes the KM for both substrates were similar. 5. These results suggest a sequential incorporation of the G-6-Pase system components in the endoplasmic reticulum.

Thioacetamide effects on the polypeptidic and nucleoporin pattern and chemical composition of rat liver nuclear envelope subfractions.

The effect of the administration of seven doses of the hepatocarcinogen thioacetamide on the chemical composition of rat liver nuclear envelope subfractions: associated chromatin, nuclear membranes and pore complex-lamina fraction, is analyzed. No alteration in DNA, RNA or phospholipid content is observed after the hepatocarcinogen treatment. Electrophoretic studies of each subfraction from thioacetamide treated rats show differences in the relative proportions of some polypeptides when compared with the controls. Examination of the wheat germ agglutinin binding polypeptides of each subfraction reveals a decrease in the stain of two pore complex-lamina nucleoporins of 85 and 164 kDa and an increase in one of 93 kDa; this observation can be due to changes in the quantity and/or in the agglutinin binding capacity of the nucleoporin as a result of thioacetamide administration. In view of the participation of nucleoporins in the nucleocytoplasmic transport, the changes observed suggest a relationship between changes of some O-linked N-acetyl glucosamine polypeptides components of the nuclear pore complex and the altered transport of some RNA species observed after thioacetamide administration.

Proteins from different classes of liver nuclei in normal and thioacetamide-treated rats.

1. In normal rats the amounts of each of the main types of nuclear protein, i.e. soluble proteins, histones, non-histone chromosomal proteins and residual proteins, vary within the different classes of rat liver nuclei fractionated by zonal centrifugation. 2. Heterogeneity is observed in the non-histone chromosomal proteins prepared from different classes of liver nuclei. These differences were observed by analysis of the proteins both by sodium dodecyl sulphate-polyacrylamide-gel electrophoresis and electrofocusing electrophoresis. They are most evident between the non-histone chromosomal proteins obtained from stromal and parenchymal nuclei. However, some differences are also found for the parenchymal nuclei, between the diploid parenchymal and the tetraploid parenchymal, and between them and the nuclei involved in the synthesis of DNA respectively. 3. Drastic alterations in the nuclear proteins are found after the administration of thioacetamide. The changes observed are complex and not uniform. They vary with the age of the animal and the type of nucleus. In general an increase in the soluble proteins and non-histone chromosomal proteins and a decrease in the residual proteins is observed. There is a decrease in the specific radioactivity of soluble and residual proteins. 4. Electrophoretic analysis of the non-histone chromosomal proteins showed that specific changes occurred after administration of thioacetamide, which are different in adolescent and young adult rats.

Studies of nuclei separated by zonal centrifugation from liver of rats treated with thioacetamide.

1. The effects of the inclusion of thioacetamide in the diet on the properties of rat liver nuclei were studied both in adolescent rats, in which the parenchymal cells contain diploid nuclei, and in young adult rats, with a high proportion of tetraploid nuclei. 2. These investigations included a survey of the sedimentation properties of the nuclei, the nuclear volumes, content of DNA, RNA and protein, the incorporation in vivo of [(3)H]thymidine into DNA and [(14)C]orotate into RNA, and measurements of the activity of RNA polymerase and ribonuclease. These studies were conducted on nuclei fractionated by zonal centrifugation. 3. In both groups of animals, exposure to thioacetamide produced large numbers of nuclei that were abnormal in their chemical composition and enzymic activity. The changes were complex as regards both the types of nuclei that were affected and in their variation with time. 4. In adolescent rats two waves of synthesis of DNA and RNA were observed, one at 3 days and the other after 2 weeks of treatment. The first decline in the incorporations into both DNA and RNA coincided with a decrease in the pool sizes of some of the precursors. The activity of RNA polymerase was not substantially altered. A marked increase in the content of protein was observed before the first wave of synthesis. The normal progressive increase in tetraploid nuclei was prevented. 5. In young adult rats two waves of DNA synthesis were detected. Each was preceded by a large increase in the amount of protein per nucleus but was not accompanied by increased RNA synthesis. After 4 weeks of treatment, the diploid stromal nuclei appeared mainly unaffected and large numbers of tetraploid nuclei with a greatly increased quantity of protein were observed.