The mechanism of biliary secretion of reduced glutathione. Analysis of transport process in isolated rat-liver canalicular membrane vesicles

Transport of reduced glutathione (GSH) was studied in isolated rat liver canalicular membrane vesicles by a rapid filtration technique. The membrane vesicles exhibit uptake of [2-3H]glycine--labeled GSH into an osmotically reactive intravesicular space. Although the canalicular membrane vesicles possess gamma-glutamyltransferase and aminopeptidase M, enzymes that hydrolyze glutathione into component amino acids, inactivation of the vesicle-associated transferase by affinity labeling with L-(alpha S,5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (AT-125) had no effect on the initial rate of GSH transport. Chemical analysis revealed that intact GSH accounted for most of vesicle-associated radioactivity. The initial rate of transport followed saturation kinetics with respect to GSH concentration; an apparent Km of 0.33 mM and V of 1.47 nmol/mg protein in 20 s were calculated. These results indicate that transport of GSH across the canalicular membranes is a carrier-mediated process. Replacement of NaCl in the transport medium by KCl, LiCl or choline chloride had no effect on the transport activity of the vesicles. The rate of GSH uptake by the vesicles was enhanced by valinomycin-induced K+-diffusion potential (vesicle inside-positive) and was inhibited by probenecid, indicating that GSH transport across the canalicular membranes is electrogenic and involves the transfer of negative charge. The transport of GSH was inhibited by oxidized glutathione or S-benzyl-glutathione. This transport system in canalicular plasma membranes may function in biliary secretion of GSH and its derivatives which are synthesized in hepatocytes by oxidative processes or glutathione S-transferase.     

Accuracy of endometrial aspiration in the diagnosis of endometrial cancer

A study was performed to evaluate the cytologic criteria for recognizing endometrial cancer and to determine the accuracy of endometrial aspiration in its detection. In addition to the conventional cytologic criteria for the diagnosis of endometrial cancer, the irregular chromatin distribution around the macronucleoli proved to be a useful criterion. Using all criteria, cytology was reported as positive in 18 of 19 patients (94.7%) with endometrial cancer. In 856 of 12,563 high-risk outpatients at Kinki University Hospital, aspiration using the Masubuchi apparatus was carried out in screening for endometrial cancer. Cancer was detected in 18 patients (2.1%), with 94.7% of the cancers detected by the cytologic screening. This result indicates that endometrial aspiration using the Masubuchi apparatus is a reliable and safe method of screening for endometrial cancer.     

Identification of Cytokinins in Root Exudate of the Rice Plant

Cytokinins, cis-zeatin and cis- and (trans-ribosylzeatin, wereidentified in the root exudate of the rice plant (Oryza sativa,indica cultivar IR-24) after several chromatographic separationsand combined gas-liquid chromatography-selected ion monitoring(GC-SIM) analysis. The presence of trans-zeatin ribotide wassuggested by enzyme hydrolysis, subsequent chromatographic separationand GC-SIM. The comparatively high content of the ribotide inthe root exudate suggests the form of cytokinins to be transportedfrom roots to other parts in the rice plant. (Received July 22, 1982; Accepted November 25, 1982)     

Changes in polyamine content during limb regeneration in adult Xenopus laevis

Polyamine contents in the regenerates were determined at various stages after amputation of the forelimbs of the adult female Xenopus laevis. Putrescine, spermidine, spermine, and sym-homospermidine were detected in all the specimens examined. Cadaverine was detected only in a limited number of samples. At 5 days after amputation of forelimbs, well before the formation of regenerates, the putrescine content in the stump tissues increased, followed by the increase in spermidine content. The putrescine level in the forelimb regenerates was highest between 30 and 50 days after amputation, and then decreased. The spermidine concentration in the regenerates was about 20 times greater than that in intact forelimbs all throughout the experiments. The concentration of spermine was initially lower than that of both putrescine and spermidine and further decreased soon after amputation. The concentration of sym-homospermidine was originally very low and increased slightly during regeneration. The significance of these results, with respect to the function of polyamines in forelimb regeneration of Xenopus laevis, is discussed.     

Inter- and intramolecular stacking interaction between indole and adeninium rings

Crystals of 1,9-dimethyladeninium-indole-3-acetate (1:1) complex (I) and 9-(3-indol-3-ylpropyl)-1-methyladeninium iodide (II), an inter- or intramolecular model for the stacking interaction between the tryptophanyl residue and the methylated (or protonated) adenine base, were subjected to X-ray analyses. Nearly parallel stacking and interplanar spacing near to 3.4 A were observed between the indole and adeninium rings of both crystals. In particular, one of the two stacking pairs formed in I showed the existence of a partial charge-transfer interaction in their ground states. On the basis of the molecular orbital consideration, the mutual orientation between these stacked aromatic rings is considerably governed by the orbital interaction between the highest occupied molecular orbital of the indole ring and the lowest unoccupied one of the adeninium ring. The ring stacking observed in II was stabilized by the strong coupled dipole-dipole interaction. Absorption, fluorescence, and proton nuclear magnetic resonance spectra indicated the existence of a stacking interaction in the aqueous solutions of I and II, as well as in their crystalline states. The biological implication for the observed stacking interactions has been discussed.