Reactivity of sulphydryl groups of cytosolic and mitochondrial bovine aspartate aminotransferases

Summary Reactivity of sulphydryl groups of cytosolic and mitochondrial aspartate aminotransferases from ox heart has been studied. A total of 5 and 7 cysteine residues per monomer are present in cAATo and mAATo, respectively. In native conditions only a single sulphydryl group can be titrated by Nbs₂ while the catalytic activity remains unchanged, however in the mitochondrial isozyme the reactivity depends on the functional state of the enzyme. Reactivity toward NEM reveals the existence of a syncatalytic sulphydryl group in the cytosolic isozyme. Titration of cAATo with pMB at pH 8 and pH 5 confirms the existence of two exposed sulphydryl groups with a different reactivity. The results compared with those reported on the corresponding isozymes from pig and chicken heart show that syncatalytic sulphydryl groups are of general occurrence in these enzymes.     

Glycosylation and structure of the yeast MF alpha 1 alpha-factor precursor is important for efficient transport through the secretory pathway

The MF alpha 1 gene encodes a precursor, prepro-alpha-factor, that undergoes several proteolytic processing steps within the classical secretory pathway to produce the mature peptide pheromone, alpha-factor. To investigate the role of structural features of the MF alpha 1 precursor in alpha-factor production, we analyzed the effect of mf alpha 1 mutations that alter precursor structure in a number of ways. These mutations resulted in decreased alpha-factor secretion and intracellular accumulation of pro-alpha-factor. With the exception of the mutant lacking all three N glycosylation sites, the pro-alpha-factor forms that accumulated were core glycosylated but had not yet undergone the addition of outer chain carbohydrate. The delay, therefore, occurred at a step prior to the first proteolytic processing step involved in maturation of the precursor and was probably due to inefficient endoplasmic reticulum-to-Golgi transport. Elimination of all three N-glycosylation sites caused a delay in disappearance of intracellular precursor, and alpha-factor secretion was also slowed. These data indicate that N glycosylation is important but not essential for transport of the precursor through the secretory pathway. The decreased alpha-factor secretion and increased precursor accumulation seen with many different structural changes of pro-alpha-factor indicate that the secretory pathway is extremely sensitive to changes in precursor structure. This sensitivity could cause inefficient secretion of heterologous proteins and hybrids between MF alpha 1 and heterologous proteins in yeast cells.     

Age-related changes in the morphology and function of the zona glomerulosa of the rat adrenal cortex

The age-related changes in the morphology and function of rat adrenal zona glomerulosa (ZG) were investigated by coupled stereological and radioimmunological techniques. For this purpose 4-, 8-, 16- and 24-month-old rats were studied. Aging caused a notable lowering in the plasma aldosterone concentration and a marked decrease in both basal and ACTH- or angiotensin II (ANG-II)-stimulated secretion of collagenase-dispersed ZG cells. Plasma renin activity (PRA) underwent an age-dependent decrease, while the plasma level of ACTH displayed a significant rise. ZG and its parenchymal cells did not evidence any age-related morphologically demonstrable alteration in their growth, nor ZG cells showed any marked ultrastructural change, with the exception of a severe lipid-droplet repletion. This last finding is in keeping with the aging-induced decrease in the secretory activity of ZG cells, inasmuch as lipid droplets are the intra-cellular stores of cholesterol esters, the obligate precursors of steroid hormones in rat adrenals. ACTH and ANG-II are well known to be involved in the maintenance of the growth of rat ZG; thus, the combined impairment of ANG-II production (as evidenced by PRA lowering) and increase in ACTH secretion may maintain unchanged ZG growth during aging.     

Development of the ovarian surface and associated germ cells in the human fetus

Summary The ovarian surface and associated germ cells have been studied in human fetuses from 12 weeks of age until near term, using light, TEM and SEM techniques. The surface epithelium and related cords proliferate extensively, especially at midterm. The cords in the ovarian cortex appear to be linked with ingrowths from the surface epithelium, and both structures have a common basal lamina. Germ cells are always interspersed among the somatic cells of the surface epithelium and associated cords. These results indicate that both the proliferating cords and surface epithelium may contribute to the formation of early follicles. Furthermore, the occurrence, of elements having some of the features of primitive steroidogenic cells in the regions of cordsurface epithelium continuity, suggests that both structures (surface epithelium and cords) contribute somatic cells, which in addition to becoming granulosa cells, might also contribute to the provision of primitive interstitial cells.Gonocytes tend to migrate through the developing ovarian tissue towards the surface where they become extruded into the peritoneal cavity. This phenomenon might contribute to the reduction in the number of germ cells at birth and parallels the atretic processes within the ovary.     

Interrelationship between oxygen consumption,superoxide anion and hydrogen peroxide formation in phagocytosing guinea pig polymorphonuclear leucocytes

The paper presents an experimental procedure for a simultaneous assay of oxygen consumption, O2- release and H2O2 accumulation at a very early stage of the respiratory burst that is induced by phagocytosis in guinea pig polymorphonuclear leucocytes. The main findings are as follows: (a) The oxygen consumption that is measurable does not correspond to all oxygen that is reduced. The relationship between the actual oxygen consumed and the amount that is reduced depends on the fate of the intermediate products O2- and H2O2. (b) O2- is measurable extracellularly by the reduction of cytochrome c. When cytochrome c oxidizes the extracellular O2-, molecular oxygen is formed. This fact is shown by a decrease of oxygen consumption. The molar ratio between the O2- detected and the oxygen given back is 1. (c) The amount of O2- released from the cells accounts for only a small part of oxygen actually reduced. (d) H2O2 is detectable only in the presence of NaN3. In this condition almost all oxygen consumed is recovered in the form of H2O2. The molar ratio O2/H2O2 is near unity. The amount of H2O2 derived from dismutation of O2- released is only an aliquot of the total H2O2 accumulated. Thus, most of H2O2 is derived from intracellular sources. (e) In the absence of inhibitors of H2O2 degrading reactions, no detectable accumulation of peroxide occurs. Under these conditions, the main part of H2O2 formed is degraded in almost equal amount by catalase and myeloperoxidase, while only a small aliquot is degraded by NaN3 insensitive reactions.     

Three dimensional organization of mammalian adrenal cortex

Summary The adrenal cortex of different mammals was studied by SEM in order to demonstrate its actual three-dimensional organization. In the rat, as well as in the cat and pig, the adrenal cortex appeared as a tunnelled continuum of polyhedral cells arranged in plate-like structures (laminae). This laminar arrangement was more evident in the inner fasciculate and reticular zones where the cortex revealed a striking similarity to liver tissue. The polyhedral cells of all cortical zones possessed regular facets populated by small pits, larger invaginations and numerous microvilli with the exception of very short and smooth areas probably corresponding to attachment zones and/or gap junctions. This cellular architecture produced a labyrinthic system of intercellular channels or lacunae in which the capillaries were suspended.The pericapillary areas of this labyrinth contained microvilli, amorphous material, a delicate net of fibrils and occasional cells. The intercellular compartment of this lacunar system was mainly bordered by numerous microvilli arising from endocrine cells.The luminal surface of the capillary wall showed not only irregularly protruding margins (interpretable as endothelial junctions) but also clearly overlapping and flattened endothelial extensions.In all the animals and areas of the adrenal cortex examined, the endothelial wall was provided with abundant clusters of small fenestrations (about 50 nm in diameter) generally arranged in sieve plates. Larger fenestrations were noted mainly in the fasciculate and reticular zones of the cat and pig and occasionally in the rat.A final point related to the nature and significance of sinusoidal fenestrations was the occurrence of irregularly shaped and intracapillary located cells mainly noted in the deeper zones of the fasciculate and reticular zones of the gland. These elements — possessing the surface characteristics of macrophages — were observed, with their irregular and slender evaginations, in close proximity to the large fenestrations in a manner reminiscent of Kupffer cells within the lumen of liver sinusoids.     

Chromosome constitution of in vitro segregated haploid and diploid cells of the mouse

The composition in segregated haploid sets of paternal and maternal chromosomes has been studied in order to verify whether their composition is uniparental of mixed, fixed or variable. Primary cultures where prepared using kidneys from hybrids of strains of Mus musculus in which the parental chromosomes are distinguishable; the maternal set consists of 20 teleocentric chromosomes, the paternal set of 9 metacentric chromosomes, derived by Robertsonian fusion and 2 telocentrics. Applying Seabright's banding technique, an analysis of segregated haploid and diploid cells, which have originated spontaneously through polyploidisation-segregation processes was carried out. It was concluded that the haploid sets have a variable composition of paternal and maternal chromosomes.     

Prostaglandins and renal function II. The effect of prostaglandin inhibition on autoregulation of blood flow in the intact kidney of the dog

In order to evaluate the effect of prostaglandin release on renal autoregulation in the intact kidney of the dog, pressure-flow curves were obtained before and after the administration of either indomethacin or meclofenamate, two potent prostaglandin synthetase inhibitors. After drug administration renal venous prostaglandin E decreased in each of eight studies with a mean change from 286 to 141 pg/ml (p < .001). In addition, prostaglandin inhibition was associated with a 31 percent decrease in renal blood flow and a 58 percent increase in renal resistance. Yet, as renal perfusion pressure was decreased by aortic constriction, the change in flow per pressure reduction and the percent change in renal resistance were not significantly different after prostaglandin inhibition when compared to control values in the same animals. The magnitude of the pressure range over which autoregulation was maintained was also similar in the two groups although both the initial and lowest level of autoregulation were slightly higher after prostaglandin inhibition. It is concluded that the administration of these prostaglandin synthetase inhibitors does not significantly impair renal autoregulation in the intact dog kidney.