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G E PALADE

Publications and source records attributed to G E PALADE.

At least 19 recordsLinked to original sources
PubMed1964

PROTEIN SYNTHESIS, STORAGE, AND DISCHARGE IN THE PANCREATIC EXOCRINE CELL. AN AUTORADIOGRAPHIC STUDY.

The synthesis, intracellular transport, storage, and discharge of secretory proteins in and from the pancreatic exocrine cell of the guinea pig were studied by light- and electron microscopical autoradiography using DL-leucine-4,5-H(3) as label. Control experiments were carried out to determine: (a) the length of the label pulse in the blood and tissue after intravenous injections of leucine-H(3); (b) the amount and nature of label lost during tissue fixation, dehydration, and embedding. The results indicate that leucine-H(3) can be used as a label for newly synthesized secretory proteins and as a tracer for their intracellular movements. The autoradiographic observations show that, at approximately 5 minutes after injection, the label is localized mostly in cell regions occupied by rough surfaced elements of the endoplasmic reticulum; at approximately 20 minutes, it appears in elements of the Golgi complex; and after 1 hour, in zymogen granules. The evidence conclusively shows that the zymogen granules are formed in the Golgi region by a progressive concentration of secretory products within large condensing vacuoles. The findings are compatible with an early transfer of label from the rough surfaced endoplasmic reticulum to the Golgi complex, and suggest the existence of two distinct steps in the transit of secretory proteins through the latter. The first is connected with small, smooth surfaced vesicles situated at the periphery of the complex, and the second with centrally located condensing vacuoles.

Animals↗
PubMed1964

NEW CYTOPLASMIC COMPONENTS IN ARTERIAL ENDOTHELIA.

A hitherto unknown rod-shaped cytoplasmic component which consists of a bundle of fine tubules, enveloped by a tightly fitted membrane, was regularly found in endothelial cells of small arteries in various organs in rat and man. It is about 0.1 micro thick, measures up to 3 micro in length, and contains several small tubules, approximately 150 A thick, embedded in a dense matrix, and disposed parallel to the long axis of the rod. In some of these cells, the cisternae of the endoplasmic reticulum are greatly distended by the accumulation of a dense, finely granular material. The nature and significance of these cytoplasmic components are yet unknown.

Animals↗
PubMed1964

SPECIFIC GRANULES IN ATRIAL MUSCLE CELLS.

Large populations (up to 600/cell) of spherical, electron-opaque granules approximately 0.3 to 0.4 micro in diameter are characteristically found in muscle fibers of mammalian atria. They are absent in muscle fibers of the ventricles. The granules are concentrated in the sarcoplasmic core and occur in lesser numbers in the sarcoplasmic layers between myofibrils and under the plasma membrane. Their intimate association with a central voluminous Golgi complex and the frequent occurrence of material reminiscent of the granular content within the cisternae of the Golgi complex suggest that the latter is involved in the formation of the atrial granules. Atrial granules are larger and more numerous in smaller species (rat, mouse), and generally smaller and less numerous in larger mammals (dog, cat, human); they are absent from the atrial fibers of very young fetuses (rat) but are present in those of newborn animals. A small population of bodies containing glycogen particles and remnants of the endoplasmic reticulum and mitochondria occurs in the sarcoplasmic cores of atrial as well as ventricular muscle fibers in the rat; they contain acid phosphatase and thus appear to be residual bodies of autolytic foci. Their frequency increases with the age of the animal. Typical lipofuscin pigment granules, which are known to contain acid phosphatase and are found in the sarcoplasmic cores in old animals (cat, dog and human), are presumed to arise by progressive aggregation and fusion of small residual bodies.

Animals↗
PubMed1964

LYTIC ACTIVITIES IN RENAL PROTEIN ABSORPTION DROPLETS. AN ELECTRON MICROSCOPICAL CYTOCHEMICAL STUDY.

The digestive cycle following reabsorption of hemoglobin by cells of the proximal convoluted tubules in mouse kidney and the uptake of ferritin by glomerular mesangial cells in the kidney of normal and nephrotic rats were investigated by electron microscopical histochemical procedures. Mouse kidneys, sampled at closely spaced time points between 1 to 48 hours after intraperitoneal injection of hemoglobin, and rat (normal and nephrotic) kidneys, sampled at 30 minutes, 2 hours, and 48 hours after intravenous injection of ferritin, were fixed in glutaraldehyde, cut at 50 micro on a freezing microtome, incubated for acid phosphatase and thiolacetate-esterase, and postfixed in OsO(4). Satisfactory preservation of fine structure permitted the localization of the enzymatic reaction products on cell structures involved in uptake and digestion of exogenous proteins. The latter were identified either by their density (hemoglobin) or their molecular structure (ferritin). It was found that lysosomal enzymic activities and incorporated exogenous proteins occur together in the same membrane-bounded structures. In the cells of the proximal convolution, lytic activities become demonstrable within 1 hour after hemoglobin injection, appear first in apical vacuoles filled with hemoglobin, and persist in fully formed protein absorption droplets. At the end of the lytic cycle ( approximately 48 hours post injection), the cells have an increased population of polymorphic bodies which exhibit lytic activities. In smaller numbers, identical bodies occur in controls. It is concluded that they represent remnants of previous digestive events. The means by which the resorptive vacuoles acquire hydrolytic activities remain unknown. Fusion of newly formed vacuoles with residual bodies was not seen, and hemoglobin incorporation into such bodies was only occasionally encountered. Acid phosphatase activity was found sometimes in the Golgi complex, but enzyme transport from the complex to the resorbing vacuoles could not be established. Autolytic vacuoles containing mitochondria or mitochondrial remnants were frequently found during the early stages of hemoglobin resorption, but no definite conclusions about the mechanism involved in the segregation of endogenous material were obtained. In nephrotic rats ferritin was segregated in membrane-bounded bodies mainly in the mesangial cells and to a lesser extent in epithelial and endothelial cells. Most of these sites were marked by the reaction products of acid phosphatase and organophosphorus-resistant esterase and therefore identified as lysosomes connected with the digestion of incorporated exogenous proteins.

Acid Phosphatase↗
PubMed1963

Junctional complexes in various epithelia.

The epithelia of a number of glands and cavitary organs of the rat and guinea pig have been surveyed, and in all cases investigated, a characteristic tripartite junctional complex has been found between adjacent cells. Although the complex differs in precise arrangement from one organ to another, it has been regularly encountered in the mucosal epithelia of the stomach, intestine, gall bladder, uterus, and oviduct; in the glandular epithelia of the liver, pancreas, parotid, stomach, and thyroid; in the epithelia of pancreatic, hepatic, and salivary ducts; and finally, between the epithelial cells of the nephron (proximal and distal convolution, collecting ducts). The elements of the complex, identified as zonula occludens (tight junction), zonula adhaerens (intermediary junction), and macula adhaerens (desmosome), occupy a juxtaluminal position and succeed each other in the order given in an apical-basal direction. The zonula occludens (tight junction) is characterized by fusion of the adjacent cell membranes resulting in obliteration of the intercellular space over variable distances. Within the obliterated zone, the dense outer leaflets of the adjoining cell membranes converge to form a single intermediate line. A diffuse band of dense cytoplasmic material is often associated with this junction, but its development varies from one epithelium to another. The zonula adhaerens (intermediate junction) is characterized by the presence of an intercellular space ( approximately 200 A) occupied by homogeneous, apparently amorphous material of low density; by strict parallelism of the adjoining cell membranes over distances of 0.2 to 0.5 micro; and by conspicuous bands of dense material located in the subjacent cytoplasmic matrix. The desmosome or macula adhaerens is also characterized by the presence of an intercellular space ( approximately 240 A) which, in this case, contains a central disc of dense material; by discrete cytoplasmic plaques disposed parallel to the inner leaflet of each cell membrane; and by the presence of bundles of cytoplasmic fibrils converging on the plaques. The zonula occludens appears to form a continuous belt-like attachment, whereas the desmosome is a discontinuous, button-like structure. The zomula adhaerens is continuous in most epithelia but discontinuous in some. Observations made during experimental hemoglobinuria in rats showed that the hemoglobin, which undergoes enough concentration in the nephron lumina to act as an electron-opaque mass tracer, does not penetrate the intercellular spaces beyond the zonula occludens. Similar observations were made in pancreatic acini and ducts where discharged zymogen served as a mass tracer. Hence the tight junction is impervious to concentrated protein solutions and appears to function as a diffusion barrier or "seal." The desmosome and probably also the zonula adhaerens may represent intercellular attachment devices.

Animals↗
PubMed1963

STUDIES ON ISOLATED NUCLEI. I. ISOLATION AND CHEMICAL CHARACTERIZATION OF A NUCLEAR FRACTION FROM GUINEA PIG LIVER.

This article describes a method for the isolation of nuclei from guinea pig liver. It involves the homogenization of the tissue in 0.88 M sucrose-1.5 mM CaCl(2) followed by centrifugation in a discontinuous density gradient in which the upper phase is the homogenate and the lower phase is 2.2 M sucrose-0.5 mM CaCl(2). Based on DNA recovery, the isolated fraction contains 25 to 30 per cent of the nuclei of the original homogenate. Electron microscopical observations showed that approximately 88 per cent of the isolated nuclei come from liver cells (the rest from von Kupffer cells and leucocytes) and that approximately 90 per cent of the nuclei appear intact, with well preserved nucleoli, nucleoplasm, nuclear envelope, and pores. Cytoplasmic contamination is minimal and consists primarily of the nuclear envelope and its attached ribosomes. The nuclear fraction consists of approximately 22.3 per cent DNA, approximately 4.7 per cent RNA, and approximately 73 per cent protein, the DNA/RNA ratio being 4.7. Data on RNA extractibility by phosphate and salt and on the base composition of total nuclear RNA are included.

Animals↗
PubMed1963

STUDIES ON ISOLATED NUCLEI. II. ISOLATION AND CHEMICAL CHARACTERIZATION OF NUCLEOLAR AND NUCLEOPLASMIC SUBFRACTIONS.

This paper describes the subfractionation of nuclei isolated from guinea pig liver by the procedure presented in the first article of the series (8). Centrifugation in a density gradient system of nuclear fractions disrupted by sonication permits the isolation of the following subfractions: (a) a nucleolar subfraction which consists mainly of nucleoli surrounded by a variable amount of nucleolus-associated chromatin and contaminated by chromatin blocks derived primarily from von Kupffer cell nuclei; (b) and (c), two nucleoplasmic subfractions (I and II) which consist mainly of chromatin threads in a coarser (I) or finer (II) degree of fragmentation. The protein, RNA, and DNA content of these subfractions was determined, and their RNA's characterized in terms of NaCl-solubility, nucleotide composition, and in vivo nucleotide turnover, using inorganic (32)P as a marker. The results indicate that there are at least three types of RNA in the nucleus (one in the nucleolus and two in the nucleoplasm or chromatin), which differ from one another in NaCl-solubility, nucleotide composition, turnover, and possibly sequence. Possible relations among these RNA's and those of the cytoplasm are discussed.

Animals↗
PubMed1962

Cytochemical study on the pancreas of the guinea pig. VII. Effects of spermine on ribosomes.

Pancreatic ribosomes (guinea pig) aggregate and lose upon treatment with polyamines, particularly spermine, their bound secretory enzymes. Spermine, at 0.5 mM, for example, causes the release of about 85 per cent of the chymotrypsinogen and RNase, and from 85 to 100 per cent of the ribosomal amylase. At the same time, the particles lose about 10 per cent of their RNA, 7 to 24 per cent of their total protein, and from 75 to 100 per cent of their Mg(++). Observations with the electron microscope confirm the heavy agglutinating of the ribosomes but otherwise show little change in the structure of the particles. Using radioactive spermine it was found that, concomitant with the loss of bound enzymes and Mg(++) from the ribosomes, spermine became bound to the particle. The extent of binding ranged from 0.29 to 1.49 micromoles per 10micromoles RNA-P. The bound radioactive spermine can be removed by subsequent treatment of the ribosomes with GTP, ATP, or P-P, which treatment also removes most of the RNA of the particles, leaving behind ribosomes with a much lower RNA/protein ratio. From this evidence it was inferred that spermine, in releasing the Mg(++) of the particle, becomes salt-linked to the free phosphate hydroxyl groups of the RNA. Freshly isolated pancreatic and hepatic ribosomes contain very little spermine, about 0.1 to 0.2 micromoles polyamine/10 micromoles RNA-P. The results are discussed in terms of the linkages between the structural protein, the bound secretory enzymes, and the RNA of the ribosomes.

Guinea Pigs↗
PubMed1961

Glomerular permeability. II. Ferritin transfer across the glomerular capillary wall in nephrotic rats.

Ferritin was used as a tracer to investigate glomerular permeability in the nephrotic rat. The results were compared with those previously obtained in normal animals. A nephrotic syndrome was induced by 9 daily injections of the aminonucleoside of puromycin. Ferritin was administered intravenously on the 10th day, and kidney tissue was fixed at intervals of 5 minutes to 44 hours after injection of the tracer and examined by electron microscopy. The observations confirmed that at this stage of the experimental nephrotic syndrome the changes affect predominantly the visceral epithelium (loss of foot processes, reduction and modification of urinary slits, and intracellular accumulation of vacuoles and protein absorption droplets). Less extensive changes were found in other layers (reduction of endothelial fenestrae, an increase in the population of "deep" cells, and a thinning and "loosening" of the basement membrane.) At short intervals (5 to 15 minutes) after ferritin administration, the tracer was found at high concentration in the lumen and endothelial fenestrae, and at decreasing concentrations embedded throughout the basement membrane and incorporated into the epithelium (within cytoplasmic vesicles and within invaginations of the plasmalemma facing the basement membrane). After longer intervals (1 to 3 hours) the distribution of the tracer within the capillary wall was similar except that its concentration in the epithelium was higher, and, in addition to plasma membrane invaginations and small vesicles, ferritin also marked larger vacuoles, dense bodies, and intermediate forms. Large accumulations of tracer typically occurred in the spongy areas of the basement membrane, especially in the axial regions. Ferritin also appeared in the endothelium within membrane-limited vacuoles and dense bodies, particularly in the deep cells. After 6 to 44 hours the tracer still occurred in the lumen and throughout the basement membrane. The ferritin deposits in the spongy areas as well as the ferritin-containing vacuoles of the deep endothelium were larger and more numerous. In the epithelium ferritin was found not only within various membrane-limited bodies, but also "free" within the cytoplasmic matrix. These observations indicate that in the nephrotic glomerulus, as in the normal, the basement membrane functions as the main filtration barrier; however, in nephrosis, the basement membrane is defective and allows leakage of increased quantitites of ferritin and presumably plasma proteins. The basement membrane defect appears to be fine and widespread, occurring at or near the molecular level of organization of the filter. The accumulation of unfiltered ferritin in axial regions together with the demonstration of its subsequent phagocytosis by the "deep" endothelial cells suggest that the latter may function in the removal of filtration residues. Finally, the findings indicate that in the nephrotic, as in the normal animal, the epithelium acts as a monitor that recovers, at least in part, the protein which leaks through the filter, and that in nephrosis, the recovering activities of the epithelium are greatly enhanced because of the increased permeability of the basement membrane.

Animals↗
PubMed1961

Glomerular permeability. I. Ferritin transfer across the normal glomerular capillary wall.

Ferritin was used as a tracer to investigate pathways and mechanisms for transfer across the various layers of the glomerular capillary wall. Kidney tissue, fixed at intervals of 2 minutes to 2 hours following an intravenous injection of ferritin, was examined by electron microscopy. The observations confirmed the existence of three distinct and successive layers in the glomerular capillary wall (the endothelium, the basement membrane, and the visceral epithelium). In addition, they demonstrated a number of new structural features: namely (a) discrete fibrils in the subendothelial spaces; (b) a characteristic, highly elaborate, cytoplasmic organization in the visceral epithelium; and (c) special structures resembling "desmosomes" in the slits between foot processes. In animals sacrificed at short time intervals (2 to 15 minutes) following ferritin administration, ferritin molecules were found at high concentration in the lumen and endothelial fenestrae, at low concentration in the basement membrane, and in very small numbers within the epithelium. Later (1 to 2 hours), the tracer particles were still present in the lumen and within endothelial fenestrae, and, in addition, had accumulated on the luminal side of the basement membrane, especially in the axial regions of the vessels. Larger numbers of ferritin molecules were also found in the epithelium-in invaginations of the cell membrane at the base of the foot processes, and in various membrane-limited bodies (vesicles, multivesicular bodies, vacuoles, and dense bodies) present within the cytoplasm. These observations suggest that the endothelial fenestrae are patent and that the basement membrane is the main filtration barrier. Since the basement membrane has no demonstrable pores, it is probably not a simple sieve but presumably is a gel-like structure with two fine fibrillar components embedded in an amorphous matrix. Both the epithelium and endothelium may be concerned with building and maintaining this structure. Finally, the intracellular accumulation of particles in the epithelium suggests that the latter acts as a monitor that recovers, at least in part, the small amounts of protein which normally leak through the filter.

Animals↗
PubMed1961

Studies on inflammation. 1. The effect of histamine and serotonin on vascular permeability: an electron microscopic study.

The mechanism, whereby histamine and serotonin increase the permeability of blood vessels, was studied in the rat by means of the electron microscope. The drugs were injected subcutaneously into the scrotum, whence they diffused into the underlying (striated) cremaster muscle. An intravenous injection of colloidal HgS was also given, in order to facilitate the identification of leaks by means of visible tracer particles. After intervals varying from 1 minute to 57 days the animals were killed; the cremaster was fixed, embedded in methacrylate, and examined with the electron microscope. One to 12 minutes after the injection, the blood vessels of the smallest caliber (3 to 5 micra as measured on electron micrographs) appeared intact. Numerous endothelial openings were present in blood vessels with a diameter of 7 to 8 micra or more. These gaps were 0.1 to 0.8 micra in width; portions of intercellular junctions were often present in one or both of the margins. The underlying basement membrane was morphologically intact. An accumulation of tracer particles and chylomicra against the basement membrane indicated that the latter behaved as a filter, allowing fluid to escape but retaining and concentrating suspended particulate matter of the size used. Uptake of tracer particles by endothelial vesicles was minimal. Phagocytosis by endothelial cells became more prominent at 3 hours, but as a secondary occurrence; the pericytes were actively phagocytic at all stages. At the 3-hour stage no leaks were found. The changes induced by histamine and serotonin were indistinguishable, except that the latter was more potent on a mole-to-mole basis. In control animals only small accumulations of tracer particles were found in the wall of a number of blood vessels. With regard to the pathogenesis of the endothelial leaks, the electron microscopic findings suggested that the endothelial cells become partially disconnected along the intercellular junctions. Supporting evidence was provided at the level of the light microscope, by demonstrating-in the same preparation-the leaks with appropriate tracer particles(1), and the intercellular junctions by the silver nitrate method. The lipid nature of the chylomicron deposits observed in electron micrographs was also confirmed at the level of the light microscope, using cremasters fixed in formalin and stained in toto with sudan red.

Animals↗
PubMed1961

Studies on inflammation. II. The site of action of histamine and serotonin along the vascular tree: a topographic study.

While it is an established fact that histamine and serotonin increase the permeability of blood vessels, the exact portion of the vascular tree which is so affected has not been conclusively demonstrated. The present study was undertaken to clarify this point. Our experiments were based on a method to which we refer as "vascular labeling," and which permits one to identify leaking vessels by means of visible accumulations of foreign particles within their walls. The mechanism of the labeling, elucidated by previous electron microscopic studies, is the following. Histamine and serotonin cause the endothelial cells of certain vessels to separate, and thus to create discrete intercellular gaps. Plasma escapes through these gaps, and filters through the basement membrane. If the plasma has been previously loaded (by intravenous injection) with colloidal particles of a black material such as carbon or mercuric sulfide, these particles-too large to pass through the basement membrane-will be retained and accumulate in visible amounts within the wall of the leaking vessel. This method is used to maximal advantage if the tissue is cleared and examined by transillumination in toto, so that leaking vessels can be accurately identified in their relationship to the vascular tree. As a test tissue we used the rat cremaster, a laminar striated muscle which can be easily excised with its vascular supply virtually intact. The rats were prepared with an intravenous injection of carbon or HgS, and a subcutaneous injection into the scrotum of histamine, serotonin, or NaCl (as a control). The injected drug diffused into the underlying cremaster and the vessels became labeled. One hour later, when the carbon had been cleared from the blood stream, the animal was killed. The cremaster was excised, stretched, fixed in formalin, cleared in glycerin, and examined by transillumination under a light microscope. The lesions induced by histamine and serotonin were identical. The leaking vessels, as indicated by the carbon deposits, always belonged to the venous side of the circulation. The heaviest deposits were found in venules 20 to 30 micra in diameter. The deposits decreased towards larger venules up to a maximum diameter of 75 to 80 micra, and towards the finer vessels until the caliber reached approximately 7 micra. Essentially spared by the deposits were the finest vessels, 4 to 7 micra in diameter, and constituting an extensive network oriented along the muscular fibers. By killing animals at varying intervals after the injections, it was found that the carbon particles were slowly removed from the vascular walls by the action of phagocytic cells. After 10 months there was still enough carbon locally to be recognized by the naked eye.

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