scholarly journals An electron microscope autoradiographic study of the carbohydrate recognition systems in rat liver. II. Intracellular fates of the 125I-ligands.

1979 ◽  
Vol 83 (1) ◽  
pp. 65-81 ◽  
Author(s):  
A L Hubbard ◽  
H Stukenbrok
Keyword(s):  
Electron Microscope ◽  
Cell Types ◽  
Autoradiographic Study ◽  
Dense Bodies ◽  
Golgi Region ◽  
Recognition Systems ◽  
Secondary Lysosomes ◽  
Different Cell Types ◽  
Intracellular Pathway ◽  
Peripheral Cytoplasm

Electron microscope autoradiographic and biochemical methods were used to study the intracellular fates of several 125I-glycoproteins, known to be specifically bound and internalized by the different cell types in the liver. At the earliest times examined (1--2 min), 125I-glycoproteins (ASGP) were localized predominantly along the sinusoidal front of hepatocytes. Analysis of the distribution of autoradiographic grains indicated that: (a) approximately 40--60% of the 125I-ligand could be ascribed to the plasmalemma; (b) a significant fraction had already been internalized; yet (c) very little 125I-ligand was present in the lysosome-Golgi region. Between 4 and 15 min after administration of 125I-ASGPs, there was a dramatic redistribution of autoradiographic grains from regions of the plasmalemma and peripheral cytoplasm (30% decrease) to the lysosome-Golgi region (30% increase). At longer times (30 min), there was continued drainage of 125I-ASGP into this region. The grain density over secondary lysosomes was 60--90 times higher than that over recognizable Golgi elements, clearly indicating that lysosomes were the ultimate destination of the 125I-ASGP. However, no more than 60% of the total 125I-ligand could be localized to lysosome-rich regions of the hepatocyte, with the remaining 40% primarily in the intermediate cytoplasm. Biochemical evidence for proteolysis of the internalized 125I-ASGP (presumably within lysosomes) was obtained when [125I]-mono-iodotyrosine was found in the liver (i.e., hepatocytes) at times later than 15 min. The temporal redistribution observed for mannose and N-acetylglucosamine-terminated glycoproteins (ahexosamino-orosomucoid and agalacto-orosomucoid, respectively) in endothelial cells indicated that the 125I-ligands resided in macropinocytic vesicles (1--15 min) before their ultimate residence in dense bodies (15 min). The same 125I-ligands were also localized to structures resembling secondary lysosomes in Kupffer cells. The lysosomal nature of "these organelles" was implied from the appearance of [125I]mono-iodotyrosine in the liver at later times. 125I-beta-glucuronidase followed the same intracellular pathway in both cell types but was not degraded.

scholarly journals MIGRATION OF GLYCOPROTEIN FROM THE GOLGI APPARATUS TO THE SURFACE OF VARIOUS CELL TYPES AS SHOWN BY RADIOAUTOGRAPHY AFTER LABELED FUCOSE INJECTION INTO RATS

1974 ◽  
Vol 60 (1) ◽  
pp. 258-284 ◽  
Author(s):  
Gary Bennett ◽  
C. P. Leblond ◽  
Antonio Haddad
Keyword(s):  
Plasma Membrane ◽  
Electron Microscope ◽  
Golgi Apparatus ◽  
Early Time ◽  
Cell Types ◽  
Cell Type ◽  
Time Intervals ◽  
Cell Coat ◽  
Golgi Region ◽  
Silver Grains

A single intravenous injection of L-[3H]fucose, a specific glycoprotein precursor, was given to young 35–45 g rats which were sacrificed at times varying between 2 min and 30 h later. Radioautography of over 50 cell types, including renewing and nonrenewing cells, was carried out for light and electron microscope study. At early time intervals (2–10 min after injection), light microscope radioautography showed a reaction over nearly all cells investigated in the form of a discrete clump of silver grains over the Golgi region. This reaction varied in intensity and duration from cell type to cell type. Electron microscope radioautographs of duodenal villus columnar cells and kidney proximal and distal tubule cells at early time intervals revealed that the silver grains were restricted to Golgi saccules. These observations are interpreted to mean that glycoproteins undergoing synthesis incorporate fucose in the saccules of the Golgi apparatus. Since fucose occurs as a terminal residue in the carbohydrate side chains of glycoproteins, the Golgi saccules would be the site of completion of synthesis of these side chains. At later time intervals, light and electron microscope radioautography demonstrated a decrease in the reaction intensity of the Golgi region, while reactions appeared over other parts of the cells: lysosomes, secretory material, and plasma membrane. The intensity of the reactions observed over the plasma membrane varied considerably in various cell types; furthermore the reactions were restricted to the apical surface in some types, but extended to the whole surface in others. Since the plasma membrane is covered by a "cell coat" composed of the carbohydrate-rich portions of membrane glycoproteins, it is concluded that newly formed glycoproteins, after acquiring fucose in the Golgi apparatus, migrate to the cell surface to contribute to the cell coat. This contribution implies turnover of cell coat glycoproteins, at least in nonrenewing cell types, such as those of kidney tubules. In the young cells of renewing populations, e.g. those of gastro-intestinal epithelia, the new glycoproteins seem to contribute to the growth as well as the turnover of the cell coat. The differences in reactivity among different cell types and cell surfaces imply considerable differences in the turnover rates of the cell coats.

Cytoplasmic fine-structure

1958 ◽  
Vol 148 (932) ◽  
pp. 290-308 ◽  
Keyword(s):  
Fine Structure ◽  
Electron Microscope ◽  
Large Body ◽  
Cell Types ◽  
General Terms ◽  
Thin Sectioning ◽  
Cytoplasmic Change ◽  
Cellular Specialization ◽  
Different Cell Types ◽  
Made In

In any attempt to reach an integrated conception of the cytoplasm in variation and development, a study of the fine-structure of the cytoplasm and its relation to the nucleus must take its proper place. It is the object of our paper to survey, as adequately as we are able in a limited space, selected data on cytoplasmic fine-structure and we hope that this will provide the Discussion with a background against which to consider a morphological basis for that variation which genetical studies may show to be due to changes in the organization of the cytoplasm. It is possibly too early as yet to hope that examination of the morphology of cells by means of the electron microscope will reveal cytoplasmic differences between organisms which differ from one another in the characteristics studied in genetical experiments; it would be useful to the future study of the problem of Cytoplasmic change, however, to know within what limits speculation must be reasonably confined by the nature of the fine-structure of the cytoplasm. It is now becoming apparent that though cells of organisms widely separated phylogenetically have basic similarities, cellular specialization has led to some diversity in the fine-structure. In the first part of the paper we shall briefly consider the development of electron-microscope methods, e.g. the thin-sectioning procedures, which have made it possible to examine biological material at a resolution which allows comparatively small macromolecular units to be distinguished (10 to 50 Å); at the same time we shall emphasize the danger of overestimating the significance of the observations that have been made. In the second part we shall consider certain selected features of the cell in some detail; in view of the large body of literature on cell fine-structure that is now available (publications numbered over 100 during the last 6 months of 1956) no attempt will be made to review all the findings which have been published during the last few years. Rather we will consider, in general terms, the structure of each component, then compare the variations in structural form noted in different cell types and indicate where there is direct disagreement in the findings of various authorities.

scholarly journals The rubella virus E2 and E1 spike glycoproteins are targeted to the Golgi complex.

1993 ◽  
Vol 121 (2) ◽  
pp. 269-281 ◽  
Author(s):  
T C Hobman ◽  
L Woodward ◽  
M G Farquhar
Keyword(s):  
Membrane Proteins ◽  
Golgi Complex ◽  
Rubella Virus ◽  
Cell Types ◽  
Vero Cells ◽  
Type I ◽  
Surface Transport ◽  
Golgi Stack ◽  
Golgi Region ◽  
Different Cell Types

Rubella virus (RV) has been reported to bud from intracellular membranes in certain cell types. In this study the intracellular site of targeting of RV envelope E2 and E1 glycoproteins has been investigated in three different cell types (CHO, BHK-21 and Vero cells) transfected with a cDNA encoding the two glycoproteins. By indirect immunofluorescence, E2 and E1 were localized to the Golgi region of all three cell types, and their distribution was disrupted by treatment with BFA or nocodazole. Immunogold labeling demonstrated that E2 and E1 were localized to Golgi cisternae and indicated that the glycoproteins were distributed across the Golgi stack. Analysis of immunoprecipitates obtained from stably transfected CHO cells revealed that E2 and E1 become endo H resistant and undergo sialylation without being transported to the cell surface. Transport of RV glycoproteins to the Golgi complex was relatively slow (t1/2 = 60-90 min). Coprecipitation experiments indicated that E2 and E1 form a heterodimer in the RER. E1 was found to fold much more slowly than E2, suggesting that the delay in transport of the heterodimer to the Golgi may be due to the slow maturation of E1 in the ER. These results indicate that RV glycoproteins behave as integral membrane proteins of the Golgi complex and thus provide a useful model to study targeting and turnover of type I membrane proteins in this organelle.

scholarly journals An electron microscope autoradiographic study of the carbohydrate recognition systems in rat liver. I. Distribution of 125I-ligands among the liver cell types.

1979 ◽  
Vol 83 (1) ◽  
pp. 47-64 ◽  
Author(s):  
A L Hubbard ◽  
G Wilson ◽  
G Ashwell ◽  
H Stukenbrok
Keyword(s):  
Endothelial Cells ◽  
Electron Microscope ◽  
Kupffer Cells ◽  
Cellular Localization ◽  
Cell Types ◽  
Autoradiographic Study ◽  
Rnase A ◽  
Electron Microscope Autoradiography ◽  
A Cell

Electron microscope autoradiography was used to study the cellular localization of seven glycoproteins rapidly cleared from the circulating plasma of rats and taken up by the liver. 1 and 15 min after intravenous administration of the 125I-glycoproteins, livers were fixed in situ by perfusion and processed for autoradiography. Autoradiographic grains in the developed sections were found to represent the intact 125I-ligand. A quantitative analysis of the distribution and concentration (density) of autoradiographic grains over the three major cell types of the liver was then performed. Three molecules, asialo-fetuin, asialo-orosomucoid, and lactosaminated RNase A dimer, the oligosaccharide chains of which terminate in galactose residues, were bound and internalized almost exclusively (greater than 90%) by hepatocytes. Conversely, four molecules, the oligosaccharide chains of which terminate in either N-acetyl-glucosamine (agalacto-orosomucoid) or mannose (ahexosamino-orosomucoid, preputial beta-glucuronidase, and mannobiosaminated RNase A dimer), were specifically bound and internalized by cells lining the blood sinusoids--that is, by Kupffer cells and endothelial cells. Endothelial cells were two to six times more active (on a cell volume basis) than were Kupffer cells in the internalization of these four 125I-ligands. Mannose and N-acetylglucosamine-terminated glycoproteins competed with each other for uptake into either endothelial cells or Kupffer cells, indicating that a single system recognized mannose or N-acetyl-glucosamine residues. Finally, agalacto-orosomucoid and ahexosamino-orosomucoid were also associated with hepatocytes, but competition experiments utilizing excess asialo-orosomucoid demonstrated that residual galactosyl residues were responsible for this association.

Synthesis and Deposition of Oocyte Envelopes (Vitelline Membrane, Chorion) and the Uptake of Yolk in the Dragonfly (Odonata: Aeschnidae)

1969 ◽  
Vol 4 (1) ◽  
pp. 241-264
Author(s):  
H. W. BEAMS ◽  
R. G. KESSEL
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Electron Microscope ◽  
Follicle Cells ◽  
Vitelline Membrane ◽  
Extracellular Material ◽  
Dense Bodies ◽  
Golgi Region ◽  
Inner Surface ◽  
Oocyte Envelopes

Light and electron-microscope studies on dragonfly ovarioles reveal evidence that the precursor vitelline membrane and chorion secretions are synthesized within the follicle cells. It is suggested that the sequence of synthesis and deposition of the vitelline membrane occurs as follows. The vitelline membrane presecretion appears to be synthesized by the rough surfaced endoplasmic reticulum, giving rise to intracisternal granules. These appear to migrate in the cisternae to the region of the Golgi complex where the endoplasmic reticulum loses most of its ribosomes and the intracisternal granules move into the Golgi region where they appear within small vesicles. These seem to find their way into the Golgi cisternae where they may be incorporated with the secretions from the Golgi cisternae to produce the definitive previtelline secretion. The previtelline secretion bodies are eventually discharged into the space between the oocyte and follicle cells, forming rows of secretion bodies between the microvilli. These fuse into progressively larger bodies until a complete membrane is established. Follicle cells actively secreting precursor vitelline membrane substance show many disk-shaped, relatively clear vesicles in the cytoplasm. After the vitelline membrane is laid down, the follicle cells take on an entirely different function; namely, the synthesis and deposition of the chorion. The first visible chorion secretion appears in profile as elongate dense bodies within the Golgi cisternae which tend to coil, and in so doing, expand the cisternae. As this occurs, the enlarged cisterna, loaded with concentric coiled secretion material, separates from the remainder of the Golgi cisternae and becomes free in the cytoplasm as a prechorion secretion body. These migrate to, and collect below, the surface of the cell where they are eventually ejected between the surface folds and become incorporated into the developing chorion. Uptake of yolk in the dragonfly seems to be predominantly by micropinocytosis. The oocyte surface during active vitellogenesis bears many pits which contain an extracellular material closely applied to the outer surface of the plasma membrane. Thin, radially oriented bristles are continuous with the inner surface of the plasma membrane in this region. The pits continue to invaginate until they are cut off from the plasma membrane and come to lie in the oocyte cortex as coated vesicles. These appear to lose their coats gradually and fuse with one another to produce definitive yolk spheres.

Study of the Molecular Architecture of Keratin Filaments by Electron Microscopy

1982 ◽  
Vol 40 ◽  
pp. 118-119
Author(s):  
U. Aebi ◽  
P. Rew ◽  
T.-T. Sun
Keyword(s):  
Electron Microscopy ◽  
Structural Organization ◽  
Cell Types ◽  
Structural Features ◽  
Molecular Architecture ◽  
The Past ◽  
Keratin Filaments ◽  
Different Types ◽  
10 Nm Filaments ◽  
Different Cell Types

Various types of intermediate-sized (10-nm) filaments have been found and described in many different cell types during the past few years. Despite the differences in the chemical composition among the different types of filaments, they all yield common structural features: they are usually up to several microns long and have a diameter of 7 to 10 nm; there is evidence that they are made of several 2 to 3.5 nm wide protofilaments which are helically wound around each other; the secondary structure of the polypeptides constituting the filaments is rich in ∞-helix. However a detailed description of their structural organization is lacking to date.

Copper Localization in Cirrhotic Rat Liver by Scanning Electron Microscopy

1969 ◽  
Vol 27 ◽  
pp. 38-39 ◽  
Author(s):  
J. C. Russ ◽  
E. McNatt
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Electron Microscope ◽  
Copper Acetate ◽  
Lead Citrate ◽  
Dense Bodies ◽  
Transmission Electron ◽  
Electron Mode ◽  
Scanning Electron

In order to study the retention of copper in cirrhotic liver, rats were made cirrhotic by carbon tetrachloride inhalation twice weekly for three months and fed 0.2% copper acetate ad libidum in drinking water for one month. The liver tissue was fixed in osmium, sectioned approximately 2000 Å thick, and stained with lead citrate. The section was examined in a scanning electron microscope (JEOLCO JSM-2) in the transmission electron mode.Figure 1 shows a typical area that includes a red blood cell in a sinusoid, a disse, and a portion of the cytoplasm of a hepatocyte which contains several mitochondria, peribiliary dense bodies, glycogen granules, and endoplasmic reticulum.

Processing and Characterization of Recombinant von Willebrand Factor Expressed in Different Cell Types Using a Vaccinia Virus Vector

1992 ◽  
Vol 67 (01) ◽  
pp. 154-160 ◽  
Author(s):  
P Meulien ◽  
M Nishino ◽  
C Mazurier ◽  
K Dott ◽  
G Piétu ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Von Willebrand Factor ◽  
Human Fibroblasts ◽  
Active Form ◽  
Cell Types ◽  
Biologically Active ◽  
L Cells ◽  
Von Willebrand ◽  
Different Cell Types ◽  
Willebrand Factor

SummaryThe cloning of the cDNA encoding von Willebrand factor (vWF) has revealed that it is synthesized as a large precursor (pre-pro-vWF) molecule and it is now clear that the prosequence or vWAgll is responsible for the intracellular multimerization of vWF. We have cloned the complete vWF cDNA and expressed it using a recombinant vaccinia virus as vector. We have characterized the structure and function of the recombinant vWF (rvWF) secreted from five different cell types: baby hamster kidney (BHK), Chinese hamster ovary (CHO), human fibroblasts (143B), mouse fibroblasts (L) and primary embryonic chicken cells. Forty-eight hours after infection, the quantity of vWF antigen found in the cell supernatant varied from 3 to 12 U/dl depending on the cell type. By SDS-agarose gel electrophoresis, the percentage of high molecular weight forms of vWF varied from 39 to 49% relative to normal plasma for BHK, CHO, 143B and chicken cells but was less than 10% for L cells. In all cell types, the two anodic subbands of each multimer were missing. The two cathodic subbands were easily detected only in BHK and L cells. By SDS-PAGE of reduced samples, pro-vWF was present in similar quantity to the fully processed vWF subunit in L cells, present in moderate amounts in BHK and CHO and in very low amounts in 143B and chicken cells. rvWF from all cells bound to collagen and to platelets in the presence of ristocetin, the latter showing a high correlation between binding efficiency and degree of multimerization. rvWF from all cells was also shown to bind to purified FVIII and in this case binding appeared to be independent of the degree of multimerization. We conclude that whereas vWF is naturally synthesized only by endothelial cells and megakaryocytes, it can be expressed in a biologically active form from various other cell types.

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