Biomedical subjects

C Zurzolo

Publications and source records attributed to C Zurzolo.

24 records · Page 2Linked to original sources

PubMed1990

Human CD8 alpha glycoprotein is expressed at the apical plasma membrane domain in permanently transformed MDCK II clones.

Madin-Darby canine kidney cells (MDCK II) have been cotransfected with plasmids expressing the human CD8 alpha glycoprotein and the bacterial gene which confers resistance to neomycin. Stable transformants have been isolated in the presence of G-418 in the culture medium and screened for CD8 alpha expression by immunofluorescence. The three clones we have characterized showed: 1) high level of synthesis and efficient surface expression of glycosylated, homodimeric CD8 alpha and 2) preferential apical deposition of CD8 alpha in confluent monolayers. This polar distribution has been measured in cells grown on a plastic substratum as well as on nitrocellulose filter by means of EM immunocytochemistry and surface radioimmunoassay. CD8 alpha was 6 to 11-fold enriched on the apical membrane whereas the 58 kDa protein, a basolateral marker in MDCK II cells, resulted about 9-fold enriched on the basolateral membrane of the three clones. We believe these permanently transformed clones could prove to be a useful tool with which to study cell polarity.

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PubMed1990

Morphological changes induced by prolonged TSH stimulation or starvation in the rat thyroid cell line FRTL.

Subconfluent FRTL cells were examined in the electron microscope after the following treatments: i) continuous TSH stimulation; ii) starvation from TSH for 3 or 7 days; iii) seven days of TSH deprivation followed by two days of TSH stimulation. The organelle complement of cells grown in the presence of TSH appeared to be consistent with their secretory properties. Rough endoplasmic reticulum (RER), in the form of round vesicles, and Golgi apparatus, were quite prominent. Cells were not properly polarized. Their dorsal surface was covered with microvilli and occasional pseudopods. After TSH withdrawal the cells flattened on the dish, lost most of their plasma membrane specializations and reorganized actin stress fibers. RER shape was modified from round vesicles to flat cisternae which thereafter almost completely disappeared. The Golgi apparatus did not seem to be modified. Autophagic vacuoles became more prominent. All the modifications were fully reversed after TSH replacement indicating an hormonal regulation in the amount and morphology of some organelle, in particular of the RER.

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PubMed1988

Synthesis of extracellular matrix glycoproteins by a differentiated thyroid epithelial cell line.

We examined the synthesis of extracellular matrix macromolecules by the differentiated rat thyroid epithelial cell line FRTL-5. As shown by electron microscopy, the extracellular material produced by these cells is deposited at the basolateral surface and focally organized in the form of a basement membrane. Biochemical and biosynthetic studies demonstrated that laminin, type IV collagen, and fibronectin are synthesized and deposited in the culture monolayer. Secretion of fibronectin into the culture medium also occurred. By immunofluorescence we observed some peculiarities in the distribution patterns of the basement membrane glycoproteins; while fibronectin and laminin had an almost superimposable distribution, type IV collagen displayed a rather different pattern. Type IV collagen and laminin localization at sites where extracellular material was detected was confirmed by immuno electronmicroscopy using the protein A-colloidal gold technique. The results indicate that under appropriate culture conditions the differentiated thyroid epithelial cell line FRTL-5 synthesizes, secretes and organizes an extracellular matrix where some basement membrane glycoproteins are present.

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PubMed1986

In vivo biosynthesis of clathrin and other coated vesicle proteins from rat liver.

A biosynthetic study of rat liver coated vesicle (CV) proteins was undertaken by using in vivo labeling with L-[35S]methionine. CVs were isolated and purified by using standard procedures and characterized by electron microscopy, sedimentation, and sodium dodecyl sulfate polyacrylamide gel electrophoresis followed by fluorography, or by gel slicing and liquid scintillation counting. After 5 1/2 min of labeling (the earliest time examined), incorporation of radioactive clathrin heavy-chain (180-kD (kilodalton] subunits as well as a 90-kD CV-associated protein into purified CVs was demonstrated. The level of labeled 180-kD clathrin in coated vesicles increased rapidly during the first 2 hr of labeling and then continued to rise at a slower rate between 4 and 16 hr. This slow accumulation of labeled clathrin heavy chains in the CV pool may reflect early compartmental sequestration of a fraction of newly synthesized clathrin with delayed assembly into free CVs. By 16 hr of labeling, clathrin 180-kD chains and the 90-kD CV-associated protein accounted for approximately 48 and 26%, respectively, of the radioactivity in all CV proteins. Two proteins of MWa 68 kD and 53 kD showed marked declines in cpm/unit protein between 30 min and 4 hr, raising the possibility that these species may be transferred out of CVs during or after transport without loss of the other CV proteins. The possibility is also raised that clathrin heavy chains may be recycled during CV formation. Possible heterogeneity within individual CV preparations with respect to protein composition and derivation from both plasma membrane and Golgi regions are proposed.

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PubMed1986

Functional properties of normal and inverted rat thyroid follicles in suspension culture.

Rat thyroid follicles in both the normal and the inverted configuration have been cultured in suspension. It has been found that normal follicles (i.e., those having a structural organization similar to the follicles in vivo) express most functional properties of the thyroid gland. They trap iodide, synthesize, secrete, and iodinate thyroglobulin, and secrete thyroid hormone into the culture medium. On the contrary, in inverted follicles (i.e., those having a cell polarity reversed with respect to normal) we were unable to detect expression of some of these functions. We conclude that both the follicular organization and the proper cell polarity are needed for the full expression of thyroid functional properties.

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PubMed1985

Coated vesicles from the thyroid gland: isolation, characterization, and a search for a possible role in thyroglobulin transport.

We report the first isolation of purified coated vesicles (CVs) from thyroid gland. Bovine thyroid CVs were isolated by differential centrifugation, including a step through sucrose-D2O, using a modification of the method described by Nandi et al. (1) for bovine brain CVs. The CVs were characterized by electron microscopy, sedimentation properties, and SDS-PAGE of the protein components. Thyroglobulin (Tg) was found to be associated with the purified CVs. When the thyroid CVs were exposed to conditions known to remove the protein coat from brain CVs, such as low ionic strength at pH 8.5, most of the Tg dissociated from the vesicles along with the coat proteins. Moreover, the Tg remaining with the uncoated vesicles (UVs) was trypsin sensitive, and therefore judged to be associated with the external surface of the vesicle. Since ligand-receptor complexes are normally located within CVs and not on their outer surface, no evidence was found for Tg-receptor complexes within thyroid CVs. Thyroid slices were incubated in the presence of [35S] methionine with subsequent isolation of labeled CVs in order to study the incorporation of newly-synthesized proteins into these structures. At 0.5 and 2 hours of incubation, the 180K MW subunit of clathrin, as well as other proteins, but not Tg, had become labeled in the purified CVs. Extracellular 19S-[35S] thyroglobulin was isolated from the incubation medium, however, demonstrating release of newly-synthesized Tg (presumably into cut follicles). It is concluded that thyroid CVs do not seem to be involved in the secretion of newly-synthesized Tg from the rough endoplasmic reticulum into the follicular lumen. While a possible role of thyroid CVs in the reabsorption of small quantities Tg by micropinocytosis cannot be completely excluded, the present data do not support a primary role for thyroid CVs in either endocytosis or exocytosis of Tg.

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