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51.
Extracellular Fluid Proteins of Goldfish Brain: Evidence for the Presence of Proteases and Esterases 总被引:1,自引:1,他引:0
Preparations of enriched fractions of extracellular fluid (ECF) proteins from goldfish brain were found to contain protease(s) and esterase(s). The N-substituted furanacryloyl (FA) peptides FA-Phe-Gly-Gly and FA-Phe-OMe were used as model substrates for determining protease and esterase activity, respectively, in a spectrophotometric assay. Studies of the profile of substrate specificity and identification of the types of compounds that were effective as inhibitors showed that these ECF enzymes have some distinctive properties. GSH, but not GSSG, and EDTA inhibited the protease(s) without influencing the esterase(s), whereas L-1-tosylamide-2-phenylethylchloromethyl ketone blocked both protease and esterase activities of ECF. Most of the protease and esterase properties of ECF could be bound to concanavalin A-Sepharose affinity chromatographic columns in association with ependymin--a brain extracellular protein. These observations indicate that ECF may contain a metalloprotease(s) and raise the possibility that the ependymins might be a substrate for these ECF enzymes. 相似文献
52.
Philip Skehan James E. Thomas Susan J. Friedman 《In vitro cellular & developmental biology. Plant》1986,22(11):632-636
Summary Sarcoma 180 monolayers spontaneously shed single cells and small multicellular aggregates into the surrounding medium to produce
a dual population of floating and substratum-attached cells. Shedding was a motility-associated event that occurred when cells
attempted to migrate over one another. It resulted from a combination of cell shape change and active motility, which increased
sensitivity to fluid shear dislodgement by reducing a cell's surface area of adhesive contact and increasing strain tension
at its adhesive contact points. Shedding occurred at all phases of the cell cycle. Extracellular matrix but not conditioned
medium enhanced the floating subpopulation by slowing the kinetics of rattachment to plastic and cellular substrata. Although
sarcoma 180 cells are anchorage independent in the sense that they grow readily in single cell suspension, they nevertheless
exhibited anchorage modulation of their cell cycle. Short periods in suspension produced a mild G1 accumulation, whereas longer periods of anchorage deprivation led to a mild G2 accumulation which appeared to result from an interference with cytokinesis.
This work was supported by grants from the Medical Research Council of Canada, The National Cancer Institute of Canada, the
Alberta Heritage Savings and Trust Fund for Applied Cancer Research, and the Alberta Heritage Fund for Medical Research. 相似文献
53.
Michael M. Lipsky Talia R. Sheridan Richard O. Bennett Eric B. May 《In vitro cellular & developmental biology. Plant》1986,22(6):360-362
Summary Comparisons were made of attachment and viability of rainbow trout (Salmo gairdneri) hepatocytes in short-term (2 days), primary culture on plastic, collagen-coated or extracellular matrix (ECM) coated dishes.
Hepatocyte isolation routinely yielded cells with good viability (96%). Cells plated on ECM attached with high efficiency
(93%) in contrast to cells cultured on plastic or collagen (∼20%). The cells plated on ECM flattened out and formed monolayers,
while the cells on plastic and collagen rounded up and formed multi-cell aggregates in suspension. Viability of cells in all
substrates remained high over the 2 day culture period. ECM is the first substrate to support trout-hepatocyte attachment
in primary culture. Differentiated liver function was maintained in cells cultured on ECM as evidence by the induction of
tyrosine aminotransferase by hydrocortisone (200%).
This work was supported in part by research grant R809599010 from the U. S. Environmental Protection Agency.
Editor's Statement This paper reports improved methods for culture of trout liver-derived cells that make in vitro investigations
of fish metabolism, carcinogenesis and chemical toxicity more feasible than previously applied techniques. Recent interest
in fish as models for study and indicators of effects of envionmental and food-related toxins make this work timely, poarticularly
since many of the compounds of interest are primarily metabolized by hepatocytes or act on liver as a major target. David
W. Barnes 相似文献
54.
Microfibrils, elastic anchoring components of the extracellular matrix, are associated with fibronectin in the zonule of Zinn and aorta 总被引:3,自引:0,他引:3
Microfibrils are striated tubules that play a role in the formation of elastin fibers by providing a scaffold upon which newly synthesized elastin is deposited. Ultrastructural and staining studies also demonstrate microfibrils that terminate where elastin is sparse or absent in basal laminae, plasma membranes, and the collagenous matrix. The most striking accumulation of microfibrils is found in the zonule of Zinn, the transparent and elastic suspensory ligament of the lens, which contains no elastin. Application of immunocytochemical staining with a peroxidase-antiperoxidase (PAP) procedure demonstrates that fibronectin is associated with the microfibrils of the zonule and aorta. Aggregates of microfibrils are identical to oxytalan ('acid enduring') fibers that have been described in peridontal membranes and other sites subject to mechanical stress and they can be found in sites as disparate as the rabbit zonule, rat hepatic stroma and human cardiac papillary muscle, indicating that microfibrils are a widely distributed connective tissue element with a function that extends beyond elastogenesis; their association with fibronectin and localization suggests that they serve as an elastic anchoring component of the extracellular matrix. 相似文献
55.
Degradation of extracellular matrix by the trophoblastic cells of first-trimester human placentas 总被引:6,自引:0,他引:6
S J Fisher M S Leitch M S Kantor C B Basbaum R H Kramer 《Journal of cellular biochemistry》1985,27(1):31-41
First-trimester human placental villi were cultured on 3H-leucine-labeled extracellular matrices isolated from the PF HR9 and PYS-2 cell lines. Both cell lines produced an extracellular matrix that contained basement membrane-specific macromolecules, including type IV collagen, laminin and proteoglycan. Both matrices promoted outgrowth of cells from the villi which, according to morphological criteria, were identified as cytotrophoblastic cells. As the cells migrated from the attachment site, they caused a marked focal dissolution of the matrix which was accompanied by a concomitant release of 3H-labeled material into the media. Approximately half of this material chromatographed near the inclusion volume of Sephadex G-50, indicating that the labeled matrix components had been degraded. This phenomenon was dependent on the age of the placenta. Second-trimester placental villi also adhered to the matrix, but no areas of dissolution were formed and no significant amounts of radioactivity were released into the medium. These results suggest that culture of first-trimester human placental villi on extracellular matrices may be useful for the study of some of the early embryonic events leading to human implantation, during which the trophoblastic cells erode the uterine epithelium. 相似文献
56.
R Cancedda O Capasso P Castagnola F Descalzi-Cancedda N Quarto 《Journal of cellular biochemistry》1985,28(1):7-14
In cultured chick embryo chondrocytes, type X collagen is preferentially deposited in the extracellular matrix, the ratio between type II and type X collagen being about 5 times higher in the culture medium than in the cell layer. When the newly synthesized collagens deposited in slices from the epiphyseal cartilage of 17-day-old embryo tibiae were isolated, type X collagen was always the major species. In agreement with this result the mRNA for type X collagen was the predominant mRNA species purified from the same tissue. When the total collagen (unlabeled) deposited in the epiphyseal cartilage was analyzed, it was observed that type X collagen represented only 1/15 of the type II collagen recovered in the same preparation. The possible explanations for these differences are discussed. 相似文献
57.
Michel Delarue Thierry Darribere Christian Aimar Jean-Claude Boucaut 《Development genes and evolution》1985,194(5):275-280
Summary Most hybrids betweenBufo bufo andB. calamita obtained by nuclear transplantation become arrested at the early gastrula stage. In both parental controls and the hybrid embryos, the presence and distribution of extracellular matrix was analysed with fluorescent wheat germ agglutinin and by immunolabelling with antibodies directed against fibronectin. InB. bufo andB. calamita gastrulae and in the few hybrids that complete gastrulation, the inner surface of the blastocoel roof is covered by a fibronectin-rich fibrillar matrix. In nucleocytoplasmic hybrids whose development was arrested at the gastrula stage, the fibronectin-containing extracellular matrix was either totally absent or poorly developed and disorganized. 相似文献
58.
Abstract: Extracellular protein fractions were obtained (1) by mild, isotonic irrigation of freshly perfused brain tissue; (2) by collection of proteins released into super-fusing medium by physiologically viable slices of rat hippocampus; and (3) by sampling the CSF of anesthetized rats. Analysis of the S-100 protein content of these fractions gave values of 2.8, 4.2, and 1.8 μg S-100/mg protein, respectively. These values were three- to sixfold higher than the S-100 content of the soluble cytoplasmic protein fractions from the same tissue. This several-fold higher S-100 content of the extracellular protein fractions relative to the intracellular cytoplasmic protein fractions indicates that S-100 is selectively released into the extracellular spaces of the brain. We suggest that the biological function of this CNS protein may involve intercellular transfer. 相似文献
59.
Characterization, Localization, and Biosynthesis of an Interstitial Retinol-Binding Glycoprotein in the Human Eye 总被引:7,自引:0,他引:7
S.-L. Fong G. I. Liou R. A. Landers R. A. Alvarez F. Gonzalez-Fernandez P. A. Glazebrook D. M. K. Lam C. D. B. Bridges 《Journal of neurochemistry》1984,42(6):1667-1676
Abstract: Human eyes contain an Mr 135K retinol-binding protein that is analogous to interstitial retinol-binding protein (IRBP) in the subretinal space of bovine eyes. It is a glycoprotein, because it binds 125 I-concanavalin A, 125 I-wheat germ agglutinin and 125 I- Lens culinaris hemagglutinin. It does not bind Ricinus communis agglutinin I. After desialation, it binds Ricinus communis agglutinin I, but loses its capacity to bind wheat germ agglutinin. These observations, coupled with the known specificities of these lectins, suggest that at least one of the oligosaccharide chains is a sialated, biantennary complex type containing fucose. Both by direct analysis of dissected ocular tissues and by immunocytochemistry it was shown that human interstitial retinol binding protein is an extracellular protein that is confined predominantly to the subretinal space. Monkey retinas incubated in vitro in medium containing [3 H]leucine were shown to synthesize and secrete this protein into the medium, a conclusion that was confirmed by immunoprecipitation with an immunoglobulin fraction prepared from rabbit antibovine IRBP serum. Virtually no other labeled proteins were detectable in the medium. It is concluded that interstitial retinol-binding protein meets many of the requirements for a putative transport protein implicated in the transfer of retinol between the pigment epithelium and retina during the visual cycle, and that the neural retina may play an important role in regulating its amount in the subretinal space. 相似文献
60.
A comparison of the effects of different substrata on chondrocyte morphology and the synthesis of collagen types IX and X 总被引:2,自引:0,他引:2
G. P. Bates S. L. Schor M. E. Grant 《In vitro cellular & developmental biology. Plant》1987,23(5):374-380
Summary Embryonic chick sternal chondrocytes were cultured either within three dimensional gels of type I collagen, type II collagen
or agar, or as monolayers on plastic dishes coated with air-dried films of these matrix macromolecules. It was observed that
cell shape and cell growth varied markedly between the different culture conditions. Flattened monolayers of cells on plastic
or films of type I or type II collagen, proliferated more rapidly and reached a higher final cell density per culture than
the more rounded cells found in the cultures on agar films or within three-dimensional gels. Biosynthetic studies demonstrated
that in addition to the synthesis of type II collagen, all the cultures were producing collagen types IX and X. Chondrocytes
cultured on plastic or films of the different matrix macromolecules all showed a similar expression of types IX and X collagen,
independent of whether they displayed a flattened or round cell morphology. In contrast, marked variations in the proportions
of the minor collagens, particularly type X collagen, were observed when the cells were cultured within three-dimensional
gels. The data suggest that direct interaction of the cell surface with matrix constituents displaying a particular spatial
array could be an important aspect in the control of type IX and X collagen expression by chondrocytes.
The financial support of the Arthritis & Rheumatism Council and the Medical Research Council is gratefully acknowledged. 相似文献