bluuebunny/crossref_metadata_2025_split huggingface dataset:
Production of matrix metalloproteinase 3 (stromelysin) by cultured ovine endometrial cells
Authors: Salamonsen Lois A. , Nagase Hideaki , Woolley David E. 
Published: April 2021

ABSTRACT
Ovine endometrial cells (stromal plus epithelial) in primary culture release matrix metalloproteinase 3 (stromelysin, MMP-3), the enzyme that degrades various extracellular components including the basement membrane components collagen IV and laminin. The enzyme was detected in tissue culture medium by specific enzyme assay. A low level of MMP-3 activity was released from the cells (0.12±0.02 unit per 106 cells per 48 h; 1 unit degrades l/&lt;g of reduced, carboxymethylated transferrin min−1 at 37 °C), but following stimulation by phorbol myristate acetate (PMA) a mean 4.4-fold increase in MMP-3 production was observed (to 0.53±0.13 unit per 108 cells per 48 h). The enzyme was released primarily in the form of its proenzyme, which could be activated in vitro by (4-aminophenyl)mercuric acetate (APMA).
Stimulation of enzyme release from cells by PMA was not seen before 24 h and required PMA at a dose of 10 nM or higher. This finding was substantiated by Western blot analysis. Immunocytochemistry of cells that had been treated with PMA for 48 h, and with the ionophore, monensin, for 4 h, confirmed the potential of the endometrial cells to produce stromelysin. The release of MMP-3 from ovine endometrial cells suggests that this enzyme may have a role in assisting the passage of syncytial processes through the basement membrane during the process of implantation.
Basket scales of the green alga, Mesostigma viride: Chemistry and ultrastructure
Authors: Domozych David S. , Wells Brian , Shaw Peter J. 
Published: April 2021

ABSTRACT
The unicellular green algal flagellate, Mesostigma viride, possesses an extracellular matrix consisting of three layers of highly distinct scales. This study focused upon the elaborate basket scales. The basket scale is approximately 450–500 nm in height and consists of a solid base, two distinct lattices, two upper rims and various struts and roots. Pure preparations of isolated basket scales were obtained and used for subsequent chemical and immunological analyses. X-ray microanalyses revealed the scale as being mineralized with both calcium and phosphorus. Fourier Transform Infrared Spectroscopic analyses revealed the presence of phosphate, suggesting in turn that the basket scale is complexed with calcium phosphate. SDS-gel electrophoresis of partially digested scales revealed the presence of two protein components, a high Mr, non-migrating form and a form with a Mr of approximately 48 000. Further biochemical analyses of scales showed the presence of two major sugars: glucose and the unusual keto sugar acid, 3-deoxy-lyxo-2-heptulosaric acid, or DHA. Antibodies to scales were raised in rats and used in immunolabeling studies. It is estimated from immunofluorescence studies that a cell of 8/cm contains about 800 basket scales. The scales are in a close-packed arrangement resulting in quasi-crystalline arrays upon the cell surface.
Bone-marrow-derived chondrogenesis in vitro
Authors: Berry L. , Grant M. E. , McClure J. , Rooney P. 
Published: April 2021

ABSTRACT
Bone marrow stromal cells from embryonic, neo-natal and adult chickens were grown in vitro over a 21-day period. Marrow stromal cells from embryonic and neonatal chicks produced clonally derived chondrocytic colonies. The cells within the colonies were surrounded by a refractile, Alcian-blue-positive matrix and their cartilagenous nature was shown biochemically and immunocytochemically by the synthesis of collagen types n and X. The ability of chick bone marrow cells to form chondrocytic colonies decreased during development and was lost by adulthood. In addition to chondrocytic colonies, fat cells and fibroblasts were also observed in the cultures.
Our data demonstrate that chick bone marrow stroma contains cells that are capable of differentiating along different pathways Within the same culture, providing further evidence for the presence in bone marrow of a stromal stem cell.
Vimentin expression as a late event in the in vitro differentiation of human promonocytic cells
Authors: Rius Carlos , Aller Patricio 
Published: April 2021

ABSTRACT
The administration of either 12-O-tetradeeanoyl phor-bol-13-acetate (TPA, 3xlO−8M), sodium butyrate (SB, 10−3M), A6,2’-O-dibutyryladenosine-3’:5’-cyclic monophosphate (dbcAMP, 10−3M), cytosine arabinoside (ara-C, 10−7M), amsacrine (mAMSA, 10−7M) or retinoic acid (RA, 10−6M) inhibits the growth activity of human promonocytic U-937 cells, by arresting them at G1 or at the G1/S border (SB, RA, ara-C), at G2 (mAMSA) or at G1 and G2 (dbcAMP). All these agents trigger cell differentiation, as proved by the increased expression of the maturation-associated CDllb and CD 11c surface antigens, and induce the expression of the vimentin gene at both the protein and the mRNA levels. TPA, SB and dbcAMP behave as “early” inducers, in the sense that vimentin mRNA levels are rapidly increased (hour 6) upon drug administration. In contrast, mAMSA and RA behave as “late” inducers, since they do not increase vimentin mRNA levels until 48 to 72 hours, following the stimulation of surface antigen expression. The action of RA is characterized by an initial inhibition period, in which the basal level of vimentin mRNA is abolished (hour 24). Nevertheless, this RNA is later re-induced, to reach at 72 hours higher levels than in untreated cells. Moreover, RA is capable of delaying the early induction of vimentin expression caused by TPA and SB, without affecting the normal expression of differentiation markers. Taken together, these results strongly suggest that vimentin expression is not required at the initial stages of promonocytic cell differentiation, although it could play a role at an advanced stage.
Organisation of glycoproteins into plasma membrane domains on Fucus serratas eggs
Authors: Stafford C. J. , Green J. R. , Callow J. A. 
Published: April 2021

ABSTRACT
Fertilisation in the brown alga Fucus involves the interaction of motile, biflagellate sperm with spherical eggs. The eggs differ from those of animals in not having the equivalent of a vitelline layer, jelly coat or zona pellucida outside the plasma membrane, and in addition they are not surrounded by a cell wall. Previous studies on Fucus eggs have shown that the lectin Concanavalin A (ConA) binds in patches on the egg surface, suggesting that there is a non-uniform distribution of plasma membrane glycoproteins. In this paper we have further investigated the occurrence of domains on the Fucus egg plasma membrane using monoclonal antibodies (mAbs) and the lectins ConA and Fucose Binding Protein (FBP). Confocal laser scanning microscopy (CLSM) has been used to observe the binding of probes to the Fucus egg cell surface. Four mAbs (FS2, FS4, FS5 and FS9) raised to Fucus serratas sperm have previously been shown to cross-react with crude egg membrane vesicles. Three of these mAbs (FS2, FS4 and FS5) have now been shown to bind to the egg cell surface and they recognise glycoproteins which are organised into domains. mAb FS4 labels large areas of the egg surface, whereas mAbs FS2 and FS5 bind to smaller patches. The lectins ConA and FBP also bind to smaller, discrete domains. Western blotting results and competition binding assays have shown that mAbs FS2 and FS5 compete for binding to the same set of glycoproteins, and FS5 is used in subsequent experiments; FS4 also binds to several glycoproteins but produces a different pattern of labelling on Western blots compared to FS5, though there may be some common components. ConA labels a subset of the glycoproteins recognised by mAb FS5, and FBP recognises one major glycoprotein which is also recognised by ConA and FS5. Double labelling experiments using the CLSM, with FITC- and Au-labelled probes, have shown that the regions on the egg surface labelled by FS4 and FS5 are mainly exclusive, with small areas of overlap. There are also areas which are not labelled by either of these antibodies. The domains recognised by mAb FS5 contain smaller areas which are labelled by ConA. Overall the results show that the Fucus egg surface is heterogeneous with different sets of glycoproteins being organised into domains. With the probes used it is possible to distinguish between FS4+ FS5−, FS4− FS5+, FS4+ FS5+ and FS4− FS5” regions. Within the FS5+ domains smaller sets of glycoproteins are recognised by ConA and within these latter regions there are glycoproteins recognised by FBP.
Histamine decreases the permeability of an endothelial cell monolayer by stimulating cyclic AMP production through the H2-receptor
Authors: Takeda Taichi , Yamashita Yuko , Shimazaki Syuji , Mitsui Youji 
Published: April 2021

ABSTRACT
To determine if histamine acts directly on the vascular endothelium, the effect of histamine on the permeability of cultured human endothelial cell monolayers and the role of second messengers were examined. The addition of 10−6 to 10−4 M histamine to the culture medium decreased the endothelial cell monolayer permeability and increased both cyclic AMP and free-calcium levels. The decrease in permeability and the increase in cyclic AMP mediated by histamine were prevented by an H2-blocker (famotidine) while the increase in free-calcium was inhibited by an H1-blocker (diphenhydramine). These results suggest that histamine decreases the permeability of endothelial cell monolayers through the H2-receptor, and cyclic AMP plays a more important role than calcium ion as a second messenger.
A monoclonal antibody recognizing nuclear matrix-associated nuclear bodies
Authors: Stuurman Nico , Graaf Arjan De , Floore Arno , Josso Arthur , Humbel Bruno , Jong Luitzen De , Driel Roel Van 
Published: April 2021

ABSTRACT
We have isolated a monoclonal antibody, 5E10, that labels discrete spots in the interphase nucleus. By immunoblotting mAb SE10 recognized predominantly a 126 kDa polypeptide with an isoelectric point of 5.5. Indirect immunofluorescence showed that mAb 5E10 labeled spots in many cell lines and tissues from rat or human origin, but not in cells from mouse, chicken, African green monkey, or the lower eukaryotes Saccharomyces and Dictyostelium. In the human bladder carcinoma cell line T24 the number of nuclear spots was found to be 21 ± 10 (n=132). In many cells spots were found also in the cytoplasm. In a small fraction of T24 cells the mAb revealed thread-like structures in addition to spots. Throughout mitosis the antigen was found to be clustered in the cytoplasm, not associated with metaphase chromosomes. The spherical structures that contain the antigen were tightly bound to the nuclear matrix. Immunogold labeling with mAb 5E10 showed that the antigen is localized in 0.3 /mi diameter spherical, electron-dense structures, reminiscent of nuclear bodies. Double-labeling experiments showed that these spots do not colocalize with U1 snRNPs and centromeres. The spots did colocalize with nuclear speckles recognized by a primary biliary cirrhosis autoimmune serum, which is thought to recognize nuclear bodies. On the basis of these observations we conclude that mAb 5E10 recognizes discrete nuclear substructures, most likely nuclear bodies.
Tubulin in sea urchin embryonic cilia: post-translational modifications during regeneration
Authors: Stephens R. E. 
Published: April 2021

ABSTRACT
Tubulin is the major protein found in the membrane/periaxonemal matrix fraction of mature sea urchin embryonic cilia but its distribution and possible function during ciliary assembly are unknown. Hypertonic salt may be used to deciliate the embryos, allowing synchronous regrowth of cilia and subsequent deciliation of the regenerating embryos at various times. During the earliest stages of regeneration, the amounts of tubulin in the axoneme and membrane/matrix fractions are nearly equal, but the proportion of tubulin in the axoneme fraction increases coincident with the quasi-linear growth phase while the membrane/matrix tubulin remains constant. Antibodies to tyrosinated and detyro-sinated α-tubulin show that both the membrane/matrix and axonemal tubulin fractions are primarily unmodified (i.e. tyrosinated) at the earliest stages of regeneration but are progressively and equally detyrosinated coincident with regeneration, approaching a final level of 50% C-terminal Glu. A monoclonal antibody to acetylated a-tubulin reveals that both tubulin fractions are equally and maximally acetylated at relatively early stages of regeneration. In contrast, three-times-repoly-merized tubulin from either unfertilized eggs or mid-gastrula embryos is primarily tyrosinated (&gt;97%) and not detectably acetylated. These data suggest that membrane/matrix tubulin is a precursor to axonemal tubulin and that acetylation and detyrosination may be involved in partitioning tubulin among cytoplasmic, ciliary membrane/matrix, and 9+2 compartments.
Tripeptide RGD-dependent adhesion of articular chondrocytes to synovial fibroblasts
Authors: Ramachandrula Anand , Tiku Katherine , Tiku Moti L. 
Published: April 2021

ABSTRACT
Cell-cell interactions play an important role in the development of cartilage. Heterologous and homologous cell-cell interactions are critical for chondrogenic differentiation during development. Cell-cell interactions in the formation of fracture callus and cartilage neoplasia also invoke the process of cartilage differentation. We have investigated cell-cell interactions between articular chondrocytes and synovial fibroblasts and show that there was enhanced binding between these two cell types compared to background binding of the labelled cells to the tissue culture plastic surface. The binding of chondrocytes to fibroblasts was temperature- and calcium-dependent, suggesting ligand-integrin involvement. The peptide, GRGDSP, which competes with the ligand-integrin through the tripeptide RGD (arginineglycine-aspartic acid), almost completely inhibited chondrocyte attachment to synovial fibroblasts. The control peptide, GRGESP, had no Inhibitory effect on binding. Antibodies to fibronectin (Fn) inhibited chondrocyte attachment by about 50%. Monoclonal antibodies to the α and β chains of the fibronectin receptor (FnR) interfered with the attachment of chondrocytes to synovial fibroblasts. A combination of antibodies to Fn and to FnR did not completely abrogate chondrocyte binding, suggesting that other ligand-receptors were involved in the adhesion process. Chondrocytes and fibroblasts were shown to express membrane-associated Fn and FnR, by immunofluorescence. The α and β chains of FnR, migrating at 110 and 140 kDa, respectively, could be immunoprecipitated from [35S]methionine-labelled synovial fibroblasts and chondrocytes. Northern blots showed the presence of mRNA for the α and β chains of fibronectin receptors in fibroblasts and chondrocytes. Changes in cell shape were observed in chondrocytes on attachment to fibroblasts, i.e. the chondrocytes appeared fibroblast-like, suggesting that the chondrocytes had dedifferentiated. These studies suggest that chondrocytes specifically bind to synovial fibroblasts through RGD-dependent receptors. β1 Integrins are involved in this adhesion process and these heterlogous cell interactions appear to have a negative influence on chondrogenic differentiation.
Stable and slow-turning-over microtubules characterize the processes of motile epithelial cells treated with scatter factor
Authors: Prescott A. R. , Dowrick P. G. , Warn R. M. 
Published: April 2021

ABSTRACT
The turnover of microtubules was studied in the processes of PtK2 cells, after treatment with the cytokine scatter factor (SF), using micro-injected biotin-tubulin as a reporter of new microtubule growth. Cells treated with SF became dispersed and fibroblast-like in morphology, showing one or more elongated processes. These processes contained bundles of microtubules, a significant proportion of which did not turn over during incubation times of up to an hour. Short broken pieces of microtubule were frequently found in all parts of the cell, particularly after longer incubation times, suggesting that more-stable microtubules were cut into pieces, which were subsequently degraded. From about half an hour after injection small tangles of stable microtubules were found. Some of these were clearly within the cell bodies. Others were usually larger in size and seemingly located outside the injected cells. These were considered to have formed part of small ‘feet’ presumed to be broken off during the retraction of trailing processes.
The microtubules within the processes were resistant to the effects of both microtubule-depolymerizing drugs and cold under conditions where the processes were maintained. When these microtubules disappeared as the result of longer drug treatment the processes were also lost although, rarely, short processes lacking microtubules were found. It is concluded that the stable microtubules have a major role in process maintenance, although one that is indirect rather than a structural relationship.
Retinoic acid receptor expression in human skin keratinocytes and dermal fibroblasts in vitro
Authors: Redfern Christopher P. F. , Todd Carole 
Published: April 2021

ABSTRACT
Retinoic acid is essential for the normal differentiation of epithelia but its cellular function is obscure. The expression patterns of retinoic acid receptors (RARs) in skin cell types may give an insight into the role of retinoic acid in skin. We have compared the patterns of RAR expression in human keratinocytes and dermal fibroblasts in vitro, and studied the effects of retinoic acid on RAR expression. RAR-α and RAR-γ were expressed in keratinocytes and fibroblasts: RAR-γ was expressed at similar levels in both cell types but RAR-α was more abundant in fibroblasts. There were no differences in expression of either RAR-α or RAR-γ between stratifying (high-calcium medium) and proliferating (low-calcium medium) keratinocytes and expression of these RARs was unaffected by retinoic acid. RAR-β was undetectable in keratinocytes. In the majority of fibroblast cell lines, RAR-β transcripts were either undetectable or expressed at a low level. Retinoic acid at low concentrations (10−10 to 10−9 M) rapidly induced the expression of RAR-γ. Cyclic adenosine monophosphate (cAMP) analogues inhibit RAR-β induction in teratocarcinoma cells. However, dibutyryl-cAMP did not affect RAR-β induction in fibroblasts. Forskolin, an adenylate cyclase activator, and the phosphodiesterase inhibitor 3-isobutyl-l-methylxanthine (IBMX) decreased constitutive RAR-β mRNA levels but did not block induction of RAR-β by retinoic acid. Since intracellular cAMP levels were only increased detectably in response to forskolin, the reduction in constitutive levels of RAR-β mRNA may be mediated by mechanisms other than via cAMP.
Identification of a large pre-lysosomal compartment in the pathogenic protozoon Trypanosoma cruzi
Authors: Soares Maurilio J. , Souto-Padrón Thais , Souza Wanderley De 
Published: April 2021

ABSTRACT
Epimastigote forms of the pathogenic parasite Trypanosoma cruzi were used to study the endocytic process in a protozoon. These elongated unicellular organisms are highly polarized cells: endocytosis occurs only at the anterior region through the cytostome and the flagellar pocket membrane, areas of the plasma membrane where the cell cytoskeleton, formed by sub-pellicular microtubules, is absent. When the cells were incubated at 4°C or 28°C with gold-labeled transferrin, fixed and processed for routine transmission electron microscopy our observations show that this ligand initially binds to the cytostome and the membrane lining the flagellar pocket and is subsequently ingested through a clathrin-independent receptor-mediated endocytotic process, with formation of uncoated pits and vesicles. Ingested complexes are carried in uncoated vesicles to the reservosomes, large membrane-bound organelles found mostly at the posterior end of the cell. Immunocytochemical data from Lowicryl-embedded cells demonstrated that the reservosomes are acidic compartments (pH 6.0, as shown using DAMP as a pH probe) with no acid phosphatase or typical lysosome-associated membrane proteins (LAMP 1, LAMP 2 and Igp 120), but rich in cysteine proteinase. These data suggest that the reservosome is a pre-lysosomal compartment. Since cysteine proteinase of T. cruzi contains no phosphorylated mannose residues and the cation-independent mannose 6-phosphate receptor could not be immunocytochemically detected in the reservosomes, it is possible that lysosomal enzymes in the epimastigote forms of T. cruzi are targeted to compartments related to the endocytic pathway through a mechanism different from that which occurs in other eukaryotic cells.
Vertebrate p34cdc2 phosphorylation site mutants: effects upon cell cycle progression in the fission yeast Schizosaccharomyces pombe
Authors: Krek Wilhelm , Marks John , Schmitz Nicole , Nigg Erich A. , Simanis Viesturs 
Published: April 2021

ABSTRACT
We have used the fission yeast Schizosaccharomyces pombe to analyse the effects of in vitro mutagenesis of the four known phosphorylation sites in the chicken p34cdc2 protein, Thr14, Tyr15, Thr161 and Ser277, upon cell cycle progression. We have studied both the effect of overexpression of mutant proteins in a cdc2+ background and assayed their ability to rescue null and temperature-sensitive alleles of cdc2. Mutations of Thrl4 and Tyrl5 within the ATP binding domain of p34cdc2 that mimic constitutive phosphorylation cause dominant negative cell cycle arrest when overexpressed. In contrast, some substitutions that simulate permanent dephosphorylation of the corresponding sites advance mitosis. These data confirm the model that p34cdc2 function is negatively regulated by phosphorylation of residues in the ATP binding site. Mutagenesis of the conserved residue Thr161 functionally inactivates p34cdc2, and our data suggest that both phosphorylation and dephosphorylation events at Thr161 are required for progression through the cell cycle. Mutations at the fourth site of phosphorylation, Ser277, lead to coldsensitive cell cycle arrest, in minimal but not rich growth medium, suggesting that this site is involved in monitoring the nutritional status of the cell.
PCNA and total nuclear protein content as markers of cell proliferation in pea tissue
Authors: Citterio Sandra , Sgorbati Sergio , Levi Marisa , Colombo Bruno Maria , Sparvoli Elio 
Published: April 2021

ABSTRACT
The identification of cell proliferation markers has been shown to be a useful tool with which to study basic mechanisms of cell cycle progression. The use of immunofluorescence techniques revealed the presence of the proliferating cell nuclear antigen (PCNA) in pea tissue, where we observed a high PCNA expression in proliferating cells of the root meristem compared to noncycling cells of the differentiated leaf. The presence of PCNA was monitored also during the time-course of seed germination, before, during and after the cell cycle resumption of the embryo cells. PCNA is present in embryo cells not only during and after resumption of the cell cycle but also before, when cells have not yet begun replicating their genome. A bivariate flow cytometric analysis of DNA and nuclear protein content was used to localize precisely the cells of the examined pea tissues in different cell cycle phase subcompartments. A high correlation was found between the degree of cell proliferation and the protein content of G1 nuclei, on the one hand, and the percentage of PCNA positive cells on the other.