bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mechanical compression modulates matrix biosynthesis in chondrocyte/agarose culture
Authors: Buschmann Michael D. , Gluzband Yehezkiel A. , Grodzinsky Alan J. , Hunziker Ernst B. 
Published: April 2021

ABSTRACT
This study focuses on the effect of static and dynamic mechanical compression on the biosynthetic activity of chondrocytes cultured within agarose gel. Chondrocyte/agarose disks (3 mm diameter) were placed between impermeable platens and subjected to uniaxial unconfined compression at various times in culture (2-43 days). [35S]sulfate and [3H]proline radiolabel incorporation were used as measures of proteoglycan and protein synthesis, respectively. Graded levels of static compression (up to 50%) produced little or no change in biosynthesis at very early times, but resulted in significant decreases in synthesis with increasing compression amplitude at later times in culture; the latter observation was qualitatively similar to that seen in intact cartilage explants. Dynamic compression of ∼3% dynamic strain amplitude (≃30 μm displacement amplitude) at 0.01-1.0 Hz, superimposed on a static offset compression, stimulated radiolabel incorporation by an amount that increased with time in culture prior to loading as more matrix was deposited around and near the cells. This stimulation was also similar to that observed in cartilage explants. The presence of greater matrix content at later times in culture also created differences in biosynthetic response at the center versus near the periphery of the 3 mm chondrocyte/agarose disks.The fact that chondrocyte response to static compression was significantly affected by the presence or absence of matrix, as were the physical properties of the disks, suggested that cell-matrix interactions (e.g. mechanical and/or receptor mediated) and extracellular physicochemical effects (increased [Na+], reduced pH) may be more important than matrix-independent cell deformation and transport limitations in determining the biosynthetic response to static compression. For dynamic compression, fluid flow, streaming potentials, and cell-matrix interactions appeared to be more significant as stimuli than the small increase in fluid pressure, altered molecular transport, and matrix-independent cell deformation. The qualitative similarity in the biosynthetic response to mechanical compression of chondrocytes cultured in agarose gel and chondrocytes in intact cartilage further indicates that gel culture preserves certain physiological features of chondrocyte behavior and can be used to investigate chondrocyte response to physical and chemical stimuli in a controlled manner.
MHC class I molecules are an essential cell surface component involved in Theileria parva sporozoite binding to bovine lymphocytes
Authors: Shaw Michael K. , Tilney Lewis G. , Musoke Antony J. , Teale Alan J. 
Published: April 2021

ABSTRACT
The major histocompatibility complex (MHC) class I molecules are ubiquitous cell surface molecules involved in the cell-mediated immune response. We show here, using a number of different, independent approaches, that these proteins are an essential component of the host cell surface receptor involved in Theileria parva sporozoite invasion. Monoclonal antibodies (mAbs) reactive with common determinants on MHC class I molecules and with β-2 microglobulin inhibited sporozoite entry by specifically preventing the initial binding event. However, in experiments using lymphocytes from heterozygous cattle in which at least four MHC class I gene products are expressed, mAbs which reacted with only one of these products did not inhibit entry. Using a series of bovine deletion mutant cell lines from which one or both MHC class I haplotypes had been lost, sporozoite binding and entry clearly correlated with the level of class I surface expression. While the level of sporozoite entry into cells in which one of the MHC class I haplotypes was lost was only slightly lower than into the parent cells, in a double deletion cell line having less than 5% of the class I expression of the parent cells the level of infection was only 4.3% of that into the parent cells. Furthermore, sporozoite entry into cells from a spontaneously arising mutant cell line exhibiting low levels of class I expression was correspondingly low. Treatment of lymphocytes with IL-2 produced a significant increase in host cell susceptibility and sporozoite entry and this increase correlated with either an increase in the number of target molecules per host cell, or in the binding of bovine MHC class I molecules to the mAbs. In particular, a significant increase in the level of reactivity with mAb W6/32 was observed. Lastly, we show that parasite entry can be competitively inhibited with an isolated sporozoite surface protein, p67. However, p67 binds weakly to lymphocyte surface molecules and initial attempts to use p67 to isolate the relevant host cell molecule(s) have not been successful.
Nuclear distribution of Drosophila DNA topoisomerase II is sensitive to both RNase and DNase
Authors: Meller Victoria H. , Fisher Paul A. 
Published: April 2021

ABSTRACT
The nuclear distribution of Drosophila DNA topoisomerase II was determined by immunoblot analysis after nuclease digestion and cell fractionation. About 60% of DNA topoisomerase II could be removed from nuclei by RNase A, about 70% by DNase I, and about 90% by incubation with both enzymes together or with micrococcal nuclease. Nuclease treatment of nuclei did not affect the distribution of lamins Dm1 and Dm2 or other nuclear proteins similarly. Nuclease-mediated solubilization of DNA topoisomerase II from Drosophila nuclei was also dependent on NaCl concentration. Solubilization was not efficient below 100 mM NaCl. Sucrose velocity gradient ultracentrifugation demonstrated that DNA topoisomerase II solubilized from nuclei by either RNase A or DNase I migrated at about 9 S, as expected for the homodimer. Results of chemical crosslinking supported this observation. We conclude that DNA topoisomerase II has both RNA- and DNA-dependent anchorages in Drosophila embryo nuclei.
The role of the microtubular system in the cell response to HGF/SF
Authors: Dugina V. B. , Alexandrova A. Y. , Lane K. , Bulanova E. , Vasiliev J. M. 
Published: April 2021

ABSTRACT
The effects of the microtubular drugs colcemid and taxol on the morphological changes induced by hepatocyte growth factor/scatter factor (HGF/SF) in MDCK cells were studied. Dynamic changes in the area and shape of individual cells were assessed by morphometric methods whereas alterations of the cytoskeleton were assessed by immunomorphological methods. The results suggest that there are two components in the response to HGF/SF: (a) activation of the extension of lamellae leading to cell spreading; and (b) reorganization of microtubules leading to polarization of cell shape. The latter response is highly sensitive to microtubular drugs, especially taxol. HGF/SF induced spreading in taxol-treated MDCK cells but these cells retained a non-polarized discoid shape and a pattern of actin microfilament bundles characteristic of the untreated cells. Colcemid and taxol did not prevent HGF/SF-induced migration of cells in Boyden chambers but completely inhibited the outgrowth of multicellular strands and tubules from cell aggregates in collagen gels. These results show that enhanced lamella formation in response to HGF/SF without polarization of cell shape is sufficient to induce cell motility. In contrast, microtubule-dependent polarization is essential for complex morpho-genetic responses such as tubulogenesis in collagen gels.
Modulation of microtubule dynamic instability in vivo by brain microtubule associated proteins
Authors: Dhamodharan Rama , Wadsworth Patricia 
Published: April 2021

ABSTRACT
Heat-stable brain microtubule associated proteins (MAPs) and purified microtubule associated protein 2 (MAP-2) were microinjected into cultured BSC-1 cells which had been previously injected with rhodamine-labeled tubulin. The dynamic instability behavior of individual micro-tubules was then examined using low-light-level fluorescence microscopy and quantitative microtubule tracking methods. Both MAP preparations suppressed microtubule dynamics in vivo, by reducing the average rate and extent of both growing and shortening events. The average duration of growing events was not affected. When measured as events/unit time, heat-stable MAPs and MAP-2 did not significantly alter the frequency of rescue; the frequency of catastrophe was decreased approximately two-fold by heat-stable MAPs and MAP-2. When transition frequencies were calculated as events/unit distance, both MAP preparations increased the frequency of rescue, without altering the frequency of catastrophe. The percentage of total time spent in the phases of growth, shrink and pause was determined. Both MAP-2 and heat-stable MAPs decreased the percentage of time spent shortening, increased the percentage of time spent paused, and had no effect on percentage of time spent growing. Heat-stable MAPs increased the average pause duration, decreased the average number of events per minute per microtubule and increased the probability that a paused microtubule would switch to growing rather than shortening. The results demonstrate that addition of MAPs to living cells reduces the dynamic behavior of individual microtubules primarily by suppressing the magnitude of dynamic events and increasing the time spent in pause, where no change in the microtubule length can be detected. The results further suggest that the expression of MAPs directly contributes to cell type-specific microtubule dynamic behavior.
Regulation of growth and dissemination of a human lymphoma by CD44 splice variants
Authors: Bartolazzi Armando , Jackson David , Bennett Kelly , Aruffo Alejandro , Dickinson Richard , Shields John , Whittle Nigel , Stamenkovic Ivan 
Published: April 2021

ABSTRACT
CD44 is a polymorphic cell surface glycoprotein, currently proposed to be the principal cell surface receptor for hyaluronan. However, different isoforms of CD44, expressed in human lymphoid tumor cells, appear to have distinct effects on the ability of the cells to attach to hyaluronan-coated surfaces and on their capacity to form tumors in vivo. In the present study, we address the mechanisms that may regulate CD44 isoform-dependent adhesion to hyaluronan. We use a human Burkitt lymphoma, stably transfected with six different alternatively spliced human CD44 isoforms, to determine their potential hyaluronan binding and tumor growth promoting roles. We show that transfectants expressing CD44 splice variants that contain variable exons 6-10, 7-10 and 8-10 adhere to hyaluronan-coated surfaces weakly and that corresponding tumor formation in vivo is delayed with respect to CD44-negative parental cell-derived tumors. Abundant shedding of these three isoforms may play a significant role in determining the rate of tumor development. Transfectants expressing variable exon 3, on the other hand, fail to display CD44-mediated adhesion to hyaluronan, but form bone marrow tumors rapidly following intravenous injection. These observations suggest that different mechanisms regulate CD44-mediated adhesion and tumor growth, and provide evidence that expression of exon v3 may confer novel ligand-binding properties.
Rapid intracellular assembly of tenascin hexabrachions suggests a novel co-translational process
Authors: Redick Sambra D. , Schwarzbauer Jean E. 
Published: April 2021

ABSTRACT
Tenascin, an extracellular matrix protein that modulates cell adhesion, exists as a unique six-armed structure called a hexabrachion. The human hexabrachion is composed of six identical 320 kDa subunits and the structure is stabilized by inter-subunit disulfide bonds between amino-terminal segments. We have examined the biosynthesis of tenascin and its assembly into hexabrachions using pulsechase labeling of U-138 MG human glioma cells. Newly synthesized tenascin hexamers are secreted within 60 minutes of translation initiation. Intracellularly, as early as full length tenascin can be detected in pulse-labeled cell lysates, it is already in hexameric form. No precursors, such as monomers, dimers, or trimers, were identified that could be chased into hexamers. This lack of assembly intermediates suggests that nascent tenascin polypeptides associate prior to completion of translation. In contrast, fibronectin monomers in the same lysates are gradually formed into disulfide-bonded dimers. Although hexamer assembly is rapid, the rate-limiting step in secretion appears to be transport to the medial Golgi as endoglycosidase H-resistance was not detected until after a 30 minute chase. These results provide evidence for a novel co-translational mechanism of tenascin assembly which would be facilitated by its length and by the amino-terminal location of the assembly domain.
Morphological polarization of human polymorphonuclear leucocytes in response to three different chemoattractants: an effector response independent of calcium rise and tyrosine kinases
Authors: Lepidi H. , Zaffran Y. , Ansaldi J. L. , Mege J. L. , Capo C. 
Published: April 2021

ABSTRACT
Chemoattractants such as interleukin-8, C5a and N-formylmethionyl-leucyl-phenylalanine induce a cytosolic calcium rise involved in triggering the secretory functions of human polymorphonuclear leucocytes. We studied the possible role of calcium rise in membrane ruffling, actin polymerization, filamentous actin distribution, and morphological polarization, which are all events contributing to chemotaxis. Membrane ruffling was assessed by right-angle light-scatter changes, the cellular content of polymerized actin by fluorescence of bodipy phallacidin, the intracellular distribution of filamentous actin by fluorescence microscopy and image digitization, and morphological polarization by scanning electron microscopy. Pretreatment of polymorphonuclear leucocytes with 50 μM BAPTA/AM, an intracellular calcium chelator, lowered the basal level in cell calcium and inhibited the transient calcium rise stimulated by 2 nM interleukin-8, 2 nM C5a, and 10 nM N-formylmethionyl-leucyl-phenylalanine. However, BAPTA pretreatment of polymorphonuclear leucocytes did not modify membrane ruffling, actin polymerization, filamentous actin distribution, and morphological polarization stimulated by chemoattractants. Downstream effectors may be protein tyrosine kinases. However, the tyrosine kinase inhibitor tyrphostin did not affect the cytoskeletal characteristics elicited by chemoattractants. Taken together, our results suggest that the transductional pathway leading to cytoskeleton organization and morphological polarization of polymorphonuclear leucocytes is different from that leading to secretion.
On the possible role of cytoskeletal filamentous networks in intracellular signaling: An approach based on percolation
Authors: Forgacs G. 
Published: April 2021

ABSTRACT
Signaling in multicellular organisms is a challenging problem, for it requires the understanding of processes at drastically varying scales, at molecular, cellular and multicellular levels. As an embryo develops it must coordinate cell division, cellular adhesion and migration, and ultimately form a properly functioning organism. Extracellular morphogens and direct cell-to-cell interactions are used to establish and redefine the pattern of the developmental process. The associated extracellular signals must be sensed by the cells and transmitted eventually to the nucleus to coordinate with internal signals to induce the appropriate changes in gene activities. The cell-to-cell or environment-to-cell communication requires intricate signal transduction mechanisms whose importance is clearly not restricted to development.
Regulation of mesenchymal extracellular matrix protein synthesis by transforming growth factor-β and glucocorticoids in tumor stroma
Authors: Talts Jan Fredrik , Weller Andreas , Timpl Rupert , Ekblom Marja , Ekblom* Peter 
Published: April 2021

ABSTRACT
We have here studied the composition and regulation of stromal extracellular matrix components in an experimental tumor model. Nude mice were inoculated with WCCS- 1 cells, a human Wilms’ tumor cell line. In the formed tumors the stroma was found to contain mesenchymal extracellular matrix proteins such as tenascin-C, fibulins-1 and 2 and fibronectin, but no nidogen. Nidogen was confined to basement membranes of tumor blood vessels. Since glucocorticoids have been shown to downregulate tenascin-C expression in vitro, we tested whether dexamethasone can influence biosynthesis of extracellular matrix components during tumor formation in vivo. A downregulation of tenascin-C mRNA and an upregulation of fibronectin mRNA expression by dexamethasone was noted. Transforming growth factor-β1 mRNA levels were unaffected by the dexamethasone treatment. Glucocorticoids can thus downregulate tenascin-C synthesis although local stimulatory growth factors are present. The competition between a negative and a positive extrinsic factor on synthesis of stromal extracellular matrix components was studied in a fibroblast/preadipocyte cell line. Transforming growth factor-β1 stimulated tenascin-C synthesis but did not affect fibronectin or fibulin-2 synthesis. Dexamethasone at high concentrations could completely suppress the effect of transforming growth factor-β1 on tenascin-C mRNA expression. Transforming growth factor-β1 could in turn overcome the downregulation of tenascin-C mRNA expression caused by a lower concentration of dexamethasone. We therefore suggest that the limited expression of tenascin-C in part is due to a continuous suppression by physiological levels of glucocorticoids, which can be overcome by local stimulatory growth factors when present in sufficient amounts.
Emergence of salivary gland cell lineage diversity suggests a role for androgen-independent epidermal growth factor receptor signaling
Authors: Durban Elisa M. , Nagpala Pablito G. , Barreto Patricia D. , Durban Egon 
Published: April 2021

ABSTRACT
Diversity of cell lineages within glandular organs is generated postnatally by differentiation of committed progenitor cells. Fundamental regulatory aspects of this process are not understood. The mouse submandibular salivary gland (SSG) served as model to assess the role of epidermal growth factor (EGF) receptor signaling during emergence of cell lineage diversity. Temporal fluctuations in EGF receptor mRNA levels coincident with crucial differentiative cell lineage transitions were revealed by RNase protection analyses. Between days 2 and 5, when proacinar cells are maturing and striated duct cells emerge, EGF receptor mRNA levels were highest and all differentiating cells exhibited EGF receptor immunoreactivity. EGF receptor mRNA levels then declined sharply and immunoreactivity became confined to ductal cells. At day 11 in male mice, and days 11 and 16 in females, a second increase in EGF receptor mRNA was detected coincident with emergence of granular convoluted tubule (GCT) cells. With completion of androgen-dependent GCT cell differentiation at the onset of puberty, EGF receptor mRNA levels and intensity of immunoreactivity decreased. Androgen effects on EGF receptor mRNA or immunoreactivity could not be detected. These temporally distinct patterns of EGF receptor expression suggest that this signaling pathway is a mechanism of potential importance in emergence of cell lineage diversity in a glandular organ.
Focal adhesion formation by f9 embryonal carcinoma cells after vinculin gene disruption
Authors: Volberg Tova , Geiger Benjamin , Kam Zvi , Pankov Roumen , Simcha Inbal , Sabanay Helena , Coll Jean-Luc , Adamson Eileen , Ben-Ze’ev Avri 
Published: April 2021

ABSTRACT
The assembly of focal adhesions was investigated in F9 embryonal carcinoma cells in which the expression of vinculin was eliminated by a targeted disruption of the vinculin gene. Vinculin-deficient F9 cells were capable of adhering to fibronectin-coated surfaces, though they displayed a reduced spreading compared to the parental cells. Transmission electron microscopy as well as inter-ference reflection microscopy of live cells showed that vinculin-null F9 cells formed focal adhesions that were indistinguishable from those of the control cells. Fluores-cent labeling for actin, talin, α-actinin, paxillin and phos-photyrosinated components indicated that the organization of all these focal contact-associated components was essen-tially identical in the vinculin-containing and vinculin-null cells. However, quantitative, digitized microscopy indicated that the intensity of fluorescence labeling in focal adhesions for α-actinin, talin and paxillin was significantly higher in cells lacking vinculin. The results suggest that there are multiple molecular mechanisms for the formation of focal adhesions in the absence of vinculin.
Differential expression of bcl-2 in intestinal epithelia correlation with attenuation of apoptosis in colonic crypts and the incidence of colonic neoplasia
Authors: Merritt Anita J. , Potten Christopher S. , Watson Alastair J. M. , Loh Dennis Y. , Nakayama Kei-ichi , Nakayama Keiko , Hickman John A. 
Published: April 2021

ABSTRACT
The cell-positional incidence of both spontaneous and damage-induced apoptosis of epithelial cells was assessed in longitudinal sections of the crypts of small intestine and colon of BDF1 mice. This was compared, using immuno-histochemistry, with the pattern of expression of bcl-2, a suppressor of apoptosis. In the small intestine, apoptosis was maximal around cell position 4 from the base of the crypt; this closely corresponds to the position considered to contain the stem cells. In the colon, however, apoptosis was not confined to the area considered to harbour the stem cells (position 1 and 2). Instead, apoptosis was attenuated and distributed along the length of the crypt. Some cells at the base of murine colonic crypts expressed bcl-2 protein, whereas bcl-2 was absent in the crypts of the small intestine. Most pertinently, bcl-2 was absent from small intestinal crypt cells at positions 4-5 (the stem cell region). The importance of the expression of bcl-2 to the attenuation of apoptosis in stem cells was confirmed by analysis of the levels of both spontaneous and induced apoptosis in homozygously bcl-2 null C57BL/6 mice: in colonic crypts the level of spontaneous apoptosis rose significantly, and selectively at the base of the crypt, in comparison with crypts from wild-type animals. In contrast, there was no rise in spontaneous apoptosis in the small intestinal crypts from the bcl-2 null animals. Analysis of sections of human colon and small intestine also showed that expression of bcl-2 was confined to the base of the colonic crypt. The attenuation of apoptosis by bcl-2 in the region of the stem cells of the colonic crypts may dispose these to neoplastic transformation. Indeed, analysis of human carcinomas revealed expression of bcl-2, which in some samples was reciprocal with the expression of p53.