bluuebunny/crossref_metadata_2025_split huggingface dataset:
Astrocytes induce neural microvascular endothelial cells to form capillary‐like structures in vitro
Authors: Laterra John , Guerin Christopher , Goldstein Gary W. 
Published: February 2005

AbstractAstrocytes maintain a unique association with the central nervous system microvasculature and are thought to play a role in neural microvessel formation and differentiation. We investigated the influence of astroglial cells on neural microvascular endothelial differentiation in vitro. Using an astroglial‐endothelial coculture system, rat brain astrocytes and C6 cells of astroglial lineage are shown to induce bovine retinal microvascular endothelial (BRE) cells to form capillarylike structures. Light microscopic evidence for endothelial reorganization began within 48 hours and was complete 72‐96 hours following the addition of BRE cells to 1‐day‐old astroglial cultures. The extent of BRE reorganization was quantitated by computer‐assisted analysis and shown to be dependent upon the density of both the BRE and C6 cells within the cocultures. Coculture conditions in which BRE cells were separated from C6 cells by porous membranes failed to generate this endothelial cell change. Likewise, C6‐conditioned media and C6‐endothelial coculture conditioned media did not induce BRE cell reorganization. Extracellular laminin within the C6‐endothelial cocultures, identified by indirect immunofluorescence, was concentrated at the endothelial‐astroglial interface of capillary‐like structures consistent with incipient basement membrane formation. Astroglial cells accumulated adjacent to capillary‐like structures suggesting the presence of bidirectional influences between the reorganized endothelial cells and astroglia. This is the first demonstration of astroglial induction of angiogenesis in vitro and these findings support a functional role for perivascular astrocytes in the vascularization of neural tissue such as retina and brain.
Synthesis of basic fibroblast growth factor upon differentiation of rat mammary epithelial to myoepithelial‐like cells in culture
Authors: Barraclough Roger , Fernig David G. , Rudland Philip S. , Smith John A. 
Published: February 2005

AbstractAcidic fibroblast growth factor (aFGF) mRNA was detected in a rat mammary fibroblastic cell line, but not in rat mammary epithelial cell lines or myoepithelial‐like cell lines. Basic FGF (bFGF) mRNA was detected in both the fibroblasts and the myoepithelial‐like cells, but was absent from the epithelial cells. A series of cell lines representing stages in the differentiation pathway of epithelial cells to a myoepithelial‐like morphology showed an increase in the amount of bFGF mRNA and activity present and the FGF from the myoepithelial‐like rat mammary 29 cells was able to displace [125I]‐bFGF specifically bound to rat mammary fibroblasts. FGF activity was also present in an extract of rat mammary gland. Analysis of cell extracts and conditioned medium indicated that FGF activity was cell‐associated. The cell‐associated bFGF was resistant to degradation by trypsin. Extraction of myoepithelial‐like cells with Triton X‐100 and 2 M NaCl showed that 50–65% of the cell‐associated bFGF was in a detergent‐resistant but 2 M NaCl− labile structure. Thus, the synthesis of bFGF is developmentally regulated in rat mammary cell lines, and at least 50% is present in the extracellular matrix.
Pedersen fetuin contains three adipogenic factors with distinct biochemical characteristics
Authors: Zaitsu Hirokazu , Serrero Ginette 
Published: February 2005

AbstractCrude Pedersen fetuin, derivea from fetal bovine serum, contains adipogenic activity, Biochemical characterization was undertaken by following the differentiation of the 1246 adipogenic cell line. The present paper provides evidence that crude fetuin contains distinct proteins with adipogenic activity. By molecular sieve fractionation using Sephacryl S‐300, the majority of adipogenic activity eluted in two distinct peaks, F1 (molecular weight greater than 669 kDa) and Fii (molecular weight ranging from 445 and 232 kDa). In addition a minor activity was found in a third peak, Fiii (molecular weight around 69 kDa). Partial purification and biochemical characterization indicate that FI and FII are two distinct factors. F1 has a PI higher than 9.4, is destroyed by alkaline treatment, and is stable when treated with acid. Fii has a PI lower than 4.0, is alkali stable, but is destroyed completely by treatment with acid. Moreover, our data show that adipogenic factors are distinct from another protein α2 macroglobulin known to be found in crude Pedersen fetuin. These results suggest that serum contains two large molecular weight proteins bearing adipogenic activity which could play an important role in the control of the adipose differentiation process.
Synergism between gangliosides and basic fibroblastic growth factor in favouring survival, growth, and motility of capillary endothelium
Authors: Alessandri G. , De Cristan G. , Cappa A. P. M. , Gullino P. M. , Morbidelli L. , Ziche M. 
Published: February 2005

AbstractThe experiments reported were motivated by the observation that in vivo gangli‐osides promoted angiogenesis when the dose of the angiogenic factor was too low to be effective (Ziche et al.: Laboratory Investigation 61:629–634, 1989). As an approach to understanding the mechanism of this modulatory effect, we analysed the influence that gangliosides have on survival, growth, and migration of capillary endothelium when and angiogenesis factor like basic fibroblast growth factor (bFGF) was present in the culture medium. Clones of bovine capillary endothelium were cultivated in media unable to sustain survival over a 72 h period. With this experimental approach, cell survival was evaluated after addition of either bFGF or gangliosides or both to the medium. The Boyden chamber procedure was utilized to measure the influence of bFGF or gangliosides on cell mobilization across a micropore filter. Low doses of both molecules, ineffective when added singly to the culture media, improved all three parameters when added in combination. A synergic effect between bFGF and the gangliosides (GM1, GD1b, GT1b) was observed for the improvement of survival or growth and for the acceleration of endothelial cell migration. The removal of sialic acid from the ganglioside molecule prevented any effect on all three parameters. The addition of sialic acid alone to cultures was also totally ineffective. In the aduit organism most angiogenic events occur under conditions of tissue damage. The synergism between gangliosides and bFGF can be interpreted as the initial phase of a process for which endothelial cell survival is the indispensable first step in the formation of a new vascular network.
PDGF‐stimulated fibroblast proliferation is enhanced synergistically by receptor‐recognized α2‐Macroglobulin
Authors: Bonner James C. , Badgett Annette , Osornio‐Vargas Alvaro R. , Hoffman Maureane , Brody Arnold R. 
Published: February 2005

Abstractα‐Macroglobulins derived from plasma or secreted by macrophages are plateletderived growth factor (PDGF) binding proteins that compete with cell‐surface receptors on fibroblasts for PDGF binding. α2‐Macroglobulin (α2M) derived from bovine plasma was tested for its ability to modulate the PDGF‐induced proliferation of primary passage rat lung fibroblasts (RLFs) and a human skin fibroblast cell line (CRL 1508). Fibroblasts were grown in 10% fetal bovine serum (FBS) for 24 hr, then washed with serum‐free medium before adding serum‐free defined medium (SFDM) containing insulin and transferrin. To this medium were added varying concentrations of human plasma‐derived AB‐PDGF and α2M, alone or in combination. Receptor‐recognized α2M was prepared by treatment with methylamine. Both native α2M and the α2M‐methylamine (α2M‐MA) were tested for growth promoting activity in the absence or presence of PDGF. After 3 days, a concentration‐dependent growth curve of fibroblast proliferation was demonstrated for PDGF alone, with near maximal stimulation reached at 15–20 ng/ml PDGF. α2M and α2M‐MA alone had no effect on cell proliferation. However, α2M‐MA concentrations above 32 μg/ml synergistically enhanced PDGF‐stimulated proliferation >100% in the presence of 15 ng/ml PDGF. Native α2M enhanced PDGF‐stimulated growth 80–100% above PDGF controls only at low concentrations (32–64 μg/ml α2M). High concentrations of native α2M (128–256 μg/ml) either had no effect on growth or were inhibitory to PDGF‐stimulated growth, depending on the cell type tested. Rat lung fibroblasts were shown to secrete a factor(s) that inhibited the trypsin‐binding capacity of native α2M. We further demonstrated that early passage RLFs possess specific cell‐surface receptors for [125I]‐PDGF and [125I]‐α2M‐MA, and preincubation of RLFs with α2M‐MA increased the specific binding of [125I]‐PDGF to the cell surface of these fibroblasts. Considered together, these data support the view that receptor‐recognized α2M synergistically enhances the proliferative capacity of PDGF. We postulate that receptor‐recognized αMs enhance PDGF‐stimulated growth by increasing the local concentration of PDGF at the cell surface, where the PDGF could be released in close proximity to its own receptors.
Transcellular processing of disulfide‐and thioether‐linked peroxidase‐polylysine conjugates in cultured MDCK epithelial cells
Authors: Wan Jiansheng , Persiani Stefano , Shen Wei‐Chiang 
Published: February 2005

AbstractHorseradish peroxidase (HRP) was conjugated to nondegradable polycationic poly(D‐lysine) (PDL) through either a thioether (HRP‐S‐PDL) or a disulfide (HRP‐SS‐PDL) linkage. The binding and transcytosis of these conjugates was studied in Madin‐Darby canine kidney (MDCK) cell monolayers grown on 3‐μm microporous polycarbonate filters. Conjugation of HRP to PDL with both linkages markedly increased the binding of this protein onto the cell monolayers. However, an enhancement of the transcellular transport of HRP in both apical‐to‐basal and basal‐to‐apical directions was observed only in HRP‐SS‐PDL, but not in HRP‐S‐PDL. HRP‐SS‐PDL transport was inhibited by colchicine and by 4oC incubation. The transport of 14C‐sucrose was not affected by the presence of conjugates. These results indicate that the transport of the conjugate across the cell monolayers was due to a transcellular process rather than to any leakage of the cell junction caused by polycations. The disulfide linkage between HRP and PDL was cleaved rapidly at the basal and, to a lesser extent, at the apical surface of the cell. Neuraminidase treatment decreased the binding of the conjugates onto the cell surface, but did not decrease the transcellular transport, suggesting that not all surface‐bound conjugates were available for transcytosis. These results demonstrate that disulfide linkages can be cleaved during transcytosis in MDCK cells. The cleavage, however, occurs mostly at the binding site on the cell surface, which may prevent the cellular uptake of the intact conjugate.
Thrombospondin gene expression is associated with mitogenesis in 3T3 cells: Induction by basic fibroblast growth factor
Authors: Donoviel Dorit B. , Amacher Sharon L. , Judge Kevin W. , Bornstein Paul 
Published: February 2005

AbstractGrowth factor‐depleted Swiss 3T3 cells responded to basic fibroblast growth factor (bFGF) with a burst of mitogenesis and with a rapid and marked increase in thrombospondin (TS) mRNA levels. mRNA levels for the α1 chain of type I collagen and for fibronectin were unaffected. At early times following stimulation (0–2 h), “superinduction” of TS mRNA by inhibition of protein synthesis with cycloheximide was not observed and the increase in TS mRNA could be attributed primarily to an increase in transcription rate of the TS gene. However, at later times (4–8 h) the combination of cycloheximide and bFGF superinduced TS mRNA levels, suggesting the existence of a labile inhibitor of transcription or a short‐lived RNase that might be produced in response to prolonged treatment with bFGF. In contrast to its stimulatory effect on 3T3 cells, bFGF did not stimulate the proliferation of mouse muscle BC3H1 cells nor did it cause an increase in TS mRNA levels, but BC3H1 cells do respond to bFGF by inhibition of myogenic differentiation. We propose, on the basis of these and other findings, that TS facilitates the progression of some anchorage‐dependent cells through the cell cycle.
A factor contained in plasma is required for IGF binding protein‐1 to potentiate the effect of IGF‐I on smooth muscle cell DNA synthesis
Authors: Clemmons D. R. , Gardner L. I. 
Published: February 2005

AbstractOne of the forms of the insulin‐like growth factor (IGF) binding proteins present in human amniotic fluid has been shown to potentiate the growth‐promoting effect of IGF‐I markedly. This study was undertaken to determine the cellular and hormonal factors that modulate this potentiation and to determine whether this protein would potentiate the effects of other mitogens. Although the combination of the IGFBP‐1 (20 ng/ml) and IGF‐I (10 ng/ml) induced a five‐ to sixfold increase in DNA synthesis compared with IGF‐I alone, this response required the simultaneous addition of IGF‐I with 0.1% platelet‐poor plasma (PPP). If PPP was omitted from the incubation medium, no increase above the effect that was obtained with IGF‐I alone was noted. Substitution of cerebrospinal fluid (CSF) for PPP permitted a full mitogenic response, although substitution with amniotic fluid resulted in no enhancement. The factor contained in PPP was heat and acid stable. If the binding protein was co‐incubated with fibroblast growth factor (FGF), platelet‐derived growth factor (PDGF), or epidermal growth factor (EGF), a slight inhibition of the cellular response to each of these factors was detected. Co‐incubation of IGF‐I with the IGF‐binding protein plus these other peptide growth factors resulted in no further enhancement of DNA synthesis above the level observed with IGF‐I and the binding protein alone. Likewise, addition of plasma proteins such as transferrin or albumin did not result in a further enhancement of the DNA synthesis response to IGF‐I plus binding protein, and these proteins could not substitute for PPP or IGFBP‐1. Transient exposure of the cultures (2 hr) to the binding protein plus IGF‐I resulted in a submaximal DNA synthesis response, and the binding protein had to be present continuously to achieve a maximal effect. These studies indicate that a factor contained in plasma and CSF is required for a maximal cellular response to IGFBP‐1 plus IGF‐I, and this factor does not appear to be a well‐defined mitogen.
Evidence for involvement of a nuclear envelope‐associated RNA helicase activity in nucleocytoplasmic RNA transport
Authors: Schröder Heinz C. , Ugarkovic Durdica , Langen Peter , Bachmann Michael , Dorn August , Kuchino Yoshiyuki , Müller Werner E.G. 
Published: February 2005

AbstractIt seems well established that translocation of at least some mRNAs through the nuclear pore is (1) an energy‐dependent process, and (2) dependent on the presence of the poly(A) segment attached to most mRNA species. We describe that RNA helicase (RNA duplex unwindase) activity is present in a nuclear envelope (NE) preparation, which also appears to be involved in nucleocytoplasmic RNA transport. This activity unwinds RNA : RNA hybrids. The helicase has a pH optimum of 7.5 and a temperature optimum of 30°C. Applying the sealed NE vesicle system, it was shown that duplex RNA species are readily released from the vesicles in an unidirectional manner, in contrast to single‐stranded RNA, which is much slower transported into the extravesicular space. Attachment of a poly(A) segment to the RNA duplex additionally increases the efflux rate of this RNA. Efflux of duplex RNA but not efflux of single‐stranded RNA was strongly inhibited by formycin B 5′‐triphosphate. Our results suggest that, besides poly(A), duplex structures, if present in a given RNA, modulate and control the export of RNA.
Stromal influences on transformation of human mammary epithelial cells overexpressing c‐myc and SV40T
Authors: Valverius Eva M. , Ciardiello Fortunato , Heldin Nils‐Erik , Blondel Bruno , Merlo Giorgio , Smith Gilbert , Stampfer Martha R. , Lippman Marc E. , Dickson Robert B. , Salomon David S. 
Published: February 2005

AbstractThe proto‐oncogene c‐myc and the oncogene SV40T, both of which have been implicated in the process of cellular immortalization in vitro, have been introduced via amphotropic retroviral expression vectors into the human mammary epithelial cell (HMEC) line 184A1N4 (A1N4). Two stable cell lines were established by growth in selective medium and were found to overexpress either c‐myc (A1N4‐myc) or SV40T antigen (A1N4‐T). Neither the A1N4, A1N4‐myc, or A1N4‐T cells will grow in soft agar or form tumors in nude mice. However, A1N4‐T or A1N4‐myc cells, but not the parental A1N4 cells, form colonies in soft agar in response to either epidermal growth factor (EGF), transforming growth factor α (TGFα), or basic fibroblast growth factor (bFGF). Like EGF and TGFα, bFGF is moderately mitogenic for the anchorage‐dependent growth (ADG) of all three cell lines. Further, co‐cultivation of A1N4‐T or A1N4‐myc cells with primary diploid mammary fibroblasts can also induce the anchorage‐independent growth (AIG) and stimulate the ADG of A1N4‐T or A1N4‐myc. In addition, conditioned medium obtained from these mammary fibroblasts also stimulated the AIG of the A1N4‐T and A1N4‐myc cells and was found to contain immunoreactive TGFα and bioactive FGF. The mammary fibroblasts express specific mRNA transcripts for bFGF and acidic FGF (aFGF). These results suggest that growth factors such as TFGα or FGF, which may be derived from the adjacent mammary stroma, might influence in a paracrine manner the phenotypic characteristics of a population of human mammary epithelial cells toward transformation.
The Na+,K+,2Cl−‐cotransport system in HeLa cells: Aspects of its physiological regulation
Authors: Kort Jens J. , Koch Gebhard 
Published: February 2005

AbstractWe have previously reported on the biochemical properties of a Na+,K+,2Cl−‐cotransport in HeLa cells and here we deal with aspects of its physiological regulation. Na+,K+,2Cl−‐cotransport in HeLa cells was studied by 86Rb+ influx and 86Rb+/22Na+ efflux measurements. The effects of rat atrial natriuretic peptide (ANP), isoproterenol, and amino acids on 86Rb+ flux, mediated by the bumet‐anide‐sensitive Na+, K+, 2Cl−‐cotransport system and the ouabain‐sensitive Na+/K+‐pump, were investigated. ANP reduced bumetanide‐sensitive 86Rb+ influx under isotonic as well as under hypertonic conditions. Similar decrease of bumetanide‐sensitive 86Rb+ influx was observed in the presence of 8‐bromo‐cGMP, while neither isoproterenol as a β‐receptor agonist nor 8‐bromo‐cAMP‐could alter bumetanide‐sensitive 86Rb+ influx. Furthermore, efflux of 86Rb+ and 22Na+ was greatly reduced in the presence of bumetanide and ANP. Together with our recent findings, showing functionally active, high affinity receptors for ANP on HeLa cells (Kort and Koch, Biochim. Biophys. Res. Commun. 168:148–154, 1990), this study indicates that ANP participates in the regulation of the Na+, K+, 2Cl−‐cotransport system in HeLa cells. Further measurements revealed that amino acids as present in the growth medium (Joklik's minimal essential medium) and the amino acid derivative α‐methyl‐aminoisobutyric acid (metAlB, 1 and 5 mM, respectively) also reduced Na+, K+, 2Cl−‐cotransport‐mediated 86Rb+ uptake and diminished the stimulatory effect of hypertonicity on the cotransporter. In addition, the Na+/K+‐pump was markedly stimulated in the presence of amino acids, while neither ANP and 8‐Br‐cGMP nor isoproterenol and 8‐Br‐cAMP had a significant effect on the activity of the Na+/K+‐pump.
Extracellular matrix derived from Trypanosoma cruzi infected endothelial cells directs phenotypic expression
Authors: Morris Stephen A. , Wittner Murray , Weiss Louis , Hatcher Victor B. , Tanowitz Herbert B. , Bilezikian John P. , Gordon Portia B. 
Published: February 2005

AbstractInfection of confluent human umbilical vein endothelial cells by the parasite Trypanosoma cruzi results in the appearance of an altered heparan sulfate proteoglycan (HSPG) isolated from the extracellular matrix of infected endothelial cells (ECMi). HSPG from ECMi differed from HSPG obtained from the extracellular matrix of uninfected endothelial cells (ECMu) by virtue of an 8–10‐fold increase in sulfation and a different elution pattern using DEAE Sepharose chromatography. Analysis of the HSPG that binds to acidic fibroblast growth factor (aFGF) revealed that infection increased the proportion of HSPG that binds to aFGF by 35%. Heparitinase and alkaline borohydride treatment of aFGF‐binding HSPG and chromatographic resolution on Sepharose CL4B column revealed an infection‐associated 10‐fold increase in sulfation of the GAG side chain with no significant change in the migration of the core protein. In addition, the aFGF binding HSPG isolated from ECMi demonstrated a markedly attenuated synergistic mitogenic activity with aFGF in a cell proliferation assay. All of the infection associated changes in HSPG could be demonstrated in HSPG obtained from uninfected endothelial cell cultures grown on ECMi. Hence, the ECMi is associated with signals capable of modulating the ECM associated metabolism of uninfected endothelial cells. This facility of ECMi was also shown to extend to patterns of Gs protein synthesis as revealed by Western blot analysis. The observation that the ECM produced by infected endothelial cells can direct the synthetic patterns of uninfected endothelial cells in a manner uniquely observed in infected endothe‐lial cells suggests a plausible pathway by which infection of only a few cells can ultimately result in the coordinate responses of neighboring uninfected cells.