bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of platelet‐derived growth factor on DNA synthesis and gene expression in bone marrow stromal cells derived from adult and old rats
Authors: Tanaka Hiroshi , Liang C. Tony 
Published: February 2005

AbstractThe effects of platelet‐derived growth factor (PDGF) on DNA synthesis and mRNA expression of osteoblast markers in marrow stromal cells derived from adult (6 months) and old (24 months) rats were examined. Treatment of stromal cells from adult rats with dexamethasone induced the appearance of osteoblast‐like cells. PDGF partially also inhibited the differentiation of stromal cells induced by dexamethasone. In cultures of serum‐starved stromal cells, PDGF stimulated [3H]‐thymidine incorporation into DNA in a dose‐dependent manner with a maximum stimulation of 15‐fold at 500 ng/ml. By comparison, insulin‐like growth factor (IGF‐I) has a small effect on [3H] ‐thymidine incorporation. The effect of PDGF and IGF‐I on DNA synthesis was additive. Treatment of the confluent stromal cells from adult rats with PDGF increased the mRNA level of osteopontin fourfold without any significant effect on alkaline phosphatase and type I collagen mRNAs. In contrast, dexamethasone stimulated the mRNA expression of alkaline phosphatase, type I collagen, and osteopontin 2.1‐, 2.3‐, and 14‐fold, respectively. Addition of PDGF to dexamethasone‐treated cells failed to induce any further increase in osteopontin expression whereas the expression of alkaline phosphatase and type I collagen was partially reduced. The expression of osteocalcin mRNA was negligible in stromal cells but stimulated several fold by dexamethasone and 1,25(OH)2D3. PDGF inhibited drastically the elevation of osteocalcin mRNA. In contrast, IGF‐I stimulated type I collagen expression 100% without any appreciable effect on the expression of osteopontin and alkaline phosphatase. The stimulatory effect of PDGF on osteopontin expression was augmented by IGF‐I. Furthermore, PDGF attenuated the stimulatory effect of IGF‐I on type I collagen expression. The responses of cultured cells from old rats to growth factors were also examined. PDGF or PDGF plus IGF‐I increased [3H]‐thymidine incorporation in stromal cells from old rats but to a lesser extent. However, PDGF was equally effective in stimulating osteopontin expression in cells from both adult and old rats. We concluded that PDGF is a potent mitogen but that the response of stromal cells from old rats is impaired. In addition, PDGF stimulates osteopontin expression in stromal cells and this effect is not age dependent. © 1995 Wiley‐Liss, Inc.
Heat‐Induced bFGF gene expression in the absence of heat shock element correlates with enhanced AP‐1 binding activity
Authors: Erdös Géza , Lee Yong J. , Cho Joong M. , Corry Peter M. 
Published: February 2005

AbstractBasic fibroblast growth factor (bFGF) has been shown to be a potent mitogen and a promoter of angiogenesis. It has been hypothesized that the expression of the bFGF gene may be induced by stress of various types. To test that hypothesis, we investigated the expression of the bFGF gene during heat treatment in adriamycin‐resistant (MCF‐7/ADR) and‐sensitive (MCF‐7) human breast carcinoma cells. Under normal growth conditions, the bFGF mRNA was detected in MCF‐7/ADR cells, while it was not detectable in MCF‐7 cells by Northern blot analysis. During heating at 41°C, the level of bFGF mRNA increased in MCF‐7/ADR cells and the message became detectable in the MCF‐7 cell line. However, after continuous heating at 41°C for 24 h, the bFGF mRNA level decreased to control level in MCF‐7/ADR cells. Interestingly, simultaneous treatment with heat and 60 m̈g/ml H‐7 (1‐(isoquinolinylsulfonyl)‐2‐methylpiperzine, a potent PKC inhibitor) decreased the level of bFGF mRNA in MCF‐7/ADR cells. These results suggest that a protein kinase, likely PKC, is involved in the transcriptional regulation of the heat‐enhanced bFGF gene expression in human breast carcinoma cells. Although no heat shock element can be identified in the promoter of the bFGF gene, we observed that the AP‐1 binding activity to a TPA responsive element (TRE)‐like sequence in the promoter of bFGF gene was enhanced by heat, as tested by mobility shift assay. Antibody developed against the c‐Jun and c‐Fos proteins inhibited the AP‐1 binding activity to TRE. Therefore, the AP‐1 complex appears to be responsible for the heat‐enhanced binding to the TRE‐like motif of the bFGF gene. Furthermore, the increased AP‐1 binding activity does not require new protein synthesis but activation of the preexisting c‐Jun proteins. © 1995 Wiley‐Liss, Inc.
Regulation of platelet‐derived growth factor A and B chain gene expression in bone marrow stromal cells
Authors: Abboud Sherry L. 
Published: February 2005

AbstractMBA‐2, bone marrow‐derived endothelial stromal cells, express platelet‐derived growth factor (PDGF) A and B chain mRNAs and secrete PDGF activity that is induced by TGF‐beta. Either chain of the PDGF molecule could modulate hematopoiesis and stromal cell growth. Intracellular pathways that regulate PDGF expression in the marrow microenvironment are unknown. In the present study, we examined the mechanisms that mediate PDGF A and B chain mRNA induction by TGF‐beta and the role of protein kinase C (PKC) and cyclic AMP in PDGF regulation. TGF‐beta was tested in parallel with PMA, an activator of phorbol ester‐dependent PKC isoforms. Both PMA (10−7M) and TGF‐beta (2.5 ng/ml) increased PDGF A and B chain mRNA levels. The serine/threonine protein kinase inhibitor, H7, blocked PDGF A and B chain mRNA induction in response to TGF‐beta. However, down‐regulation of PKC by prolonged incubation with PMA failed to abolish TGF‐beta induction of PDGF A and B chain mRNAs. These findings indicate that induction of PDGF A and B chain mRNAs can be mediated via phorbol ester‐dependent PKC pathway. In contrast, H7‐sensitive protein kinase(s) other than phorbol ester‐sensitive protein kinase C mediate the effect of TGF‐beta. Agents that increase cAMP were also tested for their effect on PDGF gene expression. TGF‐beta‐mediated induction of PDGF A and B chain mRNAs was markedly inhibited by cAMP. cAMP also blocked stimulation of PDGF A chain mRNA by PMA. The positive and negative signaling mechanisms involved in modulating PDGF in the microenvironment may be important for determining hematopoietic and stromal cell responses in vivo. © 1995 Wiley‐Liss, Inc.
Binding and endocytosis of 39 kDa protein by mdbk cells
Authors: Vettenranta Kim , Bu Guojun , Schwartz Alan L. 
Published: February 2005

AbstractA 39 kDa protein copurifies with the low density lipoprotein receptor‐related protein (LRP) and regulates ligand interactions with LRP. In our recent studies on the clearance of the 39 kDa protein in vivo, we demonstrated that once the liver LRP receptors were saturated, the kidney necame the major organ responsible for the 39 kDa protein clearance (Warshawsky et al., 1993, J. Clin. Invest., 92:937‐944). The current study was undertaken in order to investigate the potential binding and cellular processing of the 39 kDa protein by kidney‐derived MDBK cells. Herein we demonstrate specific, high‐affinity, saturable, and Ca2+‐dependent binding of the 125I‐39 kDa protein to MDBK cells (Kd ∼ 10‐15 nM, 50‐70,000 bindings sites per cell). Cellular uptake and degradation of the 125I‐39 kDa protein by MDBK cells was also demonstrated with kinetics typical of receptor‐mediated endocytosis. Using chemical crosslinking we show that LRP in part mediates the binding of 125I‐39 kDa protein to the MDBK cell surface. In addition, the presence of functional LRP on the MDBK cell surface was confirmed by the specific binding of activated α2‐macroglobulin, another ligand of LRP. Our data thus demonstrate the ability of kidney‐derived MDBK cells to specifically bind, endocytose, and degrade the 39 kDa protein. © 1995 Wiley‐Liss, Inc.
Functional characterization of lymphoid cells generated in serum‐deprived culture stimulated with stem cell factor and interleukin 7 from normal and autoimmune mice
Authors: Ashany Dalit , Elkon Keith B. , Migliaccio Giovanni , Migliaccio Anna Rita 
Published: February 2005

AbstractWe have investigated the phenotypic and functional characteristics of murine pre‐B cells obtained in semisolid and liquid culture with stem cell factor (SCF) and interleukin 7 (IL‐7). Both serum‐supplemented and serum‐deprived culture conditions were used. The source of bone marrow cells was either normal mice (CD1 and C3H) or the lupus strain of mice MRL/Ipr and its congenic strain MRL/+. SCF (100 ng/ml) and IL‐7 (250 ng/ml) supported murine B cell proliferation in vitro from all the murine strains analyzed both in serum‐supplemented and serum‐deprived conditions. Maximal colony growth was observed in both cases when the factors were used in combination. The growth factors alone induced some colony growth in serum‐supplemented cultures but were either ineffective or had modest activity in serum‐deprived cultures. Cells harvested from the colonies or generated in liquid cultures and stimulated with SCF + IL‐7 in the absence of serum had almost exclusively a pre‐B cell phenotype (BP‐1+, B220+, slg‐, CD4‐, CD8‐, Mac‐1, RB‐6‐). Both the maximal colony growth in semisolid culture and the maximal number of cells in liquid culture were observed at day 12–14. At this time, the pre‐B cells failed to differentiate further and started to die. Pre‐B cells generated in vitro were, however, capable of differentiating in vivo. SCID mice injected with 2 × 106 pre‐B cells had readily detectable serum levels of IgM (54 ± 26 m̈g/ml) and IgG (60 ± 95 m̈g/ml) at 4 weeks and 6 weeks posttransplantation, respectively. Mature B and T cells of the donor major histocompatibility complex type were detected in the SCID mice at sacrifice 14 weeks posttransplantation. These data indicate that purified (>80% BP‐1+) populations of functional pre‐B cells can be grown from murine bone marrow of normal mice as well as of lupus mice in serum‐deprived cultures stimulated with SCF and IL‐7. These cultures, therefore, provide a highly enriched source of pre‐B cells but also contain T cell precursors that differentiate upon adoptive transfer into SCID mice. © 1995 Wiley‐Liss, Inc.
Evidence for the presence of a Hg‐inhibitable water‐permeability pathway and aquaporin 1 in A5 salivary epithelial cells
Authors: Lazowski Krzysztof W. , Li Jun , Delporte Christine , Baum Bruce J. 
Published: February 2005

AbstractThe molecular basis of water‐permeability in salivary and other exocrine glands is not understood. We have examined two well‐studied salivary epithelial cell lines for evidence of a Hg‐inhibitable water‐permeability pathway. A5 and HSG cells are derived from rat and human submandibular glands, respectively. Only A5 cells are derived from rat and human submandibular glands, respectively. Only A5 cells demonstrated such a pathway. The rate of A5 cell osmotic shrinkage was inhibited about fivefold in the presence of 300 m̈M HgCl2. To determine if this activity was associated with the expression of the prototypical water channel (aquaporin, AQP) AQP1, we used three separate experimental approaches; Northern analysis and reverse transcription‐polymerase chain reaction (RT‐PCR) analysis of isolated mRNA, and Western analysis of cell membranes. All three methods yielded positive results with A5 cells and negative results with HSG cells. The ∼800 bp product of RT‐PCR was analyzed further by sequencing and restriction enzyme digestion. The results were consistent with the previously reported coding region sequence for rat kidney AQP1. The aggregate data demonstrate that marked differences in water‐permeability and water channel expression exist in these two salivary epithelial cell lines. © 1995 Wiley‐Liss, Inc.
Nucleolar p120 is expressed as a delayed early response gene and is inducible by DNA‐damaging agents
Authors: Fonagy Anna , Swiderski Carol , Freeman James W. 
Published: February 2005

AbstractRegulation of the expression of the growth‐related nucleolar p120 protein was examined in serum‐deprived and stimulated nontransformed and SV40‐transformed WI‐38 human fibroblasts. In quiescent cells, transcriptional activity of the p120 gene was very low or undetectable, and the steady‐state levels of the p120 mRNA and the p120 protein were also negligible. The transient expression of the p120 gene in the cell cycle was detected in middle G1‐phase after the expression of the early response genes and before the expression of the DNA‐synthesis genes. Protein synthesis was required for the induction of p120 expression in serumstimulated cells. The increased level of p120 mRNA in middle G1‐phase was attributed to an increased transcription rate of the p120 gene, and not to a change in p120 mRNA stability. The calculated half‐life of p120 mRNA was unchanged (1.8 ± 0.2 hr) in all four cell conditions tested; i.e., in middle G1‐ or S‐phase cells and in exponentially growing normal or transformed cells. Transcription rate of the p120 gene was correlated with the steady‐state levels of either p120 protein or p120 mRNA. A sharp increase in p120 mRNA level occurred in both normal and transformed cells treated with actinomycin D used to examine p120 mRNA stability. This induction of p120 mRNA expression was seen in early G1‐phase, but not in quiescent cells, or in middle to late G1‐phase when cells expressed the highest level of p120 mRNA. The same expression pattern was seen by treatment with chlorambucil, another DNA‐damaging agent. The conclusions of these studies are that the expression of p120 (1) is serum inducible in a fashion characteristic of the delayed early response gene products, (2) requires the presence of newly synthesized proteins, (3) is regulated transcriptionally, and (4) can be induced by DNA‐damaging agents. © 1995 Wiley‐Liss, Inc.
Secretin potentiates cholecystokinin‐stimulated amylase release by AR4‐2J cells via a stimulation of phospholipase C
Authors: Bold Richard J. , Ishizuka Jin , Townsend Courtney M. , Thompson James C. 
Published: February 2005

AbstractAlterations in the activity of phospholipase C (PLC) are thought to be the primary intracellular events leading to pancreatic acinar cell exocytosis of zymogen granules. When multiple hormones, each of which may stimulate different signal transduction pathways, bind to cell surface receptors, the cell must integrate these signals into a common response through communication (cross‐talk) among intracellular second messengers. We show that cholecystokinin (CCK) induces amylase secretion from AR4‐2J pancreatic acinar cells via stimulation of PLC activity. Secretin indirectly stimulated the PLC pathway through cross‐talk of the activated cAMP pathway to potentiate the CCK‐stimulated amylase secretion. Therefore, secretin potentiated the acinar cell secretory response to CCK by cAMP‐mediated cross‐talk with the PLC signal transduction pathway. © 1995 Wiley‐Liss Inc.
Changes in membrane potential during the progression of MCF‐7 human mammary tumor cells through the cell cycle
Authors: Wonderlin William F. , Woodfork Karen A. , Strobl Jeannine S. 
Published: February 2005

AbstractWe previously reported that MCF‐7 cells were arrested in the G0/G1 phase of the cell cycle by agents known to block the activity of ATP‐sensitive potassium channels (Woodfork et al., 1995, J. Cell Physiol. 162:163–171). The goal of our current study was to determine if MCF‐7 cells undergo changes in membrane potential during the cell cycle that might be linked to changes in K permeability. The resting membrane potentials of unsynchronized MCF‐7 cells during exponential growth phase were measured using sharp glass microelectrodes, and they ranged from −58.6 mV to −2.7 mV. The distribution of membrane potentials was best fitted by the sum of four Gaussian distributions with means of −9.0 mV, −17.4 mV, −24.6 mV, and −40.4 mV. These membrane potential groups were designated D (depolarized), ID (intermediate depolarized), IH (intermediate hyperpolarized), and H (hyperpolarized), respectively. The membrane potential was sensitive to the substitution of external K and Na but not Cl. The K:Na permeability ratio increased in proportion to the negativity of the membrane potential. MCF‐7 cells pharmacologically arrested in G0/G1 phase were depolarized compared to control, with cells shifted from the H and IH groups to the D group. Tamoxifen‐arrested cells chased from G0/G1 into S phase by the addition of mitogenic concentrations of 17β‐estradiol were not depolarized, and these cells were shifted from the D group back to the IH and H groups. We conclude that MCF‐7 cells hyperpolarize during passage through G0/G1 and into S phase, and this hyperpolarization probably results from an increase in the relative permeability of the plasma membrane to K. © 1995 Wiley‐Liss Inc.
Short circuiting stress protein expression via a tyrosine kinase inhibitor, herbimycin A
Authors: Hegde Ramanujan S. , Zuo Jianru , Voellmy Richard , Welch William J. 
Published: February 2005

AbstractWe set out to identify pharmacological means by which to activate the so‐called heat shock or stress response and thereby harness the protective effect afforded to the cell by its acquisition of a thermotolerant phenotype. An earlier report by Murakami et al. (1991, Exp. Cell Res., 195: 338–344) described the increased expression of the 70 kDa heat shock proteins in human A431 cells exposed to Herbimycin A (HA), a benzoquinoid ansamycin antibiotic. We show here that treatment of cells with HA results in the increased expression of all of the constitutively expressed stress proteins and confers upon the cells a thermotolerant‐like phenotype. Increases in the expression of the stress proteins continued for as long as the cells were exposed to the drug and was independent of the pre‐existing levels of the stress proteins. Unlike heat shock or other metabolic stressors, we did not observe any adverse cellular effects following HA exposure. For example, unlike most agents/treatments that elicit the stress response HA‐treated cells exhibited no obvious abnormalities with respect to protein maturation, protein insolubility, the integrity of the intermediate filament cytoskeleton, or overall cell viability. In addition, unlike other metabolic stressors, HA treatment did not result in the translocation of hsp 73 into the nucleus/nucleolus. Finally, for at least rodent cells, HA exposure did not result in any obvious activation of the heat shock transcription factor. Based on these findings, we suggest that HA treatment of cells results in a “short‐circuiting” of the pathway(s) that normally regulates the expression of the stress proteins. These results are discussed as they pertain to the potential use of HA in animals as a way to harness the protective effects afforded by the stress response. © 1995 Wiley‐Liss Inc.
Ouabain induces inhibition of the progression phase in human T‐cell proliferation
Authors: Brodie Chaya , Tordai Atilla , Saloga Joachim , Domenico Joanne , Gelfand Erwin W. 
Published: February 2005

AbstractOuabain, a specific inhibitor of the Na‐K ATPase, has been shown to exert immunosuppressive effects. The goals of this study were to define the stage of the proliferative response which is sensitive to ouabain and to correlate the inhibitory action of ouabain on cell proliferation with its effect on Na‐K ATPase activity. We found that ouabain inhibited T‐cell proliferation in a dose‐dependent manner and this inhibition was similar in CD4+ and CD8+ T cells. To define the role of the Na‐K ATPase in early activation of T lymphocytes, we examined the effects of ouabain on the induction of competence (acquisition of responsiveness to interleukin (IL)‐2 or IL‐4) by phytohemagglutinin (PHA) or the combination of phorbol dibutyrate/ionomycin. Ouabain, at concentrations that completely inhibited the enzyme activity, did not interfere with the induction of competence, suggesting that although activated cells express increased activity of Na‐K ATPase, this enzyme activity does not play a role in early activation pathways. In contrast, ouabain inhibited the progression phase to DNA synthesis in a dose‐dependent manner even at concentrations that had little or no effect on NaK ATPase activity. This inhibition was not due to a decrease in the production of IL‐2 but rather to an inhibition of the expression of the p55 and p75 subunits of the IL‐2 receptor (IL‐2R). The inhibition of p55 appeared to occur at the mRNA level. These results indicate that the activity of the NaK ATPase is not essential for the induction of competence or early activation. On the other hand, inhibition of cell proliferation and transcription of IL‐2R subunits by low concentrations of ouabain may be related to changes in intracellular K+ concentrations or to inhibition of membranal phospholipid metabolism secondary to alteration in NaK ATPase activity. © 1995 Wiley‐Liss, Inc.
Stem cell factor can overcome inhibition of highly purified human burst‐forming units‐erythroid by interferon γ
Authors: Dai Chun‐Hua , Krantz Sanford B. , Kollar Kathleen , Price James O. 
Published: February 2005

AbstractHighly purified human blood burst‐forming units‐erythroid (BFU‐E) were used to study the effects of interferon γ (IFNγ). IFNγ inhibited erythroid colony formation, cell proliferation, and differentiation of day 3 to day 6 mature BFU‐E in a dose‐dependent manner. The primitive BFU‐E (day 1 and day 2 cells) and later day 7 cells were less affected. IFNγ dose‐response experiments demonstrated that the number and size of erythroid colonies were reduced at a concentration of 500 U/ml with more complete inhibition at 1,000 U/ml. Inhibition of day 4 to day 6 erythroid progenitors was first noted by 72 h of incubation with IFNγ, and target cell growth and differentiation continued to decrease with further incubation. IFNγ also induced erythroblast apoptosis which was demonstrated by both nuclear condensation and fragmentation plus flow cytometry with in situ end‐labelling. Because day 3 to day 6 cells need stem cell factor (SCF) for development in serum‐free culture, the relationship of IFNγ inhibition to this growth factor was investigated. The reduction in the number of erythroid colonies by IFNγ was reversed by SCF although the colony size was not completely re‐established. In contrast, interleukin‐3 did not have the capacity to overcome the inhibitory effects of IFNγ. Since IFNγ blood levels are elevated in some anemias of chronic disease, IFNγ may have a role in promoting this anemia and its inhibitory effect might be better overcome by SCF plus EP. However, the mechanism by which these growth factors overcome the inhibition of IFNγ, or vice versa, is unknown at the present time. © 1995 Wiley‐Liss, Inc.