bluuebunny/crossref_metadata_2025_split huggingface dataset:
Release from quiescence stimulates the expression of integrin α5β1 which regulates DNA synthesis in human fibrosarcoma HT1080 cells
Authors: Wang Danhui , Birkenmeier Thomas M. , Yang Junhua , Venkateswarlu Srinivas , Humphrey Lisa , Brattain Michael G. , Sun Luzhe 
Published: February 2005

AbstractWe show that integrin α5 subunit expression is stimulated when human fibrosarcoma HT1080 cells are released from quiescence. The α5 subunit mRNA level in quiescent HT1080 cells was increased 24 hr after their release by 10% fetal bovine serum‐containing medium reaching a maximum of 2.5 fold on day 2. Similar levels of induction of cell‐surface α5 subunit protein as well as b̃1 subunit protein were also observed. This resulted in a significant increase of cell attachment to fibronectin. The serum stimulation also increased α5 subunit promoter activity by twofold which was protein synthesis independent. Subsequent deletion of α5 subunit promoter DNA showed that the cis‐element responsible for the activation is located between ‐ 92 bp and the transcription start site. The promoter activity was not induced until 12 hr after the release. Comparison of the effect of a serum‐free medium and a 10% fetal bovine serum‐supplemented medium revealed that both the DNA synthesis and α5 subunit induction were independent of exogenous growth factors. The increased integrin α5b̃1 appears to function by reducing mitogenic activity since blockade of fibronectin binding to its receptor with a RGD peptide, a monoclonal anti‐fibronectin antibody, or a monoclonal anti‐α5 subunit antibody during the release from quiescence significantly stimulated DNA synthesis. On the other hand, stable overexpression of the α5 subunit resulted in decreased DNA synthesis. © 1995 Wiley‐Liss, Inc.
Cytoskeletal regulation of Caco‐2 intestinal monolayer paracellular permeability
Authors: Ma Thomas Y. , Hollander Daniel , Tran Le Thuy , Nguyen Don , Hoa Neil , Bhalla Deepak 
Published: February 2005

AbstractAn abnormal increase in intestinal paracellular permeability may be an important pathogenic factor in various intestinal diseases. The intracellular factors and processes that regulate and cause alteration of intestinal paracellular permeability are not well understood. The purpose of this study was to examine some of the intracellular processes involved in cytoskeletal regulation of intestinal epithelial paracellular permeability using the filter‐grown Caco‐2 intestinal epithelial monolayers. Cytochalasin‐b and colchicine were used to disrupt the cytoskeletal elements, actin microfilaments, and microtubules. Cytochalasin‐b (5 m̈g/ml) and colchicine (2 × 10−5M) at the doses used caused marked depolymerization and disruption of actin microfilaments and microtubules, respectively. Cytochalasin‐b‐induced disruption of actin microfilaments resulted in perturbation of tight junctions and desmosomes and an increase in Caco‐2 monolayer paracellular permeability. The cytochalasin‐b‐induced disruption of actin microfilaments and subsequent changes in intercellular junctional complexes and paracellular permeability were not affected by inhibitors of protein synthesis (actinomycin‐D or cycloheximide) or microtubule function (colchicine), but were inhibited by metabolic energy inhibitors (2,4‐dinitrophenol or sodium azide). The cytochalasin‐b‐induced disturbance in Caco‐2 actin microfilaments and intercellular junctional complexes and increase in paracellular permeability were rapidly reversed. The paracellular pathway “re‐tightening” following cytochalasin‐b removal was not affected by actinomycin‐D, cycloheximide, or colchicine, but was inhibited by 2,4‐dinitrophenol and sodium azide. The colchicine‐induced disruption of microtubules did not have significant effect on actin microfilaments, intercellular junctions, or paracellular permeability. These findings suggest that cytochalasin‐b‐induced increase in Caco‐2 monolayer paracellular permeability was due to actin microfilament mediated perturbation of intercellular junctional complexes. The re‐tightening of paracellular pathways (following removal of cytochalasin‐b) resulted from energy‐mediated re‐assembly of pre‐existing actin microfilaments and intercellular junctional complexes. This re‐closure process did not require protein synthesis or microtubule‐mediated shuttling process. © 1995 Wiley‐Liss, Inc.
Two major pathways of zinc(II) acquisition by human placental syncytiotrophoblast
Authors: Bax Christopher M. R. , Bloxam David L. 
Published: February 2005

AbstractUptake of zinc into placental villous syncytiotrophoblast is the first step in its transfer from mother to fetus. To help characterise physiologically significant pathways of zinc accumulation by these cells, we incubated cultured layers of syncytiotrophoblast cells derived from human near‐term placental tissue with serum ultrafiltrate (containing the zinc complexed with low molecular mass serum constituents), dialysed serum (containing the zinc bound to the serum proteins) and whole serum, each of whose endogenous zinc was tracer‐labelled with 65Zn(II). Zinc label from both fractions of serum readily entered a rapidly labelled EDTA‐sensitive cellular compartment, probably representing zinc bound to the outside cell surface and in accumulative fashion, an EDTA‐resistant compartment, probably consisting largely of internalised cellular zinc. Movement of zinc into the EDTA‐resistant pool was strongly temperature‐dependent and did not occur via the EDTA‐sensitive pool from either serum source. Transfer of zinc from the low molecular mass serum fraction into the EDTA‐resistant pool was saturable, the concentration giving half‐maximal rate being 1.2 m̈mol/l nonprotein‐bound zinc. No nonsturable component was detected. Zinc from the serum protein‐bound fraction entered by a saturable component, already saturated at physiological total protein‐bound zinc concentration, and by an apparently nonsaturable component, not appreciably accounted for by nonspecific fluid‐phase endocytosis. The results show that zinc is acquired by placental syncytiotrophoblast from the low molecular mass serum zinc pool probably by a carrier‐mediated process, and at least as importantly, from the zinc bound to serum protein, possibly by an endocytic mechanism. © 1995 Wiley‐Liss, Inc. © 1995 Wiley‐Liss, Inc.
Members of the 70 kDa heat shock protein family specifically recognize sulfoglycolipids: Role in gamete recognition and mycoplasma‐related infertility
Authors: Boulanger J. , Faulds D. , Eddy E. M. , Lingwood C. A. 
Published: February 2005

AbstractWe have previously shown that several mycoplasma species associated with infertility bind specifically to sulfated glycolipids isolated from the mammalian reproductive tract. We now show that a germ cell‐specific sulfoglycolipid binding protein (SLIP 1), which is a potent inhibitor of sperm/egg binding in vitro, is immunologically related to the heat shock protein (Hsp) 70 family of stress proteins and that Hsps are surface antigens in male germ cells. Our present data demonstrate that several mycoplasma and mammalian Hsps share this glycolipid binding specificity in vitro, and suggest that surface Hsps can function as adhesins which mediate sulfoglycolipid recognition in infectious disease and normal reproductive physiology. © 1995 Wiley‐Liss Inc.
Protein kinase A regulates the degradation rate of Rs acetylcholine receptors
Authors: Xu Rufeng , Salpeter Miriam M. 
Published: February 2005

AbstractAcetylcholine receptors at the neuromuscular junction of innervated vertebrate muscle (called Rs AChRs) have a stable degradation rate (t1/2∼8–12 days) which accelerates after denervation to a half‐life of ∼3 days, but can be restabilized by reinnervation or by cAMP. We examined the mechanism by which cAMP regulates the Rs degradation rate. When dibutyryl cAMP (DB‐cAMP) was applied to denervated mouse diaphragms in organ culture, it stabilized the accelerated degradation rate of the Rs. We found that this stabilization is reversible upon removal of the DB‐cAMP, is cAMP specific and is mediated by intracellular cAMP. A major observation of this study is that the cAMP‐induced stabilization of Rs AChRs is via protein kinase A (PKA), since H89, a PKA inhibitor, blocked the DB‐cAMP induced stabilization of Rs, and H85, an analog of H89, which does not inhibit PKA but does inhibit other kinases as efficiently as H89, did not prevent the DB‐cAMP‐induced stabilization of Rs degradation. These results suggest that the cAMP messenger system via a PKA‐dependent pathway could be among the mechanisms whereby the nerve regulates AChR degradation. © 1995 Wiley‐Liss Inc.
Significant induction of spermidine/spermine N1‐acetyltransferase without cytotoxicity by the growth‐supporting polyamine analogue 1,12‐dimethylspermine
Authors: Yang Jianing , Xiao Lei , Berkey Kimberly A. , Tamez Pamela A. , Coward James K. , Casero Robert A. 
Published: February 2005

AbstractThe superinduction of the polyamine catabolic enzyme spermidine/spermine N1‐acetyltransferase (SSAT) has been implicated in the cell type‐specific cytotoxic activity of some polyamine analogues. We now report that one polyamine analogue, 1, 12‐dimethylspermine (DMSpm), produces a large induction of SSAT with no significant effects on growth in the human large cell lung carcinoma line, NCI H157. This cell line has been demonstrated to respond to other analogues with SSAT superinduction and cell death. Treatment of the lung cancer cell line with DMSpm produces a rapid increase in SSAT activity and a near complete depletion of the natural polyamines. Additionally, DMSpm supports cell growth in cells which have been depleted of their natural polyamines by the ornithine decarboxylase inhibitor, 2‐difluoromethylornithine. The current results suggest that significant induction of SSAT can occur in the absence of cytotoxicity when the inducing polyamine analogue can support growth and that increased SSAT activity alone is not sufficient for cytotoxicity to occur. © 1995 Wiley‐Liss Inc.
Interleukin‐4 inhibits lipopolysaccharide‐induced expression of prostaglandin H synthase‐2 in human alveolar macrophages
Authors: Yano Tatsutoshi , Hopkins Harvey A. , Hemplel Stephen L. , Monick Martha , Hunninghake Gary W. 
Published: February 2005

AbstractSeveral studies have shown that interleukin‐4 (IL‐4) down‐regulates synthesis of prostaglandin E2 (PGE2). We evaluated the mechanisms for this suppression in human alveolar macrophages (HAMs). Normal HAMs were obtained from healthy nonsmoking volunteers. The cells either remained unstimulated, or were exposed to 10 μg/ml of lipopolysaccharide (LPS) and/or various amounts of IL‐4. LPS alone induced the synthesis of large amounts of PGE2 and prostaglandin H synthase‐2 (PGHS‐2) protein. This effect of LPS was suppressed by increasing amounts of IL‐4. Expression of LPS‐induced PGHS‐2 mRNA was also inhibited by IL‐4. In addition, IL‐4 inhibited expression of CD14, which is a receptor for LPS bound to the LPS‐binding protein (LBP). We conclude that IL‐4 down‐regulates LPS‐induced release of PGE2, by reducing expression of the enzyme, PGHS‐2. One potential mechanism for this effect of IL‐4 is a reduced expression of CD14, which is the LPS‐LBP receptor. © 1995 Wiley‐Liss Inc.
Plasminogen activators augment endothelial cell organization in vitro by two distinct pathways
Authors: Schnaper H. William , Barnathan Elliot S. , Mazar Andrew , Maheshwari Shailendra , Ellis Stefanie , Cortez Shirley L. , Baricos William H. , Kleinman Hynda K. 
Published: February 2005

AbstractEndothelial cell differentiation into capillary structures is a complex process that requires the concerted effects of several extracellular matrix proteases, including plasminogen activators. Here, the role of tissue‐type plasminogen activator (tPA) and urokinase‐type plasminogen activator (uPA) was evaluated in an in vitro model of endothelial morphogenesis involving organization of human umbilical vein endothelial cells into tubular structures when they are cultured on the basement membrane preparation, Matrigel. Both uPA and tPA were detected in HUVEC cultures on Matrigel, and inhibitors of plasminogen activators or of serine proteases decreased the extent of the tube network formed by the cells. The decrease resulting from serine protease inhibitors was additive to that from matrix metalloproteinase inhibitors which have previously been shown to decrease tube formation in this model, suggesting that the two classes of proteases modulate tube formation by distinct mechanisms. Plasminogen activator inhibitor (PAl)‐1 decreased tube formation by 50% when added up to 4.5 h after the initiation of an 18 h assay and caused 25% inhibition when added 9.5 h after culture initiation, indicating that the effects of plasminogen activators are not limited to an early event in the differentiation process. Steady‐state expression of mRNA for uPA increased during the first several hours of culture on Matrigel, further supporting a role for PA activity throughout the process of tube formation. These findings suggested that PAs may affect multiple events during tube‐forming activity. A fucosylated peptide comprising the amino‐terminal domain of uPA that binds to the uPA receptor (uPAR) but lacking proteolytic activity enhanced tube formation. In contrast, a defucosylated form of the same peptide had no effect. Since fucosylation of this fragment has been shown to be essential in other models of cell stimulation by uPA‐uPAR interaction, these data support the hypothesis that uPA enhances endothelial morphogenesis both through proteolytic activity and via uPAR occupancy. Plasminogen activators could facilitate angiogenesis in vivo. © 1995 Wiley‐Liss Inc.
Stimulation of protein and DNA synthesis in mouse C2C12 satellite cells: Evidence for phospholipase d‐dependent and ‐independent pathways
Authors: Morrison Kenneth S. , Mackie Steven C. , Palmer Robert M. , Thompson Michael G. 
Published: February 2005

AbstractIn C2C12 myoblasts, 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) stimulated a phospholipase D (PLD) to degrade phosphatidylcholine (PC) as measured by the release of choline and an increase in the formation of phosphatidic acid (PA) (or phosphatidylbutanol [PtdBuOH] in the presence of 0.5% butanol). Exogenous PLD also stimulated choline release, PA and PtdBuOH formation. The protein kinase C (PKC) inhibitor, Ro‐31‐8220, and PKC downregulation significantly inhibited the effects of TPA but Ro‐31‐8220 had no effect on PLD action. Neither basic Fibroblast Growth Factor (bFGF) or Epidermal Growth Factor (EGF) increased PLD activity. All agonists stimulated protein synthesis during both a 90 min and a 6 hr incubation and increased RNA accretion after 6 hr. The response at 90 min was not inhibited by the transcription inhibitor, actinomycin D. Ro‐31‐8220 and PKC downregulation significantly inhibited all the effects of TPA. In contrast, Ro‐31‐8220 significantly inhibited the increase in RNA accretion elicited by PLD but had no effect on the ability of agonists other than TPA to enhance protein synthesis. All agonists also stimulated thymidine incorporation into DNA. The effects of EGF, bFGF, and PLD were rapid and transient whereas that of TPA was delayed and sustained. Ro‐31‐8220 and PKC downregulation significantly inhibited the response due to TPA. Furthermore, Ro‐31‐8220 also significantly inhibited the effects elicited by EGF and PLD but not that induced by bFGF. In differentiated myotubes, TPA and PLD, but not bFGF or EGF, again stimulated choline release and PtdBuOH formation. However, all agents failed to stimulate protein synthesis and RNA accretion. The data demonstrate the presence in C2C12 myoblasts, but not differentiated myotubes, of both a PLD‐dependent and PLD‐independent pathway(s) leading to the stimulation of protein synthesis, RNA accretion, and DNA synthesis. © 1995 Wiley‐Liss, Inc.
Melanoma cell spreading on fibronectin induced by 12(S)‐HETE involves both protein kinase C‐ and protein tyrosine kinase‐dependent focal adhesion formation and tyrosine phosphorylation of focal adhesion kinase (pp125FAK)
Authors: Tang Dean G. , Tarrien Mark , Dobrzynski Philip , Honn Kenneth V. 
Published: February 2005

AbstractOur previous work demonstrated that 12(S)‐HETE, a lipoxygenase metabolite of arachidonic acid, promoted B16 amelanotic melanoma (B16a) cell spreading on fibronectin. In the current study, we investigated the biochemical mechanisms of the 12(S)‐HETE induced response. 12(S)‐HETE treatment resulted in a time‐dependent increase in B16a cell spreading on fibronectin, which was blocked by either calphostin C or by genistein but not by H8. Two hours following cell plating, both spontaneous and 12(S)‐HETE promoted cell spreading reached their maximum (nearly 100%). Spontaneous cell spreading was inhibited by the select 12‐lipoxygenase inhibitor, BHPP, whose inhibitory effect could be overcome by increasing doses of exogenous 12(S)‐HETE. 12(S)‐HETE‐treated B16a cells plated on either fibronectin or cultured on their own extracellular matrix demonstrated increased vinculin and tyrosine‐phosphorylated proteins, which were colocalized at focal adhesions. The increase in vinculin localization to focal adhesions appeared to be a post‐transcriptional process, since 12(S)‐HETE treatment did not alter the overall protein level of vinculin in tumor cells, but resulted in a specific enrichment of vinculin to focal adhesions. Pretreatment of B16a cells with either calphostin C or genistein abolished 12(S)‐HETE‐increased formation of vinculin‐and phosphotyrosine‐containing focal adhesions. Immunoblotting using antiphosphotyrosine antibody 4G10 demonstrated, following 12(S)‐HETE stimulation, an increased tyrosine phosphorylation of several proteins in focal adhesions; most prominently, a ∼︁ 155 kd protein, a 120‐130 kd protein cluster, a 76 kd protein, and a 42/44 kd complex. Immunoprecipitation with anti‐phosphotyrosine antibody PY20 revealed increased tyrosine phosphorylation, post 12(S)‐HETE stimulation, of proteins migrating at 120, 76, and 42/44 kd, of which the 120 kd protein co‐migrated with pp125FAK. Immunoprecipitation with anti‐FAK antibody BC‐3 followed by immunoblotting with anti‐phosphotyrosine antibody RC20H demonstrated a time‐dependent hyperphosphorylation of pp125FAK. The present study suggests that 12(S)‐HETE promoted melanoma cell spreading on fibronectin involves tyrosine phosphorylation of pp125FAK and protein kinase C‐ and tyrosine kinase‐dependent focal adhesion formation. © 1995 Wiley‐Liss, Inc.
Hepatocyte growth factor activates quiescent skeletal muscle satellite cells in vitro
Authors: Allen Ronald E. , Sheehan Shannon M. , Taylor Richard G. , Kendall Teresa L. , Rice Glenna M. 
Published: February 2005

AbstractThe effect of hepatocyte growth factor (HGF) on the activation of quiescent rat skeletal muscle satellite cells was evaluated in vitro. Satellite cells from 9‐month‐old adult rats are quiescent in vivo and when cultured, display a protracted lag phase prior to division that is not present in satellite cells from neonatal or regenerating muscle. Under normal growth conditions, satellite cells divide for the first time between 42 and 60 hr. Hepatocyte growth factor increased proliferation in a dose‐dependent fashion prior to 48 hr with half‐maximal stimulation at approximately 3 ng/ml; in addition, heparin enhanced this activity. The time course of cyclin‐D1 and proliferating cell nuclear antigen (PCNA) expression was accelerated in HGF‐treated satellite cells, indicating that cells entered the cell cycle earlier. No significant effects on muscle‐derived fibroblast proliferation was observed. The signalling receptor for HGF is the product of the c‐met protooncogene, and rtPCR analysis of satellite cells 0–72 hr in culture demonstrated the presence of this message throughout this time period. The presence of c‐met in quiescent satellite cells, the ability of HGF to stimulate precocious entry into the cell cycle, and the previously described localization of HGF message in regenerating muscle (Jennische et al., 1993) indicate that HGF could act as an activator of quiescent satellite cells in vivo. © 1995 Wiley‐Liss, Inc.
Changes in expression and subcellular localization of annexin IV in rabbit kidney proximal tubule cells during primary culture
Authors: Massey‐Harroche D. , Traverso V. , Mayran N. , Francou V. , Vandewalle A. , Maroux S. 
Published: February 2005

AbstractIn the present study, we investigated the polarized expression of annexin IV at various stages in the growth of rabbit kidney proximal tubule cells (PTC) in primary cultures. The results of immunoblotting analysis and indirect immunofluorescence studies using a specific anti‐annexin IV monoclonal antibody, indicated that annexin IV is expressed in proximal tubule cultured cells, although it was not detected in the proximal tubules present in frozen sections of kidney cortex and freshly isolated proximal tubule cells. In either non‐confluent or confluent cells which remained attached to the collagen‐coated support, annexin IV was mainly concentrated around the nucleus, whereas in PTC forming the monolayer of domes, it was restricted to the basolateral membrane domain. This basolateral localization was identical to that observed in other polarized epithelial cell types such as enterocytes. When the domes burst, the cells returned to the collagen‐coated support and the annexin IV was again localized around the nuclei. The fact that the change of localization was very rapid suggested the existence of a considerable difference between the differentiation states of dome forming and adherent confluent cells. Moreover, a transient association of annexin IV with the basal body of apically located cilia also seemed to be correlated with a particular polarization state and/or differentiation states of adherent cultured cells, corresponding to the beginning of the polarized expression of aminopeptidase N, a hydrolase located in the apical brush border membrane, and to the falling of cells onto the support, subsequent to the bursting of the domes. In conclusion, these results provide evidence that annexin IV may constitute a new marker of the basolateral membrane domain of polarized epithelial renal cells in primary cultures. © 1995 Wiley‐Liss, Inc.
Characterization of transforming growth factor‐β growth regulatory effects and receptors on bovine mammary cells
Authors: Woodward T. L. , Dumont N. , O'Connor‐McCourt M. , Turner J. D. , Philip A. 
Published: February 2005

AbstractTransforming growth factor‐β (TGF‐β) has been shown to inhibit mammary morphogenesis, growth, and differentiation in murine studies. We have characterized TGF‐β receptors and their autoregulation, and the growth response to TGF‐β1 and TGF‐β2 in cultured bovine mammary epithelium (MAC‐T) and fibroblasts. Affinity labelling studies revealed that fibroblast and epithelial cells contained type I, II, and III (betaglycan) receptors, with the type III receptor being the predominant binding component. On both fibroblasts and epithelial cells, TGF‐β1 and TGF‐β2 had equal binding affinities for the type I and II receptors, but TGF‐β2 had a higher affinity for the type III receptor. Also, preincubation of MAC‐T cells with 50 pM TGF‐β1 or TGF‐β2 markedly downregulated TGF‐β receptors. Proliferative response was measured using both total DNA and 3H‐thymidine incorporation. Both TGF‐β isoforms were effective in inhibiting proliferation of MAC‐T cells and fibroblasts. Inhibition of proliferation was not altered following immortalization of fibroblasts with SV‐40 Large‐T‐antigen (LT), even when the cells acquired a transformed phenotype. Inhibition of proliferation was not a result of cytotoxicity, as TGF‐β at concentrations 1,000‐fold higher than ED50 levels did not increase cell death. Moreover, the inhibition was reversible as shown by return of cellular proliferation to control levels following TGF‐β removal. Although growth inhibition was not transient as culture of MAC‐T cells in TGF‐β resulted in sustained inhibition of proliferation for at least 144 h. © 1995 Wiley‐Liss, Inc.