bluuebunny/crossref_metadata_2025_split huggingface dataset:
Endothelin‐1 acts as an autocrine growth factor for normal human keratinocytes
Authors: Tsuboi Ryoji , Sato Chiyo , Shi Chong‐Ming , Nakamura Tsukasa , Sakurai Takeshi , Ogawa Hideoki 
Published: February 2005

AbstractEndothelin‐1 (ET‐1) is an endothelium‐derived 21 amino acid vasoconstrictor peptide possessing two intrachain disulfide bridges. Recently it has become evident that isoforms of ET (ET‐1, ‐2, and ‐3) have a wide range of pharmacological effects in various tissues and act as autocrine/paracrine factors. We demonstrate here that ET‐1 is secreted from normal human keratinocytes and may work as an autocrine growth factor through a specific receptor. In this study, human foreskin keratinocytes were cultured in serum‐free MCDB 153 medium. Cell growth and [3H] thymidine incorporation in low and high Ca++ concentration media was stimulated by ET‐1, ‐2, and ‐3 with similar potencies. The strongest response was observed at 10 nM ETs, whereas stimulatory activity was reduced at 100 nM. ETs suppressed keratinocyte differentiation as measured by reactivity with involucrin antibody. Plasminogen activator activity (mainly urokinase) in the medium was also stimulated by the addition of 10 nM ETs. ET‐1–like immunoreactivity measured by radioimmunoassay was 1.4 fmol/day/106 cells in non‐treated condition medium. Among the various cytokines, tumor necrosis factor–α (TNF‐α), interleukin‐1α, and transforming growth factor–β stimulated ET‐1 secretion in a dose‐dependent manner. The strongest response (ten‐fold) was observed upon the addition of 10 ng/ml TNF‐α. Scatchard plot analysis of [125I] ET‐1 binding to keratinocytes revealed the presence of a single class of high affinity receptors (KD 50 pM, 9 x 103 sites/cell). Binding was competitively inhibited by the addition of unlabeled ET‐1 and ‐2 with similar affinities and by ET‐3 with weaker affinity. ET‐1 mRNA expression in keratinocytes was detected by reverse transcription‐polymerase chain reaction and was increased by treatment with 10 ng/ml TNF‐α. These results suggest that ET‐1 acts as an autocrine growth factor for keratinocytes through a specific receptor. © 1994 wiley‐Liss, Inc.
Retinoic acid enhances adhesiveness, laminin and integrin β1 synthesis, and retinoic acid receptor expression in F9 teratocarcinoma cells
Authors: Ross Sharon A. , Ahrens Richard A. , De Luca Luigi M. 
Published: February 2005

AbstractThe teratocarcinoma‐derived F9 cells respond to retinoic acid (RA) and RA plus dibutyrylcyclic adenosine monophosphate (dcAMP) by differentiating into endoderm cells, which elaborate a laminin and type IV collagen‐rich matrix. We found that the induction of differentiation is accompanied by a small but consistent increase in cell adhesiveness to a variety of substrates, including laminin. Therefore we investigated biochemical mechanisms involved in this phenomenon. Endoglycosidase treatment showed that laminin contains complex and hybrid oligosaccharide structures. RA enhanced general biosynthesis of laminin without a specific increase in galactose incorporation: this sugar was mainly in polylactosamine structures in the A chain of laminin and as terminal galactose α 1,3 galactose in the B chain. Laminin receptor analysis showed that RA decreased laminin binding protein‐37 (LBP‐37) but increased the amount of β1 integrin, suggesting the involvement of β1 integrin in the attachment process. Northern blot analysis showed increased expression of retinoid receptors within hours of RA exposure. These studies demonstrate that RA increases cell to substrate interactions by increasing the biosynthesis of laminin and β1 integrin. These effects are most likely subsequent to the RA‐induced biosynthesis of the retinoid receptors. © 1994 wiley‐Liss, Inc.
Increased expression of heme oxygenase‐1 in human retinal pigment epithelial cells by transforming growth factor‐β
Authors: Krishnan Kutty R. , Nagineni Chandrasekharam N. , Kutty Geetha , Hooks John J. , Chader Gerald J. , Wiggert Barbara 
Published: February 2005

AbstractAntibodies specific for heme oxygenase‐1 (HO‐1) were produced in rabbits, using the multiple antigen peptide (MAP) technique, and were employed to investigate the ability of transforming growth factor‐β1 (TGF‐β1) to induce the HO‐1 protein in cultured human retinal pigment epithelial (RPE) cells. Western blot analyses showed that the cytokine induced HO‐1 in these cells in a time‐ and dose‐dependent manner. TGF‐β1 also increased the mRNA for HO‐1 in treated cells prior to the increase in HO‐1 protein. The induction was effectively blocked by a neutralizing antibody preparation against TGF‐β1. When tested under similar conditions, other growth factors such as basic fibroblast growth factor‐I, plateletderived growth factor, insulin‐like growth factor, transforming growth factor‐α, and epidermal growth factor did not show appreciable induction of HO‐1. Lipopolysaccharide, tumor necrosis factor‐α, and interferon‐γ were also not inducers, although TGF‐β2 effectively induced HO‐1. Heavy metal ions and thiol reagents were also highly potent inducers of HO‐1 in human RPE cells. The induction of HO‐1 by TGF‐β1 was also observed in bovine choroid fibroblasts, but not in HELA, HEL or bovine corneal fibroblasts. Our results demonstrate for the first time that HO‐1 can be induced by an important cytokine, TGF‐β1, causing an increase in the expression of both HO‐1 message and protein in specific neuroepithelial and fibroblast cells. © 1994 wiley‐Liss, Inc.This article is a US Government work and, as such, is in the public domain in the United States of America.
Peripheral and central vascular smooth muscle cells from rat lung exhibit different cytoskeletal protein profiles but similar growth factor requirements
Authors: Davies Paul , Patton Wayne 
Published: February 2005

AbstractIn pulmonary vascular remodelling, the lining smooth muscle cells undergo various forms of growth involving cellular hypertrophy and hyperplasia. Differences in the growth pattern between central and peripheral regions suggested that cells from both should be obtained when investigating the cellular basis for the remodelling. Accordingly, we have obtained two smooth muscle cell types in culture: a cell from the central pulmonary artery (CC) and a cell morphologically similar to a pericyte (PC), from the periphery of the lung. Both cell types gave positive immunostaining for α‐smooth muscle isoactin. In vivo, the α‐isoactin was immunolocalized in the extracapillary vasculature. Quantitative two‐dimensional gel electrophoresis of cell extracts showed that PC express more vimentin and gelsolin than CC. Despite the differences between PC and CC in the expression of cytoskeletal proteins, their response to growth factors was similar. Both cell types increased DNA synthesis when stimulated by exogenous PDGF‐AB. This occurred in the absence of exogenous progression factors, but depended on a post‐competence, suramin‐sensitive mechanism that probably represents an autocrine progression factor. The cells were also stimulated by IGF‐1 alone, in the absence of exogenous competence factors. At an IGF‐1 concentration of 1 ng/ml, this response appeared specific for the IGF‐1 receptor and was sensitive to pretreatment with pertussis toxin, thus implicating a role for a G protein. © 1994 wiley‐Liss, Inc.
Critical role for intracellular calcium in tight junction biogenesis
Authors: Stuart Robert O. , Sun Adam , Panichas Michael , Hebert Steven C. , Brenner Barry M. , Nigam Sanjay K. 
Published: February 2005

AbstractUsing the Madin Darby canine kidney (MDCK) cell “calcium switch,” we have previously demonstrated that, as MDCK cells establish contact and ultimately form tight junctions, there are marked global and localized changes in intracellular calcium at the sites of cell‐cell contact (Nigam et al., 1992, Proc. Natl. Acad. Sci. USA, 89:6162–6166). We have now examined whether intracellular Ca++ is critical to the biogenesis of tight junctions by chelating this ion and monitoring the formation of junctions by electrical, immunocytochemical, and biochemical criteria. Intracellular Ca++ was chelated with the cell‐permeant chelators, dimethyl‐BAPTA‐AM and BAPTA‐AM. By digital imaging of fura‐2 loaded cells, it was demonstrated that both agents efficiently chelated Ca++ during the “switch” in a dose‐dependent manner which paralleled their respective in vitro affinities for Ca++. Chelation of Ca++ during the switch markedly attenuated the development of transepithelial electrical resistance (TER), a measure of tight junction assembly. Immunofluorescent staining of the tight junctional protein, zonula occludens‐1 (ZO‐1), revealed that chelation of intracellular Ca++ retarded the movement of ZO‐1 from intracellular sites to the plasma membrane during the switch. During the development of tight junctions, a fraction of ZO‐1 redistributed from the Triton X‐100 soluble to the Triton X‐100 insoluble pool; chelation of Ca++ during the induction of cell–cell contact prevented this stabilization into the Triton X‐100 insoluble fraction. Taken together, these data indicate an important role for intracellular Ca++ in tight junction biogenesis and suggest a specific role for calcium in the early sorting and possible cytoskeletal association of tight junction components. © 1994 wiley‐Liss, Inc.
Responses of salivary acinar cells to intracellular alkalinization
Authors: Seagrave Jc. , Curry M. , Martinez J. R. 
Published: February 2005

AbstractResponses of rat submandibular acini to intracellular alkalinization were investigated. Intracellular alkalinization was induced by addition of NH4Cl or methylamines, or by prepulse with Na butyrate. Only partial recovery occurred following Na butyrate prepulse or methylated amine addition, but full recovery was observed following addition of NH4Cl. The latter recovery was DIDS and dimethylamiloride‐insensitive but was inhibited by bumetanide or high [K+] and stimulated in Na+ free buffer and by ouabain. Acetylcholine stimulated recovery from NH4Cl‐ or Na butyrate pre‐pulse‐induced alkalinization and reduced the extent of alkalinization induced by methylated amines. Acetylcholine‐stimulated recovery from NH4Cl‐induced alkalinization was mimicked by substance P or ionomycin and was partially Ca2+‐dependent. This stimulated recovery was bumetanide‐insensitive but was partially sensitive to charybdotoxin. Taken together, these data indicate that in unstimulated cells, recovery from alkalinization induced by NH4Cl occurs by bumetanide‐sensitive transport of the NH4+ ion, that DIDS‐inhibitable anion transport contributes little to this recovery, and that acetylcholine and other Ca2+‐elevating agents accelerate recovery from NH4Cl‐induced alkaline challenge by a mechanism insensitive to bumetanide, DIDS, ouabain, and dimethylamiloride but sensitive to extracellular Ca2+ and to charybdotoxin. Partial recovery from alkaline challenge can also occur in the absence of NH4+ ions, and acetylcholine also stimulates this mode of recovery. Together, these data suggest that these cells have little intrinsic ability to recover from intracellular alkalinization and that the NH4+ ion may be a surrogate for K+ in at least two ion transport pathways. © 1994 wiley‐Liss, Inc.
Characterization of 3′‐azido‐3′‐deoxythymidine inhibition of ricin and pseudomonas exotoxin A toxicity in CHO and vero cells
Authors: Wellner Robert B. , Pless Dorothy D. , Thompson William L. 
Published: February 2005

AbstractRicin (RIC), modeccin (MOD), Pseudomonas exotoxin A (PE), and diphtheria toxin (DT) are protein toxins that enter cells by receptor‐mediated endocytosis. After intracellular transport and membrane translocation to the cytosol, these toxins inhibit protein synthesis by enzymatically removing a specific adenine residue from ribosomal RNA (RIC, MOD), or by ADP‐ribosylation of elongation factor‐2 (PE, DT). Recently, Thompson and Pace (1992) reported that AZT (3′‐azido‐3′‐deoxythymidine) inhibited RIC toxicity in Vero cells, and this inhibition was not due to a block of RIC enzymatic activity. This paper extends these findings and examines the effects of AZT treatment on the toxicities of other protein toxins in Chinese hamster ovary (CHO) and Vero cell lines. AZT treatment did not significantly alter the toxicity of DT or MOD in either cell line, but it markedly reduced RIC and PE toxicity in both cell lines. The ID50 values (concentration of toxin required to inhibit protein synthesis by 50%) for RIC and PE in CHO cells increased approximately 6.5‐ and 12.5‐fold, respectively; while in Vero cells the ID50 values increased ca. 8.5‐and 4.5‐fold, respectively. Results of further studies revealed differences in the mechanisms by which AZT inhibited RIC and PE toxicity. Results of cell‐free translation indicated that, unlike its effects on RIC, AZT blocked the ability of PE to perform its enzymatic activity. As AZT did not block RIC enzymatic activity, we examined the effects of AZT on earlier steps in the RIC intoxication process. AZT treatment did not inhibit cell‐surface binding or internalization of [125I]‐RIC. Results of kinetic studies showed that when AZT was incubated with cells at the time of RIC exposure, it caused no major change in the lag phase, during which RIC reaches the site of translocation. However, it clearly reduced the subsequent first‐order reduction in the rate of protein synthesis, suggesting an effect on translocation. Monensin (an ionophore that perturbs intracellular trafficking and increases the toxicities of RIC and PE) reduced AZT protection against both toxins. Nocodazole and colchicine (agents that disrupt microtubules and some routes of intracellular trafficking) reduced the ability of AZT to inhibit RIC, but not PE, toxicity. In summary, our results suggest that (1) AZT acts within the cytosol to inhibit (directly or indirectly) the enzymatic action of PE, and (2) the AZT inhibition of RIC cytotoxicity does not involve perturbations of RIC cell‐surface binding, internalization, or enzymatic activity but might result from an alteration in RIC translocation. © 1994 wiley‐Liss, Inc.
Thapsigargin‐induced calcium entry in FRTL‐5 cells: Possible dependence on phospholipase A2 activation
Authors: Törnquist Kid , Ekokoski Elina , Forss Lisbeth 
Published: February 2005

AbstractStimulating rat thyroid FRTL‐5 cells with agonists that activate the inositol phosphate cascade results in the release of sequestered calcium and influx of extracellular calcium. In addition, phospholipase A2 (PLA2) is activated. Since PLA2 is a calcium‐dependent enzyme we wanted to investigate the interrelationships between PLA2 activity and the entry of calcium. Stimulating 3H‐arachidonic acid (3H‐AA)‐labelled cells with thapsigargin resulted in a substantial release of 3H‐AA. This release was totally abolished in a calcium‐free buffer. Pretreatment of Fura 2 loaded cells with 4‐bromophenacyl bromide, an inhibitor of PLA2 activity, decreased the thapsigargin‐induced entry of calcium, suggesting a role for PLA2 in the regulation of calcium entry. In cells treated with nordihydroguaiaretic acid (NDGA), clotramizole, or econazole, compounds with lipoxygenase and cytochrome P‐450 inhibitory actions, the thapsigargin‐induced entry of calcium was decreased in a dose‐dependent manner. However, treatment of the cells with indomethacin, a cyclooxygenase inhibitor, had no effect on the thapsigargin‐induced calcium entry. We also showed that stimulation of the cells with arachidonic acid released sequestered calcium, apparently from the same intracellular pool as did thapsigargin. The results suggested that the calcium‐induced PLA2 activation and the metabolism of the produced arachidonic acid by a noncyclooxygenase pathway may be of importance in maintaining calcium entry after releasing sequestered Ca2+ in FRTL‐5 cells. © 1994 Wiley‐Liss, Inc.
Recovery of nuclear matrix ultrastructure of interphase CHO cells after heat shock
Authors: Wachsberger Phyllis R. , Coss Ronald A. 
Published: February 2005

AbstractHeat shock induces changes in G1 CHO cell nuclear matrix (NM) ultrastructure that may be related to heat‐induced nuclear protein accumulation (Wachsberger and Coss, 1993, J. Cell. Physiol., 155:615–634). The present study quantitates recovery of alterations in NM fine structure in CHO cells heated in G1 and compares structural recovery with recovery of bulk RNA synthesis and surviving fraction (SF). Morphology of NM preparations was quantified 30 min and 20 hr following heat shock by 1) measurement of the number of fiber anastomosing points per unit area per NM, and 2) measurement of the length of fibers between points of anastomoses within individual NMs. Architectural recovery was nearly complete within 20 hr in cells heated at 43°C or 45°C with SFs of 0.27 or greater. No recovery of architecture was observed in heated cells with SFs of approximately 0.01 or less. The residual damage to NMs was associated with RNA‐containing fiber networks as determined by means of RNase gold labeling. Recovery from inhibition of RNA synthesis following heat shock was related to recovery of NM architecture. It is suggested that 1) repair of NM architecture does not require full recovery of bulk RNA synthesis, and 2) partial or complete irreversible collapse of the NM may be responsible, in part, for heat‐induced, interphase cell death. © 1994 Wiley‐Liss, Inc.
SR 25989 inhibits healing of a mechanical wound of confluent human saphenous vein endothelial cells which is modulated by standard heparin and growth factors
Authors: Klein‐Soyer C. , Cazenave J.‐P. , Herbert J.‐M. , Maffrand J.‐P. 
Published: February 2005

AbstractThe thienopyridine, ticlopidine, a potent platelet antiaggregating agent and SR 25989, an esterified derivative of ticlopidine, devoid of antiplatelet activity, were tested in an in vitro model of healing of a mechanical wound in confluent endothelium. This model allows exploration of substances involved in wound healing and angiogenesis. These two compounds inhibited both cell proliferation and cell migration during lesion repair in a dose‐dependent manner (18–150 μM), SR 25989 being twice as active as ticlopidine. Its effect was not inhibited by acidic or basic fibroblast growth factor or by platelet derived growth factor. In contrast, it exerted a conjugated inhibition with standard heparin and was able to totally reverse the healing increase induced by a mixture of acidic fibroblast growth factor and heparin. The mechanism of action of SR 25989 is not yet elucidated, but it does not seem to involve competition with fibroblast growth factors since these substances were not able to alter their binding to receptors on the endothelial cell surface. SR 25989 therefore appears as a promising new candidate for inhibition of angiogenesis. © 1994 Wiley‐Liss, Inc.
Role for autocrine TGF‐β1 in regulating differentiation of a human leukemic cell line toward osteoclast‐like cells
Authors: Fiorelli G. , Gori F. , Masi L. , Frediani U. , Brandi M. L. , Tanini A. , Bernabei P. A. , Ballock R. T. , Wakefield L. M. , Sporn M. B. 
Published: February 2005

AbstractIncreasing evidence suggests that transforming growth factor‐β (TGF‐β) is involved in bone formation during remodeling. Using a recently cloned human leukemic cell line (FLG 29.1 cells) we demonstrate that these cells synthesize and secrete TGF‐β1 and that exogenous or autocrine TGF‐β1 can induce the same features of osteoclastic‐like cells, exerting its effects through the binding to TGF‐β specific receptors. Scatchard analysis of 125I‐labeled TGF‐β1 to FLG 29.1 cells revealed the presence of a single high affinity binding site with a Kd value of ∼25 pM and a binding capacity of ∼900 sites/cell. Affinity labeling experiments showed that FLG 29.1 cells express type I and type II TGF‐β receptors. Stimulation of FLG 29.1 cells with low TGF‐β1 doses reduced cell proliferation and increased cell adhesion and tartrate resistant acid phosphatase (TRAcP) activity. Pretreatment of FLG 29.1 cells with TGF‐β1 caused a significant and dose‐dependent response to calcitonin. Northern blot of total mRNA and analysis of the conditioned media (CM) showed that TGF‐β1 was synthesized by FLG 29.1 cells. TPA treatment, which induces partial differentiation of these cells, markedly increased TGF‐β1 mRNA expression and growth factor release. The majority of TGF‐β1 secreted by TPA‐treated cells was in its latent form. However, anti‐TGF‐β antibodies inhibited TGF‐β1 and TPA‐induced growth inhibition, calcitonin responsiveness, and TRAcP activity, suggesting that the TPA effect is mediated in part by autocrine TGF‐β1 and indicating that the cells can activate and respond to the TGF‐β that they secrete. These findings support a potential autocrine role for TGF‐β1 in osteoclast differentiation. © 1994 Wiley‐Liss, Inc.
Mouse fibroblasts defective in thrombin mitogenesis possess functional proteolytically activated receptor for thrombin: Requirement for a second signaling pathway
Authors: Kim Dennis W. , Wang Fang , Ramakrishnan Shyam , Scott David L. , Hensler Theresa M. , Thompson William C. , Carney Darrell H. 
Published: February 2005

AbstractThrombin mitogenesis in fibroblasts requires two distinguishable subsets of signals; one generated by proteolytic cleavage, the other by high‐affinity cell surface binding. Characterizing two closely related mouse embryo (ME) cell lines with high numbers of thrombin binding sites, we found that one line, B11‐A, responds mitogenically to thrombin, epidermal growth factor (EGF), and serum, whereas the B11‐B cell line is responsive to EGF and serum, but not to thrombin. The B11‐B defect responsible for loss of thrombin responsiveness is not due to differences in the number of high‐affinity binding sites, the affinity of thrombin binding to these sites, or to differences in cell surface expression of proteolytically activated receptors for thrombin (PART). The defect is also not associated with an inability of thrombin to activate PART since thrombin stimulates the cleavage‐dependent induction of the proto‐oncogene c‐fos in both B11‐A and B11‐B cells. Various combinations of thrombin, synthetic thrombin receptor peptide, TRP‐14 (SFFLRNPGENTFEL), platelet‐derived growth factor (PDGF), and phorbol 12‐myristate 13‐acetate (PMA) were used to better define the defect in thrombin‐mediated mitogenesis in B11‐B cells. Direct activation of protein kinase C with PMA in combination with thrombin did not overcome B11‐B nonresponsiveness. However, mitogenic responsiveness was regained in B11‐B cells by simultaneous addition of PDGF and either thrombin or TRP‐14. Therefore, the B11‐B defect may involve a set of signals initiated by nonproteolytic thrombin interactions distinct from those initiated by PART, but related to the downstream signals initiated by the tyrosine kinase‐associated growth factors, EGF and PDGF. © 1994 Wiley‐Liss, Inc.
Apoptotic cell death induced by serum and its prevention by thiols
Authors: Kurita Takeshi , Namiki Hideo 
Published: February 2005

AbstractWe recently reported that serum contains low molecular weight factors that inhibit growth and cause cell death in vitro. The present study focused on identifying components of basal media that counteract the toxic effects of serum. Amino acids L‐cyst(e)ine and L‐tryptophan were found to prevent serum‐induced cell death of TIG‐1 human fetal lung fibroblasts and other cell types. In addition to L‐cysteine, other thiol‐bearing and dithiol‐cleaving compounds showed a similar ability to rescue the cells. Various inhibitors of protein or RNA synthesis also prevented the cell death. By contrast, nonthiol‐containing reducing agents and super oxide dismutase (SOD), an active oxygen‐eliminating enzyme, were ineffective. Thiol compounds appeared to exert a supportive level in TIG‐1 cells cultured in FBS, whereas protein synthesis inhibitors did not alter the reduced intracellular thiol content. Fragmentation of DNA occurred prior to the plasma membrane breakdown of dying cells. Taken together, these data suggest that serum‐induced cell death represents a form of apoptosis in which molecules containing thiol groups are active participants. © 1994 Wiley‐Liss, Inc.