bluuebunny/crossref_metadata_2025_split huggingface dataset:
CCAAT/enhancer binding protein δ (C/EBPδ) regulation and expression in human mammary epithelial cells: II. Analysis of activating signal transduction pathways, transcriptional, post‐transcriptional, and post‐translational control
Authors: Sivko G.S. , Sanford D.C. , Dearth L.D. , Tang D. , DeWille J.W. 
Published: September 2004

AbstractCCAAT/enhancer binding protein δ (C/EBPδ) plays a key role in mammary epithelial cell G0 growth arrest. C/EBPδ gene expression is down‐regulated in rodent mammary tumorigenesis and in human breast cancer, suggesting that “loss of function” alterations in C/EBPδ gene expression are common in mammary gland malignancies. The goal of this study was to systematically investigate the mechanisms controlling C/EBPδ gene expression in MCF‐10A and MCF‐12A human mammary epithelial cell lines. The results demonstrate that G0 growth arrest conditions (i.e., serum and growth factor withdrawal or Oncostatin M (OSM) treatment) result in the activation of JAK1, JAK2, and Tyk 2, members of the Janus kinase family of non‐receptor tyrosine kinases, in MCF‐10A and MCF‐12A cells. Growth arrest or OSM treatment also specifically increases activated (phosphorylated) signal transduction and activators of transcription 3 (STAT3) levels, demonstrating that STAT3, not STAT1 or STAT5, is the downstream target of the activated Janus kinases in MCF‐10A and MCF‐12A cells. Whole cell lysates from G0 growth arrested (GA) and OSM‐treated MCF‐12A cells exhibit increased acute phase response element (APRE) binding compared to lysates from growing (GR) MCF‐12A cells. Transient transfection using C/EBPδ promoter‐luciferase constructs demonstrated that the APRE (STAT3) consensus binding site is essential for growth arrest or OSM induction of the C/EBPδ promoter. Mutation of the C/EBPδ promoter STAT3 site or expression of a dominant negative STAT3 construct (STAT3β) reduces C/EBPδ promoter activity in response to growth arrest conditions. The human C/EBPδ promoter also contains an Sp1 site at −61 bp (relative to the transcriptional start site) which is required for basal transcriptional activation. Mutation or deletion of the Sp1 site decreases promoter activity in response to growth arrest conditions. Treatment with the transcriptional inhibitor actinomycin D demonstrated that the C/EBPδ mRNA exhibits a relatively short half‐life (∼40 min). Similarly, treatment with the translational inhibitor anisomysin demonstrated that the C/EBPδ protein half‐life was also relatively short (∼160 min). These results indicate that the human C/EBPδ gene is controlled at multiple levels, consistent with a role for C/EBPδ in cell cycle control and/or cell fate determination. © 2004 Wiley‐Liss, Inc.
Molecular pathways involved in the anti‐apoptotic effect of 1,25‐dihydroxyvitamin D3 in primary human keratinocytes
Authors: De Haes Petra , Garmyn Marjan , Carmeliet Geert , Degreef Hugo , Vantieghem Katleen , Bouillon Roger , Segaert Siegfried 
Published: September 2004

AbstractWe previously reported that 1,25‐dihydroxyvitamin D3 [1,25(OH)2D3] protects primary human keratinocytes against ultraviolet (UV)B‐induced apoptosis. Here, we confirmed the anti‐apoptotic effect of 1,25(OH)2D3 in keratinocytes, using cisplatin and doxorubicin as apoptotic triggers. We further showed that 1,25(OH)2D3 activates two survival pathways in keratinocytes: the MEK/extracellular signal regulated kinase (ERK) and the phosphatidylinositol 3‐kinase (PI‐3K)/Akt pathway. Activation of ERK and Akt by 1,25(OH)2D3 was transient, required a minimal dose of 10−9 M and could be blocked by actinomycin D and cycloheximide. Moreover, inhibition of Akt or ERK activity with respectively a PI‐3K inhibitor (LY294002) or MEK inhibitors (PD98059, UO126), partially or totally suppressed the anti‐apoptotic capacity of 1,25(OH)2D3. Finally, 1,25(OH)2D3 changed the expression of different apoptosis regulators belonging to the Bcl‐2 family. Indeed, 1,25(OH)2D3 treatment increased levels of the anti‐apoptotic protein Bcl‐2 and decreased levels of the pro‐apoptotic proteins Bax and Bad in a time‐ and dose‐dependent way. Induction of Bcl‐2 by 1,25(OH)2D3 was further shown to be mediated by ERK and, to a lesser extent, by Akt. In conclusion, 1,25(OH)2D3 clearly protects keratinocytes against apoptosis (1) by activating the MEK/ERK and the PI‐3K/Akt survival pathways and (2) by increasing the Bcl‐2 to Bax and Bad ratio. © 2004 Wiley‐Liss, Inc.
Identification of alternative transcripts of the TRF1/Pin2 gene
Authors: Silva Lages Céline , Etienne Olivier , Comte Julien , Gauthier Laurent R. , Granotier Christine , Pennarun Gaëlle , Boussin François D. 
Published: September 2004

AbstractTRF1 and Pin2 play an essential role in telomere homeostasis, by regulating telomere maintenance. They are generated from the same gene, TRF1/Pin2, by alternative splicing but no functional differences between these proteins have been demonstrated. We report here the detection of new alternative transcripts of the TRF1/Pin2 gene in peripheral blood lymphocytes resulting from a 76 nt insertion. Real‐time RT‐PCR showed that these transcripts were also produced in various normal human cells and tissues and in immortalized cell lines, but at levels lower (by a factor of 8–111) than those for the TRF1 and Pin2 transcripts. These new transcripts are predicted to encode polypeptides identical to TRF1/Pin2 at the C‐terminal end but entirely lacking the acid domain and the amino‐terminal part of the homodimerization domain of TRF1/Pin2. These proteins, fused at their N‐terminal ends to enhanced green fluorescent protein (EGFP), were found to be located at telomeres and to induce apoptosis in cell lines with short telomeres, thereby displaying similar activity to TRF1/Pin2. However, these putative proteins lack regions important for interactions with other proteins and for homodimerization. Unlike TRF1/Pin2, they were unable to interact with tankyrase 1, suggesting that these proteins may play a role in telomere homeostasis different from those of TRF1/Pin2. The production of these alternative transcripts was down‐regulated in peripheral blood lymphocytes following PHA‐p activation, suggesting a possible role in resting lymphocytes. © 2004 Wiley‐Liss, Inc.
Vascular smooth muscle cell‐specific regulation of cyclin‐dependent kinase inhibitor p21WAF1/Cip1 transcription by Sp1 is mediated via distinct cis‐acting positive and negative regulatory elements in the proximal p21WAF1/Cip1 promoter
Authors: Kavurma Mary M. , Khachigian Levon M. 
Published: September 2004

AbstractSmooth muscle cells (SMC) play a central role in common vascular pathologies such as atherosclerosis and restenosis. Understanding the molecular regulation of SMC proliferation at a transcriptional level may provide important clues for the targeted control of vascular hyperplasia. We recently reported the capacity of the transcription factor Sp1 to down‐regulate p21WAF1/Cip1 production thereby reducing p21WAF1/Cip1–cyclin D1–Cdk4 complex formation and inhibiting vascular SMC proliferation (Kavurma and Khachigian [2003] J. Biol. Chem. 278, 32537–32543). We have now localized the Sp1‐response elements in the p21WAF1/Cip1 promoter responsible for p21WAF1/Cip1 repression in WKY12‐22 SMCs. The proximal region of the p21WAF1/Cip1 promoter contains five distinct Sp1‐binding elements that we have termed A, B, C, D, and E. Electrophoretic mobility shift analysis revealed that SMC nuclear Sp1 interacts with all five Sp1‐binding sites, and each of these sites is critical for Sp1 repression of the p21WAF1/Cip1 promoter, since mutation in any one element ablates repression, and in some cases results in activation. In contrast, only elements C, D, and E are bound by Sp1 in endothelial cells. Sp1 overexpression activates the p21WAF1/Cip1 promoter in this cell type. Furthermore, mutation in any of these five elements is not sufficient to prevent activation of the p21WAF1/Cip1 promoter by Sp1 in endothelial cells. Surprisingly, double mutations of elements C and E facilitates superactivation by Sp1 in both cell types, whereas triple mutations of C, D, and E inactivate the promoter. These findings demonstrate cell type‐specific regulation of p21WAF1/Cip1 transcription by Sp1 via distinct cis‐acting positive and negative regulatory elements in the proximal p21WAF1/Cip1 promoter. © 2004 Wiley‐Liss, Inc.
Production of the chemokine eotaxin‐1 in osteoarthritis and its role in cartilage degradation
Authors: Hsu Yi‐Hsin , Hsieh Ming‐Shium , Liang Yu‐Chih , Li Chao‐Yi , Sheu Ming‐Thau , Chou Der‐Tsay , Chen Tzeng‐Fu , Chen Chien‐Ho 
Published: September 2004

AbstractThe expression of the chemokine, eotaxin‐1, and its receptors in normal and osteoarthritic human chondrocytes was examined, and its role in cartilage degradation was elucidated in this study. Results indicated that plasma concentrations of eotaxin‐1 as well as the chemokines, RANTES, and MCP‐1α, were higher in patients with osteoarthritis (OA) than those in normal humans. Stimulation of chondrocytes with IL‐1β or TNF‐α significantly induced eotaxin‐1 expression. The production of eotaxin‐1 induced expression of its own receptor of CCR3 and CCR5 on the cell surface of chondrosarcomas, suggesting that an autocrine/paracrine pathway is involved in eotaxin‐1's action. In addition, eotaxin‐1 markedly increased the expressions of MMP‐3 and MMP‐13 mRNA, but had no effect on TIMP‐1 expression in chondrocytes. However, pretreatment of anti‐eotaxin‐1 antibody significantly decreased the MMP‐3 expression induced by IL‐1β. These results first demonstrate that human chondrocytes express the chemokine, eotaxin‐1, and that its expression is induced by treatment with IL‐1β and TNF‐α. The cytokine‐triggered induction of eotaxin‐1 further results in enhanced expressions of its own receptor of CCR3, CCR5, and MMPs, suggesting that eotaxin‐1 plays an important role in cartilage degradation in OA. © 2004 Wiley‐Liss, Inc.
Adenoviral down‐regulation of osteopontin inhibits human osteoclast differentiation in vitro
Authors: Aitken Cathy J. , Hodge Jason M. , Nicholson Geoffrey C. 
Published: September 2004

AbstractAlthough osteopontin (OPN) is highly expressed in osteoclasts, OPN‐deficient mice have a near‐normal bone phenotype and its role in osteoclast differentiation and function remains uncertain. We used an adenoviral OPN‐antisense vector (AdOPN‐AS) to down‐regulate OPN expression in a human in vitro osteoclastogenesis model employing CFU‐GM precursors treated with RANKL and M‐CSF. Cultures infected with AdOPN‐AS showed reduced secretion of OPN compared to cultures infected with a control adenoviral vector expressing β‐galactosidase. Infection with AdOPN‐AS co‐incident with exposure to RANKL was associated with substantial (approximately 50%) inhibition of osteoclast formation with a concomitant reduction in dentine resorption. There was also a small reduction in the size of generated osteoclasts but no significant effect on the size of resorption pits/tracks nor on the amount of resorption per osteoclast. When the cultures were infected with AdOPN‐AS after 4 days exposure to RANKL only minor effects on osteoclastogenesis were seen. Our data demonstrate that early down‐regulation of OPN in vitro inhibits human osteoclastogenesis. Since mice totally lacking OPN do not have reduced osteoclast numbers our results imply the existence in vivo of an alternative molecular pathway(s). © 2004 Wiley‐Liss, Inc.
Etiology of Paget's disease and osteoclast abnormalities
Authors: Reddy Sakamuri V. 
Published: September 2004

AbstractPaget's disease of bone is a chronic focal skeletal disorder that affects up to 2–3% of the population over the age of 60 years. Paget's disease is primarily a disease of the osteoclast. The pathologic abnormality in patients with Paget's disease involves increased bone resorption by the osteoclasts, followed by abundant new bone formation that is of poor quality. Genetic linkage analysis indicated that 40% of patients with Paget's disease have an affected first degree relative and 1% of patients develop osteosarcoma. Paget's disease is an autosomal dominant trait with genetic heterogeneity. Recurrent mutations in the ubiquitin‐associated (UBA) domain of sequestosome 1 (SQSTM1/p62) are identified in patients with Paget's disease. Osteoclasts and osteoclast precursors from patients with Paget's disease contain paramyxoviral transcripts and appear hyperresponsive to 1,25‐(OH)2D3 and RANK ligand (RANKL). It has been suggested that the enhanced sensitivity of osteoclast precursors for 1,25‐(OH)2D3 in Paget's disease results from increased expression of coactivators of vitamin D receptor (VDR). However, a cause and effect relationship for the paramyxoviral infection and SQSTM1/p62 gene mutations associated with this disease and osteoclast abnormalities are unclear. Therefore, the etiology of Paget's disease remains uncertain. © 2004 Wiley‐Liss, Inc.
Transactivation of the epidermal growth factor receptor mediates muscarinic stimulation of focal adhesion kinase in intestinal epithelial cells
Authors: Calandrella Sean O. , Barrett Kim E. , Keely Stephen J. 
Published: September 2004

AbstractWe have previously shown that the Gq protein coupled receptor (GqPCR) agonist, carbachol (CCh), transactivates and recruits epidermal growth factor receptor (EGFr)‐dependent signaling mechanisms in intestinal epithelial cells. Increasing evidence suggests that GqPCR agonists can also recruit focal adhesion‐dependent signaling pathways in some cell types. Therefore, the aim of the present study was to investigate if CCh stimulates activation of the focal adhesion‐associated protein, focal adhesion kinase (FAK), in intestinal epithelia and, if so, to examine the signaling mechanisms involved. Experiments were carried out on monolayers of T84 cells grown on permeable supports. CCh rapidly induced tyrosine phosphorylation of FAK in T84 cells. This effect was accompanied by phosphorylation of another focal adhesion‐associated protein, paxillin, and association of FAK with paxillin. CCh‐stimulated FAK phosphorylation was inhibited by a chelator of intracellular Ca2+, BAPTA/AM (20 μM), and was mimicked by thapsigargin (2 μM), which mobilizes intracellular Ca2+ in a receptor‐independent fashion. CCh also induced association of FAK with the EGFr and FAK phosphorylation was attenuated by an EGFr inhibitor, tyrphostin AG1478, and an inhibitor of Src family kinases, PP2. The actin cytoskeleton disruptor, cytochalasin D (20 μM), abolished FAK phosphorylation in response to CCh but did not alter CCh‐induced EGFr or ERK MAPK activation. In summary, these data demonstrate that agonists of GqPCRs have the ability to induce FAK activation in intestinal epithelial cells. GqPCR‐induced FAK activation is mediated by via a pathway involving transactivation of the EGFr and alterations in the actin cytoskeleton. © 2004 Wiley‐Liss, Inc.
Mechanistic understanding of the fermentative L‐glutamic acid overproduction by Corynebacterium glutamicum through combined metabolic flux profiling and transmembrane transport characteristics
Authors: Tryfona Theodora , Bustard Mark T 
Published: September 2004

AbstractSince the 1950s when Micrococcus glutamicus later renamed Corynebacterium glutamicum was discovered, the production of amino acids by fermentative methods has become an important aspect of industrial microbiology. Numerous studies to understand and improve the metabolic conditions leading to amino acid overproduction have been carried out. Most amino acids are currently produced by use of mutants that contain combinations of auxotrophic and regulatory mutations. L‐Glutamic acid is the amino acid produced in the greatest quantities (106 tonnes per year) and Corynebacteria are central to its industrial production. However, further improvements to strain performance are difficult to obtain by empirical optimization and a more rational approach is required. The use of metabolic flux analysis provides valuable information regarding bottlenecks in the formation of desired metabolites. Such techniques have found application in elucidating flux control, provided insight into metabolic network function and developed methods to amplify or redirect fluxes in engineered bioprocesses. Hence, branch points in biosynthesis, precursor supply in fuelling reactions and export of metabolites can be manipulated, resulting in high glutamic acid overproduction by Corynebacterium glutamicum fermentations. In this review, in addition to reviewing the state of play in metabolic flux analysis for glutamate overproduction, the metabolic pathways involved in the production of L‐glutamic acid, the mechanisms mediating its efflux and secretion as well as their manipulation to achieve higher glutamate production, are discussed. The link between metabolic flux and transmembrane transport of glutamic acid are also considered. Copyright © 2004 Society of Chemical Industry
New method for the detection of intraperitoneal free air by sonography: Scissors maneuver
Authors: Karahan Okkes Ibrahim , Kurt Alibekir , Yikilmaz Ali , Kahriman Guven 
Published: September 2004

AbstractPurposeThis study was conducted to investigate the diagnostic value of a new sonographic technique for the detection of intraabdominal free air.MethodsSeventy‐two patients with a suspected gastrointestinal tract perforation were included in the study and prospectively evaluated by sonography and abdominal and chest radiography for the detection of intraperitoneal free air. A new sonographic technique (the scissors maneuver) was used to detect intraperitoneal free air superficial to the liver; the maneuver consists of applying and then releasing slight pressure onto the abdominal wall with the caudal part of a parasagittaly oriented linear‐array probe. The results of the imaging studies were correlated with surgical findings when possible.ResultsSixteen patients had a surgically proven gastrointestinal tract perforation causing pneumoperitoneum. Sonography and radiography each showed pneumoperitoneum in 15 patients, with 1 false‐negative result for each modality. The sensitivity and specificity values of sonography and radiography were identical; sensitivity was 94% and specificity was 100% for both imaging modalities. The scissors maneuver was positive in all patients with sonographically detected pneumoperitoneum.ConclusionsSonography is an effective tool in the diagnosis of pneumoperitoneum, with sensitivity and specificity equal to those of radiography. The scissors maneuver may be a useful adjunct for improving the diagnostic yield of sonography. © 2004 Wiley Periodicals, Inc. J Clin Ultrasound 32:381–385, 2004
Neurogenic and non‐neurogenic placodes in ascidians
Authors: Manni Lucia , Lane Nancy J. , Joly Jean‐Stéphane , Gasparini Fabio , Tiozzo Stefano , Caicci Federico , Zaniolo Giovanna , Burighel Paolo 
Published: September 2004

AbstractThe late differentiation of the ectodermal layer is analysed in the ascidians Ciona intestinalis and Botryllus schlosseri, by means of light and electron microscopy, in order to verify the possible presence of placodal structures. Cranial placodes, ectodermal regions giving rise to nonepidermal cell types, are classically found exclusively in vertebrates; however, data are accumulating to demonstrate that the nonvertebrate chordates possess both the genetic machinery involved in placode differentiation, and ectodermal structures that are possible homologues of vertebrate placodes. Here, the term “placode” is used in a broad sense and defines thickenings of the ectodermal layer that can exhibit an interruption of the basal lamina where cells delaminate, and so are able to acquire a nonepidermal fate. A number of neurogenic placodes, ones capable of producing neurons, have been recognised; their derivatives have been analysed and their possible homologies with vertebrate placodes are discussed. In particular, the stomodeal placode may be considered a multiple placode, being composed of different sorts of placodes: part of it, which differentiates hair cells, is discussed as homologous to the octavo‐lateralis placodes, while the remaining portion, giving rise to the ciliated duct of the neural gland, is considered homologous to the adenohypophyseal placode. The neurohypophyseal placode may include the homologues of the hypothalamus and vertebrate olfactory placode; the rostral placode, producing the sensorial papillae, may possibly be homologous to the placodes of the adhesive gland of vertebrates. J. Exp. Zool. (Mol. Dev. Evol.) 302B:1–22, 2004. © 2004 Wiley‐Liss, Inc.
A synthetic peptide containing loop 4 of nerve growth factor for targeted gene delivery
Authors: Zeng Jieming , Too Heng‐Phon , Ma Yuexia , Luo Elizabeth S. E. , Wang Shu 
Published: September 2004

AbstractBackgroundGene delivery vectors that restrict the expression of a therapeutic gene to a particular type of cells are critical to gene therapy in a complex structure, such as the central nervous system. We constructed a nonviral vector for targeted gene transfer to cells expressing nerve growth factor (NGF) receptor TrkA.Methods and resultsThe vector was a synthetic chimeric peptide composed of a targeting moiety derived from NGF loop 4 and a DNA‐binding moiety of 10 lysine residues. The peptide activated signal transduction pathways of the NGF receptor TrkA in PC12 cells and supported the survival of the cells after serum deprivation. After forming complexes with plasmid DNA, the peptide dose‐dependently increased reporter gene expression in PC12 cells, which could be inhibited by excess NGF. The peptide‐mediated gene expression was not affected in PC12 cells by co‐incubation with a blocking antibody against the low‐affinity NGF receptor p75 and was significantly enhanced in NIH3T3 cells stably transfected with TrkA cDNA, suggesting the involvement of the high‐affinity NGF receptor TrkA without the participation of p75. Moreover, the peptide did not assist gene transfer in TrkA‐poor, but TrkB‐ and/or TrkC‐positive primary cerebellar granule neurons and primary cortical glial cells.ConclusionsThe chimeric peptide reported will be useful in gene delivery to and gene therapy of the nervous system and other tissues/organs with cells expressing TrkA. Copyright © 2004 John Wiley & Sons, Ltd.
Recovery of functional cytotoxic T lymphocytes during lamivudine therapy by acquiring multi‐specificity
Authors: Kondo Yasuteru , Asabe Shinichi , Kobayashi Koju , Shiina Masaaki , Niitsuma Hirofumi , Ueno Yoshiyuki , Kobayashi Tomoo , Shimosegawa Tooru 
Published: September 2004

AbstractTo characterize cytotoxic T lymphocytes (CTLs) that appeared in circulation during lamivudine therapy, we analyzed HBV‐specific CTLs using HLA‐A24 tetramer and HBcAg‐specific Th1 cells in patients receiving lamivudine therapy. Six patients (HLA‐A24+) with chronic hepatitis B, six patients (HLA‐A24−) with chronic hepatitis B, and six patients (HLA‐A24+) with chronic hepatitis C were studied. In addition to known CTL epitopes (C117 and P756), three epitopes were confirmed as CTL epitopes (C23, S89, S226) by chromium release assay and by staining intracellular perforin. CTLs specific for P756 were most frequently found at pre‐treatment. During lamivudine therapy, increase in the frequencies of HLA‐tetramer+ cells was found for C117, S89, and S226. Recovery of CTLs was observed earlier in patients with HBeAg−/anti‐HBe+ compared with those with HBeAg+/anti‐HBe−. HBcAg‐specific Th1 cells did not increase significantly up to 8 weeks. T cell lines from patients with chronic hepatitis B had a lower level of proliferation (0‐ to 24.9‐fold expansion by in vitro stimulation) and a higher ability to produce interferon‐γ (0–84% except for S89), while perforin‐positive cells showed low frequencies (0–50% except for S89). In conclusion, these results suggest that lamivudine therapy induces mainly CTLs that were less frequent before the therapy. Since recovered CTLs maintained the ability to produce interferon‐γ in response to peptides, these CTLs apparently contribute to the efficacy of lamivudine therapy in patients with hepatitis B. J. Med. Virol. 74:425–433, 2004. © 2004 Wiley‐Liss, Inc.