bluuebunny/crossref_metadata_2025 huggingface dataset:
Anticoagulant effect of sodium dehydroacetate (DHA‐S) in rats
Authors: Sakaguchi Yuka , Suga Sachiko , Oshida Keiyu , Miyamoto‐Kuramitsu Keiko , Ueda Kohei , Miyamoto Yohei
Published: October 2007

Sodium dehydroacetate (DHA‐S) is used as a food additive, preservative and antimicrobial agent. Repeated oral administration of DHA‐S in rats induced severe hemorrhage in multiple organs and prolongation of blood coagulation factors. To determine the mechanism of hemorrhage, the protective effect of vitamin K (VK) was investigated. Increased VK‐dependent blood coagulation parameters, a prolonged prothrombin time (PT) and an activated partial thromboplastin time (APTT) were observed in rats when DHA‐S alone was administered, while only a slight change was observed in animals that received a single injection of vitamin K2 following the DHA‐S dosing. These results suggest that DHA‐S‐induced hemorrhage is caused by a deficiency of vitamin K. In addition, the inhibitory effect of DHA‐S on vitamin K1 epoxide reductase (VKOR) activity was measured with an in vitro system using liver microsomes of normal male rats. DHA‐S concentration‐dependently inhibited VKOR activity similar to warfarin, but the inhibitory concentration was high. Therefore, it was concluded that the DHA‐S‐induced hemorrhage was caused by a depletion of blood VK, associated with any factors including VKOR inhibition. Copyright © 2007 John Wiley & Sons, Ltd.
Influence of surface microroughness by plasma deposition and chemical erosion followed by TiO2 coating upon anticoagulation, hydrophilicity, and corrosion resistance of NiTi alloy stent
Authors: Wang Gui‐Xue , Shen Yang , Zhang He , Quan Xue‐Jun , Yu Qing‐Song
Published: October 2007

Two different surface modification techniques were used to change the surface morphology and roughness of stents at the micrometer level, and eventually improve their surface adhesion properties with respect to endothelial cells. One was chemical erosion followed by sol–gel TiO2 coating, and the other was low temperature gas plasma deposition. After surface modification, the biocompatibility including the anticoagulation properties, hydrophilicity, and corrosion resistance of these stents was evaluated. It was found that both techniques could change the surface morphology of the stents with microroughness. In comparison with the control, the treated NiTi alloy intravascular stents showed increased surface hydrophilicity and enhanced anticoagulation properties. However, the corrosion properties of the stents were not improved significantly. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Influences of mechanical properties and permeability on chitosan nano/microfiber mesh tubes as a scaffold for nerve regeneration
Authors: Wang Wei , Itoh Soichiro , Matsuda Atsushi , Ichinose Shizuko , Shinomiya Kenichi , Hata Yuiro , Tanaka Junzo
Published: October 2007

We have developed chitosan nonwoven micro/nanofiber mesh tubes and examined the effects of their mechanical strength and permeability on nerve regeneration. Chitosan nano/microfibrous tubes with a deacetylation rate (DAc) of 78% or 93% were prepared by electrospinning. A chitosan film tube with a DAc of 93% was also fabricated and combined with the nano/microfibrous tubes to form bilayered tubes with a nano/microfiber mesh inner structure and a film outer layer. Nano/microfiber mesh tubes with a DAc of 78% or 93%, bilayered tubes with a nano/microfiber mesh inner structure with a DAc of 78% or 93% and a film outer layer with a DAc of 93%, and film tubes with a DAc of 93% were each tested as bridge grafts into injured rat sciatic nerve. Isografting was performed as a control. Although the functional recovery of motor activity was delayed in each group, sensory function reemerged first in the isograft group followed by the group receiving nano/microfiber mesh tubes with a DAc of 93%. Histological analysis was consistent with these results. The chitosan nano/microfiber mesh tubes with a DAc of 93% have sufficient mechanical properties to preserve tube space, provide a better scaffold for cell migration and attachment, and facilitate humoral permeation to enhance nerve regeneration. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Folate‐functionalized polymeric micelles for tumor targeted delivery of a potent multidrug‐resistance modulator FG020326
Authors: Yang Xiaoqiang , Deng Wenjing , Fu Liwu , Blanco Elvin , Gao Jinming , Quan Daping , Shuai Xintao
Published: October 2007

To overcome multidrug resistance (MDR) existing in tumor chemotherapy, polymeric micelles encoded with folic acid on the micelle surface were prepared with the encapsulation of a potent MDR modulator, FG020326. The micelles were fabricated from diblock copolymers of poly(ethylene glycol) (PEG) and biodegradable poly(ε‐caprolactone) (PCL) with folate attached to the distal ends of PEG chains. The folate‐conjugated copolymers, folate‐PEG‐PCL, were synthesized by multistep chemical reactions. First, allyl‐terminated copolymer (allyl‐PEG‐PCL) was synthesized through a ring‐opening polymerization of ε‐caprolactone in bulk employing monoallyl‐PEG as a macroinitiator. Second, the allyl terminal groups of copolymers were converted into primary amino groups by a radical addition reaction, followed by conjugation of the carboxylic group of folic acid. In vitro studies at 37°C demonstrated that FG020326 release from micelles at pH 5.0 was faster than that at pH 7.4. Cytotoxicity studies with MTT assays indicated that folate‐functionalized and FG020326‐loaded micelles resensitized the cells approximately five times more than their folate‐free counterparts (p < 0.01) in human KBv200 cells treated with vincristine (VCR). The in vitro Rhodamine 123 efflux experiment using MDR KBv200 cells revealed that when cells were pretreated with folate‐attached and FG020326‐loaded micelles, the P‐glycoprotein (P‐gp) drug efflux function was significantly inhibited. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res 2008
Bioactive glass‐induced osteoblast differentiation: A noninvasive spectroscopic study
Authors: Jell G. , Notingher I. , Tsigkou O. , Notingher P. , Polak J.M. , Hench L.L. , Stevens M.M.
Published: October 2007

Here, we report on a rapid, noninvasive biophotonics system using Raman spectroscopy to detect real‐time biochemical changes in foetal osteoblasts (FOBs) following exposure to 45S5 Bioglass® (BG)‐conditioned media. Bio‐Raman spectroscopy, combined with multivariate statistical analysis techniques (principal component analysis and least squares analysis), was able to noninvasively identify biochemical differences in FOBs cultured for different time periods and between FOBs exposed/or not to BG‐conditioned media. Gene and protein expression studies were also performed for known markers of osteoblastic differentiation, namely, alkaline phosphatase, bone sialoprotein, and collagen type I. Quantitative RT‐PCR confirmed upregulation of genes associated with osteoblast differentiation after exposure to BG‐conditioned media. These results suggest that Raman spectroscopy can noninvasively detect biochemical changes in FOBs associated with differentiation. This technique could have important applications in the field of regenerative medicine by enabling rapid characterization of cell or organoid behavior on novel bioactive scaffolds without damage to either cell or biomaterial. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res 2008
Osteoblast differentiation onto different biometals with an endoprosthetic surface topography in vitro
Authors: Jäger M. , Urselmann F. , Witte F. , Zanger K. , Li X. , Ayers D. C. , Krauspe R.
Published: October 2007

In this in vitro study, we compared the cytocompatibility and osteoblast promoting potency on human bone marrow cell culture with three different alloys (surgical steel, CoCr, Ti6Al4V) and three different surface structures (polished, sandblasted, porous coated). These biometals were specifically chosen because of their current applications in clinical orthopedic practices. Human mononuclear bone marrow cells were cultivated onto the surface of the different biomaterials and stimulated by dexamethasone, L‐ascorbic‐acid‐2‐phoshpate and beta‐glycerolphosphate over a 3‐week period. Immunfluorescent stainings against several antigens (ALP, RANKL, osteopontin, collagen I), mRNA‐expression of collagen (Col) I/II, BSP, osteopontin, osteocalcin, TRAP, light and scanning electron microscopy evaluation were used to evaluate cellular growth and osteoblast differentiation. For surface roughness and energy analysis of the specimen, roughness profile (Ra, Rz) and contact angle (CA) measurements were performed. We found differences between the different biometals and surface structures. Steel showed potential cytotoxic effects whereas CoCr and more Ti6Al4V showed an excellent cytocompatibility. There were no qualitative differences in mRNA expression between each of the tested biomaterials. In terms of antigen expression, a sandblasted Ti6Al4V surface showed enhanced osteoblastic differentiation. A porous‐coated surface improved the osteoconductivity of CoCr when compared to a polished surface. In contrast to controls all cells cultivated with biometals induced a RANKL expression. Cells increased the implant roughness with the exception of sandblasted Ti6Al4V. Our data show that surface topography and physicochemical properties of biometals influence osteoblast differentiation in vitro. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Evaluation of a highly‐radiopaque iodine‐containing acrylic bone cement for use in augmentation of vertebral compression fractures
Authors: Boelen Erik J.H. , Lewis Gladius , Xu Jie , Slots Tristan , Koole Leo H. , van Hooy‐Corstjens Catharina S.J.
Published: October 2007

Vertebroplasty and balloon kyphoplasty are widely used for the augmentation of osteoporosis‐induced vertebral compression fractures. Almost invariably, an injectable poly(methyl methacrylate) (PMMA) bone cement that contains a large amount of BaSO4particles is used in these procedures. The deleterious effects of this radiopacifier on various properties of PMMA cement have been detailed in the literature. The objective of the present study was therefore to avoid such high levels of inorganic contrast agent and thus to develop an all‐polymeric bone cement, for which radiopacity was provided by 60 wt % of an iodine‐containing methacrylic copolymer, incorporated into the powder (IO cement), ultimately leading to 6.6 wt % of iodine in the cement. A large array of properties of this cement were determined, ranging from setting time and injectability to fatigue life under fully‐reversed tension‐compression loading and cytotoxicity, and a comprehensive comparison with a cement containing 30 wt % BaSO4in the powder component (BA cement) has been made (11 wt % of Ba in the cement). Statistical analyses of the results showed that, for the majority of the properties, the difference between the means for the two cements was not significant. It is therefore suggested that the IO cement is a suitable alternative to the BA cement for use in the aforementioned procedures. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res 2008
Effects of calcium ion incorporation on osteoblast gene expression in MC3T3‐E1 cells cultured on microstructured titanium surfaces
Authors: Park Jin‐Woo , Suh Jo‐Young , Chung Hyun‐Ju
Published: October 2007

The surface characteristics of a calcium ion (Ca)‐incorporated titanium (Ti) surface, produced by hydrothermal treatment using an alkaline Ca‐containing solution, and its effects on osteoblastic differentiation were investigated. MC3T3‐E1 pre‐osteoblastic cells were cultured on machined or grit‐blasted Ti surfaces with and without Ca incorporation. The MTT assay was used to determine cell proliferation, and real‐time PCR was used for quantitative analysis of osteoblastic gene expression. Hydrothermal treatment with a Ca‐containing solution produced a crystalline CaTiO3 nanostructure of approximately 100 nm in dimension, preserving original micron‐scaled surface topographies and microroughness caused by machining, blasting, or blasting and etching treatments. After immersion in Hank's balanced salt solution, considerable apatite formation was observed on all surfaces of the Ca‐incorporated samples. Significantly more cell proliferation was found on Ca‐incorporated Ti surfaces than on untreated Ti surfaces (p < 0.001). Quantitative real‐time PCR analysis showed notably higher alkaline phosphatase, osteopontin, and osteocalcin mRNA levels in cells grown on Ca‐incorporated blasted surfaces than on other surfaces at an early time point. Thus, Ca incorporation may have a beneficial effect on osseointegration of microstructured Ti implants by accelerating osteoblast proliferation and differentiation during the early healing phase following implantation. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res 2008
A methodology based on the “anterior chamber of rabbit eyes” model for noninvasively determining the biocompatibility of biomaterials in an immune privileged site
Authors: Lu Pei‐Lin , Lai Jui‐Yang , Tabata Yasuhiko , Hsiue Ging‐Ho
Published: October 2007

In this study, a novel methodology based on the anterior chamber of rabbit eyes model was developed to evaluate the in vivo biocompatibility of biomaterials in an immune privileged site. The 7‐mm‐diameter membrane implants made from either a biological tissue material (amniotic membrane, AM group) or a biomedical polymeric material (gelatin, GM group) were inserted in rabbit anterior chamber for 36 months and characterized by biomicroscopic examinations, intraocular pressure measurements, and corneal thickness measurements. The noninvasive ophthalmic parameters were scored to provide a quantitative grading system. In this animal model, both AM and GM implants were visible in an ocular immune privileged site during clinical observations. The implants of the AM group appeared as soft tissue patches and have undergone a slow dissolution process resulting in a partial reduction of their size. Additionally, the AM implants did not induce any foreign body reaction or change in ocular tissue response for the studied period. By contrast, in the GM groups, significant corneal edema, elevated intraocular pressure, and increased corneal thickness were noted in the early postoperative phase (within 3 days), but resolved rapidly with in vivo dissolution of the gelatin. The results from the ocular grading system showed that both implants had good long‐term biocompatibility in an ocular immune privileged site for up to 3 years. It is concluded that the anterior chamber of rabbit eyes model is an efficient method for noninvasively determining the immune privileged tissue/biomaterial interactions. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Organic‐solvent extraction of model biomaterials for use in the in vitro chromosome aberration test
Authors: Matsuoka Atsuko , Haishima Yuji , Hasegawa Chie , Matsuda Yoshie , Tsuchiya Toshie
Published: October 2007

We prepared polyurethane (PU) containing 0.4% or 4% 4,4′‐methylenedianiline (MDA) as model materials to investigate the effectiveness of sample preparation by organic‐solvent extraction for the in vitro chromosome aberration (CA) test. MDA itself (0.4 mg/mL) was positive only in the presence of an exogenous metabolizing system (S9 mix). The culture medium extract of PU containing 4% MDA (PU/4% MDA) was negative with and without S9 mix. Methanol and acetone extracts, on the other hand, induced structural CAs without S9 mix, which we did not expect because MDA requires S9 mix for activity. On chemical analysis, however, we found that the ratio of MDA extracted by the organic solvents to that extracted by the culture medium of PU/4% MDA was about 15:1. Interestingly, oligomers consisting of poly(tetramethyleneglycol) derivatives (OTMG) were also extracted by the organic solvents. The data suggest that the induction of structural CAs in the absence of S9 mix may have been partly due to synergism of MDA and OTMG. CA tests of MDA and PTMG‐1000 in combination confirmed that to be the case. Thus, organic‐solvent extraction may be more effective than medium extraction in evaluating the biological safety of biomaterials. Detailed chemical analysis of extracts was performed. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Wear‐corrosion performance of Si‐DLC coatings on Ti‐6Al‐4V substrate
Authors: Kim Jung‐Gu , Lee Kwang‐Ryeol , Yang Seok‐Jo
Published: October 2007

Si‐incorporated diamond‐like carbon (Si‐DLC) coatings ranging from 0 to 2 at % Si were deposited on Ti‐alloy substrate by means of radio frequency plasma‐assisted chemical vapor deposition (r.f. PACVD) technique, using a mixture of benzene (C6H6) and silane (SiH4) as the reaction gas. The synergy in wear and corrosion of Si‐DLC coatings was investigated by tribological and electrochemical techniques. The electrolyte used in this test to simulate the corrosive environment of body fluid was a 0.89 wt % NaCl solution of pH 7.4 at 37°C. This study provides quantitative data for the assessment of the effect of Si incorporation on the synergistic effect between wear and corrosion in the simulated body fluid environment. In conclusion, tribological and electrochemical measurements showed that the Si‐DLC films could improve wear‐corrosion resistance in the simulated body fluid environment owing to the lower friction coefficient, corrosion rate, delamination area, and water uptake. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Characterization of porous injectable poly‐(propylene fumarate)‐based bone graft substitute
Authors: Kim Choll W. , Talac Robert , Lu Lichun , Moore Michael J. , Currier Bradford L. , Yaszemski Michael J.
Published: October 2007

The use of bone grafts for orthopedic applications have increased steadily over the past decade. With improvements in surgical technique, combined with an increasing aged population requiring orthopedic treatment, the need for bone grafts substitutes have also increased. To be useful clinically, the bone graft substitute must be biocompatible, bioabsorbable, and have convenient handling properties. In addition, it must possess a microarchitecture that allows cellular ingrowth and remodeling while simultaneously providing mechanical strength. Poly(propylene fumarate) (PPF) has been investigated as an injectable, biodegradable scaffold for orthopedic applications. Various methods to create a porous, interconnected polymer scaffold are available. The foaming technique is a convenient method to accomplish this task. Reactions between bicarbonate salts and weak acids generate CO2 gas, causing a bubbling reaction during the polymerization process. This technique allows the porosity of the scaffold to be modulated. Image analysis and mechanical testing of porous PPF fabricated using the foaming technique shows that a highly porous, interconnected scaffold can be produced. At ∼50% porosity, the scaffold has excellent handling properties, contains pore sizes ranging from 50 to 500 μm with an elastic modulus ranging from 20 to 40 MPa. The foaming technique provides an additional method by which clinically useful polymers can be fabricated for use in various bone tissue engineering applications. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008
Is HMGB1 an osteocyte alarmin?
Authors: Bidwell Joseph P. , Yang Jieping , Robling Alexander G.
Published: October 2007

The death of osteocytes, the terminally differentiated cells of the osteoblast lineage that are embedded in bone and regulate remodeling, is significant to both normal and pathological bone resorption. Apoptotic osteocytes putatively release a clarion signal that enhances the development of the bone‐resorbing osteoclasts and targets their migration to the breach in the osteocyte network. This phenomenon is thought to underlie normal repair of bone microdamage and contribute to the etiologies of inflammatory bone loss. The chromatin protein high mobility group box 1 protein (HMGB1) has been identified as an “alarmin” in other tissues. An alarmin is an endogenous molecule released by dead and dying cells that alert the innate immune system to damage and the need for tissue repair. Wang and colleagues presented evidence in a landmark 1999 study showing that released HMGB1 is a lethal mediator of sepsis. Extracellular HMGB1 is a ligand for the toll‐like receptors (TLRs) and for the receptor for advanced glycation end products (RAGE) all of which amplify inflammation. Recent studies by our lab and others have shown that HMGB1 is a bone‐active cytokine. It is released by apoptotic osteoblasts in vitro, including the MLO‐Y4 osteocyte‐like cells. Extracellular HMGB1 enhances the expression of RANKL, TNFα, and IL6 in osteoblastogenic bone marrow stromal cell cultures, and it is chemotactic to osteoclasts. In this prospectus we will review HMGB1 activity at the immune‐bone interface and propose a role for HMGB1 as an osteocyte alarmin and mediator of normal remodeling and inflammatory bone loss. J. Cell. Biochem. 103: 1671–1680, 2008. © 2007 Wiley‐Liss, Inc.
Activation or inactivation of cardiac Akt/mTOR signaling diverges physiological from pathological hypertrophy
Authors: Kemi Ole Johan , Ceci Marcello , Wisloff Ulrik , Grimaldi Serena , Gallo Paolo , Smith Godfrey L , Condorelli Gianluigi , Ellingsen Oyvind
Published: October 2007

Cardiomyocyte hypertrophy differs according to the stress exerted on the myocardium. While pressure overload‐induced cardiomyocyte hypertrophy is associated with depressed contractile function, physiological hypertrophy after exercise training associates with preserved or increased inotropy. We determined the activation state of myocardial Akt signaling with downstream substrates and fetal gene reactivation in exercise‐induced physiological and pressure overload‐induced pathological hypertrophies. C57BL/6J mice were either treadmill trained for 6 weeks, 5 days/week, at 85–90% of maximal oxygen uptake (VO2max), or underwent transverse aortic constriction (TAC) for 1 or 8 weeks. Total and phosphorylated protein levels were determined with SDS‐PAGE, and fetal genes by real‐time RT‐PCR. In the physiologically hypertrophied heart after exercise training, total Akt protein level was unchanged, but Akt was chronically hyperphosphorylated at serine 473. This was accompanied by activation of the mammalian target of rapamycin (mTOR), measured as phosphorylation of its two substrates: the ribosomal protein S6 kinase‐1 (S6K1) and the eukaryotic translation initiation factor‐4E binding protein‐1 (4E‐BP1). Exercise training did not reactivate the fetal gene program (β‐myosin heavy chain, atrial natriuretic factor, skeletal muscle actin). In contrast, pressure overload after TAC reactivated fetal genes already after 1 week, and partially inactivated the Akt/mTOR pathway and downstream substrates after 8 weeks. In conclusion, changes in opposite directions of the myocardial Akt/mTOR signal pathway appears to distinguish between physiological and pathological hypertrophies; exercise training associating with activation and pressure overload associating with inactivation of the Akt/mTOR pathway. J. Cell. Physiol. 214: 316–321, 2008. © 2007 Wiley‐Liss, Inc.
Extremely low‐frequency electromagnetic fields promote in vitro neurogenesis via upregulation of Cav1‐channel activity
Authors: Piacentini Roberto , Ripoli Cristian , Mezzogori Daniele , Azzena Gian Battista , Grassi Claudio
Published: October 2007

We previously reported that exposure to extremely low‐frequency electromagnetic fields (ELFEFs) increases the expression and function of voltage‐gated Ca2+ channels and that Ca2+ influx through Cav1 channels plays a key role in promoting the neuronal differentiation of neural stem/progenitor cells (NSCs). The present study was conducted to determine whether ELFEFs influence the neuronal differentiation of NSCs isolated from the brain cortices of newborn mice by modulating Cav1‐channel function. In cultures of differentiating NSCs exposed to ELFEFs (1 mT, 50 Hz), the percentage of cells displaying immunoreactivity for neuronal markers (β‐III‐tubulin, MAP2) and for Cav1.2 and Cav1.3 channels was markedly increased. NSC‐differentiated neurons in ELFEF‐exposed cultures also exhibited significant increases in spontaneous firing, in the percentage of cells exhibiting Ca2+ transients in response to KCl stimulation, in the amplitude of these transients and of Ca2+ currents generated by the activation of Cav1 channels. When the Cav1‐channel blocker nifedipine (5 µM) was added to the culture medium, the neuronal yield of NSC differentiation dropped significantly, and ELFEF exposure no longer produced significant increases in β‐III‐tubulin‐ and MAP2‐immunoreactivity rates. In contrast, the effects of ELFEFs were preserved when NSCs were cultured in the presence of either glutamate receptor antagonists or Cav2.1‐ and Cav2.2‐channel blockers. ELFEF stimulation during the first 24 h of differentiation caused Cav1‐dependent increases in the number of cells displaying CREB phosphorylation. Our data suggest that ELFEF exposure promotes neuronal differentiation of NSCs by upregulating Cav1‐channel expression and function. J. Cell. Physiol. 215: 129–139, 2008. © 2007 Wiley‐Liss, Inc.
Adrenomedullin is anti‐apoptotic in osteoblasts through CGRP1 receptors and MEK‐ERK pathway
Authors: Uzan Benjamin , Villemin Aude , Garel Jean‐Michel , Cressent Michèle
Published: October 2007

Adrenomedullin (ADM) has been shown to mediate multifunctional responses in cell culture and animal system such as regulation of growth and apoptosis. ADM stimulates the proliferation of osteoblasts in vitro and promotes bone growth in vivo. The ability of ADM to influence osteoblastic cell number through inhibition of apoptosis has not yet been studied. To address this question we have investigated its effect on the apoptosis of serum‐deprived osteoblastic cells using mouse MC3T3‐E1 cells which express both ADM and ADM receptors. Treatment with ADM significantly blunted apoptosis, evaluated by caspase‐3 activity, DNA fragmentation quantification and annexin V‐FITC labeling. This effect was abolished by the subtype‐1 CGRP receptor antagonist, CGRP(8–37). Both ADM and its specific receptor antagonist, the (22–52) ADM fragment exhibited a similar anti‐apoptotic effect. Thus, our data suggest that ADM exerts anti‐apoptotic effects through CGRP1 receptors. This was substantiated by a similar protective effect of CGRP on MC3T3‐E1 cells apoptosis. Accordingly, neutralization of endogenous ADM by a specific antibody enhanced apoptosis. Finally, the selective inhibitor of MAPK kinase (MEK), PD98059, abolished the apoptosis protective effect of ADM and prevented ADM activation of ERK1/2. These data show that ADM acts as a survival factor in osteoblastic cells via a CGRP1 receptor‐MEK‐ERK pathway, which provides further understanding on the physiological function of ADM in osteoblasts. J. Cell. Physiol. 215: 122–128, 2008. © 2007 Wiley‐Liss, Inc.
GHRH proliferative action on somatotrophs is cell‐type specific and dependent on Pit‐1/GHF‐1 expression
Authors: Solloso A , Barreiro L , Seoane R , Nogueira E , Cañibano C , Alvarez C.V. , Zalvide J , Diéguez C , Pombo C.M.
Published: October 2007

To investigate the mechanisms by which the hypothalamic peptide GHRH influences cell division, we analyzed its effects on the proliferation of two different cell lines: CHO‐4, an ovary‐derived cell line, and GH3, a pituitary‐derived cell line. We found that GHRH induces the proliferation of pituitary‐derived cells but inhibits the proliferation of ovary‐derived cells. We further characterized this dual effect of GHRH to find that the cytoplasmic signals induced by this hormone are similar in both cell lines. Moreover, in CHO‐4 cells GHRH stimulates two well‐known positive cell cycle regulators, c‐myc and cyclin D1, but is unable to induce the degradation of the negative cell cycle regulator p27Kip1. Significantly, when the Pit‐1/GHF‐1 gene is exogenously expressed in CHO‐4 cells, the negative effect of GHRH on the proliferation of these cells is attenuated. Furthermore, when the levels of Pit‐1 are downregulated by siRNA in GH3‐GHRHR cells, the positive effects of GHRH on the proliferation of these cells are diminished. These findings add to our understanding of the molecules involved in the regulation of cell proliferation by GHRH, as we demonstrate for the first time that Pit‐1 is not only required to drive the expression of the GHRH receptor, as previously described, but is also needed for the downstream effects that occur after its activation to modulate cell proliferation. These data suggest that the regulation of cell proliferation in response to a specific growth factor depends in certain cell populations on the presence of a tissue‐specific transcription factor. J. Cell. Physiol. 215: 140–150, 2008. © 2007 Wiley‐Liss, Inc.
All‐trans retinoic acid prevents angiotensin II‐ and mechanical stretch‐induced reactive oxygen species generation and cardiomyocyte apoptosis
Authors: Choudhary Rashmi , Baker Kenneth M. , Pan Jing
Published: October 2007

Cardiomyocyte apoptosis has an important role in the transition from compensatory cardiac remodeling to heart failure. All‐trans retinoic acid (RA), a bioactive vitamin A derivative, prevents stretch‐ and angiotensin II (Ang II)‐induced cardiac hypertrophy. However, the anti‐apoptotic potential of RA in the heart remains unexplored. Here, we demonstrate that stretch‐ and Ang II‐induced apoptosis is prevented by RA in neonatal cardiomyocytes. RA improved mitochondrial function by inhibiting the stretch‐ and Ang II‐induced reduction in mitochondrial membrane potential, cytochrome c release and by increasing the Bcl2/Bax ratio. RA inhibited stretch‐ and Ang II‐induced intracellular reactive oxygen species (ROS) generation and upregulated the SOD2 level. Hydrogen peroxide‐induced increases in the number of TUNEL‐positive cells and percentage of Annexin V positive cells, were dose‐dependently inhibited by RA. The thiol antioxidant, N‐acetyl cysteine (NAC), completely inhibited stretch‐ and Ang II‐induced apoptosis. Using diazoxide (mitochondrial ATP‐sensitive K+ channel opener) and SDS (NADPH oxidase activator), we confirmed that RA suppressed both mitochondrial‐ and NADPH oxidase‐derived ROS. We also observed that both RAR and RXR were involved in preventing Ang II‐ and stretch‐induced ROS production and apoptosis, by using selective retinoid receptor agonists and antagonists. Our data provide the first evidence that RA prevents Ang II and stretch induced apoptosis, by inhibiting ROS generation and increasing the anti‐oxidant defense system, suggesting that RA‐mediated signaling may provide a new therapeutic target for the prevention of the cardiac remodeling process. J. Cell. Physiol. 215: 172–181, 2008. © 2007 Wiley‐Liss, Inc.
Transcriptional responses of human epidermal keratinocytes to cytokine interleukin‐1
Authors: Yano Shoichiro , Banno Tomohiro , Walsh Rebecca , Blumenberg Miroslav
Published: October 2007

Interleukin‐1 is a proinflammatory and immunomodulatory cytokine that plays a crucial role in inflammatory diseases of the skin, including bacterial infections, bullous diseases, UV damage, and especially psoriasis. To characterize the molecular effects of IL‐1 in epidermis, we defined the transcriptional changes in human epidermal keratinocytes 1, 4, 24, and 48 h after treatment with IL‐1α. IL‐1 significantly regulated 388 genes, including genes associated with proteolysis, adhesion, signal transduction, proliferation, and epidermal differentiation. IL‐1 induces many genes that have antimicrobial function. Secreted cytokines, chemokines, growth factors, and their receptors are the prominent targets of IL‐1 regulation, including IL‐8, IL‐19, elafin, C3, and S100A proteins, which implicate IL‐1 in the pathogenesis of inflammatory diseases. IL‐1 induced not only proliferation‐associated genes but also differentiation marker genes such as transglutaminase‐1 and involucrin, which suggests that IL‐1 plays an important role in the aberrant proliferation and differentiation seen in psoriasis. Correlation of IL‐1 regulated genes with the TNFα and IFNγ regulated ones showed more similarities between IL‐1 and TNFα than IL‐1 and IFNγ, whereas Oncostatin‐M (OsM) affected a largely unrelated set of genes. IL‐1 regulates many genes previously shown to be specifically over‐expressed in psoriasis. In summary, IL‐1 regulates a characteristic set of genes that define its specific contribution to inflammation and aberrant differentiation in skin diseases. J. Cell. Physiol. 214:1–13, 2008. © 2007 Wiley‐Liss, Inc.
Etoposide‐resistant HT‐29 human colon carcinoma cells during glucose deprivation are sensitive to piericidin A, a GRP78 down‐regulator
Authors: Hwang Ji‐Hwan , Kim Ju‐Young , Cha Mi‐Ran , Ryoo In‐Ja , Choo Soo‐Jin , Cho Sung‐Min , Tsukumo Yoshinori , Tomida Akihiro , Shin‐Ya Kazuo , Hwang Yong‐Il , Yoo Ick‐Dong , Park Hae‐Ryong
Published: October 2007

Glucose deprivation, a pathophysiological cell condition, causes up‐regulation of GRP78 and induction of etoposide resistance in human cancer cells. The induction of drug resistance can be partly explained by the fact that GRP78 can block activation of caspase‐7 induced by treatment with etoposide. Therefore, downregulating GRP78 expression may be a novel strategy anticancer drug development. Based on that premise, we established a screening program for anticancer agents that exhibit preferential cytotoxic activity for etoposide‐resistant cancer cells under glucose‐deprived conditions. We recently isolated an active compound, AR‐054, from the culture broth of Streptomyces sp., which prevents stress‐induced etoposide resistance in vitro. AR‐054 was identified as piericidin A, a prototypical compound, by ESI‐MS analysis and various NMR spectroscopic methods. Here, we showed that piericidin A suppressed the accumulation of GRP78 protein and was also highly toxic to etoposide‐resistant HT‐29 cells, with IC50 values for colony formation of 6.4 and 7.7 nM under 2‐deoxyglucose supplemented and glucose‐deprived conditions, respectively. Interestingly, piericidin A had no effect under normal growth conditions. Therefore, we suggest that piericidin A prevents up‐regulation of GRP78, and exhibits cytotoxicity in glucose‐deprived HT‐29 cells that are resistant to etoposide. J. Cell. Physiol. 215: 243–250, 2008. © 2007 Wiley‐Liss, Inc.
Evidence that 3‐hydroxy‐3‐methylglutaric acid promotes lipid and protein oxidative damage and reduces the nonenzymatic antioxidant defenses in rat cerebral cortex
Authors: Leipnitz Guilhian , Seminotti Bianca , Haubrich Josué , Dalcin Manuela B. , Dalcin Karina B. , Solano Alexandre , de Bortoli Giorgia , Rosa Rafael B. , Amaral Alexandre U. , Dutra‐Filho Carlos S. , Latini Alexandra , Wajner Moacir
Published: October 2007

In the present work we investigated the in vitro effect of 3‐hydroxy‐3‐methylglutarate (HMG) that accumulates in 3‐hydroxy‐3‐methylglutaryl‐CoA lyase deficiency (HMGLD) on important parameters of oxidative stress in rat cerebral cortex. It was observed that HMG induced lipid peroxidation by significantly increasing chemiluminescence and levels of thiobarbituric acid‐reactive substances (TBA‐RS). This effect was prevented by the antioxidants α‐tocopherol, melatonin, N‐acetylcysteine, and superoxide dismutase plus catalase, suggesting that free radicals were involved in the lipid oxidative damage. On the other hand, HMG did not change TBA‐RS levels in intact or disrupted mitochondrial preparations, indicating that generation of oxidants by this organic acid was dependent on cytosolic mechanisms. HMG also induced protein oxidative damage in cortical supernatants, which was reflected by increased carbonyl content and sulfhydryl oxidation. Furthermore, HMG significantly reduced the nonenzymatic antioxidant defenses total‐radical trapping antioxidant potential, total antioxidant reactivity, and reduced glutathione (GSH) levels in rat cerebral cortex. HMG‐induced GSH reduction was totally blocked by melatonin pretreatment. We also verified that the decrease of GSH levels provoked by HMG in cortical supernatants was not due to a direct oxidative effect of this organic acid, because exposition of commercial GSH and purified membrane protein‐bound thiol groups to HMG in the absence of cortical supernatants did not decrease the reduced sulfhydryl groups. Finally, the activities of the main antioxidant enzymes were not altered by HMG exposure. Our data indicate that oxidative stress elicited in vitro by HMG may possibly contribute at least in part to the pathophysiology of the brain injury in HMGLD. © 2007 Wiley‐Liss, Inc.
Eicosapentaenoic acid stimulates the expression of myelin proteins in rat brain
Authors: Salvati Serafina , Natali Francesco , Attorri Lucilla , Di Benedetto Rita , Leonardi Fabiana , Di Biase Antonella , Ferri Federica , Fortuna Stefano , Lorenzini Paola , Sanchez Massimo , Ricceri Laura , Vitelli Luigi
Published: October 2007

We have previously demonstrated that, in C6 glioma cells, eicosapentaenoic acid (EPA) stimulates the expression of proteolipid protein (PLP) via cAMP‐mediated pathways. In this study, we investigated whether n‐3 polyunsaturated fatty acids can affect myelinogenesis in vivo. A single dose of either EPA or docosahexaenoic acid (DHA) was injected intracerebroventricularly into 2‐day‐old rats, which were then killed after 3 days post‐injection (p.i.). Total RNA was isolated from the medulla, cerebellum, and cortex, and the expression of myelin‐specific mRNAs was analyzed by real‐time PCR. The levels of PLP, myelin basic protein, and myelin oligodendrocyte protein mRNAs increased in nearly all brain regions of DHA‐ and EPA‐treated animals, but the effect was more pronounced in EPA‐treated rats. The enhancement in PLP transcript levels was followed by an increase in PLP translation in EPA‐treated rats. A further indicator of accelerated myelination was the increase in 2′‐3′‐cyclic nucleotide 3′‐phosphodiesterase (CNPase) protein levels. In EPA‐treated rats, the increased expression of myelin genes coincided with a decrease of cAMP‐response element‐binding protein (CREB)‐DNA binding in the cerebellum and cortex (1 hr p.i.). After 16 hr, this effect was still present in the same cerebral regions even though the decrease in EPA‐treated rats was less pronounced than in controls. The down‐regulation of CREB activity was due to a decrease in the levels of CREB phosphorylation. In conclusion, our data suggest that EPA stimulates the expression of specific myelin proteins through decreased CREB phosphorylation. These results corroborate the clinical studies of the n‐3 PUFA beneficial effects on several demyelinating diseases. © 2007 Wiley‐Liss, Inc.
Spermine induces cell death in cultured human embryonic cerebral cortical neurons through N‐methyl‐D‐aspartate receptor activation
Authors: de Vera Nuria , Martínez Emili , Sanfeliu Coral
Published: October 2007

The polyamines putrescine, spermidine, and spermine play important roles in cell proliferation, differentiation, and modulation of ion channel receptors. However, the function of increased concentrations of these compounds in brain injury and disease is unclear, in that they have been proposed as being both neuroprotective and neurotoxic. The effects of spermine and putrescine were studied in human primary cerebral cortical cultures containing both neurons and glia. No toxic effects were induced at 8 days in vitro (DIV) by either of the two polyamines at concentrations ranging from 0.3 μM to 2 mM. However, when the oxidative metabolism of spermine that generates toxic byproducts was induced by the presence of fetal calf serum, spermine caused cellular death with an LC50 of approximately 50 μM. At 14 DIV, the coapplication of spermine 2 mM and glutamate 5 mM induced neuron cell death, but the effect of applying both components separately was null. Both spermine and glutamate were toxic to older neurons (26–42 DIV cultures), and here the coapplication of glutamate was found always to intensify the effect of spermine. Spermine showed greater toxicity than glutamate in neurons. Another effect observed is that glutamate, but not spermine, induced astrocyte swelling. Spermine toxicity was inhibited by both MK801 and ifenprodil, indicating a mechanism involving N‐methyl‐D‐aspartate (NMDA) receptor activation. Moreover, a strong spermine modulation of the NMDA receptor was demonstrated by the inhibition of glutamate toxicity by ifenprodil. Putrescine induced minor effects also as a neurotoxic agent. In conclusion, neuronal death by spermine can be induced by its toxic byproducts as well as through NMDA receptor action. The present results confirm the potentially harmful role of the polyamines in excitotoxicity‐related human disorders. © 2007 Wiley‐Liss, Inc.
Ethanol promotes endoplasmic reticulum stress‐induced neuronal death: Involvement of oxidative stress
Authors: Chen Gang , Ma Cuiling , Bower Kimberly A. , Shi Xianglin , Ke Zunji , Luo Jia
Published: October 2007

One of the most devastating effects of ethanol exposure during development is the loss of neurons in selected brain areas. The underlying cellular/molecular mechanisms remain unclear. The endoplasmic reticulum (ER) is involved in posttranslational protein processing and transport. The accumulation of unfolded or misfolded proteins in the ER lumen triggers ER stress, which is characterized by translational attenuation, synthesis of ER chaperone proteins such as GRP78, and activation of transcription factors such as ATF4, ATF6, and CHOP. Sustained ER stress ultimately leads to cell death. ER stress response can be induced experimentally by treatment with tunicamycin and thapsigargin. Using SH‐SY5Y neuroblastoma cells and primary cerebellar granule neurons as in vitro models, we demonstrated that exposure to ethanol alone had little effect on the expression of markers for ER stress; however, ethanol drastically enhanced the expression of GRP78, CHOP, ATF4, ATF6, and phosphorylated PERK and eIF2α when induced by tunicamycin and thapsigargin. Consistently, ethanol promoted tunicamycin‐ and thapsigargin‐induced cell death. Ethanol rapidly caused oxidative stress in cultured neuronal cells; antioxidants blocked ethanol's potentiation of ER stress and cell death, suggesting that the ethanol‐promoted ER stress response is mediated by oxidative stress. CHOP is a proapoptotic transcription factor. We further demonstrated that CHOP played an important role in ethanol‐promoted cell death. Thus, the effect of ethanol may be mediated by the interaction between oxidative stress and ER stress. © 2007 Wiley‐Liss, Inc.
Dye coupling in developing vestibular nuclei
Authors: Shao M. , Gottesman‐Davis A. , Popratiloff A. , Peusner K.D.
Published: October 2007

The chick tangential nucleus is a major vestibular nucleus whose principal cells participate in the vestibular reflexes. During development, most mature vestibular nucleus neurons must acquire repetitive firing of action potentials on depolarization and spontaneous spike activity to process signals effectively. In the chicken, these properties emerge gradually in the embryo, starting the week before (E13, E16) and continuing through the first week after hatching (H7). Since gap junction‐mediated cell coupling may influence the emergence of neuronal excitability, we investigated whether neuron–neuron and neuron–glia coupling are present in this morphologically distinctive vestibular nucleus during the period for establishing signal processing. In brain slices, principal cells were injected with biocytin in the whole‐cell configuration and visualized via confocal imaging at E13, E16, and H7. The incidence of dye coupling between the injected principal cell and neurons was 42% at E13, 75% at E16, and 7% at H7, whereas the incidence of dye coupling with glia was 100% at both embryonic ages but decreased to 27% by H7. For each injected principal cell at E13, one coupled neuron and 35 coupled glia were detected, whereas three coupled neurons and 12 coupled glia were observed at E16, and few if any coupled neurons and glia were detected at H7. These results suggest that neuron–neuron and neuron–glia coupling are developmentally regulated and present before, but not after, the onset of mature signal processing by these neurons. Thus, transient neuron–neuron and neuron–glia coupling may both play roles in establishing excitability in vestibular nucleus neurons during development. © 2007 Wiley‐Liss, Inc.
Increasing gamma‐ and delta‐tocopherols in oils improves oxidative stability
Authors: Warner Kathleen
Published: October 2007

Over the past two decades, plant geneticists have revolutionized how fatty acid compositions of vegetable oils are optimized to improve oxidative stability and functionality. Now, the expertise of plant geneticists is reaching beyond altering fatty acids to changing other oil components such as tocopherols. Basic lipid research on optimizing tocopherol profiles and ratios in vegetable oils is providing information for geneticists to breed the next generation of oxidatively stable vegetable oils. This review will discuss three studies; first, a basic study to determine the oxidative stability provided by the addition of pure gamma and delta tocopherols to oils treated to remove natural tocopherols; second, a practical study to evaluate the oxidative stability of mid‐oleic sunflower oil from seeds modified by plant breeding to contain high amounts of γ‐ and δ‐tocopherols; and third, a frying test to determine the effects of gamma tocopherol addition.
3D CT angiography of abdominal wall vascular perforators to plan DIEAP flaps
Authors: Rosson Gedge D. , Williams Christopher G. , Fishman Elliot K. , Singh Navin K.
Published: October 2007

Purpose: Since the first report of TRAM flap reconstruction, there have been numerous studies to reduce complications of elective breast reconstruction. Current methods of preoperative perforator localization can be time‐consuming, inaccurate, and imprecise. Thus, we sought to evaluate ultra‐high resolution 3D CT angiography for the preoperative mapping of DIEAP flap perforating vessels. Methods: We reviewed all perforator‐based breast reconstructions performed over a 5‐month period. Candidates for DIEAP flap reconstruction were sent for a focused CT scan of the abdominal wall, using the 64 slice multi‐detector CT scanner. Results: This article presents our first 23 flaps in 17 patients with preoperative ultra‐high resolution 3D CT angiography. The reconstruction plan changed in three patients (18%). There was one take‐back for venous congestion, but no partial or total flap loss. Conclusions: Preoperative perforator flap planning for breast reconstruction utilizing 3D CT angiogram is safe, easy to read, and can change the operative plan. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.