bluuebunny/crossref_metadata_2025 huggingface dataset:
Osteogenecity of octacalcium phosphate coatings applied on porous metal implants
Authors: Barrère Florence , van der Valk Chantal M. , Dalmeijer Remco A. J. , Meijer Gert , van Blitterswijk Clemens A. , de Groot Klaas , Layrolle Pierre
Published: August 2003

The biomimetic route allows the homogeneous deposition of calcium phosphate (Ca‐P) coatings on porous implants by immersion in simulated physiologic solution. In addition, various Ca‐P phases, such as octacalcium phosphate (OCP) or bone‐like carbonated apatite (BCA), which are stable only at low temperatures, can be deposited. In this pilot study, experiments were designed with a twofold‐purpose: (1) to investigate the osteoinduction of OCP‐coated and noncoated porous tantalum cylinders and of dense titanium alloy cylinders (5 mm in diameter and 10 mm in length) in the back muscle of goats at 12 and 24 weeks (n = 4); and (2) to compare the osteogenic potentials of BCA‐coated, OCP‐coated, and bare porous tantalum cylinders in a gap of 1 mm created in the femoral condyle of a goat at 12 weeks (n = 2). In the goat muscle, after 12 weeks the OCP‐coated porous cylinder had induced ectopic bone as well as bone within the cavity of the OCP‐coated dense titanium cylinder. In the femoral condyle, bone did not fill the gap in any of the porous implants. In contrast with the two other groups, OCP‐coated porous cylinders exhibited bone formation in the center of the implant. The nature of the Ca‐P coating, via its microstructure, its dissolution rate, and its specific interactions with body fluids, may influence the osteogenecity of the Ca‐P biomaterial. © 2003 Wiley Periodicals, Inc. J Biomed Mater Res 66A: 779–788, 2003
Corrosion resistance of stressed NiTi and stainless steel orthodontic wires in acid artificial saliva
Authors: Huang Her‐Hsiung
Published: August 2003

The purpose of this study was to investigate the corrosion resistance of stressed NiTi and stainless steel orthodontic wires using cyclic potentiodynamic and potentiostatic tests in acid artificial saliva at 37°C. An atomic force microscope was used to measure the 3‐D surface topography of as‐received wires. Scanning electron microscope observations were carried out before and after the cyclic potentiodynamic tests. The surface chemical analysis was characterized using X‐ray photoelectron spectroscopy and Auger electron spectroscopy after the potentiostatic tests. The cyclic potentiodynamic test results showed that the pH had a significant influence on the corrosion parameters of the stressed NiTi and stainless steel wires (p < 0.05). The pitting potential, protection potential, and passive range of stressed NiTi and stainless steel wires decreased on decreasing pH, whereas the passive current density increased on decreasing pH. The load had no significant influence on the above corrosion parameters (p > 0.05). For all pH and load conditions, stainless steel wire showed higher pitting potential and wider passive range than NiTi wire (p < 0.001), whereas NiTi wire had lower passive current density than stainless steel wire (p < 0.001). The corrosion resistance of the stressed NiTi and stainless steel wires was related to the surface characterizations, including surface defect and passive film. © 2003 Wiley Periodicals, Inc. J Biomed Mater Res 66A: 829–839, 2003
The effect of hyaluronan on osteoblast proliferation and differentiation in rat calvarial‐derived cell cultures
Authors: Huang L. , Cheng Y. Y. , Koo P. L. , Lee K. M. , Qin L. , Cheng J. C. Y. , Kumta S. M.
Published: August 2003

Hyaluronan (or hyaluronic acid, HA) is an essential component of extracellular matrices. It interacts with other macromolecules and plays a predominant role in tissue morphogenesis, cell migration, differentiation, and adhesion. The cell signaling functions of HA are mediated through the CD‐44 receptor and are dependent upon the molecular weight of the polymer. We hypothesized that an HA of appropriate molecular weight alone in optimal concentration may induce osteoblast differentiation and bone formation. Enzyme‐digested calvarial‐derived mesenchymal cells from 2‐day‐old newborn rats were cultured with the addition of HA of three different molecular weights (2300, 900, and 60 kDa). We added, 0.5, 1.0, and 2.0 mg/mL HA for each molecular weight to the medium at the first plating of cells. After 7 to 20 days in culture, cell proliferation and differentiation were evaluated by measuring thymidine incorporation, alkaline phosphatase activity, and osteocalcin gene expression. The effects of HA on bone formation were examined by using Alizarin red staining for mineralization. The results showed that low molecular weight HA (60 kDa) significantly stimulated cell growth, increased osteocalcin mRNA expression in a dose‐dependent manner, but showed no apparent effects on alkaline phosphatase activity and bone mineralization. On the other hand, high‐weight HA (900 and 2300 kDa) significantly increased all the parameters examined, particularly alkaline phosphatase activity, in a dose‐dependent manner and stimulated cell mineralization to 126% and 119% of the controls, respectively, in the 1.0 mg/mL dose. Our findings suggest that HA has a molecular weight‐specific and dose‐specific mode of action that may enhance the osteogenic and osteoinductive properties of bone graft materials and substitutes due to its stimulatory effects on osteoblasts. © 2003 Wiley Periodicals, Inc. J Biomed Mater Res 66A: 880–884, 2003
Influence of dentinal polyelectrolytes on wet demineralized dentin, a bonding substrate
Authors: Piemjai M. , Iwasaki Y. , Nakabayashi N.
Published: August 2003

The objective of this study was to show the influence of dissolved dentinal polyelectrolytes on the characteristics of dentin (bonding substrate) demineralized by citric acid in the absence or presence of ferric chloride. The demineralizing agent was an aqueous mixture of 0, 1, 3, or 10% ferric chloride in 10% citric acid (10–0, 10–1, 10–3, 10–10, respectively). The hypothesis was that the concentration of dissolved dentinal noncollagenous substances, mainly polyelectrolytes soluble in water, must be decreased by their aggregation with ferric ions, which changes the characteristics of demineralized dentin, the rates of demineralization, and dehydration. Cervical bovine dentin was prepared in 3 × 2 × 2‐mm blocks, each weighing 20.0 ± 0.5 mg. The rate of demineralization was investigated by measuring the weight loss resulting from demineralization by immersion in 10 mL of conditioner at 2‐h intervals. The dehydration rate of wet demineralized dentin was determined using two methods: (1) weight loss in a desiccator under 263 Pa pressure and (2) differential scanning calorimetry (DSC). Twenty, 12, 8, and 4 h were required to complete demineralization of the blocks with the 10–0, 10–1, 10–3, and 10–10 solutions, respectively. The 10–10 wet demineralized dentin showed the highest rate of dehydration, followed in descending order by the 10–3, 10–1, and 10–0 specimens. Ferric chloride in dentin conditioners provided both a higher rate of dentin demineralization and a higher dehydration rate of wet demineralized dentin. These results suggest that in the presence of ferric chloride, a decreasing amount of dissolved polyelectrolytes aggregated with ferric ions in the substrates may increase the permeability of dentin to water and citric acid. Improvement of monomer permeability is essential to the preparation of good hybridized dentin, providing a more stable and reliable bonding and also protecting the dentin and pulp from infection. A further study of bonding substrates is required in order to understand the role of hybridized dentin in improved dental treatment. © 2003 Wiley Periodicals, Inc. J Biomed Mater Res 66A: 789–794, 2003
High‐affinity augmentation of endothelial cell attachment: Long‐term effects on focal contact and actin filament formation
Authors: Mathur Anshu B. , Chan Bernard P. , Truskey George A. , Reichert William M.
Published: August 2003

Coadsorption of high‐affinity avidin with lower affinity cell adhesion protein fibronectin has been shown to significantly augment short‐term (1 h) adhesion and spreading of endothelial cells; however, the longer term persistence of avidin binding and its effect on endothelial cell adhesion have not been addressed. In this study, the presence of avidin–biotin bonds 24 h after cell adhesion to the dual ligand surfaces was verified by laser confocal microscopy of a fluorescent avidin analog, streptavidin. Total internal reflection microscopy showed that the focal contact area, focal contact density, and cell spreading all increased significantly at 24 h compared to fibronectin‐treated control surfaces. Focal contact area was identical when measured with cells that were labeled with either the fluorescent streptavidin or a carbocyanine dye incorporated in the cell membrane. Confocal images of stress fibers formed in cells adherent to dual ligand surfaces after 24 h were thicker and more numerous compared to cells adherent to fibronectin controls. The results indicate that 24 h after initial attachment avidin–biotin is localized to focal contacts on the basal surface and affects cell spreading, actin filament organization, and focal contact density. © 2003 Wiley Periodicals, Inc. J Biomed Mater Res 66A: 729–737, 2003
The profits and perils of international outsourcing
Authors: Bean LuAnn
Published: August 2003

More and more companies are outsourcing work abroad. This trend is transforming everything from call centers
to credit cards. But there are both benefits and dangers for your company. The author investigates how companies
are using this strategy today and gives some solid advice to help you choose the right outsourcing model for your
particular company. © 2003 Wiley Periodicals, Inc.
Key trends in Southeast Asia
Authors: Jasmon Abadan , Nordin Mohd Ridzuan
Published: August 2003

Tempted to do business in Southeast Asia (SEA)? Many SEA countries offer very attractive
incentives—in addition to a low‐cost, high‐quality labor force. They also allow 100 percent
foreign ownership, tax holidays, and capital allowances. But there are important differences compared to developed
countries. Government agencies and corporations play a different role, and business decisions may not be made in
the way you expect. What must you know to avoid potential problems? Where are the best opportunities?
© 2003 Wiley Periodicals, Inc.
An insider's guide to doing business in India
Authors: Rathinasamy R. S. , Mantripragada Krishna G. , Krishnan Rishikesha T. , Shivaswamy Melkote K.
Published: August 2003

Many U.S. companies are searching for foreign business opportunities. And the world's largest
democracy—India—has become a favorite of U.S. multinational companies. But what is the Indian business
climate really like? What risks do you face? What pitfalls should you avoid? Where are the best
fast‐growth opportunities? Here is a comprehensive, hands‐on guide to doing business in India
that will answer those vital questions and more. © 2003 Wiley Periodicals, Inc.
How to handle the threat of catastrophe
Authors: Sánchez Carol , Goldberg Stephen R.
Published: August 2003

One major task CEOs face is to minimize risk and vulnerability to catastrophic events. Since 9/11, we know
that simply having insurance—and conventional emergency planning—is not enough. And in addition to
global terrorism, other catastrophes threaten—including cyber crime and new diseases like SARS. Traditional
risk management strategies dealt with two types of danger: known and unknown risk. But now we also have a third
kind: unknowable risk! So how should we handle catastrophic threats today? © 2003 Wiley
Periodicals, Inc.
Managing risk in global operations
Authors: Massoud Marc F. , Raiborn Cecily A.
Published: August 2003

Companies doing business abroad face two tough questions: How can we assess our risks? And what are the
best strategies to minimize those risks? The authors explain how to develop a risk control strategy and how to
lessen your risk using financial mechanisms. © 2003 Wiley Periodicals, Inc.
Don't be a victim of international fraud
Authors: Byington J. Ralph , Christensen Jo Ann
Published: August 2003

American companies conducting business abroad must be wary about the risk of international fraud. Why and
how has this become a problem? What are the different types of international fraud? And what
measures can reduce your risk? The authors provide some answers. © 2003 Wiley Periodicals,
Inc.
How will Sarbanes‐Oxley affect foreign companies?
Authors: Rouse Robert W.
Published: August 2003

The Sarbanes‐Oxley Act (SOA) will impact far more than just U.S. domiciled companies, says
the author. © 2003 Wiley Periodicals, Inc.
Euro M&A: Dealmakers rethink their strategies
Authors: Metwalli Ali M. , Tang Roger Y.W.
Published: August 2003

M&A (merger and acquisition) activities in the European Union expanded steadily in the 1990s,
reaching a peak in 1999 and 2000. But there was a big slowdown after that, due to a faltering global economy, the
9/11 attacks, and fears of terrorism and regulatory changes. Many firms were forced to rethink their M&A
strategies. © 2003 Wiley Periodicals, Inc.
Hedge accounting: What's the problem with IASC standards?
Authors: Hernández Francisco Gabriel Hernández
Published: August 2003

The International Accounting Standards Committee (IASC) requires companies use IAS 39 and IAS 32 in
accounting for financial instruments and hedging activities. But this causes problems, says the author. Instead,
he offers an alternative approach. © 2003 Wiley Periodicals, Inc.
Pro‐ethics activities: Do they really reduce fraud?
Authors: Singleton Tommie , King Brett , Messina Frank M. , Turpen Richard A.
Published: August 2003

We have seen a growing number of extraordinary financial frauds over the past few years. So critics call for
more focus on ethics, and more pro‐ethics activities. But do ethics programs really reduce fraud? The
results of one study may surprise you. © 2003 Wiley Periodicals, Inc.
International auditing and managing exports
Authors: Goldberg Stephen R. , Godwin Joseph H.
Published: August 2003

O'Regan, David. 2003. International Auditing, Practical Resource Guide (New York:
John Wiley & Sons).

Reynolds, Frank. 2003. Managing Exports: Navigating the Complex Rules, Controls, Barriers, and
Laws (New York: John Wiley & Sons).
How much does bad data cost your company?
Authors: Jankovsky Lynn , Grimme Julie
Published: August 2003

A recent project at Motorola that addressed the cost of bad data accomplished several important goals. Among
these, the project identified the cost of bad data for a certain data element and then calculated the percentage
of cost of the element in terms of total cost. This contributed to the knowledge of reducing cost by understanding
the cost drivers; it also helped managers in prioritization, resource management, and implementation of
enterprisewide systems. © 2003 Wiley Periodicals, Inc.
Effect of dexamethasone withdrawal on osteoblastic differentiation of bone marrow stromal cells
Authors: Porter Ryan M. , Huckle William R. , Goldstein Aaron S.
Published: August 2003

Dexamethasone is capable of directing osteoblastic differentiation of bone marrow stromal cells (BMSCs) in vitro, but its effects are not lineage‐specific, and sustained exposure has been shown to down‐regulate collagen synthesis and induce maturation of an adipocyte subpopulation within BMSC cultures. Such side effects might be reduced if dexamethasone is applied in a regimented manner, but the discrete steps in osteoblastic maturation that are stimulated by dexamethasone are not known. To examine this, dexamethasone was added to medium to initiate differentiation of rat BMSCs cultures and then removed after a varying number of days. Cell layers were analyzed for cell number, rate of collagen synthesis, expression of osteocalcin (OC), bone sialoprotein (BSP) and lipoprotein lipase (LpL), and matrix mineralization. Withdrawal of dexamethasone at 3 and 10 days was found to enhance cell number relative to continuous exposure, but did not affect to decrease collagen synthesis slightly. Late markers of osteoblastic differentiation, BSP expression and matrix mineralization, were also sensitive to dexamethasone and increased systematically with exposure while LpL systematically decreased. These results indicate that dexamethasone acts at both early and late stages to direct proliferative osteoprogenitor cells toward terminal maturation. J. Cell. Biochem. 90: 13–22, 2003. © 2003 Wiley‐Liss, Inc.
Immune cell proliferation is suppressed by the interferon‐γ‐induced indoleamine 2,3‐dioxygenase expression of fibroblasts populated in collagen gel (FPCG)
Authors: Sarkhosh Kourosh , Tredget Edward E. , Karami Ali , Uludag Hasan , Iwashina Takashi , Kilani Ruhangiz T. , Ghahary Aziz
Published: August 2003

Indoleamine 2,3‐dioxygenase (IDO), a tryptophan‐catabolizing enzyme, is an intracellular enzyme possessing various immunosuppressive properties. Here, we report the possible use of this enzyme to suppress proliferation of immune cells cocultured with IDO‐expressing fibroblasts of an allogenic skin substitute. Fetal skin fibroblasts embedded within bovine collagen were treated with cytokine interferon‐γ (IFN‐γ) to induce expression of IDO mRNA and protein. Expression of IDO mRNA was evaluated by Northern analysis. IDO enzyme activity was evaluated by measurement of kynurenine and tryptophan levels in the IFN‐γ untreated and treated fibroblasts. The results of Northern analysis showed a dose‐dependent increase in expression of IDO mRNA in response to various concentrations of IFN‐γ used. The levels of kynurenine and tryptophan measured, as the bioactivity of IDO, were significantly different in the IFN‐γ treated fibroblasts, compared to those of controls (P < 0.001). In a lasting effect experiment, the expression of IDO mRNA was gradually reduced to an undetectable level within 32 h of IFN‐γ removal. The results of Western blot analysis, however, revealed a significantly longer (192 h) lasting effect of IFN‐γ on IDO protein level, relative to that of mRNA expression. To demonstrate immunosuppressive effects of IDO on proliferation of immune cells, IDO‐expressing fibroblasts were cocultured with peripheral blood mononuclear cells (PBMC) for a period of 5 days. The results of 3H‐thymidine incorporation showed a significant reduction in proliferation of PBMC when cocultured with IDO‐expressing fibroblasts, compared to those cocultured with non‐IDO‐expressing fibroblasts (P < 0.001). Furthermore, addition of IDO‐inhibitor (1‐methyl‐d‐tryptophan) reversed the suppressive effects of IDO on PBMC proliferation in a dose‐dependant fashion. To test the viability of immune cells cocultured with IDO‐expressing fibroblasts, FACS analysis of the PI stained PBMC was conducted and no significant difference was found between these cells and the controls. In another set of experiments, we showed that migration rate and subsequent proliferation of IDO‐expressing fibroblasts are also the same as those of control cells. In conclusion, IDO‐expressing allogenic fibroblasts embedded within collagen gel suppress the proliferation of allogenic immune cells, while they still remain viable in this IDO‐induced tryptophan‐deficient culture environment. J. Cell. Biochem. 90: 206–217, 2003. © 2003 Wiley‐Liss, Inc.
Fluorescence activated cell sorting reveals heterogeneous and cell non‐autonomous osteoprogenitor differentiation in fetal rat calvaria cell populations
Authors: Purpura Kelly A. , Zandstra Peter W. , Aubin Jane E.
Published: August 2003

Identification of osteoblast progenitors, with defined developmental capacity, would facilitate studies on a variety of parameters of bone development. We used expression of alkaline phosphatase (ALP) and the parathyroid hormone/parathyroid hormone‐related protein receptor (PTH1R) as osteoblast markers in dual‐color fluorescence activated cell sorting (FACS) to fractionate rat calvaria (RC) cells into ALP−PTH1R−, ALP+PTH1R−, ALP−PTH1R+, and ALP+PTH1R+ populations. These fractionated populations were seeded clonally (n = 96) or over a range of cell densities (∼150–8,500 cell/cm2; n = 3). Our results indicate that colony forming unit‐osteoblast (CFU‐O)/bone nodule‐forming cells are found in all fractions, but the frequency of CFU‐O and total mineralized area is different across fractions. Analysis of these differences suggests that ALP−PTH1R−, ALP−PTH1R+, ALP+PTH1R−, and ALP+PTH1R+ cell populations are separated in order of increasing bone formation capacity. Dexamethasone (dex) differentially increased the CFU‐O number in the four fractions, with the largest stimulation in the ALP− cell populations. However, there was no significant difference in the number or size distribution of CFU‐F (fibroblast) colonies that formed in vehicle versus dex. Finally, both cell autonomous and cell non‐autonomous (i.e., inhibitory/stimulatory effects of cell neighbors) differentiation of osteoprogenitors was seen. Only the ALP−PTH1R− population was capable of forming nodules at the clonal level, at approximately 3‐ or 12‐times the predicted frequency of unfractionated populations in dex or vehicle, respectively. These data suggest that osteoprogenitors can be significantly enriched by fractionation of RC populations, that assay conditions modify the osteoprogenitor frequencies observed and that fractionation of osteogenic populations is useful for interrogation of their developmental status and osteogenic capacity. J. Cell. Biochem. 90: 109–120, 2003. © 2003 Wiley‐Liss, Inc.
How is Type I procollagen synthesis regulated at the gene level during tissue fibrosis
Authors: Cutroneo Kenneth R.
Published: August 2003

In response to tissue injury connective tissue synthesis occurs either normally or abnormally, which is mediated by transforming growth factor‐β (TGF‐β) and other growth factors. This article will be primarily concerned with the response of injured tissues at the gene level of Type I procollagen synthesis in response to TGF‐β. This leads to provisional repair, which in turn may lead to involution, remodeling, regeneration, and ultimately repair. Alternately, continuation of provisional repair may lead to fibrosis and ultimately scarring. Scarring of internal organs such as the liver and the lung leads to loss of function and ultimately death. In the case of scarring of skin, this is a cosmetic problem and can be rectified by surgery. Type I procollagen is synthesized by two genes, proα1 (Type I) and proα2 (Type I) collagen genes. This article will focus on DNA binding sites on these two genes, which regulate the transcription of the specific gene. This article will also define specific cell signaling pathways for the turning on of the proα1 and proα2 (Type I) collagen genes. This article will address several questions. First, what is the major cytokine acting extracellularly which stimulates the transcription of the proα1 and proα2 (Type I) collagen genes during tissue fibrosis? Secondly, how are the signals transmitted by the extracellular profibrotic cytokine TGF‐β from the cellular membrane to the nucleus for transcription of the proα1 (Type I) and proα2 (Type I) collagen genes? Thirdly, what signaling pathways cross‐talk with the signaling pathways resulting in the expression of the Type I collagen genes? Fourthly, how does TGF‐β affect extracellular matrix homeostasis? Fifthly, what are the nuclear factors corresponding to the DNA elements required for the promotion of the proα1 (Type I) and proα2 (Type I) collagen genes? Finally, how are the proα1 (Type I) and proα2 (Type I) collagen genes coordinately regulated? Strategies will also be presented for reducing fibrosis, which is the result of overexpression of TGF‐β. J. Cell. Biochem. 90: 1–5, 2003. © 2003 Wiley‐Liss, Inc.
Modulation of VE‐cadherin and PECAM‐1 mediated cell–cell adhesions by mitogen‐activated protein kinases
Authors: Wu Jianqiang , Sheibani Nader
Published: August 2003

Endothelial cell transition from a differentiated, quiescent phenotype to a migratory, proliferative phenotype is essential during angiogenesis. This transition is dependent on alterations in the balanced production of stimulatory and inhibitory factors, which normally keep angiogenesis in check. Activation of MAPK/ERKs is essential for endothelial cell migration and proliferation. However, its role in regulation of endothelial cell adhesive mechanisms requires further delineation. Here, we show that sustained activation of MAPK/ERKs results in disruption of cadherin‐mediated cell–cell adhesion, down‐regulation of PECAM‐1 expression, and enhanced cell migration in microvascular endothelial cells. Expression of a constitutively active MEK‐1 in mouse brain endothelial (bEND) cells resulted in down‐regulation of VE‐cadherin and catenins expression concomitant with down‐regulation of PECAM‐1 expression. In contrast, inhibition of MEK‐1 restored parental morphology, cadherin/catenins expression and localization. These data are further supported by our observation that sustained activation of MAPK/ERKs in phorbol myristate acetate incubated HUVEC lead to disruption of cadherin‐mediate cell–cell interactions and enhanced capillary formation on Matrigel. Thus, sustained activation of MAPK/ERKs plays an important role in disruption of cell–cell adhesion and migration of endothelial cells. J. Cell. Biochem. 90: 121–137, 2003. © 2003 Wiley‐Liss, Inc.
Skeletal muscle metabolism in physiology and in cancer disease
Authors: Giordano Anna , Calvani Menotti , Petillo Orsolina , Carteni' Maria , Melone Maria Rosa A.B. , Peluso Gianfranco
Published: August 2003

Skeletal muscle is a tissue of high demand and it accounts for most of daily energy consumption. The classical concept of energy metabolism in skeletal muscle has been profoundly modified on the basis of studies showing the influence of additional factors (i.e., uncoupling proteins (UCPs) and peroxisome proliferator activated receptors (PPARs)) controlling parameters, such as substrate availability, cellular enzymes, carrier proteins, and proton leak, able to affect glycolysis, nutrient oxidation, and protein degradation. This extremely balanced system is greatly altered by cancer disease that can induce muscle cachexia with significant deleterious consequences and results in muscle wasting and weakness, delaying or preventing ambulation, and rehabilitation in catabolic patients. J. Cell. Biochem. 90: 170–186, 2003. © 2003 Wiley‐Liss, Inc.
Pitavastatin alters the expression of thrombotic and fibrinolytic proteins in human vascular cells
Authors: Markle Ronald A. , Han Jihong , Summers Barbara D. , Yokoyama Toru , Hajjar Katherine A. , Hajjar David P. , Gotto Antonio M. , Nicholson Andrew C.
Published: August 2003

In addition to lowering blood lipids, clinical benefits of 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG Co‐A; EC 1.1.1.34) reductase inhibitors may derive from altered vascular function favoring fibrinolysis over thrombosis. We examined effects of pitavastatin (NK‐104), a relatively novel and long acting statin, on expression of tissue factor (TF) in human monocytes (U‐937), plasminogen activator inhibitor‐1 (PAI‐1), and tissue‐type plasminogen activator (t‐PA) in human aortic smooth muscle cells (SMC) and human umbilical vein endothelial cells (HUVEC). In monocytes, pitavastatin reduced expression of TF protein induced by lipopolysaccharide (LPS) and oxidized low‐density lipoprotein (OxLDL). Similarly, pitavastatin also reduced expression of TF mRNA induced by LPS. Pitavastatin reduced PAI‐1 antigen released from HUVEC under basal, OxLDL‐, or tumor necrosis factor‐alpha (TNF‐α)‐stimulated conditions. Reductions of PAI‐1 mRNA expression correlated with decreased PAI‐1 antigen secretion and PAI‐1 activity as assessed by fibrin–agarose zymography. In addition, pitavastatin decreased PAI‐1 antigen released from OxLDL‐treated and untreated SMC. Conversely, pitavastatin enhanced t‐PA mRNA expression and t‐PA antigen secretion in untreated OxLDL‐, and TNF‐α‐treated HUVEC and untreated SMC. Finally, pitavastatin increased t‐PA activity as assessed by fibrin–agarose zymography. Our findings demonstrate that pitavastatin may alter arterial homeostasis favoring fibrinolysis over thrombosis, thereby reducing risk for thrombi at sites of unstable plaques. J. Cell. Biochem. 90: 23–32, 2003. © 2003 Wiley‐Liss, Inc.
Identification of an IQGAP1/AKAP79 complex in β‐cells
Authors: Nauert J. Brian , Rigas Johanna D. , Lester Linda B.
Published: August 2003

IQGAP1, is a recently discovered scaffold protein proposed to regulate membrane cytoskeleton events through protein–protein interactions with F‐actin, E‐cadherin, β‐catenin, and CLIP170. The binding of IQGAP1 to its partners is regulated by calcium/calmodulin (Ca++/CaM) and the small molecular weight guanine nucleotide triphosphatases (GTPases), Cdc42, and Rac1. Here we identify a novel IQGAP1 scaffolding function by isolating the cyclic AMP dependent kinase (PKA) with IQGAP1. IQGAP1 was co‐purified with PKA using 5′‐cyclic AMP (cAMP) affinity chromatography and PKA activity was co‐immunoprecipitated with IQGAP1 using an anti‐IQGAP1 antibody. The association of IQGAP1 with PKA was shown to occur through a direct interaction between A kinase anchoring protein 79 (AKAP79) and the carboxyl‐terminal domain of IQGAP1. This suggests that cAMP/PKA may be coupled with Ca++/CaM and GTPases through an IQGAP1/AKAP79 complex. J. Cell. Biochem. 90: 97–108, 2003. © 2003 Wiley‐Liss, Inc.
Vitamin D and androgen regulation of prostatic growth
Authors: Leman Eddy S. , Arlotti Julie A. , Dhir Rajiv , Getzenberg Robert H.
Published: August 2003

Vitamin D has been reported to inhibit the growth of prostate cancer cells and model systems. In this study, we examined the interaction between 1,25‐dihydroxyvitamin D3 (1,25 D) in the presence or absence of endogenous testosterone on the growth and development of the adult rat prostate. Male Sprague–Dawley rats (165 days old) were either kept intact or castrated. Seven days after castration, the rats were treated with vehicle (control) or 1,25 D for 3 weeks and then sacrificed. Both ventral and dorsal lateral prostates were harvested; whole tissue lysates were collected and AR and VDR protein levels were analyzed by immunoblot analyses. Administration of 1,25 D in the intact animals decreased the prostatic size by 40%, compared to control animals, whereas 1,25 D did not influence the size of the prostate in castrated rats. 1,25 D administration in intact groups also increased both the AR and VDR protein levels by ∼twofold, whereas in castrated groups, 1,25 D only increased the AR protein level by 1.5–2.5‐fold. 1,25 D in the presence of endogenous testosterone inhibits prostatic growth, whereas 1,25 D in the absence of endogenous testosterone does not affect prostatic growth. The growth inhibitory activity of 1,25 D in the presence of testosterone may be mediated through the ligand activated AR and VDR pathways. These studies may reveal important information about the potential efficacy of 1,25 D as well as hormonal status in understanding the development of prostate diseases. J. Cell. Biochem. 90: 138–147, 2003. © 2003 Wiley‐Liss, Inc.
Estrogen regulates cytokine production and apoptosis in PMA‐differentiated, macrophage‐like U937 cells
Authors: Carruba Giuseppe , D'Agostino Pietro , Miele Monica , Calabrò Maurizio , Barbera Caterina , Bella Gloria Di , Milano Salvatore , Ferlazzo Viviana , Caruso Rosalba , Rosa Marzia La , Cocciadiferro Letizia , Campisi Ildegarda , Castagnetta Luigi , Cillari Enrico
Published: August 2003

We have investigated the effects of sex steroids, estradiol (E2), and testosterone (T) on the synthesis of tumor necrosis factor alpha (TNF‐α) and interleukin‐10 (IL‐10) in phorbol‐myristate‐acetate (PMA)‐differentiated human monoblastic U937 cells. The ability of both hormones to modulate the viability and programmed cell death of macrophage‐like PMA‐differentiated U937 cells was also inspected. E2 increased TNF‐α synthesis, whereas T had no effect on the production of this cytokine. The combination of E2 and its antagonist tamoxifen or ICI‐182,789 completely abolished the induction of TNF‐α, while combination of T and its antagonist Casodex (CSDX) did not significantly affect TNF‐α production by U937 cells. Exposure of cells to E2 resulted in a dose‐dependent decrease of IL‐10 synthesis, while again T did not show any detectable effect. In addition, E2 induced a significant increase of apoptosis in macrophage‐like U937 cells and this increase was inhibited by the simultaneous addition of either tamoxifen or ICI‐182. In contrast, T alone or in combination with CSDX did not modify apoptotic rates of U937 cells. This evidence, taken together, suggests that estrogens, but not androgens, exert a pro‐inflammatory action through the modulation of TNF‐α and IL‐10, and regulate the immune effector cells by the induction of programmed cell death. J. Cell. Biochem. 90: 187–196, 2003. © 2003 Wiley‐Liss, Inc.
Cyclic tensile stretch modulates proteoglycan production by intervertebral disc annulus fibrosus cells through production of nitrite oxide
Authors: Rannou François , Richette Pascal , Benallaoua Mourad , François Mathias , Genries Valérie , Korwin‐Zmijowska Carla , Revel Michel , Corvol Maité , Poiraudeau Serge
Published: August 2003

Degeneration of the intervertebral disc is the main pathophysiological process implicated in low back pain and is a prerequisite to disc herniation. Clinically, mechanical forces are important modulators of the degeneration, but the underlying molecular mechanism is not known and needs investigation to identify the biological target. The aim of this work was to study, at the molecular level, the effects of cyclic tensile stretch (CTS) on the production of proteoglycan by intervertebral disc annulus fibrosus cells since proteoglycans seem to be implicated in the dynamic process of intervertebral disc degeneration. Such cells of rabbit were cultured at high density on plates with a flexible bottom. CTS was applied with use of a pressure‐operated instrument to deform the plates. With CTS at 1% elongation (1 Hz frequency), the level of 35S‐labeled neosynthesized proteoglycans that accumulated in the cellular pool or were secreted in the culture medium did not change, but at 5% elongation, the level was significantly reduced after 8 h of stimulation (30 and 21%, respectively) and further reduced at 24 h (43 and 41%, respectively). Introducing the protein synthesis inhibitor cycloheximide had no effect on this result. Neither aggrecan and biglycan expression nor proteoglycan physical properties were modified. The level of nitrite oxide production significantly increased by 3.5 times after 8 h of 5% elongation. Introducing the nitric oxide synthase (NOS) inhibitors NG‐methyl‐l‐arginine or N‐omega nitro‐l‐arginine diminished the effects of CTS on the production of nitrite oxide and proteoglycans. By contrast, introducing N‐iminoethyl‐l‐lysine (a more specific inhibitor of inductible NOS [iNOS]) had little or no effect. Taken together, these results suggest that cNOS activation seems to be more implicated in the 5% CTS modulation of proteoglycan production than iNOS activation. These results suggest that CTS can help regulate the intervertebral disc matrix by decreasing proteoglycan production through a post‐translational regulation involving nitrite oxide. This result could be of interest in the development of local therapeutic strategies aimed at controlling intervertebral disc degeneration. J. Cell. Biochem. 90: 148–157, 2003. © 2003 Wiley‐Liss, Inc.
Identification and characterization of a novel enhancer for the human MCT‐1 oncogene promoter
Authors: Shi Bo , Levenson Victor , Gartenhaus Ronald B.
Published: August 2003

Cloning and characterization of the promoter region for the MCT‐1 oncogene is described. We used luciferase assays to identify cis‐acting elements responsible for human MCT‐1 promoter function. The MCT‐1 promoter is TATA‐less with a consensus initiator element located at the transcription start site and facilitated by two Sp1 sites that directs basal transcription. Deletion of a region of the MCT‐1 promoter (−133 to −122) resulted in significant decrease in luciferase activity, suggesting that this region contains a positive cis‐acting element. Using mobility shift assays with a 26‐mer oligonucleotide, which contains this fragment and its flanking regions, we demonstrated the presence of sequence‐specific DNA‐binding protein in both Jurkat and Hela nuclear extracts that we designated as LMBF (for lymphoid MCT‐1 binding factor). This 26‐mer oligonucleotide containing the LMBF binding site is required for maximum transcriptional activity of the MCT‐1 promoter. Although the 26‐mer oligonucleotide contains a sequence with strong homology to a heat‐shock factor consensus, competitive electrophoretic mobility shift assay (EMSA) analysis demonstrated that the binding protein is not a known member of heat shock family. Furthermore, this sequence when placed in reverse orientation downstream of the luciferase gene was able to enhance luciferase activity driven by a minimal promoter. These data are consistent with this sequence behaving as an enhancer. Finally, Southwestern blot analysis revealed a 96‐kDa protein capable of binding a probe containing the LMBF binding site. J. Cell. Biochem. 90: 68–79, 2003. © 2003 Wiley‐Liss, Inc.
Effects of 18β‐Glycyrrhetinic acid on the junctional complex and steroidogenesis in rat adrenocortical cells
Authors: Huang Shih‐Horng , Wu Jiahn‐Chun , Hwang Ra‐Der , Yeo Hui‐Lin , Wang Seu‐Mei
Published: August 2003

Cellular junctions play important roles in cell differentiation, signal transduction, and cell function. This study investigated their function in steroid secretion by adrenal cells. Immunofluorescence staining revealed the presence of gap junctions and adherens junctions between adrenal cells. The major gap junction protein, connexin43, was seen as a linear dotted pattern of the typical gap junction plaques, in contrast to α‐, β‐, and γ‐catenin, which were seen as continuous, linear staining of cell–cell adherens junction. Treatment with 18β‐glycyrrhetinic acid, a gap junction inhibitor, reduced the immunoreactivity of these proteins in a time‐ and dose‐dependent manner, and caused the gap junction and adherens junction to separate longitudinally from the cell–cell contact sites, indicating the structural interdependency of these two junctions. Interestingly, 18β‐glycyrrhetinic acid stimulated a two‐ to three–fold increase in steroid production in these adrenal cells lacking intact cell junctions. These data raise the question of the necessity for cell communication for the endocrine function of adrenal cells. Pharmacological analyses indicated that the steroidogenic effect of 18β‐glycyrrhetinic acid was partially mediated by extracellular signal‐related kinase and calcium/calmodulin‐dependent kinase, a pathway distinct from the protein kinase A signaling pathway already known to mediate steroidogenesis in adrenal cells. J. Cell. Biochem. 90: 33–41, 2003. © 2003 Wiley‐Liss, Inc.
Regulation of nitric oxide and bcl‐2 expression by shear stress in human osteoarthritic chondrocytes in vitro
Authors: Lee Mel S. , Trindade Michael C.D. , Ikenoue Takashi , Goodman Stuart B. , Schurman David J. , Smith Robert Lane
Published: August 2003

Onset and progression of cartilage degeneration is associated with shear stress occurring in diarthrodial joints subjected to inappropriate loading. This study tested the hypothesis that shear stress induced nitric oxide is associated with altered expression of regulatory onco‐proteins, bcl‐2, and Fas (APO‐1/CD95) and apoptosis in primary human osteoarthritic chondrocyte cultures. Shear stress induced membrane phosphatidylserine and nucleosomal degradation were taken as evidence of chondrocyte apoptosis. Application of shear stress upregulated nitric oxide in a dose‐dependent manner and was associated with increases in membrane phosphatidylserine and nucleosomal degradation. Increasing levels of shear stress decreased expression of the anti‐apoptotic factor, bcl‐2, from 44 to 10 U/ml. Addition of the nitric oxide antagonists, l‐N5‐(1‐iminoethyl) ornithine and Nω‐nitro‐l‐arginine methyl ester (L‐NAME), reduced shear stress induced nucleosomal degradation by 62% and 74%, respectively. Inhibition of shear stress induced nitric oxide release by l‐NAME coincided with a 2.7‐fold increase of bcl‐2, when compared to chondrocytes exposed to shear stress in the absence of l‐NAME. These data suggest that shear stress induced nitric oxide is associated with changes in apoptotic regulatory factors that alter chondrocyte metabolism and may contribute to joint degeneration. J. Cell. Biochem. 90: 80–86, 2003. © 2003 Wiley‐Liss, Inc.
TRPC3‐like protein is involved in the capacitative cation entry induced by 1α,25‐dihydroxy‐vitamin D3 in ROS 17/2.8 osteoblastic cells
Authors: Baldi Carolina , Vazquez Guillermo , Calvo Juan Carlos , Boland Ricardo
Published: August 2003

In ROS 17/2.8 rat osteoblastic‐like cells a capacitative Ca2+ entry (CCE) pathway operates which is activated by either 1α,25‐dihydroxy‐vitamin D3 (1α,25(OH)2D3) or thapsigargin (Tpg)‐induced depletion of Ca2+ stores (Baldi et al. [2002]: J. Cell. Biochem. 86:678–687). In view of recent evidence favoring a role for transient receptor potential (TRP) proteins in mediating CCE, we investigated if channels involved in the 1α,25(OH)2D3‐sensitive CCE in rat osteoblasts were related to an endogenous TRP‐canonical (TRPC) isoform homologue. By reverse transcription (RT)‐PCR using mRNA from ROS 17/2.8 cells and primers based on conserved regions within the mammalian TRPC3/6/7 subfamily, two fragments were amplified of 390 and 201 bp with 100 and 94% sequence identity, respectively, with human TRPC3. Northern blot analysis showed the presence of a 3.5 kb transcript and both immunobloting and immunocytochemistry using a specific anti‐TRPC3 antibody confirmed endogenous expression of a TRPC3‐like protein (∼110 kDa) with membrane localization. In ROS 17/2.8 cells intranuclearly microinjected with anti‐TRPC3 antisense oligodeoxynucleotides (ODN), both the initial rate and magnitude of CCE activated by either 1α,25(OH)2D3 or Tpg were markedly reduced, whereas no changes were detected in control‐injected cells. The present findings constitute the first evidence to date suggesting that an endogenous TRPC3‐like protein is functionally involved in the CCE route activated by 1α,25(OH)2D3 in a secosteroid target cell. We anticipate TRPC3 as a candidate for mediating store‐operated non‐selective cation entry into osteoblasts. J. Cell. Biochem. 90: 197–205, 2003. © 2003 Wiley‐Liss, Inc.
Activin A stimulates IκB‐α/NFκB and rank expression for osteoclast differentiation, but not AKT survival pathway in osteoclast precursors
Authors: Sugatani T. , Alvarez U.M. , Hruska K.A.
Published: August 2003

Recent studies have reported that activin A enhances osteoclastogenesis in cultures of mouse bone marrow cells stimulated with receptor activator of nuclear factor‐κB ligand (RANKL) and macrophage colony‐stimulating factor (M‐CSF). However, the exact mechanisms by which activin A functions during osteoclastogenesis are not clear. RANKL stimulation of RANK/TRAF6 signaling increases nuclear factor‐κB (NFκB) nuclear translocation and activates the Akt/PKB cell survival pathway. Here we report that activin A alone activates IκB‐α, and stimulates nuclear translocation of NFκB and receptor activator of nuclear factor‐κB (RANK) expression for osteoclastogenesis, but not Akt/PKB survival signal transduction including BAD and mammalian target of rapamycin (mTOR) for survival in osteoclast precursors in vitro. Activin A alone failed to activate Akt, BAD, and mTOR by immunoblotting, and it also failed to prevent apoptosis in osteoclast precursors. While activin A activated IκB‐α and induced nuclear translocation of phosphorylated‐NFκB, and it also enhanced RANK expression in osteoclast precursors. Moreover, activin A enhanced RANKL‐ and M‐CSF‐stimulated nuclear translocation of NFκB. Our data suggest that activin A enhances osteoclastogenesis treated with RANKL and M‐CSF via stimulation of RANK, thereby increasing the RANKL stimulation. Activin A alone activated the NFκB pathway, but not survival in osteoclast precursors in vitro, but it is, thus, insufficient as a sole stimulus to osteoclastogenesis. J. Cell. Biochem. 90: 59–67, 2003. © 2003 Wiley‐Liss, Inc.
Differentiation potential of a mouse bone marrow stromal cell line
Authors: Allan Elizabeth H. , Ho Patricia W.M. , Umezawa Akihro , Hata Jun‐Ichi , Makishima Fusao , Gillespie Matthew T. , Martin T. John
Published: August 2003

In order to study osteoblast differentiation we subcloned a cell derived from a mouse a bone marrow stromal cell line, Kusa O, and obtained a number of clones representative of three different phenotypes. One that neither differentiated into osteoblasts nor into adipocytes, a second that differentiated into osteoblasts but not adipocytes, and a third that differentiated into both osteoblasts and adipocytes. Four subclones were selected for further characterization according to their ability to mineralize and/or differentiate into adipocytes. The non‐mineralizing clone had no detectable alkaline phosphatase activity although some alkaline phosphatase mRNA was detected after 21 days in osteoblast differentiating medium. Alkaline phosphatase activity and mRNA in the three mineralizing clones were comparable with the parent clones. Osteocalcin mRNA and protein levels in the non‐mineralizing clone were low and non‐detectable, respectively, while both were elevated in the parent cells and mineralizing subclones after 21 days in differentiating medium. PTH receptor mRNA and activity increased in the four subclones and parent cells with differentiation. mRNA for two other osteoblast phenotypic markers, osteopontin and bone sialoprotein, were similarly expressed in the parent cells and subclones while mRNAs for the transcription factors, Runx2 and osterix, were detectable in both parent and subclone cells. Runx2 was unchanged with differentiation while osterix was increased. Interestingly, PPARγ mRNA expression did not correlate with cell line potential to differentiate into adipocytes. Indian hedgehog mRNA and its receptor (patched) mRNA levels both increased with differentiation while mRNA levels of the Wnt pathway components β‐catenin and dickkopf also increased with differentiation. Although we have focussed on characterizing these clones from the osteoblast perspective it is clear that they may be useful for studying both osteoblast and adipocyte differentiation as well as their transdifferentiation. J. Cell. Biochem. 90: 158–169, 2003. © 2003 Wiley‐Liss, Inc.
Mitogen‐activated protein kinase pathway mediates DBP–maf‐induced apoptosis in RAW 264.7 macrophages
Authors: Gumireddy Kiranmai , Reddy C. Damodar , Swamy Narasimha
Published: August 2003

Vitamin D‐binding protein–macrophage‐activating factor (DBP–maf) is derived from serum vitamin D binding protein (DBP) by selective deglycosylation during inflammation. In the present study, we investigated the effect of DBP–maf on RAW 264.7 macrophages and the underlying intracellular signal transduction pathways. DBP–maf increased proapoptotic caspase‐3, ‐8, and ‐9 activities and induced apoptosis in RAW 264.7 cells. However, DBP, the precursor to DBP–maf did not induce apoptosis in these cells. Cell cycle analysis of DBP–maf‐treated RAW 264.7 cells revealed growth arrest with accumulation of cells in sub‐G0/G1 phase. We also investigated the role of mitogen‐activated protein kinase (MAPK) pathways in the DBP–maf‐induced apoptosis of RAW264.7 cells. DBP–maf increased the phosphorylation of p38 and JNK1/2, while it decreased the ERK1/2 phosphorylation. Treatment with the p38 MAPK inhibitor, SB202190, attenuated DBP–maf‐induced apoptosis. PD98059, a MEK specific inhibitor, did not show a significant inhibition of apoptosis induced by DBP–maf. Taken together, these results suggest that the p38 MAPK pathway plays a crucial role in DBP–maf‐mediated apoptosis of macrophages. Our studies indicate that, during inflammation DBP–maf may function positively by causing death of the macrophages when activated macrophages are no longer needed at the site of inflammation. In summary, we report for the first time that DBP–maf induces apoptosis in macrophages via p38 and JNK1/2 pathway. J. Cell. Biochem. 90: 87–96, 2003. © 2003 Wiley‐Liss, Inc.