bluuebunny/crossref_metadata_2025 huggingface dataset:
Study of the stability of fuzzy controllers by an estimation ofthe attraction regions: A Vector Norm approach
Authors: Dieulot J.-Y. , El Kamel A. , Borne P.
Published: September 2003

A fuzzy controller with singleton defuzzification can be considered as the association of a regionwise constant term and of a regionwise non linear term, the latter being bounded by a linear controller. Based on the regionwise structure of fuzzy controller, the state space is partitioned into a series of disjoint sets. The fuzzy controller parameters are tuned in order to ensure that the ith set is included into the domain of attraction of the preceding sets of the series. If the first set of the series is included into the region of attraction of the equilibrium point, the overall fuzzy controlled system is stable. The attractors are estimated with the help of the comparison principle, using Vector Norms, which ensures the robustness with respect to uncertainties and perturbations of the open loop system.
Design of a decentralized detection of interacting LTI systems
Authors: Shankar Shamanth , Darbha Swaroop , Datta Aniruddha
Published: September 2003

In this paper, the problem of designing a decentralized detection filter for a large homogeneous collection of LTI systems is considered. The collection of systems considered here draws inspiration from platoons of vehicles, and the considered interactions amongst systems in the collection are banded and lower triangular, mimicking the typical “look‐ahead” nature of interactions in a platoon of vehicles. A fault in a system propagates to other systems in the collection via such interactions.

The decentralized detection filter for the collection is composed of interacting detection filters, one for each system. The feasibility of communicating the state estimates to other systems in the collection is assumed here. An important concern is the propagation of state estimation errors. In order that the state estimation errors not amplify as they propagate, a ℋ∞ constraint on the state estimation error propagation dynamics is imposed. A sufficient condition for constructing a decentralized detection filter for the collection is presented. An example is provided to illustrate the design procedure.
Schwartz′ distributions in nonlinear setting: Applications to differential equations, filtering and optimal control
Authors: Orlov Y.
Published: September 2003

The paper is intended to be of tutorial value for Schwartz′ distributions theory in nonlinear setting. Mathematical models are presented for nonlinear systems which admit both standard and impulsive inputs. These models are governed by differential equations in distributions whose meaning is generalized to involve nonlinear, non single‐valued operating over distributions. The set of generalized solutions of these differential equations is defined via closure, in a certain topology, of the set of the conventional solutions corresponding to standard integrable inputs. The theory is exemplified by mechanical systems with impulsive phenomena, optimal impulsive feedback synthesis, sampled‐data filtering of stochastic and deterministic dynamic systems.
Sliding mode control on electro‐mechanical systems
Authors: Utkin Vadim I. , Chang Hao-Chi
Published: September 2003

The first sliding mode control application may be found in the papers back in the 1930s in Russia. With its versatile yet simple design procedure the methodology is proven to be one of the most powerful solutions for many practical control designs. For the sake of demonstration this paper is oriented towards application aspects of sliding mode control methodology. First the design approach based on the regularization is generalized for mechanical systems. It is shown that stability of zero dynamics should be taken into account when the regular form consists of blocks of second‐order equations. Majority of applications in the paper are related to control and estimation methods of automotive industry. New theoretical methods are developed in the context of these studies: sliding made nonlinear observers, observers with binary
measurements, parameter estimation in systems with sliding mode control.
On fast path‐finding algorithms in AND‐OR graphs
Authors: Adelson-Velsky George M. , Gelbukh Alexander , Levner Eugene
Published: September 2003

We present a polynomial‐time path‐finding algorithm in AND‐OR graphs Given p arcs and n nodes, the complexity of the algorithm is O(np), which is superior to the complexity of previously known algorithms.
An LQG approach to systems with saturating actuators andanti‐windup implementation
Authors: Eun Y. , Gökçek C. , Kabamba P. T. , Meerkov S. M.
Published: September 2003

This paper extends the LQG design methodology to systems with saturating actuators and shows that resulting linear controllers do not require an anti‐windup implementation.
Conclusions from Two Model Concepts on Germinal Center Dynamics and Morphology
Authors: Meyer-Hermann Michael , Beyer Tilo
Published: September 2003

Germinal centers (GC) are an essential part of the humoral immune response. They develop a clear structure during maturation: Centroblasts and centrocytes are separated into two zones, the dark and the light zone. The mechanisms leading to this specific morphology as well as the reason for zone‐depletion during a later phase of the GC reaction have not clearly been revealed in experiment. We discuss and weigh possible mechanisms of dark and light zone development in the framework of two mathematical models. In a comparative approach we formulate constraints on typical lymphocyte velocities in GCs which are characteristic for the different proposed mechanisms.
Marginal Zone B Cells in Neonatal Rats Express Intermediate Levels of CD90 (Thy‐1)
Authors: Dammers Peter M. , Lodewijk Monique E. , Zandvoort André , Kroese Frans G. M.
Published: September 2003

Here we show that marginal zone (MZ)‐B cells in rats can already be detected in neonatal spleen from two days after birth. At this time point, morphologically distinct MZs are not present yet and the vast majority of B cells in spleen are located in a concentric area surrounding the T cell zones (PALS). Before MZs are obviously detectable in spleen (14 days after birth), MZ‐B cells seem to be enriched at the outer zones of the concentric B cell areas. Similar to adult rats, neonatal MZ‐B cells are intermediate‐sized cells that express high levels of surface (s)IgM and HIS57 antigen, and low levels of sIgD and CD45R (HIS24). We show here, however, that in contrast to adult MZ‐B cells, MZ‐B cells (and also recirculating follicular (RF)‐B cells) in neonatal rats express higher levels of CD90 (Thy‐1). In adult rats, expression of CD90 on the B cell lineage is confined to immature B cells. We speculate that the expression of CD90 on neonatal MZ‐B cells may have implications for their responsiveness to polysaccharide (T cell‐independent type 2) antigens.
Analysis of the IDDM Candidate Gene Prss16 in NOD and NON Mice
Authors: Cheunsuk Saijai , Hsu Tom , Gershwin M. Eric , Bowlus Christopher L.
Published: September 2003

The thymus‐specific serine protease Prss16 is highly expressed by the epithelial cells in the thymic cortex. It has been suggested to play an important role in the positive selection of T cells through the antigen presention pathway of the cortical antigen presenting cells. Recently, the gene ecoding Prss16 has been linked to insulin dependent diabetes mellitus (IDDM) susceptibility independent of HLA‐DR3 suggesting the Prss16 may be involved in the development of autoimmune disease. Due to the similarities of the gene structure and expression pattern between the human and mouse genes, we compared Prss16 between non‐obese diabetic (NOD) and non‐obese non‐diabetic (NON) mice. Analysis of the Prss16 coding region failed to identify any differences in sequence. Northern analysis and semi‐quantitative reverse transcriptase polymerase chain reaction showed that the mRNA was equal in size and abundance in the two strains. In situ hybridization showed similar patterns of staining. Therefore, our data suggests that there is no significant different in the gene structure, transcription level, and expression pattern of Prss16 gene between NOD and NON mice.
T Cells of Different Developmental Stages Differ in Sensitivity to Apoptosis Induced by Extracellular NAD
Authors: Haag Friedrich , Freese Dunja , Scheublein Felix , Ohlrogge Wiebke , Adriouch Sahil , Seman Michel , Koch-Nolte Friedrich
Published: September 2003

Extracellular nucleotides such as ATP and NAD can profoundly affect the functions of lymphocytes, macrophages, and other cells. We have recently shown that extracellular NAD induces rapid apoptosis in naive T cells by a mechanism involving the ADP‐ribosylation of cell surface molecules. In the present paper, we describe that T cells of different developmental stages differ in their sensitivity to NAD‐induced apoptosis. Thymocytes were less susceptible than peripheral lymph node T cells, and freshly activated cells were more resistant than resting cells. Sensitivity to NAD‐induced apoptosis generally correlated with expression of the ADP‐ribosyltransferase ART2.2, which is not expressed on thymocytes and shed from peripheral T cells upon activation. Our findings suggest that NAD‐induced apoptosis does not play a role during thymic selection of T cells, but rather may play a role by preventing the activation of unwanted bystander T cells during an immune response, and thus may participate in the control of autoimmunity.
Proliferative Responses of Harbor Seal (Phoca vitulina) T Lymphocytes to Model Marine Pollutants
Authors: Neale Jennifer C. C. , Van de Water Judith A. , Harvey James T. , Tjeerdema Ronald S. , Gershwin M. Eric
Published: September 2003

In recent years, population declines related to viral outbreaks in marine mammals have been associated with polluted coastal waters and high tissue concentrations of certain persistent, lipophilic contaminants. Such observations suggest a contributing role of contaminant‐induced suppression of cell‐mediated immunity leading to decreased host resistance. Here, we assessed the effects of the prototypic polycyclic aromatic hydrocarbon (PAH), benzo[a]pyrene (B[a]P), and two polychlorinated biphenyls (PCBs), CB‐156 and CB‐80, on the T‐cell proliferative response to mitogen in harbor seal peripheral lymphocytes. Despite the variability associated with our samples from free‐ranging harbor seals, we observed a clear suppressive effect of B[a]P (10 uM) exposure on T cell mitogenesis. Exposures to 10 uM CB‐156 and CB‐80, and 1.0 and 0.1 uM B[a]P, did not produce significant depression in lymphoproliferation. Exposure to the model PAH at 10 uM resulted in a 61% (range 34‐97%) average reduction in lymphoproliferation. We were able to rule out a direct cytotoxic effect of B[a]P, indicating that observed effects were due to altered T cell function. Based on our in vitro results, we hypothesize that extensive accumulation of PAH by top‐trophic‐level marine mammals could alter T cell activation in vivo and impaired cell‐mediated immunity against viral pathogens.
In Vivo ESR Study on Hepatic Reduction of a Nitroxide Radical after Administration of Glucose in Rats
Authors: Tada Mika , Yokoyama Hidekatsu , Toyoda Yoshiko , Ohya Hiroaki , Ogata Tateaki , Kamada Hitoshi
Published: September 2003

The in vivo reducing ability of a nitroxide radical, 4‐hydroxyl‐2,2,6,6‐tetramethylpiperidin‐1‐oxyl (TEMPOL), in the liver of rats was estimated by using an electron spin resonance spectrometer equipped with a surface‐coil‐type resonator following administration of glucose. Both oral and intravenous administration of glucose significantly enhanced the reduction capacity. In vitro studies show that the reduction site of TEMPOL in the hepatic homogenate is located in the mitochondrial respiratory chain and microsomal electron transport system. These findings indicate that the enhancement of activity of these systems caused by the glucose administration prompts reduction of TEMPOL in the liver.
Thiol‐Oxidizing Agent Diamide and Acidic pH Enhance Lipid Peroxidation of Rat Heart Mitochondria and Cardiolipin? Cytochrome c Complex
Authors: Iwase Hirotaro , Sakurada Koichi , Ikegaya Hiroshi , Hatanaka Kazuhito , Takeichi Hisako , Kobayashi Masahiko , Yoshida Kenichi
Published: September 2003

In this study, we investigated lipid peroxidation in rat heart mitochondria hydrolyzed by phospholipase A2 (PLA2) and lipid peroxidation in a mitochondrial‐mimetic lipid peroxidation system, where phospholipids such as cardiolipin (CL) and cytochrome c (Cyt c) were first mixed together and then PLA2 and calcium chloride were added to the mixture (CL‐Cyt c‐PLA2 system). Production of hydroperoxy and hydroxy compounds of linoleic acid (LA)1
Do Rodents Have a Gene Encoding Glycogenin‐2, the Liver Isoform of the Self‐Glucosylating Initiator of Glycogen Synthesis?
Authors: Zhai Lanmin , Schroeder Jill , Skurat Alexander V. , Roach Peter J.
Published: September 2003

The discovery of a second human gene, GYG2, encoding a liverspecific isoform of glycogenin, the self‐glucosylating initiator of glycogen biosynthesis, raised the possibility for differential controls of this protein in liver and muscle. The new protein, glycogenin‐2, had several properties similar biochemically to the muscle isoform, glycogenin‐1, but unlike glycogenin‐1, stable expression in fibroblasts led to a significant overaccumulation of glycogen. Ensuing attempts to generate reagents suitable for use with rodents, to examine the physiological regulation of glycogenin‐2 by nutritional and hormonal factors, have been unsuccessful. Proof of a negative is difficult but the weight of the evidence is beginning to mitigate against the existence of a second glycogenin gene in rodents leading us to hypothesize that the presence of the GYG2 gene is limited to primates.
Antioxidants in the Treatment of Graves Disease
Authors: Guerra Liliana N. , Moiguer Silvia , Karner Mirta , de Molina Ma del Carmen Rios , Sreider Claudia M. , Burdman José A.
Published: September 2003

An antioxidant mixture (LAROTABE) was evaluated in the treatment of Graves disease. Fifty‐six hyperthyroid patients were treated with methimazol (MMI) (A), LAROTABE (B), or MMI plus LAROTABE (C). According to a clinical score, improvement was obtained at 8 weeks in A and 4 weeks in B and C. Group A diminished their thyroid hormone concentration to normal levels, while patients with LAROTABE did not reduce T3 and T4 unless MMI was introduced. Hyperthyroid patients had increased malondialdehyde (MDA) content and SOD activity and decreased catalase activity compared to controls. Within group A, MDA decreased to control values while SOD was reduced 38.3% and catalase increased 21.6%. Similar results were obtained for MDA and for both enzymes after treatment with LAROTABE. Signs and symptoms of Graves disease might be related to an increase in free radicals; antioxidants could be a new therapeutic tool to improve the clinical manifestation of this illness.
Antioxidant Enzyme Activity and Oxidative Stress in Bovine Oocyte In Vitro Maturation
Authors: Cetica P. D. , Pintos L. N. , Dalvit G. C. , Beconi M. T.
Published: September 2003

Reactive oxygen species (ROS) production is a normal process of cell metabolism. In vitro environments usually increase cell production of ROS, which has been implicated as a main cause of cell damage. Nevertheless, the role of ROS in oocyte in vitro maturation (IVM) is controversial. In most cells, enzymatic antioxidant systems can attenuate the effect of oxidative stress by scavenging ROS. The aim of this work was to determine whether: (1) standard conditions of bovine oocyte IVM are responsible for oxidative stress; (2) cumulus cells participate in protection against oxidative stress of the oocyte; and (3) enzymatic antioxidant activity is present in oocytes and cumulus cells. Cumulus‐oocyte complexes (COCs) were matured in TCM‐199 10% steer serum for 24 h at 39 °C in 5% CO2:95% humidified air. Oxidative stress was determined by the 2', 7'‐dichlorofluorescein diacetate assay. Superoxide dismutase (SOD), glutathione peroxidase, and catalase activities were measured spectrophotometrically.Under standard conditions of in vitro maturation, there was no increase in ROS production per COC (P > 0.05), but ROS level per cumulus cell diminished. There was no modification in ROS levels in oocytes matured in the presence versus the absence of their surrounding cumulus cells (P > 0.05). To the best of our knowledge, the presence of SOD, glutathione peroxidase and catalase activities were detected in oocytes and cumulus cells for the first time. Enzymatic units were lower in denuded oocytes with respect to cumulus (P < 0.05), accounting for 37% for SOD, 25% for glutathione peroxidase, and 11% for catalase of the total COC units. Specific enzyme activity diminished in cumulus cells (P > 0.05) and increased in oocytes due to maturation (P > 0.05). The presence of activity of an enzymatic antioxidant system in the bovine oocyte would regulate in part ROS levels during IVM. Oocytes could be capable of controlling the increase in ROS because of the presence of their own enzymatic antioxidant system, SOD having the highest specific activity with respect to cumulus cells.
Transient Up‐Regulation of Liver Mitochondrial Thymidine Kinase Activity in Proliferating Mitochondria
Authors: Elholm Morten , HollÅs Hanne , Issalene Carol , Barroso Joao Filipe Viegas , Berge Rolf Kristian , Flatmark Torgeir
Published: September 2003

Administration of the fatty acid analogue tetradecylthioacetic acid (TTA) to rodents up‐regulates peroxisomal and mitochondrial lipid‐metabolizing enzymes and induces a proliferation of these organelles in hepatocytes. We show here that male NMRI mice fed a diet containing 0.3% (w/w) TTA revealed a transient two‐fold increase in the incorporation of [3
Intra‐ and Intercellular Distribution of Mitochondrial Probes and Changes after Treatment with MDR Modulators
Authors: Diaz Giacomo , Diana Andrea , Falchi Angela Maria , Gremo Fulvia , Pani Alessandra , Batetta Barbara , Dessì Sandra , Isola Raffaella
Published: September 2003

Fluorescent probes are currently used to evaluate the mitochondrial transmembrane potential in situ. However, in parallel experiments using the probes JC‐1 and TMRM in different cell types (human astrocytes, HEp‐2, Vero, KB, and HeLa cells), we found that the distribution of JC‐1 and TMRM is highly variable not only in different cell types but also in different cells of the same cell type, a condition that has never been documented until our work. This phenomenon depends on a hidden, widespread multidrug resistance (MDR) phenotype that can be recognized only by comparative assays with MDR inhibitors (progesterone, verapamil, and cyclosporin A) and represents a serious risk of error in the evaluation of the mitochondrial potential.
Internally Located Signal Peptides Direct Hepatitis C Virus Polyprotein Processing in the ER Membrane
Authors: Wu Jim Zhen
Published: September 2003

An endoplasmic reticulum (ER) signal peptide is an amino acid sequence motif that directs the translocation of nascent polypeptides to the lumen of ER membrane. Most of known ER signal peptides are either N‐terminal cleavable or internally uncleavable. In the structural protein region of hepatitis C virus (HCV) polyprotein, however, four internally located cleavable signal peptides are arranged in a tandem array. The published experimental results indicated that the nascent HCV polyprotein is processed in the ER membrane by host signal peptidase(s) to the respective viral proteins. Here we propose that the four ER signal peptides lead the nascent HCV polyprotein to ER membrane, and the four internally located cleavable signal peptides are the sole determinant for the compartment localization of the matured viral proteins. After cleavage at the C‐terminus, the signal peptides retain at the C‐terminus of mature proteins, and serve as ER membrane anchors. The signal peptide directed polyprotein processing in the ER membrane preludes the virion assembly and budding from the ER membrane. This unique processing may be a general mechanism adopted by many types of virus for virion assembly and replication. The revelation of signal peptidase involved in HCV polyprotein processing presents a novel drug target to suppress HCV viral replication for the much needed HCV therapy.
Precursor of a Novel Scorpion Venom Peptide (BmKn1) with no Disulfide Bridge from Buthus martensii Karsch
Authors: Zeng Xian‐Chun , Li Wen‐Xin , Wang San‐Xia , Zhu Shun‐Yi , Luo Feng
Published: September 2003

A full‐length cDNA sequence encoding the precursor of a novel venom peptide (named BmKn1) with no disulfide bridge was first isolated from the venom gland cDNA library of Chinese scorpion Buthus martensii Karsch. The encoded precursor consisted of 70 amino acid residues including two parts: a signal peptide of 23 residues, and a putative mature venom peptide (BmKn1) of 47 residues. The sequence of BmKn1 showed no similarity to those of other scorpion venom peptides. BmKn1 may be the first member of a new venom peptide family from scorpion. Future research will be interesting to unravel further the pharmacological function of this novel scorpion venom peptide.
Ironing Out Disease: Inherited Disorders of Iron Homeostasis
Authors: Anderson Gregory J.
Published: September 2003

Iron presents us with a paradox. Without it, cells simply cannot survive because iron is an essential cofactor for many enzymes in critical biochemical pathways. However, when iron is present in excess, it can be highly cytotoxic due to its propensity to catalyze the formation of reactive oxygen radicals. To cater for this dual nature, cells and organisms have developed elaborate mechanisms for regulating iron intake and efflux. When these mechanisms are disrupted, as is the case in a number of inherited disorders of iron metabolism, the pathological consequences can be severe. Many of these disorders are characterized by iron overload and include relatively common diseases such as hereditary hemochromatosis, rare abnormalities of plasma protein synthesis (atransferrinemia and aceruloplasminemia), and the neuromuscular disease Friedreich ataxia. The few described inherited anemias in humans have yet to yield to molecular dissection, but the investigation of several rodent anemias has proved highly rewarding. This review will provide a summary of some of these disorders and describe how their analysis has provided important new insights into iron trafficking pathways and their regulation.
Does Inhibition of Trypanosoma cruzi Key Enzymes Affect Parasite Life Cycle and Geographic Distribution?
Authors: Salvati Luca , Bolognesi Martino , Ascenzi Paolo
Published: September 2003

The hemoflagellate protozoan parasite Trypanosoma cruzi is the causative agent of the Chagas disease, a progressive fatal cardiomyopathy that afflicts more than 20 million people in Central and South America. The T. cruzi life cycle is affected by changes in temperature and pH, the parasite replication being facilitated in mammalian hosts rather than in insect vectors. Here, we postulate that the modulation of key enzymes by pH‐ and temperature‐dependent substrate inhibition may affect the T. cruzi life cycle and limit the geographic range covered by the parasite.
Isosteric Replacement of A3 Val of Porcine Insulin by allo‐Thr
Authors: Li Moyi , Cui Dafu , Zhang Youshang
Published: September 2003

A3 Val is important for insulin activity. It is invariant in insulins from different species studied thus far. Based on the three dimensional structure of insulin, it was thought to be involved in receptor binding. Its replacement by Leu resulted in remarkable lowering of insulin activity, indicating the crucial requirement of the side chain geometry at this position. When A3 Val was replaced by Thr, which is hydrophilic but isosteric with Val, substantial insulin activity was retained. Therefore, the isosteric requirement for receptor binding at this site is more stringent than the hydrophobic requirement. Here we report the replacement of A3 Val of porcine insulin by the unnatural allo‐Thr. The in vivo biological activity of A3 allo‐Thr insulin is similar to that of A3 Thr insulin or native insulin, but its receptor binding activity is 7.6% instead of 50% for A3 Thr insulin, indicating that at the A3 position the hydrophilic OH group of Thr could be more tolerated in receptor binding than the OH group of allo‐Thr. The retention of insulin activity by substituting A3 Val with the unnatural isosteric allo‐Thr demonstrates again the importance of isosteric interaction in the binding of insulin with its receptor.
The Structure and Function of Catalytic Domains Within Inositol Polyphosphate 5‐Phosphatases
Authors: Whisstock J. C. , Wiradjaja F. , Waters J. E. , Gurung R.
Published: September 2003

Phosphoinositide signaling pathways regulate many essential cellular functions including proliferation, differentiation and survival, cytoskeletal organization, and vesicular trafficking. The inositol polyphosphate 5‐phosphatases regulate the cellular levels of several bioactive phosphoinositide species. This review describes the structure and function of the 5‐phosphatase and Sac1 catalytic domains of these enzymes. The crystal structure of the 5‐phosphatase domain has been solved and shares homology with members of the AP endonuclease family. The phosphoinositide polyphosphatase activity of the Sac1 domain, found in some inositol polyphosphate 5‐phosphatases, is defined by a motif, CX 5 R(T/S), also found in both protein and lipid phosphatases.
Structure and Function of Collagen‐Derived Endostatin Inhibitors of Angiogenesis
Authors: Sasaki Takako , Timpl Rupert , Hohenester Erhard
Published: September 2003

Endostatins are inhibitors of endothelial cell migration and angiogenesis and have been shown to reduce tumor growth in animal models. They are derived from the nontriplehelical C ‐terminal NC1 domains of collagens XV and XVIII, which are released proteolytically in trimeric form and further converted to monomeric endostatins of about 20 kDa. Both endostatin isoforms share a compact globular fold, but differ in certain binding properties for proteins and cells, as well as in tissue distribution. Differences in activity were found between NC1 domains and endostatins and are related to the oligomerization state. Endostatin effects are not restricted to endothelial cells, but also control renal epithelial cells and neuronal guidance in C. elegans . Cellular receptors are still insufficiently characterized and include for endostatin‐XVIII heparan sulfate proteoglycans. Receptor engagement elicits various downstream effects including tyrosine kinase and gene activation. Much remains to be learned, however, about details of the signal transduction cascades and how they interfer with pro‐angiogenic factors under physiological conditions and during therapeutic treatment.
Structural Basis of Perturbed pKa Values of Catalytic Groups in Enzyme Active Sites
Authors: Harris Thomas K. , Turner George J.
Published: September 2003

In protein and RNA macromolecules, only a limited number of different side‐chain chemical groups are available to function as catalysts. The myriad of enzyme‐catalyzed reactions results from the ability of most of these groups to function either as nucleophilic, electrophilic, or general acid‐base catalysts, and the key to their adapted chemical function lies in their states of protonation. Ionization is determined by the intrinsic p K a of the group and the microenvironment created around the group by the protein or RNA structure, which perturbs its intrinsic p K a to its functional or apparent p K a . These p K a shifts result from interactions of the catalytic group with other fully or partially charged groups as well as the polarity or dielectric of the medium that surrounds it. The electrostatic interactions between ionizable groups found on the surface of macromolecules are weak and cause only slight p K a perturbations (<2 units). The sum of many of these weak electrostatic interactions helps contribute to the stability of native or folded macromolecules and their ligand complexes. However, the p K a values of catalytic groups that are found in the active sites of numerous enzymes are significantly more perturbed (>2 units) and are the subject of this review. The magnitudes of these p K a perturbations are analyzed with respect to the structural details of the active‐site microenvironment and the energetics of the reactions that they catalyze.
The Focal Adhesion Kinase‐‐A Regulator of Cell Migration and Invasion
Authors: Hauck Christof R. , Hsia Datsun A. , Schlaepfer David D.
Published: September 2003

Cell migration plays an important role in embryonic development, wound healing, immune responses, and in pathological phenomena such as tissue invasion and metastasis formation. In this review, we summarize recent reports that connect the focal adhesion kinase (FAK) to cell migration and invasion. FAK is a nonreceptor protein tyrosine kinase involved in signal transduction from integrin‐enriched focal adhesion sites that mediate cell contact with the extracellular matrix. Multiple protein‐protein interaction sites allow FAK to associate with adapter and structural proteins allowing for the modulation of mitogen‐activated protein (MAP) kinases, stress‐activated protein (SAP) kinases, and small GTPase activity. FAK‐enhanced signals have been shown to mediate the survival of anchorage‐dependent cells and are critical for efficient cell migration in response to growth factor receptor and integrin stimulation. Elevated expression of FAK in human tumors has been correlated with increased malignancy and invasiveness. Because recent findings show that FAK contributes to the secretion of matrix‐metalloproteinases, FAK may represent an important checkpoint in coordinating the dynamic processes of cell motility and extracellular matrix remodeling during tumor cell invasion.
Duchenne and Becker Muscular Dystrophy: From Gene Diagnosis to Molecular Therapy
Authors: Matsuo Masafumi
Published: September 2003

Duchenne and Becker muscular dystrophy (DMD/BMD) are X‐linked muscular dystrophies. The isolation of the defective gene in DMD/BMD has led to a better understanding of the disease process and has promoted studies regarding the application of molecular therapy. The purpose of this review is to present the progress made in this area of research with particular reference to dystrophin Kobe. Based on the results from the molecular analysis of dystrophin Kobe, we propose a novel molecular therapeutic method for DMD in which antisense oligonucleotides transform DMD into a milder phenotype by inducing exon skipping. In addition, current proposals for the molecular therapy of DMD are discussed.
Synergistic Effect of Vitamin D Derivatives and Retinoids on C2C12 Skeletal Muscle Cells
Authors: Stio Maria , Celli Alessandra , Treves Cristina
Published: September 2003

The response of C 2 C 12 myoblasts to 1 nM 1,25‐dihydroxyvitamin D 3 [1,25(OH) 2 D 3 ], two vitamin D analogues (KH 1060 and EB 1089, which are 20‐epi‐22‐oxa and 22,24‐diene‐analogues, respectively), 100 nM retinoids (9‐ cis retinoic acid, all‐ trans retinoic acid) and to combination treatments, after 72 h incubation, was studied. The incubation with 1,25(OH) 2 D 3 was ineffective on either cell proliferation or [ 3 H]thymidine incorporation (expressed as DPM per cell) or protein content per cell. On the contrary, all the other treatments inhibited cell proliferation, this inhibition being synergistic when the vitamin D derivatives were combined with 9‐ cis or all‐ trans retinoic acid, and increased [ 3 H]thymidine incorporation and protein content per cell. The levels of the VDR protein remarkably increased in comparison with control cells, except for the incubation with 9‐ cis retinoic acid. This increase was particularly accentuated in C 2 C 12 cells treated with KH 1060 and 9‐ cis retinoic acid in combination. These results, taken together, suggest a role for vitamin D derivatives and retinoids on C 2 C 12 cells.
Role of Ca2+‐Dependent Metalloprotease‐2 in Stimulating Ca2+ATPase Activity Under Peroxynitrite Treatment in Bovine Pulmonary Artery Smooth Muscle Membrane
Authors: Chakraborti Tapati , Das Sudip , Mandal Malay , Mandal Amritlal , Chakraborti Sajal
Published: September 2003

We have determined effect of the oxidant peroxynitrite (ONOO ‐) on Ca 2+ ‐dependent matrix metalloprotease‐2 (MMP‐2) activity and the role of the protease on Ca 2+ ATPase activity in bovine pulmonary vascular smooth muscle plasma membrane under ONOO ‐‐triggered conditions. The smooth muscle plasma membrane possesses a 72‐kDa protease activity in a gelatin‐containing zymogram. The 72‐kDa protease activity has been found to be inhibited by tissue inhibitor of metalloprotease‐2 (TIMP‐2), indicating that the protease is the matrix metalloprotease‐2 (MMP‐2). Treatment of the membrane suspension with ONOO ‐ caused stimulation of the MMP‐2 activity (as evidenced by 14 C‐gelatin degradation) and also increased Ca 2+ ATPase activity. The ONOO ‐‐triggered protease activity and the Ca 2+ ATPase activity were found to be inhibited by the antioxidants: vitamin E, thiourea, and mannitol. Pretreatment with catalase and superoxide dismutase did not significantly alter ONOO ‐‐stimulated MMP‐2 activity and Ca 2+ ATPase activity, indicating that peroxide and superoxide are not present in appreciable amount in ONOO ‐. Under both basal and ONOO ‐ triggered conditions, the MMP‐2 activity and the Ca 2+ ATPase activity were also inhibited by EGTA, 1:10‐phenanthroline, and TIMP‐2. However, the ONOO ‐‐stimulated MMP‐2 activity and the Ca 2+ ATPase activity were found to be insensitive to phenylmethylsulfonylfluoride, Bowman‐Birk inhibitor, chymostatin, leupeptin, antipain, N ‐ethylmaleimide, and pepstatin. These results suggest that ONOO ‐ caused stimulation of MMP‐2 activity and that the increased MMP‐2 activity subsequently played a pivotal role in stimulating Ca 2+ ATPase activity in bovine pulmonary vascular smooth muscle plasma membrane.
HIV Epidemics in Africa: What Explains the Variations in HIV Prevalence?
Authors: Buvé Anne
Published: September 2003

There are large differences in the prevalence of HIV infection between different regions in sub‐Saharan Africa, ranging from less than 10% in pregnant women in most of West Africa, to over 25% in pregnant women in large cities in Eastern and Southern Africa. These differences in HIV prevalence are in many instances due to differences in rate of spread of the virus. The multicenter study on factors determining the differential spread of HIV in four African cities tried to identify factors that could explain differences in spread of HIV between different regions in sub‐Saharan Africa. The study was conducted in four cities, including two cities with a relatively low HIV prevalence (Cotonou in Bénin and Yaoundé in Cameroon) and two cities with a high HIV prevalence (Kisumu in Kenya and Ndola in Zambia). The difference in HIV prevalence between the four cities could not be explained by differences in sexual behavior. Any differences in sexual behavior were outweighed by differences in factors that influence HIV transmission, i.e. male circumcision and HSV‐2 infection. These findings have important implications for the design of interventions.
Kaposi's Sarcoma Biology
Authors: Boshoff Chris
Published: September 2003

Cancer remains a major burden for HIV‐infected individuals. The majority of cancers associated with HIV infection are driven by oncogenic viruses like Epstein‐Barr virus, Kaposi's sarcoma‐associated herpesvirus and human papillomavirus. Kaposi's sarcoma is a tumour of endothelium and is the most common malignancy in HIV infected individuals. AIDS‐related cancers are providing critical insight into cancer immunity and viral oncogenesis.
The Development of HIV‐1 Subtype C Vaccines for Southern Africa
Authors: Williamson Anna‐Lise
Published: September 2003

One in nine people in South Africa are estimated to be HIV‐1 infected, with the majority of these infections being due to HIV‐1 subtype C. Until recently, most HIV‐1 candidate vaccines were not based on subtype C genes. In response to this epidemic, therefore, the South African AIDS Vaccine Initiative (SAAVI) was established to facilitate the development and testing of candidate HIV‐1 subtype C vaccines. The first HIV‐1 subtype C candidate vaccine is due to be tested at the end of 2002, and is based on Venezuelan encephalitis virus replicons expressing Gag protein. The next candidate vaccines to be tested will be DNA and modified vaccinia Ankara vaccines expressing subtype C genes.
Transcriptomics and Proteomics: Tools for the Identification of Novel Drug Targets and Vaccine Candidates for Tuberculosis
Authors: Betts Joanna C.
Published: September 2003

The availability of the complete genome sequence of Mycobacterium tuberculosis has revolutionised many areas of tuberculosis research and facilitated functional genomics studies. Most notably, transcriptomics and proteomics have become important tools in gaining increased understanding of the biology of M. tuberculosis and offer the promise of being able to deliver novel drug targets and vaccine candidates.
Molecular Epidemiology of TB: Challenging Dogmas and Asking New Questions
Authors: van Helden Paul D.
Published: September 2003

Traditional epidemiological methods provide insight into the dynamics of diseases such as tuberculosis. These traditional techniques have limitations and rely on a number of assumptions. The application of molecular techniques to the study of epidemiology has allowed us to gain new insights into the biology of the organism Mycobacterium tuberculosis and the dynamics of the disease. We have been enabled to push the limits of understanding of the epidemiology of this disease, allowing us to challenge the old clinical dogmas, ask new questions, design new strategies, and measure the efficacy of such new interventions to combat this age‐old scourge. Among the dogmas challenged are that infection outside the home is commonplace, so‐called relapse cases may in fact be largely reinfection, and active transmission may be more common than previously thought and reactivation disease relatively uncommon. These findings alone demand urgent attention and the design of optimal intervention strategies to reduce the burden of disease.
A Chemical Approach Towards Understanding the Mechanism and Reversal of Drug Resistance in Plasmodium falciparum: Is it Viable?
Authors: Chibale Kelly
Published: September 2003

Genetic and biochemical approaches to studies of drug resistance mechanisms in Plasmodium falciparum have raised controversies and contradictions over the past several years. A different and novel chemical approach to this important problem is desirable at this point in time. Recently, the molecular basis of drug resistance in P. falciparum has been associated with mutations in the resistance genes, Chloroquine Resistance Transporter (PfCRT) and the P‐glycoprotein homologue (Pgh1). Although not the determinant of chloroquine resistance in P. falciparum, mutations in Pgh1 have important implications for resistance to other antimalarial drugs. Because it is mutations in the aforementioned resistance genes rather than overexpression that has been associated with drug resistance in malaria, studies on mechanisms of drug resistance and its reversal by chemosensitisers should benefit from a chemical approach. Target‐oriented organic synthesis of chemosensitisers against proteins implicated in drug resistance in malaria should shed light on mechanism of drug resistance and its reversal in this area. The effect of structurally diverse chemosensitisers should be examined on several putative resistance genes in P. falciparum to deal with antimalarial drug resistance in the broadest sense. Therefore, generating random mutations of these resistance proteins and subsequent screening in search of a specific phenotype followed by a search for mutations and/or chemosensitisers that affect a specific drug resistance pathway might be a viable strategy. This diversity‐oriented organic synthesis approach should offer the means to simultaneously identify resistance proteins that can serve as targets for therapeutic intervention (therapeutic target validation) and chemosensitisers that modulate the functions of these proteins (chemical target validation).
Neutralizing Antibody Responses to HIV‐1 Infection
Authors: Morris Lynn
Published: September 2003

Neutralizing antibodies represent an important component of immune control in many viral infections. In HIV‐1 infection, almost all individuals develop antibodies capable of neutralizing autologous viruses in vitro; however, the role of these antibodies in vivo still remains unclear. Their absence during the acute phase of infection, when the viral levels are brought under control, suggests they play a minor role in immune control and that cellular immune responses are more critical during this time. However, during chronic infection these antibodies may be important in preventing cell‐to‐cell spread and they still represent our best hope of providing sterilizing immunity (i.e., prevention of infection) by vaccination. Significant advances over the last few years in understanding the structure of the envelope glycoproteins have renewed interest in the role of neutralizing antibodies and the possibility that immunogens capable of stimulating a neutralizing antibody response can be developed.
Nutrient‐Gene Interactions in Mitochondrial Function: Vitamin A Needs Are Increased in BHE/Cdb Rats
Authors: Everts Helen B. , Berdanier Carolyn D.
Published: September 2003

The BHE/Cdb rat has a maternally inherited mutation in the ATPase 6 mitochondrial gene that associates with impaired oxidative phosphorylation (OXPHOS) and glucose intolerance. A longevity study revealed that feeding an egg‐rich (vitamin A‐rich) diet delayed the onset of impaired glucose tolerance. Two experiments were conducted to test the hypothesis that BHE/Cdb rats require more dietary vitamin A than normal rats. Experiment 1 was a dose‐response study examining OXPHOS in BHE/Cdb rats fed one of six levels of vitamin A. In experiment 2 BHE/Cdb and Sprague‐Dawley rats were used. The rats were depleted of retinol stores, then repleted with 4 or 12 IU vitamin A/g diet. Vitamin A status was assessed in depleted, never depleted, and depleted/repleted rats. OXPHOS was optimized at 4 IU/g diet for the Sprague‐Dawley rats and 12 IU/g diet for the BHE/Cdb rats. These results suggested that the criteria for vitamin intake adequacy in the BHE/Cdb rats is the optimization of mitochondrial OXPHOS. Using this criteria, we conclude that diabetes‐prone BHE/Cdb rats require more dietary vitamin A than normal rats.
Nutrient‐Gene Interactions: Dietary Vitamin A and Mitochondrial Gene Expression
Authors: Everts Helen B. , Claassen Daniel O. , Hermoyian Christina L. , Berdanier Carolyn D.
Published: September 2003

The BHE/Cdb rat is a model for mitochondrial diabetes due to a mutation in the ATPase 6 gene. These rats require more dietary vitamin A to optimize mitochondrial function than do normal Sprague‐Dawley rats. To determine a possible mechanism for this effect, cultured hepatocytes and hepatic tissues were studied. ATPase 6 (F 0 ATPase subunit a), retinoic acid receptors (RARs), and mitochondrial transcription factor A (mtTFA) gene products were determined using Western blot analysis. Northern analysis was used to determine ATPase 6, ATPase 6,8, and ND1 mRNA. Mitochondrial density was determined using confocal microscopy. Dose response studies using primary hepatocyte cultures showed that both ATPase 6 gene product and mRNA were optimized with additions of 10 ‐9 M retinoic acid. Retinoic acid receptors were found in the mitochondrial compartment. MtTFA levels were increased by vitamin A. Mitochondrial density was greater in the BHE/Cdb tissue than in Sprague‐Dawley tissue. These results show that vitamin A affects mitochondrial function via an effect on both nuclear and mitochondrial encoded genes.
Protease‐Activated Receptors: A Means of Converting Extracellular Proteolysis into Intracellular Signals
Authors: Mackie E. J. , Pagel C. N. , Smith R. , de Niese M. R. , Song S. ‐ J. , Pike R. N.
Published: September 2003

Protease‐activated receptors (PARs) mediate cellular responses to a variety of extracellular proteases. The four known PARs constitute a subgroup of the family of seven‐transmembrane domain G protein‐coupled receptors and activate intracellular signalling pathways typical for this family of receptors. Activation of PARs involves proteolytic cleavage of the extracellular domain, resulting in formation of a new N terminus, which acts as a tethered ligand. PAR‐1, ‐3, and ‐4 are relatively selective for activation by thrombin whereas PAR‐2 is activated by a variety of proteases, including trypsin and tryptase. Recent studies in mice genetically incapable of expressing specific PARs have defined roles for PAR‐1 in vascular development, and for PAR‐3 and ‐4 in platelet activation, which plays a fundamental role in blood coagulation. PAR‐1 has also been implicated in a variety of other biological processes including inflammation, and brain and muscle development. Responses mediated by PAR‐2 include contraction of intestinal smooth muscle, epithelium‐dependent smooth muscle relaxation in the airways and vasculature, and potentiation of inflammatory responses. The area of PAR research is rapidly expanding our understanding of how cells communicate and control biological functions, in turn increasing our knowledge of disease processes and providing potential targets for therapeutic intervention.
What Nuclease Cleaves Pre‐mRNA in the Process of Polyadenylation?
Authors: Zarudnaya Margarita I. , Kolomiets Iryna M. , Hovorun Dmytro M.
Published: September 2003

A transcript‐specific cleavage by a large set of proteins is the first stage of eukaryotic pre‐mRNA polyadenylation. The main participant of this reaction‐‐endonuclease‐‐has not been discovered until now. However, mammalian CPSF‐30 and yeast Yth 1p proteins are known to be homologues to Drosophila Clipper (CLP) protein, which possesses endoribonucleolytic activity. In the N ‐terminal region, all three proteins contain five copies of the CCCH zinc finger motif associated with nucleolytic activity in the case of CLP. The literature data on these proteins are reviewed here. These data were shown not to contradict the hypothesis that CPSF‐30 and its homologues are the actual nucleases that cleave pre‐mRNA in the process of polyadenylation.
Direct Visualization by Confocal Fluorescent Microscopy of the Permeation of Myristoylated Peptides Through the Cell Membrane
Authors: Enseñat‐Waser Roberto , Martin Franz , Barahona Fernando , Vazquez Jesús , Soria Bernat , Reig Juan A.
Published: September 2003

To study the permeability through the cellular membrane of synthetic peptides containing an hydrofobic moiety, we used a 13‐mer myristoylated peptide labeled with a N ‐terminal fluorescent probe. After 2 h of incubation, the subcellular distribution was analyzed in intact chromaffin cells by confocal fluorescent microscopy. Our results demonstrate that myristoylated peptides diffuse into intact cells, showing an heterogeneous distribution, but they do not reach the cellular nucleous, at least during the time range used.