bluuebunny/crossref_metadata_2025 huggingface dataset:
Cellular studies of auditory hair cell regeneration in birds
Authors: Stone Jennifer S. , Rubel Edwin W
Published: July 2002

A decade ago it was discovered that mature birds are able to
regenerate hair cells, the receptors for auditory perception. This
surprising finding generated hope in the field of auditory neuroscience
that new hair cells someday may be coaxed to form in another class of
warm-blooded vertebrates, mammals. We have made considerable progress
toward understanding some cellular and molecular events that lead to
hair cell regeneration in birds. This review discusses our current
understanding of avian hair cell regeneration, with some comparisons to
other vertebrate classes and other regenerative systems.
The subcellular localization of acetyl-CoA carboxylase 2
Authors: Abu-Elheiga Lutfi , Brinkley William R. , Zhong Ling , Chirala Subrahmanyam S. , Woldegiorgis Gebretateos , Wakil Salih J.
Published: July 2002

Animals, including humans, express two isoforms of acetyl-CoA carboxylase (EC
6.4.1.2
), ACC1 (
M
r
= 265 kDa) and ACC2 (
M
r
= 280 kDa). The predicted amino acid sequence of ACC2 contains an additional 136 aa relative to ACC1, 114 of which constitute the unique N-terminal sequence of ACC2. The hydropathic profiles of the two ACC isoforms generally are comparable, except for the unique N-terminal sequence in ACC2. The sequence of amino acid residues 1–20 of ACC2 is highly hydrophobic, suggesting that it is a leader sequence that targets ACC2 for insertion into membranes. The subcellular localization of ACC2 in mammalian cells was determined by performing immunofluorescence microscopic analysis using affinity-purified anti-ACC2-specific antibodies and transient expression of the green fluorescent protein fused to the C terminus of the N-terminal sequences of ACC1 and ACC2. These analyses demonstrated that ACC1 is a cytosolic protein and that ACC2 was associated with the mitochondria, a finding that was confirmed further by the immunocolocalization of a known human mitochondria-specific protein and the carnitine palmitoyltransferase 1. Based on analyses of the fusion proteins of ACC–green fluorescent protein, we concluded that the N-terminal sequences of ACC2 are responsible for mitochondrial targeting of ACC2. The association of ACC2 with the mitochondria is consistent with the hypothesis that ACC2 is involved in the regulation of mitochondrial fatty acid oxidation through the inhibition of carnitine palmitoyltransferase 1 by its product malonyl-CoA.
Organizational communication and the symbolic construction of police murder investigations 1
Authors: Innes Martin
Published: July 2002

This paper draws upon a range of empirical data to consider the ways in which police murder investigations are symbolically constructed, both within and outside of the police organization. It is argued that a range of communicative formats serve to produce the activities associated with police murder investigations in a way that serves to legitimate the police function to both members of the public and police officers alike. A particular emphasis is placed upon understanding the connections between informal and formal communications, and the instrumental and expressive objectives that variously underpin them.
Culture in the Mediation of International Disputes
Authors: Carnevale Peter J. , Choi Dong‐Won
Published: July 2002

Unlike most treatments of culture in international diplomacy, this article suggests that culture can play a positive role in the mediation of international disputes. Cultural ties between the mediator and one or both of the disputants can facilitate mediation by, among other things, enhancing the mediator's acceptability to the parties, and enhancing the belief that the mediator can deliver concessions and agreements. Moreover, a mediator who is closer to one side than the other can be effective in mediation, especially when the mediator acts in an even‐handed manner. Data from laboratory research on mediation, as well as anecdotal evidence, support this view.
Local Economic Development: A Review and Assessment of its Current Status in South Africa
Authors: Nel Etienne
Published: July 2002

In recent years, local economic development (LED) has become a widely practised development strategy in the countries of the North at both the local government and community levels. LED is less widely implemented in the South where, in most instances, it appears to be still in an incipient phase. This paper investigates the current status of LED in South Africa, where, over the past decade, local governments, community groups and non-governmental organisations (NGOs) have become significantly more active in locality-based economic development. Several local governments have established comprehensive LED programmes including the establishment of LED units and the pursuit of a range of developmental strategies, whilst in parallel, an array of community and NGO initiatives are in place. In almost all cases, however, results are still of a rather limited nature and this paper assesses some of the reasons for this situation.
The Effects of the Crisis of 1994/95 on the Mexican Labour Market: The Case of the City of Puebla
Authors: Fuchs Martina
Published: July 2002

In 1994/95, the economic crisis in Mexico strongly affected not only the Mexican economy but also the everyday lives of the Mexican population. The weaker sectors of the economy and the poorer segments of the Mexican population were particularly affected. This paper attempts to link these global trends to a specific location: Mexico's fourth-largest city, Puebla. In order better to understand the relationship between international developments, like a financial crisis, and local issues, like employment and the household economy, relevant aspects of the political and social spheres must be considered. This relationship will be discussed on a theoretical basis employing regulation theory. The paper proposes that regulation is not only important at the macro level (government, parties and trade unions), but also at the micro level (households). Puebla serves as an example useful for portraying the conditions of human labour and the strategies of survival of people as 'micro regulation' in a Mexican urban environment. The households suffered greatly during the economic crisis in Mexico because the major actors in the government and the administration did not support the people very effectively.
Family Structure, Educational Achievement and the Inner City
Authors: Gordon Ian
Published: July 2002

Current rates of success in the GCSE exams (taken at age 16 years in England and Wales) are particularly associated with location away from the inner cities. This paper uses aggregate data on success rates and socio-economic characteristics at the level of LEAs to assess the contributions of social, economic and schooling factors to this pattern of spatial disparity. Regression analyses indicate that the strongest influence is the spatial distribution of lone-parent families, and that rates of school absenteeism are an important intervening variable. The strength of the geographical relationship reflects the fact that British inner city areas are now distinguished more by the prevailing family structure than by their ethnic or class mix, or their level of unemployment. While high levels of (male) unemployment contribute to this situation, it is argued that more attention ought to be given to the social dimension of urban problems and policy than has been in the past 20 years.
Congestion and Safety on Highways: Towards an Analytical Model
Authors: Shefer Daniel , Rietveld Piet
Published: July 2002

Congestion and accidents are important components of the externalities created by road users in metropolitan areas. In the present paper, we investigate the relationship between these two components. Among the factors which influence the number of fatalities on highways are: speed, speed differences and traffic composition. We pay special attention to the impact of congestion on the number of fatalities. The lower speeds which are caused by congestion would lead to lower numbers of fatal accidents. As a result, we expect a parabolic relationship between density and fatal accidents on highways. When densities increase, we would first have a positive relationship due to the increase in the numbers of cars in the system. However, when density becomes so high that speeds are influenced negatively, the number of accidents will decrease. The conclusion would be that in addition to the negative impact of congestion in terms of time losses, we also have a positive impact, since fatalities are reduced. Some supporting evidence is found for a number of countries where relatively low numbers of fatalities are observed during the morning peak.
Recent School Effectiveness Counter‐critiques: Problems and possibilities
Authors: Thrupp Martin
Published: July 2002

Abstract As part of continuing debate over school effectiveness research (SER), this article examines the recent responses of several school effectiveness researchers to the concerns of their critics. Issues examined include the complexity of the debate over SER, the value of pragmatism, the impact of social class, the significance of methodological problems, the value of current attempts to theorise SER, the relationship of SER to educational politics and policy‐making, the politics of doing SER, and the claimed achievements of SER. The discussion highlights a number of continuing tensions surrounding SER but also suggests that, if taken at face value, the counter‐critiques are often not taking the arguments of critics seriously. Nevertheless, with substantial concerns about SER now being aired from positions within as well as beyond SER, a shift in the nature of SER seems likely.
Adolescents’ ideas about individual and social responsibility in relation to children’s household work: Some international comparisons
Authors: Bowes Jennifer M. , Flanagan Constance , Taylor Alan J.
Published: July 2002

Cultural values of individual and social responsibility were investigated through adolescents’ ideas about children’s household work. A total of 4627 adolescents from six countries completed questionnaire items about the age children should begin chores, the value of children’s participation, and the appropriateness of payment. Differences were found between countries for all questions but notably for the question of payment. Adolescents in the USA, Australia, and Sweden were more likely to support a general payment associated with children’s household work, whereas adolescents in Bulgaria, the Czech Republic and Hungary were more likely to oppose payment. All differences interacted with type of task (self-care and family-care), corresponding to a relative emphasis on individual and social responsibility.
Developmental Expectations of Dutch, Turkish-Dutch, and Zambian Mothers: Towards an Explanation of Cross-cultural Differences
Authors: Willemsen Madde E. , van de Vijver Fons. J. R.
Published: July 2002

In this study, three models of cross-cultural differences of developmental expectations by parents were examined. The domain dependence model holds that differences vary across psychological domains such as perception and cognition. The cumulative differences model states that cross-cultural differences increase with age. The context variables model holds that cross-cultural differences in developmental expectations are mainly a function of background variables such as differences in parental education. The expectations of mothers in three different cultures were examined. Sixty eight Dutch, 50 Turkish-Dutch (Turkish women living in The Netherlands), and 69 Zambian mothers were asked to indicate the expected age of mastery for each of 77 skills, covering a broad range of behaviours. Zambian mothers expected most skills to develop at a later age than did Dutch and Turkish-Dutch mothers. Partial support was obtained for each model. Tentative evidence was obtained that the domain dependence model could explain most and the cumulative differences model the least cross-cultural differences.
Infant-Mother Attachment of Internationally Adopted Children in the Netherlands
Authors: Juffer Femmie , Rosenboom Lizette G.
Published: July 2002

In the Netherlands, 80 mothers and their infants, adopted from Sri Lanka, South Korea and Colombia, were observed at home at 6 and 12 months to rate the adoptive mother’ssensitivity, and in the Strange Situation at 12 and 18 months to assess the infant-mother attachment relationship. All inter-racially adopted infants were placed before the age of 6 months, with a mean age of 11 weeks, in adoptive families with or without biological children. Coded with Ainsworth’sclassification scheme the results reveal 74% secure attachment relationships, a percentage comparable to that of normative studies. The results indicate no differences regarding the child’scountry of origin, or the (non)presence of biological children. The results contradict findings from a study that revealed an over-representation of insecure infant-mother attachment relationships in a sample of American mothers with an interracially adopted infant. In the current study the adoptive mother’ssensitivity seems comparable to the sensitivity of nonadoptive mothers, a finding that concurs with the attachment results. It is suggested that the outcomes in this study may be partly explained by the fact that these infants were placed for adoption at a rather young age, with relatively favourable circumstances prior to the placement. This may well indicate that adoption placement per se, without the cumulative effects of understimulation and lack of personal affection that older placed children often experience in institutions, does not inevitably lead to a disturbed parent-infant relationship.
Determining the Meaning of Parameters in Multilevel Models for Longitudinal Data
Authors: Schuster Christof , von Eye Alexander
Published: July 2002

This paper is concerned with parameter interpretation in longitudinal, multilevel models. Models are described that consider repeated observations nested within individuals. These models typically first estimate subject-specific parameters for growth curves that describe the development of some observed variable overtime. Examples of such descriptors include polynomials. It is shown that interpretation of polynomial parameters can be facilitated by linear transformations. Examples of such transformations include centring (i.e. subtracting the mean from raw data). When parameters are specified such that they have no straightforward meaning at the first level of analysis, interpretation problems carry over to the second and higher levels. Therefore, proper specification of models at the first level is of utmost importance. Methods of transformation are introduced. Examples illustrate the method using data that describe children’s vocabulary development in the second year of life.
The Role of Dreams in Protecting Psychological Well-being in Traumatic Conditions
Authors: Punamäki Raija-Leena
Published: July 2002

The mental health function of dreaming was studied among Palestinian children and adolescents in a trauma group ( N= 268) and a comparison ( N= 144) group. The subjects were 6- to 15-year-old boys and girls, the mean age being 11.2± 2.64. They used a seven-day dream diary to record the dreams they could recall every morning. The results suggest that compensatory dreams could moderate between trauma and psychological symptoms. Traumatic events were not associated with psychological symptoms among children whose dreams were bizarre, vivid and active, and involved joyful feelings and happy endings. A mediating model suggested that exposure to traumatic events was associated with mundane persecution and unpleasant repetitious dreams. These dysfunctional dreams were, in turn, associated with poor psychological adjustment. The dynamics of mastery and compensation dreams in traumatic conditions are discussed.
The Transaction between Parents’ Perceptions of their Children’s Shyness and their Parenting Styles
Authors: Rubin Kenneth H. , Nelson Larry J. , Hastings Paul , Asendorpf Jens
Published: July 2002

In recent years, researchers have examined factors that “determine” parenting beliefs, styles, and behaviours. One potential determinant of parenting is the child him/herself. Child characteristics, such as temperament, have been cited as evocative influences on parenting beliefs and behaviours. The primary purpose of this study was to investigate the longitudinal relations between children’s social wariness/inhibition and parents’ beliefs about how to best socialise their children. Questionnaire data on child temperament and parenting practices were collected from the parents (mothers and fathers) of sixty 2-year-olds; identical data were collected 2 years later. Observations of inhibited behaviour were taken at two years. Results indicated that few differences existed between mothers’ and fathers’ expressed parenting styles at ages 2 and 4 years. Second, parental perceptions of child shyness at age 2 were: (a) stable to age 4; and (b) predicted a lack of encouragement of independence at age 4. Third, parents’ expressed lack of encouragement of independence, although stable from 2 to 4 years, failed to predict child shyness at age 4. The findings support the conjecture that young children’s dispositional characteristics predict subsequent maternal and paternal behaviour.
International Multifoods – Case B
Authors: Desanctis Gerardine , Price Robert M.
Published: July 2002

The International Multifoods company (IMC) confronted a series of crises in the 1990s that are described in this case. The Renaissance information system, which was intended to bring enormous cash flow to IMC's subsidiary, Vending Services of America (VSA), was behind schedule and over budget. Furthermore, VSA was experiencing serious managerial problems. IMC had cash flow problems that were reaching crisis levels due to, among other things, economic volatility in Venezuela where IMC had a major food business. IMC struggled with organization structure issues and was pressed by its investors to reorganize. As Renaissance-related and other problems brewed within VSA, the CEO of IMC resigned, and an interim CEO had to step in and somehow move the company out of the crises at hand.
Cytokine changes in patients with heatstroke during pilgrimage to Makkah
Authors: Hashim I. A. , Al-Zeer A. , Al-Shohaib S. , Al-Ahwal M. , Shenkin A.
Published: July 2002

Circulating levels and role of IL‐6, IL‐1ra, TNFsr‐II and CRP in patients with heatstroke is not fully known. This study correlated levels of these mediators with outcome in 26 patients. In survivors (n=20), IL‐6 concentration declined on cooling, whereas in non‐survivors levels continued to increase at 6 h following admission before declining. Admission TNFsr‐II concentrations in survivors were significantly lower than non‐survivors and levels continued to rise in both groups. IL‐1ra levels were markedly elevated in both groups. Changes in cytokine levels were not influenced by renal function. Elevated C‐reactive protein levels were observed for both groups and remained so despite cooling, furthermore, there was no correlation with alanine aminotransferase levels. The study demonstrated the elevation of the above mediators and suggested a role in the pathogenesis of heatstroke. Markedly elevated levels or those that remained elevated despite cooling were associated with mortality.
Ozone oxidative preconditioning: a protection against cellular damage by free radicals
Authors: León O. S. , Menéndez S. , Merino N. , Castillo R. , Sam S. , Pérez L. , Cruz E. , Bocci V.
Published: July 2002

There is some anecdotal evidence that oxygen‐ozone therapy may be beneficial in some human diseases . However so far only a few biochemical and pharmacodynamic mechanisms have been elucidated. On the basis of preliminary data we postulated that controlled ozone administration would promote an oxidative
preconditioning preventing the hepatocellular damage mediated by free radicals. Six groups of rats were classified as follows: (1) negative control, using intraperitoneal sunflower oil; (2) positive control using carbon tetrachloride (CCl4) as an oxidative challenge; (3) oxygen‐ozone, pretreatment via rectal insufflation (15 sessions ) and after it, CCl4; (4) oxygen, as group 3 but using oxygen only; (5) control oxygen‐ozone, as group 3, but without CCl4 ; group (6) control oxygen, as group 5, but using oxygen only. We have evaluated critical biochemical parameters such as levels of transaminase , cholinesterase , super ‐ oxide dismutase, catalase , phospholipase A, calcium dependent ATPase, reduced glutathione , glucose 6 phosphate dehydrogenase and lipid peroxidation. Interestingly, in spite of CCl4 adminis tration, group 3 did not differ from group 1, while groups 2 and 4 showed significant differences from groups 1 and 3 and displayed hepatic damage . To our knowledge these are the first experimental results showing that repeated administration of ozone in atoxic doses is able to induce an adaptation to oxidative stress thus enabling the animals to maintain hepatocellular integrity after CCl4 poisoning.
Prevention of Renal Injury After Induction of Ozone Tolerance in Rats Submitted to Warm Ischaemia
Authors: Barber E. , Menéndez S. , León O. S. , Barber M. O. , Merino N. , Calunga J. L. , Cruz E. , Bocci V.
Published: July 2002

On the basis that ozone (O3) can upregulate cellular antioxidant enzymes, a morphological, biochemical and functional renal study was performed in rats undergoing a prolonged treatment with O3 before renal ischaemia. Rats were divided into four groups: (1) control, a medial abdominal incision was performed to expose the kidneys; (2) ischaemia, in animals undergoing a bilateral renal ischaemia (30 min), with subsequent reperfusion (3 h); (3) O3 + ischaemia, as group 2, but with previous treatment with O3 (0.5 mg/kg per day given in 2.5 ml O2) via rectal administration for 15 treatments; (4) O2 + ischaemia, as group 3, but using oxygen (O2) alone. Biochemical parameters as fructosamine level, phospholipase A, and superoxide dismutases (SOD) activities, as well as renal plasma flow (RPF) and glomerular filtration rate (GFR), were measured by means of plasma clearance of p‐amino‐hippurate and inulin, respectively. In comparison with groups 1 and 3, the RPF and GFR were significantly decreased in groups 2 and 4. Interestingly, renal homogenates of the latter groups yielded significantly higher values of phospholipase A activity and fructosamine level in comparison with either the control (1) and the O3 (3) treated groups. Moreover renal SOD activity showed a significant increase in group 3 without significant differences among groups 1, 2 and 4. Morphological alterations of the kidney were present in 100%, 88% and 30% of the animals in groups 2, 4 and 3, respectively. It is proposed that the O3 protective effect can be ascribed to the substantial possibility of upregulating the antioxidant defence system capable of counteracting the damaging effect of ischaemia. These findings suggest that, whenever possible, ozone preconditioning may represent a prophylactic approach for minimizing renal damage before transplantation.
Subchronic Toxicity of Thalidomide in Rodents After 13 Weeks of Oral Administration
Authors: Teo Steve K. , Trigg Nancy J. , Shaw Mary E. , Morgan J. Michael , Thomas Steve D.
Published: July 2002

The subchronic toxicology of thalidomide was determined in CD-1 mice and F344 rats. Animals (10/sex/dose) were orally dosed at 30,300, and 3000 mg/kg/day over 13 weeks. Control animals were given 1% carboxymethylcellulose. No thalidomide-related mortality occurred throughout the study. Some species and sex differences were seen. In mice, thalidomide had no effect on body weight, food consumption, ophthalmic function or clinical chemistry/hematology, but a dose-dependent orange-pink urine was observed in both sexes. The discoloration was probably due to chromogenic breakdown products of thalidomide. The only significant finding in the mouse study was dose-related hepatic centrilobular hypertrophy in the males. This appeared only at the highest dose in the females. The hypertrophy was correlated with increased liver weight for the high dose of both sexes suggesting enzyme induction. In rats, thalidomide produced lower body weights in both sexes compared to control with a dose-response more evident in males. Male rats dosed at 30, 300, and 3000 mg/kg had body weights that were 8, 11, and 19% below control weight just before necropsy. Corresponding female rats were only 6–7% below control weights at all dose levels. Lower food consumption was observed in male rats and varied between 6–13% below control with no dose-response. Decreased forelimb strength was noted in males and could be due to the lower body weights. Functional observational battery tests and histopathology of the sural nerve and lumbar spinal cord sections suggested that the rat did not develop thalidomide-induced peripheral neuropathy. Mild anemia and leukopenia were seen only in some treated males. A decrease in total and free T4 was more consistent in females. Both sexes had lower thymus weights with no histological correlate compared to control. The no-observed-adverse-effect level for mice and female rats were 3000 mg/kg and 30 mg/kg for male rats.
Developmental Toxicity Studies in Rats and Rabbits and Two-Generation Reproduction Study in Rats on 4-(2,4-DichIorophenoxy)butyric Acid
Authors: Charles Jeffrey M. , Henwood Susan M. , Leeming Nicholas M.
Published: July 2002

4-(2,4-Dichlorophenoxy)butyric acid (2,4-DB) is principally used in the USA on peanuts, soybeans, and alfalfa. The developmental toxicity in rats (25/group) was studied at dose levels of 0, 31.25, 62.5, and 125 mg/kg/day. The maximum tolerated dose (MTD) was clearly exceeded at 125 mg/kg/day, as evidenced by a net weight loss in the dams during the dosing period, resulting in four dams totally resorbing their fetuses. The no-observed-effect level (NOEL) for maternal toxicity was 31.25 mg/kg/day based on a slight, but consistent, body weight effect at 62.5 mg/kg/day. The NOEL for embryo/fetal toxicity was 31.25 mg/kg/day based on increases in fetal skeletal variations at 62.5 mg/kg/day. The developmental toxicity in rabbits (16/group) was evaluated at dose levels of 0, 15, 30, or 60 mg/kg/day. The MTD was clearly exceeded at 60 mg/kg/day as evidenced by severe weight-gain depression during the dosing period in the does, resulting in two does aborting and two does being sacrificed in a moribund condition. The NOEL for maternal toxicity was 30 mg/kg/day based on the above noted effects at 60 mg/kg/day. The NOEL for embryo/fetal toxicity was 60 mg/kg/day due to the lack of any effects noted. The reproductive toxicity of 2,4-DB was investigated in a two-generation study with two litters bred per generation. Rats (28/sex/group for the F0 generation and 24/sex/group for the F1 generation) were fed dietary concentrations of 0, 60,300, and 1500 ppm during the first generation, but because it proved impossible to rear a filial generation at 1500 ppm, the second generation was continued at doses of 0, 60, and 300 ppm. Mating performance, fertility, and pregnancy rate were unimpaired. A NOEL for reproductive toxicity was 1500 ppm (111.8 mg/kg/day for males and 110.6 mg/kg/day for females); however, severe postnatal toxicity was produced at this level. The NOEL for postnatal toxicity was 300 ppm (22.5-32.6 mg/kg/day for males and 26.4-36.7 mg/kg/day for females depending on the generation). The NOELs for developmental and reproductive toxicity are in general agreement with those reported for 2,4-dichlorophenoxyacetic acid (2,4-D), the major metabolite of 2,4-DB.
Endocrine-Modulating Chemicals as a Case Study of Science in the Courtroom
Authors: Chrostowski Paul C. , Durda Judi L. , Estreicher Herbert
Published: July 2002

In contemporary society, many scientific issues involving toxi-cologists, epidemiologists, and other risk assessment professionals have become the topics of hotly contested legal actions. Nowhere is this more apparent than in toxic tort cases where judgements involving millions of dollars can hinge on the expert testimony provided by a toxicologist. Many of these cases involve scientific opinions that have not yet entered the mainstream of scientific thought, thus it becomes necessary for courts to place a great deal of emphasis on the reliability of science. In this paper, we analyze the scientific and legal bases of disease causation in the toxic tort context and illustrate how the emerging issue of endocrine modulation may evolve in this context. We conclude that there is an inadequate scientific basis to show personal injuries associated with exposure to a class of putative endocrine modulators. This paper is not intended to illustrate a specific case; it is intended to be a heuristic for toxicologists, risk assessors, and attorneys, who may be forced to confront these and similar issues in the course of their professional careers.
Assessment of Proposed Cyclohexanone Limits for Containers
Authors: Cammack Jon N. , White Randy D. , Wilson Daniel M. , Gonder Janet C. , Hoet Patrick
Published: July 2002

A limit of 5 mg was recently proposed as the European Pharmacopoeia limit for residual cyclohexanone in plastic containers for parenteral medical use with a nominal volume greater than 250 ml. Comparison of the proposed limit to published toxicological end-points and current governmental regulations was used to assess the potential risk following exposure. Based on the comparison, no adverse effects would be predicted from clinical exposures to 5 mg residual cyclohexanone in containers with a nominal volume of greater than 250 ml.
MDMA toxicity: no evidence for a major influence of metabolic genotype at CYP2D6
Authors: O'DONOHOE ALAN , O'FLYNN KAREN , SHIELDS KEVIN , HAWI ZIARIH , GILL MICHAEL
Published: July 2002

3,4 Methylenedioxymethamphetamine (MDMA or ecstasy) has become a major drug of abuse over the last decade. It produces a mixture of systemic and neuropsychological effects. Animal studies show a range of short‐ and long‐term toxic effects, both systemically and neurochemically. In humans, toxicity and death due to the drug have been attributed to a variety of causes, with ‘idiosyncratic’, or non‐dose‐related, reactions often cited. It has recently been established that MDMA is metabolized via the cytochrome P450 enzyme, debrisoquine hydroxylase. This enzyme is coded by the gene CYP2D6. This gene contains mutations which effect the function of the enzyme, and individuals homozygous for these mutations are known as poor metabolizers. Between 3 and 10% of the Caucasian population are thus affected, and therefore may be less able to metabolize MDMA. In this paper we examine the hypothesis that individuals selected on the basis of having had an adverse reaction to MDMA will be more likely than the general population to have homozygous mutations at CYP2D6. We obtained retrospectively seven cases of toxicity or death thought to be due to MDMA. DNA was extracted from these patients, and their genotype ascertained. None of this small sample was shown to be homozygous for the mutation at CYP2D6. Three possible explanations are offered for these results. (1) The non‐dose‐related nature of MDMA toxicity may be due, either alone or in combination, to contaminants in the drug, or ambient environmental/physiological factors. (2) Our genotyping methods may have missed one of the rare additional mutations which effect gene function at CYP2D6. (3) Our study sample may be too small to demonstrate a statistically significant result. A larger study is currently under way.
Isolated P-selectin Glycoprotein Ligand-1 Dynamic Adhesion to P- and E-selectin
Authors: Goetz Douglas J. , Greif Daniel M. , Ding Han , Camphausen Raymond T. , Howes Steven , Comess Kenneth M. , Snapp Karen R. , Kansas Geoffrey S. , Luscinskas Francis W.
Published: July 2002

Leukocyte adhesion to vascular endothelium under flow involves an adhesion cascade consisting of multiple receptor pairs that may function in an overlapping fashion. P-selectin glycoprotein ligand-1 (PSGL-1) and L-selectin have been implicated in neutrophil adhesion to P- and E-selectin under flow conditions. To study, in isolation, the interaction of PSGL-1 with P-and E-selectin under flow, we developed an in vitro model in which various recombinant regions of extracellular PSGL-1 were coupled to 10-μm-diameter microspheres. In a parallel plate chamber with well defined flow conditions, live time video microscopy analyses revealed that microspheres coated with PSGL-1 attached and rolled on 4-h tumor necrosis factor-α–activated endothelial cell monolayers, which express high levels of E-selectin, and CHO monolayers stably expressing E-or P-selectin. Further studies using CHO-E and -P monolayers demonstrate that the first 19 amino acids of PSGL-1 are sufficient for attachment and rolling on both E- and P-selectin and suggest that a sialyl Lewis x–containing glycan at Threonine-16 is critical for this sequence of amino acids to mediate attachment to E- and P-selectin. The data also demonstrate that a sulfated, anionic polypeptide segment within the amino terminus of PSGL-1 is necessary for PSGL-1–mediated attachment to P- but not to E-selectin. In addition, the results suggest that PSGL-1 has more than one binding site for E-selectin: one site located within the first 19 amino acids of PSGL-1 and one or more sites located between amino acids 19 through 148.
An Intralysosomal hsp70 Is Required for a Selective Pathway of Lysosomal Protein Degradation
Authors: Agarraberes Fernando A. , Terlecky Stanley R. , Dice J. Fred
Published: July 2002

Previous studies have implicated the heat shock cognate (hsc) protein of 73 kD (hsc73) in stimulating a lysosomal pathway of proteolysis that is selective for particular cytosolic proteins. This pathway is activated by serum deprivation in confluent cultured human fibroblasts. We now show, using indirect immunofluorescence and laser scanning confocal microscopy, that a heat shock protein (hsp) of the 70-kD family (hsp70) is associated with lysosomes (ly-hsc73). An mAb designated 13D3 specifically recognizes hsc73, and this antibody colocalizes with an antibody to lgp120, a lysosomal marker protein. Most, but not all, lysosomes contain ly-hsc73, and the morphological appearance of these organelles dramatically changes in response to serum withdrawal; the punctate lysosomes fuse to form tubules.

Based on susceptibility to digestion by trypsin and by immunoblot analysis after two-dimensional electrophoresis of isolated lysosomes and isolated lysosomal membranes, most ly-hsc73 is within the lysosomal lumen. We determined the functional importance of the ly-hsc73 by radiolabeling cellular proteins with [3H]leucine and then allowing cells to endocytose excess mAb 13D3 before measuring protein degradation in the presence and absence of serum. The increased protein degradation in response to serum deprivation was completely inhibited by endocytosed mAb 13D3, while protein degradation in cells maintained in the presence of serum was unaffected. The intralysosomal digestion of endocytosed [3H]RNase A was not affected by the endocytosed mAb 13D3. These results suggest that ly-hsc73 is required for a step in the degradative pathway before protein digestion within lysosomes, most likely for the import of substrate proteins.
Dynamics of Myoblast Transplantation Reveal a Discrete Minority of Precursors with Stem Cell–like Properties as the Myogenic Source
Authors: Beauchamp Jonathan R. , Morgan Jennifer E. , Pagel Charles N. , Partridge Terence A.
Published: July 2002

Myoblasts, the precursors of skeletal muscle fibers, can be induced to withdraw from the cell cycle and differentiate in vitro. Recent studies have also identified undifferentiated subpopulations that can self-renew and generate myogenic cells (Baroffio, A., M. Hamann, L. Bernheim, M.-L. Bochaton-Pillat, G. Gabbiani, and C.R. Bader. 1996. Differentiation. 60:47–57; Yoshida, N., S. Yoshida, K. Koishi, K. Masuda, and Y. Nabeshima. 1998. J. Cell Sci. 111:769–779). Cultured myoblasts can also differentiate and contribute to repair and new muscle formation in vivo, a capacity exploited in attempts to develop myoblast transplantation (MT) for genetic modification of adult muscle. Our studies of the dynamics of MT demonstrate that cultures of myoblasts contain distinct subpopulations defined by their behavior in vitro and divergent responses to grafting. By comparing a genomic and a semiconserved marker, we have followed the fate of myoblasts transplanted into muscles of dystrophic mice, finding that the majority of the grafted cells quickly die and only a minority are responsible for new muscle formation. This minority is behaviorally distinct, slowly dividing in tissue culture, but rapidly proliferative after grafting, suggesting a subpopulation with stem cell–like characteristics.
Quantitative Changes in Integrin and Focal Adhesion Signaling Regulate Myoblast Cell Cycle Withdrawal
Authors: Sastry Sarita K. , Lakonishok Margot , Wu Stanley , Truong Tho Q. , Huttenlocher Anna , Turner Christopher E. , Horwitz Alan F.
Published: July 2002

We previously demonstrated contrasting roles for integrin α subunits and their cytoplasmic domains in controlling cell cycle withdrawal and the onset of terminal differentiation (Sastry, S., M. Lakonishok, D. Thomas, J. Muschler, and A.F. Horwitz. 1996. J. Cell Biol. 133:169–184). Ectopic expression of the integrin α5 or α6A subunit in primary quail myoblasts either decreases or enhances the probability of cell cycle withdrawal, respectively. In this study, we addressed the mechanisms by which changes in integrin α subunit ratios regulate this decision. Ectopic expression of truncated α5 or α6A indicate that the α5 cytoplasmic domain is permissive for the proliferative pathway whereas the COOH-terminal 11 amino acids of α6A cytoplasmic domain inhibit proliferation and promote differentiation. The α5 and α6A cytoplasmic domains do not appear to initiate these signals directly, but instead regulate β1 signaling. Ectopically expressed IL2R-α5 or IL2R-α6A have no detectable effect on the myoblast phenotype. However, ectopic expression of the β1A integrin subunit or IL2R-β1A, autonomously inhibits differentiation and maintains a proliferative state. Perturbing α5 or α6A ratios also significantly affects activation of β1 integrin signaling pathways. Ectopic α5 expression enhances expression and activation of paxillin as well as mitogen-activated protein (MAP) kinase with little effect on focal adhesion kinase (FAK). In contrast, ectopic α6A expression suppresses FAK and MAP kinase activation with a lesser effect on paxillin. Ectopic expression of wild-type and mutant forms of FAK, paxillin, and MAP/erk kinase (MEK) confirm these correlations. These data demonstrate that (a) proliferative signaling (i.e., inhibition of cell cycle withdrawal and the onset of terminal differentiation) occurs through the β1A subunit and is modulated by the α subunit cytoplasmic domains; (b) perturbing α subunit ratios alters paxillin expression and phosphorylation and FAK and MAP kinase activation; (c) quantitative changes in the level of adhesive signaling through integrins and focal adhesion components regulate the decision of myoblasts to withdraw from the cell cycle, in part via MAP kinase.
Relationship between Arp2/3 Complex and the Barbed Ends of Actin Filaments at the Leading Edge of Carcinoma Cells after Epidermal Growth Factor Stimulation
Authors: Bailly Maryse , Macaluso Frank , Cammer Michael , Chan Amanda , Segall Jeffrey E. , Condeelis John S.
Published: July 2002

Using both light and high resolution electron microscopy, we analyzed the spatial and temporal relationships between the Arp2/3 complex and the nucleation activity that is required for lamellipod extension in mammary carcinoma cells after epidermal growth factor stimulation. A rapid two- to fourfold increase in filament barbed end number occurs transiently after stimulation and remains confined almost exclusively to the extreme outer edge of the extending lamellipod (within 100–200 nm of the plasma membrane). This is accompanied by an increase in filament density at the leading edge and a general decrease in filament length, with a specific loss of long filaments. Concomitantly, the Arp2/3 complex is recruited with a 1.5-fold increase throughout the entire cortical filament network extending 1–1.5 μm in depth from the membrane at the leading edge. The recruitment of the Arp2/3 complex at the membrane of the extending lamellipod indicates that Arp2/3 may be involved in initial generation of growing filaments. However, only a small subset of the complex present in the cortical network colocalizes near free barbed ends. This suggests that the 100–200-nm submembraneous compartment at the leading edge of the extending lamellipod constitutes a special biochemical microenvironment that favors the generation and maintenance of free barbed ends, possibly through the locally active Arp2/3 complex, severing or decreasing the on-rate of capping protein. Our results are inconsistent with the hypothesis suggesting uncapping is the dominant mechanism responsible for the generation of nucleation activity. However, they support the hypothesis of an Arp2/3-mediated capture of actin oligomers that formed close to the membrane by other mechanisms such as severing. They also support pointed-end capping by the Arp2/3 complex, accounting for its wide distribution at the leading edge.
Localization of Putative Stem Cells in Dental Epithelium and Their Association with Notch and Fgf Signaling
Authors: Harada Hidemitsu , Kettunen Päivi , Jung Han-Sung , Mustonen Tuija , Wang Y. Alan , Thesleff Irma
Published: July 2002

The continuously growing mouse incisor is an excellent model to analyze the mechanisms for stem cell lineage. We designed an organ culture method for the apical end of the incisor and analyzed the epithelial cell lineage by 5-bromo-2′-deoxyuridine and DiI labeling. Our results indicate that stem cells reside in the cervical loop epithelium consisting of a central core of stellate reticulum cells surrounded by a layer of basal epithelial cells, and that they give rise to transit-amplifying progeny differentiating into enamel forming ameloblasts. We identified slowly dividing cells among the Notch1-expressing stellate reticulum cells in specific locations near the basal epithelial cells expressing lunatic fringe, a secretory molecule modulating Notch signaling. It is known from tissue recombination studies that in the mouse incisor the mesenchyme regulates the continuous growth of epithelium. Expression of Fgf-3 and Fgf-10 were restricted to the mesenchyme underlying the basal epithelial cells and the transit-amplifying cells expressing their receptors Fgfr1b and Fgfr2b. When FGF-10 protein was applied with beads on the cultured cervical loop epithelium it stimulated cell proliferation as well as expression of lunatic fringe. We present a model in which FGF signaling from the mesenchyme regulates the Notch pathway in dental epithelial stem cells via stimulation of lunatic fringe expression and, thereby, has a central role in coupling the mitogenesis and fate decision of stem cells.
Drosophila β Spectrin Functions Independently of α Spectrin to Polarize the Na,k Atpase in Epithelial Cells
Authors: Dubreuil Ronald R. , Wang Ping , Dahl Steve , Lee John , Goldstein Lawrence S.B.
Published: July 2002

Spectrin has been proposed to function as a sorting machine that concentrates interacting proteins such as the Na,K ATPase within specialized plasma membrane domains of polarized cells. However, little direct evidence to support this model has been obtained. Here we used a genetic approach to directly test the requirement for the β subunit of the αβ spectrin molecule in morphogenesis and function of epithelial cells in Drosophila. β Spectrin mutations were lethal during late embryonic/early larval development and they produced subtle defects in midgut morphology and stomach acid secretion. The polarized distributions of αβH spectrin and ankyrin were not significantly altered in β spectrin mutants, indicating that the two isoforms of Drosophila spectrin assemble independently of one another, and that ankyrin is upstream of αβ spectrin in the spectrin assembly pathway. In contrast, β spectrin mutations had a striking effect on the basolateral accumulation of the Na,K ATPase. The results establish a role for β spectrin in determining the subcellular distribution of the Na,K ATPase and, unexpectedly, this role is independent of α spectrin.
Death Commitment Point Is Advanced by Axotomy in Sympathetic Neurons
Authors: Fletcher Graham C. , Xue Luzheng , Passingham Shareta K. , Tolkovsky Aviva M.
Published: July 2002

Axotomized neurons have several characteristics that are different from intact neurons. Here we show that, unlike established cultures, the axotomized sympathetic neurons deprived of NGF become committed to die before caspase activation, since the same proportion of NGF-deprived neurons are rescued by NGF regardless of whether caspases are inhibited by the pan-caspase inhibitor Boc-Asp(O-methyl)-CH2F (BAF). Despite prolonged Akt and ERK signaling induced by NGF after BAF treatment has prevented death, the neurons fail to increase protein synthesis, recover ATP levels, or grow. Within 3 d, all the mitochondria disappear without apparent removal of any other organelles or loss of membrane integrity. Although NGF does rescue intact BAF-treated 6-d cultures after NGF deprivation, rescue by NGF fails when these neurons are axotomized before NGF deprivation and BAF treatment. Moreover, cytosolic cytochrome c rapidly kills axotomized neurons. We propose that axotomy induces signals that make sympathetic neurons competent to die prematurely. NGF cannot repair these NGF-deprived, BAF-treated neurons because receptor signaling (which is normal) is uncoupled from protein renewal, and the mitochondria (which are damaged) go on to be eliminated. Hence, the order of steps underlying neuronal death commitment is mutable and open to regulation.
Identification and Characterization of an Escorter for Two Secretory Adhesins in Toxoplasma gondii
Authors: Reiss Matthias , Viebig Nicola , Brecht Susan , Fourmaux Marie-Noelle , Soete Martine , Di Cristina Manlio , Dubremetz Jean François , Soldati Dominique
Published: July 2002

The intracellular protozoan parasite Toxoplasma gondii shares with other members of the Apicomplexa a common set of apical structures involved in host cell invasion. Micronemes are apical secretory organelles releasing their contents upon contact with host cells. We have identified a transmembrane micronemal protein MIC6, which functions as an escorter for the accurate targeting of two soluble proteins MIC1 and MIC4 to the micronemes. Disruption of MIC1, MIC4, and MIC6 genes allowed us to precisely dissect their contribution in sorting processes. We have mapped domains on these proteins that determine complex formation and targeting to the organelle. MIC6 carries a sorting signal(s) in its cytoplasmic tail whereas its association with MIC1 involves a lumenal EGF-like domain. MIC4 binds directly to MIC1 and behaves as a passive cargo molecule. In contrast, MIC1 is linked to a quality control system and is absolutely required for the complex to leave the early compartments of the secretory pathway. MIC1 and MIC4 bind to host cells, and the existence of such a complex provides a plausible mechanism explaining how soluble adhesins act. We hypothesize that during invasion, MIC6 along with adhesins establishes a bridge between the host cell and the parasite.