bluuebunny/crossref_metadata_2025 huggingface dataset:
“What are food and air like inside our bodies?”: Children’s thinking about digestion and respiration
Authors: Toyama Noriko
Published: July 2002

A series of five experiments evaluated whether young children are aware of biological transformations associated with eating and breathing. In Experiment 1, children aged 4, 5, 7, and 8 years predicted that biological damage results from lack of eating and breathing. Children also recognised that food changes inside the body, but seldom referred spontaneously to biological transformation. In Experiments 2 and 2A, children were presented with several alternative explanations of what food and air would be like inside the human body. Both preschoolers and elementary schoolchildren assumed that air would acquire warmth and colour inside the body. The older children consistently understood biological transformation of food. Preschoolers accepted the idea that food undergoes a transformation necessary for health and growth, but did not think so when the transformation was expressed in a material sense. In Experiments 3 and 3A, some preschoolers predicted the transformation of resources inside familiar and unfamiliar living things, but not inside nonliving things. In addition, some 4- and 5-year-olds recognised the sun’s contribution to “digestive” processes for plants, but not for mammals. Finally, the question of whether early understanding of digestion can be termed “theory-like” was discussed.
Pathology of Ocular Irritation with Bleaching Agents in the Rabbit Low-Volume Eye Test
Authors: Maurer James K. , Molai AL , Parker Ron D. , Li Li  , Carr Gregory J. , Petroll W. Matthew , Cavanagh H. Dwight , Jester James V.
Published: July 2002

Despite differences in the processes leading to tissue damage, the ocular irritation response to various surfactants, two concentrations of an acid and an alkali, and an acetone, alcohol, aromatic amine, and aldehyde has been shown to depend on the extent of initial injury. The purpose of this study was to assess the extent to which this fundamental relationship exists for bleaching agents in the rabbit low-volume eye test. Ten μl of sodium perborate monohydrate (NaBO 3), sodium hypochlorite (NaOCl), 10% hydrogen peroxide (H2O 2), and 15% H2O2 was applied directly to the cornea of the right eye of each rabbit. Macroscopic assessments for irritation were made 3 hours after dosing and periodically until 35 days. Light microscopic examinations were conducted on tissues obtained at 3 hr and on 1, 3, and 35 days. In vivo confocal microscopy (CM) and measurements of dead corneal epithelial cells and keratocytes at 3 hours and 1 day were used to characterize quantitatively initial corneal injury, while in vivo CM performed at 3 hours and 1, 3, 7, 14, and 35 days was used to characterize quantitatively the corneal changes over time. The changes with NaBO3 and NaOCl were consistent with mild irritancy. For both, corneal injury was limited to the epithelium and superficial stroma. The changes with 10% H2O2 and 15% H2O2 were consistent with severe irritation. Both concentrations affected the epithelium and deep stroma, with 15% H2O 2 also at times affecting the endothelium. However, unlike other irritants previously studied, with 10% H2O2 and 15% H2O 2 there was an incongruity between the extent of epithelial and stromal injury, with stromal injury being more extensive than epithelial injury. A similar, although less dramatic, effect was observed with NaBO3. Additionally, there was still significant keratocyte loss at 35 days with 10% H2O2 and 15% H2O2 even though the eyes at times were considered to be macroscopically normal. These observations highlight the need to include both epithelial and stromal components in an ex vivo or in vitro alternative assay. In conclusion, these results continue to support and extend our hypothesis that ocular irritation is principally defined by the extent of initial injury despite clear differences in the means by which irritants cause tissue damage. Importantly, we have identified unique differences in the ocular injury and responses occurring with bleaching agents that are important to consider in the development and validation of alternative ocular irritation tests to characterize a broad range of materials differing in type and irritancy.
Toxicologic Lesions Associated with Two Related Inhibitors of Oxidosqualene Cyclase in the Dog and Mouse
Authors: Pyrah Ian T. , Kalinowski Adam , Jackson David , Davies Wendy , Davis Stewart , Aldridge Andrew , Greaves Peter
Published: July 2002

Two novel hypolipidaemic agents, both members of the aminopyrimidine series, with a mode of action of inhibition of oxidosqualene cyclase (OSC), were administered orally to dogs and mice for 14 and 28 days. Both compounds produced a similar spectrum of pathologic changes. In dogs, the agents produced equatorial single cell necrosis and cataract in the lens (also observed clinically); atrophy, ulceration, and infl ammation of the cornea; hyperkeratosis, acanthosis, hair papillary atrophy, and infl ammation of the skin; and epithelial degeneration and sperm granuloma in the epididymides. One female dog showed signs of liver toxicity. In mice, severe cataract formation was seen with both compounds, and liver toxicity was produced by one of the compounds. The severity and speed of onset of the cataract formation were very marked. The changes seen were dissimilar to those reported with the most commonly used class of hypolipidaemic agents in the clinic, the hydroxymethy l glutaryl coenzyme A (HMGCoA) reductase inhibitors but were reminiscent of those reported for the hypolipidaemic agent Triparanol, which was predictive of toxicity seen in man.
Pathology of Ocular Irritation with Acetone, Cyclohexanol, Parafl uoroaniline, and Formaldehyde in the Rabbit Low-Volume Eye Test
Authors: Maurer James K. , Molai AL , Parker Ron D. , Li Li , Carr Gregory J. , Petroll W. Matthew , Cavanagh H. Dwight , Jester James V.
Published: July 2002

The ocular irritation responses to 11 different surfactants and two concentrations of acetic acid and sodium hydroxide have been shown to depend on the extent of initial injury, despite marked differences in the processes leading to tissue damage. The purpose of these studies was to determine the extent to which this fundamental relationship applies to other nonsurfactants. Ten μl of acetone (ACT), cyclohexanol (CY), parafluoroaniline (PF), or 37% formaldehyde (FA) was directly applied to the cornea of the right eye of each rabbit. Eyes and eyelids were macroscopically scored for signs of irritation beginning 3 hours after dosing and periodically until recovery or 35 days. Tissues were obtained for light microscopic examination after 3 hours and on days 1, 3, and 35. Initial corneal injury was characterized quantitatively at 3 hours and 1 day using in vivo confocal microscopy (CM) and by postmortem quantitation of dead corneal epithelial cells and keratocytes using a Live—Dead Assay (L/D, Molecular Probes) and scanning laser CM. Corneal changes over time were characterized quantitatively using in vivo CM performed at 3 hours and 1, 3, 7, 14, and 35 days. The changes with ACT were consistent with mild irritation. Corneal injury was limited to the epithelium and superfi cial stroma, with the mean normalized depth of injury (NDI) being less than 10% with the majority of regions showing no stromal injury. Changes with CY and PF were consistent with moderate to severe irritation, and FA caused severe irritation. Specifi cally, corneal injury by CY and PF tended to involve the epithelium and anterior stroma, with the mean NDI being 10.4% to 23.8%, while injury with FA involved the epithelium, deep stroma, and at times the endothelium. Interestingly, with FA significantly less injury was observed at 3 hours with a dramatic increase in injury observed at 1 day and thereafter. In conclusion, these results continue to support and extend our hypothesis that ocular irritation is principally defined by the extent of initial injury despite clear differences in the means by which irritants cause tissue damage. We believe this approach can be applied to developing alternative assays based on injury to ex vivo eyes or injury to an in vitro corneal equivalent system.
Helping Those With Mental Health Problems Access Open Employment - A Glasgow Case Study
Authors: Hayton Keith
Published: July 2002

Mental health problems are relatively common within the population. For some, the problems may be so severe that they have to give up paid work. However, there is evidence that, not only do the mentally ill have a strong desire to return to paid work but, such work can improve mental health. This paper looks at the varied ways that have evolved to help the mentally ill return to work. Drawing on research undertaken in Glasgow, it finds that mental health projects have developed in isolation from one an other and from the wider local economic development network. It argues that a more effective service can be provided if there is improved networking between projects and if they concentrate upon interventions which make optimum use of their particular knowledge and skills. This would involve mental health projects providing the social and therapeutic aspects of support, whilst the economic development projects dealt with vocation al training and labour market links.
Non‐prescription drug use by gay men in Sydney, Melbourne and Brisbane
Authors: KNOX STEPHANIE , KIPPAX SUSAN , CRAWFORD JUNE , PRESTAGE GARRETT , VAN DE VEN PAUL
Published: July 2002

This study investigated the prevalence of non‐prescription and injecting drug use among a nonclinical sample of gay and homosexually active men and examined what distinguished men who had recently injected drugs from other gay men. Gay and homosexually active men from three cohort studies in Brisbane, Melbourne and Sydney, Australia (N = 1438) were interviewed between June 1995 and July 1996. Three‐quarters of the men reported using drugs in the 6 months prior to interview and 10% had injected drugs in this period. It was concluded that drug use is normative among gay men in Australian cities. Recent drug use without injecting was associated with high socio‐economic status and gay community attachment. Men who had recently injected drugs were as attached to gay community as men who had used without injecting. Multivariate analysis found that injecting drugs was more common among younger men and men who were HIV positive. Socio‐economic differences between the men who had injected recently and other men were possibly related to the higher rates of HIV infection among the men who injected and injecting drugs may be part of an HIV positive subculture. Men who injected drugs were more likely to have received money for sex and there was a significant minority of injectors who did not identify as gay, suggesting that there is a small subgroup of men who are likely to be injecting in contexts outside gay community. [Knox S, Kippax S, Crawford J, Prestage G, Van de Ven P. Non‐prescription drug use by gay men in Sydney, Melbourne and Brisbane. Drug Alcohol Rev 1999:18;425–433]
The inhibitory effect of anti‐allergic agent suplatast tosilate (IPD–1151T) on methacholine‐ and allergen‐induced bronchoconstriction in sensitized mice
Authors: Asano Kazuhito , Mizutani Tetsuya , Shimane Toshikazu , Hisano Masataka , Hisamitsu Tadashi , Suzaki Harumi
Published: July 2002

The influence of an anti‐allergic agent, suplatast tosilate (IPD‐1151T; (±)‐[2‐[4‐(3‐ethoxy–2‐hydroxypropoxy)phenyl‐carbamoyl]‐ethyl] dimethylsulfonium p‐toluenesulfonate) on allergic bronchoconstriction induced by allergen and methacholine (MCh) were examined in mice. BALB/c mice were sensitized by intraperitoneal injection of dinitrophenylated‐keyhole limpet hemocyanin (DNP‐KLH) mixed with
Al(OH)3 (DNP‐KLH). IPD‐1151T was administered orally once a day for either 5 or 14 days in doses of 10, 30 or 100 mg/kg. Bronchoconstriction was measured 24 h after the final drug administration. IPD‐1151T inhibited both antigen‐ and MCh‐mediated bronchoconstriction in actively sensitized mice. The inhibition induced was closely related to the dose and frequency of oral administration of the agent. We also examined the effect of IPD‐1151T on IgE production in response to DNP‐KLH immunization. IPD‐1151T inhibited dose‐dependently both total and specific IgE concentrations in serum prepared from mice 15 days after im munization. These results strongly indicate that IPD‐1151T inhibits IgE
production in vivo and results in attenuating effect on bronchoconstriction.
The role of selective cyclooxygenase isoforms in human intestinal smooth muscle cell stimulated prostanoid formation and proliferation
Authors: Longo Walter E. , Erickson Brian , Panesar Ninder , Mazuski John E. , Robinson Sandra , Kaminski Donald L.
Published: July 2002

Intestinal smooth muscle plays a major role in the repair of injured intestine and contributes to the prostanoid pool during intestinal inflammatory states. Cyclooxygenase (COX), which catalyzes the conversion of arachidonic acid to prostanoids exists in two isoforms, COX‐1 and COX‐2. The purpose of this study was to determine the relative contributions of COX‐1 and COX‐2 in the production of prostanoids by human intestinal smooth muscle (HISM) cells when stimulated by interleukin‐1β (IL‐1β) and lipopolsaccharide (LPS). Furthermore the effects of specific COX‐1 and COX‐2 inhibitors on the proliferation of smooth muscle cells was also evaluated. Confluent monolayer cultures of HISM cells were incubated with IL‐1β or LPS for 0‐24 h while control cells received medium alone. PGE2 and PGI2 as 6‐keto‐PGF1α and LTB4 were measured by a specific radioimmunoassay. COX enzymes were evaluated by Western immunoblotting. Unstimulated and stimulated cells were exposed to the specific COX‐1 inhibitor valerylsalicylic acid (VSA) and the COX‐2 inhibitors NS‐398 and SC‐58125. The effects of serum on proliferation were then evaluated in the presence of each of the specific COX inhibitors by incorporation of 3H‐thym idine into DNA. IL‐1β and LPS increased both PGE2 and 6‐ketoPGF1α in a dose dependent fashion with enhanced production detected two hours following exposure. Neither stimulus stimulated LTB4 release. Immunoblot analysis using isoform‐specific antibodies showed that both COX‐1 and COX‐2 were present constitutively. Furthermore, COX‐1 was upregulated by each inflammatory stimulus. In a separate set of experiments cells were pretreated with either the selective COX‐1 inhibitor VSA or the selective COX‐2 inhibitors NS‐398 or SC‐58125 prior to treatment with IL‐1β or LPS. The COX‐1 and COX‐2 inhibitors decreased both basal and IL‐1β and LPS stimulated prostanoid release. Spontaneous DNA synthesis was present and serum consistently increased proliferation. 3H‐thymidine incorporation, stimulated by serum, was inhibited by both COX‐1 and COX‐2 inhibitors. This study suggests that the prostanoid response stimulated by proinflammatory agents of gut‐derived smooth muscle cells appears to be mediated by both COX‐1 and COX‐2 enzymes. Proliferation of smooth muscles cells also appears to be influenced by both COX‐1 and COX‐2.
Effect of platelet‐activating factor on the growth of human erythroid and myeloid CD34+ progenitors
Authors: Dupuis F. , Gachard N. , Allegraud A. , Dulery C. , Praloran V. , Denizot Y.
Published: July 2002

We have assessed the effect of platelet‐activating factor (PAF), a biologically active phospholipid present in the human marrow, on the growth of human marrow and blood CD34+ progenitors. While the metabolization rate of PAF by CD34+ cells is low (weak acetylhydrolase and acylation processes) it is readily catabolized by the acetylhydrolase activity present in the growth medium (10% fetal calf serum + 10% 5637‐conditioned medium). Treatment of marrow CD34+ cells with the non‐metabolizable PAF agonist C‐PAF (1 nM to 100 nM) immediately before semi‐solid culture significantly (p < 0.01) decreased the number of BFU‐E but not of CFU‐GM colonies. Treatment of marrow or blood CD34+ cells with C‐PAF (10‐100 nM) for 3 days in liquid medium before semi‐solid culture significantly (p < 0.01) decreased the number of BFUE and CFU‐GM colonies. Treatment of blood CD34+ cells with the two PAF receptor antagonists CV 3988 and BN 52021 (1 μ M) had no significant effect on the number of BFU‐E and CFU‐GM colonies suggesting no role of endogenous PAF in these processes. These results show that exogenous PAF downregulates human erythropoiesis and myelopoiesis, a result that might be of importance during inflammatory states.
(1 →3)‐ β ‐D‐Glucan and endotoxin modulate immune response to inhaled allergen
Authors: Rylander R. , Holt P. G.
Published: July 2002

Exposure to dust may involve co‐exposure to agents which are allergens, together with those which are pro‐inflamm atory. To study the effects of such a coexposure, the humoral and inflammatory responses were studied in guinea pigs inhaling the T‐cell dependent antigen ovalbumin (OVA) and the inflammatory agents (1 → 3)‐β ‐D‐glucan and lipopolysaccharide (LPS). The effects were evaluated as inflammatory cells in the lung and serum antibodies to OVA. LPS caused a stimulation of the OVA‐induced antibody production which was abolished by simultaneous exposure to (1 → 3)‐β‐D‐glucan. An increase of eosinophils after OVA exposure was decreased by coexposure to (1 → 3)‐β‐D‐glucan. The results demonstrate a complex interaction between adaptive and innate immune mechanisms in the lung, determined by exposure to common contaminants in airborne dust.
Aspirin and Some Other Nonsteroidal Anti‐Inflammatory Drugs Inhibit Cystic Fibrosis Transmembrane Conductance Regulator Protein Gene Expression in T‐84 Cells
Authors: Tondelier Danielle , Brouillard Franck , Lipecka Joanna , Labarthe Régis , Bali Moëz , De Beauregard Marie-Ayliette Costa , Torossi Tanja , Cougnon Marc , Edelman Aleksander , Baudouin-Legros Maryvonne
Published: July 2002

Cystic fibrosis (CF) is caused by mutations in the CF gene, which encodes CF transmembrane conductance regulator protein (CFTR), a transmembrane protein that acts as a cAMP‐regulated chloride channel. The disease is characterized by inflammation but the relationship between inflammation, abnormal transepithelial ion transport, and the clinical manifestations of CF are uncertain. The present study was undertaken to determine whether three nonsteroidal anti‐inflammatory drugs (NSAIDs) (aspirin, ibuprofen, and indomethacin) modulate CFTR gene expression in T‐84 cells. Treatment with NSAIDs reduced CFTR transcripts, and decreased cAMP‐stimulated anion fluxes, an index of CFTR function. However, the two phenomena occurred at different concentrations of both drugs. The results indicate that NSAIDs can regulate both CFTR gene expression and the function of CFTR‐related chloride transport, and suggest that NSAIDs act via multiple transduction pathways.
Production of Secretory Leucocyte Protease Inhibitor (SLPI) in Human Pancreatic β‐Cells
Authors: Nyström Max , Bergenfeldt Magnus , Ljungcrantz Irena , Lindeheim Èsa , Ohlsson Kjell
Published: July 2002

Secretory leucocyte protease inhibitor (SLPI) is a potent inhibitor of granulocyte elastase and cathepsin G, and also an inhibitor of pancreatic enzymes like trypsin, chymotrypsin and pancreatic elastase. SLPI has also been shown to inhibit HIV‐1 infections by blocking viral DNA synthesis. Since SLPI is an inhibitor of pancreatic proteases we wished to investigate whether SLPI was also actually produced in the pancreas. M‐RNA from human pancreatic tissue showed evidence of SLPI production using the reverse transcriptase polymer chain reaction technique (RTPCR). Using immunohistochemical methods SLPI was demonstrated in the β‐ce1ls of the islets of Langerhans. The function could be local protease/antiprotease regulation or antiviral/antibacterial defence in the close vicinity of the cell surface, or even inside the β‐cell itself.
Extracorporeal Circulation Causes Release of Neutrophil Gelatinase‐Associated Lipocalin (NGAL)
Authors: Jönsson Per , Ståhl Marie-Louise , Ohlsson Kjell
Published: July 2002

Extracorporeal circulation (ECC) used during cardiac surgery causes activation of several inflammatory systems. These events are not fully understood but are responsible for complications during the immediate postoperative period. Neutrophil gelatinase‐associated lipocalin (NGAL), a member of the expanding lipocalin family, has recently been described as an inflammatory protein. In this study, the release of NGAL into the circulation in 41 patients undergoing heart surgery with ECC was evaluated. A 4‐ to 5‐fold elevation of the concentration of NGAL in plasma was observed during the immediate postoperative course with a rapid elimination during the first postoperative day. Four patients undergoing lung surgery (without ECC) were also studied. The plasma concentration of NGAL only increased with a factor of 1.1‐2.2 over the operation. We conclude that NGAL is released into the circulation during heart surgery, probably as a result of the inflammatory activation of leukocytes initiated by the extracorporeal circulation.
Book Reviews
Authors: GOSSOP MICHAEL
Published: July 2002

Books Reviewed in this Article:

Clinical Guide to Alcohol Treatment. The Community Reinforcement Approach ROBERT J. MEYERS & JANE ELLEN SMITH

Tobacco and Health KAREN SLAMA (Ed.)

The Cigarette Papers STANTON A. GLANZ, JOHN SLADE, LISA A. BERO, PETER HANAUER & DEBORAH E. BARNES

Snow Job? The War against International Cocaine Trafficking KEVIN JACK RILEY

Substance and Shadow: Women and Addiction in the United States STEPHEN R. KANDALL
What We Have Learned about Addiction from Animal Models of Drug Self‐Administration
Authors: Gardner Eliot L.
Published: July 2002

Self‐administration of addictive drugs in laboratory animals has been widely used for decades as a tool for studying behavioral, neurobiological, and genetic factors in addiction. From such studies has come an enormous amount of information on brain mechanisms involved in addiction, on vulnerability factors in the addictive process, and on possible pharmacotherapeutic treatments for addiction. Modifications of the laboratory animal self‐administration paradigmö including progressive ratio break‐point models and the “reinstatement” model of relapse to drug‐seeking behaviorö are currently increasing our knowledge of incentive motivational factors in addiction and of the mechanisms underlying relapse to drug self‐administration behavior.
Effect of Gender, Treatment Site and Psychiatric Comorbidity on Quality of Life Outcome in Substance Dependence
Authors: Garg Nick , Yates William R. , Jones Rick , Zhou Miranda , Williams Sally
Published: July 2002

Patients receiving treatment for substance dependence frequently endorse high rates of psychological impairment and other measures of reduced quality of life. We conducted a baseline and six‐month follow‐up study of a series of one hundred and three unselected patients receiving treatment for a substance abuse or dependence problem. Women and patients requiring in‐patient detoxification demonstrated the most psychological impairment at baseline, as measured by the mental component summary of the SF‐36. Inpatient site of treatment was associated with continued psychological impairment six months following treatment. More aggressive psychiatric and psychological interventions may be indicated for women and for inpatient substance dependence populations.
Immune Effects of Oral 2,4,6-Trinitrotoluene (TNT) Exposure to the White-Footed Mouse, Peromyscus leucopus
Authors: Johnson Mark S. , Ferguson Jennifer W. , Holladay Steven D.
Published: July 2002

Immune toxicity associated with exposure to 2,4,6-trinitrotoluene (TNT) in a 14-day feeding study was examined in male and female white-footed mice ( Peromyscus leucopus). Mice (10/group/sex) were exposed to 0, 0.042, 0.083, 0.165, and 0.330% TNT in feed for 14 days. Based upon average feed consumption and body weight, these diets resulted in approximate daily doses of 66, 145, 275, and 601 mg TNT/kg body weight (bw) for males and 70, 142, 283, and 550 mg/kg/day for females. At the end of the exposure period the mice were euthanized and several indicators of nonspecific immunity were examined. These indicators included primary and secondary lymphoid organ/body weight ratio (spleen and thymus), and characterization of nonspecific immune responses (phagocytesis and radical oxygen intermediate [ROI] production). No deaths occurred, even though the high-dose group approached the reported LD50 in Swiss-Webster mice (Dilley et al. 1982. Toxicol. Environ. Health 9:565–585). Spleen weight was significantly increased in the high-dose group (0.330% TNT) for both sexes, whereas a dose-related trend in thymus cellularity was suggestive for males, but not females. In addition, males, but not females, displayed inhibited splenic macrophage phagocytosis and ROI production. Splenic congestion and extramedullary hematopoiesis were observed in both sexes in the two highest dose groups. These results suggest species-specific differences in relative subacute toxicity between laboratory (genus Mus) and Nearctic (genus Peromyscus) mice. In addition, these immunological indicators appear more sensitive than other toxicological endpoints that have been reported as most descriptive of TNT-related effects in mammals.
Silicone Breast Implants, Breast Cancer and Specific Connective Tissue Diseases: A Systematic Review of the Data in the Epidemiological Literature
Authors: Lamm Steven H.
Published: July 2002

Unanswered concerns about the systemic safety of silicone breast implants (BI) underlay the Food and Drug Administration's moratorium pronouncement in 1992. Since then, many epidemiological studies have been reported that examined either the association between BI and cancer, particularly breast cancer, or the association between BI and connective tissue diseases (CTD), particularly scleroderma. These studies are reviewed, and their data are synthesized. Three breast cancer easel control studies that examine BI as a risk factor show no association between BI and breast cancer. Nor do four BI cohort studies. The data appear to show a reduced risk. No association has been seen between Bl and either breast sarcomas or total cancers. Case-control studies do not show an association between BI and scleroderma (four studies), rheumatoid arthritis (three studies), systemic lupus erythematosus (two studies), or other connective tissue diseases. Eight cohort studies of women with breast implants sought an association between BI and CTD. Seven had negative results. One found a statistically significant risk of self-reported CTD of 1.24 (upper confidence limit = 1.41), but medical record review for diagnostic confirmation has not yet been performed. In toto, the epidemiological studies do not indicate an association between breast implants and breast cancer, though they suggest possibly a negative association. In toto, the epidemiological studies do not indicate an association between breast implants and specific connective tissue diseases, though one study's current results present a small statistically significant association with self-reported CTD.
Does Photosensitivity Predict Photococarcinogenicity?
Authors: Jacobs Abigail , Avalos Javier , Brown Paul , Wilkin Jonathan
Published: July 2002

Assessment of short-term and long-term effects of light (pho—totesting) is part of the safety evaluation of drugs. Results are incorporated into drug package inserts to advise patients and health care providers about the use of drug products on sun—exposed skin. We undertook an exhaustive literature search and a search of archived studies at the Center for Drug Evaluation and Research (CDER), Food and Drug Administration (FDA), in order to evaluate the potential of short—term photoassays to predict long—term effects of drugs used in sunlight (280–700 nm). The correlation between the findings from the photococarcinogenicity assays in mice that used exposure to simulated sunlight and those from photogenotoxicity and photosensitivity studies was examined. Results indicated that photosensitivity and photogenotoxicity assays did not necessarily predict effects in photococarcinogenicity studies in mice. Effects of drugs on skin that are not due to photoactivation of drug can be important factors in enhancement of UV—induced skin carcinogenesis.
The ubiquitin‐proteasome system and its role in ethanol‐induced disorders
Authors: Donohue Terrence M.
Published: July 2002

Abstract Some of the most fundamental yet important cellular activities such as cell division and gene expression are controlled by short‐lived regulatory proteins. The levels of these proteins are controlled by their rates of degradation. Similarly, protein catabolism plays a crucial role in prolonging cellular life by destroying damaged proteins that are potentially cytotoxic. A major player in these catabolic reactions is the ubiquitin‐proteasome system, a novel proteolytic system that has become the primary proteolytic pathway in eukaryotic cells. Ubiquitin‐mediated proteolysis is now regarded as the major pathway by which most intracellular proteins are destroyed. Equally important, from a toxicological standpoint, is that the ubiquitin‐proteasome system is also widely considered to be a cellular defense mechanism, since it is involved in the removal of damaged proteins generated by adduct formation and oxidative stress. This review describes the history and the components of the ubiquitin‐proteasome system, its regulation and its role in pathological states, with the major emphasis on ethanol‐induced organ injury. The available literature cited here deals mainly with the effects of ethanol consumption on the ubiquitin‐proteasome pathway in the liver. However, since this proteolytic system is an essential pathway in all cells it is an attractive experimental model and therapeutic target in extrahepatic organs such as the brain and heart that are also affected by excessive alcohol consumption.
Disruption of the MyoD/p21 Pathway in Rhabdomyosarcoma
Authors: Weintraub Michael , Kalebic Thea , Helman Lee J. , Bhatia Kishor G.
Published: July 2002

Purpose. Rhabdomyosarcoma (RMS) is an embryonal tumor thought to arise from skeletal muscle cells that fail to
differentiate terminally. The majority of RMSs express MyoD, a protein essential to the differentiation of skeletal muscle.
It was recently shown that during myogenesis, MyoD activates the expression of the cyclin‐dependent kinase inhibitor
(CDKi), p21, which itself plays a critical role in normal muscle development. To investigate the integrity of the MyoD/p21
pathway in RMS, we analyzed p21 and its relationship to MyoD expression in RMS.
Methods. A panel of RMS samples was assembled from primary biopsies and from cell lines. Integrity of p21 was analyzed
by single‐strand conformation polymorphism (SSCP) and sequencing. Expression of p21 and MyoD was determined by
Northern blot analysis, and the ability of exogenous p21 to arrest the cell cycle of RMS cell line was determined by
transfection studies.
Results. Our analysis indicates that although p21 is wild type in RMS, there is an inverse correlation between the levels
of p21 and MyoD in these tumors. Tumors that express significant amounts of MyoD fail to express p21. This does not
appear to be the result of mutations within the potential CACGTG sites present in the p21 promoter region or in the
coding region of p21. An additional group of RMSs express very high levels of p21 but express little, if any, MyoD.
Furthermore, RD, a RMS cell line which expresses high levels of endogenous p21, undergoes withdrawal from the cell
cycle following forced expression of p21, suggesting that the pathway which would lead to G1 arrest from endogenous p21 activity is defective.
Discussion. These data suggest that the interaction between p21 and MyoD is defective in RMS although the precise
nature of the defect remains to be elucidated.
Amplification and Over‐Expression of the MDM2 Gene in Human Soft Tissue Tumours
Authors: Patterson Helen , Barnes Diana , Gill Sandra , Spicer James , Fisher Cyril , Thomas Merion , Grimer Robert , Fletcher Chris , Gusterson Barry , Cooper Colin
Published: July 2002

Purpose. Amplification of genetic sequences on chromosome 12q13 is frequently found in soft tissue tumours. However,
for the MDM2 gene, over‐expression of the MDM2 protein has not always been shown to accompany gene amplification,
raising the possibility that amplification of genetic sequences targets alternative genes on chromosome 12q13 for
over‐expression. To investigate this discrepancy, we have examined 129 soft tissue tumours for amplification of the MDM2
gene using Southern analysis, and 39 of these tumours were also examined by immunohistochemical staining for MDM2
over‐expression.

Results. Gene amplification was identified in 14/114 (12.3%) of the malignant tumours, but was not identified in any of
the benign tumours; 21/39 (54%) of the malignant tumours also demonstrated MDM2 over‐expression. Within this group
the MDM2 gene was over‐expressed in every tumour in which the gene amplification was found, and over‐expression in
the absence of gene amplification was also found in an additional 10 tumours.

Discussion. These data demonstrate a clear correlation between the presence of MDM2 amplification and MDM2
over‐expression, and provide persuasive evidence therefore that the amplification of genetic sequences on chromosome
12q13 in soft tissue sarcomas targets the MDM2 gene for over‐expression. These data also indicate that alternative
mechanisms may contribute to MDM2 over‐expression within some tumours.
The Activity of Collagenase-1 Is Required for Keratinocyte Migration on a Type I Collagen Matrix
Authors: Pilcher Brian K. , Dumin Jo Ann , Sudbeck Barry D. , Krane Stephen M. , Welgus Howard G. , Parks William C.
Published: July 2002

We have shown in a variety of human wounds that collagenase-1 (MMP-1), a matrix metalloproteinase that cleaves fibrillar type I collagen, is invariably expressed by basal keratinocytes migrating across the dermal matrix. Furthermore, we have demonstrated that MMP-1 expression is induced in primary keratinocytes by contact with native type I collagen and not by basement membrane proteins or by other components of the dermal or provisional (wound) matrix. Based on these observations, we hypothesized that the catalytic activity of MMP-1 is necessary for keratinocyte migration on type I collagen. To test this idea, we assessed keratinocyte motility on type I collagen using colony dispersion and colloidal gold migration assays. In both assays, primary human keratinocytes migrated efficiently on collagen. The specificity of MMP-1 in promoting cell movement was demonstrated in four distinct experiments. One, keratinocyte migration was completely blocked by peptide hydroxymates, which are potent inhibitors of the catalytic activity of MMPs. Two, HaCaTs, a line of human keratinocytes that do not express MMP-1 in response to collagen, did not migrate on a type I collagen matrix but moved efficiently on denatured type I collagen (gelatin). EGF, which induces MMP-I production by HaCaT cells, resulted in the ability of these cells to migrate across a type I collagen matrix. Three, keratinocytes did not migrate on mutant type I collagen lacking the collagenase cleavage site, even though this substrate induced MMP-1 expression. Four, cell migration on collagen was completely blocked by recombinant tissue inhibitor of metalloproteinase-1 (TIMP-1) and by affinity-purified anti–MMP-1 antiserum. In addition, the collagen-mediated induction of collagenase-1 and migration of primary keratinocytes on collagen was blocked by antibodies against the α2 integrin subunit but not by antibodies against the α1 or α3 subunits. We propose that interaction of the α2β1 integrin with dermal collagen mediates induction of collagenase-1 in keratinocytes at the onset of healing and that the activity of collagenase-1 is needed to initiate cell movement. Furthermore, we propose that cleavage of dermal collagen provides keratinocytes with a mechanism to maintain their directionality during reepithelialization.
An Endothelial Storage Granule for Tissue-Type Plasminogen Activator
Authors: Emeis J.J. , Eijnden-Schrauwen Y. van den , Hoogen C.M. van den , Priester W. de , Westmuckett A. , Lupu F.
Published: July 2002

In previous studies we have shown that, after stimulation by a receptor ligand such as thrombin, tissue-type plasminogen activator (tPA) and von Willebrand factor (vWf) will be acutely released from human umbilical vein endothelial cells (HUVEC). However, the mechanisms involved in the secretion of these two proteins differ in some respects, suggesting that the two proteins may be stored in different secretory granules.

By density gradient centrifugation of rat lung homogenates, a particle was identified that contained nearly all tPA activity and antigen. This particle had an average density of 1.11–1.12 g/ml, both in Nycodenz density gradients and in sucrose density gradients. A similar density distribution of tPA was found for a rat endothelial cell line and for HUVEC. After thrombin stimulation of HUVEC to induce tPA secretion, the amount of tPA present in high-density fractions decreased, concomitant with the release of tPA into the culture medium and a shift in the density distribution of P-selectin.

vWf, known to be stored in Weibel-Palade bodies, showed an identical distribution to tPA in Nycodenz gradients. In contrast, the distribution in sucrose gradients of vWf from both rat and human lung was very different from that of tPA, suggesting that tPA and vWf were not present in the same particle.

Using double-immunofluorescence staining of HUVEC, tPA- and vWf-containing particles showed a different distribution by confocal microscopy. The distribution of tPA also differed from the distribution of tissue factor pathway inhibitor, endothelin-1, and caveolin. By immunoelectronmicroscopy, immunoreactive tPA could be demonstrated in small vesicles morphologically different from the larger Weibel-Palade bodies. It is concluded that tPA in endothelial cells is stored in a not-previously-described, small and dense (d = 1.11– 1.12 g/ml) vesicle, which is different from a Weibel-Palade body.
Defect in Synaptic Vesicle Precursor Transport and Neuronal Cell Death in KIF1A Motor Protein–deficient Mice
Authors: Yonekawa Yoshiaki , Harada Akihiro , Okada Yasushi , Funakoshi Takeshi , Kanai Yoshimitsu , Takei Yosuke , Terada Sumio , Noda Tetsuo , Hirokawa Nobutaka
Published: July 2002

The nerve axon is a good model system for studying the molecular mechanism of organelle transport in cells. Recently, the new kinesin superfamily proteins (KIFs) have been identified as candidate motor proteins involved in organelle transport. Among them KIF1A, a murine homologue of unc-104 gene of Caenorhabditis elegans, is a unique monomeric neuron– specific microtubule plus end–directed motor and has been proposed as a transporter of synaptic vesicle precursors (Okada, Y., H. Yamazaki, Y. Sekine-Aizawa, and N. Hirokawa. 1995. Cell. 81:769–780). To elucidate the function of KIF1A in vivo, we disrupted the KIF1A gene in mice. KIF1A mutants died mostly within a day after birth showing motor and sensory disturbances. In the nervous systems of these mutants, the transport of synaptic vesicle precursors showed a specific and significant decrease. Consequently, synaptic vesicle density decreased dramatically, and clusters of clear small vesicles accumulated in the cell bodies. Furthermore, marked neuronal degeneration and death occurred both in KIF1A mutant mice and in cultures of mutant neurons. The neuronal death in cultures was blocked by coculture with wild-type neurons or exposure to a low concentration of glutamate. These results in cultures suggested that the mutant neurons might not sufficiently receive afferent stimulation, such as neuronal contacts or neurotransmission, resulting in cell death. Thus, our results demonstrate that KIF1A transports a synaptic vesicle precursor and that KIF1A-mediated axonal transport plays a critical role in viability, maintenance, and function of neurons, particularly mature neurons.
Mass Transport of Proform of a Kdel-Tailed Cysteine Proteinase (Sh-EP) to Protein Storage Vacuoles by Endoplasmic Reticulum–Derived Vesicle Is Involved in Protein Mobilization in Germinating Seeds
Authors: Toyooka Kiminori , Okamoto Takashi , Minamikawa Takao
Published: July 2002

A vacuolar cysteine proteinase, designated SH-EP, is expressed in the cotyledon of germinated Vigna mungo seeds and is responsible for the degradation of storage proteins. SH-EP is a characteristic vacuolar proteinase possessing a COOH-terminal endoplasmic reticulum (ER) retention sequence, KDEL. In this work, immunocytochemical analysis of the cotyledon cells of germinated V. mungo seeds was performed using seven kinds of antibodies to identify the intracellular transport pathway of SH-EP from ER to protein storage vacuoles. A proform of SH-EP synthesized in ER accumulated at the edge or middle region of ER where the transport vesicle was formed. The vesicle containing a large amount of proSH-EP, termed KV, budded off from ER, bypassed the Golgi complex, and was sorted to protein storage vacuoles. This massive transport of SH-EP via KV was thought to mediate dynamic protein mobilization in the cotyledon cells of germinated seeds. We discuss the possibilities that the KDEL sequence of KDEL-tailed vacuolar cysteine proteinases function as an accumulation signal at ER, and that the mass transport of the proteinases by ER-derived KV-like vesicle is involved in the protein mobilization of plants.
In Vivo Analysis of Cajal Body Movement, Separation, and Joining in Live Human Cells
Authors: Platani Melpomeni , Goldberg Ilya , Swedlow Jason R. , Lamond Angus I.
Published: July 2002

Cajal bodies (also known as coiled bodies) are subnuclear organelles that contain specific nuclear antigens, including splicing small nuclear ribonucleoproteins (snRNPs) and a subset of nucleolar proteins. Cajal bodies are localized in the nucleoplasm and are often found at the nucleolar periphery. We have constructed a stable HeLa cell line, HeLaGFP-coilin, that expresses the Cajal body marker protein, p80 coilin, fused to the green fluorescent protein (GFP-coilin). The localization pattern and biochemical properties of the GFP-coilin fusion protein are identical to the endogenous p80 coilin. Time-lapse recordings on 63 nuclei of HeLaGFP-coilin cells showed that all Cajal bodies move within the nucleoplasm. Movements included translocations through the nucleoplasm, joining of bodies to form larger structures, and separation of smaller bodies from larger Cajal bodies. Also, we observed Cajal bodies moving to and from nucleoli. The data suggest that there may be at least two classes of Cajal bodies that differ in their size, antigen composition, and dynamic behavior. The smaller size class shows more frequent and faster rates of movement, up to 0.9 μm/min. The GFP-coilin protein is dynamically associated with Cajal bodies as shown by changes in their fluorescence intensity over time. This study reveals an unexpectedly high level of movement and interactions of nuclear bodies in human cells and suggests that these movements may be driven, at least in part, by regulated mechanisms.
Differential Localization of Rho Gtpases in Live Cells
Authors: Michaelson David , Silletti Joseph , Murphy Gretchen , D'Eustachio Peter , Rush Mark , Philips Mark R.
Published: July 2002

Determinants of membrane targeting of Rho proteins were investigated in live cells with green fluorescent fusion proteins expressed with or without Rho-guanine nucleotide dissociation inhibitor (GDI)α. The hypervariable region determined to which membrane compartment each protein was targeted. Targeting was regulated by binding to RhoGDIα in the case of RhoA, Rac1, Rac2, and Cdc42hs but not RhoB or TC10. Although RhoB localized to the plasma membrane (PM), Golgi, and motile peri-Golgi vesicles, TC10 localized to PMs and endosomes. Inhibition of palmitoylation mislocalized H-Ras, RhoB, and TC10 to the endoplasmic reticulum. Although overexpressed Cdc42hs and Rac2 were observed predominantly on endomembrane, Rac1 was predominantly at the PM. RhoA was cytosolic even when expressed at levels in vast excess of RhoGDIα. Oncogenic Dbl stimulated translocation of green fluorescent protein (GFP)-Rac1, GFP-Cdc42hs, and GFP-RhoA to lamellipodia. RhoGDI binding to GFP-Cdc42hs was not affected by substituting farnesylation for geranylgeranylation. A palmitoylation site inserted into RhoA blocked RhoGDIα binding. Mutations that render RhoA, Cdc42hs, or Rac1, either constitutively active or dominant negative abrogated binding to RhoGDIα and redirected expression to both PMs and internal membranes. Thus, despite the common essential feature of the CAAX (prenylation, AAX tripeptide proteolysis, and carboxyl methylation) motif, the subcellular localizations of Rho GTPases, like their functions, are diverse and dynamic.
Cotyledon cells of Vigna mungo seedlings use at least two distinct autophagic machineries for degradation of starch granules and cellular components
Authors: Toyooka Kiminori , Okamoto Takashi , Minamikawa Takao
Published: July 2002

α-Amylase is expressed in cotyledons of germinated Vigna mungo seeds and is responsible for the degradation of starch that is stored in the starch granule (SG). Immunocytochemical analysis of the cotyledon cells with anti–α-amylase antibody showed that α-amylase is transported to protein storage vacuole (PSV) and lytic vacuole (LV), which is converted from PSV by hydrolysis of storage proteins. To observe the insertion/degradation processes of SG into/in the inside of vacuoles, ultrastructural analyses of the cotyledon cells were conducted. The results revealed that SG is inserted into LV through autophagic function of LV and subsequently degraded by vacuolar α-amylase. The autophagy for SG was structurally similar to micropexophagy detected in yeast cells. In addition to the autophagic process for SG, autophagosome-mediated autophagy for cytoplasm and mitochondria was detected in the cotyledon cells. When the embryo axes were removed from seeds and the detached cotyledons were incubated, the autophagosome-mediated autophagy was observed, but the autophagic process for the degradation of SG was not detected, suggesting that these two autophagic processes were mediated by different cellular mechanisms. The two distinct autophagic processes were thought to be involved in the breakdown of SG and cell components in the cells of germinated cotyledon.