bluuebunny/crossref_metadata_2025 huggingface dataset:
β-Endorphin blocks luteinizing hormone-releasing hormone release by inhibiting the nitricoxidergic pathway controlling its release
Authors: Faletti Alicia G. , Mastronardi Claudio A. , Lomniczi Alejandro , Seilicovich Adriana , Gimeno Martha , McCann Samuel M. , Rettori Valeria
Published: July 2002

β-Endorphin blocks release of luteinizing hormone (LH)-releasing
hormone (LHRH) into the hypophyseal portal vessels by stimulating
μ-opiate receptors, thereby inhibiting secretion of LH. LHRH release
is controlled by release of nitric oxide from nitricoxidergic (NOergic)
neurons in the basal tuberal hypothalamus. To determine whether
β-endorphin exerts its inhibitory action on this NOergic pathway,
medial basal hypothalami (MBH) from male rats were incubated with
β-endorphin (10
−8
M). β-Endorphin decreased basal
secretion of LHRH, and significantly inhibited the release of
prostaglandin E
2
(PGE
2
), a known stimulant of
LHRH release. Incubation of MBH with β-endorphin at various
concentrations (10
−9
–10
−6
M)
in
vitro
decreased the activity of NO synthase (NOS) (measured by
the conversion of [
14
C]arginine to labeled citrulline).
Conversely, the activity of NOS was increased by the μ-receptor
antagonist, naltrexone (10
−8
M). Not only was the
inhibitory action of β-endorphin on LHRH and PGE
2
release
blocked by naltrexone (10
−8
M), but it increased NOS
activity and LHRH and PGE
2
release. β-Endorphin also
stimulated γ-aminobutyric acid (GABA) release. Because GABA inhibits
both nitroprusside (NP-induced PGE
2
and LHRH release by
blocking the activation of cyclooxygenase by NO, this is another
mechanism by which β-endorphin inhibits NP-induced PGE
2
and LHRH release. The results indicate that β-endorphin stimulates
μ-opioid receptors on NOergic neurons to inhibit the activation and
consequent synthesis of NOS in the MBH. β-Endorphin also blocks the
action of NO on PGE
2
release and, consequently, on LHRH
release, by stimulating GABAergic inhibitory input to LHRH terminals
that blocks NO-induced activation of cyclooxygenase and consequent
PGE
2
secretion.
Lysozyme and RNases as anti-HIV components in β-core preparations of human chorionic gonadotropin
Authors: Lee-Huang Sylvia , Huang Paul L. , Sun Yongtao , Huang Philip L. , Kung Hsiang-fu , Blithe Diana L. , Chen Hao-Chia
Published: July 2002

Human chorionic gonadotropin (hCG) preparations contain activity against HIV type 1 (HIV-1). However, there has been controversy about whether some biological activities of hCG β-subunit (hCGβ) preparations are caused by the β-subunit itself or other proteins present in the preparations. We report here the purification, characterization, and identification of three enzymes with anti-HIV activity present in the β-core fraction of hCGβ prepared from the urine of pregnant women. The N-terminal amino acid sequence of one protein is identical to human urinary lysozyme C, and those of the other two are identical to human RNase A and urinary RNase U. We thus refer to these proteins as AVL (antiviral lysozyme) and AVR (antiviral RNases). In addition to HIV-1 inhibition, AVL is capable of lysing
Micrococcus lysodeikticus
.
AVR digests a variety of RNA substrates, including RNA from HIV-1-infected cells. We also find that lysozyme from chicken egg white, human milk, and human neutrophils and RNase A from bovine pancreas possess activity against HIV-1. These findings may offer additional strategies for the treatment of HIV-1 infection.
Three-dimensional structure of poliovirus receptor bound to poliovirus
Authors: Belnap David M. , McDermott Brian M. , Filman David J. , Cheng Naiqian , Trus Benes L. , Zuccola Harmon J. , Racaniello Vincent R. , Hogle James M. , Steven Alasdair C.
Published: July 2002

Poliovirus initiates infection by binding to its cellular receptor (Pvr). We have studied this interaction by using cryoelectron microscopy to determine the structure, at 21-Å resolution, of poliovirus complexed with a soluble form of its receptor (sPvr). This density map aided construction of a homology-based model of sPvr and, in conjunction with the known crystal structure of the virus, allowed delineation of the binding site. The virion does not change significantly in structure on binding sPvr in short incubations at 4°C. We infer that the binding configuration visualized represents the initial interaction that is followed by structural changes in the virion as infection proceeds. sPvr is segmented into three well-defined Ig-like domains. The two domains closest to the virion (domains 1 and 2) are aligned and rigidly connected, whereas domain 3 diverges at an angle of ≈60°. Two nodules of density on domain 2 are identified as glycosylation sites. Domain 1 penetrates the “canyon” that surrounds the 5-fold protrusion on the capsid surface, and its binding site involves all three major capsid proteins. The inferred pattern of virus–sPvr interactions accounts for most mutations that affect the binding of Pvr to poliovirus.
T cell immunity to copolymer 1 confers neuroprotection on the damaged optic nerve: Possible therapy for optic neuropathies
Authors: Kipnis Jonathan , Yoles Eti , Porat Ziv , Cohen Avi , Mor Felix , Sela Michael , Cohen Irun R. , Schwartz Michal
Published: July 2002

We recently reported that the posttraumatic spread of degeneration in the damaged optic nerve can be attenuated by the adoptive transfer of autoimmune T cells specific to myelin basic protein. However, it would be desirable to obtain immune neuroprotection free of any possible autoimmune disease. In an attempt to obtain disease-free immune neuroprotection, we used the synthetic four-amino acid polymer copolymer 1 (Cop-1), which is known not to be encephalitogenic despite its cross-reactivity with myelin basic protein. We show here that active immunization with Cop-1 administered in adjuvant, as well as adoptive transfer of T cells reactive to Cop-1, can inhibit the progression of secondary degeneration after crush injury of the rat optic nerve. These results have implications for the treatment of optic neuropathies.
Glucose stimulates protein modification by
O
-linked GlcNAc in pancreatic β cells: Linkage of
O
-linked GlcNAc to β cell death
Authors: Liu Kan , Paterson Andrew J. , Chin Edward , Kudlow Jeffrey E.
Published: July 2002

The pancreatic β cell can respond in the long term to hyperglycemia both with an increased capacity for insulin production and, in susceptible individuals, with apoptosis. When glucose-induced apoptosis offsets the increasing β cell capacity, type 2 diabetes results. Here, we tested the idea that the pathway of glucose metabolism that leads to the modification of intracellular proteins with the
O
-linked monosaccharide
N-
acetylglucosamine (
O
-GlcNAc) is involved in the glucose-induced apoptosis. This idea is based on two recent observations. First, the β cell expresses much more
O
-GlcNAc transferase than any other known cell, and second, that the β cell-specific toxin, streptozotocin (STZ), itself a GlcNAc analog, specifically blocks the enzyme that cleaves
O
-GlcNAc from intracellular proteins. As a consequence, we now show that hyperglycemia leads to the rapid and reversible accumulation of
O
-GlcNAc specifically in β cells
in vivo
. Animals pretreated with STZ also accumulate
O
-GlcNAc in their β cells when hyperglycemic, but this change is sustained upon re-establishment of euglycemia. In concert with the idea that STZ toxicity results from the sustained accumulation of
O
-GlcNAc after a hyperglycemic episode, we established a low-dose STZ protocol in which the β cells' toxicity of STZ was manifest only after glucose or glucosamine administration. Transgenic mice with impaired β cell glucosamine synthesis treated with this protocol are resistant to the diabetogenic effect of STZ plus glucose yet succumb to STZ plus glucosamine. This study provides a causal link between apoptosis in β cells and glucose metabolism through glucosamine to
O
-GlcNAc, implicating this pathway of glucose metabolism with β cell glucose toxicity.
High metastatic potential in mice inheriting a targeted p53 missense mutation
Authors: Liu Geng , McDonnell Timothy J. , Montes de Oca Luna Roberto , Kapoor Mini , Mims Betsy , El-Naggar Adel K. , Lozano Guillermina
Published: July 2002

To understand the relevance of
p53
missense mutations
in vivo
, we generated a mouse containing an arg-to-his substitution at p53 amino acid 172, which corresponds to the R175H hot-spot mutation in human tumors by homologous recombination. Inadvertently, this mouse contains the additional deletion of a G nucleotide at a splice junction that attenuates levels of mutant p53 to near wild-type levels. Mice heterozygous for the mutant allele differed from
p53
+/−
mice in tumor spectrum, with a significant increase in the number of carcinomas and a slight decrease in the number of lymphomas. More importantly, the osteosarcomas and carcinomas that developed in these mutant mice frequently metastasized (69% and 40%, respectively). In contrast, metastasis is rare in osteosarcomas of
p53
+/−
mice. Loss of heterozygosity studies of tumors indicated loss of heterozygosity in only 1 of 11 tumors. These data indicate clear differences between a
p53
missense mutation and a null allele in tumorigenesis
in vivo
and suggest that the p53R172HΔg mutant represents a gain-of-function allele.
Noise versus Access: The Impact of an Airport in an Urban Property Market
Authors: Tomkins J. , Topham N. , Twomey J. , Ward R.
Published: July 2002

The effects of a major airport are unlikely to exhibit a uniform spatial distribution. The benefits to industries and individual households may extend well beyond the local economy, whereas many of the costs are spatially concentrated in the immediate environment. In particular, the problems of noise and traffic generation can be expected to fall principally upon adjacent populations. This paper addresses the general question of whether the costs to local economies of airport proximity, which are in the nature of externalities, outweigh the benefits of access, employment and improved infrastructure. Based on data relating to Manchester airport and its surrounding areas, the specific approach adopted in the paper involves an investigation of the extent to which such proximity effects are capitalised into residential property prices. Our results provide some evidence to suggest that circumstances may exist where positive attributes, such as improved access and employment opportunities, may be more highly valued by local residents than the negative externality effects of airport proximity.
Positivity in the aged’s perceptions of intergenerational relationships: A “stake” or “leniency” effect?
Authors: Winkeler Markus , Filipp Sigrun-Heide , Boll Thomas
Published: July 2002

The “developmental stake hypothesis” has been proposed for the frequent finding that aged parents consistently report higher levels of closeness to and consensus with their (adult) children than these children do themselves. This study investigated an alternative hypothesis: Drawing on research on prosocial behaviour in old age, it proposes that the aged tend to construe all social relationships in a positively biased manner (“leniency hypothesis”). Using a quasi-experimental (vignette) approach, scenarios describing two family members discussing a controversial issue were presented to 809 middle-aged (aged 40 to 50 years) and aged subjects (over-65s). The lineage composition of the dyads of family members in the scenarios (i.e. aged parent and adult child vs. two adult siblings) was varied systematically as a between-subjects factor, and the controversial issue was varied as a within-subjects factor. Dependent variables were the participants’ evaluative and prescriptive judgements on the protagonists’ behaviour and the quality of their relationship. Overall, results showed that the aged perceived all scenarios in a significantly more positive light than middle-aged participants, regardless of their lineage composition. Thus, a “positivity bias” was observed in judgements of both intergenerational as well as intragenerational dyads, and it is concluded that the leniency hypothesis provides a better account of these findings than the stake hypothesis.
Interpersonal relationships in Brazilian adolescents
Authors: Van Horn K. Roger , Marques Juracy Cunegatto
Published: July 2002

Interpersonal relationships were evaluated in 260 middle-class Brazilian youths aged 11–12, 15–16, and 19–20 years, using the Network of Relationships Inventory (Furman & Buhrmester, 1992). Participants rated four dimensions (support, conflict, relative power, and punishment) of relationships with five social network members (mother, father, teacher, sibling, and same-sex friend). Ratings were consistent with ecological models predicting culture-specific characteristics of relationships. In contrast to previous research on US adolescents, Brazilian adolescents reported high levels of both support and conflict and there was no shift from parents to peers as the primary source of support. Differences between early and late adolescents were not consistent with the processes of individuation and rapprochement. One of the few significant sex differences resulted from females rating support higher than males in relationships with siblings and friends.
Taxonomic structure in early to middle childhood: A longitudinal study with Zimbabwean schoolchildren
Authors: Mpofu Elias , van de Vijver Fons J.R.
Published: July 2002

Children’s classification reasoning was examined with longitudinal data for 103 Zimbabwean Black (47) and White (56) children attending a randomly selected sample of public schools. The children varied by gender, social class membership (lower, middle, upper) and race (black, white). The children attempted a set of classification tasks at ages 7, 9, and 11. Responses to the classification tasks were scored in terms of interpretive strategy used to engage the tasks (taxonomic vs. instrumental). Repeated measures MANOVA and post-hoc orthogonal contrasts yielded significant differences in interpretive strategies by age or level of schooling, and social class. Higher social class membership was significantly related to more frequent use of taxonomic rather than functional classification strategies. Results support age/schooling-related effects in the development of taxonomic structure in a non-Western society.
Problems Reported by Parents of Romanian Orphans Adopted to British Columbia
Authors: Fisher Lianne , Ames Elinor W. , Chisholm Kim , Savoie Lynn
Published: July 2002

Behaviour problems in Romanian orphans adopted to Canada were examined through parents’ interview reports of specific problems, and children’s scores on the Child Behavior Checklist (CBCL) (Achenbach, Edelbrock, & Howell, 1987) completed by their parents. Three groups of children were studied. Romanian orphanage (RO) children had spent at least 8 months in a Romanian orphanage. Parents’ reports of RO children’s problems were compared to parent reports from 2 comparison groups: (1) Canadian-born children (CB) who were not adopted and never institutionalised; and (2) children who would have gone to a Romanian orphanage had they not been adopted before 4 months of age (Romanian Comparison: RC). RO children scored higher than CB and RC children for Total problems and Internalizing problems on the CBCL. No significant differences were found for any group comparison on Externalizing problems. CBCL scores were positively correlated with RO children’s total time in orphanage. According to parent interview, RO children had more eating problems, medical problems, and stereotyped behaviour problems than both CB and RC children. These problems were distinctive ones, rarely if ever being reported for CB or RC children. It is suggested that these distinctive RO problems arise out of a normal developmental base, and reflect either continuations of orphanage behaviours, reactions to stimuli different from those experienced in orphanage, or lack of opportunity for development or learning within the orphanage.
Gastric Neuroendocrine Tumors in a 2-Year Oncogenicity Study With CD-1 Mice
Authors: Thoolen Bob , Koster Henk , Van Kolfschoten Anton , De Haan Martin
Published: July 2002

Descriptions of two rare gastric neuroendocrine tumors (carcinoids) of enterochromaffin (ECL) cells in CD-1 mice (2/50) from a 104-week oncogenicity study of a serotonergic/dopaminergic compound are presented. These tumors were detected at necropsy and confirmed by histopathology in hematoxylin and eosin- and Chromogranin A-stained slides. ECL cell counts of the glandular stomachs were determined by quantitative image analysis and did not reveal any hyperplastic changes as possible predisposing lesions for carcinoid formation. To investigate the possibility of drug-induced hypergastrinemi a as the cause of tumor formation of ECL cells, gastrin blood levels were measured after treating mice for 7 days with the test substance. In this study, Omeprazole, the positive control, raised gastrin levels, while the test material did not. It was concluded that these two tumors were an example of “late-life”-occurring, spontaneous neuroendocrine tumors in the stomachs of aged CD-1 mice.
An Immunohistochemical Label to Facilitate Counting of Ovarian Follicles
Authors: Muskhelishvili Levan , Freeman Lisa D. , Latendresse John R. , Bucci Thomas J.
Published: July 2002

U.S. and internationally harmonized Health Effects Test Guidelines for Reproduction and Fertility Effects include enumeration of primordial and developing ovarian follicles as endpoints of safety tests, and the number of these structures is also of interest for other aspects of reproductive biology. Performing the counts microscopically on representative hematoxylin and eosin (H&E)-stained sections of ovary is tedious and error-prone. The ability to mark oocyte nuclei distinctly with an antibody significantly increases speed and accuracy of counting. We have identifi ed a rabbit polyclonal antibody directed against a synthetic 14-amino acid sequence from human cytochrome P-450 1B1 (CYP1B1) that unequivocally marks rodent oocyte nuclei, in addition to nuclei of some ovarian granulosa and theca cells. Follicles of all degrees of maturity are easily distinguished from ovarian background ; ability to detect and identify primordial follicles is particularly enhanced. High-contrast and high-resolution labeling was achieved with routine immunohistochemical procedures using an avidin—biotin —peroxidase method on rat and mouse tissues fixed in 10% neutral buffered formalin.
Indigenous Australians who inject drugs: results from a Brisbane study
Authors: LARSON ANN , SHANNON CINDY , ELDRIDGE CHRIS
Published: July 2002

Injecting drug use by Aboriginal and Torres Strait Islander people has recently generated considerable concern in communities and by public health services and planners. However, there has been almost no research on the use of illicit drugs by indigneous people. This study is the first step in filling this information gap. Through a process of extensive community consultation the important issues were identified and a culturally appropriate research method was developed. A questionnaire was administered by peer interviewers to 77 Aboriginal and Torres Strait Islander peoples who had injected an illegal drug in the past 12 months and who lived in the Brisbane metropolitan area. Speed is the most commonly injected drug but 56% had tried heroin and for about one‐third of the sample it was the last drug injected. Sharing injecting equipment is common in this group, with 39% reporting that they had shared a needle in the past month; among participants under 20 years old the percentage rises to 63. Youth and/or inexperience is also associated with unsafe needle disposal and having never personally obtained injecting equipment from a chemist or needle exchange. There is widespread ignorance about safe procedures for cleaning needles. Knowledge about HIV/AIDS is not universal and less than one‐quarter personally know someone living with HIV/AIDS; 67% rank their own chances of becoming infected with HIV as very low or low. The results have been used to develop programmes and strategies for harm reduction locally.
The Role of Social Support Following Short‐Term Inpatient Treatment
Authors: Broome Kirk M. , Simpson D. Dwayne , Joe George W.
Published: July 2002

The intensive, time‐limited, short‐term inpatient modality treatment for substance abuse appears to have positive outcomes despite its brevity. This study examined patient characteristics and posttreatment experiences to understand who is likely to benefit from this treatment and under what circumstances. Our sample included 748 patients in 12 short‐term inpatient programs. Twenty‐two percent of patients used cocaine at least weekly in the 1‐year follow‐up period, and an additional 9% drank frequently (compared with pretreatment rates of 69% and 15% respectively). Overall, patients' social support networks following treatment were more important factors than the pre‐ or during‐treatment variables examined.
Hepatic haemodynamics and reticuloendothelial function in the rat in response to chronic ethanol administration
Authors: PREEDY V. R. , NOTT D. M. , YATES J. , VENKATESAN S. , JENKINS S. A. , PETERS T. J.
Published: July 2002

Abstract This study investigated the effects of chronic ethanol intake on hepatic haemodynamics and reticuloendothelial system function in the rat. Comparisons were also made with blood flow to pancreas, kidney, spleen, lung and skin. Male Wistar rats, approximately 0.15 kg initial body weight, were fed a diet containing 35% of the total calories as ethanol. Controls were pair‐fed identical amounts of the same diet in which ethanol was replaced by isocaloric glucose. The hepatic perfusion index and reticuloendothelial function was determined with [99m Tc]‐labelled sulphur colloid [99m Tc]‐SC) and blood flow with radiolabelled microspheres under anaesthesia. After 4‐5 weeks the weights of liver and skin of alcohol fed rats decreased by 10% (p= 0.040) and 23% (p= 0.024), respectively, compared to controls and there was a small increase in kidney weight (15%, p = 0.001). Blood flow to liver, pancreas, kidney, spleen, lung and skin was not altered significantly by chronic alcohol administration, irrespective of whether the data were expressed as a percentage of cardiac output, blood flow per minute per organ or blood flow per minute per g tissue weight (p > 0.113 in all instances). However, there was a significant increase in splenic reticuloendothelial system activity (+ 121%, p = 0.018). Hepatic reticuloendothelial system activity was also increased (+ 22%, p = 0.061). Chronic alcohol administration resulted in significant increases in portal pressure (+ 55%, p = 0.042) and portal venous resistance (+ 66%, p = 0.001), but portal venous inflow and hepatic perfusion index were not altered compared to controls The results of this study indicated that chronic alcohol administration did not alter visceral blood flow significantly, but did increase portal pressure, portal vascular resistance and reticuloendothelial system activity.
Effect of cessation of drug misuse on the immune status of HIV‐infected injecting drug users
Authors: RADKOWSKI MAREK , WEREZYNSKA TERESA , LASKUS TOMASZ , KAMINSKA EWA , KULIGOWSKI STANISLAW
Published: July 2002

Abstract The effects of cessation of i.v. drug use on CD4 lymphocyte count and serum levels of beta2 ‐microglobulin (beta2 m) and soluble interleukin‐2 receptor (sIL‐2R) were prospectively investigated in 41 HIV‐positive drug users enrolled in a methadone treatment programme. Fifteen HIV‐positive matched non‐abstainers served as controls. During the follow‐up of 12 months the mean CD4 lymphocyte count remained unchanged in the methadone‐treated group but fell significantly in controls. beta2 m and sIL‐2R values decreased in abstainers at the beginning of the treatment but rose in non‐abstaining group. We conclude that cessation of illicit drug abuse and methadone treatment in HIV‐infected injecting drug users may exert a beneficial effect on the immune status.
Continuous cocaine treatment and monoamine transmission measured by microdialysis in the rat ventral tegmental area
Authors: Li Ming‐YA , Reith Maarten E. A.
Published: July 2002

Rats were treated on a continuous cocaine administration schedule (85 mumol/kg/day s.c. by osmotic minipumps for 2 weeks followed by a 1‐week drug‐free period) that makes them tolerant to the locomotor‐activating effect of cocaine. Subsequently, extracellular serotonin (5‐HT) and dopamine (DA) were assayed in the ventral tegmental area (VTA) by microdialysis in the awake animal. Local infusion of the 5‐HT 1A, B, D, 2, 6, 7 receptor antagonist methiothepin (50 muM for 3 hours) into the VTA caused a smaller increase in dialysate 5‐HT in cocaine‐ than saline‐pretreated rats (902±189 versus 1630±210%; area under the curve, p<0.02), whereas the increase in DA was small and unchanged (281±50 versus 290±76%; area under the curve, p=0.09). The results provide preliminary support for a cocaine‐induced subsensitivity of 5‐HT 1B autoreceptors in the VTA in agreement with our previous results obtained ex vivo.
Expression of brain‐derived neurotrophic factor (BDNF) mRNA in rat retrosplenial cortex following administration of phencyclidine
Authors: HASHIMOTO KENJI , MINABE YOSHIO , IYO MASAOMI
Published: July 2002

The non‐competitive NMDA receptor antagonists such as phencyclidine (PCP) cause neurotoxicity in the retrosplenial cortex of rat brain. However, the precise mechanism(s) underlying the neurotoxicity of NMDA receptor antagonists is currently unclear. Using an in situ hybridization technique, we studied the effects of PCP on expression of brain‐derived neurotrophic factor (BDNF) mRNA in the rat brain. No expression of BDNA mRNA was observed in the retrosplenial cortex of rats treated with vehicle, although a high basal level of BDNF mRNA was detected in the hippocampus of control rats. Administration of PCP (12.5, 25 or 50 mg/kg, i.p., 6 hours) caused marked induction of BDNF mRNA in the retrosplenial cortex, in a dosedependent manner. These results suggest that the expression of BDNF mRNA may occur as a trophic response to the neurotoxicity of NMDA receptor antagonists such as PCP.
Periosteal Ewing′s Sarcoma: Report of Two New Cases and Review of the Literature
Authors: Kollender Yehuda , Shabat Shay , Nirkin Alexander , Issakov Josephine , Flusser Gideon , Merimsky Ofer , Meller Isaac
Published: July 2002

Background. The origin of Ewing′s sarcoma in a periosteal location is
rare and not clearly documented. Other malignant bone tumors appear to have a
somewhat better prognosis when confined between periosteum and bone. Is it the
same for periosteal Ewing′s sarcoma?

Methods. We describe two new cases and comprehensively
review the literature consisting of 18 documented cases since the condition was first
described in 1986 (S.M. Bator.Cancer 58:1781– 4).

Results. Periosteal Ewing′s sarcoma differs from the other forms
of Ewing′s sarcoma in terms of sex predominance, location of tumor, surgical stage at
presentation and typical imaging studies. Eighteen out of the 20 patients were
reported to be alive with no evidence of disease.

Conclusions. It seems that the prognosis of this rare variant of
Ewing′s sarcoma family of tumors might be better but the small number of cases
precludes such a firm conclusion.
Speech and language therapy provision for bilingual children: aspects of the current service
Authors: Winter Kirsten
Published: July 2002

The findings are presented here of a questionnaire survey which asked speech and language therapy managers in England about the caseloads of the therapists in their trust as regards bilingual children. The survey was the first part of a larger research project aiming to explore why there appears to be overrepresentation of bilingual children in speech and language therapy in some areas and under‐representation in others (compared to the proportion of bilingual children in the population). This main research question is discussed in relation to the literature and to the survey aims. The results describe aspects of the context in which the speech and language therapy service for bilingual children is operating: the lack of information about bilingual children is highlighted. The staffing and training implications of bilingual clients' distribution over therapists' caseloads in areas with different levels of minority ethnic population are also considered. The finding that the majority of speech and language therapists in England who work with children (59%) see at least one bilingual child speaks of the extent to which issues relating to bilingualism affects therapists.
Visualization of the Peroxisomal Compartment in Living Mammalian Cells: Dynamic Behavior and Association with Microtubules
Authors: Wiemer Erik A.C. , Wenzel Thibaut , Deerinck Thomas J. , Ellisman Mark H. , Subramani Suresh
Published: July 2002

Peroxisomes in living CV1 cells were visualized by targeting the green fluorescent protein (GFP) to this subcellular compartment through the addition of a COOH-terminal peroxisomal targeting signal 1 (GFP–PTS1). The organelle dynamics were examined and analyzed using time-lapse confocal laser scanning microscopy. Two types of movement could be distinguished: a relatively slow, random, vibration-like movement displayed by the majority (∼95%) of the peroxisomes, and a saltatory, fast directional movement displayed by a small subset (∼5%) of the peroxisomes. In the latter instance, peak velocities up to 0.75 μm/s and sustained directional velocities up to 0.45 μm/s over 11.5 μm were recorded. Only the directional type of motion appeared to be energy dependent, whereas the vibrational movement continued even after the cells were depleted of energy. Treatment of cells, transiently expressing GFP–PTS1, with microtubule-destabilizing agents such as nocodazole, vinblastine, and demecolcine clearly altered peroxisome morphology and subcellular distribution and blocked the directional movement. In contrast, the microtubule-stabilizing compound paclitaxel, or the microfilament-destabilizing drugs cytochalasin B or D, did not exert these effects. High resolution confocal analysis of cells expressing GFP–PTS1 and stained with anti-tubulin antibodies revealed that many peroxisomes were associated with microtubules. The GFP–PTS1–labeled peroxisomes were found to distribute themselves in a stochastic, rather than ordered, manner to daughter cells at the time of mitosis.
Detection of Dimers of Dimers of Human Leukocyte Antigen (HLA)–DR on the Surface of Living Cells by Single-Particle Fluorescence Imaging
Authors: Cherry Richard J. , Wilson Keith M. , Triantafilou Kathy , O'Toole Peter , Morrison Ian E.G. , Smith Patricia R. , Fernández Nelson
Published: July 2002

The technique of single-particle fluorescence imaging was used to investigate the oligomeric state of MHC class II molecules on the surface of living cells. Cells transfected with human leukocyte antigen (HLA)–DR A and B genes were labeled at saturation with a univalent probe consisting of Fab coupled to R-phycoerythrin. Analysis of the intensities of fluorescent spots on the cell surface revealed the presence of single and double particles consistent with the simultaneous presence of HLA-DR heterodimers and dimers of dimers. The proportion of double particles was lower at 37°C than at 22°C, suggesting that the heterodimers and dimers of dimers exist in a temperature-dependent equilibrium. These results are discussed in the context of a possible role for HLA-DR dimers of dimers in T cell receptor–MHC interactions. The technique is validated by demonstrating that fluorescence imaging can distinguish between dimers and tetramers of human erythrocyte spectrin deposited from solution onto a solid substrate. The methodology will have broad applicability to investigation of the oligomeric state of immunological and other membrane-bound receptors in living cells.
Why Are Two Different Cross-linkers Necessary for Actin Bundle Formation In Vivo and What Does Each Cross-link Contribute?
Authors: Tilney Lewis G. , Connelly Patricia S. , Vranich Kelly A. , Shaw Michael K. , Guild Gregory M.
Published: July 2002

In developing Drosophila bristles two species of cross-linker, the forked proteins and fascin, connect adjacent actin filaments into bundles. Bundles form in three phases: (a) tiny bundles appear; (b) these bundles aggregate into larger bundles; and (c) the filaments become maximally cross-linked by fascin. In mutants that completely lack forked, aggregation of the bundles does not occur so that the mature bundles consist of <50 filaments versus ∼700 for wild type. If the forked concentration is genetically reduced to half the wild type, aggregation of the tiny bundles occurs but the filaments are poorly ordered albeit with small patches of fascin cross-linked filaments. In mutants containing an excess of forked, all the bundles tend to aggregate and the filaments are maximally crossbridged by fascin. Alternatively, if fascin is absent, phases 1 and 2 occur normally but the resultant bundles are twisted and the filaments within them are poorly ordered. By extracting fully elongated bristles with potassium iodide which removes fascin but leaves forked, the bundles change from being straight to twisted and the filaments within them become poorly ordered. From these observations we conclude that (a) forked is used early in development to aggregate the tiny bundles into larger bundles; and (b) forked facilitates fascin entry into the bundles to maximally cross-link the actin filaments into straight, compact, rigid bundles. Thus, forked aligns the filaments and then directs fascin binding so that inappropriate cross-linking does not occur.
Visualization of Phosphoinositides That Bind Pleckstrin Homology Domains: Calcium- and Agonist-induced Dynamic Changes and Relationship to Myo-[3H]inositol-labeled Phosphoinositide Pools
Authors: Várnai Péter , Balla Tamás
Published: July 2002

Phosphatidylinositol 4,5-bisphosphate (PtdIns[4,5]P2) pools that bind pleckstrin homology (PH) domains were visualized by cellular expression of a phospholipase C (PLC)δ PH domain–green fluorescent protein fusion construct and analysis of confocal images in living cells. Plasma membrane localization of the fluorescent probe required the presence of three basic residues within the PLCδ PH domain known to form critical contacts with PtdIns(4,5)P2. Activation of endogenous PLCs by ionophores or by receptor stimulation produced rapid redistribution of the fluorescent signal from the membrane to cytosol, which was reversed after Ca2+ chelation. In both ionomycin- and agonist-stimulated cells, fluorescent probe distribution closely correlated with changes in absolute mass of PtdIns(4,5)P2. Inhibition of PtdIns(4,5)P2 synthesis by quercetin or phenylarsine oxide prevented the relocalization of the fluorescent probe to the membranes after Ca2+ chelation in ionomycin-treated cells or during agonist stimulation. In contrast, the synthesis of the PtdIns(4,5)P2 imaged by the PH domain was not sensitive to concentrations of wortmannin that had been found inhibitory of the synthesis of myo-[3H]inositol– labeled PtdIns(4,5)P2. Identification and dynamic imaging of phosphoinositides that interact with PH domains will further our understanding of the regulation of such proteins by inositol phospholipids.
The Lis1-Related Nudf Protein of Aspergillus nidulans Interacts with the Coiled-Coil Domain of the Nude/Ro11 Protein
Authors: Efimov Vladimir P. , Morris N. Ronald
Published: July 2002

The nudF gene of the filamentous fungus Aspergillus nidulans acts in the cytoplasmic dynein/dynactin pathway and is required for distribution of nuclei. NUDF protein, the product of the nudF gene, displays 42% sequence identity with the human protein LIS1 required for neuronal migration. Haploinsufficiency of the LIS1 gene causes a malformation of the human brain known as lissencephaly. We screened for multicopy suppressors of a mutation in the nudF gene. The product of the nudE gene isolated in the screen, NUDE, is a homologue of the nuclear distribution protein RO11 of Neurospora crassa. The highly conserved NH2-terminal coiled-coil domain of the NUDE protein suffices for protein function when overexpressed. A similar coiled-coil domain is present in several putative human proteins and in the mitotic phosphoprotein 43 (MP43) of X. laevis. NUDF protein interacts with the Aspergillus NUDE coiled-coil in a yeast two-hybrid system, while human LIS1 interacts with the human homologue of the NUDE/RO11 coiled-coil and also the Xenopus MP43 coiled-coil. In addition, NUDF coprecipitates with an epitope-tagged NUDE. The fact that NUDF and LIS1 interact with the same protein domain strengthens the notion that these two proteins are functionally related.
Drosophila Atypical Protein Kinase C Associates with Bazooka and Controls Polarity of Epithelia and Neuroblasts
Authors: Wodarz Andreas , Ramrath Andreas , Grimm Alexandra , Knust Elisabeth
Published: July 2002

The establishment and maintenance of polarity is of fundamental importance for the function of epithelial and neuronal cells. In Drosophila, the multi-PDZ domain protein Bazooka (Baz) is required for establishment of apico-basal polarity in epithelia and in neuroblasts, the stem cells of the central nervous system. In the latter, Baz anchors Inscuteable in the apical cytocortex, which is essential for asymmetric localization of cell fate determinants and for proper orientation of the mitotic spindle. Here we show that Baz directly binds to the Drosophila atypical isoform of protein kinase C and that both proteins are mutually dependent on each other for correct apical localization. Loss-of-function mutants of the Drosophila atypical isoform of PKC show loss of apico-basal polarity, multilayering of epithelia, mislocalization of Inscuteable and abnormal spindle orientation in neuroblasts. Together, these data provide strong evidence for the existence of an evolutionary conserved mechanism that controls apico-basal polarity in epithelia and neuronal stem cells. This study is the first functional analysis of an atypical protein kinase C isoform using a loss-of-function allele in a genetically tractable organism.
Identification of Ubiquilin, a Novel Presenilin Interactor That Increases Presenilin Protein Accumulation
Authors: Mah Alex L. , Perry George , Smith Mark A. , Monteiro Mervyn J.
Published: July 2002

Mutations in the highly homologous presenilin genes encoding presenilin-1 and presenilin-2 (PS1 and PS2) are linked to early-onset Alzheimer's disease (AD). However, apart from a role in early development, neither the normal function of the presenilins nor the mechanisms by which mutant proteins cause AD are well understood. We describe here the properties of a novel human interactor of the presenilins named ubiquilin. Yeast two-hybrid (Y2H) interaction, glutathione S-transferase pull-down experiments, and colocalization of the proteins expressed in vivo, together with coimmunoprecipitation and cell fractionation studies, provide compelling evidence that ubiquilin interacts with both PS1 and PS2. Ubiquilin is noteworthy since it contains multiple ubiquitin-related domains typically thought to be involved in targeting proteins for degradation. However, we show that ubiquilin promotes presenilin protein accumulation. Pulse-labeling experiments indicate that ubiquilin facilitates increased presenilin synthesis without substantially changing presenilin protein half-life. Immunohistochemistry of human brain tissue with ubiquilin-specific antibodies revealed prominent staining of neurons. Moreover, the anti-ubiquilin antibodies robustly stained neurofibrillary tangles and Lewy bodies in AD and Parkinson's disease affected brains, respectively. Our results indicate that ubiquilin may be an important modulator of presenilin protein accumulation and that ubiquilin protein is associated with neuropathological neurofibrillary tangles and Lewy body inclusions in diseased brain.
Adenovirus E4orf4 protein induces PP2A-dependent growth arrest in Saccharomyces cerevisiae and interacts with the anaphase-promoting complex/cyclosome
Authors: Kornitzer Daniel , Sharf Rakefet , Kleinberger Tamar
Published: July 2002

Adenovirus early region 4 open reading frame 4 (E4orf4) protein has been reported to induce p53-independent, protein phosphatase 2A (PP2A)–dependent apoptosis in transformed mammalian cells. In this report, we show that E4orf4 induces an irreversible growth arrest in Saccharomyces cerevisiae at the G2/M phase of the cell cycle. Growth inhibition requires the presence of yeast PP2A-Cdc55, and is accompanied by accumulation of reactive oxygen species. E4orf4 expression is synthetically lethal with mutants defective in mitosis, including Cdc28/Cdk1 and anaphase-promoting complex/cyclosome (APC/C) mutants. Although APC/C activity is inhibited in the presence of E4orf4, Cdc28/Cdk1 is activated and partially counteracts the E4orf4-induced cell cycle arrest. The E4orf4–PP2A complex physically interacts with the APC/C, suggesting that E4orf4 functions by directly targeting PP2A to the APC/C, thereby leading to its inactivation. Finally, we show that E4orf4 can induce G2/M arrest in mammalian cells before apoptosis, indicating that E4orf4-induced events in yeast and mammalian cells are highly conserved.
Selective Suppression of Interleukin-12 Induction after Macrophage Receptor Ligation
Authors: Sutterwala Fayyaz S. , Noel Gary J. , Clynes Raphael , Mosser David M.
Published: July 2002

Interleukin (IL)-12 is a monocyte- and macrophage-derived cytokine that plays a crucial role in both the innate and the acquired immune response. In this study, we examined the effects that ligating specific macrophage receptors had on the induction of IL-12 by lipopolysaccharide (LPS). We report that ligation of the macrophage Fcγ, complement, or scavenger receptors inhibited the induction of IL-12 by LPS. Both mRNA synthesis and protein secretion were diminished to near-undetectable levels following receptor ligation. Suppression was specific to IL-12 since IL-10 and tumor necrosis factor-α (TNF-α) production were not inhibited by ligating macrophage receptors. The results of several different experimental approaches suggest that IL-12 downregulation was due to extracellular calcium influxes that resulted from receptor ligation. First, preventing extracellular calcium influxes, by performing the assays in EGTA, abrogated FcγR-mediated IL-12(p40) mRNA suppression. Second, exposure of macrophages to the calcium ionophores, ionomycin or A23187, mimicked receptor ligation and inhibited IL-12(p40) mRNA induction by LPS. Finally, bone marrow–derived macrophages from FcR γ chain–deficient mice, which fail to flux calcium after receptor ligation, failed to inhibit IL-12(p40) mRNA induction. These results indicate that the calcium influxes that occur as a result of receptor ligation are responsible for inhibiting the induction of IL-12 by LPS. Hence, the ligation of phagocytic receptors on macrophages can lead to a dramatic decrease in IL-12 induction. This downregulation may be a way of limiting proinflammatory responses of macrophages to extracellular pathogens, or suppressing the development of cell-mediated immunity to intracellular pathogens.
Altered Immune Responses in Interleukin 10 Transgenic Mice
Authors: Hagenbaugh Amy , Sharma Sherven , Dubinett Steven M. , Wei Sherry H.-Y. , Aranda Richard , Cheroutre Hilde , Fowell Deborah J. , Binder Scott , Tsao Betty , Locksley Richard M. , Moore Kevin W. , Kronenberg Mitchell
Published: July 2002

Interleukin (IL)-10 is a pleiotropic cytokine which inhibits a broad array of immune parameters including T helper cell type 1 (Th1) cytokine production, antigen presentation, and antigenspecific T cell proliferation. To understand the consequences of altered expression of IL-10 in immune models of autoimmune disease, the response to infectious agents, and the response to tumors, we developed transgenic mice expressing IL-10 under the control of the IL-2 promoter. Upon in vitro stimulation, spleen cells from unimmunized transgenic mice secrete higher levels of IL-10 and lower amounts of IFN-γ than do controls, although no gross abnormalities were detected in lymphocyte populations or serum Ig levels. Transfer of normally pathogenic CD4+ CD45RBhigh splenic T cells from IL-10 transgenic mice did not cause colitis in recipient severe combined immunodeficiency mice. Furthermore, co-transfer of these transgenic cells with CD4+ CD45RBhigh T cells from control mice prevented disease. Transgenic mice retained their resistance to Leishmania major infection, indicating that their cell-mediated immune responses were not globally suppressed. Lastly, in comparison to controls, IL-10 transgenic mice were unable to limit the growth of immunogenic tumors. Administration of blocking IL-10 mAbs restored in vivo antitumor responses in the transgenic mice. These results demonstrate that a single alteration in the T cell cytokine profile can lead to dramatic changes in immune responses in a manner that is stimulus dependent. These mice will be useful in defining differences in inflammatory conditions and cellular immunity mediated by IL-10.
Eotaxin-2, a Novel CC Chemokine that Is Selective for the Chemokine Receptor CCR3, and Acts Like Eotaxin on Human Eosinophil and Basophil Leukocytes
Authors: Forssmann Ulf , Uguccioni Mariagrazia , Loetscher Pius , Dahinden Clemens A. , Langen Hanno , Thelen Marcus , Baggiolini Marco
Published: July 2002

A novel human CC chemokine consisting of 78 amino acids and having a molecular mass of 8,778.3 daltons (VVIPSPCCMF FVSKRIPENR VVSYQLSSRS TCLKAGVIFT TKKGQQ SCGD PKQEWVQRYM KNLDAKQKKA SPRARAVA) was isolated together with three minor COOH-terminally truncated variants with 73, 75, and 76 residues. The new chemokine was termed eotaxin-2 because it is functionally very similar to eotaxin. In terms of structure, however, eotaxin and eotaxin-2 are rather distant, they share only 39% identical amino acids and differ almost completely in the NH2-terminal region. Eotaxin-2 induced chemotaxis of eosinophils as well as basophils, with a typically bimodal concentration dependence, and the release of histamine and leukotriene C4 from basophils that had been primed with IL-3. In all assays, eotaxin-2 had the same efficacy as eotaxin, but was somewhat less potent. The migration and the release responses were abrogated in the presence of a monoclonal antibody that selectively blocks the eotaxin receptor, CCR3, indicating that eotaxin-2, like eotaxin, acts exclusively via CCR3. Receptor usage was also studied in desensitization experiments by measuring [Ca2+]i changes in eosinophils. Complete cross-desensitization was observed between eotaxin-2, eotaxin and MCP-4 confirming activation via CCR3. No Ca2+ mobilization was obtained in neutrophils, monocytes and lymphocytes, in agreement with the lack of chemotactic responsiveness. Intradermal injection of eotaxin-2 in a rhesus monkey (100 or 1,000 pmol per site) induced a marked local infiltration of eosinophils, which was most pronounced in the vicinity of postcapillary venules and was comparable to the effect of eotaxin.