The gendered nature of South African teachers' discourse on sex education.

In South Africa, high pregnancy and infection rates show that many teenagers are having sex, and that they are not adequately protecting themselves against undesired pregnancies and disease. Sex education is usually taught as part of the subject area Life Orientation. In a qualitative study of 25 Life Orientation teachers in the South African Free State Province, we used semi-structured interviews to explore the ways in which these teachers understand gender to be a factor in learners' experiences of sexuality. Our analysis draws upon the conceptual framework of heteronormativity, a key aspect of which is that girls and boys are socialized into different gender roles in ways that propagate the patriarchy, and these are largely viewed as part of the natural order of things. Our data revealed a tendency for teachers to cast boys as largely predatory and girls as victims of sexual predation, either by their peers or by older boys or men. Although these assumptions reflect some of the everyday experiences in South Africa and many other countries, these expectations may be transmitted and reinforced unconsciously in well-meaning educational interventions meant to protect girls.

Glatiramer acetate (Copaxone).

Glatiramer acetate (Copaxone) is a novel preparation of synthetic peptides composed of four amino acids. Laboratory studies have shown that it prevents, or modifies, experimental allergic encephalomyelitis, the animal model for multiple sclerosis (MS), in several mammalian species. Its mode of action has not been fully elucidated but it is known to induce suppresser T-cells, known to be deficient in MS, and competitively inhibits the effect of CNS myelin antigens, thought to be important in the pathogenesis of MS, through MHC blockade. Controlled clinical trials have shown it to improve the natural history of MS by reducing both the relapse rate and the resultant disability. GA shows similar efficacy to interferon-beta (IFN-beta) but with fewer systemic side-effects and appears to be better tolerated by patients. It has thus justified its place in the new era of disease-modifying treatments for MS. While the evidence suggests GA should be considered as first-line therapy in selected patients, its differing mechanism of action also gives patients and doctors the option of an alternative agent when the efficacy of IFN-beta is waning or side-effects predominate.

CTLA4 in multiple sclerosis. Lack of genetic association in a European Caucasian population but evidence of interaction with HLA-DR2 among Shanghai Chinese.

In the present study we searched for an association between multiple sclerosis (MS) and the gene encoding the cytotoxic T lymphocyte antigen 4 (CTLA4). Our experimental approach involved amplification of DNA fragments of the promoter and exon 1 of this gene containing single nucleotide polymorphisms followed by treatment of the amplified fragments with restriction enzymes for allele determination. Included in the study were 84 MS patients and 125 healthy control subjects from a population of white Caucasians. We also examined 42 MS patients and 86 healthy control subjects of Shanghai Chinese origin. Significant differences in the distribution of genotypes or haplotypes of the CTLA4 gene were not observed between MS patients and control subjects in either of the two populations (P>0.05). Moreover, we were not able to confirm a previous finding of an association between relapsing-remitting MS and the heterozygous genotype A/G of CTLA4 exon 1. There was no evidence to suggest that interaction between HLA-DR2 and CTLA4 is involved in the development of MS among European Caucasians (P>0.05). Opposed to this, analysis of the Shanghai Chinese suggested presence of such interaction (P=0.02). Our results do not support the assumption that CTLA4 influences susceptibility to MS in European Caucasians. On the other hand, they raise the possibility that the development of MS in other ethnic groups involves interaction between CTLA4 and DR2.

Papillomavirus E2 induces senescence in HPV-positive cells via pRB- and p21(CIP)-dependent pathways.

A hallmark of human papillomavirus (HPV) associated carcinogenesis is the integration of the viral DNA into the cellular genome, usually accompanied by the loss of expression of the viral E2 gene. E2 binds to and represses the viral promoter directing expression of the E6 and E7 oncogenes. The re-introduction and expression of exogenous E2 in HPV-positive cancer cells results in cellular growth arrest, while growth in the context of exogenous E2 can be restored through the expression of exogenous E6 and E7. Here we examine the individual contributions of the viral E6 and E7 genes to this phenotype. E6 alone displays moderate activity, whereas both E7 and adenovirus E1A display high activity in reversing E2-mediated cellular growth suppression. Using defined mutants of E7 and E1A, we show that an intact retinoblastoma interaction domain is required for this function. In addition, we show that the E2-mediated growth arrest of HPV-positive cells results in cellular senescence, and implicate the cyclin/cdk inhibitor p21(CIP) as a downstream E2 effector in this phenotype.

The effect of oral and intravenous methylprednisolone treatment on subsequent relapse rate in multiple sclerosis.

We investigated the effect of oral and intravenous methylprednisolone treatment on subsequent relapse rate in patients with multiple sclerosis. Following a double blind trial designed to compare the effect of oral and intravenous methylprednisolone treatment on promoting recovery from acute relapses of multiple sclerosis, 80 patients were followed for two years with six-monthly assessments during which all subsequent relapses were recorded. The annual relapse rate was slightly higher in the oral compared with the intravenous methylprednisolone-treated patients (1.06 vs. 0.78), but the adjusted difference between the two groups was not statistically significant (0.18; 95% CI -0.19 to 0.55, P=0.3). The time to onset and the severity of the first relapse after treatment, the number of relapse free patients at the end of the follow-up period, and the severity of the relapses during the follow-up period were similar in the two groups. This trial did not show a statistically significant difference in relapse rate during the first two years following oral compared with intravenous methylprednisolone treatment.

Repression of the integrated papillomavirus E6/E7 promoter is required for growth suppression of cervical cancer cells.

The human papillomavirus (HPV) E2 protein is an important regulator of viral E6 and E7 gene expression. E2 can repress the viral promoter for E6 and E7 expression as well as block progression of the cell cycle in cancer cells harboring the DNA of "high-risk" HPV types. Although the phenomenon of E2-mediated growth arrest of HeLa cells and other HPV-positive cancer cells has been well documented, the specific mechanism by which E2 affects cellular proliferation has not yet been elucidated. Here, we show that bovine papillomavirus (BPV) E2-induced growth arrest of HeLa cells requires the repression of the E6 and E7 promoter. This repression is specific for E2TA and not E2TR, a BPV E2 variant that lacks the N-terminal transactivation domain. We demonstrate that expression of HPV16 E6 and E7 from a heterologous promoter that is not regulated by E2 rescues HeLa cells from E2-mediated growth arrest. Our data indicate that the pathway of E2-mediated growth arrest of HeLa cells requires repression of E6 and E7 expression through an activity specified by the transactivation domain of E2TA.

Association between the endogenous retrovirus HRES-1 and multiple sclerosis in the United Kingdom--evidence of genetically different disease subsets?

In the present study we determined the frequencies of four haplotypes of the human T-cell lymphotropic virus-related endogenous sequence, HRES-1, in 110 multiple sclerosis (MS) patients and 100 healthy control subjects from the United Kingdom. We found evidence of an association between this endogenous retrovirus and MS (p < 0.01), in particular reflecting an increased frequency of HRES-1 haplotype 1 in the group of patients. There was no significant difference in the distribution of HRES-1 haplotypes between relapsing-remitting MS and the primary progressive form of the disease. The odds ratio for HRES-1 haplotype 1 and MS did not differ significantly between individuals positive for HLA-DR2 and DR2-negative individuals. Comparison of the observations from the present study with previous results implicated HRES-1 as a marker of genetic heterogeneity in MS.

Receptor-specific induction of NF-kappaB components in primary B cells.

The NF-kappaB transcription factor complex plays a key role in the expression of genes involved in immune responses. Nuclear NF-kappaB is induced in B lymphocytes by engagement of either the antigen receptor (sIg) or the CD40 receptor for a T cell activation antigen, although different intracellular pathways appear to be involved. In the present study the protein composition of NF-kappaB complexes triggered by sIg and CD40 was probed by electrophoretic mobility shift, supershift, shift-Western, and Western blot analyses. At the time of peak NF-kappaB induction (2 h), the NF-kappaB components detected in the complexes induced through sIg and through CD40 were the same. However, with continued stimulation RelB completely disappeared from anti-Ig-stimulated kappaB binding material, but remained a component of CD40L-induced NF-kappaB. The loss of DNA-binding RelB from anti-Ig-induced NF-kappaB did not result from depletion of RelB from B cell nuclei, suggesting specific regulation of RelB function which is not directly attributed to IkappaB function. These results indicate that NF-kappaB complexes may undergo protein-specific alterations in a time- and receptor-dependent fashion that may be associated with differences in the outcomes of B cell stimulation through sIg and CD40.

The interaction of arginine 106 of human prostaglandin G/H synthase-2 with inhibitors is not a universal component of inhibition mediated by nonsteroidal anti-inflammatory drugs.

The three-dimensional cocrystal structures of ovine prostaglandin G/H synthase-1 (PGHS-1) with S-flurbiprofen and murine PGHS-2 with S-flurbiprofen and indomethacin reveal that the carboxylate acid groups of these nonsteroidal anti-inflammatory drugs (NSAIDs) form a salt bridge with the guanidinium group of Arg120 in PGHS-1 and Arg106 in PGHS-2. Mutagenesis studies confirmed that the Arg120 residue of PGHS-1 is critical for binding of substrate and inhibitors through ionic interactions of its guanidinium group with the carboxylate moieties of arachidonic acid and certain NSAIDs. We report here that the analogous R106E substitution in human PGHS-2 results in a catalytically active enzyme with a 30-fold higher Km value for arachidonic acid. Comparison of the inhibition of hPGHS-2(R106E) with wild-type hPGHS-2 by 11 structurally diverse selective and nonselective PGHS inhibitors revealed a 0-1000-fold decrease in inhibitory potency on the mutant enzyme. The loss of inhibitory potency of NSAIDs on hPGHS-2(R106E) could not be correlated with the presence or absence of a carboxylate functional group in the inhibitor, as was demonstrated previously for the PGHS-1(R120E) mutant, or with the selective or nonselective nature of the PGHS inhibitor. The decreases in the inhibitory potencies on hPGHS-2(R106E) by the carboxylate-containing NSAIDs flurbiprofen, indomethacin, meclofenamic acid, and diclofenac on hPGHS-2(R106E) were 965-, 48-, 5.5-, and 4.5-fold, respectively. The nonuniversal requirement for interaction of the carboxylate group of certain NSAIDs with the Arg106 residue in hPGHS-2 is supported by the observation that the methyl ester derivative of indomethacin was a more potent inhibitor than indomethacin on both hPGHS-2 and hPGHS-2(R106E). The greatest loss of potency for inhibition of hPGHS-2(R106E) was observed with the hPGHS-2-selective sulfonamide-containing inhibitors NS-398 and flosulide. The PGHS-2-selective inhibitor DuP697 and a desbromo-sulfonamide analogue of DuP697 displayed equivalent potency on hPGHS-2(R106E) and hPGHS-2. The change in inhibitory potency of NS-398 on hPGHS-2(R106E) was due to a difference in the kinetics of inhibition, with NS-398 displaying time-dependent inhibition of hPGHS-2 but time-independent inhibition of PGHS-2(R106E). The time-dependent inhibition of hPGHS-2 by DuP697 was not affected by the presence of the R106E mutation. We conclude that the Arg106 residue of hPGHS-2 is involved in binding arachidonic acid and certain NSAIDs, but interactions with Arg106 are not a universal requirement for inhibition by either carboxylate-containing NSAIDs or PGHS-2-selective inhibitors.

Randomised trial of oral and intravenous methylprednisolone in acute relapses of multiple sclerosis.

BACKGROUND: An intravenous rather than oral course of methylprednisolone is often prescribed for treating acute relapses in multiple sclerosis (MS) despite the lack of evidence to support this route of administration. Our double-blind placebo-controlled randomised trial was designed to compare the efficacy of commonly used intravenous and oral steroid regimens in promoting recovery from acute relapses in MS. METHODS: 42 patients with clinically definite relapse in MS received oral, and 38 intravenous, methylprednisolone. Clinical measurements at entry and at 1 week, 4 weeks, 12 weeks, and 24 weeks included Kurtzke's expanded disability status scale (EDSS), Hauser's Ambulatory Index, and an arm-function index. The primary outcome criterion was a difference between the two treatment groups of one or more EDSS grades at 4 weeks. FINDINGS: There were no significant differences between the two groups at any stage of the study in any measurement taken: the mean difference in EDSS at 4 weeks (adjusted for baseline level) was 0.07 grades more in those taking oral steroids (95% CI -0.46 to 0.60). The most optimistic outcome for intravenous therapy is an average benefit of less than half a grade improvement on EDSS over oral treatment. INTERPRETATION: Since our study did not show any clear advantage of the intravenous regime we conclude that it is preferable to prescribe oral rather than intravenous steroids for acute relapses in MS for reasons of patient convenience, safety, and cost.

Determination of tumor cell procoagulant activity by Sonoclot analysis in whole blood.

Coagulation activation in cancer may be due to expression of procoagulant activity (PCA) by tumor cells. Some PCA activate coagulation, while others influence platelet aggregation. Thus, clotting time assessments of PCA may not reflect the potential for hypercoagulability. We therefore studied the effect of various malignant and non-malignant cells on whole blood coagulation using the Sonoclot Analyzer. Five human (HT29 colon, J82 bladder, MCF-7 breast, BT-474 breast and A375 malignant melanoma) and three rodent (MC28, WEHI-164 and Neuro2a) cell lines were used. Rat thymocytes and peritoneal macrophages and human endotoxin-stimulated mononuclear cells and umbilical vein endothelial cells (HUVEC) were used as non-malignant controls. All tumor cells markedly shortened the recalcification time of citrated blood and the most potent (HT29 and J82) also increased clot rate (P < 0.01). The breast cancer cells MCF-7 and BT-474 expressed only weak PCA and did not affect clotting rate. None of the non-malignant cells significantly affected clotting time or rate in whole blood. J82 and HT29 cells grown in serum-rich media shortened the recalcification time of both normal and FVII-deficient plasmas. However, cells grown in serum-free conditions had significantly less PCA in FVII-deficient plasma. We conclude that the Sonoclot Analyzer is useful for determining cellular PCA; significant PCA is a feature of malignant cells and cells grown in medium containing serum supplements cannot be reliably evaluated for PCA.

Mechanism of selective inhibition of human prostaglandin G/H synthase-1 and -2 in intact cells.

Selective inhibitors of prostaglandin synthase-2 (PGHS-2) possess potent anti-inflammatory, antipyretic, and analgesic properties but demonstrate reduced side-effects (e.g. gastrotoxicity) when compared with nonselective inhibitors of PGHS-1 and -2. We investigated the mechanism of the differential inhibition of human PGHS-1 (hPGHS-1) and -2 (hPGHS-2) in intact cells by nonsteroidal anti-inflammatory drugs (NSAIDs) and examined factors that contribute to the increased potency of PGHS inhibitors observed in intact cells versus cell-free systems. In intact Chinese hamster ovary (CHO) cell lines stably expressing the hPGHS isozymes, both PGHS isoforms exhibited the same affinity for arachidonic acid. Exogenous and endogenous arachidonic acid were used as substrates by both CHO [hPGHS-1] and CHO [hPGHS-2] cell lines. However, differences were observed in the ability of the hPGHS isoforms to utilize endogenous arachidonic acid released intracellularly following calcium ionophore stimulation or released by human cytosolic phospholipase A2 transiently expressed in the cells. Cell-based screening of PGHS inhibitors demonstrated that the selectivities and potencies of PGHS inhibitors determined using intact cells are affected by substrate concentration and differ from that determined in cell-free microsomal or purified enzyme preparations of PGHS isozymes. The mechanism of inhibition of PGHS isozymes by NSAIDs in intact cells involved difference in their time-dependent inhibition. Indomethacin displayed time-dependent inhibition of cellular hPGHS-1 and -2. In contrast, the selective PGHS-2 inhibitor NS-398 exhibited time-independent inhibition of hPGHS-1 but time-dependent inhibition of hPGHS-2 in intact cells. Reversible inhibition of cellular CHO [hPGHS-1] and CHO [hPGHS-2] was observed with the nonselective NSAIDs ibuprofen and indomethacin, whereas inhibition by the selective PGHS-2 inhibitor DuP-697 was reversible against hPGHS-1 but irreversible against hPGHS-2.

A study of oligoclonal band negative multiple sclerosis.

OBJECTIVES: To determine whether oligoclonal band (OCB) negative multiple sclerosis is a reliable diagnosis and, if so, whether it has a distinctive prognosis. METHODS: Retrospective and matched prospective comparison of the clinical and laboratory features of patients with clinical definite multiple sclerosis with and without intrathecal synthesis of oligoclonal IgG. RESULTS: Thirty four patients were identified with apparent OCB negative clinically definite multiple sclerosis. The results of oligoclonal banding proved to have been equivocal in 14 of 34; the clinical diagnosis of multiple sclerosis was questionable in 8 of 34. The remaining 12 patients with "true" OCB negative multiple sclerosis were significantly less disabled than matched OCB positive controls. Re-examination of CSF-serum pairs from six OCB negative patients showed that three remained OCB negative while three showed evidence of intrathecal synthesis of OCBs. CONCLUSIONS: OCB negative clinically definite multiple sclerosis is rare and should be diagnosed with caution; in unequivocal cases it seems to have a relatively benign prognosis.

The importance of platelets in the expression of monocyte tissue factor antigen measured by a new whole blood flow cytometric assay.

Previous methods for the determination of monocyte tissue factor (TF) have been technically complex, difficult to standardize, prone to spuriously elevated results and difficult to implement in a clinical laboratory environment. We report the development of a two-color whole blood cytometric technique that overcomes many of these disadvantages. The assay uses small volumes of citrated blood (1.0 ml), can be performed in under one hour (if endotoxin stimulation is not performed), is reproducible (CV = 5%) and uses methodology commonly available in clinical laboratories. Baseline (mean +/- SD) expression of monocyte TF in normal subjects was very low (1.1 +/- 0.95%, Mean Fluorescence [Mean FL] 0.20 +/- 0.01) making relatively small increases easy to detect. Monocyte TF expression following endotoxin (LPS) stimulation for 1 h was 34.6 +/- 11.2% (Mean FL 0.32 +/- 0.04). LPS-stimulated activity varied between subjects (21-68%) but was remarkably consistent for individual subjects (CV = 5.4%). Stimulated monocyte TF expression was directly proportional to the platelet count and was reduced by platelet protective anticoagulants and by ingestion of aspirin. Non LPS-stimulated monocyte TF was markedly increased, in a dose-dependent manner, by adding collagen to whole blood. This was apparently associated with platelet-monocyte binding and could be abolished by anti-P-Selectin. We conclude that the whole blood flow cytometric assay of monocyte TF may be a valuable tool for clinical use and a useful model system for evaluating the humoral and cellular factors governing monocyte TF expression in a natural environment.

The clinical value of tissue factor assays.

Tissue factor (TF) is now considered to be the primary physiologic activator of the blood coagulation system. Coupled with recent advances in our understanding of the biochemistry of TF this has heightened interest in measuring aspects of TF activity in disease states. Expression of TF by blood monocytes in various diseases is an established trigger for intravascular coagulation and there is now a considerable body of experience with its measurement. This has considerable clinical potential although more widespread application awaits a consensus on the most appropriate methodologic approach to its measurement. TF can be detected in urine and may reflect the activation state of renal macrophages. Urinary TF is increased in cancer and could have diagnostic and prognostic value in a variety of malignant diseases. Finally, it is now possible to measure soluble TF in plasma. One such assay is commercially available and is technically simple to perform. The clinical value of such assays, however, must await better understanding of the source and function of soluble TF in plasma.

Genetic susceptibility to multiple sclerosis in the Shanghai Chinese is not linked to the myelin basic protein gene microsatellite.

Aim-To investigate the role of myelin basic protein (MBP) gene polymorphisms in determining susceptibility to multiple sclerosis in a Shanghai Chinese population.Methods-Forty seven unrelated patients with multiple sclerosis and 94 healthy control subjects were included in the study. Genomic DNA was extracted from peripheral blood lymphocytes and amplified using the polymerase chain reaction to characterise two adjacent tetranucleotide repeats ([ATGG](12) and [TGGA](9)) located 5' to exon 1 of the MBP gene.Results-Two polymorphic loci were identified: locus A, comprising both repeats, and locus B, comprising the [ATGG](12) repeat only. Nine allelic variants were identified at locus A and six at locus B, ranging from 212 to 244 and 122 to 146 base pairs, respectively. The 244 base pair allele at locus A has not been reported before. The allele frequencies observed in the controls differed from those seen in normal white populations.Conclusions-The present study demonstrates a race specific pattern of allelic distribution within the tetranucleotide repeat of the MBP gene. Further studies are needed to fully elucidate the role of the MBP gene in inherited susceptibility to multiple sclerosis.

Genetic susceptibility to multiple sclerosis in a Shanghai Chinese population. The role of the HLA class II genes.

The association of MS with the HLA class II loci DR and DQ was investigated in subjects of Shanghai Chinese and British Caucasian origin. Our aim was to determine whether common alleles predispose to the disease in both races. In the Caucasian population MS was significantly positively associated with the putative haplotype DRB1*1501, DQA1*0102, DQB1*0602. In contrast, HLA class II alleles were not found to predispose to the disease in the Shanghai Chinese, suggesting that this haplotype is unlikely to be a universal susceptibility determinant. The absence of a disease association with the HLA-DR and -DQ genes in the Chinese population has a number of possible explanations. Our study suggests that other genetic and/or environmental components may be more important in determining susceptibility to MS in this race.

An investigation of HLA-encoded genetic susceptibility to multiple sclerosis in subjects of Asian Indian and Afro-Caribbean ethnic origin.

The association of multiple sclerosis (MS) with the HLA class II loci DR and DQ was investigated in populations of Asian Indian and Afro-Caribbean ethnic origin, resident in the United Kingdom. The putative haplotype, DRB1*1501.DQA1*0102.DQB1*0602, was weakly positively associated with MS in both races. The overall contribution to disease susceptibility of this marker was small. Over 80% of the MS patients in both racial groups did not possess this haplotype. The data suggest that other genetic and/or environmental factors may be more important in predisposing to MS in these two races. Our study also raises the possibility that genetically distinct forms of the disease may be expressed in white Caucasian and non-Caucasian populations.