140 research outputs found

    Reliability of Human Fresh and Frozen Gingiva Explant Culture in Assessing Dental Materials Cytocompatibility

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    An explant culture of human gingival epithelium has been set up in order to provide a valuable test for evaluating the cytocompatibility of dental material s. In an attempt to supply a bank of gingiva ex plants, frozen and freshly excised specimens were cultured in parallel. Optical and scanning electron microscopy showed an early release of cuboidal cells forming a dense layer around the explants. Afterwards, cultures evolved differently . Spread cells grew and migrated more rap idly in fresh than in frozen explant cultures but their adhesion to substratum increased earlier in frozen ones. Epithelial phenotype of cells had been immunologically characterized by using a battery of monoclonal antibouies to cytokeratins (CKs). We found a time increasing expression of CKs 5, 6, 13 , 14115 , 16 and 17, whereas amounts of CKs 1, 2, 10 and 11 , specific for terminal differentiation , remained constant. The freezing procedure decreased the yield of CKs but did not modify the electrophoretic pattern . These results suggested that the differentiation of epithelial cells might proceed as in vivo. As an application, the cytocompatibility of precious (Au, Pd, Ag) and non-precious (Ni-Cr) alloys was assessed, the reference metals being Ti, which was chosen for its cytocompatibility and Cu, which was chosen for its cytotoxicity. Alloys differed by their ability to modulate cell proliferation and migration . Pd and Au exhibited a high migration potential, whereas Au-Pd and Ti allowed efficient cell proliferation but restricted migration . Reduced migration and proliferation attested the low cytocompatibility of Ag. The toxicity of Cu and Ni-Cr prevented cell migration. These result s showed the availability of this method for selecting biomaterials

    Protecting women from economic shocks to fight HIV in Africa (POWER): A study protocol for a randomised controlled trial

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    Introduction: HIV/AIDS is the leading cause of mortality among women aged 15 – 44 years. In Africa is there a huge gender disparity in the acquisition of HIV. African women aged 15 – 24 are twice as likely to be infected with HIV than their male counterparts. However, in Cameroon, adolescent girls are five times more likely to be infected with HIV than boys of the same age. Evidence suggests that risks taken during transactional and commercial sexual relationships account for this gender inequality. Recent studies demonstrate that a key driver of a woman’s engagement in risky sexual behaviours is to cope with negative income shocks (e.g., agricultural and climatic shocks, illness or death of family members), suggesting that economic shocks are a substantial piece of the HIV puzzle in Africa. This study is the first to estimate the effectiveness of a formal coping strategy, health insurance, to protect women from economic shocks to prevent STIs and HIV. Methods and Analysis: This study is a stratified cluster randomised controlled superiority trial with two parallel groups. 1,508 unmarried girls and young women engaging in transactional (753) and commercial (755) sex have been recruited onto a baseline study in Yaoundé, Cameroon. Participants are randomised into treatment (receiving health insurance for them and their economic dependents) and control (receiving nothing) groups using participatory randomisation. A target of a 1:1 allocation was stratified by those who engage in commercial or transactional sex. Midline data will be collected at 6 months and 12 months after randomisation. Ethics and Dissemination: This trial has been prospectively registered with the ISRCTN Registry. We obtained ethical approval from Cameroon National Ethics Committee and the UCL ethics committee before data collection. The findings of the trial will be published in peer-reviewed journals irrespective of the final results

    Defining fair and acceptable randomisation procedure in trials targeting vulnerable populations: Qualitative evidence from the POWER trial in Cameroon

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    Context and aims: Important ethical issues have been consistently highlighted in randomised controlled trials (RCTs). Although RCTs are widely used in non-clinical health research, there is little research on participants' perceptions of the fairness and transparency of RCTs. / Methods: Data were collected as part of the POWER trial, which aims to test the effectiveness of health shock prevention for HIV prevention among women who engage in commercial and transactional sex in urban Cameroon. This qualitative study was carried out in 2 phases. In the pre-randomisation phase (phase 1), we conducted 25 focus groups and 8 in-depth semi-structured interviews to determine the most acceptable randomisation strategy for our study participants. In the post-randomisation phase (phase 2), 41 in-depth semi-structured interviews were conducted with participants to assess their perceptions and satisfaction with their group allocation status (e.g. treatment or control). / Results: We found that participants understood the rationale for randomisation of the intervention and were satisfied with the randomisation strategy. This latter finding was mainly due to i) the involvement of participants in the chosen randomisation method and ii) the fact that the control group would receive the intervention at the end of the study. / Conclusion: It is important that researchers conduct similar studies before designing RCTs in vulnerable populations to minimise ethical issues related to the conduct of RCTs. Conducting pre- and post-randomisation qualitative research is an effective approach to improve RCT design and to assess and address potential problems with RCTs

    Protection from Experimental Cerebral Malaria with a Single Dose of Radiation-Attenuated, Blood-Stage Plasmodium berghei Parasites

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    BACKGROUND: Whole malaria parasites are highly effective in inducing immunity against malaria. Due to the limited success of subunit based vaccines in clinical studies, there has been a renewed interest in whole parasite-based malaria vaccines. Apart from attenuated sporozoites, there have also been efforts to use live asexual stage parasites as vaccine immunogens. METHODOLOGY AND RESULTS: We used radiation exposure to attenuate the highly virulent asexual blood stages of the murine malaria parasite P. berghei to a non-replicable, avirulent form. We tested the ability of the attenuated blood stage parasites to induce immunity to parasitemia and the symptoms of severe malaria disease. Depending on the mouse genetic background, a single high dose immunization without adjuvant protected mice from parasitemia and severe disease (CD1 mice) or from experimental cerebral malaria (ECM) (C57BL/6 mice). A low dose immunization did not protect against parasitemia or severe disease in either model after one or two immunizations. The protection from ECM was associated with a parasite specific antibody response and also with a lower level of splenic parasite-specific IFN-γ production, which is a mediator of ECM pathology in C57BL/6 mice. Surprisingly, there was no difference in the sequestration of CD8+ T cells and CD45+ CD11b+ macrophages in the brains of immunized, ECM-protected mice. CONCLUSIONS: This report further demonstrates the effectiveness of a whole parasite blood-stage vaccine in inducing immunity to malaria and explicitly demonstrates its effectiveness against ECM, the most pathogenic consequence of malaria infection. This experimental model will be important to explore the formulation of whole parasite blood-stage vaccines against malaria and to investigate the immune mechanisms that mediate protection against parasitemia and cerebral malaria

    CNS Recruitment of CD8+ T Lymphocytes Specific for a Peripheral Virus Infection Triggers Neuropathogenesis during Polymicrobial Challenge

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    Although viruses have been implicated in central nervous system (CNS) diseases of unknown etiology, including multiple sclerosis and amyotrophic lateral sclerosis, the reproducible identification of viral triggers in such diseases has been largely unsuccessful. Here, we explore the hypothesis that viruses need not replicate in the tissue in which they cause disease; specifically, that a peripheral infection might trigger CNS pathology. To test this idea, we utilized a transgenic mouse model in which we found that immune cells responding to a peripheral infection are recruited to the CNS, where they trigger neurological damage. In this model, mice are infected with both CNS-restricted measles virus (MV) and peripherally restricted lymphocytic choriomeningitis virus (LCMV). While infection with either virus alone resulted in no illness, infection with both viruses caused disease in all mice, with ∼50% dying following seizures. Co-infection resulted in a 12-fold increase in the number of CD8+ T cells in the brain as compared to MV infection alone. Tetramer analysis revealed that a substantial proportion (>35%) of these infiltrating CD8+ lymphocytes were LCMV-specific, despite no detectable LCMV in CNS tissues. Mechanistically, CNS disease was due to edema, induced in a CD8-dependent but perforin-independent manner, and brain herniation, similar to that observed in mice challenged intracerebrally with LCMV. These results indicate that T cell trafficking can be influenced by other ongoing immune challenges, and that CD8+ T cell recruitment to the brain can trigger CNS disease in the apparent absence of cognate antigen. By extrapolation, human CNS diseases of unknown etiology need not be associated with infection with any particular agent; rather, a condition that compromises and activates the blood-brain barrier and adjacent brain parenchyma can render the CNS susceptible to pathogen-independent immune attack

    IP-10-Mediated T Cell Homing Promotes Cerebral Inflammation over Splenic Immunity to Malaria Infection

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    Plasmodium falciparum malaria causes 660 million clinical cases with over 2 million deaths each year. Acquired host immunity limits the clinical impact of malaria infection and provides protection against parasite replication. Experimental evidence indicates that cell-mediated immune responses also result in detrimental inflammation and contribute to severe disease induction. In both humans and mice, the spleen is a crucial organ involved in blood stage malaria clearance, while organ-specific disease appears to be associated with sequestration of parasitized erythrocytes in vascular beds and subsequent recruitment of inflammatory leukocytes. Using a rodent model of cerebral malaria, we have previously found that the majority of T lymphocytes in intravascular infiltrates of cerebral malaria-affected mice express the chemokine receptor CXCR3. Here we investigated the effect of IP-10 blockade in the development of experimental cerebral malaria and the induction of splenic anti-parasite immunity. We found that specific neutralization of IP-10 over the course of infection and genetic deletion of this chemokine in knockout mice reduces cerebral intravascular inflammation and is sufficient to protect P. berghei ANKA-infected mice from fatality. Furthermore, our results demonstrate that lack of IP-10 during infection significantly reduces peripheral parasitemia. The increased resistance to infection observed in the absence of IP-10-mediated cell trafficking was associated with retention and subsequent expansion of parasite-specific T cells in spleens of infected animals, which appears to be advantageous for the control of parasite burden. Thus, our results demonstrate that modulating homing of cellular immune responses to malaria is critical for reaching a balance between protective immunity and immunopathogenesis

    CD8+ T Cells and IFN-γ Mediate the Time-Dependent Accumulation of Infected Red Blood Cells in Deep Organs during Experimental Cerebral Malaria

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    Background: Infection with Plasmodium berghei ANKA (PbA) in susceptible mice induces a syndrome called experimental cerebral malaria (ECM) with severe pathologies occurring in various mouse organs. Immune mediators such as T cells or cytokines have been implicated in the pathogenesis of ECM. Red blood cells infected with PbA parasites have been shown to accumulate in the brain and other tissues during infection. This accumulation is thought to be involved in PbA–induced pathologies, which mechanisms are poorly understood. Methods and Findings: Using transgenic PbA parasites expressing the luciferase protein, we have assessed by real-time in vivo imaging the dynamic and temporal contribution of different immune factors in infected red blood cell (IRBC) accumulation and distribution in different organs during PbA infection. Using deficient mice or depleting antibodies, we observed that CD8 + T cells and IFN-c drive the rapid increase in total parasite biomass and accumulation of IRBC in the brain and in different organs 6–12 days post-infection, at a time when mice develop ECM. Other cells types like CD4 + T cells, monocytes or neutrophils or cytokines such as IL-12 and TNF-a did not influence the early increase of total parasite biomass and IRBC accumulation in different organs. Conclusions: CD8 + T cells and IFN-c are the major immune mediators controlling the time-dependent accumulation of P. berghei-infected red blood cells in tissues

    CD4+ Natural Regulatory T Cells Prevent Experimental Cerebral Malaria via CTLA-4 When Expanded In Vivo

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    Studies in malaria patients indicate that higher frequencies of peripheral blood CD4+ Foxp3+ CD25+ regulatory T (Treg) cells correlate with increased blood parasitemia. This observation implies that Treg cells impair pathogen clearance and thus may be detrimental to the host during infection. In C57BL/6 mice infected with Plasmodium berghei ANKA, depletion of Foxp3+ cells did not improve parasite control or disease outcome. In contrast, elevating frequencies of natural Treg cells in vivo using IL-2/anti-IL-2 complexes resulted in complete protection against severe disease. This protection was entirely dependent upon Foxp3+ cells and resulted in lower parasite biomass, impaired antigen-specific CD4+ T and CD8+ T cell responses that would normally promote parasite tissue sequestration in this model, and reduced recruitment of conventional T cells to the brain. Furthermore, Foxp3+ cell-mediated protection was dependent upon CTLA-4 but not IL-10. These data show that T cell-mediated parasite tissue sequestration can be reduced by regulatory T cells in a mouse model of malaria, thereby limiting malaria-induced immune pathology

    Pathogenic Roles of CD14, Galectin-3, and OX40 during Experimental Cerebral Malaria in Mice

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    An in-depth knowledge of the host molecules and biological pathways that contribute towards the pathogenesis of cerebral malaria would help guide the development of novel prognostics and therapeutics. Genome-wide transcriptional profiling of the brain tissue during experimental cerebral malaria (ECM ) caused by Plasmodium berghei ANKA parasites in mice, a well established surrogate of human cerebral malaria, has been useful in predicting the functional classes of genes involved and pathways altered during the course of disease. To further understand the contribution of individual genes to the pathogenesis of ECM, we examined the biological relevance of three molecules – CD14, galectin-3, and OX40 that were previously shown to be overexpressed during ECM. We find that CD14 plays a predominant role in the induction of ECM and regulation of parasite density; deletion of the CD14 gene not only prevented the onset of disease in a majority of susceptible mice (only 21% of CD14-deficient compared to 80% of wildtype mice developed ECM, p<0.0004) but also had an ameliorating effect on parasitemia (a 2 fold reduction during the cerebral phase). Furthermore, deletion of the galectin-3 gene in susceptible C57BL/6 mice resulted in partial protection from ECM (47% of galectin-3-deficient versus 93% of wildtype mice developed ECM, p<0.0073). Subsequent adherence assays suggest that galectin-3 induced pathogenesis of ECM is not mediated by the recognition and binding of galectin-3 to P. berghei ANKA parasites. A previous study of ECM has demonstrated that brain infiltrating T cells are strongly activated and are CD44+CD62L− differentiated memory T cells [1]. We find that OX40, a marker of both T cell activation and memory, is selectively upregulated in the brain during ECM and its distribution among CD4+ and CD8+ T cells accumulated in the brain vasculature is approximately equal
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