Microbicides Development Programme: design of a phase III trial to measure the efficacy of the vaginal microbicide PRO 2000/5 for HIV prevention

Background

Non-pharmacological, non-surgical interventions are recommended as the first line of treatment for osteoarthritis (OA) of the hip and knee. There is evidence that exercise therapy is effective for reducing pain and improving function in patients with knee OA, some evidence that exercise therapy is effective for hip OA, and early indications that manual therapy may be efficacious for hip and knee OA. There is little evidence as to which approach is more effective, if benefits endure, or if providing these therapies is cost-effective for the management of this disorder. The MOA Trial (Management of OsteoArthritis) aims to test the effectiveness of two physiotherapy interventions for improving disability and pain in adults with hip or knee OA in New Zealand. Specifically, our primary objectives are to investigate whether: 1. Exercise therapy versus no exercise therapy improves disability at 12 months; 2. Manual physiotherapy versus no manual therapy improves disability at 12 months; 3. Providing physiotherapy programmes in addition to usual care is more cost-effective than usual care alone in the management of osteoarthritis at 24 months.

Methods

This is a 2 × 2 factorial randomised controlled trial. We plan to recruit 224 participants with hip or knee OA. Eligible participants will be randomly allocated to receive either: (a) a supervised multi-modal exercise therapy programme; (b) an individualised manual therapy programme; (c) both exercise therapy and manual therapy; or, (d) no trial physiotherapy. All participants will continue to receive usual medical care. The outcome assessors, orthopaedic surgeons, general medical practitioners, and statistician will be blind to group allocation until the statistical analysis is completed. The trial is funded by Health Research Council of New Zealand Project Grants (Project numbers 07/199, 07/200).

Discussion

The MOA Trial will be the first to investigate the effectiveness and cost-effectiveness of providing physiotherapy programmes of this kind, for the management of pain and disability in adults with hip or knee OA.

Trial registration

Australian New Zealand Clinical Trials Registry ref: ACTRN12608000130369.     

Mitochondria in the programmed death phenomena; a principle of biology: "it is better to die than to be wrong"

The very fact that mitochondria participate in amplification of the cell suicide signals has stimulated interest in the mechanism of this and related phenomena. It seems probable that mitochondria possess an autonomic system that allows them to commit suicide. This mitoptosis is mediated by reactive oxygen species (ROS), causing opening of the permeability transition pores (PTP) in the inner mitochondrial membrane. Mitoptosis can purify the mitochondrial population in a cell from the ROS-overproducing organelles. Massive mitoptosis can result in apoptosis (programmed cell death) because of the release of proapoptotic proteins from the mitochondrial intermembrane space, a mechanism purifying tissues from the ROS-overproducing and other unwanted cells. Large-scale apoptosis can be used by organisms to eliminate some organs during ontogenesis (organoptosis). In adult organisms, organoptosis of organs of vital importance may entail a programmed death of individuals (phenoptosis). This mechanism might purify kins, communities, and populations from individuals becoming dangerous because of, for example, heavy infection (septic shock). It is hypothesized that aging represents a slow ROS-linked phenoptosis that eliminates individuals with damaged genomes and gives reproductive advantage to those who succeeded in a better preservation of their genomes from damage.     

Uptake of Home-Based HIV Testing,Linkage to Care,and Community Attitudes about ART in Rural KwaZulu-Natal,South Africa: Descriptive Results from the First Phase of the ANRS 12249 TasP Cluster-Randomised Trial

BackgroundThe 2015 WHO recommendation of antiretroviral therapy (ART) for all immediately following HIV diagnosis is partially based on the anticipated impact on HIV incidence in the surrounding population. We investigated this approach in a cluster-randomised trial in a high HIV prevalence setting in rural KwaZulu-Natal. We present findings from the first phase of the trial and report on uptake of home-based HIV testing, linkage to care, uptake of ART, and community attitudes about ART.ConclusionsHome-based HIV testing was well received in this rural population, although men were less easily contactable at home; immediate ART was acceptable, with good viral suppression and retention. However, only about half of HIV-positive people accessed care within 6 mo of being identified, with nearly two-thirds accessing care by 12 mo. The observed delay in linkage to care would limit the individual and public health ART benefits of universal testing and treatment in this population.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01509508","term_id":"NCT01509508"}}NCT01509508     

Cytochrome oxidase I sequences reveal possible cryptic diversity in the cosmopolitan symbiotic copepod Nesippus orientalis Heller, 1868 (Pandaridae: Siphonostomatoida) on elasmobranch hosts from the KwaZulu-Natal coast of South Africa

Over the past decade, numerous molecular phylogenetic studies uncovered cryptic diversity within the Copepoda, yet very few investigations focused on symbiotic copepods. Here we report mitochondrial DNA cytochrome oxidase I diversity in the cosmopolitan elasmobranch symbiont Nesippus orientalis off the KwaZulu-Natal coast of South Africa. Analysis of partial COI sequences of copepods sampled from a diversity of shark hosts, revealed the presence of two divergent clades. Diversity within the clades does not appear to be structured based on host species, host individual, geographic locality or time of sampling. However, divergence between the two clades seems to be related to host species. Phylogenetic analyses of representatives from the two clades, along with Nesippus spp., Caligus spp. and Lepeophtheirus spp. outgroups, further supports the distinction between the two clades. Future molecular phylogenetic investigations of widespread copepod symbionts most likely will reveal far greater levels of biodiversity than currently recognized.     

Nse2, a component of the Smc5-6 complex, is a SUMO ligase required for the response to DNA damage

The Schizosaccharomyces pombe SMC proteins Rad18 (Smc6) and Spr18 (Smc5) exist in a high-M(r) complex which also contains the non-SMC proteins Nse1, Nse2, Nse3, and Rad62. The Smc5-6 complex, which is essential for viability, is required for several aspects of DNA metabolism, including recombinational repair and maintenance of the DNA damage checkpoint. We have characterized Nse2 and show here that it is a SUMO ligase. Smc6 (Rad18) and Nse3, but not Smc5 (Spr18) or Nse1, are sumoylated in vitro in an Nse2-dependent manner, and Nse2 is itself autosumoylated, predominantly on the C-terminal part of the protein. Mutations of C195 and H197 in the Nse2 RING-finger-like motif abolish Nse2-dependent sumoylation. nse2.SA mutant cells, in which nse2.C195S-H197A is integrated as the sole copy of nse2, are viable, whereas the deletion of nse2 is lethal. Smc6 (Rad18) is sumoylated in vivo: the sumoylation level is increased upon exposure to DNA damage and is drastically reduced in the nse2.SA strain. Since nse2.SA cells are sensitive to DNA-damaging agents and to exposure to hydroxyurea, this implicates the Nse2-dependent sumoylation activity in DNA damage responses but not in the essential function of the Smc5-6 complex.     

Cytoplasmic expression of SSTR2 and 5 by immunohistochemistry and by RT/PCR is not associated with the pharmacological response to octreotide

ObjectiveTo evaluate expression of somatostatin receptor subtypes 2 and 5 (SSTR 2 and 5) by RT/PCR and immunohistochemistry (IHC) in GH-secreting adenomas, seeking correlations with response to octreotide.MethodsSSTR2 and 5 expression was tested by IHC (n = 37), RT/PCR (n = 36) or both (n = 13) in GH-secreting adenomas from 60 patients with acromegaly who had undergone pituitary surgery; 36 had been treated preoperatively with octreotide LAR for 3–6 months, and were categorized as responders (achievement of GH <2.5 ng/mL and a normal age-adjusted IGF-1), partial responders (GH and IGF-1 reduction >50% and >30%, respectively) or non-responders. IHC was performed on a tissue microarray using specific antibodies directed to the carboxyl terminus of SSTR2 and 5.ResultsSSTR5 was the predominantly expressed receptor subtype by both IHC and RT/PCR in all tumors tested, regardless of whether they came from octreotide-naïve, octreotide-responsive, or octreotide-resistant patients. Immunostaining was concentrated in the cytoplasm. Neither SSTR2 nor SSTR5 expression correlated with baseline or post-octreotide GH or IGF-1 levels or tumor volume by either method. The agreement rate between RT/PCR and IHC was 77% in all 13 adenomas in which both methods were used.ConclusionExpression of these receptors does not guarantee an adequate response to somatostatin analogs; other functional aspects of this interaction, such as receptor homo- and heterodimerization, and the resulting signaling cascade, probably play a role in determining whether a patient will respond or not to these agents.     

5-HT1-, but not 5-HT2-serotonergic, M1-, M2-muscarinic cholinergic or dopaminergic receptors mediate the ACTH/cortisol response to metoclopramide in man

The possible mediation of dopaminergic, muscarinic cholinergic and/or serotonergic receptors in the response of ACTH/cortisol to metoclopramide (MCP) was evaluated in 27 normal men. All subjects were tested with MCP (10 mg in an intravenous bolus plus placebo or saline, NaCl 0.9%, control test). For the other tests (experimental tests), the men were divided into three groups of 9 subjects each. One group was tested with MCP in the presence of the dopaminergic agonist bromocriptine (5 mg p.o. 3 h before MCP), another group was tested with MCP plus the M1- and M2-muscarinic-cholinergic antagonist atropine (1.2 mg in an intravenous bolus, just before MCP) or the M1-muscarinic receptor blocker pirenzepine (40 mg in an intravenous bolus 10 min before MCP). The third group was tested with MCP after treatment with the selective 5-HT1-serotonergic receptor blocker metergoline (10 mg/day p.o. in 5 divided doses for 4 days before MCP) or the 5-HT2-serotonergic receptor antagonist ketanserin (10 mg as a slow 3-min intravenous injection, 5 min before MCP). ACTH and cortisol rose by 45 and 55%, respectively, in response to MCP. The basal levels of ACTH and cortisol were not modified by bromocriptine, atropine, pirenzepine, metergoline or ketanserin treatment. Both ACTH and cortisol responses to MCP did not change significantly after bromocriptine, atropine, pirenzepine or ketanserin administration, whereas they were completely abolished by pretreatment with metergoline. Additional experiments were performed in order to evaluate whether the effect of metergoline on the ACTH/cortisol response to MCP depends on the amount of the serotonergic antagonist (dose-response study).(ABSTRACT TRUNCATED AT 250 WORDS)     

PINCH2 is a new five LIM domain protein, homologous to PINCHand localized to focal adhesions

PINCH is a five LIM domain protein involved in the regulation of integrin-mediated cell adhesion. It has been shown that PINCH interacts with integrin-linked kinase and Nck2. Here we describe a new isoform of PINCH, which we call PINCH2. Therefore, we rename PINCH to PINCH1. PINCH2 has an overall similarity of 92% to PINCH1 and contains five LIM domains like PINCH1. While protein and gene structure of the PINCH homologues are very similar and well conserved during evolution, we observed differential expression pattern of the mRNAs. Based on northern hybridization of mouse embryo RNA, PINCH1 is already detectable at E8.5. It is highly expressed during later stages of development and in all adult mouse tissues analyzed, with the highest levels in heart, lung, bladder, skin, and uterus. In contrast, significant PINCH2 expression starts at E14.5. In adult mice it is widely expressed, similar to PINCH1, but absent from spleen and thymus. In situ hybridization confirmed the Northern data and showed differential expression of PINCH1 and PINCH2 in embryonic intestine. Finally, we demonstrate that PINCH2 localizes to focal adhesions in NIH 3T3 cells and to Z-disks in primary rat cardiomyocytes.     

Movement response of Orange-Vaal largemouth yellowfish (<Emphasis Type="Italic">Labeobarbus kimberleyensis</Emphasis>) to water quality and habitat features in the Vaal River,South Africa

Many threatened fish species that utilize riverine habitats are faced with habitat degradation and subsequent deterioration in their ecological surroundings. Habitat degradation is a consequence of water quality parameters associated with anthropogenic activities including mining, industrial, agricultural and urban activities. We examined how the movement behaviour of radio-tracked Orange-Vaal largemouth yellowfish (Labeobarbus kimberleyensis) responded to a suite of water quality chemical parameters and habitat features in the Vaal River, South Africa. We found that the probability of their movement increased with a decrease in water clarity, presence of emergent and overhanging marginal vegetation and fast flowing rapids. High mobility in conditions of low water clarity was probably related to low prey capture success of this piscivorous fish. High movement of largemouth yellowfish in emergent and overhanging marginal vegetation areas and rapid habitat biotopes were attributed to accessibility of prey within these important cover features. When water quality parameters were considered, the probability of largemouth yellowfish movement increased with increasing levels of dissolved chloride (Cl) and silicon (Si), whereas movement decreased with high phosphate (PO₄) concentrations and increased with alkalinity levels in the river. High nutrient levels associated with eutrophication caused reductions in the movement of largemouth yellowfish. The association of increased movement of largemouth yellowfish with increasing Cl and Si is indicative of degraded habitat condition in the Vaal River system. Our study showed the importance of monitoring fish behavioural movement patterns to multiple environmental parameters, as these fish are important ecological indicators when appropriate conservation and management plans of freshwater ecosystems are required.     

Specificity of the mouse cytotoxic T lymphocyte response to adenovirus 5. E1A is immunodominant in H-2b, but not in H-2d or H-2k mice

The Ag specificity and MHC restriction of the CTL response to adenovirus 5 (Ad5) in three strains of mice, C57BL/10 (H-2b), BALB/c (H-2d), and C3H/HeJ (H-2k), were tested. Polyclonal Ad5-specific CTL were prepared by priming mice in vivo with live Ad5 virus followed by secondary in vitro stimulation of the spleen cells with virus-infected syngeneic cells. The Ad5-specific CTL were Db restricted in C57BL/10 and Kk restricted in C3H/HeJ. In BALB/c mice both Kd- and Dd/Ld-restricted CTL were detected. The polyclonal Ad5-specific CTL response in C57BL/10 mice is directed exclusively against the products of the E1A region, which comprises only 5% of the Ad5 genome. In BALB/c mice E1A is at best a very minor target Ag and in C3H/HeJ mice E1A is not recognized at all. Using the H-2 congenic mouse strains B10.BR (H-2k) and C3H.SW (H-2b) it was shown that the immunodominance of E1A is H-2 dependent. The 19-kDa glycoprotein encoded in the E3 region of Ad5, which binds to class I MHC in the endoplasmic reticulum and prevents its translocation to the cell surface, does not affect the specificity of the CTL response in C57BL/10 mice toward E1A. However, it affects the MHC restriction of the Ad5-specific response in BALB/c mice, selectively inhibiting generation of Kd-restricted CTL.     

<Emphasis Type="Italic">Betula</Emphasis> pollen: One of the most important aeroallergens in Ourense,Spain. Aerobiological studies from 1993 to 2000

This study presents the results obtained for airborne Betula pollen between 1992 and 2000 in Ourense, Spain, sampled by volumetric spore-trap (LANZONI VPPS2000). Annual and year-on-year variations were analyzed, and a statistical study of the correlation between daily counts and several meteorological parameters was performed. Birch pollen is present in the atmosphere during March and April in Ourense. Significant differences were observed among the different years. Values obtained for the correlation coefficient between Betula pollen counts and the various meteorological parameters studied indicate, for Ourense, a positive correlation between pollen count and both temperature and sunlight. A negative correlation was recorded for relative humidity. Temperature is thus the determining factor for flowering onset and intensity. Regression equations included values for the days prior to pollen concentration measurement in order to optimize results.     

The proinflammatory cytokine response to Chlamydia trachomatis elementary bodies in human macrophages is partly mediated by a lipoprotein, the macrophage infectivity potentiator, through TLR2/TLR1/TLR6 and CD14

Chlamydiae components and signaling pathway(s) responsible for the production of proinflammatory cytokines by human monocytes/macrophages are not clearly identified. To this aim, Chlamydia trachomatis-inactivated elementary bodies (EB) as well as the following seven individual Ags were tested for their ability to induce the production of proinflammatory cytokines by human monocytes/macrophages and THP-1 cells: purified LPS, recombinant heat shock protein (rhsp)70, rhsp60, rhsp10, recombinant polypeptide encoded by open reading frame 3 of the plasmid (rpgp3), recombinant macrophage infectivity potentiator (rMip), and recombinant outer membrane protein 2 (rOmp2). Aside from EB, rMip displayed the highest ability to induce release of IL-1beta, TNF-alpha, IL-6, and IL-8. rMip proinflammatory activity could not be attributed to Escherichia coli LPS contamination as determined by the Limulus Amoebocyte lysate assay, insensitivity to polymyxin B (50 microg/ml), and different serum requirement. We have recently demonstrated that Mip is a "classical" bacterial lipoprotein, exposed at the surface of EB. The proinflammatory activity of EB was significantly attenuated in the presence of polyclonal Ab to rMip. Native Mip was able to induce TNF-alpha and IL-8 secretion, whereas a nonlipidated C20A rMip variant was not. Proinflammatory activity of rMip was unaffected by heat or proteinase K treatments but was greatly reduced by treatment with lipases, supporting a role of lipid modification in this process. Stimulating pathways appeared to involve TLR2/TLR1/TLR6 with the help of CD14 but not TLR4. These data support a role of Mip lipoprotein in pathogenesis of C. trachomatis-induced inflammatory responses.     

Cytochrome bd-I in Escherichia coli is less sensitive than cytochromes bd-II or bo′' to inhibition by the carbon monoxide-releasing molecule,CORM-3: N-acetylcysteine reduces CO-RM uptake and inhibition of respiration

Background: CO-releasing molecules (CO-RMs) are potential therapeutic agents, able to deliver CO – a critical gasotransmitter – in biological environments. CO-RMs are also effective antimicrobial agents; although the mechanisms of action are poorly defined, haem-containing terminal oxidases are primary targets. Nevertheless, it is clear from several studies that the effects of CO-RMs on biological systems are frequently not adequately explained by the release of CO: CO-RMs are generally more potent inhibitors than is CO gas and other effects of the molecules are evident. Methods: Because sensitivity to CO-RMs cannot be predicted by sensitivity to CO gas, we assess the differential susceptibilities of strains, each expressing only one of the three terminal oxidases of E. coli — cytochrome bd-I, cytochrome bd-II and cytochrome bo′, to inhibition by CORM-3. We present the first sensitive measurement of the oxygen affinity of cytochrome bd-II (K_m 0.24 μM) employing globin deoxygenation. Finally, we investigate the way(s) in which thiol compounds abolish the inhibitory effects of CORM-2 and CORM-3 on respiration, growth and viability, a phenomenon that is well documented, but poorly understood. Results: We show that a strain expressing cytochrome bd-I as the sole oxidase is least susceptible to inhibition by CORM-3 in its growth and respiration of both intact cells and membranes. Growth studies show that cytochrome bd-II has similar CORM-3 sensitivity to cytochrome bo′. Cytochromes bo′ and bd-II also have considerably lower affinities for oxygen than bd-I. We show that the ability of N-acetylcysteine to abrogate the toxic effects of CO-RMs is not attributable to its antioxidant effects, or prevention of CO targeting to the oxidases, but may be largely due to the inhibition of CO-RM uptake by bacterial cells. Conclusions: A strain expressing cytochrome bd-I as the sole terminal oxidase is least susceptible to inhibition by CORM-3. N-acetylcysteine is a potent inhibitor of CO-RM uptake by E. coli. General significance: Rational design and exploitation of CO-RMs require a fundamental understanding of their activity. CO and CO-RMs have multifaceted effects on mammalian and microbial cells; here we show that the quinol oxidases of E. coli are differentially sensitive to CORM-3. This article is part of a Special Issue entitled: Oxygen Binding and Sensing Proteins.