Posters. P11 Positive predictive values for high and low‐grade cytological abnormalities as surrogates for specificity and sensitivity

Introduction Positive predictive value (PPV), measuring the percentage of moderate dyskaryosis or worse confirmed as CIN2 or worse, is used as a measure of accuracy in cervical screening. However, it relates more to specificity than sensitivity because the denominator includes false positives rather than false negatives. Low values reflect over‐reporting of high‐grade dyskaryosis but high values may reflect under‐reporting. Sensitivity is impossible to measure from correlation of cytology with outcome because women with negative cytology are rarely referred for colposcopy. Rates of CIN3 resulting from referrals for low‐grade cytology may be used as a surrogate for sensitivity, as high values may reflect under‐reporting (ref). Study design Outcome of colposcopy referrals was monitored during a period of 4 years, using a fail‐safe database. Results PPV at Guy's & St Thomas rose from 54% in 1998/1999 to 69% in 2001/2002. The former was below the NHSCSP recommended range. During the same period of time CIN1 rates for moderate dyskaryosis fell from 37% to 24%, reflecting the main source of discrepancy. While specificity increased (as reflected by increasing PPV) sensitivity remained constant in that CIN3 rates for mild dyskaryosis and borderline remained below 6%: average rates in England have fallen over the last 3 years and were 7.4% in 2000/2001 (ref). CIN2 rates for mild dyskaryosis also remained constant at 11% to 12%. Conclusion Correlation of biopsy results with high‐ and low‐grade cytological abnormalities is a useful method of monitoring accuracy of cytology reporting, and can be used to measure over‐ and under‐reporting as surrogates for specificity and sensitivity.     

Linkage of interactions in sickle hemoglobin fiber assembly: inhibitory effect emanating from mutations in the AB region of the alpha-chain is annulled by a mutation at its EF corner

The AB and GH regions of the alpha-chain are located in spatial proximity and contain a cluster of intermolecular contact residues of the sickle hemoglobin (HbS) fiber. We have examined the role of dynamics of AB/GH region on HbS polymerization through simultaneous replacement of non-contact Ala(19) and Ala(21) of the AB corner with more flexible Gly or rigid alpha-aminoisobutyric acid (Aib) residues. The polymerization behavior of HbS with Aib substitutions was similar to the native HbS. In contrast, Gly substitutions inhibited HbS polymerization. Molecular dynamics simulation studies of alpha-chains indicated that coordinated motion of AB and GH region residues present in native (Ala) as well as in Aib mutant was disrupted in the Gly mutant. The inhibitory effect due to Gly substitutions was further explored in triple mutants that included mutation of an inter-doublestrand contact (alphaAsn(78) --> His or Gln) at the EF corner. Although the inhibitory effect of Gly substitutions in the triple mutant was unaffected in the presence of alphaGln(78), His at this site almost abrogated its inhibitory potential. The polymerization studies of point mutants (alphaGln(78) --> His) indicated that the inhibitory effect due to Gly substitutions in the triple mutant was synergistically compensated for by the polymerization-enhancing activity of His(78). Similar synergistic coupling, between alphaHis(78) and an intra-double-strand contact point (alpha16) mutation located in the AB region, was also observed. Thus, two conclusions are made: (i) Gly mutations at the AB corner inhibit HbS polymerization by perturbing the dynamics of the AB/GH region, and (ii) perturbations of AB region (through changes in dynamics of the AB/GH region or abolition of a specific fiber contact site) that influence HbS polymerization do so in concert with alpha78 site at the EF corner. The overall results provide insights about the interaction-linkage between distant regions of the HbS tetramer in fiber assembly.     

Review and new insights on Wegener granulomatosis

The purpose of this paper is to present Wegener granulomatosis manifesting in its periocular form. Review with new developments in understanding of the etiopathogenesis, clinical and laboratory findings as well as therapy modalities are shown through a series of patients gathered in collaboration with colleagues from UMC Utrecht, Netherlands. In the period from 1992 until 2004 the group of 54 patients with established diagnosis of Mb. Wegener were observed. 13 patients developed periocular form but only 2 presented it as initial symptoms. Lacrimal stenosis and orbital infiltration were predominant periocular symptoms while nasal manifestations were predominant systemic symptoms of the disease. Different treatment modalities were employed showing that orbital disease is difficult to treat in spite of satisfying systemic answer to immunosuppressives which calls for alternative solutions.     

Intra-articular knee implantation of autologous bone marrow–derived mesenchymal stromal cells in rheumatoid arthritis patients with knee involvement: Results of a randomized,triple-blind,placebo-controlled phase 1/2 clinical trial

Background

In this study, we intend to assess the safety and tolerability of intra-articular knee implantation of autologous bone marrow–derived mesenchymal stromal cells (MSCs) in patients with rheumatoid arthritis (RA) and to determine the preliminary clinical efficacy data in this population. The trial registration numbers are as follows: Royan Institute Ethics Committee: AC/91/1133; NCT01873625.

Strong responses of Drosophila melanogaster microbiota to developmental temperature

Physiological responses to changes in environmental conditions such as temperature may partly arise from the resident microbial community that integrates a wide range of bio-physiological aspects of the host. In the present study, we assessed the effect of developmental temperature on the thermal tolerance and microbial community of Drosophila melanogaster. We also developed a bacterial transplantation protocol in order to examine the possibility of reshaping the host bacterial composition and assessed its influence on the thermotolerance phenotype. We found that the temperature during development affected thermal tolerance and the microbial composition of male D. melanogaster. Flies that developed at low temperature (13°C) were the most cold resistant and showed the highest abundance of Wolbachia, while flies that developed at high temperature (31°C) were the most heat tolerant and had the highest abundance of Acetobacter. In addition, feeding newly eclosed flies with bacterial suspensions from intestines of flies developed at low temperatures changed the heat tolerance of recipient flies. However, we were not able to link this directly to a change in the host bacterial composition.     

<Emphasis Type="Italic">Lactobacillus salivarius</Emphasis> NK02: a Potent Probiotic for Clinical Application in Mouthwash

A specific strain of naturally occurring oral lactobacilli was isolated and identified based on morphological, biochemical, and 16S rRNA gene sequencing. The phylogenetic affiliation of the isolate confirmed that the NK02 strain had close association with the Lactobacillus salivarius. An effective mouthwash was developed for treatment of periodontitis and suppression of the indicator bacterium Aggregatibacter actinomycetemcomitans which is an obvious pathogen of periodontal disease. The mouthwash containing L. salivarius NK02 was tested at a dose level of 10⁸ (colony forming units (CFU) ml^?1), monitoring over a period of 4 weeks. The study was a randomized double-blind placebo control trial, and the patients were treated in two groups of control and test by using scaling and root planing (SRP) + placebo and scaling and root planing (SRP) + probiotic, respectively. It appeared that the probiotic mouthwash was able to inhibit the bacterial growth on both saliva and sub-gingival crevice and exhibited antibacterial activity against A. actinomycetemcomitans. The results also showed that SRP+ probiotic treatment led to a significant decrease of gingival index (GI) and bleeding on probing (BOP) compared with that of SRP + placebo for the probiotic group. The rate of decrease in pocket depth was displayed in the group with SRP + probiotic treatment equal to 1/2 mm, and probing pocket depth (PPD) value was decreased in the probiotic bacteria treatment group that can explain the decrease in inflammation in gingiva. Our findings suggest that probiotic mouthwash is healthy for daily use as an alternative for maintaining dental and periodontal health.     

A qPCR assay that specifically quantifies <Emphasis Type="Italic">Tricholoma matsutake</Emphasis> biomass in natural soil

Tricholoma matsutake is an ectomycorrhizal basidiomycete that produces prized, yet uncultivable, “matsutake” mushrooms along densely developed mycelia, called “shiro,” in the rhizosphere of coniferous forests. Pinus densiflora is a major host of this fungus in Japan. Measuring T. matsutake biomass in soil allows us to determine the kinetics of fungal growth before and after fruiting, which is useful for analyzing the conditions of the shiro and its surrounding mycorrhizosphere, predicting fruiting timing, and managing forests to obtain better crop yields. Here, we document a novel method to quantify T. matsutake mycelia in soil by quantifying a single-copy DNA element that is uniquely conserved within T. matsutake but is absent from other fungal species, including close relatives and a wide range of ectomycorrhizal associates of P. densiflora. The targeted DNA region was amplified quantitatively in cultured mycelia that were mixed with other fungal species and soil, as well as in an in vitro co-culture system with P. densiflora seedlings. Using this method, we quantified T. matsutake mycelia not only from shiro in natural environments but also from the surrounding soil in which T. matsutake mycelia could not be observed by visual examination or distinguished by other means. It was demonstrated that the core of the shiro and its underlying area in the B horizon are predominantly composed of fungal mycelia. The fungal mass in the A or A₀ horizon was much lower, although many white mycelia were observed at the A horizon. Additionally, the rhizospheric fungal biomass peaked during the fruiting season.     

Experimental and theoretical studies of cadmium ions absorption by a new reduced recombinant defensin

Heavy metal pollutants such as Cd, Hg, Pb, As, and Se are considered as both a global problem and a growing threat to the humanity. Being strongly poisonous to the metal-sensitive enzymes and leading to the growth inhibition and death of organisms, these metals have a toxic impact on the plants and animals. Inducing the metal-binding cysteine-rich peptides such as metallothioneins, phytochelatins, and defensins, higher organisms like plants and animals usually react to the heavy metal stress. In this study, a recombinant defensin protein was expressed in bean and its ability in the cadmium absorption was determined. Experimental studies revealed that this protein was able to absorb cadmium ions in reduced form more than oxide one. Molecular dynamics simulations were carried out in order to evaluation of experimental studies, using a model of Cd²⁺ or Na⁺ and Cl^– ions enclosed in a fully hydrated simulation box with the recombinant defensin. The theoretical results also suggested that the reduced recombinant defensin was more powerful in the absorption of Cd²⁺ than its oxide form. The present study is the first report of Cd²⁺ absorption potential of this new reduced recombinant defensin. The results unraveled that this recombinant defensin can be adopted as a molecular switch in the cadmium pollution of the environment and also the important role of sulfur groups in the absorption of cadmium ions.     

Ameliorating Effect of Osteopontin on H<Subscript>2</Subscript>O<Subscript>2</Subscript>-Induced Apoptosis of Human Oligodendrocyte Progenitor Cells

Recently our group used oligodendrocyte progenitor cells (OPCs) as appropriate model cells to pinpoint the mechanism of the progress of neurodegenerative disorders. In the present study, we focused on the therapeutic role of osteopontin (OPN), a secreted glycosylated phosphoprotein, involved in a number of physiological events including bone formation and remodeling, immune responses, and tumor progression. Protective role of OPN, as a negative regulator of tumorigenesis, has already been clarified. Human embryonic stem cell-derived OPCs were pretreated with OPN before induction of apoptosis by H₂O₂. Data indicated that OPN prohibited cell death and enhanced OPC viability. This effect is achieved through reduction of apoptosis and induction of anti-apoptosis markers. In addition OPN induces expression of several integrin subunits, responsible for OPN interaction. Notably, our findings showed that expression of αV β1/β3/β5 and β8 integrins increased in response to OPN, while treatment with H₂O₂ down-regulated αV β1/β5 and β8 integrins expression significantly. In conclusion, OPN may act via αV integrin signaling and trigger suppression of P53-dependent apoptotic cascades. Therefore OPN therapy may be considered as a feasible process to prevent progress of neurodegenerative diseases in human.