RENEB Inter-Laboratory comparison 2017: limits and pitfalls of ILCs.

PURPOSE: In case of a mass-casualty radiological event, there would be a need for networking to overcome surge limitations and to quickly obtain homogeneous results (reported aberration frequencies or estimated doses) among biodosimetry laboratories. These results must be consistent within such network. Inter-laboratory comparisons (ILCs) are widely accepted to achieve this homogeneity. At the European level, a great effort has been made to harmonize biological dosimetry laboratories, notably during the MULTIBIODOSE and RENEB projects. In order to continue the harmonization efforts, the RENEB consortium launched this intercomparison which is larger than the RENEB network, as it involves 38 laboratories from 21 countries. In this ILC all steps of the process were monitored, from blood shipment to dose estimation. This exercise also aimed to evaluate the statistical tools used to compare laboratory performance. MATERIALS AND METHODS: Blood samples were irradiated at three different doses, 1.8, 0.4 and 0 Gy (samples A, C and B) with 4-MV X-rays at 0.5 Gy min-1, and sent to the participant laboratories. Each laboratory was requested to blindly analyze 500 cells per sample and to report the observed frequency of dicentric chromosomes per metaphase and the corresponding estimated dose. RESULTS: This ILC demonstrates that blood samples can be successfully distributed among laboratories worldwide to perform biological dosimetry in case of a mass casualty event. Having achieved a substantial harmonization in multiple areas among the RENEB laboratories issues were identified with the available statistical tools, which are not capable to advantageously exploit the richness of results of a large ILCs. Even though Z- and U-tests are accepted methods for biodosimetry ILCs, setting the number of analyzed metaphases to 500 and establishing a tests' common threshold for all studied doses is inappropriate for evaluating laboratory performance. Another problem highlighted by this ILC is the issue of the dose-effect curve diversity. It clearly appears that, despite the initial advantage of including the scoring specificities of each laboratory, the lack of defined criteria for assessing the robustness of each laboratory's curve is a disadvantage for the 'one curve per laboratory' model. CONCLUSIONS: Based on our study, it seems relevant to develop tools better adapted to the collection and processing of results produced by the participant laboratories. We are confident that, after an initial harmonization phase reached by the RENEB laboratories, a new step toward a better optimization of the laboratory networks in biological dosimetry and associated ILC is on the way.

Regulation of NF-κB- and STAT1-mediated plasmacytoid dendritic cell functions by A20.

Dendritic cells (DCs) are professional antigen presenting cells involved in the induction of T cell-mediated adaptive immunity. Plasmacytoid DCs (pDCs) originate from lymphoid precursors and produce type I interferons (IFNs) in response to pathogens. A20 is considered as a negative regulator of toll-like receptor (TLR) signaling pathways, in which Toxoplasma gondii- derived profilin (TgPRF) is a TLR11/12 ligand recognised by DCs to stimulate their maturation/activation. Little is known about contributions of A20 to changes in biological properties of pDCs. The present study, therefore, explored whether pDC functions are influenced by A20. To this end, bone marrow cells were isolated and cultured with Flt3L to attain CD8DCs, CD11bDCs and pDCs and followed by challenge with TgPRP in the presence or absence of A20 siRNA. Expression of maturation markers were analysed by flow cytometry, and secretion of inflammatory cytokines by ELISA, cell migration by a transwell migration assay and expression of signalling molecules by western blotting. As a result, treatment with A20 siRNA enhanced activations of IκB-α and STAT-1, leading to increases in expressions of maturation markers and cytokine productions as well as migration of TgPRP-treated pDCs, while mature CD11bDCs produced at higher levels of TNF-α and IL-6 only. In addition, functions of CD8DCs remained unaltered following A20 silencing. The effects of A20 on pDC maturation and activation were completely abolished by IKK inhibitor and partially blunted by fludarabine. In conclusion, the inhibitory effects of A20 on pDC functions are expected to affect the immune response in T. gondii infection.

RENEB intercomparisons applying the conventional Dicentric Chromosome Assay (DCA).

PURPOSE: Two quality controlled inter-laboratory exercises were organized within the EU project 'Realizing the European Network of Biodosimetry (RENEB)' to further optimize the dicentric chromosome assay (DCA) and to identify needs for training and harmonization activities within the RENEB network. MATERIALS AND METHODS: The general study design included blood shipment, sample processing, analysis of chromosome aberrations and radiation dose assessment. After manual scoring of dicentric chromosomes in different cell numbers dose estimations and corresponding 95% confidence intervals were submitted by the participants. RESULTS: The shipment of blood samples to the partners in the European Community (EU) were performed successfully. Outside the EU unacceptable delays occurred. The results of the dose estimation demonstrate a very successful classification of the blood samples in medically relevant groups. In comparison to the 1st exercise the 2nd intercomparison showed an improvement in the accuracy of dose estimations especially for the high dose point. CONCLUSIONS: In case of a large-scale radiological incident, the pooling of ressources by networks can enhance the rapid classification of individuals in medically relevant treatment groups based on the DCA. The performance of the RENEB network as a whole has clearly benefited from harmonization processes and specific training activities for the network partners.

Calibration Curve for Dicentric Chromosomes Induced in Human Blood Lymphocytes Exposed to Gamma Rays at a Dose Rate of 12.5 mGy/s.

To develop a calibration curve for induction of dicentric chromosomes by radiation, we have used a 60Co gamma-ray source with dose rate of 12.5 mGy/s. Whole blood from 15 healthy donors was collected. Whole blood from each donor was divided equally into 8 parts for exposing to supposedly physical doses 0, 0.30, 0.50, 1.00, 1.50, 2.00, 3.00 and 4.00 Gy for a independent calibration curve. Whole blood from 15 donors was used to calibrate dose - effect and statistical for general calibration curve. Using Poisson test (U-test) for the distribution of dicentric chromosomes in the metaphases to determine the uniformity of the radiation field. The average from 15 independent calibration curves of linear correlated coefficient was determined to be r (y, d) = 0.5136 ± 0.0038. The model equation derived is y = aD + bD2 + C. The calibration equation of dose-effect was y = 1.01D + 4.43D2 + 0.56.