Immunogenecity of <i>Mycobacterium avium</i> subsp. <i>paratuberculosis</i> epitopes cross-reacting with human ZnT8 and proinsulin peptides in autoimmune diabetes

Although numerous studies put in evidence the increasing incidence of type 1 diabetes (T1D), its cause remains unclear. The role of Mycobacterium avium subsp. paratuberculosis (MAP) as a putative environmental agent triggering or accelerating the disease has been previously hypothesized in Sardinian and Italian T1D populations. The present thesis further sustains this association by reporting an elevated seroreactivity to MAP-derived epitopes and homologous human peptides corresponding to proinsulin and ZnT8 fragments in populations at different T1D stages and originating from distinct biogeographic backgrounds. Anti-MAP antibodies (Abs) resulted detectable in the first months of life before the appearance of classical autoantigens and, in most cases, were maintained in time making the selected peptides good candidates for early biomarkers. Likewise, Abs responses to the same antigens were observed among LADA patients and subjects affected by Hashimoto’s thyroiditis which frequently complicates T1D. Validation with a MAP-specific lipopentapeptide confirmed these results in coincidence with a stable Abs status. In PBMC primary culture, ZnT8 peptide and its MAP homolog induced the expression of proinflammatory cytokines along with increased cell-mediated responses and apoptosis. Good correlation between values obtained for the homologous MAP/human peptide pairs point at cross-reactivity through which mechanisms of self tolerance may be disrupted leading to autoimmunity

Hypotensive efficacy of eprosartan in postmenopausal women

Wstęp Celem pracy była ocena skuteczności eprosartanu w monoterapii u kobiet w okresie pomenopauzalnym z pierwotnym nadciśnieniem tętniczym. Materiał i metody Badania przeprowadzono u 52 kobiet z nadciśnieniem tętniczym pierwotnym,w wieku 52,8 &#177; 3,4 rż., u których ostatnie krwawienie miesięczne wystąpiło przed dwoma laty. W momencie włączenia do leczenia eprosartanu (Teveten) w dawce 600 mg/dobę w godzinach 6.30-7.00 pozostawały one co najmniej 1 miesiąc bez leków hipotensyjnych. Wykonano u nich całodobową rejestrację ciśnienia aparatem SpaceLabs 90207 przed włączeniem leku. Skuteczność terapii oceniano po 6 i 12 tygodniach leczenia. Wyniki Najniższe wartości w ciągu doby obu ciśnień SBP i DBP zanotowano po 12 tygodniach leczenia eprosartanem. Po 6-tygodniowym stosowaniu eprosartanu zaobserwowano istotne statystycznie obniżenie się wartości SBP i DBP zarówno w ciągu dnia, jak i w nocy w porównaniu z wartościami sprzed leczenia eprosartanem. Kontynuując leczenie przez następne 6 tygodni, uzyskano dalsze istotne statystycznie obniżenie SBP tylko w ciągu dnia. Najniższe wartości w ciągu doby HT zanotowano po 12. tygodniu leczenia eprosartanem. Różnice były istotne statystycznie w ciągu dnia i nocy w porównaniu z HT zanotowanej przed leczeniem, a w porównaniu z wartościami uzyskanymi w 6. tygodniu leczenia istotne statystycznie - tylko w ciągu dnia. Wnioski Eprosartan jest skutecznym lekiem hipotensyjnym w monoterapii u kobiet w okresie pomenopauzalnym i wykazuje intensywniejsze działanie hipotensyjne po 12-tygodniowym stosowaniu w niezmienionej dawce niż po 6-tygodniowym leczeniu. Eprosartan wpływa korzystnie na częstość rytmu serca przez całą dobę zarówno w dzień, jak i szczególnie w nocy. Nadciśnienie Tętnicze 2007, tom 11, nr 4, strony 328&#8211;334.Background The aim of the study was to evaluate the efficacy of eprosartan monotherapy treatment in postmenopausal women with essential hypertension. Material and methods 52 women with essential hypertension aged 52.8 &#177; 3.4 years who had last menstrual period 2 years ago. The women were not treated with any hypertensive medication for at least 1 month. Women received eprosartan (Teveten) 600 mg/daily between 6.30-7.00 am. Ambulatory blood pressure monitoring was performed with SpaceLabs 90207 before treatment, 6 and 12 weeks of therapy with eprosartan. Results Twelve weeks of eprosartan treatment resulted in the lowest values of both SBP and DBP during day and night. After 6 weeks of eprosartan treatment in comparison with the period before treatment the significant reduction of systolic blood pressure (SBP) and diastolic blood pressure (DBP) during day and night was observed. During next 6 weeks further, statistically significant reduction of SBP only during day was observed. After 12 weeks of eprosartan treatment the lowest HT was also observed. The differences were statistically significant when compared to HT noted before treatment during day and night. The following 6 weeks of treatment showed statistically significant decreased values only during day. Conclusions Eprosartan is an effective hypotensive monotherapy in postmenopausal women and showes more intensive hypotensive effects after 12 weeks of treatment with a stable dose than after 6 weeks of treatment. Eprosartan presents a good effect on the heart rate during day but especially during night.Arterial Hypertension 2007, vol. 11, no 4, pages 328-334

Microbiological parameters under the Drinking Water Directive.

In November 1998, the European Council adopted a directive, the Drinking Water Directive (DWD), concerning the quality of water intended for human consumption. It includes a certain number of microbiological, chemical or physical criteria or parameters to monitor, to ensure that i) it is “clean”, ii) the distribution network is safe and iii) to react promptly in case of contamination (Directive 98/83/EC)1. The Directive has been implemented by Member States, but its approach to monitoring quality at the point of consumption uses parameters determined over 20 years ago. After the submission of the European’s citizens’ initiative “Right2Water” to the Commission in December 2013, the Commission invited Member States to improve the access to a minimum water supply and the management of water in a sustainable manner. In 2017, following the WHO recommendations2, the Drinking Water Directive (DWD) was revised either for the microbiological or chemical parameters (RECAST DWD). Among the first ones, somatic coliphage (virus infecting Escherichia coli) has been proposed as new parameter, while bacterium Clostridium perfringens (C. perfringens) and its spores are already included in the Directive. The present report provides an overview on the current knowledge of these two microbiological parameters, their biological characterisations, relevance and suitability as indicators for human faecal contamination in the drinking water treatment. Finally, the report illustrates the available and standardised methods for their detection in water, listing as well the new and most promising ones with advantages/disadvantages and costs. Furthermore, the report provides a list of recommendations in order to elucidate the role of the two microbiological parameters for drinking water quality management.JRC.D.2-Water and Marine Resource

Lack of humoral response against <i>Helicobacter pylori</i> peptides homologous to human ZnT8 in Hashimoto's thyroiditis patients

Introduction: The Helicobacter pylori (HP) reinfection rate seems to be higher in developing countries than in developed ones. An increased seroprevalence of HP has also been reported in patients with type 1 diabetes (T1D) and Hashimoto’s thyroiditis (HT). Mycobacterium avium subsp. paratuberculosis (MAP) has been linked to both T1D and HT. Quite a few lines of evidence indicate that autoantibodies against several epitopes belonging to human zinc transporter 8 (ZnT8) cross-recognize the homologous MAP3865c epitopes in both T1D and HT patients. HP may play a role in HT disease, most likely acting through a molecular mimicry mechanism that targets ZnT8 as reported for MAP and the two autoimmune diseases. Methodology: An enzyme-linked immunosorbent assay (ELISA) has been developed for the detection of antibodies against several epitopes deriving from HP proteins, which are highly homologous to the immunodominant ZnT8 peptides previously identified: ZnT8178–186 and ZnT8186–194. Results: None of the HP peptides tested were significantly recognized when the humoral responses of 92 HT patients and 91 healthy volunteers were analyzed. Conclusions: These findings do not support a triggering role for HP (through ZnT8 mimicking) in HT. If a molecular mimicry phenomenon is taking place, it involves a different self-antigen. Moreover, the negative outcome of the experiments performed stresses the fact that sharing stretches of sequence homology is relevant, but not enough to trigger an antibody-mediated cross-recognition

Modes of action of the current Priority Substances list under the Water Framework Directive and other substances of interest

The Water Framework Directive 2000/60/EC (WFD) has established a strategy for water protection that includes specific measures for pollution control to achieve good chemical and ecological status at European level. There is a need to review the approach to the current listing of priority substances (PS) under the WFD and to the current assessment of the chemical status, and consider eventually a wider range of chemical substances that could be covered in future monitoring programmes. Overall, the aim is to assess the water status more holistically and understand which the real effects are caused by the sum of the chemical substances present in the aquatic environment (including emerging pollutants /other substances of interest, metabolites and transformation products). The assessment of chemical status should be improved and linked with ecological status where relevant. Hundreds of different substances may co-occur, and even if most are present at very small concentrations they could exert a toxic effect on aquatic organisms (Carvalho et al. 2014) exposed for their entire life cycle and indirectly on human health (via food and drinking water consumption). Some of the substances in the current list of Priority Substances and in the first Watch List are considered in groups (e.g. brominated diphenylethers, neonicotinoid insecticides), but the overall approach to chemical pollution is otherwise based on the regulation of single substances. It has become increasingly clear that the risks from the vast number of chemical substances present in the environment cannot be adequately controlled on this basis. The Commission acknowledges the need to consider the potential toxic effects of mixtures of chemicals (EC COM(2012)252, 7th EAP). The challenge is to find a way of capturing a true picture of the chemical status of water bodies based on standards and methods that assess the presence of an adequate range of representative chemical effect types or modes of action (MoA), for example. The knowledge on the MoA is an important driver for linking exposure to chemicals to their effects in the aquatic environment, and therefore for development and application of the scientific methodologies for the assessment of combined effects of chemicals - the effect-based methods (EBM). The EBM, including biomarkers and bioassays, can target different levels of biological organisation in the aquatic environment, such as individual and/or sub-organism, community, and population levels (Carvalho et al. 2014, Ann-Sofie Wernersson et al. 2014). It is however much less clear how these EBM can be used to capture (predictively) the indirect effects that might occur in humans following long-term chronic exposure to pollutants via the aquatic environment. The use of effect-based monitoring approaches, complementary to chemical analysis, could allow assessing chemical status more holistically (rather than with a limited but ever-growing list of individual substances). The use of the EBM offers also the advantage of overcoming analytical difficulties (Kunz et al. 2015) and reducing monitoring costs by screening. To become a credible complement to chemical monitoring information, however, a better understanding of the capabilities and gaps of available EBM is needed. This report, based on a comprehensive literature study, reviews the current PS list and other substances of interest, considering their MoA(s). The review of data from the open sources clearly identified few groups of toxicological endpoints, with the majority driven by non-specific mechanisms (e.g. oxidative stress, activation of metabolizing / detoxifying pathways, histopathology, and others), and few groups with more specific biochemical / physiological pathways (photosynthesis inhibition, acetylcholinesterase inhibition, presence of PAHs metabolites, expression of metallothioneins). The majority of current PS and other substances of interest can be grouped, based on few common toxicological endpoints, and biomarkers are available for determining the concentrations and/or effects of some groups of substances. The identified biomarkers of effect seem to be however in general not very specific. There is clearly no “one size fits all” bioassay / EBM that could provide the toxicological potency of every PS and other substances of interest and their mixture toward all aquatic organisms in all water bodies, but rather a battery of bioassays that should be selected as “fit for purpose”. In addition, the present report allowed identification of uncertainty and inconsistency in observations, and thus identified areas where future investigations can be best directed. The present knowledge about MoA(s) remains limited, especially for the emerging substances of concern, such as pyrethroids and neonicotinoides.JRC.D.2-Water and Marine Resource

Increased seroreactivity to proinsulin and homologous mycobacterial peptides in latent autoimmune diabetes in adults

Latent Autoimmune Diabetes in Adults (LADA) is a slowly progressing form of immune-mediated diabetes that combines phenotypical features of type 2 diabetes (T2D) with the presence of islet cell antigens detected in type 1 diabetes (T1D). Heterogeneous clinical picture have led to the classification of patients based on the levels of antibodies against glutamic acid decarboxylase 65 (GADA) that correlate with clinical phenotypes closer to T1D or T2D when GADA titers are high or low, respectively. To date, LADA etiology remains elusive despite numerous studies investigating on genetic predisposition and environmental risk factors. To our knowledge, this is the first study aimed at evaluation of a putative role played by Mycobacterium avium subsp. paratuberculosis (MAP) as an infective agent in LADA pathogenesis. MAP is known to cause chronic enteritis in ruminants and has been associated with autoimmune disorders in humans. We analyzed seroreactivity of 223 Sardinian LADA subjects and 182 healthy volunteers against MAP-derived peptides and their human homologs of proinsulin and zinc transporter 8 protein. A significantly elevated positivity for MAP/proinsulin was detected among patients, with the highest prevalence in the 32-41-year-old T1D-like LADA subgroup, supporting our hypothesis of a possible MAP contribution in the development of autoimmunity

Type 1 Diabetes at-risk children highly recognize <i>Mycobacterium avium</i> subspecies <i>paratuberculosis</i> epitopes homologous to human Znt8 and Proinsulin

Mycobacterium avium subspecies paratuberculosis (MAP) has been previously associated to T1D as a putative environmental agent triggering or accelerating the disease in Sardinian and Italian populations. Our aim was to investigate the role of MAP in T1D development by evaluating levels of antibodies directed against MAP epitopes and their human homologs corresponding to ZnT8 and proinsulin (PI) in 54 T1D at-risk children from mainland Italy and 42 healthy controls (HCs). A higher prevalence was detected for MAP/ZnT8 pairs (62,96% T1D vs. 7,14% HCs; p &lt; 0.0001) compared to MAP/PI epitopes (22,22% T1D vs. 9,52% HCs) and decreasing trends were observed upon time-point analyses for most peptides. Similarly, classical ZnT8 Abs and GADA decreased in a time-dependent manner, whereas IAA titers increased by 12%. Responses in 0–9 year-old children were stronger than in 10–18 age group (75% vs. 69,1%; p &lt; 0.04). Younger age, female sex and concomitant autoimmune disorders contributed to a stronger seroreactivity suggesting a possible implication of MAP in multiple autoimmune syndrome. Cross-reactivity of the homologous epitopes was reflected by a high correlation coefficient (r2 &gt; 0.8) and a pairwise overlap of positivity (&gt;83% for MAP/ZnT8)

Testing comparability of existing and innovative bioassays for water quality assessment

The JRC led a consortium of seventeen research Institutes from eleven countries in EU and associated countries to evaluate the suitability of the current paradigm in environmental risk assessment that considers the risk of single chemicals for assessing water quality. Combined effects of chemical mixtures of concern were measured on different aquatic organisms and different levels of biological organisation using existing and innovative bioassays. Aquatic organisms in most European surface waters were exposed to many chemical pollutants simultaneously. However, the current paradigm in water quality assessment under the Water Framework Directive (WFD) still considers the effects of single substances instead of evaluating the combined action of environmentally relevant mixtures.The potential effects of combinations of chemicals are equally relevant to the risk assessment of consumer products and of drinking water to humans. In this EU-wide exercise, we could show that exposure to mixtures of dissimilarity acting substances at concentrations considered environmentally acceptable can exert significant effects on the biota. Therefore, chemical monitoring of a few substances may be insufficient to assess the quality status of water impacted by complex anthropogenic mixtures. The study highlighted an urgent need to revise methods and paradigms used to assess the safety of chemicals to the environment. Bioassays as part of a multi-tier approach to water quality monitoring can fill the gap between chemical and ecological assessments for a more holistic characterisation of water quality. Considering the upcoming revision of the WFD in 2019, it is timely to introduce the issue of risk posed by mixtures of pollutants into the discussion table and find innovative ways to assess water quality in a more holistic way than the mere assessment of biological and chemical indicators.JRC.D.2-Water and Marine Resource

Seroreactivity against specific L5P antigen from <i>Mycobacterium avium</i> subsp. <i>paratuberculosis</i> in children at risk for T1D

Aims/Hypothesis: Although numerous environmental agents have been investigated over the years as possible triggers of type 1 diabetes (T1D), its causes remain unclear. We have already demonstrated an increased prevalence of antibodies against peptides derived from Mycobacterium avuim subsp. paratuberculosis (MAP) homologous to human zinc transporter 8 protein (ZnT8) and proinsulin in Italian subjects at risk for or affected by T1D. In this study, we compared titers of the previously detected antibodies with seroreactivity to MAP lipopentapetide (L5P) that recently emerged as a strong immunogenic component able to specifically distinguish MAP from other mycobacteria. Methods: Plasma of 32 children and youth at risk for T1D including follow-up samples and 42 age-matched healthy controls (HC) recruited at the Tor Vergata University Hospital in Rome was analyzed by indirect ELISA for the presence of antibodies against MAP-derived epitopes MAP3865c133–141, MAP3865c125-133, MAP2404c70-85 and MAP1,4αgbp157-173 along with their ZnT8 and proinsulin homologs. The data were analyzed through two-tailed Mann-Whitney U test and relation between variables was determined by principal component analysis. Results: Responses to L5P were not detectable in subjects whose initial seroreactivity to MAP peptides and their human homologs was lost in follow-up samples, whereas anti-L5P antibodies appeared constantly in individuals with a stable immunity against MAP antigens. The overall coincidence in positivity to L5P and the four MAP epitopes both in children at risk for T1D and HC exceeded 90%. Conclusions: MAP-derived homologs may cross-react with ZnT8 and proinsulin peptides inducing immune responses at a young age in subjects predisposed for T1D. Thus, L5P may have a diagnostic value to immediately indicate the presence of anti-MAP seroreactivity when evaluation of a more complex antibody status is not required. Almost complete coincidence in responses to both types of antigens lends support to the involvement of MAP in T1D

Rheumatoid arthritis patient antibodies highly recognize IL-2 in the immune response pathway involving IRF5 and EBV antigens

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by a progressive joint damage due to largely unknown environmental factors acting in concert with risk alleles conferring genetic susceptibility. A major role has been attributed to viral infections that include past contacts with Epstein-Barr virus (EBV) and, more recently, to non-protein coding sequences of human endogenous retrovirus K (HERV-K) integrated in the human genome. Molecular mimicry between viral and self proteins is supposed to cause the loss of immune tolerance in predisposed hosts. There are evidences that anti-IL-2 antibodies (Abs) are present in subjects affected by autoimmune diseases and may be responsible for alterations in regulatory T cell responses. In this study, we evaluated the levels of Abs against IL-2, viral epitopes and interferon regulatory factor 5 (IRF5) in 140 RA patients and 137 healthy controls (HCs). Ab reactivity reached the highest levels for IRF5, EBV and IL-2 (56%, 44% and 39%, respectively) in RA with significantly lower values among HCs (7–9%, p &lt;  0.0001), which suggests a possible cross-reaction between IRF5/EBV homologous antigens and shifts in T cell balance disrupted by anti-IL-2 Abs