Early queen infection shapes developmental dynamics and induces long-term disease protection in incipient ant colonies

Infections early in life can have enduring effects on an organism's development and immunity. In this study, we show that this equally applies to developing ‘superorganisms’––incipient social insect colonies. When we exposed newly mated Lasius niger ant queens to a low pathogen dose, their colonies grew more slowly than controls before winter, but reached similar sizes afterwards. Independent of exposure, queen hibernation survival improved when the ratio of pupae to workers was small. Queens that reared fewer pupae before worker emergence exhibited lower pathogen levels, indicating that high brood rearing efforts interfere with the ability of the queen's immune system to suppress pathogen proliferation. Early-life queen pathogen exposure also improved the immunocompetence of her worker offspring, as demonstrated by challenging the workers to the same pathogen a year later. Transgenerational transfer of the queen's pathogen experience to her workforce can hence durably reduce the disease susceptibility of the whole superorganism.     

Certolizumab pegol plus methotrexate 5-year results from the rheumatoid arthritis prevention of structural damage (RAPID) 2 randomized controlled trial and long-term extension in rheumatoid arthritis patients

IntroductionAs patients with rheumatoid arthritis (RA) receive treatment with anti-tumour necrosis factors over several years, it is important to evaluate their long-term safety and efficacy. The objective of this study was to examine the safety and benefits of certolizumab pegol (CZP)+methotrexate (MTX) treatment for almost 5 years in patients with RA.MethodsPatients who completed the 24-week Rheumatoid Arthritis Prevention of Structural Damage (RAPID) 2 randomized controlled trial (RCT; {"type":"clinical-trial","attrs":{"text":"NCT00160602","term_id":"NCT00160602"}}NCT00160602), or who were American College of Rheumatology (ACR) 20 non-responders at Week 16, entered the open-label extension (OLE; {"type":"clinical-trial","attrs":{"text":"NCT00160641","term_id":"NCT00160641"}}NCT00160641). After ≥6 months treatment with CZP 400 mg every two weeks (Q2W), dose was reduced to 200 mg Q2W, the approved maintenance dose. Safety data are presented from all patients who received ≥1 dose CZP (Safety population, n=612). Efficacy data are presented to Week 232 for the intent-to-treat (ITT, n=492) and Week 24 CZP RCT Completer (n=342) populations, and through 192 weeks of dose-reduction for the Dose-reduction population (patients whose CZP dose was reduced to 200 mg, n=369). Radiographic progression (modified total Sharp score change from RCT baseline >0.5) to Week 128 is reported for the Week 24 CZP Completers.ResultsIn the RCT, 619 patients were randomized to CZP+MTX (n=492) or placebo+MTX (n=127). Overall, 567 patients (91.6%) entered the OLE: 447 CZP and 120 placebo patients. Of all randomized patients, 358 (57.8%) were ongoing at Week 232. Annual drop-out rates during the first four years ranged from 8.4–15.0%. Event rates per 100 patient-years were 163.0 for adverse events (AEs) and 15.7 for serious AEs. Nineteen patients (3.1%) had fatal AEs (incidence rate=0.8). Clinical improvements in the RCT were maintained to Week 232 in the CZP Completers: mean Disease Activity Score 28 (Erythrocyte Sedimentation Rate) change from baseline was −3.4 and ACR20/50/70 responses 68.4%/47.1%/25.1% (non-responder imputation). Similar improvements observed in the ITT were maintained following dose-reduction. 73.2% of CZP Completers had no radiographic progression at Week 128.ConclusionsIn patients with active RA despite MTX therapy, CZP was well tolerated, with no new safety signals identified. CZP provided sustained improvements in clinical outcomes for almost 5 years.

Trial registration

ClinicalTrials.gov, {"type":"clinical-trial","attrs":{"text":"NCT00160602","term_id":"NCT00160602"}}NCT00160602 and {"type":"clinical-trial","attrs":{"text":"NCT00160641","term_id":"NCT00160641"}}NCT00160641. Registered 8 September 2005.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0767-2) contains supplementary material, which is available to authorized users.     

Serum calprotectin (S100A8/9): an independent predictor of ultrasound synovitis in patients with rheumatoid arthritis

IntroductionCalprotectin, a heterodimeric complex of S100A8/9 (MRP8/14), has been proposed as an important serum biomarker that reflects disease activity and structural joint damage in rheumatoid arthritis (RA). The objective of this cross-sectional study was to test the hypothesis that calprotectin is associated with clinical and ultrasound-determined disease activity in patients with RA.MethodsA total of 37 patients with RA (including 24 females, a mean disease duration of 20 months) underwent a clinical examination and 7-joint ultrasound score (German US-7) of the clinically dominant hand and foot to assess synovitis by grey-scale (GS) and synovial vascularity by power Doppler (PD) ultrasound using semiquantitative 0–3 grading. The levels of serum calprotectin and C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were determined at the time of the ultrasound assessment. We analysed the relationship between serum calprotectin level, traditional inflammatory markers, and ultrasound-determined synovitis.ResultsThe levels of serum calprotectin were significantly correlated with swollen joint count (r = 0.465, p < 0.005), DAS28-ESR (r = 0.430, p < 0.01), ESR (r = 0.370, p < 0.05) and, in particular, CRP (r = 0.629, p < 0.001). Calprotectin was significantly associated with GS (r = 0.359, p < 0.05) and PD synovitis scores (r = 0.497, p < 0.005). Using multivariate regression analysis, calprotectin, adjusted for age and sex, was a better predictor of PD synovitis score (R² = 0.765, p < 0.001) than CRP (R² = 0.496, p < 0.001).ConclusionsThe serum levels of calprotectin are significantly associated with clinical, laboratory and ultrasound assessments of RA disease activity. These results suggest that calprotectin might be superior to CRP for monitoring ultrasound-determined synovial inflammation in RA patients.     

FireProt: Energy- and Evolution-Based Computational Design of Thermostable Multiple-Point Mutants

There is great interest in increasing proteins’ stability to enhance their utility as biocatalysts, therapeutics, diagnostics and nanomaterials. Directed evolution is a powerful, but experimentally strenuous approach. Computational methods offer attractive alternatives. However, due to the limited reliability of predictions and potentially antagonistic effects of substitutions, only single-point mutations are usually predicted in silico, experimentally verified and then recombined in multiple-point mutants. Thus, substantial screening is still required. Here we present FireProt, a robust computational strategy for predicting highly stable multiple-point mutants that combines energy- and evolution-based approaches with smart filtering to identify additive stabilizing mutations. FireProt’s reliability and applicability was demonstrated by validating its predictions against 656 mutations from the ProTherm database. We demonstrate that thermostability of the model enzymes haloalkane dehalogenase DhaA and γ-hexachlorocyclohexane dehydrochlorinase LinA can be substantially increased (ΔT _m = 24°C and 21°C) by constructing and characterizing only a handful of multiple-point mutants. FireProt can be applied to any protein for which a tertiary structure and homologous sequences are available, and will facilitate the rapid development of robust proteins for biomedical and biotechnological applications.     

Evolutionarily Conserved 5’-3’ Exoribonuclease Xrn1 Accumulates at Plasma Membrane-Associated Eisosomes in Post-Diauxic Yeast

Regulation of gene expression on the level of translation and mRNA turnover is widely conserved evolutionarily. We have found that the main mRNA decay enzyme, exoribonuclease Xrn1, accumulates at the plasma membrane-associated eisosomes after glucose exhaustion in a culture of the yeast S. cerevisiae. Eisosomal localization of Xrn1 is not achieved in cells lacking the main component of eisosomes, Pil1, or Sur7, the protein accumulating at the membrane compartment of Can1 (MCC) - the eisosome-organized plasma membrane microdomain. In contrast to the conditions of diauxic shift, when Xrn1 accumulates in processing bodies (P-bodies), or acute heat stress, in which these cytosolic accumulations of Xrn1 associate with eIF3a/Rpg1-containing stress granules, Xrn1 is not accompanied by other mRNA-decay machinery components when it accumulates at eisosomes in post-diauxic cells. It is important that Xrn1 is released from eisosomes after addition of fermentable substrate. We suggest that this spatial segregation of Xrn1 from the rest of the mRNA-decay machinery reflects a general regulatory mechanism, in which the key enzyme is kept separate from the rest of mRNA decay factors in resting cells but ready for immediate use when fermentable nutrients emerge and appropriate metabolism reprogramming is required. In particular, the localization of Xrn1 to the eisosome, together with previously published data, accents the relevance of this plasma membrane-associated compartment as a multipotent regulatory site.     

GRACE Score among Six Risk Scoring Systems (CADILLAC,PAMI, TIMI,Dynamic TIMI,Zwolle) Demonstrated the Best Predictive Value for Prediction of Long-Term Mortality in Patients with ST-Elevation Myocardial Infarction

Aim

To compare the prognostic accuracy of six scoring models for up to three-year mortality and rates of hospitalisation due to acute decompensated heart failure (ADHF) in STEMI patients.

Methods and Results

A total of 593 patients treated with primary PCI were evaluated. Prospective follow-up of patients was ≥3 years. Thirty-day, one-year, two-year, and three-year mortality rates were 4.0%, 7.3%, 8.9%, and 10.6%, respectively. Six risk scores—the TIMI score and derived dynamic TIMI, CADILLAC, PAMI, Zwolle, and GRACE—showed a high predictive accuracy for six- and 12-month mortality with area under the receiver operating characteristic curve (AUC) values of 0.73–0.85. The best predictive values for long-term mortality were obtained by GRACE. The next best-performing scores were CADILLAC, Zwolle, and Dynamic TIMI. All risk scores had a lower prediction accuracy for repeat hospitalisation due to ADHF, except Zwolle with the discriminatory capacity for hospitalisation up to two years (AUC, 0.80–0.83).

Conclusions

All tested models showed a high predictive value for the estimation of one-year mortality, but GRACE appears to be the most suitable for the prediction for a longer follow-up period. The tested models exhibited an ability to predict the risk of ADHF, especially the Zwolle model.     

Biochemical,Histopathological and Morphological Profiling of a Rat Model of Early Immune Stimulation: Relation to Psychopathology

Perinatal immune challenge leads to neurodevelopmental dysfunction, permanent immune dysregulation and abnormal behaviour, which have been shown to have translational validity to findings in human neuropsychiatric disorders (e.g. schizophrenia, mood and anxiety disorders, autism, Parkinson’s disease and Alzheimer’s disease). The aim of this animal study was to elucidate the influence of early immune stimulation triggered by systemic postnatal lipopolysaccharide administration on biochemical, histopathological and morphological measures, which may be relevant to the neurobiology of human psychopathology. In the present study of adult male Wistar rats we examined the brain and plasma levels of monoamines (dopamine, serotonin), their metabolites, the levels of the main excitatory and inhibitory neurotransmitters glutamate and γ-aminobutyric acid and the levels of tryptophan and its metabolites from the kynurenine catabolic pathway. Further, we focused on histopathological and morphological markers related to pathogenesis of brain diseases - glial cell activation, neurodegeneration, hippocampal volume reduction and dopaminergic synthesis in the substantia nigra. Our results show that early immune stimulation in adult animals alters the levels of neurotransmitters and their metabolites, activates the kynurenine pathway of tryptophan metabolism and leads to astrogliosis, hippocampal volume reduction and a decrease of tyrosine hydroxylase immunoreactivity in the substantia nigra. These findings support the crucial pathophysiological role of early immune stimulation in the above mentioned neuropsychiatric disorders.     

Overaccumulation of γ-Glutamylcysteine in a Jasmonate-Hypersensitive Arabidopsis Mutant Causes Jasmonate-Dependent Growth Inhibition

Glutathione (GSH) is essential for many aspects of plant biology and is associated with jasmonate signaling in stress responses. We characterized an Arabidopsis (Arabidopsis thaliana) jasmonate-hypersensitive mutant (jah2) with seedling root growth 100-fold more sensitive to inhibition by the hormone jasmonyl-isoleucine than the wild type. Genetic mapping and genome sequencing determined that the mutation is in intron 6 of GLUTATHIONE SYNTHETASE2, encoding the enzyme that converts γ-glutamylcysteine (γ-EC) to GSH. The level of GSH in jah2 was 71% of the wild type, while the phytoalexin-deficient2-1 (pad2-1) mutant, defective in GSH1 and having only 27% of wild-type GSH level, was not jasmonate hypersensitive. Growth defects for jah2, but not pad2, were also seen in plants grown to maturity. Surprisingly, all phenotypes in the jah2 pad2-1 double mutant were weaker than in jah2. Quantification of γ-EC indicated these defects result from hyperaccumulation of this GSH precursor by 294- and 65-fold in jah2 and the double mutant, respectively. γ-EC reportedly partially substitutes for loss of GSH, but growth inhibition seen here was likely not due to an excess of total glutathione plus γ-EC because their sum in jah2 pad2-1 was only 16% greater than in the wild type. Further, the jah2 phenotypes were lost in a jasmonic acid biosynthesis mutant background, indicating the effect of γ-EC is mediated through jasmonate signaling and not as a direct result of perturbed redox status.Glutathione (GSH) is an essential thiol of most higher organisms, including plants. Primarily found in the reduced form, its roles in maintaining a reduced intracellular state are numerous and well characterized (Foyer and Noctor, 2011; Noctor et al., 2011). Additionally, GSH is involved in detoxifying reactive oxygen species, heavy metal detoxification through phytochelatins, elimination of xenobiotics, and signaling of plant development and stress responses (Rouhier et al., 2008).GSH is synthesized in two steps. The first links Cys to the γ-carboxyl group of Glu through an amide bond catalyzed by γ-glutamylcysteine (γ-EC) synthetase, encoded by the single gene GSH1 in Arabidopsis (Arabidopsis thaliana). Gly is then added by GSH synthetase (GSH-S), also encoded by a single gene (GSH2). GSH is typically present at millimolar levels in plants, and although γ-EC is normally present at only a few percent of this amount, there is evidence that γ-EC has redox activities in Arabidopsis (Pasternak et al., 2008).Insertional knockouts of GSH1 are embryo lethal, and rootmeristemless1, with only 5% of wild-type GSH level, lacks a root apical meristem due to cell cycle arrest (Vernoux et al., 2000; Cairns et al., 2006). Other mutants producing 25% to 50% of wild-type GSH levels grow normally but exhibit defects under various stress conditions. For example, phytoalexin-deficient2-1 (pad2-1) and cadmium sensitive2 mutants are susceptible to pathogens and hypersensitive to Cd, respectively, while regulator of axillary meristems1 causes elevated expression of ASCORBATE PEROXIDASE2 under non-photooxidative-stress conditions (Glazebrook and Ausubel, 1994; Cobbett et al., 1998; Ball et al., 2004).GSH2 null alleles (gsh2-1 and gsh2-2) are also lethal, although plants survive to the early seedling stage (Pasternak et al., 2008). Survival past the embryo stage was attributed to partial complementation of GSH activity by γ-EC, which accumulates to excessive levels in gsh2-1, and the mutant is partially rescued by GSH supplementation. Missense and nonsense GSH2 alleles of membrane trafficking mutants (gsh2-3–gsh2-5) disrupt endoplasmic reticulum (ER) organization and also arrest growth in early seedling development, while a weaker allele (gsh2-6) reached maturity but was smaller than the wild type (Au et al., 2012). A screen for reduced response to Cd also yielded a viable missense mutant of GSH2 (nonresponse or reduced response to Cd2) with approximately 75% of the wild-type GSH level (Jobe et al., 2012).Plant oxidative stress responses involve both redox signaling through GSH and jasmonate hormonal signaling, and gene expression studies have clearly linked these two signaling systems. GSH biosynthesis and metabolism genes are induced by jasmonate, while manipulating GSH level or redox status in various mutants alters expression of genes for jasmonate biosynthesis and signaling (Xiang and Oliver, 1998; Mhamdi et al., 2010; Han et al., 2013). GSH and jasmonate are also associated with protective glucosinolate production in response to insect feeding (Noctor et al., 2011). For example, pad2-1 is deficient in glucosinolates and more susceptible to insects, while several studies have shown jasmonate induces glucosinolates (Brader et al., 2001; Mikkelsen et al., 2003; Sasaki-Sekimoto et al., 2005; Schlaeppi et al., 2008). Liu et al. (2010) isolated jasmonic acid hypersensitive1 (jah1), an Arabidopsis mutant with greater inhibition of root growth than the wild type in the presence of jasmonic acid (JA). The affected gene encodes a cytochrome P450 (CYP82C3) involved in indole glucosinolate production, and this mutant was more susceptible to Botrytis cinerea.The basic mechanism of jasmonate signal transduction and some of the downstream responses emanating from it are now well understood (Browse, 2009; Wasternack and Hause, 2013). However, the mechanisms by which jasmonate and GSH coordinate their activities to mediate oxidative stress and other responses are not known. This study characterized, to our knowledge, a new jasmonate-hypersensitive mutant that accumulates excess γ-EC due to a defect in GSH2, but GSH is only modestly reduced. Results show that elevated γ-EC is deleterious to plant growth through a jasmonate-dependent mechanism.