Heating and microwave assisted SPPS of C-terminal acid peptides on trityl resin: the truth behind the yield

Despite correct purity of crude peptides prepared on trityl resin by Fmoc/tBu microwave assisted solid phase peptide synthesis, surprisingly, lower yields than those expected were obtained while preparing C-terminal acid peptides. This could be explained by cyclization/cleavage through diketopiperazine formation during the second amino acid deprotection and third amino acid coupling. However, we provide here evidence that this is not the case and that this yield loss was due to high temperature promoted hydrolysis of the 2-chlorotrityl ester, yielding premature cleavage of the C-terminal acid peptides.     

Barth syndrome: Cellular compensation of mitochondrial dysfunction and apoptosis inhibition due to changes in cardiolipin remodeling linked to tafazzin (TAZ) gene mutation

Cardiolipin is a mitochondrion-specific phospholipid that stabilizes the assembly of respiratory chain complexes, favoring full-yield operation. It also mediates key steps in apoptosis. In Barth syndrome, an X chromosome-linked cardiomyopathy caused by tafazzin mutations, cardiolipins display acyl chain modifications and are present at abnormally low concentrations, whereas monolysocardiolipin accumulates. Using immortalized lymphoblasts from Barth syndrome patients, we showed that the production of abnormal cardiolipin led to mitochondrial alterations. Indeed, the lack of normal cardiolipin led to changes in electron transport chain stability, resulting in cellular defects. We found a destabilization of the supercomplex (respirasome) I + III₂ + IV_n but also decreased amounts of individual complexes I and IV and supercomplexes I + III and III + IV. No changes were observed in the amounts of individual complex III and complex II. We also found decreased levels of complex V. This complex is not part of the supercomplex suggesting that cardiolipin is required not only for the association/stabilization of the complexes into supercomplexes but also for the modulation of the amount of individual respiratory chain complexes. However, these alterations were compensated by an increase in mitochondrial mass, as demonstrated by electron microscopy and measurements of citrate synthase activity. We suggest that this compensatory increase in mitochondrial content prevents a decrease in mitochondrial respiration and ATP synthesis in the cells. We also show, by extensive flow cytometry analysis, that the type II apoptosis pathway was blocked at the mitochondrial level and that the mitochondria of patients with Barth syndrome cannot bind active caspase-8. Signal transduction is thus blocked before any mitochondrial event can occur. Remarkably, basal levels of superoxide anion production were slightly higher in patients' cells than in control cells as previously evidenced via an increased protein carbonylation in the taz1Δ mutant in the yeast. This may be deleterious to cells in the long term. The consequences of mitochondrial dysfunction and alterations to apoptosis signal transduction are considered in light of the potential for the development of future treatments.     

Colonic Inflammation in Mice Is Improved by Cigarette Smoke through iNKT Cells Recruitment

Cigarette smoke (CS) protects against intestinal inflammation during ulcerative colitis. Immunoregulatory mechanisms sustaining this effect remain unknown. The aim of this study was to assess the effects of CS on experimental colitis and to characterize the intestinal inflammatory response at the cellular and molecular levels. Using the InExpose® System, a smoking device accurately reproducing human smoking habit, we pre-exposed C57BL/6 mice for 2 weeks to CS, and then we induced colitis by administration of dextran sodium sulfate (DSS). This system allowed us to demonstrate that CS exposure improved colonic inflammation (significant decrease in clinical score, body weight loss and weight/length colonic ratio). This improvement was associated with a significant decrease in colonic proinflammatory Th1/Th17 cytokine expression, as compared to unexposed mice (TNF (p = 0.0169), IFNγ (p<0.0001), and IL-17 (p = 0.0008)). Smoke exposure also induced an increased expression of IL-10 mRNA (p = 0.0035) and a marked recruitment of iNKT (invariant Natural Killer T; CD45+ TCRβ+ CD1d tetramer+) cells in the colon of DSS-untreated mice. Demonstration of the role of iNKT cells in CS-dependent colitis improvement was performed using two different strains of NKT cells deficient mice. Indeed, in Jα18KO and CD1dKO animals, CS exposure failed to induce significant regulation of DSS-induced colitis both at the clinical and molecular levels. Thus, our study demonstrates that iNKT cells are pivotal actors in the CS-dependent protection of the colon. These results highlight the role of intestinal iNKT lymphocytes and their responsiveness to environmental stimuli. Targeting iNKT cells would represent a new therapeutic way for inflammatory bowel diseases.     

Intra-Seasonal Flexibility in Avian Metabolic Performance Highlights the Uncoupling of Basal Metabolic Rate and Thermogenic Capacity

Stochastic winter weather events are predicted to increase in occurrence and amplitude at northern latitudes and organisms are expected to cope through phenotypic flexibility. Small avian species wintering in these environments show acclimatization where basal metabolic rate (BMR) and maximal thermogenic capacity (M_SUM) are typically elevated. However, little is known on intra-seasonal variation in metabolic performance and on how population trends truly reflect individual flexibility. Here we report intra-seasonal variation in metabolic parameters measured at the population and individual levels in black-capped chickadees ( Poecile atricapillus ). Results confirmed that population patterns indeed reflect flexibility at the individual level. They showed the expected increase in BMR (6%) and M_SUM (34%) in winter relative to summer but also, and most importantly, that these parameters changed differently through time. BMR began its seasonal increase in November, while M_SUM had already achieved more than 20% of its inter-seasonal increase by October, and declined to its starting level by March, while M_SUM remained high. Although both parameters co-vary on a yearly scale, this mismatch in the timing of variation in winter BMR and M_SUM likely reflects different constraints acting on different physiological components and therefore suggests a lack of functional link between these parameters.     

Novel extracellular and nuclear caspase-1 and inflammasomes propagate inflammation and regulate gene expression: a comprehensive database mining study

Background

Caspase-1 is present in the cytosol as an inactive zymogen and requires the protein complexes named “inflammasomes” for proteolytic activation. However, it remains unclear whether the proteolytic activity of caspase-1 is confined only to the cytosol where inflammasomes are assembled to convert inactive pro-caspase-1 to active caspase-1.

Methods

We conducted meticulous data analysis method?s on proteomic, protein interaction, protein intracellular localization, and gene expressions of 114 experimentally identified caspase-1 substrates and 38 caspase-1 interaction proteins in normal physiological conditions and in various pathologies.

Results

We made the following important findings: (1) Caspase-1 substrates and interaction proteins are localized in various intracellular organelles including nucleus and secreted extracellularly; (2) Caspase-1 may get activated in situ in the nucleus in response to intra-nuclear danger signals; (3) Caspase-1 cleaves its substrates in exocytotic secretory pathways including exosomes to propagate inflammation to neighboring and remote cells; (4) Most of caspase-1 substrates are upregulated in coronary artery disease regardless of their subcellular localization but the majority of metabolic diseases cause no significant expression changes in caspase-1 nuclear substrates; and (5) In coronary artery disease, majority of upregulated caspase-1 extracellular substrate-related pathways are involved in induction of inflammation; and in contrast, upregulated caspase-1 nuclear substrate-related pathways are more involved in regulating cell death and chromatin regulation.

Conclusions

Our identification of novel caspase-1 trafficking sites, nuclear and extracellular inflammasomes, and extracellular caspase-1-based inflammation propagation model provides a list of targets for the future development of new therapeutics to treat cardiovascular diseases, inflammatory diseases, and inflammatory cancers.

     

Risk of Advanced Neoplasia in First-Degree Relatives with Colorectal Cancer: A Large Multicenter Cross-Sectional Study

BackgroundFirst-degree relatives (FDR) of patients with colorectal cancer have a higher risk of developing colorectal cancer than the general population. For this reason, screening guidelines recommend colonoscopy every 5 or 10 y, starting at the age of 40, depending on whether colorectal cancer in the index-case is diagnosed at <60 or ≥60 y, respectively. However, studies on the risk of neoplastic lesions are inconclusive. The aim of this study was to determine the risk of advanced neoplasia (three or more non-advanced adenomas, advanced adenoma, or invasive cancer) in FDR of patients with colorectal cancer compared to average-risk individuals (i.e., asymptomatic adults 50 to 69 y of age with no family history of colorectal cancer).ConclusionsIndividuals having two FDR with colorectal cancer showed an increased risk of advanced neoplasia compared to those with average-risk for colorectal cancer. Men had over 2-fold higher risk of advanced neoplasia than women, independent of family history. These data suggest that screening colonoscopy guidelines should be revised in the familial-risk population.     

The Domestication Syndrome in Phoenix dactylifera Seeds: Toward the Identification of Wild Date Palm Populations

Investigating crop origins is a priority to understand the evolution of plants under domestication, develop strategies for conservation and valorization of agrobiodiversity and acquire fundamental knowledge for cultivar improvement. The date palm (Phoenix dactylifera L.) belongs to the genus Phoenix, which comprises 14 species morphologically very close, sometimes hardly distinguishable. It has been cultivated for millennia in the Middle East and in North Africa and constitutes the keystone of oasis agriculture. Yet, its origins remain poorly understood as no wild populations are identified. Uncultivated populations have been described but they might represent feral, i.e. formerly cultivated, abandoned forms rather than truly wild populations. In this context, this study based on morphometrics applied to 1625 Phoenix seeds aims to (1) differentiate Phoenix species and (2) depict the domestication syndrome observed in cultivated date palm seeds using other Phoenix species as a “wild” reference. This will help discriminate truly wild from feral forms, thus providing new insights into the evolutionary history of this species. Seed size was evaluated using four parameters: length, width, thickness and dorsal view surface. Seed shape was quantified using outline analyses based on the Elliptic Fourier Transform method. The size and shape of seeds allowed an accurate differentiation of Phoenix species. The cultivated date palm shows distinctive size and shape features, compared to other Phoenix species: seeds are longer and elongated. This morphological shift may be interpreted as a domestication syndrome, resulting from the long-term history of cultivation, selection and human-mediated dispersion. Based on seed attributes, some uncultivated date palms from Oman may be identified as wild. This opens new prospects regarding the possible existence and characterization of relict wild populations and consequently for the understanding of the date palm origins. Finally, we here describe a pipeline for the identification of the domestication syndrome in seeds that could be used in other crops.     

Identification of Tenrec ecaudatus,a Wild Mammal Introduced to Mayotte Island,as a Reservoir of the Newly Identified Human Pathogenic Leptospira mayottensis

Leptospirosis is a bacterial zoonosis of major concern on tropical islands. Human populations on western Indian Ocean islands are strongly affected by the disease although each archipelago shows contrasting epidemiology. For instance, Mayotte, part of the Comoros Archipelago, differs from the other neighbouring islands by a high diversity of Leptospira species infecting humans that includes Leptospira mayottensis, a species thought to be unique to this island. Using bacterial culture, molecular detection and typing, the present study explored the wild and domestic local mammalian fauna for renal carriage of leptospires and addressed the genetic relationships of the infecting strains with local isolates obtained from acute human cases and with Leptospira strains hosted by mammal species endemic to nearby Madagascar. Tenrec (Tenrec ecaudatus, Family Tenrecidae), a terrestrial mammal introduced from Madagascar, is identified as a reservoir of L. mayottensis. All isolated L. mayottensis sequence types form a monophyletic clade that includes Leptospira strains infecting humans and tenrecs on Mayotte, as well as two other Malagasy endemic tenrecid species of the genus Microgale. The lower diversity of L. mayottensis in tenrecs from Mayotte, compared to that occurring in Madagascar, suggests that L. mayottensis has indeed a Malagasy origin. This study also showed that introduced rats (Rattus rattus) and dogs are probably the main reservoirs of Leptospira borgpetersenii and Leptospira kirschneri, both bacteria being prevalent in local clinical cases. Data emphasize the epidemiological link between the two neighbouring islands and the role of introduced small mammals in shaping the local epidemiology of leptospirosis.     

Etiology and Epidemiology of Diarrhea in Hospitalized Children from Low Income Country: A Matched Case-Control Study in Central African Republic

Background

In Sub-Saharan Africa, infectious diarrhea is a major cause of morbidity and mortality. A case-control study was conducted to identify the etiology of diarrhea and to describe its main epidemiologic risk factors among hospitalized children under five years old in Bangui, Central African Republic.

Methods

All consecutive children under five years old hospitalized for diarrhea in the Pediatric Complex of Bangui for whom a parent’s written consent was provided were included. Controls matched by age, sex and neighborhood of residence of each case were included. For both cases and controls, demographic, socio-economic and anthropometric data were recorded. Stool samples were collected to identify enteropathogens at enrollment. Clinical examination data and blood samples were collected only for cases.

Results

A total of 333 cases and 333 controls was recruited between December 2011 and November 2013. The mean age of cases was 12.9 months, and 56% were male. The mean delay between the onset of first symptoms and hospital admission was 3.7 days. Blood was detected in 5% of stool samples from cases. Cases were significantly more severely or moderately malnourished than controls. One of the sought-for pathogens was identified in 78% and 40% of cases and controls, respectively. Most attributable cases of hospitalized diarrhea were due to rotavirus, with an attributable fraction of 39%. Four other pathogens were associated with hospitalized diarrhea: Shigella/EIEC, Cryptosporidium parvum/hominis, astrovirus and norovirus with attributable fraction of 9%, 10%, 7% and 7% respectively. Giardia intestinalis was found in more controls than cases, with a protective fraction of 6%.

Conclusions

Rotavirus, norovirus, astrovirus, Shigella/EIEC, Cryptosporidium parvum/hominis were found to be positively associated with severe diarrhea: while Giardia intestinalis was found negatively associated. Most attributable episodes of severe diarrhea were associated with rotavirus, highlighting the urgent need to introduce the rotavirus vaccine within the CAR’s Expanded Program on Immunization. The development of new medicines, vaccines and rapid diagnostic tests that can be conducted at the bedside should be high priority for low-resource countries.     

Space Use and Movement Patterns in a Semi-Free-Ranging Herd of European Bison (Bison bonasus)

The successful reintroduction and restocking of the European Bison demands a reliable knowledge of the biology of this species. Yet little is known to date about the European bison, and empirical data remains insufficient to set up a reliable plan ensuring the reintroduction, maintenance and survival of populations in habitats that have been largely modified by human activity. Studies of the ecology, social behaviour and management of bison are therefore crucial to the conservation of this species and its cohabitation with humans. To meet these challenges, we focused on movement patterns and space use in a semi-free-ranging herd of European bison living in the Réserve Biologique des Monts-d’Azur (France). Bison spend over 80% of their time foraging and resting; foraging mainly occurs around the artificial feeding sites (i.e., hay racks) or in meadows. The time of day and the presence of snow have no influence on the time budget allocated to each activity. Animals, however, spend more time at the food racks in winter. Bison also spend most of their time in small groups of individuals, confirming the occurrence of both fission-fusion dynamics and sexual segregation in this species. Bison seem to follow a Lévy walk pattern of movement, which is probably related to the geographical distribution and size of food patches in the reserve. The conclusions of this study provide a better understanding of the sociality, life habits and habitat use of bison, and also describe how the provision of hay affects all these behaviours. These results could be useful in the development of tools to select the most suitable habitats for the reintroduction, management and conservation of bison populations.