Néandertal et le feu au Paléolithique moyen ancien. Tour d’horizon des traces de son utilisation dans le Nord de la France

This article aims at drawing up balance sheet of remains of fire use by the first Neanderthals of Northern France, during the second part of the Saalian (MIS 8 to 6). This overview reminds us the rarity of fire testimonies during Early Middle Palaeolithic (300–130 ky BP) on the scale of North-Western Europe. For Northern France, only the sites of Biache-Saint-Vaast and Therdonne present remains of combustion. At Biache-Saint-Vaast, it is not less than six levels, which present clues of combustion: burnt flint and faunal remains and sometimes charcoals. At Therdonne, besides burnt numerous flint and some rare faunal remains were brought to light during the excavation of level N3 several rich zones in organic residues and micro-charcoals. All the datas collected concerning the clues of combustion at Biache-Saint-Vaast and Therdonne is compiled, analyzed and interpreted. This approach permits to establish the fire use or its absence in saalian occupations of Neanderthals of Northern France and to discuss modalities of its use, particularly at Therdonne. To conclude, fire status and its implications in first Neanderthals occupations are briefly discussed.     

High-Throughput In Vivo Genotoxicity Testing: An Automated Readout System for the Somatic Mutation and Recombination Test (SMART)

Genotoxicity testing is an important component of toxicity assessment. As illustrated by the European registration, evaluation, authorization, and restriction of chemicals (REACH) directive, it concerns all the chemicals used in industry. The commonly used in vivo mammalian tests appear to be ill adapted to tackle the large compound sets involved, due to throughput, cost, and ethical issues. The somatic mutation and recombination test (SMART) represents a more scalable alternative, since it uses Drosophila, which develops faster and requires less infrastructure. Despite these advantages, the manual scoring of the hairs on Drosophila wings required for the SMART limits its usage. To overcome this limitation, we have developed an automated SMART readout. It consists of automated imaging, followed by an image analysis pipeline that measures individual wing genotoxicity scores. Finally, we have developed a wing score-based dose-dependency approach that can provide genotoxicity profiles. We have validated our method using 6 compounds, obtaining profiles almost identical to those obtained from manual measures, even for low-genotoxicity compounds such as urethane. The automated SMART, with its faster and more reliable readout, fulfills the need for a high-throughput in vivo test. The flexible imaging strategy we describe and the analysis tools we provide should facilitate the optimization and dissemination of our methods.     

Benefits of teaching medical students how to communicate with patients having serious illness: comparison of two approaches to experiential, skill-based, and self-reflective learning

Innovative approaches are needed to teach medical students effective and compassionate communication with seriously ill patients. We describe two such educational experiences in the Yale Medical School curriculum for third-year medical students: 1) Communicating Difficult News Workshop and 2) Ward-Based End-of-Life Care Assignment. These two programs address educational needs to teach important clinical communication and assessment skills to medical students that previously were not consistently or explicitly addressed in the curriculum. The two learning programs share a number of educational approaches driven by the learning objectives, the students' development, and clinical realities. Common educational features include: experiential learning, the Biopsychosocial Model, patient-centered communication, integration into clinical clerkships, structured skill-based learning, self-reflection, and self-care. These shared features - as well as some differences - are explored in this paper in order to illustrate key issues in designing and implementing medical student education in these areas.     

Molecular Characterisation of Chikungunya Virus Infections in Trinidad and Comparison of Clinical and Laboratory Features with Dengue and Other Acute Febrile Cases

Local transmission of Chikungunya virus (CHIKV) was first documented in Trinidad and Tobago (T&T) in July 2014 preceding a large epidemic. At initial presentation, it is difficult to distinguish chikungunya fever (CHIKF) from other acute undifferentiated febrile illnesses (AUFIs), including life-threatening dengue disease. We characterised and compared dengue virus (DENV) and CHIKV infections in 158 patients presenting with suspected dengue fever (DF) and CHIKF at a major hospital in T&T, and performed phylogenetic analyses on CHIKV genomic sequences recovered from 8 individuals. The characteristics of patients with and without PCR-confirmed CHIKV were compared using Pearson’s χ² and student’s t-tests, and adjusted odds ratios (aORs) and 95% confidence intervals (CIs) were determined using logistic regression. We then compared signs and symptoms of people with RT-qPCR-confirmed CHIKV and DENV infections using the Mann-Whitney U, Pearson’s χ² and Fisher’s exact tests. Among the 158 persons there were 8 (6%) RT-qPCR-confirmed DENV and 30 (22%) RT-qPCR-confirmed CHIKV infections. Phylogenetic analyses showed that the CHIKV strains belonged to the Asian genotype and were most closely related to a British Virgin Islands strain isolated at the beginning of the 2013/14 outbreak in the Americas. Compared to persons who were RT-qPCR-negative for CHIKV, RT-qPCR-positive individuals were significantly more likely to have joint pain (aOR: 4.52 [95% CI: 1.28–16.00]), less likely to be interviewed at a later stage of illness (days post onset of fever—aOR: 0.56 [0.40–0.78]) and had a lower white blood cell count (aOR: 0.83 [0.71–0.96]). Among the 38 patients with RT-qPCR-confirmed CHIKV or DENV, there were no significant differences in symptomatic presentation. However when individuals with serological evidence of recent DENV or CHIKV infection were included in the analyses, there were key differences in clinical presentation between CHIKF and other AUFIs including DF, which can be used to triage patients for appropriate care in the clinical setting.     

Characterization of a functionally expressed dipeptidyl aminopeptidase III from Drosophila melanogaster.

A Drosophila melanogaster cDNA clone (GH01916) encoding a putative 723-residue long (82 kDa) protein (CG 7415) and displaying 50% identity with mammalian cytosolic dipeptidyl aminopeptidase (DPP) III was functionally expressed in Schneider S2 cells. Immunocytochemical studies using anti-(rat liver DPP III) Ig indicated the expression of this putative DPP III at the outer cell membrane and into the cytosol of transfected cells. Two protein bands (82 and 86 kDa) were immunologically detected after PAGE and Western blot of cytosol or membrane prepared from transfected cells. Western blot analysis of partially purified D. melanogaster DPP III confirmed the overexpression of these two protein bands into the cytosol and on the membranes of transfected cells. Despite the identification of six potential glycosylation sites, PAGE showed that these protein bands were not shifted after deglycosylation experiments. The partially purified enzyme hydrolysed the insect myotropic neuropeptide proctolin (Arg-Tyr-Leu-Pro-Thr) at the Tyr-Leu bond (Km approximately 4 micro m). In addition, low concentration of the specific DPP III inhibitor tynorphin prevented proctolin degradation (IC50 = 0.62 +/- 0.15 micro m). These results constitute the first characterization of an evolutionarily conserved insect DPP III that is expressed as a cytosolic and a membrane peptidase involved in proctolin degradation.     

Three distinct roles of aquaporin-4 in brain function revealed by knockout mice

Aquaporin-4 (AQP4) is expressed in astrocytes throughout the central nervous system, particularly at the blood-brain and brain-cerebrospinal fluid barriers. Phenotype analysis of transgenic mice lacking AQP4 has provided compelling evidence for involvement of AQP4 in cerebral water balance, astrocyte migration, and neural signal transduction. AQP4-null mice have reduced brain swelling and improved neurological outcome in models of (cellular) cytotoxic cerebral edema including water intoxication, focal cerebral ischemia, and bacterial meningitis. However, brain swelling and clinical outcome are worse in AQP4-null mice in models of vasogenic (fluid leak) edema including cortical freeze-injury, brain tumor, brain abscess and hydrocephalus, probably due to impaired AQP4-dependent brain water clearance. AQP4 deficiency or knock-down slows astrocyte migration in response to a chemotactic stimulus in vitro, and AQP4 deletion impairs glial scar progression following injury in vivo. AQP4-null mice also manifest reduced sound- and light-evoked potentials, and increased threshold and prolonged duration of induced seizures. Impaired K+ reuptake by astrocytes in AQP4 deficiency may account for the neural signal transduction phenotype. Based on these findings, we propose modulation of AQP4 expression or function as a novel therapeutic strategy for a variety of cerebral disorders including stroke, tumor, infection, hydrocephalus, epilepsy, and traumatic brain injury.     

Quantification of protein interaction in living cells by two-photon spectral imaging with fluorescent protein fluorescence resonance energy transfer pair devoid of acceptor bleed-through

Fluorescence resonance energy transfer (FRET) between fluorescent proteins (FPs) is a powerful method to visualize and quantify protein-protein interaction in living cells. Unfortunately, the emission bleed-through of FPs limits the usage of this complex technique. To circumvent undesirable excitation of the acceptor fluorophore, using two-photon excitation, we searched for FRET pairs that show selective excitation of the donor but not of the acceptor fluorescent molecule. We found this property in the fluorescent cyan fluorescent protein (CFP)/yellow fluorescent protein (YFP) and YFP/mCherry FRET pairs and performed two-photon excited FRET spectral imaging to quantify protein interactions on the later pair that shows better spectral discrimination. Applying non-negative matrix factorization to unmix two-photon excited spectral imaging data, we were able to eliminate the donor bleed-through as well as the autofluorescence. As a result, we achieved FRET quantification by means of a single spectral acquisition, making the FRET approach not only easy and straightforward but also less prone to calculation artifacts. As an application of our approach, the intermolecular interaction of amyloid precursor protein and the adaptor protein Fe65 associated with Alzheimer's disease was quantified. We believe that the FRET approach using two-photon and fluorescent YFP/mCherry pair is a promising method to monitor protein interaction in living cells.     

High Content Phenotypic Cell-Based Visual Screen Identifies Mycobacterium tuberculosis Acyltrehalose-Containing Glycolipids Involved in Phagosome Remodeling

The ability of the tubercle bacillus to arrest phagosome maturation is considered one major mechanism that allows its survival within host macrophages. To identify mycobacterial genes involved in this process, we developed a high throughput phenotypic cell-based assay enabling individual sub-cellular analysis of over 11,000 Mycobacterium tuberculosis mutants. This very stringent assay makes use of fluorescent staining for intracellular acidic compartments, and automated confocal microscopy to quantitatively determine the intracellular localization of M. tuberculosis. We characterised the ten mutants that traffic most frequently into acidified compartments early after phagocytosis, suggesting that they had lost their ability to arrest phagosomal maturation. Molecular analysis of these mutants revealed mainly disruptions in genes involved in cell envelope biogenesis (fadD28), the ESX-1 secretion system (espL/Rv3880), molybdopterin biosynthesis (moaC1 and moaD1), as well as in genes from a novel locus, Rv1503c-Rv1506c. Most interestingly, the mutants in Rv1503c and Rv1506c were perturbed in the biosynthesis of acyltrehalose-containing glycolipids. Our results suggest that such glycolipids indeed play a critical role in the early intracellular fate of the tubercle bacillus. The unbiased approach developed here can be easily adapted for functional genomics study of intracellular pathogens, together with focused discovery of new anti-microbials.