Multilayered genetic and omics dissection of mitochondrial activity in a mouse reference population

The manner by which genotype and environment affect complex phenotypes is one of the fundamental questions in biology. In this study, we quantified the transcriptome--a subset of the metabolome--and, using targeted proteomics, quantified a subset of the liver proteome from 40 strains of the BXD mouse genetic reference population on two diverse diets. We discovered dozens of transcript, protein, and metabolite QTLs, several of which linked to metabolic phenotypes. Most prominently, Dhtkd1 was identified as a primary regulator of 2-aminoadipate, explaining variance in fasted glucose and diabetes status in both mice and humans. These integrated molecular profiles also allowed further characterization of complex pathways, particularly the mitochondrial unfolded protein response (UPR(mt)). UPR(mt) shows strikingly variant responses at the transcript and protein level that are remarkably conserved among C. elegans, mice, and humans. Overall, these examples demonstrate the value of an integrated multilayered omics approach to characterize complex metabolic phenotypes.

Toll-like receptor agonists synergize with CD40L to induce either proliferation or plasma cell differentiation of mouse B cells.

In a classical dogma, pathogens are sensed (via recognition of Pathogen Associated Molecular Patterns (PAMPs)) by innate immune cells that in turn activate adaptive immune cells. However, recent data showed that TLRs (Toll Like Receptors), the most characterized class of Pattern Recognition Receptors, are also expressed by adaptive immune B cells. B cells play an important role in protective immunity essentially by differentiating into antibody-secreting cells (ASC). This differentiation requires at least two signals: the recognition of an antigen by the B cell specific receptor (BCR) and a T cell co-stimulatory signal provided mainly by CD154/CD40L acting on CD40. In order to better understand interactions of innate and adaptive B cell stimulatory signals, we evaluated the outcome of combinations of TLRs, BCR and/or CD40 stimulation. For this purpose, mouse spleen B cells were activated with synthetic TLR agonists, recombinant mouse CD40L and agonist anti-BCR antibodies. As expected, TLR agonists induced mouse B cell proliferation and activation or differentiation into ASC. Interestingly, addition of CD40 signal to TLR agonists stimulated either B cell proliferation and activation (TLR3, TLR4, and TLR9) or differentiation into ASC (TLR1/2, TLR2/6, TLR4 and TLR7). Addition of a BCR signal to CD40L and either TLR3 or TLR9 agonists did not induce differentiation into ASC, which could be interpreted as an entrance into the memory pathway. In conclusion, our results suggest that PAMPs synergize with signals from adaptive immunity to regulate B lymphocyte fate during humoral immune response.

Aging effects on selective attention-related electroencephalographic patterns during face encoding.

Previous electrophysiological studies revealed that human faces elicit an early visual event-related potential (ERP) within the occipito-temporal cortex, the N170 component. Although face perception has been proposed to rely on automatic processing, the impact of selective attention on N170 remains controversial both in young and elderly individuals. Using early visual ERP and alpha power analysis, we assessed the influence of aging on selective attention to faces during delayed-recognition tasks for face and letter stimuli, examining 36 elderly and 20 young adults with preserved cognition. Face recognition performance worsened with age. Aging induced a latency delay of the N1 component for faces and letters, as well as of the face N170 component. Contrasting with letters, ignored faces elicited larger N1 and N170 components than attended faces in both age groups. This counterintuitive attention effect on face processing persisted when scenes replaced letters. In contrast with young, elderly subjects failed to suppress irrelevant letters when attending faces. Whereas attended stimuli induced a parietal alpha band desynchronization within 300-1000 ms post-stimulus with bilateral-to-right distribution for faces and left lateralization for letters, ignored and passively viewed stimuli elicited a central alpha synchronization larger on the right hemisphere. Aging delayed the latency of this alpha synchronization for both face and letter stimuli, and reduced its amplitude for ignored letters. These results suggest that due to their social relevance, human faces may cause paradoxical attention effects on early visual ERP components, but they still undergo classical top-down control as a function of endogenous selective attention. Aging does not affect the face bottom-up alerting mechanism but reduces the top-down suppression of distracting letters, possibly impinging upon face recognition, and more generally delays the top-down suppression of task-irrelevant information.

Healthy, wealthy, wise, and happy? An exploratory analysis of the interplay between aging and subjective well-being in low and middle income countries

In this paper, we address the relationship between age and several dimension of subjective well-being. Whilst literature generally finds a U-shaped age-profile in subjective well-being, this age-pattern might only hold after controlling for objective life circumstances. The observed U-shaped age-profile might further not generalize to other dimensions of well-being and might vary across countries and cultures. Our study examines the relationship between age and several dimensions of well-being as well as the effect of objective life circumstances using the WHO Study on Global AGEing and Adult Health (SAGE). Our results suggest a decreasing age profile in the raw data associated with evaluative well-being, while experienced well-being shows a rather flat or slightly increasing pattern. However, age per se is not a cause of a decline in evaluative well-being. The negative age-profile in evaluative well-being is mainly explained by changes in life circumstances associated with aging. Controlling for socio-demographic factors, we find higher levels of well-being for older persons relative to their middle-aged counterparts. In contrast, we find that changes in life circumstances have a much smaller effect on experienced well-being.

Selective tumor localization of radiolabeled anti-human melanoma monoclonal antibody fragment demonstrated in the nude mouse model.

Monoclonal antibodies (Mab) directed against distinct epitopes of the human 240 kD melanoma-associated antigen have been evaluated for their capacity to localize in human melanoma grafted into nude mice. A favorable tumor to normal tissue ratio of 13 was obtained with intact 131I-labeled MAb Me1-14. This ratio was further increased to 43 and 23 by the use of F(ab')2 and Fab fragments, respectively. The specificity of tumor localization was demonstrated by the simultaneous injection of F(ab')2 fragments from MAb Me1-14 and anti-CEA MAb 35, each labeled with a different iodine isotope, into nude mice grafted with a melanoma and colon carcinoma. The fragments from both MAb localized with perfect selectivity in their relevant tumor as shown by differential whole body scanning and by direct measurement of the two isotopes in tumors and normal tissues. These in vivo experimental results suggest that the F(ab')2 fragment from MAb Me1-14 is suitable for melanoma detection by immunoscintigraphy in patients.

Aging and the lateralization of oscillatory activities related to external and internal motor preparation

Selection of action may rely on external guidance or be motivated internally, engaging partially distinct cerebral networks. With age, there is an increased allocation of sensorimotor processing resources, accompanied by a reduced differentiation between the two networks of action selection. The present study examines the age effects on the motor-related oscillatory patterns related to the preparation of externally and internally guided movements. Thirty-two older and 30 younger adults underwent three delayed motor tasks with S1 as preparatory and S2 as imperative cue: Full, laterality instructed by S1 (external guidance); Free, laterality freely selected (internal guidance); None, laterality instructed by S2 (no preparation). Electroencephalogram (EEG) was recorded using 64 surface electrodes. Motor-Related Amplitude Asymmetries (MRAA), indexing the lateralization of oscillatory activities, were analyzed within the S1-S2 interval in the mu (9-12 Hz) and low beta (15-20 Hz) motor-related frequency bands. Reaction times to S2 were slower in older than younger subjects, and slower in the Free than in the Full condition in older subjects only. In the Full condition, there were significant mu MRAA in both age groups, and significant low beta MRAA only in older adults. The Free condition was associated with large mu MRAA in younger adults and limited low beta MRAA in older adults. In younger subjects, the lateralization of mu activity in both Full and Free conditions indicated effective external and internal motor preparation. In older subjects, external motor preparation was associated with lateralization of low beta in addition with mu activity, compatible with an increase of motor-related resources. In contrast, absence of mu and limited low beta lateralization in internal motor preparation was concomitant with reaction time slowing and suggested less efficient cerebral processes subtending free movement selection in older adults, indicating reduced capacity for internally driven action with age.

A CRTC1 deficient mouse line study : relevance to mood disorders

Summary Mood disorders are among the most prevalent, psychosocial^ debilitating, chronic and relapsing forms of psychiatric illnesses. Despite considerable advances in their characterization, the heterogeneous nature of susceptibility factors and patient's symptoms could account for the lack of totally effective and remissive treatment. The neurobiological hypothesis of mood disorders etiology has evolved since the monoamine and neurotrophin theories and current evidence is pointing toward their integration in a broader polygenic epistatic model resulting in defective neuroplasticity of circuitries involved in emotion processing. Consequently, the unraveling of molecular underpinning pathways involved in neuronal plasticity, commonly altered among mood disorder syndromes and symptoms, should shed light on their etiology and provide new drug target. The transcription factor CREB has been critically involved in the long-lasting forms of neuronal plasticity and in the regulation of several mood disorders susceptibility genes. In addition, altered CREB activity has been associated with mood disorders pathophysiology and pharmacotherapy. Interestingly, the newly-identified protein CREB-regulated transcription coactivator 1 (CRTC1) was shown by previous studies in the laboratory to be a neuroactivity- dependent cAMP and calcium sensor, a potent activator of CREB-dependent transcription and involved in neuroplasticity mechanisms associated with long-term synaptic potentiation. Furthermore, the major mood disorder susceptibility gene Bdnf was suggested to be transcriptional regulated by CRTC1. Therefore, we aimed to investigate a role for CRTC1 in mood disorders by generating and characterizing a Crtcl deficient mouse model at the behavioral and molecular levels. Interestingly, their comprehensive characterization revealed a behavioral profile mirroring several major symptoms comorbid in mood disorders, including altered social interactions, aggressive behaviors, obesity, psychomotor retardation, increased emotional response to stress, decreased sexual drive and depression-like behaviors. To investigate the molecular mechanisms underlying these pathological behaviors and the implication of CRTC1 in the regulation of CREB-regulated genes in vivo, we also quantified transcript levels of several relevant CREB-regulated susceptibility genes in brain structures involved in the pathophysiology of mood disorders. Strikingly, we found the underexpression of primary components of the neurotrophin system: Bdnf and its cognate receptor TrkB, a marked decrease in the Nr4a family of transcription factors, implicated in neuroplasticity and associated with dopamine-related disorders, as well as in several other relevant CREB regulated genes. Moreover, neurochemical analysis revealed that Crtcl null mice presented alteration in prefrontal cortical monoamine turnover as well as in hippocampal and accumbal serotonin levels, similarly associated with mood disorders etiology and pharmacotherapy. Together, the present thesis supports the involvement of CRTC1 pathway hypofunction in the pathogenesis of mood disorders and specifically in pathological aggression, obesity and depression-related behavior comorbidities. Ultimately, CRTC1 may represent an interesting antidepressant, antiaggressive or mood stabilizer drug target candidate through the modulation of major CREB regulated susceptibility genes. Les troubles de l'humeur comptent parmi les maladies psychiatriques les plus prévalentes, psychosocialement débilitantes, chroniques et avec le plus grand risque de rechute. Malgré de considérable avancées dans leur caractérisation, la nature hétérogène des facteurs de susceptibilité et des symptômes présentés par les patients, semble justifier l'absence de traitement entraînant une rémission complète de la maladie. L'hypothèse de l'étiologie neurobiologique des troubles de l'humeur a évolué depuis la théorie des monoamines et des neurotrophines. Actuellement, elle tend à les englober dans un modèle polygénique épistatique induisant une déficience de la neuroplasticité des circuits impliqué dans la régulation des émotions. Par conséquent, il apparaît particulièrement relevant de caractériser des voies moléculaires impliquées dans la plasticité neuronale, communément altérées parmi les différents syndromes et symptômes des maladies de l'humeur, afin d'améliorer leur compréhension ainsi que de proposer de nouvelles cibles thérapeutiques potentielles. Le facteur de transcription CREB a été de façon répétée et cohérente impliqué dans les mécanismes à long terme de la plasticité neuronale, ainsi que dans la régulation de plusieurs gènes de susceptibilité aux maladies de l'humeur. De plus, une altération dans l'activité de CREB a été impliqué dans leur étiologie et pharmacothérapie. De façon intéressante, des résultats préliminaires sur la protéine récemment découverte CREB-regulated transcription coactivator 1 (CRTC1) ont indiqué que son activation était dépendante de l'activité neuronale, qu'il était un senseur du calcium et de l'AMPc, ainsi qu'un coactivateur de CREB requis et puissant impliqué dans les mécanismes de plasticité neuronale associés à la potentialisation à long terme. En outre, des résultats ont suggéré que le gène majeur de susceptibilité Bdnf est régulé par CRTC1. Ainsi, notre objectif a été d'investiguer un rôle éventuel de CRTC1 dans les maladies de l'humeur en générant et caractérisant une lignée de souris déficiente pour Crtcl, tant au niveau comportemental que moléculaire. De façon intéressante, leur caractérisation détaillée a révélé un profil comportemental reflétant de nombreux aspects des maladies de l'humeur incluant une altération des interactions sociales, une agression pathologique, l'obésité, un retard psychomoteur, une réponse émotionnelle au stress accrue, une diminution de la motivation sexuelle, et des comportements reliés à la dépression. Afin d'investiguer les mécanismes moléculaires sous- jacents cette altération du comportement, ainsi que l'implication de CRTC1 dans l'expression des gènes régulés par CREB in vivo, nous avons quantifié les niveaux de transcrits de plusieurs gènes de susceptibilité régulés par CREB et impliqués dans la physiopathologie des maladies de l'humeur. Remarquablement, nous avons trouvé la sous-expression de composants primordiaux du système neurotrophique: Bdnf et son récepteur TrkB, une diminution majeure de la famille des facteurs de transcription Nr4a, impliqués dans la neuroplasticité et associés à des désordres liés à la dopamine, ainsi que de nombreux autres gènes relevants régulés par CREB. De plus, une analyse neurochimique a révélé que les souris déficientes pour Crtcî présentent une altération du turn-over des monoamines du cortex préfrontal ainsi que des niveaux hippocampaux et accumbaux de sérotonine, associés de façon similaire dans l'étiologie et la pharmacothérapie des maladies de l'humeur. Vue dans son ensemble, la présente thèse supporte l'implication d'une sous-régulation de la voie de CRTCI dans la pathogenèse des maladies de l'humeur ainsi que dans la comorbidité de l'agression pathologique, l'obésité et la dépression. En conclusion, CRTCI pourrait représenter une cible médicamenteuse intéressante aux propriétés antidépressante, antiagressive ou stabilisatrice de l'humeur au travers de la modulation de gènes de susceptibilité majeurs régulés par CREB.

Loss of ATF2 function leads to cranial motoneuron degeneration during embryonic mouse development.

The AP-1 family transcription factor ATF2 is essential for development and tissue maintenance in mammals. In particular, ATF2 is highly expressed and activated in the brain and previous studies using mouse knockouts have confirmed its requirement in the cerebellum as well as in vestibular sense organs. Here we present the analysis of the requirement for ATF2 in CNS development in mouse embryos, specifically in the brainstem. We discovered that neuron-specific inactivation of ATF2 leads to significant loss of motoneurons of the hypoglossal, abducens and facial nuclei. While the generation of ATF2 mutant motoneurons appears normal during early development, they undergo caspase-dependent and independent cell death during later embryonic and foetal stages. The loss of these motoneurons correlates with increased levels of stress activated MAP kinases, JNK and p38, as well as aberrant accumulation of phosphorylated neurofilament proteins, NF-H and NF-M, known substrates for these kinases. This, together with other neuropathological phenotypes, including aberrant vacuolisation and lipid accumulation, indicates that deficiency in ATF2 leads to neurodegeneration of subsets of somatic and visceral motoneurons of the brainstem. It also confirms that ATF2 has a critical role in limiting the activities of stress kinases JNK and p38 which are potent inducers of cell death in the CNS.

Radioimmunotherapy of human colon carcinoma by 131I-labelled monoclonal anti-CEA antibodies in a nude mouse model.

A mixture of 3 MAbs directed against 3 different CEA epitopes was radiolabelled with 131I and used for the treatment of a human colon carcinoma transplanted s.c. into nude mice. Intact MAbs and F(ab')2 fragments were mixed because it had been shown by autoradiography that these 2 antibody forms can penetrate into different areas of the tumor nodule. Ten days after transplantation of colon tumor T380 a single dose of 600 microCi of 131I MAbs was injected i.v. The tumor grafts were well established (as evidenced by exponential growth in untreated mice) and their size continued to increase up to 6 days after radiolabelled antibody injection. Tumor shrinking was then observed lasting for 4-12 weeks. In a control group injected with 600 microCi of 131I coupled to irrelevant monoclonal IgG, tumor growth was delayed, but no regression was observed. Tumors of mice injected with the corresponding amount of unlabelled antibodies grew like those of untreated mice. Based on measurements of the effective whole-body half-life of injected 131I, the mean radiation dose received by the animals was calculated to be 382 rads for the antibody group and 478 rads for the normal IgG controls. The genetically immunodeficient animals exhibited no increase in mortality, and only limited bone-marrow toxicity was observed. Direct measurement of radioactivity in mice dissected 1, 3 and 7 days after 131I-MAb injection showed that 25, 7.2 and 2.2% of injected dose were recovered per gram of tumor, the mean radiation dose delivered to the tumor being thus more than 5,000 rads. These experiments show that therapeutic doses of radioactivity can be selectively directed to human colon carcinoma by i.v. injection of 131I-labelled anti-CEA MAbs.

Retrovirus-induced target cell activation in the early phases of infection: the mouse mammary tumor virus model.

Mouse mammary tumor virus (MMTV) infects B lymphocytes and expresses a superantigen on the cell surface after integration of its reverse-transcribed genome. Superantigen-dependent B- and T-cell activation becomes detectable 2 to 3 days after infection. We show here that before this event, B cells undergo a polyclonal activation which does not involve massive proliferation. This first phase of B-cell activation is T cell independent. Moreover, during the first phase of activation, when only a small fraction of B cells is infected by MMTV(SW), viral DNA is detected only in activated B cells. Such a B-cell activation is also seen after injection of murine leukemia virus but not after injection of vaccinia virus, despite the very similar kinetics and intensity of the immune response. Since retroviruses require activated target cells to induce efficient infection, these data suggest that the early polyclonal retrovirus-induced target cell activation might play an important role in the establishment of retroviral infections.

The HSR on chromosome 1 of the house mouse, Mus domesticus: distribution and frequency in Switzerland.

A total of 357 house mice (Mus domesticus) from 83 localities uniformly distributed throughout Switzerland were screened for the presence of a homogenously staining region (HSR) on chromosome 1. Altogether 47 mice from 11 localities were HSR/+ or HSR/HSR. One sample of 11 individuals all had an HSR/HSR karyotype. Almost all mice with the variant were collected from the Rhone valley (HSR frequency: 61%) and Val Bregaglia (HSR frequency: 81%). For samples from most of the area of Switzerland, the HSR was absent. There was no strong association between the geographic distribution of the HSR and the areas of occurrence of metacentrics. However, at Chiggiogna the HSR was found on Rb (1.3). Possible explanations for the HSR polymorphism are discussed.

Sleep, metabolism and aging

Aging is a multidimensional process of physical, psychological, and social changes. Understanding how we sleep and how this dynamic process evolves across life span will help to identify normal developmental aspects of sleep over time and to create strategies to increase awareness of sleep disturbances and their early management. In normal sleepers from HypnoLaus cohort, we evaluated the effects of age and gender on both subjective and objective sleep measurements. Our results indicate that normal aging is not accompanied by sleep complaints, and when they exist suggest the presence of underlying comorbidities. Polysomnographic data revealed that slow wave sleep was more affected with age in men, and age affected differently NREM and REM spectral power densities. Both sleep structure and spectral analysis profiles may constitute standards to delineate pathological changes in sleep, both for aging women and men. Another important aspect in the management of sleep and its disorders is a detailed characterization of sleep-inducing medications. Gamma-hydroxybutyrate (GHB) is an inhibitory neurotransmitter derivative of GABA, but its mode of action and the range of effects are not well understood. Several properties, as growth hormone stimulation in humans and the development of weight loss in treated patients suggest an unexplored metabolic effect. In different experiments we assessed the effects of acute, short term and chronic GHB administration on central (cerebral cortex) and peripheral (liver) biochemical processes involved in the metabolism of the drug, as well as the effects of the drug on metabolism in C57BL/6J, GABAB knock-out and obese (ob/ob) mice. We showed that GHB treatment affects weight gain in C57BL/6J and GABAB knock-out mice. Metabolomic analysis indicated large central and peripheral metabolic changes induced by GHB with important relevance to its therapeutic use. -- Le vieillissement est un processus multidimensionnel accompagné par de multiples changements dans les domaines physique, psychologique et social. Comprendre comment nous dormons et comment ce processus dynamique évolue sur la durée de vie nous aidera à identifier les aspects normaux du développement du sommeil au fil du temps, et à créer des stratégies pour accroître la connaissance et compréhension des troubles du sommeil et leur prise en charge précoce. Chez les sujets normaux de la cohorte HypnoLaus nous avons évalué les effets de l'âge et du sexe sur les mesures subjectives et objectives du sommeil. Nos résultats indiquent que le vieillissement normal ne s'accompagne pas de troubles du sommeil, et quand ils existent ceux-ci suggèrent la présence de comorbidités sous-jacentes. Les données polysomnographiques ont révélé que le sommeil profond était plus affecté avec l'âge chez les hommes. De plus, nous avons montré comment l'âge modifie la composition spectrale du sommeil lent et paradoxal. La structure du sommeil et les profils d'analyse spectrale peuvent donc constituer des standards permettant de définir les changements pathologiques du sommeil chez les personnes âgées. Parmi les aspects importants de la gestion du sommeil et de ses troubles, la caractérisation détaillée des médicaments hypnotiques utilisés est essentielle. L'acide gamma-hydroxybutyrique (GHB) est un acide gras à courte chaîne dérivé du GABA, principal neurotransmetteur inhibiteur du cerveau, mais son mode d'action et tous ses effets sont toujours largement méconnus. Plusieurs propriétés, comme la stimulation de la sécrétion de l'hormone de croissance chez l'homme et le développement d'une perte de poids chez les patients traités suggèrent un effet métabolique inexploré. Dans différentes expériences, nous avons évalué les effets d'une exposition aiguë, à court terme et chronique de GHB sur les processus biochimiques centraux (cortex cérébral) et périphériques (foie) impliqués dans le métabolisme du médicament. Nous avons aussi évalué les effets du médicament sur le métabolisme des souris C57BL/6J, GABAB KO et obèses (ob/ob). Nos résultats ont montré que le GHB diminue le gain de poids chez les souris C57BL/6J et GABAB KO. L'analyse métabolomique a indiqué des changements importants induits par GHB au niveau central et périphérique, et ces effets sont importants pour son utilisation thérapeutique.

Effects of mineralocorticoid and K+ concentration on K+ secretion and ROMK channel expression in a mouse cortical collecting duct cell line.

The cortical collecting duct (CCD) plays a key role in regulated K(+) secretion, which is mediated mainly through renal outer medullary K(+) (ROMK) channels located in the apical membrane. However, the mechanisms of the regulation of urinary K(+) excretion with regard to K(+) balance are not well known. We took advantage of a recently established mouse CCD cell line (mCCD(cl1)) to investigate the regulation of K(+) secretion by mineralocorticoid and K(+) concentration. We show that this cell line expresses ROMK mRNA and a barium-sensitive K(+) conductance in its apical membrane. As this conductance is sensitive to tertiapin-Q, with an apparent affinity of 6 nM, and to intracellular acidification, it is probably mediated by ROMK. Overnight exposure to 100 nM aldosterone did not significantly change the K(+) conductance, while it increased the amiloride-sensitive Na(+) transport. Overnight exposure to a high K(+) (7 mM) concentration produced a small but significant increase in the apical membrane barium-sensitive K(+) conductance. The mRNA levels of all ROMK isoforms measured by qRT-PCR were not changed by altering the basolateral K(+) concentration but were decreased by 15-45% upon treatment with aldosterone (0.3 or 300 nM for 1 and 3 h). The paradoxical response of ROMK expression to aldosterone could possibly work as a preventative mechanism to avoid excessive K(+) loss which would otherwise result from the increased electrogenic Na(+) transport and associated depolarization of the apical membrane in the CCD. In conclusion, mCCD(cl1) cells demonstrate a significant K(+) secretion, probably mediated by ROMK, which is not stimulated by aldosterone but increased by overnight exposure to a high K(+) concentration.