A New Outlook on Mental Illnesses: Glial Involvement Beyond the Glue.

Mental illnesses have long been perceived as the exclusive consequence of abnormalities in neuronal functioning. Until recently, the role of glial cells in the pathophysiology of mental diseases has largely been overlooked. However recently, multiple lines of evidence suggest more diverse and significant functions of glia with behavior-altering effects. The newly ascribed roles of astrocytes, oligodendrocytes and microglia have led to their examination in brain pathology and mental illnesses. Indeed, abnormalities in glial function, structure and density have been observed in postmortem brain studies of subjects diagnosed with mental illnesses. In this review, we discuss the newly identified functions of glia and highlight the findings of glial abnormalities in psychiatric disorders. We discuss these preclinical and clinical findings implicating the involvement of glial cells in mental illnesses with the perspective that these cells may represent a new target for treatment.

Síntese da hidroxiapatita e refinamento estrutural por difração de raios-x

A sample of hydroxyapatite was synthesized and its crystalline structure was analyzed by X-ray diffraction by means of the Rietveld method. Two functions were used to fit the peak profiles, modified Voigt (TCHZ) and Pearson VII. The occupational factors and lattice parameters obtained by both models show that the sample does not contain relevant cationic substitutions. The interatomic distances from Ca1 to oxygens O1, O2 and O3 were adequate for a pure hydroxyapatite without defect at site Ca1. Besides, the use of multiple lines in planes (300) and (002) associated with the model Pearson VII resulted in good agreement with the TCHZ model with respect to the size-strain effectswith an ellipsoidal shape of crystallites. In conclusion, the procedures adopted in the synthesis of hydroxyapatite produced a pure and crystalline material. The experimental results of transmission electron microscopy confirmed the predicted shape of crystals.

The roles of TL1A and Pno1 in the pathogenesis of rheumatoid arthritis

La polyarthrite rhumatoïde (PR) est une maladie auto-immune chronique. Elle est caractérisée par une inflammation persistante touchant de multiples petites articulations, causant douleurs, rougeurs, gonflements et déformations. Des études menées auprès de patients et d’animaux ont démontré que certains auto-anticorps, cytokines et enzymes tissue-déstructives sont des médiateurs importants dans le développement de la PR. Au cours des deux dernières décennies, les traitements de fond (DMARDs en anglais) ont été démontrés très efficaces pour traiter la PR. D'autre part, des effets secondaires ont été rapportés pour ces traitements, par exemple l'augmentation du risque d'infections opportunistes. L’objectif de ce travail est d’acquérir des connaissances sur le rôle du TL1A (TNF-like molécule 1 A; TNFSF15) et son partenaire Nob1 (Pno1 ; YOR145c) dans la pathogenèse de la PR afin de découvrir de nouveaux médicaments contre ces molécules dans l'avenir. TL1A est un membre de la famille du TNF. Il déclenche des signaux co-stimulateurs via le récepteur de mort 3 (DR3) et induit la prolifération ainsi que la production des cytokines pro inflammatoires par les lymphocytes. Des données multiples suggèrent l'implication de la cascade TL1A-DR3 dans plusieurs maladies auto-immunes. Donc, nous avons proposé les hypothèses suivantes:1) la production locale de TL1A dans les articulations est un composant d’un cercle vicieux qui aggrave la PR; 2) dans la PR, la production de TL1A dans les organes lymphoïde augmente la production d’auto-anticorps pathogénique. Au cours de ce travail, nous avons démontré que la TL1A aggrave la maladie chez les souris où l’arthrite a été induite par le collagène (AIC). Par ailleurs, nous avons constaté que l’expression de TL1A est élevée dans les tissus atteints de PR ainsi que dans les ganglions lymphatiques drainant de la souris AIC. Mécaniquement, nous avons découvert que la TL1A est induite par le TNF-α et IL-17 produits par les cellules T in vitro. Ces résultats montrent directement que les TL1A-DR3 jouent un rôle essentiel dans la pathogenèse de la PR. De plus, afin de poursuivre notre étude, la TL1A a été génétiquement supprimée dans les souris (TL1A KO). Nous avons montré que les souris TL1A KO n’ont aucune anomalie apparente et aucun dysfonctionnement du système immunitaire dans des conditions normales. Cependant, ces souris manifestent des AIC améliorées et une réduction significative des niveaux d'anticorps, anti-collagène du type II i dans le sérum. Nous avons trouvé que les ganglions lymphatiques de drainage (dLNs) de souris KO étaient plus petites avec une cellularité inférieure comparativement aux souris WT de 14 jours après l’immunisation. De plus, nous avons découvert que le DR3 a été exprimé par les cellules plasmatiques dans l’étape de la différenciation terminale et ces cellules surviennent mieux en présence de TL1A. La conclusion de cette étude apporte des nouvelles connaissances sur le rôle de TL1A qui amplifie les réponses humorales d’AIC. Nous avons suggéré que TL1A pourrait augmenter la réponse d’initiation d'anticorps contre collagène II (CII) ainsi que prolonger la survie des cellules plasmatiques. Une autre molécule qui nous intéresse est Pno1. Des études antérieures menées chez la levure ont suggéré que Pno1 est essentielle pour la néogénèse du protéasome et du ribosome Le protéasome étant crucial pour la différenciation terminale des cellules plasmatiques pendant les réponses humorales chez les mammifères, nous avons donc supposé que Pno1 joue un rôle dans la production d'anticorps pathogenique dans la PR via la voie du protéasome. Nous avons donc généré des souris génétiquement modifiées pour Pno1 afin d’étudier la fonction de Pno1 in vivo. Cependant, une mutation non-sens dans le Pno1 provoque une létalité embryonnaire à un stade très précoce chez les souris. D'autre part, une réduction de 50% de Pno1 ou une surexpression de Pno1 n’ont aucun effet ni sur le fonctionnent des cellules T et B, ni sur les activités du protéasome ainsi que sur la réponse humorale dans l’AIC. Ces résultats suggèrent que Pno1 est une molécule essentielle sans redondance. Par conséquent, il n’est pas une cible appropriée pour le développement de médicaments thérapeutiques. En conclusion, nos études ont révélé que la TL1A n’est pas essentielle pour maintenir les fonctions du système immunitaire dans des conditions normales. En revanche, il joue un rôle critique dans la pathogenèse de la PR en favorisant l'inflammation locale et la réponse humorale contre des auto-antigènes. Par conséquent, une inhibition de la TL1A pourrait être une stratégie thérapeutique pour le traitement de la PR. Au contraire, Pno1 est essentiel pour la fonction normale des cellules. Une délétion totale pourrait entraîner des conséquences graves. Il n’est pas une cible appropriée pour développer des médicaments de la PR.

Rôle et expression de l'interleukine-27 dans le contexte de la sclérose en plaques

Des études antérieures ont indiqué que l’IL-27 supprime le développement de l’encéphalomyélite auto-immune expérimentale (EAE), un modèle murin de la sclérose en plaques (SEP). L’expression en ARNm d’IL-27 est maximale au pic du développement de l’EAE. Cependant, sa contribution dans la pathogenèse de la SEP demeure irrésolue. Nous avons investigué si l’IL-27 contribue à moduler les réponses immunes dans le système nerveux central (SNC) de patients SEP. Nos résultats d’immunohistochimie sur échantillons post-mortem de cerveaux humains ont révélé que la production des deux sous-unités d’IL-27 (EBI-3 et p28) est plus élevée chez des patients comparés à des contrôles. De plus, les astrocytes (GFAP) et les microglies/macrophages (Iba1) représentent des sources biologiques importantes de l’IL-27 dans les lésions. Les lymphocytes T CD4 et CD8 qui infiltrent le SNC des patients expriment d’ailleurs le récepteur de l’IL-27 composé des chaînes gp130 et TCCR, supportant le concept que ces cellules pourraient répondre aux sources locales d’IL-27. Nous avons également démontré que des combinaisons de cytokines pro-inflammatoires (IFNγ, IL-1β et TNF) augmentent l’expression in vitro d’IL-27 par les astrocytes et macrophages humains, et que les microglies/macrophages de phénotype M1 produisent l’IL-27. Enfin, nous avons démontré que les astrocytes humains expriment aussi le récepteur à l’IL-27 et répondent à l’IL-27 par la phosphorylation de STAT1, mais pas de STAT3. Une telle signalisation dans ces cellules mène à l’augmentation d’expression de la molécule de co-inhibition PD-L1 et de la sécrétion de la chimiokine CXCL10.

New insights into trypanosomatid U5 small nuclear ribonucleoproteins

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Analysis of retrieved shoulder prostheses

In the last years the number of shoulder arthroplasties has been increasing. Simultaneously the study of their shape, size and strength and the reasons that bring to a possible early explantation have not yet been examined in detail. The research carried out directly on explants is practically nonexistent, this means a poor understanding of the mechanisms leading the patient and so the surgeon, to their removal. The analysis of the mechanisms which are the cause of instability, dislocation, broken, fracture, etc, may lead to a change in the structure or design of the shoulder prostheses and lengthen the life of the implant in situ. The idea was to analyze 22 explants through three methods in order to find roughness, corrosion and surface wear. In the first method, the humeral heads and/or the glenospheres were examined with the interferometer, a machine that through electromagnetic waves gives information about the roughness of the surfaces under examination. The output of the device was a total profile containing both roughness and information on the waves (representing the spatial waves most characteristic on the surface). The most important value is called "roughness average" and brings the average value of the peaks found in the local defects of the surfaces. It was found that 42% of the prostheses had considerable peak values in the area where the damage was caused by the implant and not only by external events, such as possibly the surgeon's hand. One of the problems of interest in the use of metallic biomaterials is their resistance to corrosion. The clinical significance of the degradation of metal implants has been the purpose of the second method; the interaction between human body and metal components is critical to understand how and why they arrive to corrosion. The percentage of damage in the joints of the prosthetic components has been calculated via high resolution photos and the software ImageJ. The 40% and 50% of the area appeared to have scratches or multiple lines due to mechanical artifacts. The third method of analysis has been made through the use of electron microscopy to quantify the wear surface in polyethylene components. Different joint movements correspond to different mechanisms of damage, which were imprinted in the parts of polyethylene examined. The most affected area was located mainly in the side edges. The results could help the manufacturers to modify the design of the prostheses and thus reduce the number of explants. It could also help surgeons in choosing the model of the prosthesis to be implanted in the patient.

Overexpression of lymphotoxin in T cells induces fulminant thymic involution

Activated lymphocytes and lymphoid-tissue inducer cells express lymphotoxins (LTs), which are essential for the organogenesis and maintenance of lymphoreticular microenvironments. Here we describe that T-cell-restricted overexpression of LT induces fulminant thymic involution. This phenotype was prevented by ablation of the LT receptors tumor necrosis factor receptor (TNFR) 1 or LT beta receptor (LTbetaR), representing two non-redundant pathways. Multiple lines of transgenic Ltalphabeta and Ltalpha mice show such a phenotype, which was not observed on overexpression of LTbeta alone. Reciprocal bone marrow transfers between LT-overexpressing and receptor-ablated mice show that involution was not due to a T cell-autonomous defect but was triggered by TNFR1 and LTbetaR signaling to radioresistant stromal cells. Thymic involution was partially prevented by the removal of one allele of LTbetaR but not of TNFR1, establishing a hierarchy in these signaling events. Infection with the lymphocytic choriomeningitis virus triggered a similar thymic pathology in wt, but not in Tnfr1(-/-) mice. These mice displayed elevated TNFalpha in both thymus and plasma, as well as increased LTs on both CD8(+) and CD4(-)CD8(-) thymocytes. These findings suggest that enhanced T cell-derived LT expression helps to control the physiological size of the thymic stroma and accelerates its involution via TNFR1/LTbetaR signaling in pathological conditions and possibly also in normal aging.

Early and Middle Holocene Hunter-Gatherer Occupations in Western Amazonia: The Hidden Shell Middens

We report on previously unknown early archaeological sites in the Bolivian lowlands, demonstrating for the first time early and middle Holocene human presence in western Amazonia. Multidisciplinary research in forest islands situated in seasonally-inundated savannahs has revealed stratified shell middens produced by human foragers as early as 10,000 years ago, making them the oldest archaeological sites in the region. The absence of stone resources and partial burial by recent alluvial sediments has meant that these kinds of deposits have, until now, remained unidentified. We conducted core sampling, archaeological excavations and an interdisciplinary study of the stratigraphy and recovered materials from three shell midden mounds. Based on multiple lines of evidence, including radiocarbon dating, sedimentary proxies (elements, steroids and black carbon), micromorphology and faunal analysis, we demonstrate the anthropogenic origin and antiquity of these sites. In a tropical and geomorphologically active landscape often considered challenging both for early human occupation and for the preservation of hunter-gatherer sites, the newly discovered shell middens provide evidence for early to middle Holocene occupation and illustrate the potential for identifying and interpreting early open-air archaeological sites in western Amazonia. The existence of early hunter-gatherer sites in the Bolivian lowlands sheds new light on the region’s past and offers a new context within which the late Holocene “Earthmovers” of the Llanos de Moxos could have emerged.

Rtr1 is the Saccharomyces cerevisiae homolog of a novel family of RNA polymerase II-binding proteins.

Cells must rapidly sense and respond to a wide variety of potentially cytotoxic external stressors to survive in a constantly changing environment. In a search for novel genes required for stress tolerance in Saccharomyces cerevisiae, we identified the uncharacterized open reading frame YER139C as a gene required for growth at 37 degrees C in the presence of the heat shock mimetic formamide. YER139C encodes the closest yeast homolog of the human RPAP2 protein, recently identified as a novel RNA polymerase II (RNAPII)-associated factor. Multiple lines of evidence support a role for this gene family in transcription, prompting us to rename YER139C RTR1 (regulator of transcription). The core RNAPII subunits RPB5, RPB7, and RPB9 were isolated as potent high-copy-number suppressors of the rtr1Delta temperature-sensitive growth phenotype, and deletion of the nonessential subunits RPB4 and RPB9 hypersensitized cells to RTR1 overexpression. Disruption of RTR1 resulted in mycophenolic acid sensitivity and synthetic genetic interactions with a number of genes involved in multiple phases of transcription. Consistently, rtr1Delta cells are defective in inducible transcription from the GAL1 promoter. Rtr1 constitutively shuttles between the cytoplasm and nucleus, where it physically associates with an active RNAPII transcriptional complex. Taken together, our data reveal a role for members of the RTR1/RPAP2 family as regulators of core RNAPII function.

Effects of butyrate on the colon cancer cell phenotype-chemosensitization and upregulation of proteins implicated in cancer progression

Colon cancer is the second leading cause of cancer mortality in the U.S. Surgery is the only truly effective human colon cancer (HCC) therapy due to marked intrinsic drug resistance. The inefficacy of therapies developed for metastatic HCC suggests that advances in colon cancer chemoprevention and chemotherapy will be needed to reduce HCC mortality. The dietary fiber metabolite butyrate (NaB) is a candidate cancer chemopreventive agent that inhibits growth, promotes differentiation and stimulates apoptosis of HCC cells. Epidemiological and experimental studies suggest that dietary fiber protects against the development of HCC, however, recent large prospective trials have not found significant protection. ^ The first central hypothesis of this dissertation project is that the diversity of phenotypic changes induced by NaB in HCC cells includes molecular alterations that oppose its chemopreventive action and thereby limit its efficacy. We investigated the effect of NaB on the expression/activity of epidermal growth factor receptor (EGFR) and cyclooxygenase-2 (COX-2) in HCC HT29 cells. NaB treatment induced a 13-fold increase in EGFR expression in concert with its chemopreventive action in vitro, i.e., induction of growth suppression and G1 arrest, apoptosis and a differentiated phenotype. NaB-induced EGFR was active based on multiple lines of evidence. The EGFR was: (1) heavily phosphorylated at Tyrosine (P-Tyr); (2) associated with the cytoskeleton; (3) localized at the cell surface, and activated in response to EGF; and (4) NaB treatment of the cells induced activation of the EGFR effector Erk1/2. NaB treatment also induced a 7-fold increase in COX-2 expression. The NaB-induced COX-2 was active based on significantly increased PGE2 production. ^ The second central hypothesis is that NaB treatment would render HCC cells more chemosensitive to chemotherapy agents based on the increased apoptotic index induced by NaB. NaB treatment chemosensitized HT29 cells to 5-FU and doxorubicin, despite increases in the expression of P-glycoprotein and a related drug resistance protein (MRP). ^ These results raise the intriguing possibility that the chemopreventive effects of fiber may require concomitant treatment with EGFR and/or COX-2 inhibitors. Similarly, NaB may be a rational drug to combine with existing chemotherapeutic agents for the management of advanced HCC patients. ^

Moving forward on facilitation research: response to changing environments and effects on the diversity, functioning and evolution of plant communities

Once seen as anomalous, facilitative interactions among plants and their importance for community structure and functioning are now widely recognized. The growing body of modelling, descriptive and experimental studies on facilitation covers a wide variety of terrestrial and aquatic systems throughout the globe. However, the lack of a general body of theory linking facilitation among different types of organisms and biomes and their responses to environmental changes prevents further advances in our knowledge regarding the evolutionary and ecological implications of facilitation in plant communities. Moreover, insights gathered from alternative lines of inquiry may substantially improve our understanding of facilitation, but these have been largely neglected thus far. Despite over 15 years of research and debate on this topic, there is no consensus on the degree to which plant–plant interactions change predictably along environmental gradients (i.e. the stress-gradient hypothesis), and this hinders our ability to predict how plant–plant interactions may affect the response of plant communities to ongoing global environmental change. The existing controversies regarding the response of plant–plant interactions across environmental gradients can be reconciled when clearly considering and determining the species-specificity of the response, the functional or individual stress type, and the scale of interest (pairwise interactions or community-level response). Here, we introduce a theoretical framework to do this, supported by multiple lines of empirical evidence. We also discuss current gaps in our knowledge regarding how plant–plant interactions change along environmental gradients. These include the existence of thresholds in the amount of species-specific stress that a benefactor can alleviate, the linearity or non-linearity of the response of pairwise interactions across distance from the ecological optimum of the beneficiary, and the need to explore further how frequent interactions among multiple species are and how they change across different environments. We review the latest advances in these topics and provide new approaches to fill current gaps in our knowledge. We also apply our theoretical framework to advance our knowledge on the evolutionary aspects of plant facilitation, and the relative importance of facilitation, in comparison with other ecological processes, for maintaining ecosystem structure, functioning and dynamics. We build links between these topics and related fields, such as ecological restoration, woody encroachment, invasion ecology, ecological modelling and biodiversity–ecosystem-functioning relationships. By identifying commonalities and insights from alternative lines of research, we further advance our understanding of facilitation and provide testable hypotheses regarding the role of (positive) biotic interactions in the maintenance of biodiversity and the response of ecological communities to ongoing environmental changes.

Emerging roles for the double strand break repair protein 53BP1 in transcriptional regulation

Disruption of the mechanisms that regulate cell-cycle checkpoints, DNA repair, and apoptosis results in genomic instability and often leads to the development of cancer. In response to double stranded breaks (DSBs) as induced by ionizing radiation (IR), generated during DNA replication, or through immunoglobulin heavy chain (IgH) rearrangements in T and B cells of lymphoid origin, the protein kinases ATM and ATR are central players that activate signaling pathways leading to DSB repair. p53 binding protein 1 (53BP1) participates in the repair of DNA double stranded breaks (DSBs) where it is recruited to or near sites of DNA damage. In addition to its well established role in DSB repair, multiple lines of evidence implicate 53BP1 in transcription which stem from its initial discovery as a p53 binding protein in a yeast two-hybrid screen. However, the mechanisms behind the role of 53BP1 in these processes are not well understood. ^ 53BP1 possesses several motifs that are likely important for its role in DSB repair including two BRCA1 C-terminal repeats, tandem Tudor domains, and a variety of phosphorylation sites. In addition to these motifs, we identified a glycine and arginine rich region (GAR) upstream of the Tudor domains, a sequence that is oftentimes serves as a site for protein arginine methylation. The focus of this project was to characterize the methylation of 53BP1 and to evaluate how methylation influenced the role of 53BP1 as a tumor suppressor. ^ Using a variety of biochemical techniques, we demonstrated that 53BP1 is methylated by the PRMT1 methyltransferase in vivo. Moreover, GAR methylation occurs on arginine residues in an asymmetric manner. We further show that sequences upstream of the Tudor domains that do not include the GAR stretch are sufficient for 53BP1 oligomerization in vivo. While investigating the role of arginine methylation in 53BP1 function, we discovered that 53BP1 associates with proteins of the general transcription apparatus as well as to other factors implicated in coordinating transcription with chromatin function. Collectively, these data support a role for 53BP1 in regulating transcription and provide insight into the possible mechanisms by which this occurs. ^

Insulin-like growth factor 1 regulates developing brain glucose metabolism

The brain has enormous anabolic needs during early postnatal development. This study presents multiple lines of evidence showing that endogenous brain insulin-like growth factor 1 (Igf1) serves an essential, insulin-like role in promoting neuronal glucose utilization and growth during this period. Brain 2-deoxy-d- [1-14C]glucose uptake parallels Igf1 expression in wild-type mice and is profoundly reduced in Igf1−/− mice, particularly in those structures where Igf1 is normally most highly expressed. 2-Deoxy-d- [1-14C]glucose is significantly reduced in synaptosomes prepared from Igf1−/− brains, and the deficit is corrected by inclusion of Igf1 in the incubation medium. The serine/threonine kinase Akt/PKB is a major target of insulin-signaling in the regulation of glucose transport via the facilitative glucose transporter (GLUT4) and glycogen synthesis in peripheral tissues. Phosphorylation of Akt and GLUT4 expression are reduced in Igf1−/− neurons. Phosphorylation of glycogen synthase kinase 3β and glycogen accumulation also are reduced in Igf1−/− neurons. These data support the hypothesis that endogenous brain Igf1 serves an anabolic, insulin-like role in developing brain metabolism.

Synaptophysin, a major synaptic vesicle protein, is not essential for neurotransmitter release.

Synaptophysin (syp I) is a synaptic vesicle membrane protein that constitutes approximately 7% of the total vesicle protein. Multiple lines of evidence implicate syp I in a number of nerve terminal functions. To test these, we have disrupted the murine Syp I gene. Mutant mice lacking syp I were viable and fertile. No changes in the structure and protein composition of the mutant brains were observed except for a decrease in synaptobrevin/VAMP II. Synaptic transmission was normal with no detectable changes in synaptic plasticity or the probability of release. Our data demonstrate that one of the major synaptic vesicle membrane proteins is not essential for synaptic transmission, suggesting that its function is either redundant or that it has a more subtle function not apparent in the assays used.

Pathogenic autoantibodies are routinely generated during the response to foreign antigen: a paradigm for autoimmune disease.

The immune system's ability to distinguish self and nonself is essential for both host defense against foreign agents and protection of self-antigens from autoimmune destruction. Such discrimination is complicated by extensive structural homology shared between foreign and self antigens. One hypothesis to explain the development of an autoimmune response is that some B cells activated by foreign antigen acquire, through somatic mutation, specificity for both the eliciting foreign antigen and self antigen. If such clones arise frequently, there must be a mechanism for their elimination. We have analyzed the extent of autoreactivity arising in a nonautoimmune host during the response to a foreign antigen. To overcome the process of apoptosis in primary B cells that might routinely eliminate autoreactive clones, we generated B-cell hybridomas from spleen cells of immunized mice by using a fusion partner constitutively expressing bcl-2. Multiple lines were obtained that recognize simultaneously the hapten phosphorylcholine and the self antigen double-stranded DNA. This dual specificity was not present early but was detected by day 10 after immunization. Some of these cross-reactive antibodies deposit in kidneys in a pattern similar to what is seen in autoimmune disease. These results demonstrate that autoantibodies arise at a high frequency as part of a response to foreign antigen. It has previously been shown that autoreactivity is regulated by central deletion; these data demonstrate a need for negative selection in peripheral lymphoid organs also, to regulate autoantibodies acquiring their self-specificity by somatic mutation.