Brain-derived neurotrophic factor-deficient mice develop aggressiveness and hyperphagia in conjunction with brain serotonergic abnormalities

Brain-derived neurotrophic factor (BDNF) has trophic effects on serotonergic (5-HT) neurons in the central nervous system. However, the role of endogenous BDNF in the development and function of these neurons has not been established in vivo because of the early postnatal lethality of BDNF null mice. In the present study, we use heterozygous BDNF+/− mice that have a normal life span and show that these animals develop enhanced intermale aggressiveness and hyperphagia accompanied by significant weight gain in early adulthood; these behavioral abnormalities are known to correlate with 5-HT dysfunction. Forebrain 5-HT levels and fiber density in BDNF+/− mice are normal at an early age but undergo premature age-associated decrements. However, young adult BDNF+/− mice show a blunted c-fos induction by the specific serotonin releaser-uptake inhibitor dexfenfluramine and alterations in the expression of several 5-HT receptors in the cortex, hippocampus, and hypothalamus. The heightened aggressiveness can be ameliorated by the selective serotonin reuptake inhibitor fluoxetine. Our results indicate that endogenous BDNF is critical for the normal development and function of central 5-HT neurons and for the elaboration of behaviors that depend on these nerve cells. Therefore, BDNF+/− mice may provide a useful model to study human psychiatric disorders attributed to dysfunction of serotonergic neurons.

Viral mediated expression of insulin-like growth factor I blocks the aging-related loss of skeletal muscle function

During the aging process, mammals lose up to a third of their skeletal muscle mass and strength. Although the mechanisms underlying this loss are not entirely understood, we attempted to moderate the loss by increasing the regenerative capacity of muscle. This involved the injection of a recombinant adeno-associated virus directing overexpression of insulin-like growth factor I (IGF-I) in differentiated muscle fibers. We demonstrate that the IGF-I expression promotes an average increase of 15% in muscle mass and a 14% increase in strength in young adult mice, and remarkably, prevents aging-related muscle changes in old adult mice, resulting in a 27% increase in strength as compared with uninjected old muscles. Muscle mass and fiber type distributions were maintained at levels similar to those in young adults. We propose that these effects are primarily due to stimulation of muscle regeneration via the activation of satellite cells by IGF-I. This supports the hypothesis that the primary cause of aging-related impairment of muscle function is a cumulative failure to repair damage sustained during muscle utilization. Our results suggest that gene transfer of IGF-I into muscle could form the basis of a human gene therapy for preventing the loss of muscle function associated with aging and may be of benefit in diseases where the rate of damage to skeletal muscle is accelerated.

Cooperative interactions among afferents govern the induction of homosynaptic long-term depression in the hippocampus.

Prolonged periods of low-frequency stimulation have been shown to produce a robust, long-term synaptic depression (LTD) in both hippocampus and visual cortex. In the present study we have examined the extent to which interactions among afferents govern the induction of homosynaptic LTD in young-adult rats in hippocampal region CA1 in vitro. Field excitatory postsynaptic potentials were assessed before and after conditioning stimulation consisting of two 10-min trains of low-frequency stimulation (LFS; 1 Hz) of the Schaffer collateral/commissural pathway. LFS at an intensity producing a 0.5-mV response did not produce significant synaptic depression. However, LFS administered at a higher intensity resulted in significant input-specific LTD of a 0.5-mV test response. Picrotoxin, which also facilitates depolarization of CA1 neurons, significantly enhanced the magnitude of LTD after LFS at 0.5 mV. In addition, LFS at 0.5 mV in normal perfusion medium (no picrotoxin) produced only small changes in synaptic efficacy when either of two converging pathways was conditioned separately but produced a robust LTD when both pathways were conditioned simultaneously. This cooperative LTD was reversibly blocked by prior administration of 100 microM DL-aminophosphonovaleric acid but not by 20 microM nimodipine. Taken together, these results suggest that cooperative interactions among afferents contribute to voltage-dependent processes underlying the induction of homosynaptic LTD.

Hematopoietic and lung abnormalities in mice with a null mutation of the common beta subunit of the receptors for granulocyte-macrophage colony-stimulating factor and interleukins 3 and 5.

Gene targeting was used to create mice with a null mutation of the gene encoding the common beta subunit (beta C) of the granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 3 (IL-3; multi-CSF), and interleukin 5 (IL-5) receptor complexes (beta C-/- mice). High-affinity binding of GM-CSF was abolished in beta C-/- bone marrow cells, while cells from heterozygous animals (beta C+/- mice) showed an intermediate number of high-affinity receptors. Binding of IL-3 was unaffected, confirming that the IL-3-specific beta chain remained intact. Eosinophil numbers in peripheral blood and bone marrow of beta C-/- animals were reduced, while other hematological parameters were normal. In clonal cultures of beta C-/- bone marrow cells, even high concentrations of GM-CSF and IL-5 failed to stimulate colony formation, but the cells exhibited normal quantitative responsiveness to stimulation by IL-3 and other growth factors. beta C-/- mice exhibited normal development and survived to young adult life, although they developed pulmonary peribronchovascular lymphoid infiltrates and areas resembling alveolar proteinosis. There was no detectable difference in the systemic clearance and distribution of GM-CSF between beta C-/- and wild-type littermates. The data establish that beta C is normally limiting for high-affinity binding of GM-CSF and demonstrate that systemic clearance of GM-CSF is not mediated via such high-affinity receptor complexes.

Functional anatomy of human eyeblink conditioning determined with regional cerebral glucose metabolism and positron-emission tomography.

Relative cerebral glucose metabolism was examined with positron-emission tomography (PET) as a measure of neuronal activation during performance of the classically conditioned eyeblink response in 12 young adult subjects. Each subject received three sessions: (i) a control session with PET scan in which unpaired presentations of the tone conditioned stimulus and corneal airpuff unconditioned stimulus were administered, (ii) a paired training session to allow associative learning to occur, and (iii) a paired test session with PET scan. Brain regions exhibiting learning-related activation were identified as those areas that showed significant differences in glucose metabolism between the unpaired control condition and well-trained state in the 9 subjects who met the learning criterion. Areas showing significant activation included bilateral sites in the inferior cerebellar cortex/deep nuclei, anterior cerebellar vermis, contralateral cerebellar cortex and pontine tegmentum, ipsilateral inferior thalamus/red nucleus, ipsilateral hippocampal formation, ipsilateral lateral temporal cortex, and bilateral ventral striatum. Among all subjects, including those who did not meet the learning criterion, metabolic changes in ipsilateral cerebellar nuclei, bilateral cerebellar cortex, anterior vermis, contralateral pontine tegmentum, ipsilateral hippocampal formation, and bilateral striatum correlated with degree of learning. The localization to cerebellum and its associated brainstem circuitry is consistent with neurobiological studies in the rabbit model of eyeblink classical conditioning and neuropsychological studies in brain-damaged humans. In addition, these data support a role for the hippocampus in conditioning and suggest that the ventral striatum may also be involved.

Metaxin, a gene contiguous to both thrombospondin 3 and glucocerebrosidase, is required for embryonic development in the mouse: implications for Gaucher disease.

We have identified a murine gene, metaxin, that spans the 6-kb interval separating the glucocerebrosidase gene (GC) from the thrombospondin 3 gene on chromosome 3E3-F1. Metaxin and GC are transcribed convergently; their major polyadenylylation sites are only 431 bp apart. On the other hand, metaxin and the thrombospondin 3 gene are transcribed divergently and share a common promoter sequence. The cDNA for metaxin encodes a 317-aa protein, without either a signal sequence or consensus for N-linked glycosylation. Metaxin protein is expressed ubiquitously in tissues of the young adult mouse, but no close homologues have been found in the DNA or protein data bases. A targeted mutation (A-->G in exon 9) was introduced into GC by homologous recombination in embryonic stem cells to establish a mouse model for a mild form of Gaucher disease. A phosphoglycerate kinase-neomycin gene cassette was also inserted into the 3'-flanking region of GC as a selectable marker, at a site later identified as the terminal exon of metaxin. Mice homozygous for the combined mutations die early in gestation. Since the same amino acid mutation in humans is associated with mild type 1 Gaucher disease, we suggest that metaxin protein is likely to be essential for embryonic development in mice. Clearly, the contiguous gene organization at this locus limits targeting strategies for the production of murine models of Gaucher disease.

Personalidade e Adesão ao tratamento em pacientes adultos jovens portadores de HIV

Objetivos: Este trabalhado teve como objetivo avaliar a relação entre características de personalidade e adesão ao tratamento antirretroviral em pacientes jovens adultos (18 a 32 anos) portadores do HIV. Metodologia: A adesão ao tratamento foi avaliada pelo \"Cuestionario para la Evaluación de la Adhesión al Tratamiento Antirretrovial (CEAT) e as características de personalidade foram avaliadas pela Bateria Fatorial de Personalidade (BFP). Resultados: A amostra foi composta por 51 pacientes e a análise dos dados sugere que o fator de personalidade associado a adesão ao tratamento é Realização. Este fator também surge como preditor independente para adesão ao tratamento. Discussão: Tais resultados reforçam a necessidade de inclusão destes aspectos na avaliação integral realizada pela equipe de saúde ao paciente jovem portador do vírus HIV. Os dados também reforçam a importância do profissional da área de psicologia como parte da equipe visto que, com o acompanhamento psicológico, é possível ajustar e modificar tais características, desenvolvendo estratégias de atendimento mais eficazes

Efeito do inibidor de proteinase de origem vegetal EcTI, sobre a inflamação pulmonar alérgica crônica em camundongos Balb/c

INTRODUÇÃO: A prevalência de asma tem crescido e a maioria dos pacientes com asma grave não obtém o controle total dos sintomas com as terapias disponíveis, fazendo-se necessária a busca por novas alternativas terapêuticas. Inibidores de proteinases têm sido estudados como tratamento de processos inflamatórios, dentre eles o Enterolobium contortisiliquum Tripsin Inhibitor (EcTI) OBJETIVO: Avaliar se o inibidor de proteinase EcTI modula a hiperresponsividade brônquica à metacolina, inflamação, remodelamento e estresse oxidativo nas vias aéreas e septos alveolares em um modelo experimental de inflamação pulmonar alérgica crônica. MÉTODOS: Vinte e quatro camundongos Balb/c machos, entre seis e sete semanas de vida, pesando em media 25 g foram divididos em quatro grupos: C (controle), OVA (sensibilizados com ovalbumina, 50 ug intraperironeal (i.p) nos dias 0 e 14 e desafiados nos dias 22, 24, 26, 28); C+EC (controle tratados com EcTI (2 mg/kg/i.p) nos dias 22 a 28); OVA+EC (sensibilizados e desafiados com ovalbumina e também tratados com EcTI (2 mg/kg -i.p) nos dias 22 a 28). No dia 29, foram realizadas realizadas: (i) hiperresponsividade à metacolina e obtidas as respostas máximas de resistência e elastância do sistema respiratório; (ii) análise histopatológica do pulmão para quantificação de eosinófilos, fibras colágenas e elásticas nas vias aéreas (VA) e nos septos alveolares (SA); e (iii) imunohistoquímica para quantificação de células positivas para IFN-y, IL-4, IL-5, IL-13, MMP-9, TIMP-1, TGF-beta, iNOS, NF-kB e fração de volume de isoprostano nas VA e nos SA. Uma semana após o dia 29 foi realizada a técnica de anafilaxia cutanea passiva(PCA) para quantificar IgE e IgG1. A significância foi considerada quando p < 0,05. RESULTADOS: Houve aumento de todos os parâmetros avaliados no grupo OVA em relação ao grupo controle (p < 0,05). Houve atenuação da resposta máxima de Rrs e Ers no grupo OVA+EC comparado as grupo OVA (p < 0,05). O tratamento com EcTI nos animais sensibilizados atenuou o número de eosinófilos, células positivas para IL-4, IL-5, IL-13,IFN-y, iNOS, MMP-9, TIMP-1, NF-kB e TGF-beta e fração de volume de isoprostano, fibras colágenas e elásticas nas vias aéreas e nos séptos alveolares quando comparado ao grupo OVA (p < 0,05).Houve reaçao de PCA nos animais sensibilizados com ovalbumina. CONCLUSÃO: EcTI atenuou a hiperresponsividade brônquica, a inflamação, o remodelamento e o estresse oxidativo nesse modelo experimental de inflamação pulmonar alérgica crônica. Embora sejam necessários mais estudos, esse inibidor pode ser considerado uma futura ferramenta farmacológica para o tratamento de asma

Dragon Tears: Highlighting the Christian Salvation Message Through the Use of Allegorical Fantasy Fiction

The recent movie versions of the Chronicles of Narnia and the Lord of the Rings Trilogy have raised in the publishing world a public demand for good quality fantasy fiction that can be used as a teaching tool for spreading the Christian salvation message. Dragon Tears was written to fulfill this need, providing young adult readers with both an engaging fantasy story and at the same time, an allegorical format through which to receive the Christian salvation story. Dragon Tears draws its foundational material from two educational backgrounds in religion and creative writing.

Moya moya disease : literature review and case report

Trabalho Final do Curso de Mestrado Integrado em Medicina, Faculdade de Medicina, Universidade de Lisboa, 2014

Bases moléculaires et cellulaires d’un trouble neurodéveloppemental causé par l’haploinsuffisance de SYNGAP1

La déficience intellectuelle est la cause d’handicap la plus fréquente chez l’enfant. De nombreuses évidences convergent vers l’idée selon laquelle des altérations dans les gènes synaptiques puissent expliquer une fraction significative des affections neurodéveloppementales telles que la déficience intellectuelle ou encore l’autisme. Jusqu’à récemment, la majorité des mutations associées à la déficience intellectuelle a été liée au chromosome X ou à la transmission autosomique récessive. D’un autre côté, plusieurs études récentes suggèrent que des mutations de novo dans des gènes à transmission autosomique dominante, requis dans les processus de la plasticité synaptique peuvent être à la source d’une importante fraction des cas de déficience intellectuelle non syndromique. Par des techniques permettant la capture de l’exome et le séquençage de l’ADN génomique, notre laboratoire a précédemment reporté les premières mutations pathogéniques dans le gène à transmission autosomique dominante SYNGAP1. Ces dernières ont été associées à des troubles comportementaux tels que la déficience intellectuelle, l’inattention, des problèmes d’humeur, d’impulsivité et d’agressions physiques. D’autres patients sont diagnostiqués avec des troubles autistiques et/ou des formes particulières d’épilepsie généralisée. Chez la souris, le knock-out constitutif de Syngap1 (souris Syngap1+/-) résulte en des déficits comme l’hyperactivité locomotrice, une réduction du comportement associée à l’anxiété, une augmentation du réflexe de sursaut, une propension à l’isolation, des problèmes dans le conditionnement à la peur, des troubles dans les mémoires de travail, de référence et social. Ainsi, la souris Syngap1+/- représente un modèle approprié pour l’étude des effets délétères causés par l’haploinsuffisance de SYNGAP1 sur le développement de circuits neuronaux. D’autre part, il est de première importance de statuer si les mutations humaines aboutissent à l’haploinsuffisance de la protéine. SYNGAP1 encode pour une protéine à activité GTPase pour Ras. Son haploinsuffisance entraîne l’augmentation des niveaux d’activité de Ras, de phosphorylation de ERK, cause une morphogenèse anormale des épines dendritiques et un excès dans la concentration des récepteurs AMPA à la membrane postsynaptique des neurones excitateurs. Plusieurs études suggèrent que l’augmentation précoce de l’insertion des récepteurs AMPA au sein des synapses glutamatergiques contribue à certains phénotypes observés chez la souris Syngap1+/-. En revanche, les conséquences de l’haploinsuffisance de SYNGAP1 sur les circuits neuronaux GABAergiques restent inconnues. Les enjeux de mon projet de PhD sont: 1) d’identifier l’impact de mutations humaines dans la fonction de SYNGAP1; 2) de déterminer si SYNGAP1 contribue au développement et à la fonction des circuits GABAergiques; 3) de révéler comment l’haploinsuffisance de Syngap1 restreinte aux circuits GABAergiques affecte le comportement et la cognition. Nous avons publié les premières mutations humaines de type faux-sens dans le gène SYNGAP1 (c.1084T>C [p.W362R]; c.1685C>T [p.P562L]) ainsi que deux nouvelles mutations tronquantes (c.2212_2213del [p.S738X]; c.283dupC [p.H95PfsX5]). Ces dernières sont toutes de novo à l’exception de c.283dupC, héritée d’un père mosaïque pour la même mutation. Dans cette étude, nous avons confirmé que les patients pourvus de mutations dans SYNGAP1 présentent, entre autre, des phénotypes associés à des troubles comportementaux relatifs à la déficience intellectuelle. En culture organotypique, la transfection biolistique de l’ADNc de Syngap1 wild-type dans des cellules pyramidales corticales réduit significativement les niveaux de pERK, en fonction de l’activité neuronale. Au contraire les constructions plasmidiques exprimant les mutations W362R, P562L, ou celle précédemment répertoriée R579X, n’engendre aucun effet significatif sur les niveaux de pERK. Ces résultats suggèrent que ces mutations faux-sens et tronquante résultent en la perte de la fonction de SYNGAP1 ayant fort probablement pour conséquences d’affecter la régulation du développement cérébral. Plusieurs études publiées suggèrent que les déficits cognitifs associés à l’haploinsuffisance de SYNGAP1 peuvent émerger d’altérations dans le développement des neurones excitateurs glutamatergiques. Toutefois, si, et auquel cas, de quelle manière ces mutations affectent le développement des interneurones GABAergiques résultant en un déséquilibre entre l’excitation et l’inhibition et aux déficits cognitifs restent sujet de controverses. Par conséquent, nous avons examiné la contribution de Syngap1 dans le développement des circuits GABAergiques. A cette fin, nous avons généré une souris mutante knockout conditionnelle dans laquelle un allèle de Syngap1 est spécifiquement excisé dans les interneurones GABAergiques issus de l’éminence ganglionnaire médiale (souris Tg(Nkx2.1-Cre);Syngap1flox/+). En culture organotypique, nous avons démontré que la réduction de Syngap1 restreinte aux interneurones inhibiteurs résulte en des altérations au niveau de leur arborisation axonale et dans leur densité synaptique. De plus, réalisés sur des coupes de cerveau de souris Tg(Nkx2.1-Cre);Syngap1flox/+, les enregistrements des courants inhibiteurs postsynaptiques miniatures (mIPSC) ou encore de ceux évoqués au moyen de l’optogénétique (oIPSC) dévoilent une réduction significative de la neurotransmission inhibitrice corticale. Enfin, nous avons comparé les performances de souris jeunes adultes Syngap1+/-, Tg(Nkx2.1-Cre);Syngap1flox/+ à celles de leurs congénères contrôles dans une batterie de tests comportementaux. À l’inverse des souris Syngap1+/-, les souris Tg(Nkx2.1-Cre);Syngap1flox/+ ne présentent pas d’hyperactivité locomotrice, ni de comportement associé à l’anxiété. Cependant, elles démontrent des déficits similaires dans la mémoire de travail et de reconnaissance sociale, suggérant que l’haploinsuffisance de Syngap1 restreinte aux interneurones GABAergiques dérivés de l’éminence ganglionnaire médiale récapitule en partie certains des phénotypes cognitifs observés chez la souris Syngap1+/-. Mes travaux de PhD établissent pour la première fois que les mutations humaines dans le gène SYNGAP1 associés à la déficience intellectuelle causent la perte de fonction de la protéine. Mes études dévoilent, également pour la première fois, l’influence significative de ce gène dans la régulation du développement et de la fonction des interneurones. D’admettre l’atteinte des cellules GABAergiques illustre plus réalistement la complexité de la déficience intellectuelle non syndromique causée par l’haploinsuffisance de SYNGAP1. Ainsi, seule une compréhension raffinée de cette condition neurodéveloppementale pourra mener à une approche thérapeutique adéquate.

Compensatory T-Cell Regulation in Unaffected Relatives of SLE Patients, and Opposite IL-2/CD25-Mediated Effects Suggested by Coreferentiality Modeling

In human systemic lupus erythematosus (SLE), diverse autoantibodies accumulate over years before disease manifestation. Unaffected relatives of SLE patients frequently share a sustained production of autoantibodies with indiscriminable specificity, usually without ever acquiring the disease. We studied relations of IgG autoantibody profiles and peripheral blood activated regulatory T-cells (aTregs), represented by CD4(+)CD25(bright) T-cells that were regularly 70-90% Foxp3(+). We found consistent positive correlations of broad-range as well as specific SLE-associated IgG with aTreg frequencies within unaffected relatives, but not patients or unrelated controls. Our interpretation: unaffected relatives with shared genetic factors compensated pathogenic effects by aTregs engaged in parallel with the individual autoantibody production. To study this further, we applied a novel analytic approach named coreferentiality that tests the indirect relatedness of parameters in respect to multivariate phenotype data. Results show that independently of their direct correlation, aTreg frequencies and specific SLE-associated IgG were likely functionally related in unaffected relatives: they significantly parallelled each other in their relations to broad-range immunoblot autoantibody profiles. In unaffected relatives, we also found coreferential effects of genetic variation in the loci encoding IL-2 and CD25. A model of CD25 functional genetic effects constructed by coreferentiality maximization suggests that IL-2-CD25 interaction, likely stimulating aTregs in unaffected relatives, had an opposed effect in SLE patients, presumably triggering primarily T-effector cells in this group. Coreferentiality modeling as we do it here could also be useful in other contexts, particularly to explore combined functional genetic effects.

Multifaceted Role of Heme during Severe Plasmodium falciparum Infections in India

Several immunomodulatory factors are involved in malaria pathogenesis. Among them, heme has been shown to play a role in the pathophysiology of severe malaria in rodents, but its role in human severe malaria remains unclear. Circulating levels of total heme and its main scavenger, hemopexin, along with cytokine/chemokine levels and biological parameters, including hemoglobin and creatinine levels, as well as transaminase activities, were measured in the plasma of 237 Plasmodium falciparum-infected patients living in the state of Odisha, India, where malaria is endemic. All patients were categorized into well-defined groups of mild malaria, cerebral malaria (CM), or severe noncerebral malaria, which included acute renal failure (ARF) and hepatopathy. Our results show a significant increase in total plasma heme levels with malaria severity, especially for CM and malarial ARF. Spearman rank correlation and canonical correlation analyses have shown a correlation between total heme, hemopexin, interleukin-10, tumor necrosis factor alpha, gamma interferon-induced protein 10 (IP-10), and monocyte chemotactic protein 1 (MCP-1) levels. In addition, canonical correlations revealed that heme, along with IP-10, was associated with the CM pathophysiology, whereas both IP-10 and MCP-1 together with heme discriminated ARF. Altogether, our data indicate that heme, in association with cytokines and chemokines, is involved in the pathophysiology of both CM and ARF but through different mechanisms.

Broadened T-cell Repertoire Diversity in ivIg-treated SLE Patients is Also Related to the Individual Status of Regulatory T-cells

Intravenous IgG (ivIg) is a therapeutic alternative for lupus erythematosus, the mechanism of which remains to be fully understood. Here we investigated whether ivIg affects two established sub-phenotypes of SLE, namely relative oligoclonality of circulating T-cells and reduced activity of CD4 + Foxp3+ regulatory T-cells (Tregs) reflected by lower CD25 surface density.

Complex genetic findings in a female patient with pyruvate dehydrogenase complex deficiency: Null mutations in the PDHX gene associated with unusual expression of the testis-specific PDHA2 gene in her somatic cells

Human pyruvate dehydrogenase complex (PDC) catalyzes a key step in the generation of cellular energy and is composed by three catalytic elements (E1, E2, E3), one structural subunit (E3-binding protein), and specific regulatory elements, phosphatases and kinases (PDKs, PDPs). The E1α subunit exists as two isoforms encoded by different genes: PDHA1 located on Xp22.1 and expressed in somatic tissues, and the intronless PDHA2 located on chromosome 4 and only detected in human spermatocytes and spermatids. We report on a young adult female patient who has PDC deficiency associated with a compound heterozygosity in PDHX encoding the E3-binding protein. Additionally, in the patient and in all members of her immediate family, a full-length testis-specific PDHA2 mRNA and a 5′UTR-truncated PDHA1 mRNA were detected in circulating lymphocytes and cultured fibroblasts, being bothmRNAs translated into full-length PDHA2 and PDHA1 proteins, resulting in the co-existence of both PDHA isoforms in somatic cells.Moreover, we observed that DNA hypomethylation of a CpG island in the coding region of PDHA2 gene is associatedwith the somatic activation of this gene transcription in these individuals. This study represents the first natural model of the de-repression of the testis-specific PDHA2 gene in human somatic cells, and raises some questions related to the somatic activation of this gene as a potential therapeutic approach for most forms of PDC deficiency.