17 research outputs found
Axon-glia interactions during central nervous system myelination
Myelination drastically speeds up action potential propagation along axons, which is
fundamental for the correct function of neuronal circuits. However, axon-oligodendrocyte
interactions regulating the onset of myelin formation remain
unclear. I sought to determine how reticulospinal axons control myelination, as they
are the first myelinated in the zebrafish spinal cord. I genetically manipulated
zebrafish in order to either remove such axons from a region of the spinal cord, or to
increase their number, and characterized oligodendrocyte-lineage cells following this
axonal loss- or gain-of-function.
In kinesin-binding protein (kbp) mutants, reticulospinal hindbrain neurons start
axonogenesis but axons fail to grow along the entire spinal cord as in wildtype,
providing an axon-deficient posterior spinal cord and an intact anterior region. I
found that early stages of oligodendrocyte development, such as the specification of
oligodendrocyte precursors, their distribution and migration were not affected in the
posterior spinal cord of these mutants. However, both the proliferation and the
survival of late precursors were impaired, resulting in a significant reduction of
mature oligodendrocytes in the posterior region of mutants at the onset of
myelination. Since the anterior spinal cord of mutants is indistinguishable from
wildtype, these results demonstrate that reticulospinal axons provide a mitogenic and
a survival signal to a subset of developing OPCs, enabling their differentiation and
lineage progression.
I then found that the absence of reticulospinal axons did not affect the timing of
oligodendrocyte differentiation, which matured on time, suggesting that this follows
an intrinsic timer, as previous studies suggested. Oligodendrocytes also did not
myelinate incorrect axonal targets, but instead adapted to the reduced axonal surface
by elaborating fewer myelin sheaths. Additionally, oligodendrocytes made shorter
sheaths, and also incorrectly ensheathed neuron somas in the mutant spinal cord,
suggesting that either kbp function or a precise amount of axonal surface are required
to prevent ectopic myelination of somas and to promote the longitudinal growth of
myelin sheaths.
In wildtype animals, the two reticulospinal Mauthner axons are the very first
myelinated in the spinal cord. In animals where Notch1a function is temporarily
abrogated or hoxb1 genes are temporarily upregulated, supernumerary Mauthner
neurons are generated. I found that these extra axons are robustly myelinated, with
no impairment of myelination of adjacent axons. Surprisingly, the number of
oligodendrocytes was not altered, but I found that each individual oligodendrocyte
elaborated more myelin sheaths, whose total length was also longer than in
wildtypes. Additionally, dorsal oligodendrocytes, which normally myelinate only
small-calibre dorsal axons, readily extended processes ventrally to myelinate the
supernumerary large-calibre Mauthner axons, in addition to small-calibre axons.
These results suggest that oligodendrocytes are plastic and are not destined to
myelinate a particular type of axon, and conversely, that axonal signals that induce
myelination are similar for different axons. The long-standing observation that
oligodendrocytes tend to myelinate either few large axons or many small axons thus
reflects local interactions of oligodendrocyte processes with the nearby axons, rather
than different subtypes of oligodendrocytes specified by an intrinsic programme of
differentiation.
Collectively, this work shows that axons extensively influence both oligodendrocyte
lineage progression and oligodendrocyte myelinating potential in vivo
CNS myelination requires VAMP2/3-mediated membrane expansion in oligodendrocytes
Myelin is required for rapid nerve signaling and is emerging as a key driver of CNS plasticity and disease. How myelin is built and remodeled remains a fundamental question of neurobiology. Central to myelination is the ability of oligodendrocytes to add vast amounts of new cell membrane, expanding their surface areas by many thousand-fold. However, how oligodendrocytes add new membrane to build or remodel myelin is not fully understood. Here, we show that CNS myelin membrane addition requires exocytosis mediated by the vesicular SNARE proteins VAMP2/3. Genetic inactivation of VAMP2/3 in myelinating oligodendrocytes caused severe hypomyelination and premature death without overt loss of oligodendrocytes. Through live imaging, we discovered that VAMP2/3-mediated exocytosis drives membrane expansion within myelin sheaths to initiate wrapping and power sheath elongation. In conjunction with membrane expansion, mass spectrometry of oligodendrocyte surface proteins revealed that VAMP2/3 incorporates axon-myelin adhesion proteins that are collectively required to form nodes of Ranvier. Together, our results demonstrate that VAMP2/3-mediated membrane expansion in oligodendrocytes is indispensable for myelin formation, uncovering a cellular pathway that could sculpt myelination patterns in response to activity-dependent signals or be therapeutically targeted to promote regeneration in disease
Myelination of neuronal cell bodies when myelin supply exceeds axonal demand
The correct targeting of myelin is essential for nervous system formation and function. Oligodendrocytes in the CNS myelinate some axons, but not others, and do not myelinate structures including cell bodies and dendrites [1]. Recent studies indicate that extrinsic signals, such as neuronal activity [2, 3] and cell adhesion molecules [4], can bias myelination toward some axons and away from cell bodies and dendrites, indicating that, in vivo, neuronal and axonal cues regulate myelin targeting. In vitro, however, oligodendrocytes have an intrinsic propensity to myelinate [5-7] and can promiscuously wrap inert synthetic structures resembling neuronal processes [8, 9] or cell bodies [4]. A current therapeutic goal for the treatment of demyelinating diseases is to greatly promote oligodendrogenesis [10-13]; thus, it is important to test how accurately extrinsic signals regulate the oligodendrocyte's intrinsic program of myelination in vivo. Here, we test the hypothesis that neurons regulate myelination with sufficient stringency to always ensure correct targeting. Surprisingly, however, we find that myelin targeting in vivo is not very stringent and that mistargeting occurs readily when oligodendrocyte and myelin supply exceed axonal demand. We find that myelin is mistargeted to neuronal cell bodies in zebrafish mutants with fewer axons and independently in drug-treated zebrafish with increased oligodendrogenesis. Additionally, by increasing myelin production of oligodendrocytes in zebrafish and mice, we find that excess myelin is also inappropriately targeted to cell bodies. Our results suggest that balancing oligodendrocyte-intrinsic programs of myelin supply with axonal demand is essential for correct myelin targeting in vivo and highlight potential liabilities of strongly promoting oligodendrogenesis
Intersectional Gene Expression in Zebrafish Using the Split KalTA4 System
In this study, we describe the adaptation of the split Gal4 system for zebrafish. The Gal4-UAS system is widely used for expression of genes-of-interest by crossing driver lines expressing the transcription factor Gal4 (under the control of the promoter of interest) with reporter lines where upstream activating sequence (UAS) repeats (recognized by Gal4) drive expression of the genes-of-interest. In the Split Gal4 system, hemi-drivers separately encode the DNA-binding domain (DBD) and the activation domain (AD) of Gal4. When encoded under two different promoters, only those cells in the intersection of the promoters' expression pattern and in which both promoters are active reconstitute a functional Gal4 and activate expression from a UAS-driven transgene. We split the zebrafish-optimized version of Gal4, KalTA4, and generated a hemi-driver encoding the KalTA4 DBD and a hemi-driver encoding KalTA4's AD. We show that split KalTA4 domains can assemble in vivo and transactivate a UAS reporter transgene and that each hemi-driver alone cannot transactivate the reporter. Also, transactivation can happen in several cell types, with similar efficiency to intact KalTA4. Finally, in transient mosaic expression assays, we show that when hemi-drivers are preceded by two distinct promoters, they restrict the expression of an UAS-driven reporter from a broader pattern (sox10) to its constituent smaller neuronal pattern. The Split KalTA4 system should be useful for expression of genes-of-interest in an intersectional manner, allowing for more refined manipulations of cell populations in zebrafish
A retroviral link to vertebrate myelination through retrotransposon RNA-mediated control of myelin gene expression
Myelin, the insulating sheath that surrounds neuronal axons, is produced by oligodendrocytes in the central nervous system (CNS). This evolutionary innovation, which first appears in jawed vertebrates, enabled rapid transmission of nerve impulses, more complex brains, and greater morphological diversity. Here, we report that RNA-level expression of RNLTR12-int, a retrotransposon of retroviral origin, is essential for myelination. We show that RNLTR12-int-encoded RNA binds to the transcription factor SOX10 to regulate transcription of myelin basic protein (Mbp, the major constituent of myelin) in rodents. RNLTR12-int-like sequences (which we name RetroMyelin) are found in all jawed vertebrates, and we further demonstrate their function in regulating myelination in two different vertebrate classes (zebrafish and frogs). Our study therefore suggests that retroviral endogenization played a prominent role in the emergence of vertebrate myelin.<br/
Rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART): Study protocol for a randomized controlled trial
Background: Acute respiratory distress syndrome (ARDS) is associated with high in-hospital mortality. Alveolar recruitment followed by ventilation at optimal titrated PEEP may reduce ventilator-induced lung injury and improve oxygenation in patients with ARDS, but the effects on mortality and other clinical outcomes remain unknown. This article reports the rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART). Methods/Design: ART is a pragmatic, multicenter, randomized (concealed), controlled trial, which aims to determine if maximum stepwise alveolar recruitment associated with PEEP titration is able to increase 28-day survival in patients with ARDS compared to conventional treatment (ARDSNet strategy). We will enroll adult patients with ARDS of less than 72 h duration. The intervention group will receive an alveolar recruitment maneuver, with stepwise increases of PEEP achieving 45 cmH(2)O and peak pressure of 60 cmH2O, followed by ventilation with optimal PEEP titrated according to the static compliance of the respiratory system. In the control group, mechanical ventilation will follow a conventional protocol (ARDSNet). In both groups, we will use controlled volume mode with low tidal volumes (4 to 6 mL/kg of predicted body weight) and targeting plateau pressure <= 30 cmH2O. The primary outcome is 28-day survival, and the secondary outcomes are: length of ICU stay; length of hospital stay; pneumothorax requiring chest tube during first 7 days; barotrauma during first 7 days; mechanical ventilation-free days from days 1 to 28; ICU, in-hospital, and 6-month survival. ART is an event-guided trial planned to last until 520 events (deaths within 28 days) are observed. These events allow detection of a hazard ratio of 0.75, with 90% power and two-tailed type I error of 5%. All analysis will follow the intention-to-treat principle. Discussion: If the ART strategy with maximum recruitment and PEEP titration improves 28-day survival, this will represent a notable advance to the care of ARDS patients. Conversely, if the ART strategy is similar or inferior to the current evidence-based strategy (ARDSNet), this should also change current practice as many institutions routinely employ recruitment maneuvers and set PEEP levels according to some titration method.Hospital do Coracao (HCor) as part of the Program 'Hospitais de Excelencia a Servico do SUS (PROADI-SUS)'Brazilian Ministry of Healt
Avaliação das Estimativas de Precipitação do Produto 3B43-TRMM do Estado do Amazonas
RESUMOPara avaliar os dados de precipitação pluvial via satélite no estado do Amazonas, compararam-se as estimativas do produto 3B43 do satélite TRMM (2004-2008) com dados de sete Estações Meteorológicas Convencionais (EMC). A comparação foi baseada nos seguintes parâmetros estatÃsticos: Erro Médio (EM), Raiz do Erro Médio Quadrático (REMQ), coeficiente de correlação linear (r) e Ãndice de concordância de Wilmott (d). As estimativas do TRMM foram similares aos dados de superfÃcie e representaram bem a variabilidade sazonal das chuvas. Os dados apresentaram alta correlação linear (r = 0,83), alto Ãndice de concordância (d = 0,85) e REMQ satisfatório (66,6 mm/mês). Dessa forma, as estimativas de precipitação pluvial do produto 3B43 podem ser utilizadas como uma fonte alternativa de dados de qualidade
Different Cell Death Responses Induced By Eupomatenoid-5 In Mcf-7 And 786-0 Tumor Cell Lines.
Natural products remain an important source of new drugs, including anticancer drugs. Recently, our group reported the anticancer activity of eupomatenoid-5 (eup-5), a neolignan isolated from Piper regnellii (Miq.) C. DC. var. regnellii leaves. In vitro studies demonstrated that MCF-7 (breast) and 786-0 (kidney) were among the cancer cell lines most sensitive to eup-5 treatment. The current results demonstrate that mitochondrial membrane depolarization and generation of reactive oxygen species are implicated in eup-5-mediated cytotoxic effects on these cancer cells lines. In MCF-7 cells, eup-5 led to phosphatidylserine externalization and caspase activation, whereas the same did not occur in 786-0 cells. Scanning electron microscopy revealed a reduction of microvilli density, as well as cell morphology alterations. Moreover, treated MCF-7 cells exhibited well-characterized apoptosis alterations, while treated 786-0 cells exhibited characteristics of programmed necroptosis process. These findings support the possibility that different mechanisms may be targeted by eup-5 in cell death response.291026-103