Saturated fatty acid alters embryonic cortical neurogenesis through modulation of gene expression in neural stem cells

A perturbed maternal metabolic environment such as chronically elevated circulating free fatty acids have been shown to affect stem cell fate during embryonic neurogenesis. However, molecular mechanisms behind this are not well defined, especially in human. Here in using directed differentiation of human embryonic stem cells (hESCs) into cortical neurons as model, we show that chronically elevated saturated fatty acid (palmitate) results in decreased proliferation of neural stem cells and increased differentiation into neurons. This phenotype could be due to palmitate mediated increased expression of key genes needed for neuronal differentiation such as EOMES, TBR1, NEUROD1 and RELN and reduced expression of SREBP regulated lipogenic genes at early stages of cortical differentiation. Furthermore, palmitate treatment increased histone acetylation globally and at select gene promoters among affected genes. We also found differential expression of several lncRNAs associated with cellular stress and metabolic diseases in the presence of palmitate including BDNF-AS suggesting the contribution of additional epigenetic regulatory mechanisms. Together, our results show that saturated fatty acid affects developmental neurogenesis through modulation of gene expression and through epigenetic regulatory mechanisms.     

Adult neurogenesis and cellular brain repair with neural progenitors, precursors and stem cells

Recent work in neuroscience has shown that the adult central nervous system (CNS) contains neural progenitors, precursors and stem cells that are capable of generating new neurons, astrocytes and oligodendrocytes. While challenging the previous dogma that no new neurons are born in the adult mammalian CNS, these findings bring with them the future possibilities for development of novel neural repair strategies. The purpose of this review is to present the current knowledge about constitutively occurring adult mammalian neurogenesis, highlight the critical differences between 'neurogenic' and 'non-neurogenic' regions in the adult brain, and describe the cardinal features of two well-described neurogenic regions-the subventricular zone/olfactory bulb system and the dentate gyrus of the hippocampus. We also provide an overview of presently used models for studying neural precursors in vitro, mention some precursor transplantation models and emphasize that, in this rapidly growing field of neuroscience, one must be cautious with respect to a variety of methodological considerations for studying neural precursor cells both in vitro and in vivo. The possibility of repairing neural circuitry by manipulating neurogenesis is an intriguing one, and, therefore, we also review recent efforts to understand the conditions under which neurogenesis can be induced in non-neurogenic regions of the adult CNS. This work aims towards molecular and cellular manipulation of endogenous neural precursors in situ, without transplantation. We conclude this review with a discussion of what might be the function of newly generated neurons in the adult brain, and provide a summary of present thinking about the consequences of disturbed adult neurogenesis and the reaction of neurogenic regions to disease.     

Nuclear receptor TLX regulates cell cycle progression in neural stem cells of the developing brain

TLX is an orphan nuclear receptor that is expressed exclusively in vertebrate forebrains. Although TLX is known to be expressed in embryonic brains, the mechanism by which it influences neural development remains largely unknown. We show here that TLX is expressed specifically in periventricular neural stem cells in embryonic brains. Significant thinning of neocortex was observed in embryonic d 14.5 TLX-null brains with reduced nestin labeling and decreased cell proliferation in the germinal zone. Cell cycle analysis revealed both prolonged cell cycles and increased cell cycle exit in TLX-null embryonic brains. Increased expression of a cyclin-dependent kinase inhibitor p21 and decreased expression of cyclin D1 provide a molecular basis for the deficiency of cell cycle progression in embryonic brains of TLX-null mice. Furthermore, transient knockdown of TLX by in utero electroporation led to precocious cell cycle exit and differentiation of neural stem cells followed by outward migration. Together these results indicate that TLX plays an important role in neural development by regulating cell cycle progression and exit of neural stem cells in the developing brain.     

Taurine induces proliferation of neural stem cells and synapse development in the developing mouse brain

Taurine is a sulfur-containing amino acid present in high concentrations in mammalian tissues. It has been implicated in several processes involving brain development and neurotransmission. However, the role of taurine in hippocampal neurogenesis during brain development is still unknown. Here we show that taurine regulates neural progenitor cell (NPC) proliferation in the dentate gyrus of the developing brain as well as in cultured early postnatal (P5) hippocampal progenitor cells and hippocampal slices derived from P5 mice brains. Taurine increased cell proliferation without having a significant effect on neural differentiation both in cultured P5 NPCs as well as cultured hippocampal slices and in vivo. Expression level analysis of synaptic proteins revealed that taurine increases the expression of Synapsin 1 and PSD 95. We also found that taurine stimulates the phosphorylation of ERK1/2 indicating a possible role of the ERK pathway in mediating the changes that we observed, especially in proliferation. Taken together, our results demonstrate a role for taurine in neural stem/progenitor cell proliferation in developing brain and suggest the involvement of the ERK1/2 pathways in mediating these actions. Our study also shows that taurine influences the levels of proteins associated with synapse development. This is the first evidence showing the effect of taurine on early postnatal neuronal development using a combination of in vitro, ex-vivo and in vivo systems.     

Enhanced cytomegalovirus infection of developing brain independent of the adaptive immune system

Cytomegalovirus (CMV) has been suggested as the most prevalent infectious agent causing neurological dysfunction in the developing brain; in contrast, CMV infections are rare in the adult brain. One explanation generally given for the developmental susceptibility to the virus is that the developing immune system is too immature to protect the central nervous system from viral infection, but as the immune system develops it can protect the brain. We suggest an alternate view: that developing brain cells are inherently more susceptible to CMV infection, independent of the immune system. We used a recombinant mouse CMV that leads to green fluorescent protein expression in infected cells. Control experiments demonstrated a high correlation between the number of cells detected with the viral GFP reporter gene and with immunocytochemical detection of the virus. After intracerebral inoculation, the number of CMV-infected cells in neonatal brains was many times greater than in mature control or mature immunodepressed SCID mice, and the mortality rate of neonates was substantially greater than SCID or control adults. Parallel experiments with live brain slices inoculated in vitro, done in the absence of the systemic immune system, generated similar data, with immature hippocampus, hypothalamus, cortex, striatum, and cerebellum showing substantially greater numbers of infected cells (100-fold) than found in adult slices in these same regions. Interestingly, in the cerebellar cortex, CMV-infected cells were more prevalent in the postmitotic Purkinje cell layer than in the mitotic granule cell layer, suggesting a selective infection of some cell types not dependent on cell division. Together, these data support the view that CMV has an intrinsic preference for infection of developing brain cells, independent, but not mutually exclusive, of the developmental status of the systemic immune system in controlling CMV infection.     

Cyclosporine inhibits mouse cytomegalovirus infection via a cyclophilin-dependent pathway specifically in neural stem/progenitor cells

The potential of neural stem and progenitor cell (NSPC) transplantation in neurodegenerative disease raises a concern about immunosuppressive agents and opportunistic neurotropic pathogens that may interfere with engraftment. Cytomegalovirus (CMV) is an important opportunistic pathogen infecting the central nervous system, where it may remain latent for life, following transplacental transmission. Cyclosporine (Cs), an immunosuppressive drug used in organ transplantation, where its use is associated with CMV reactivation, suppressed murine CMV (MCMV) infection in cultured NSPCs but not in fibroblasts. This activity of Cs appears to be mediated via cyclophilin (CyP) rather than via calcineurin. First, the calcineurin-specific inhibitor FK506 failed to suppress replication. Second, the CyP-specific inhibitor NIM811 strongly suppressed replication in NSPC. NSPCs maintained in the presence of NIM811 retained viral genomes for several weeks without detectable viral gene expression or obvious deleterious effects. The withdrawal of NIM811 reactivated viral replication, suggesting that the inhibitory mechanism was reversible. Finally, inhibition of endogenous CyP A (CyPA) by small interfering RNA also inhibited replication in NSPCs. These results show that MCMV replication depends upon cellular CyPA pathways in NSPCs (in a specific cell type-dependent fashion), that CyPA plays an important role in viral infection in this cell type, and that inhibition of viral replication via CyP leads to persistence of the viral genome without cell damage. Further, the calcineurin-signaling pathway conferring immunosuppression in T cells does not influence viral replication in a detectable fashion.     

室管膜下区神经干细胞与神经发生的研究进展

室管膜下区(subventricular zone,SVZ)存在着神经干细胞(nueral stem cells,NSCs),是成年哺乳动物脑内重要的神经发生区域。神经发生过程极为复杂,包括一系列的生物学事件。在病理状态下,SVZ区的细胞增殖,新生的神经细胞迁移到病灶处,取代或修复受损的细胞,起到保护脑组织的作用。该文就SVZ区的神经干细胞、神经发生过程及病理状态下神经发生的相关研究做一综述。     

Locating and labeling neural stem cells in the brain

The phenomenon of adult neurogenesis has been demonstrated in most mammals including humans. At least two regions of the adult brain maintain stem cells throughout life; the subgranular zone (SGZ) of the hippocampal dentate gyrus, and the subventricular zone (SVZ) of the lateral ventricle wall. Both regions continuously produce neurons that mature and become integrated into functional networks that are involved in learning and memory and odor discrimination, respectively. Apart from these well‐studied regions neurogenesis has been reported in a number of other brain regions, such as amygdala and cortex. However, these studies have been contested and there is currently no well‐postulated function for non‐SVZ/SGZ neurogenesis. The studies of the regional localization of neurogenesis in the brain have been made possible due to several methods for detecting adult neurogenesis including; bromodeoxyuridine labeling (BrdU) together with markers of mature neurons, genetic labeling, by mouse transgenesis, or with the use of viral vectors. These techniques are already put to creative use and will be essential for the discovery of the nature of the adult neural stem cells. In this mini‐review, we will discuss the localization of neural stem/progenitor cells in the brain and their implications as well as discussing the pro's and con's of stem cell labeling techniques. J. Cell. Physiol. 226: 1–7, 2010. © 2010 Wiley‐Liss, Inc.     

Atg5 and Ambra1 differentially modulate neurogenesis in neural stem cells

Neuroepithelial cells undergoing differentiation efficiently remodel their cytoskeleton and shape in an energy-consuming process. The capacity of autophagy to recycle cellular components and provide energy could fulfill these requirements, thus supporting differentiation. However, little is known regarding the role of basal autophagy in neural differentiation. Here we report an increase in the expression of the autophagy genes Atg7, Becn1, Ambra1 and LC3 in vivo in the mouse embryonic olfactory bulb (OB) during the initial period of neuronal differentiation at E15.5, along with a parallel increase in neuronal markers. In addition, we observed an increase in LC3 lipidation and autophagic flux during neuronal differentiation in cultured OB-derived stem/progenitor cells. Pharmacological inhibition of autophagy with 3-MA or wortmannin markedly decreased neurogenesis. These observations were supported by similar findings in two autophagy-deficient genetic models. In Ambra1 loss-of-function homozygous mice (gt/gt) the expression of several neural markers was decreased in the OB at E13.5 in vivo. In vitro, Ambra1 haploinsufficient cells developed as small neurospheres with an impaired capacity for neuronal generation. The addition of methylpyruvate during stem/progenitor cell differentiation in culture largely reversed the inhibition of neurogenesis induced by either 3-MA or Ambra1 haploinsufficiency, suggesting that neural stem/progenitor cells activate autophagy to fulfill their high energy demands. Further supporting the role of autophagy for neuronal differentiation Atg5-null OB cells differentiating in culture displayed decreased TuJ1 levels and lower number of cells with neurites. These results reveal new roles for autophagy-related molecules Atg5 and Ambra1 during early neuronal differentiation of stem/progenitor cells.     

Atg5 and Ambra1 differentially modulate neurogenesis in neural stem cells

Neuroepithelial cells undergoing differentiation efficiently remodel their cytoskeleton and shape in an energy-consuming process. The capacity of autophagy to recycle cellular components and provide energy could fulfill these requirements, thus supporting differentiation. However, little is known regarding the role of basal autophagy in neural differentiation. Here we report an increase in the expression of the autophagy genes Atg7, Becn1, Ambra1 and LC3 in vivo in the mouse embryonic olfactory bulb (OB) during the initial period of neuronal differentiation at E15.5, along with a parallel increase in neuronal markers. In addition, we observed an increase in LC3 lipidation and autophagic flux during neuronal differentiation in cultured OB-derived stem/progenitor cells. Pharmacological inhibition of autophagy with 3-MA or wortmannin markedly decreased neurogenesis. These observations were supported by similar findings in two autophagy-deficient genetic models. In Ambra1 loss-of-function homozygous mice (gt/gt) the expression of several neural markers was decreased in the OB at E13.5 in vivo. In vitro, Ambra1 haploinsufficient cells developed as small neurospheres with an impaired capacity for neuronal generation. The addition of methylpyruvate during stem/progenitor cell differentiation in culture largely reversed the inhibition of neurogenesis induced by either 3-MA or Ambra1 haploinsufficiency, suggesting that neural stem/progenitor cells activate autophagy to fulfill their high energy demands. Further supporting the role of autophagy for neuronal differentiation Atg5-null OB cells differentiating in culture displayed decreased TuJ1 levels and lower number of cells with neurites. These results reveal new roles for autophagy-related molecules Atg5 and Ambra1 during early neuronal differentiation of stem/progenitor cells.     

Murine cytomegalovirus infection of mouse testes.

With the aim of illustrating a mechanism of cytomegalovirus (CMV) venereal transmission, we induced murine CMV infection in the mouse testes of immunologically competent mice. Using in situ cytohybridization, we were able to show that murine CMV-specific DNA was associated with spermatocytes and mature sperm. Electron microscopy studies also supported sperm infection. The virus could be reisolated from infected epididymal sperm by cocultivation with mouse embryo fibroblasts. We found no difference in either the sexual performance or the fertilization efficiency of the sperm between infected and uninfected males.     

内源性神经干细胞与脑老化的治疗

近十几年研究发现成年人脑神经元可以再生,使人们重新认识老年脑神经细胞的可塑性,它为脑损伤的修复带来新的希望。最新研究表明,神经再生可被调控,是一种新的修复机制。这使得利用内源性神经干细胞治疗老龄相关的神经退行性疾病成为可能。     

Embryonic stem cell neurogenesis and neural specification

The prospect of using embryonic stem cell (ESC)‐derived neural progenitors and neurons to treat neurological disorders has led to great interest in defining the conditions that guide the differentiation of ESCs, and more recently induced pluripotent stem cells (iPSCs), into neural stem cells (NSCs) and a variety of neuronal and glial subtypes. Over the past decade, researchers have looked to the embryo to guide these studies, applying what we know about the signaling events that direct neural specification during development. This has led to the design of a number of protocols that successfully promote ESC neurogenesis, terminating with the production of neurons and glia with diverse regional addresses and functional properties. These protocols demonstrate that ESCs undergo neural specification in two, three, and four dimensions, mimicking the cell–cell interactions, patterning, and timing that characterizes the in vivo process. We therefore propose that these in vitro systems can be used to examine the molecular regulation of neural specification. J. Cell. Biochem. 111: 535–542, 2010. © 2010 Wiley‐Liss, Inc.     

Neural stem cells and neurogenesis in the adult

Research in the field of adult neurogenesis has seen substantial progress over recent years. Here we discuss some of the major focus areas for future investigation: neural stem cell heterogeneity, the role of latent stem cells, and the extent of neurogenesis in the adult human brain.     

Hypoxia and neural stem cells: from invertebrates to brain cancer stem cells

Oxygen is a fundamental element for all living organisms, and modifications in its concentration influence several physiological and pathological events such as embryogenesis, development and also aging. Regulation of oxygen levels is an important factor in neural stem cell biology (e.g. differentiation, growth and the capacity to generate more differentiated cells). Studies on neural stem cells in culture have deepened our knowledge of their survival, proliferation and differentiation pathways. However, traditional cell culture for neural stem cells is performed employing environmental oxygen levels of 20%, while the effective oxygen concentration in the developing and adult brain is significantly lower; this results in an important alteration of the in vivo conditions. Several data indicate that a so called "physiologic hypoxic condition" could strongly influence the growth of neural stem cells and their differentiation mechanisms both in vivo and in vitro. The present overview deals with the different mechanisms utilized by invertebrate and vertebrate organisms to respond to hypoxic conditions. It highlights how the adaptations and responses to different oxygen concentrations have changed along the developmental route and underlines the importance of oxygen concentration in neural physiology and differentiation, with a final hint to the involvement of hypoxia in brain cancer stem cells.     

Independent brain stem sites for ventilatory neurogenesis

The purpose was to determine if independent ventilatory rhythms could be generated in each half of completely separated brain stems. In decerebrate, cerebellectomized, vagotomized, paralyzed, and ventilated cats, activities of phrenic, recurrent laryngeal (RLN), and/or hypoglossal nerves were monitored. Midsaggital brain stem divisions were performed by sections and lesions. Eupnea continued following divisions of mesencephalon and pons. Hypoglossal and phrenic activities were eliminated after sections approximating the obex. In most preparations, RLNs discharged with independent rhythms after completion of midsagittal brain stem section and a C1 transection. Independent rhythms were also obtained from each half of medulla following transections at the pontomedullary junction and at C1, and midsagittal medullary divisions. In other animals with transections between pons and medulla and at C1, synchronized RLN and hypoglossal activities persisted after sagittal medullary divisions, 2.0 mm lateral to midline contralaterally. Data demonstrate that there is more than one potential brain stem site for ventilatory neurogenesis. It is hypothesized that there are many such sites, possibly having pacemaker cells, in pons and medulla.