Glycosphingolipids (GSLs) are amphipathic lipids ubiquitously expressed in all vertebrate cells and body fluids, but they are especially abundant in the nervous system. The synthesis of GSLs generally is initiated in the endoplasmic reticulum and completed in the Golgi apparatus, followed by transportation to the plasma membrane surface as an integral component. The amount and expression patterns of GSLs change drastically in brains during the embryonic to postnatal stages. Recent studies have revealed that GSLs are highly localized in cell surface microdomains and function as important components that mediate signal transduction and cell adhesion. Also in developing brains, GSLs are suggested to play important roles in nervous system formation. Disturbance of GSL expression and metabolism affects brain function, resulting in a variety of diseases, particularly lysosomal storage diseases. In this review, we describe some aspects of the roles of GSLs, especially of gangliosides, in brain development. Glycosphingolipids (GSLs) are amphipathic molecules composed of a hydrophilic carbohydrate chain and a hydrophobic ceramide moiety that contains a sphingosine and a FA residue (1). GSLs containing one or more sialic acid residues in the carbohydrate chain are referred to as gangliosides. Based on the sequences of the core carbohydrate residues, GSLs are classified into a number of series, including gala-, ganglio-, isoganglio-, lacto-, neolacto-, lactoganglio-, globo-, isoglobo-, and muco-series.

Neural stem cells (NSCs) are undifferentiated neural cells characterized by their high proliferative potential and the capacity for self-renewal with retention of multipotency. Over the past two decades, there has been a huge effort to identify NSCs morphologically, genetically, and molecular biologically. It is still controversial, however, what bona fide NSCs are. To define and characterize NSCs more systematically, it is crucial to explore novel cell-surface marker molecules of NSCs. In this study, we focused on GD3, a b-series ganglioside that is enriched in the immature brain and the subventricular zone (SVZ) of the postnatal and adult brain, and evaluated the usefulness of GD3 as a cell-surface biomarker for identifying NSCs. We demonstrated that GD3 was expressed in more than 80% of NSCs prepared from embryonic, postnatal, and adult mouse brain tissue by the neurosphere culture method. The percentage of GD3-expressing NSCs in neurospheres was nearly the same as it was in neurospheres derived from embryonic, postnatal, and adult brains but decreased drastically to about 40% after differentiation. GD3(+) cells isolated from embryonic mouse striata, postnatal, and adult mouse SVZs by fluorescence-activated cell sorting with an R24 anti-GD3 monoclonal antibody efficiently generated neurospheres compared with GD3(-) cells. These cells possessed multipotency to differentiate into neurons, astrocytes, and oligodendrocytes. These data indicate that GD3 is a unique and powerful cell-surface biomarker to identify and isolate NSCs.

Neural stem cells (NSCs) possess high proliferative potential and the capacity for self-renewal with retention of multipotency to differentiate into neuronal and glial cells. NSCs are the source for neurogenesis during central nervous system development from fetal and adult stages. Although the human natural killer-1 (HNK-1) carbohydrate epitope is expressed predominantly in the nervous system and involved in intercellular adhesion, cell migration, and synaptic plasticity, the expression patterns and functional roles of HNK-1-containing glycoconjugates in NSCs have not been fully recognized. We found that HNK-1 was expressed in embryonic mouse NSCs and that this expression was lost during the process of differentiation. Based on proteomics analysis, it was revealed that the HNK-1 epitopes were almost exclusively displayed on an extracellular matrix protein, tenascin-C (TNC), in the mouse embryonic NSCs. Furthermore, the HNK-1 epitope was found to be present only on the largest isoform of the TNC molecules. In addition, the expression of HNK-1 was dependent on expression of the largest TNC variant but not by enzymes involved in the biosynthesis of HNK-1. By knocking down HNK-1 sulfotransferase or TNC by small interfering RNA, we further demonstrated that HNK-1 on TNC was involved in the proliferation of NSCs via modulation of the expression level of the epidermal growth factor receptor. Our finding provides insights into the function of HNK-1 carbohydrate epitopes in NSCs to maintain stemness during neural development. Neural stem cells (NSCs)4 are undifferentiated neural cells characterized by their high proliferative potential and the capacity for self-renewal with retention of multipotency to differentiate into brain-forming cells, such as neurons, astrocytes, and oligodendrocytes (1-3). Environmental factors of NSCs, such as various growth factors, the extracellular matrix (ECM), and cell adhesion molecules, are known to play important roles in the maintenance of the stem cell population throughout specific cell lineage pathways (4 -7).Glycoconjugates, including glycoproteins, proteoglycans, and glycolipids, are expressed mainly on the cell surface as ECM, and they are known to regulate cell-to-cell communications. Certain glycoconjugates also serve as excellent biomarkers at various stages of the cellular differentiation of NSCs and play important functional roles in determining cell fate (8 -12). For example, stage-specific embryonic antigen-1 (SSEA-1), which is well known as a specific maker of undifferentiated cells including mouse embryonic stem cells, is expressed on NSCs and associated with cell migration (8, 10, 13). Recently, we have also demonstrated that cells positive for GD3 ganglioside (NeuAc␣2-8NeuAc␣2-3Gal␤1-4Glc␤1-1ЈCer) isolated from mouse brains of various ages possess characteristics of neural stem cells (11).The human natural killer-1 (HNK-1) carbohydrate epitope (CD57) was originally reported as a specific antigenic determinant for human natural killer cells (14) but is now widely known ...

In neural stem cells, basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) promote cell proliferation and self-renewal. In the bFGF-and EGF-responsive neural stem cells, ␤1-integrin also plays important roles in crucial cellular processes, including proliferation, migration, and apoptosis. The cross-talk of the signaling pathways mediated by these growth factors and ␤1-integrin, however, has not been fully elucidated. Here we report a novel molecular mechanism through which bFGF or EGF promotes the proliferation of mouse neuroepithelial cells (NECs). In the NECs, total ␤1-integrin expression levels and proliferation were dose-dependently increased by bFGF but not by EGF. EGF rather than bFGF strongly induced the increase of ␤1-integrin localization on the NEC surface. bFGF-and EGF-induced ␤1-integrin up-regulation and proliferation were inhibited after treatment with a mitogen-activated protein kinase kinase inhibitor, U0126, which indicates the dependence on the mitogen-activated protein kinase pathway. Involvement of ␤1-integrin in bFGF-and EGF-induced proliferation was confirmed by the finding that NEC proliferation and adhesion to fibronectin-coated dishes were inhibited by knockdown of ␤1-integrin using small interfering RNA. On the other hand, apoptosis was induced in NECs treated with RGD peptide, a small ␤1-integrin inhibitor peptide with the Arg-Gly-Asp motif, but it was independent of ␤1-integrin expression levels. Those results suggest that regulation of ␤1-integrin expression/localization is involved in cellular processes, such as proliferation, induced by bFGF and EGF in NECs. The mechanism underlying the proliferation through ␤1-integrin would not be expected to be completely identical, however, for bFGF and EGF.Neural stem cells (NSCs) 3 are defined as undifferentiated neural cells with a high potential for proliferation and the capacity for self-renewal with retention of multipotency (1-4). During development, NSCs have the capability to generate brain-forming cells, such as neurons, astrocytes, and oligodendrocytes. The application of NSCs to cell-based transplantation is a very attractive and promising strategy for regenerative and restorative medicine (5-7).The fate of NSCs (self-renewal, proliferation, differentiation, survival, and death) is regulated by intracellular programs mediated by a number of transcription factors or epigenetic modifications, including DNA methylation, histone modification, and non-coding RNA expression (8 -11). Furthermore, the signaling pathways of various growth factors, the extracellular matrix, and cell adhesion molecules present in specific microenvironments or niche in NSCs are also known to play important roles in maintenance of the stem cell population (12-18). In particular, basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) are primary mitogens to promote proliferation of NSCs through the mitogen-activated protein kinase (MAPK) pathway in vitro (19 -22). The responses of NSCs to those growth factors are thought to ...

The interaction of amyloid β-proteins (Aβs) with membrane lipids has been postulated as an early event in Aβ fibril formation in Alzheimer’s disease. We evaluated the effects of several putative bioactive Aβs and gangliosides on neural stem cells (NSCs) isolated from embryonic mouse brains or the subventricular zone of adult mouse brains. Incubation of the isolated NSCs with soluble Aβ1–40 alone did not cause any change in the number of NSCs, but soluble Aβ1–42 increased their number. Aggregated Aβ1–40 and Aβ1–42 increased the number of NSCs but soluble and aggregated Aβ25–35 decreased the number. Soluble Aβ1–40 and Aβ1–42 did not affect the number of apoptotic cells but aggregated Aβ1–40 and Aβ1–42 did. When NSCs were treated with a combination of GM1 or GD3 and soluble Aβ1–42, cell proliferation was enhanced, indicating that both GM1 and GD3 as well as Aβs are involved in promoting cell proliferation and survival of NSCs. These observations suggest the potential of beneficial effects of using gangliosides and Aβs for promoting NSC proliferation.

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Oxaliplatin is a chemotherapeutic agent that is effective against various types of cancer including colorectal cancer. Acute cold hyperalgesia is a serious side effect of oxaliplatin treatment. Although the therapeutic drug pregabalin is beneficial for preventing peripheral neuropathic pain by targeting the voltage-dependent calcium channel α2δ-1 (Cavα2δ-1) subunit, the effect of oxaliplatin-induced acute cold hypersensitivity is uncertain. To analyze the contribution of the Cavα2δ-1 subunit to the development of oxaliplatin-induced acute cold hypersensitivity, Cavα2δ-1 subunit expression in the rat spinal cord was analyzed after oxaliplatin treatment. Behavioral assessment using the acetone spray test showed that 6 mg/kg oxaliplatin-induced cold hypersensitivity 2 and 4 days later. Oxaliplatin-induced acute cold hypersensitivity 4 days after treatment was significantly inhibited by pregabalin (50 mg/kg, p.o.). Oxaliplatin (6 mg/kg, i.p.) treatment increased the expression level of Cavα2δ-1 subunit mRNA and protein in the spinal cord 2 and 4 days after treatment. Immunohistochemistry showed that oxaliplatin increased Cavα2δ-1 subunit protein expression in superficial layers of the spinal dorsal horn 2 and 4 days after treatment. These results suggest that oxaliplatin treatment increases Cavα2δ-1 subunit expression in the superficial layers of the spinal cord and may contribute to functional peripheral acute cold hypersensitivity.

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