Marjan Moreels

Cytokine-induced cell death in human oligodendroglial cell lines: I. Synergistic effects of IFN-γ and TNF-α on apoptosis

Multiple sclerosis is a chronic inflammatory disease of the central nervous system. Myelin and oligodendrocytes are considered the major targets of injury caused by a cell‐mediated immune response. There is circumstantial evidence that proinflammatory cytokines like tumor necrosis factor α (TNF‐α) and interferon γ (IFN‐γ) could have disease‐promoting roles in multiple sclerosis (MS). In the present study, the cytotoxic effects of IFN‐γ and TNF‐α on the human oligodendroglial cell lines human oligodendroglioma (HOG) and MO3.13 were analyzed. When the oligodendroglial cell lines were cultured in the presence of IFN‐γ or TNF‐α, apoptotic cell death was observed in both cell lines after >24 hr incubation. Apoptosis was evidenced by a decrease in cell viability, apoptotic changes in cell and nucleus morphology, and disruption of the membrane asymmetry. Our data show that TNF‐α and IFN‐γ induce apoptosis in a dose‐dependent fashion in both oligodendroglial cell lines and that their synergistic effect results in enhanced cell death. Understanding the regulation of cell death pathways in oligodendrocytes is critical for protecting myelin‐producing cells and their associated axons during injury in patients with MS.

Leukemia inhibitory factor is produced by myelin-reactive T cells from multiple sclerosis patients and protects against tumor necrosis factor-α-induced oligodendrocyte apoptosis

In multiple sclerosis (MS), damage to oligodendrocytes is believed to be caused by an aberrant immune response initiated by autoreactive T cells. Increasing evidence indicates that these T cells are not exclusively detrimental but might also exert protective effects. We report for the first time that myelin‐reactive T‐cell clones from eight MS patients (6/19) and five healthy controls (4/11) produce leukemia inhibitory factor (LIF), a member of the neuropoietic family of neurotrophins. In addition, T‐cell clones specific for tetanus toxoid, CD4+ and CD8+ T cells, and monocytes, but not B cells, secreted LIF. LIF‐producing T lymphocytes and macrophages were also identified immunohistochemically in both active and chronic‐active MS lesions. We further demonstrated dose‐dependent protective effects of LIF on tumor necrosis factor‐α‐induced apoptosis of oligodendrocytes. In conclusion, our data demonstrate that peripheral and CNS‐infiltrating T cells from MS patients produce LIF, a protective factor for oligodendrocytes. This study emphasizes that secretion of LIF may contribute to the neuroprotective effects of autoreactive T cells.

Ultrastructural and immunocytochemical analysis of multilineage differentiated human dental pulp- and umbilical cord-derived mesenchymal stem cells

Mesenchymal stem cells (MSCs) are one of the most promising stem cell types due to their availability and relatively simple requirements for in vitro expansion and genetic manipulation. Besides the well-characterized MSCs derived from bone marrow, there is growing evidence suggesting that dental pulp and the umbilical cord matrix both contain a substantial amount of cells having properties similar to those of MSCs. In order to assess the potential of dental pulp-derived MSCs (DPSC) and umbilical cord-derived MSCs (UCSC) in future clinical applications, it is essential to gain more insight into their differentiation capacity and to evaluate the tissues formed by these cells. In the present study, the morphological and ultrastructural characteristics of DPSC and UCSC induced towards osteogenic, adipogenic, and chondrogenic lineages were investigated. Cultured DPSC and UCSC showed a similar expression pattern of antigens characteristic of MSCs including CD105, CD29, CD44, CD146, and STRO-1. Under appropriate culture conditions, both DPSC and UCSC showed chondrogenic and osteogenic potential. Adipogenesis could be only partially induced in DPSC resulting in the de novo expression of fatty acid binding protein (FABP), whereas UCSC expressed FABP combined with a very high accumulation of lipid droplets in the cytoplasm. Our results demonstrate, at the biochemical and ultrastructural level, that DPSC display at least bilineage potential, whereas UCSC, which are developmentally more primitive cells, show trilineage potential. We emphasize that transmission electron microscopical analysis is useful to elucidate detailed structural information and provides indisputable evidence of differentiation. These findings highlight their potential therapeutic value for cell-based tissue engineering.

Alpha-smooth muscle actin (α-SMA) and nestin expression in reactive astrocytes in multiple sclerosis lesions : potential regulatory role of transforming growth factor-beta 1 (TGF-β1)

Aims: Rapid and extensive activation of astrocytes occurs subsequent to many forms of central nervous system (CNS) injury. Recent studies have revealed that the expression profile of reactive astrocytes comprises antigens present during astrocyte development. Elevated levels of the injury‐related cytokine transforming growth factor‐beta 1 (TGF‐β1) secreted by microglial cells and invading macrophages have been correlated with the reactive astrocyte phenotype and glial scar formation. Methods: In the present study, the expression profile of alpha‐smooth muscle actin (α‐SMA) and nestin, two cytoskeletal proteins expressed during astrocyte development, was studied in multiple sclerosis (MS) lesions. In addition, α‐SMA and nestin organization and expression were analysed in rat primary astrocyte cultures in response to TGF‐β1. Results: In active lesions and in the hypercellular margin of chronic active MS lesions, immunostaining for α‐SMA revealed a subpopulation of reactive astrocytes, whereas the majority of reactive astrocytes expressed nestin. α‐SMA and nestin expressing reactive astrocytes were in close relationship with TGF‐β1 expressing macrophages or microglia. In addition, TGF‐β1 expression within α‐SMA or nestin expressing astrocytes was also detected. Our in vitro experiments showed that TGF‐β1 regulated the organization and expression of α‐SMA and nestin in astrocytes. Conclusions: Reactive astrocytes in active MS lesions re‐express α‐SMA and nestin. We suggest that the in vivo re‐expression might be under regulation of TGF‐β1. These results further clarify the regulation of astrocyte activity after CNS injury, which is important for the astroglial adaptation to pathological situations.

Characterization of mature rat oligodendrocytes : a proteomic approach

Oligodendrocytes are glial cells responsible for the synthesis and maintenance of myelin in the central nervous system (CNS). Oligodendrocytes are vulnerable to damage occurring in a variety of neurological diseases. Understanding oligodendrocyte biology is crucial for the dissemination of de‐ and remyelination mechanisms. The goal of the present study is the construction of a protein database of mature rat oligodendrocytes. Post‐mitotic oligodendrocytes were isolated from mature Wistar rats and subjected to immunocytochemistry. Proteins were extracted and analyzed by means of two‐dimensional gel electrophoresis and two‐dimensional liquid chromatography, both coupled to mass spectrometry. The combination of the gel‐based and gel‐free approach resulted in confident identification of a total of 200 proteins. A minority of proteins were identified in both proteomic strategies. The identified proteins represent a variety of functional groups, including novel oligodendrocyte proteins. The results of this study emphasize the power of the applied proteomic strategy to study known or to reveal new proteins and to investigate their regulation in oligodendrocytes in different disease models.