Cytokine-induced changes in the ability of astrocytes to support migration of oligodendrocyte precursors and axon growth.

Repair of demyelination in the CNS requires that oligodendrocyte precursors (OPs) migrate, divide and then myelinate. Repair of axon damage requires axonal regeneration. Limited remyelination and axon regeneration occurs soon after injury, but usually ceases in a few days. In vivo and in vitro experiments have shown that astrocytic environments are not very permissive for migration of OPs or for axonal re-growth. Yet remyelination and axon sprouting early after injury occurs in association with astrocytes, while later astrocytes can exclude remyelination and prevent axon regeneration. A large and changing cast of cytokines are released following CNS injury, so we investigated whether some of these alone or in combination can affect the ability of astrocytes to support migration of OPs and neuritic outgrowth. Interleukin (IL) 1alpha, tumour necrosis factor alpha, transforming growth factor (TGF) beta, basic fibroblast growth factor (bFGF), platelet-derived growth factor and epidermal growth factor alone exerted little or no effect on migration of OPs on astrocytes, whereas interferon (IFN) gamma was inhibitory. The combination of IL-1alpha + bFGF was found to be pro-migratory, and this effect could be neutralized by TGFbeta. We also examined neuritic outgrowth from dorsal root ganglion explants in three-dimensional astrocyte cultures treated with cytokines and found that IL-1alpha + bFGF greatly increased axon outgrowth and that this effect could be blocked by TGFbeta and IFNgamma. All these effects were absent or much smaller when OP migration or axon growth was tested on laminin, so the main effect of the cytokines was via astrocytes. The cytokine effects did not correlate with expression on astrocytes of laminin, fibronectin, tenascin, chondroitin sulphate proteoglycan, N-cadherin, polysialyated NCAM (PSA-NCAM), tissue plasminogen activator (tPA) or urokinase (uPA).

Mechanisms of astrocyte-directed neurite guidance.

Astrocytes have recently become better recognized as playing vital roles in regulating the patterning of central nervous system neurites during development and following injury. In general, astrocytes have been shown to be supportive of neurite extension, but alterations in the biochemical properties of astrocytes in particular areas during development and in gliotic tissue may act to confine neurite outgrowth and thus provide guidance cues. In vivo studies indicate that restrictive astrocytes function through their altered expression of specific extracellular matrix molecules, including tenascin, chondroitin, and keratan sulfate proteoglycans. In addition, several in vitro models suggest that other cell surface molecules are utilized by restrictive astrocytes to direct neurite trajectories.

Inflammatory cytokines interact to modulate extracellular matrix and astrocytic support of neurite outgrowth.

Following injury to the central nervous system, an astroglial scar forms that is thought to impede neuronal regeneration and recovery of function. It is our hypothesis that inflammatory cytokines act upon astrocytes to alter their biochemical and physical properties, which may in turn be responsible for failed neuronal regeneration. We have therefore examined the interactions of two cytokines with prominent actions following injury, interferon-gamma (IFN-gamma) and basic fibroblast growth factor (FGF2), in modulating the extracellular matrix and proliferation of astrocytes in culture. We also evaluated the effects of these cytokines on the ability of astrocytes to support the growth of neurites. IFN-gamma significantly inhibited the proliferation of rat cortical astrocytes both in serum-free and serum-containing media as measured by [3H]thymidine incorporation. Furthermore, IFN-gamma also antagonized FGF2-induced proliferation. In parallel, IFN-gamma reduced the levels of the ECM molecules tenascin, laminin, and fibronectin as evaluated by Western blot analysis and immunocytochemistry. Similarly, IFN-gamma also antagonized FGF2-induced tenascin formation. While IFN-gamma-pretreated astrocyte monolayers did not differ from control in their ability to support neurite outgrowth of cortical neurons, it antagonized the enhancement of neurite outgrowth on FGF2-treated monolayers. We demonstrate that IFN-gamma did not alter signal transduction through the FGF2 receptor down to the phosphorylation of mitogen-activated protein kinase, suggesting that the interaction is at the level of transcriptional regulation or that an alternate pathway is involved. These results support the hypothesis that inflammatory cytokines interact to modulate several facets of the gliotic response and such interactions may be important in creating the biochemical and physical properties of the glial scar.

Spinal cord injury in the rat: treatment with bacterial lipopolysaccharide and indomethacin enhances cellular repair and locomotor function.

Trauma to the rat's spinal cord results in a lesion characterized by ingrowth of glial cells, accumulation of macrophages, and the progressive development of necrosis and cavitation. Since, when appropriately activated, both astrocytes and macrophages secrete growth-promoting cytokines, we examined whether treatment with drugs that stimulate the secretory activities of these cells might promote tissue repair and reduce necrosis in the traumatized spinal cord. The spinal cord of rats was crushed extradurally at T8 and the rats were injected intraperitoneally with (i) a lipopolysaccharide (LPS) or ImuVert to activate cytokine secretion, (ii) Indomethacin to reduce necrosis by inhibiting prostaglandin synthesis, (iii) a combination of LPS+Indomethacin, or (iv) vehicle. After 28 days the lesion site was examined quantitatively by light microscopical image analysis. The lesion of vehicle-treated control animals showed large cavities, extensive infiltration by debris-engorged macrophages, and relatively few axons. Treatment with LPS or ImuVert significantly reduced the degree of cavitation and increased the number of cells and axons in the lesion. Treatment with LPS+Indomethacin was significantly more effective than treatment with LPS alone, while treatment with Indomethacin alone was ineffective. To test whether the histopathological differences between treated and control rats might be reflected in functional improvement, rats were subjected to a contusion (weight-drop) injury and their walking ability was quantified by the Tarlov scale for 28 days postoperatively. Treatment with LPS+Indomethacin significantly improved locomotor function of animals subjected to a moderate (1.25 g x 20 cm) injury. We conclude that tissue repair and functional recovery after spinal cord injury are enhanced by combined treatment with agents that promote the secretory activities of the nonneuronal cells and that inhibit prostaglandin synthesis. These results indicate that the search for more effective treatments should include studies on combinations of drugs having different pharmacological specificities.