Application of Antibodies to Neuronally Expressed Nogo-A Increases Neuronal Survival and Neurite Outgrowth.

Nogo-A, a glycoprotein expressed in oligodendrocytes and central nervous system myelin, inhibits regeneration after injury. Antibodies against Nogo-A neutralize this inhibitory activity, improve locomotor recovery in spinal cord-injured adult mammals, and promote regrowth/sprouting/saving of damaged axons beyond the lesion site. Nogo-A is also expressed by neurons. Complete ablation of Nogo-A in all cell types expressing it has been found to lead to recovery in some studies but not in others. Neuronal ablation of Nogo-A reduces axonal regrowth after injury. In view of these findings, we hypothesized that, in addition to neutralizing Nogo-A in oligodendrocytes and myelin, Nogo-A antibodies may act directly on neuronal Nogo-A to trigger neurite outgrowth and neuronal survival. Here, we show that polyclonal and monoclonal antibodies against Nogo-A enhance neurite growth and survival of cultured cerebellar granule neurons and increase expression of the neurite outgrowth-promoting L1 cell adhesion molecule and polysialic acid. Application of inhibitors of signal transducing molecules, such as c-src, c-fyn, protein kinase A, and casein kinase II reduce antibody-triggered neurite outgrowth. These observations indicate that the recovery-promoting functions of antibodies against Nogo-A may not only be due to neutralizing Nogo-A in oligodendrocytes and myelin, but also to their interactions with Nogo-A on neurons.

Enhanced Neuronal Survival and Neurite Outgrowth Triggered by Novel Small Organic Compounds Mimicking the LewisX Glycan.

Glycosylation fine-tunes signal transduction of adhesion molecules during neural development and supports synaptic plasticity and repair after injury in the adult nervous system. One abundantly expressed neural glycan is LewisX (LeX). Although it is known that its expression starts at the formation of the neural tube during the second embryonic week in the mouse and peaks during the first postnatal week, its functional relevance is only rudimentarily understood. To gain better insights into the functions of this glycan, we identified small organic compounds that mimic structurally and functionally this glycan glycosidically linked to several neural adhesion molecules. Mimetic compounds were identified by competitive enzyme-linked immunosorbent assay (ELISA) using the LeX-specific monoclonal antibodies L5 and SSEA-1 for screening a library of small organic molecules. In this assay, antibody binding to substrate-coated LeX glycomimetic peptide is measured in the presence of compounds, allowing identification of molecules that inhibit antibody binding and thereby mimic LeX. Gossypol, orlistat, ursolic acid, folic acid, and tosufloxacin inhibited antibody binding in a concentration-dependent manner. With the aim to functionally characterize the molecular consequences of the compounds' actions, we here present evidence that, at nM concentrations, the mimetic compounds enhance neurite outgrowth and promote neuronal survival of cultured mouse cerebellar granule cells via, notably, distinct signal transduction pathways. These findings raise hopes that these LeX mimetics will be powerful tools for further studying the functions of LeX and its effects in acute and chronic nervous system disease models. It is worth mentioning in this context that the LeX compounds investigated in the present study have been clinically approved for different therapies.