Phosphatidylinositol transfer protein alpha and its role in neurodegeneration

Abstract

Selective neuronal loss is a prominent feature in neurodegenerative disorders. Recently, a link between neurodegeneration and a deficiency in the protein phosphatidylinositol transfer protein alpha (PI-TPalpha) has been demonstrated. In this context it is of importance that fibroblasts overexpressing PI-TPalpha (SPIalpha cells) are known to produce and secrete bio-active lipid factors that protect fibroblasts against UV-induced apoptosis. Together with the observation that PI-TPalpha expression is highest in the central nervous system (CNS), these findings imply that PI-TPalpha expression, and consequently its function, is essential in CNS maintenance. In this study, this hypothesis is elaborated and investigated in more detail. We showed that SPIalpha cells produce and secrete PI-TPalpha-dependent lipid factors capable of protecting primary rat, spinal cord-derived, motor neuron cultures. Protection was diminished in the presence of suramin, an antagonist of the G-protein coupled receptor family. Immunocytochemistry studies showed that PI-TPalpha is expressed in both primary astrocyte and motor neuron cultures. To investigate whether the high expression level of PI-TPalpha in the CNS is associated with the cellular production of PI-TPalpha-dependent survival-enhancing activity and, hence, neuroprotection, several in vitro cell models have been developed. Analysis of the PI-TPalpha level in different CNS-derived cell cultures showed that PI-TPalpha expression is relatively high in astrocytes (primary cultures as well as the C6 astroglioma cell line). The level of PI-TPalpha in neuronal cells was significantly lower. Ultimately, a new model has been introduced in which the C6 astroglioma cells served as survival factor-producing cells and the motor neuron NSC-34 cells as a target. An intermediate model was proposed with C6 astroglioma cells in combination with the apoptosis-sensitive SPIbeta fibroblast cell line as a target. This latter model was used to characterise the survival-enhancing effect of the conditioned medium of C6 astroglioma cells (CMc6). CMc6 as well as a lipid extract derived thereof, enhanced survival of SPIbeta cells under serum-free conditions. This effect was attenuated when CMc6 was co-incubated with SR141716A (rimonabant), an antagonist of the cannabinoid-1 receptor (CB1R). Mass spectrometry analysis revealed low levels eicosanoid precursors in CMc6, indicating an active eicosanoid metabolism in C6 astroglioma cells. In addition, the presence of three PI-TPalpha- and cyclooxygenase-2-dependent compounds in CMc6 was demonstrated. The nature of these compounds showed resemblance to endocannabinoids. Using RNA interference techniques, the PI-TPalpha expression level in C6 astroglioma cells was downregulated to investigate whether the survival-enhancing effect of CMc6 was a PI-TPalpha-dependent phenomenon. The survival-enhancing effect of the CM derived from these cells was tested on SPIbeta as well as NSC-34 cells and appeared to be reduced when compared to normal CMc6. Taken together, from these data it can be concluded that, at least in C6 astroglioma cells, and possibly also in astrocytes, PI-TPalpha is involved in the production of lipid factors capable of enhancing the survival of an apoptosis-sensitive fibroblast cell line and motor neuron-like cell line. As indicated above, although the identity of these factors remains largely unknown, as yet these lipid factors may be classified as arachidonic acid-derived endocannabinoids, acting on the CB1R or a CB1-like receptor