P2‐029: A Combination of Dietary Precursors and Cofactors for Membrane Synthesis Enhances Neurite Outgrowth in Vitro

Phospholipids are key components of membranes in neurites and also at the level of synapses. It has been shown that supplementation with a combination of docosahexaenoic acid (DHA), uridine and choline, precursors of phospholipid synthesis, supports membrane synthesis and neurite outgrowth by stimulating the cytidine diphosphate-choline (CDP-choline) pathway. It has also been demonstrated that the addition of other cofactors of phospholipid synthesis, can lead to beneficial synergistic effects, for example on amyloid processing or muscarinic receptor activation. Supplementation with a specific nutrient combination, known as Fortasyn® Connect (FC), which contains uridine, choline, DHA, eicosapentaenoic acid (EPA), phospholipids, folic acid, vitamins B12, B6, C, and E, and selenium, has shown beneficial effects on brain connectivity and memory performance in early Alzheimer’s disease (AD) patients. To assess the additional benefit of providing nutritional cofactors alongside precursors for membrane formation on neurite outgrowth and synapse formation compared to the effects of the precursors alone. Pheochromocytoma cells were seeded on type IV human collagen coated black colored 96-well plates and were supplemented 48 hours with medium containing Nerve Growth Factor, with or without FC nutrients. Subsequently, FITC and DAPI images were obtained for quantifying neurite outgrowth using High Content Imaging. For quantitative PCR, cells were seeded in 6-well plates and supplemented 24 hours with medium containing Nerve Growth Factor, with or without FC nutrients. RNA was extracted and real time PCR was performed in a 384 wells format using primers for various genes involved in neurite outgrowth. Supplementation with FC leads to enhanced neurite outgrowth as compared to combined supplementation with DHA, uridine and choline. FC markedly increases neurite length and the number of neurites per cell. Furthermore, quantitative PCR (qPCR) data show an upregulation of genes encoding with neurite structural proteins and pre-synaptic markers. Addition of nutritional cofactors to phospholipid precursors, as in the FC combination, has optimum effects on neurite outgrowth as compared to precursor supplementation alone.