Induction of Mitochondrial Changes Associated with Oxidative Stress on Very Long Chain Fatty Acids (C22:0, C24:0, or C26:0)-Treated Human Neuronal Cells (SK-NB-E)

Abstract

In Alzheimer's disease, lipid alterations point towards peroxisomal dysfunctions. Indeed, a cortical accumulation of saturated very long chain fatty acids (VLCFAs: C22:0, C24:0, C26:0), substrates for peroxisomal β-oxidation, has been found in Alzheimer patients. This study was realized to investigate the effects of VLCFAs at the mitochondrial level since mitochondrial dysfunctions play crucial roles in neurodegeneration. On human neuronal SK-NB-E cells treated with C22:0, C24:0, or C26:0 (0.1–20 μM; 48 h), an inhibition of cell growth and mitochondrial dysfunctions were observed by cell counting with trypan blue, MTT assay, and measurement of mitochondrial transmembrane potential (Δψm) with DiOC6(3). A stimulation of oxidative stress was observed with DHE and MitoSOX used to quantify superoxide anion production on whole cells and at the mitochondrial level, respectively. With C24:0 and C26:0, by Western blotting, lower levels of mitochondrial complexes III and IV were detected. After staining with MitoTracker and by transmission electron microscopy used to study mitochondrial topography, mass and morphology, major changes were detected in VLCFAs treated-cells: modification of the cytoplasmic distribution of mitochondria, presence of large mitochondria, enhancement of the mitochondrial mass. Thus, VLCFAs can be potential risk factors contributing to neurodegeneration by inducing neuronal damages via mitochondrial dysfunctions.

Impact of C24:0 on actin-microtubule interaction in human neuronal SK-N-BE cells: evaluation by FRET confocal spectral imaging microscopy after dual staining with rhodamine-phalloidin and tubulin tracker green

“…1 C). In agreement with previous observations, in the presence of C24:0 used at 20 μM, most of the cells were floating in the culture medium (Zarrouk et al, 2012). These functional and morphological changes were correlated with ultrastructural modifications inside the cells as shown by TEM.…”

“…In the present study, we characterized the effects of C24:0 on human neuronal SK-N-BE cells. As various cytotoxic effects associated with cell death have been observed in different cell types treated with C24:0 (Kahn et al, 2011;Baarine et al, 2012;Zarrouk et al, 2012;Hein et al, 2008), we wondered whether this fatty acid is able to induce cytoskeleton modifications. Using various flow cytometric, microscopic and biochemical methods, we demonstrated changes in actin and A.…”

“…The shape index evaluated by phase contrast microscopy was determined as previously described (Zarrouk et al, 2012). Digitized images were obtained with a camera (Axiocam ICm1, Zeiss).…”

“…Increased levels of VLCFAs have also been found in the plasma and red blood cells of patients with AD (Zarrouk, 2013). Notably, in glial and neuronal cells (Hein et al, 2008;Baarine et al, 2012;Zarrouk et al, 2012), it was previously reported that tetracosanoic acid (C24:0) (used at concentrations in the 5-20 μM range) is capable of triggering mitochondrial dysfunctions and oxidative stress associated with a non-apoptotic mode of cell death. These are regarded as important events leading to the development of AD (Kahn et al, 2011;Silva et al, 2012;von Bernhardi et al, 2012).…”

“…We focused on the activity of C24:0 for the following reasons: it is found at higher concentrations than C22:0 and C26:0 in the plasma and tissues of patients with peroxisomopathies (Kemp et al, 2011;Baarine et al, 2012), and it has also been detected at increased levels in the plasma and cortex of patients with AD (Kou et al, 2011;Zarrouk, 2013;Zarrouk et al, 2015). Whereas this fatty acid is known to have pronounced side effects, contributing to cell death of various neural cells (Hein et al, 2008;Baarine et al, 2012;Zarrouk et al, 2012), its effects on the cytoskeleton are still unknown. It was thus considered of interest to establish the precise impact of C24:0, at cytotoxic concentrations, on the organization of microtubules and actin in human neuronal SK-N-BE cells.…”