In multiple sclerosis (MS) a process of white matter degradation leading to demyelination is observed. Oxidative stress, inflammation, apoptosis, necrosis and/or autophagy result together into a progressive loss of oligodendrocytes. 7-ketocholesterol (7KC), found increased in the cerebrospinal fluid of MS patients, triggers a rupture of RedOx homeostasis associated with mitochondrial dysfunctions, aptoptosis and autophagy (oxiapoptophagy) in cultured murine oligodendrocytes (158N). α-tocopherol is able to mild the alterations induced by 7KC partially restoring the cellular homeostasis. In presence of 7KC, the amount of adherent 158N cells was decreased and oxidative stress was enhanced. An increase of caspase-3 and PARP degradation (evidences of apoptosis), and an increased LC3-II/LC3-I ratio (criterion of autophagy), were detected. These events were associated with a decrease of the mitochondrial membrane potential (ΔΨm) and by a decrease of oxidative phosphorylation revealed by reduced NAD+ and ATP. The cellular lactate was higher while pyruvate, citrate, fumarate, succinate (tricarboxylic acid (TCA) cycle intermediates) were significantly reduced in exposed cells, suggesting that an impairment of mitochondrial respiratory functions could lead to an increase of lactate production and to a reduced amount of ATP and acetyl-CoA available for the anabolic pathways. The concentration of sterol precursors lathosterol, lanosterol and desmosterol were significantly reduced together with satured and unsatured long chain fatty acids (C16:0 - C18:0, structural elements of membrane phospholipids). Such reductions were milder with α-tocopherol. It is likely that the cell death induced by 7KC is associated with mitochondrial dysfunctions, including alterations of oxidative phosphorylation, which could result from lipid anabolism dysfunctions, especially on TCA cycle intermediates. A better knowledge of mitochondrial associated dysfunctions triggered by 7KC will contribute to bring new information on the demyelination processes which are linked with oxidative stress and lipid peroxidation, especially in MS.

The brain, which is a cholesterol-rich organ, can be subject to oxidative stress in a variety of pathophysiological conditions, age-related diseases and some rare pathologies. This can lead to the formation of 7-ketocholesterol (7KC), a toxic derivative of cholesterol mainly produced by auto-oxidation. So, preventing the neuronal toxicity of 7KC is an important issue to avoid brain damage. As there are numerous data in favor of the prevention of neurodegeneration by the Mediterranean diet, this study aimed to evaluate the potential of a series of polyphenols (resveratrol, RSV; quercetin, QCT; and apigenin, API) as well as ω3 and ω9 unsaturated fatty acids (α-linolenic acid, ALA; eicosapentaenoic acid, EPA; docosahexaenoic acid, DHA, and oleic acid, OA) widely present in this diet, to prevent 7KC (50 µM)-induced dysfunction of N2a neuronal cells. When polyphenols and fatty acids were used at non-toxic concentrations (polyphenols: ≤6.25 µM; fatty acids: ≤25 µM) as defined by the fluorescein diacetate assay, they greatly reduce 7KC-induced toxicity. The cytoprotective effects observed with polyphenols and fatty acids were comparable to those of α-tocopherol (400 µM) used as a reference. These polyphenols and fatty acids attenuate the overproduction of reactive oxygen species and the 7KC-induced drop in mitochondrial transmembrane potential (ΔΨm) measured by flow cytometry after dihydroethidium and DiOC6(3) staining, respectively. Moreover, the studied polyphenols and fatty acids reduced plasma membrane permeability considered as a criterion for cell death measured by flow cytometry after propidium iodide staining. Our data show that polyphenols (RSV, QCT and API) as well as ω3 and ω9 unsaturated fatty acids (ALA, EPA, DHA and OA) are potent cytoprotective agents against 7KC-induced neurotoxicity in N2a cells. Their cytoprotective effects could partly explain the benefits of the Mediterranean diet on human health, particularly in the prevention of neurodegenerative diseases.

Argan oil is widely used in Morocco in traditional medicine. Its ability to treat cardiovascular diseases is well-established. However, nothing is known about its effects on neurodegenerative diseases, which are often associated with increased oxidative stress leading to lipid peroxidation and the formation of 7-ketocholesterol (7KC) resulting from cholesterol auto-oxidation. As 7KC induces oxidative stress, inflammation and cell death, it is important to identify compounds able to impair its harmful effects. These compounds may be either natural or synthetic molecules or mixtures of molecules such as oils. In this context: (i) the lipid profiles of dietary argan oils from Berkane and Agadir (Morocco) in fatty acids, phytosterols, tocopherols and polyphenols were determined by different chromatographic techniques; and (ii) their anti-oxidant and cytoprotective effects in 158N murine oligodendrocytes cultured with 7KC (25-50 µM; 24 h) without and with argan oil (0.1% v/v) or α-tocopherol (400 µM, positive control) were evaluated with complementary techniques of cellular and molecular biology. Among the unsaturated fatty acids present in argan oils, oleate (C18:1 n-9) and linoleate (C18:1 n-6) were the most abundant; the highest quantities of saturated fatty acids were palmitate (C16:0) and stearate (C18:0). Several phytosterols were found, mainly schottenol and spinasterol (specific to argan oil), cycloartenol, β-amyrin and citrostadienol. α- and γ-tocopherols were also present. Tyrosol and protocatechic acid were the only polyphenols detected. Argan and extra virgin olive oils have many compounds in common, principally oleate and linoleate, and tocopherols. Kit Radicaux Libres (KRL) and ferric reducing antioxidant power (FRAP) tests showed that argan and extra virgin olive oils have anti-oxidant properties. Argan oils were able to attenuate the cytotoxic effects of 7KC on 158N cells: loss of cell adhesion, cell growth inhibition, increased plasma membrane permeability, mitochondrial, peroxisomal and lysosomal dysfunction, and the induction of oxiapoptophagy (OXIdation + APOPTOsis + autoPHAGY). Altogether, our data obtained in 158N oligodendrocytes provide evidence that argan oil is able to counteract the toxic effects of 7KC on nerve cells, thus suggesting that some of its compounds could prevent or mitigate neurodegenerative diseases to the extent that they are able to cross the blood-brain barrier.

The Asteraceae family is economically very important, because many of these plants are grown mainly for their food value, such as lettuce (Lactuca), chicory (Cichorium), and sunflower (Heliantus aminus). One of the typical properties of this family, which includes milk thistle (Sylibum marianum), is the richness of the oil in various compounds (flavonoids, alkaloids, tocopherols, and unsaturated fatty acids). Currently, and for the coming decades, age-related diseases, including neurodegenerative diseases, are a major public health problem. Preventing their appearance or opposing their evolution is a major objective. In this context, the cytoprotective activities of milk thistle seed oil produced in Tunisia were studied on the 158N model using 7-ketocholesterol (7KC) and 24S-hydroxycholesterol (24S) as cytotoxic agents. 7KC and 24S were used because they can be increased in the brain and body fluids of patients with major age-related neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases. In order to evaluate the cytoprotective properties of milk thistle seed oil, complementary techniques of microscopy, flow cytometry, and biochemistry were used. The chemical composition of milk thistle seed oil has also been determined by various chromatography techniques. Milk thistle seed oils from different area of Tunisia are rich in tocopherols and are strongly antioxidant according to various biochemical tests (KRL (Kit Radicaux Libres), FRAP (Ferric Reducing Antioxidant Power), and DPPH (2,2-diphenyl-1-picrylhydrazyl)). The main fatty acids are linoleic acid (C18:2 n-6) and oleic acid (C18:1 n-9). The main polyphenols identified are homovanillic acid, p-coumaric acid, quercetin, and apigenin, with a predominance of vanillic acid. On 158N cells, milk thistle seed oil attenuates the cytotoxicity of 7KC and 24S including: loss of cell adhesion, increased plasma membrane permeability, mitochondrial dysfunction, overproduction of reactive oxygen species, induction of apoptosis, and autophagy. The attenuation of the cytotoxicity of 7KC and 24S observed with the milk thistle seed oil is in the order of that observed with α-tocopherol used as a positive control. In the presence of nigella seed oil, considered potentially cytotoxic, no cytoprotective effects were observed. Given the chemical characteristics, antioxidant properties, and cytoprotective activities of milk thistle seed oil, our results highlight the potential benefit of this oil for human health.