Polyunsaturated fatty acids in amyotrophic lateral sclerosis: role of DHA, peroxidative modifications and sexual dimorphism
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Date
2014-10-24
Authors
Cacabelos Barral, Daniel
Other authors
Portero Otín, Manuel
Pamplona Gras, Reinald
Universitat de Lleida. Departament de Medicina Experimental
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Abstract
In the present work we focus into the potential relevance of PUFAs in some models and human
samples from patients suffering amyotrophic lateral sclerosis (ALS). Due to its pathological
implication, oxidative stress was our first goal. We started from simple oxidative methodology
screening to search for an antioxidant substance (among 21 different candidates) available in a
Mediterranean diet. The results demonstrated high heterogeneity in carbonyl (measured by
DNP) accumulation, regarding the oxidative source, substrate suffering it and the antioxidant
structure. Further, thanks to GC/MS and LCQTOF, we detailed the protection over specific
accrual of protein and lipid peroxidation markers as well as lipid profile modifications (as % of
total fatty acids -FA) in oxLDL thanks to those dietary compounds. Moreover, we demonstrated
its in vitro relevance, in terms of survival, when two cell lines (HMEC-1, HepG2) were treated
with this oxidized (and protected) compounds, and finally address in vivo importance of those
findings, demonstrating decreased carbonyl and oxidative accumulation in hamsters under an
atherogenic diet supplemented with antioxidants. Once described the protective effect of
antioxidants and specific signatures found regarding lipid oxidation markers, we extend the
study focusing in different ALS samples. From previous work, we demonstrated an altered
docosohexaenoic acid (DHA) composition in different location for patients suffering sporadic
ALS. Hence, we though necessary to define whether the enzymatic machinery aimed to
synthesize DHA from its precursors, are affected in sALS. Interestingly, we found a tissue
specific variation (spinal cord vs cortex), compatible with our previous FA results. Further,
thanks to inmunohistochemistry, differential involvement was unveiled for motor neurons (MN)
and surrounding glia. Therefore, trying to depict cellular contribution, we switch to a neuronal
model (N2A under oxidative stressors and/or aggregation-prone-TDP-43) and a tissular one
(OT). There, we showed decreased desaturase (Δ6) and drebrin expression as well as increased
DHA synthesis and an unreported inverse correlation of drebrin loss and aberrant p-TDP-43
expression under oxidative conditions in the cell culture. In the OT model, lipidomic analysis
showed specific accretion of 8-iso-PGF2α and NPD1 as well as increased DHA (and
dramatically decreased precursors) and reduced AA concentrations (GC measured). Analysis of
slice O2 consumption showed decreased O2 levels under excitotoxic treatment and alleviation by
antioxidant (tocopherol) addition. Treatment of OT slices with Ω-3 precursors (better than final
products) and DHA plus tocopherol ameliorated MNs number. Finally, we wanted to disclose
PUFA’s implication and phenothype of an animal model (SODG93A) under a dietary
intervention with opposed FA unsaturation levels. Not surprisingly, FA profile was difficult to
be altered in nervous system, although subtle specific variations were found. More importantly,
differences in survival and clinical manifestations, UPR (Ubiquitin inclusions), mt-DNA (8-
oxo-dG) and protein oxidative modifications revealed sex as a relevant factor in lipid handling
for this model. Hence, whereas male under a low PUFA diet showed increased survival, females
lack this beneficial outcome. Last but not least, we wanted to dig deeper regarding this sexual
dimorphism. For this purpose, we focused in mitochondria and analyzed spinal cord oxygen
consumption, oxidative damage to proteins and lipid profile along disease progression and also
in a neuronal model (N2A overexpressing SODG93A, treated with 17β-estradiol).We could
demonstrate a clear sexual implication, with females having late onset clinical symptoms
concomitant to an upgraded mitochondrial function and lower protein and mitochondrial
damaged proteins compared with males. Finally, to further confirmed steroid potential as a
protective element in disease progression, in vitro estradiol pretreatmet of N2A showed
increased oxygen consumption, with no relation with the mitochondrial complex expression.