Combination of Secondary Plant Metabolites and Micronutrients Improves Mitochondrial Function in a Cell Model of Early Alzheimer's Disease.

Alzheimer's disease (AD) is characterized by excessive formation of beta-amyloid peptides (Aß), mitochondrial dysfunction, enhanced production of reactive oxygen species (ROS), and altered glycolysis. Since the disease is currently not curable, preventive and supportive approaches are in the focus of science. Based on studies of promising single substances, the present study used a mixture (cocktail, SC) of compounds consisting of hesperetin (HstP), magnesium-orotate (MgOr), and folic acid (Fol), as well as the combination (KCC) of caffeine (Cof), kahweol (KW) and cafestol (CF). For all compounds, we showed positive results in SH-SY5Y-APP695 cells-a model of early AD. Thus, SH-SY5Y-APP695 cells were incubated with SC and the activity of the mitochondrial respiration chain complexes were measured, as well as levels of ATP, Aß, ROS, lactate and pyruvate. Incubation of SH-SY5Y-APP695 cells with SC significantly increased the endogenous respiration of mitochondria and ATP levels, while Aß1-40 levels were significantly decreased. Incubation with SC showed no significant effects on oxidative stress and glycolysis. In summary, this combination of compounds with proven effects on mitochondrial parameters has the potential to improve mitochondrial dysfunction in a cellular model of AD.

Effects of Combining Biofactors on Bioenergetic Parameters, Aß Levels and Survival in Alzheimer Model Organisms.

Increased amyloid beta (Aß) levels and mitochondrial dysfunction (MD) in the human brain characterize Alzheimer disease (AD). Folic acid, magnesium and vitamin B6 are essential micro-nutrients that may provide neuroprotection. Bioenergetic parameters and amyloid precursor protein (APP) processing products were investigated in vitro in human neuroblastoma SH-SY5Y-APP695 cells, expressing neuronal APP, and in vivo, in the invertebrate Caenorhabditis elegans (CL2006 & GMC101) expressing muscular APP. Model organisms were incubated with either folic acid and magnesium-orotate (ID63) or folic acid, magnesium-orotate and vitamin B6 (ID64) in different concentrations. ID63 and ID64 reduced Aß, soluble alpha APP (sAPPα), and lactate levels in SH-SY5Y-APP695 cells. The latter might be explained by enhanced expression of lactate dehydrogenase (LDHA). Micronutrient combinations had no effects on mitochondrial parameters in SH-SY5Y-APP695 cells. ID64 showed a significant life-prolonging effect in C. elegans CL2006. Incubation of GMC101 with ID63 significantly lowered Aß aggregation. Both combinations significantly reduced paralysis and thus improved the phenotype in GMC101. Thus, the combinations of the tested biofactors are effective in pre-clinical models of AD by interfering with Aß related pathways and glycolysis.

Effects of Pesticides on Longevity and Bioenergetics in Invertebrates-The Impact of Polyphenolic Metabolites.

Environmentally hazardous substances such as pesticides are gaining increasing interest in agricultural and nutritional research. This study aims to investigate the impact of these compounds on the healthspan and mitochondrial functions in an invertebrate in vivo model and in vitro in SH-SY5Y neuroblastoma cells, and to investigate the potential of polyphenolic metabolites to compensate for potential impacts. Wild-type nematodes (Caenorhabditis elegans, N2) were treated with pesticides such as pyraclostrobin (Pyr), glyphosate (Gly), or fluopyram (Fluo). The lifespans of the nematodes under heat stress conditions (37 °C) were determined, and the chemotaxis was assayed. Energetic metabolites, including adenosine triphosphate (ATP), lactate, and pyruvate, were analyzed in lysates of nematodes and cells. Genetic expression patterns of several genes associated with lifespan determination and mitochondrial parameters were assessed via qRT-PCR. After incubation with environmentally hazardous substances, nematodes were incubated with a pre-fermented polyphenol mixture (Rechtsregulat®Bio, RR) or protocatechuic acid (PCA) to determine heat stress resistance. Treatment with Pyr, Glyph and Fluo leads to dose-dependently decreased heat stress resistance, which was significantly improved by RR and PCA. The chemotaxes of the nematodes were not affected by pesticides. ATP levels were not significantly altered by the pesticides, except for Pyr, which increased ATP levels after 48 h leads. The gene expression of healthspan and mitochondria-associated genes were diversely affected by the pesticides, while Pyr led to an overall decrease of mRNA levels. Over time, the treatment of nematodes leads to a recovery of the nematodes on the mitochondrial level but not on stress resistance on gene expression. Fermented extracts of fruits and vegetables and phenolic metabolites such as PCA seem to have the potential to recover the vitality of C. elegans after damage caused by pesticides.

Hesperetin Nanocrystals Improve Mitochondrial Function in a Cell Model of Early Alzheimer Disease.

Mitochondrial dysfunction represents a hallmark of both brain aging and age-related neurodegenerative disorders including Alzheimer disease (AD). AD-related mitochondrial dysfunction is characterized by an impaired electron transport chain (ETC), subsequent decreased adenosine triphoshpate (ATP) levels, and elevated generation of reactive oxygen species (ROS). The bioactive citrus flavanone hesperetin (Hst) is known to modulate inflammatory response, to function as an antioxidant, and to provide neuroprotective properties. The efficacy in improving mitochondrial dysfunction of Hst nanocrystals (HstN) with increased bioavailability has not yet been investigated. Human SH-SY5Y cells harboring neuronal amyloid precursor protein (APP695) acted as a model for the initial phase of AD. MOCK-transfected cells served as controls. The energetic metabolite ATP was determined using a luciferase-catalyzed bioluminescence assay. The activity of mitochondrial respiration chain complexes was assessed by high-resolution respirometry using a Clarke electrode. Expression levels of mitochondrial respiratory chain complex genes were determined using quantitative real-time polymerase chain reaction (qRT-PCR). The levels of amyloid ß-protein (Aß1-40) were measured using homogeneous time-resolved fluorescence (HTRF). ROS levels, peroxidase activity, and cytochrome c activity were determined using a fluorescence assay. Compared to pure Hst dissolved in ethanol (HstP), SH-SY5Y-APP695 cells incubated with HstN resulted in significantly reduced mitochondrial dysfunction: ATP levels and respiratory chain complex activity significantly increased. Gene expression levels of RCC I, IV, and V were significantly upregulated. In comparison, the effects of HstN on SY5Y-MOCK control cells were relatively small. Pure Hst dissolved in ethanol (HstP) had almost no effect on both cell lines. Neither HstN nor HstP led to significant changes in Aß1-40 levels. HstN and HstP were both shown to lower peroxidase activity significantly. Furthermore, HstN significantly reduced cytochrome c activity, whereas HstP had a significant effect on reducing ROS in SH-SY5Y-APP695 cells. Thus, it seems that the mechanisms involved may not be linked to altered Aß production. Nanoflavonoids such as HstN have the potential to prevent mitochondria against dysfunction. Compared to its pure form, HstN showed a greater effect in combatting mitochondrial dysfunction. Further studies should evaluate whether HstN protects against age-related mitochondrial dysfunction and thus may contribute to late-onset AD.