A comparative study of the effects of alpha-synuclein and S100A9 on microglial bioenergetics and inflammatory response
A comparative study of the effects of alpha-synuclein and S100A9 on microglial bioenergetics and inflammatory response
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Original abstract
The phenotypic plasticity and functional diversity of microglia are strongly influenced by changes in their energy metabolism. These metabolic alterations play a pivotal role in regulating neuroinflammation, particularly in response to pathological stimuli such as amyloid proteins, which are implicated in the pathogenesis of neurodegenerative diseases. Recent studies have shown that alpha-synuclein (αSyn) and another highly amyloidogenic protein – S100A9 are abundantly expressed in microglia and neurons intracellularly and deposited extracellularly in both Alzheimer's and Parkinson's disease patients. However, the precise molecular mechanisms of their neurotoxicity and the microglial metabolic changes during the inflammatory response remain to be elucidated. Thus, our research aimed to investigate and compare the effects of αSyn and S100A9 on the energy-producing pathways of the microglial cells and their immune response. In this study, BV-2 mouse microglial cells were cultured and exposed to nanomolar concentrations of pre-aggregated recombinant amyloid proteins – αSyn or S100A9 for 24 h. Following incubation, changes in microglial energy metabolism were assessed by measuring oxygen consumption rates and glycolytic activity using a high-resolution respirometer Oroboros O2k and O2k-pH ISE-Module, respectively. The inflammatory response of microglial cells to amyloid aggregates was evaluated by measuring TNF-α levels in the cell culture medium using ELISA, nitric oxide (NO) levels were quantified using the Griess assay, and extracellular reactive oxygen species (ROS) production was measured with the Amplex Red assay. We found that exposure of BV-2 cells to αSyn and S100A9 negatively affected mitochondrial respiration. However, the mechanisms by which they induced such effects differed. αSyn inhibited oxidative phosphorylation by decreasing oxidation of the mitochondrial complex I-linked substrates without altering uncoupled respiration. In contrast, S100A9 caused the dysfunction of the mitochondrial electron transfer system, leading to a reduction in the oxidation of both mitochondrial complex I- and II-linked substrates. Notably, BV-2 cells exhibited highly increased glycolytic activity in response to both αSyn and S100A9, suggesting a compensatory switch to anaerobic glycolysis. Moreover, S100A9 significantly increased TNF-α secretion after 24 h, while extracellular NO and H2O2 levels remained unchanged. In comparison, αSyn did not affect TNF-α concentration in the cell growth medium, NO levels, or ROS production after 24 h of incubation with BV-2 cells. Overall, our data suggest that extracellular αSyn and S100A9 oligomers cause a shift in microglial energy metabolism and induce distinct inflammatory responses. These findings have important implications for defining microglial contributions to brain energy metabolism and understanding neuroinflammation in neurodegenerative diseases.