Proteomic analysis of aged microglia: shifts in transcription, bioenergetics, and nutrient response

Antwoine Flowers, Harris Bell-Temin, Ahmad Jalloh, Stanley M. Stevens, Paula C. Bickford

Age is the primary risk factor for many diseases. As such, age is a critical co-factor for examination in order to understand the progression and potential intervention in disease progression. Studies examining both the phenotype and transcriptome of aged microglia demonstrated a propensity for the development of a pro-inflammatory phenotype. Less well studied is the concomitant blunting of anti-inflammatory aspects of microglial function with age which also impact plasticity and repair in the CNS. This study utilizes mass spectrometry-based proteomics to compare primary microglia from young and aged animals. This study revealed alterations in three clusters of inter-related proteins. The three pathways were inflammatory signaling, mitochondrial function, and cellular metabolism. Analysis of these clusters identified the protein rapamycin-insensitive companion of mTOR (RICTOR), a component of the mTORC2 complex, as a novel upstream regulator of several biological functions that are altered with age and potentially linked to phenotype development. A decrease in mTORC2-dependent AKT S473 phosphorylation, as assessed by insulin growth factor (IGF) treatment, was observed in aged microglia. This novel finding was confirmed by genetic manipulation of the microglial cell line. BV2 cells with diminished RICTOR displayed a phenotype that was strikingly similar to that of aged microglia. This finding is particularly relevant as the mTOR pathway already has a number of pharmacological modulators used clinically. The results suggest that microglia from aged mice show changes in cellular metabolism and energy regulation that might underlie the alterations in inflammatory signaling. Modulation of one pathway identified in our bioinformatic analysis, RICTOR, may provide an avenue by which deleterious aspects of the aging microglia can be attenuated. If successful, this could mean potentially delaying or diminishing the progress of diseases for which progressive inflammation is involved.