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Updated: Mar 3, 2026
Mitochondrial Preparation from Microglia for Glycan Analysis
Published on: May 30, 2025
Proteomic anaysis of aged microglia: shifts in transcription, bioenergetics, and nutrient response
Antwoine Flowers1,2, Harris Bell-Temin3, Ahmad Jalloh1,2
1Department of Neurosurgery and Brain Repair, USF Health Morsani College of Medicine, University of South Florida, 12901 Bruce B Downs Blvd., Campus Box MDC-78, Tampa, FL, 33570, USA.
Background:
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.
Methods:
This study utilizes mass spectrometry-based proteomics to compare primary microglia from young and aged animals.
Results:
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.
Conclusions:
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.
Insights
Aging microglia exhibit altered cellular metabolism and inflammatory signaling, linked to decreased RICTOR levels. Modulating RICTOR may offer therapeutic strategies for age-related CNS diseases.
Area of Science:
- Neuroscience
- Immunology
- Aging Research
Background:
- Age is a primary risk factor for numerous diseases, influencing disease progression and intervention strategies.
- Aged microglia display a pro-inflammatory phenotype and diminished anti-inflammatory functions, impacting CNS plasticity and repair.
- Understanding age-related changes in microglia is crucial for neurodegenerative disease research.
Purpose of the Study:
- To compare the proteomic profiles of primary microglia from young and aged animals.
- To identify key molecular pathways and regulators altered in aged microglia.
- To investigate the role of RICTOR in age-associated microglial dysfunction.
Main Methods:
- Mass spectrometry-based proteomics was employed to analyze primary microglia.
- Bioinformatic analysis identified altered protein clusters and upstream regulators.
- Genetic manipulation of microglial cell lines (BV2) was used to validate findings.
Main Results:
- Proteomic analysis revealed age-dependent alterations in inflammatory signaling, mitochondrial function, and cellular metabolism.
- Rapamycin-insensitive companion of mTOR (RICTOR) was identified as a novel upstream regulator of these age-related changes.
- Aged microglia showed decreased mTORC2-dependent AKT S473 phosphorylation, and diminished RICTOR in BV2 cells mimicked aged microglia phenotype.
Conclusions:
- Aging microglia exhibit metabolic and energy regulation changes potentially underlying inflammatory alterations.
- Targeting RICTOR presents a potential therapeutic avenue to attenuate detrimental aging microglia characteristics.
- Successful modulation of RICTOR could delay or reduce the progression of diseases involving chronic inflammation.
