Detection of Autophagy in Human CD4+ T cells

Authors

• L Ferrucci
• T Srinivas
• A Bektas
• R Sen

Abstract

Autophagy is a highly conserved metabolic process that plays a key role in maintaining cellular homeostasis through lysosome-mediated protein degradation, amino acid recycling, and protein biogenesis. While abnormalities in autophagic function have been associated with several age-related human diseases, a comprehensive understanding of the mechanisms governing age-related autophagic changes is still lacking. The complex and dynamic nature of autophagy renders it difficult to accurately monitor and quantify. Simple detection of autophagosome accumulation is insufficient to determine autophagic activity, as an increase in the number of autophagosomes could indicate transient induction of autophagy, inefficient clearance of cargo after lysosomal fusion, or other physiological causes. In contrast, autophagic flux provides direct evidence of lysosomal substrate degradation. However, there is no current consensus regarding the optimal methods to accurately assess autophagic flux. Using the fluorescently-tagged LC3-II protein, we aim to optimize autophagic flux detection in young and old human CD4+ T cells treated with several autophagy inhibitors. This technique may aid in understanding the mechanisms of aging and age-related diseases, potentially leading to the development of targeted interventions that promote healthy aging.

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