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Single-Cell Fluorescence Imaging Reveals Heterogeneity in Senescence Biomarkers and Identifies Rapamycin-Responsive
Vijayraghavan Seshadri1,2, Charmaine Chng1, Joel Tyler1
1Monash Biomedicine Discovery Institute, Department of Pharmacology, Monash University, Clayton, Victoria, Australia.
Aging Cell
|September 3, 2025
Summary
Cellular senescence, a state of irreversible cell cycle arrest, shows biomarker heterogeneity. This study reveals nuclear area as a robust indicator, correlating with IL-6 expression and SASP, and proposes an improved analysis method.
Area of Science:
- Cell Biology
- Aging Research
- Biomarker Discovery
Background:
- Cellular senescence is characterized by cell cycle arrest and the senescence-associated secretory phenotype (SASP).
- Existing biomarkers like SA-βgal, p21, and p16 have limitations in universally defining senescence and addressing expression heterogeneity.
- Understanding senescence heterogeneity is crucial for developing targeted therapies.
Purpose of the Study:
- To investigate cellular heterogeneity in senescence using single-cell fluorescence imaging.
- To identify robust biomarkers for cellular senescence and their correlation with SASP.
- To develop an improved method for quantifying senescence markers and evaluate senomorphic agent efficacy.
Main Methods:
- Single-cell fluorescence imaging of human fibroblasts undergoing chemotherapy-induced or oxidative stress-induced senescence.
- Assessment of multiple markers: SA-βgal activity, p21, IL-6 expression, nuclear area, and cell area.
- Development of an induction threshold method for biomarker quantification and analysis of rapamycin effects.
Main Results:
- Significant heterogeneity in SA-βgal activity and distinct senescent cell subpopulations were observed.
- Nuclear and cell area measurements proved to be robust senescence indicators with high variability.
- Specific nuclear area subpopulations correlated with IL-6 expression, linking biomarker heterogeneity to SASP.
- Rapamycin selectively targeted specific biomarker-expressing subpopulations in both senescence models.
Conclusions:
- Cellular senescence exhibits significant heterogeneity in biomarker expression, impacting the SASP.
- Nuclear area is a reliable indicator of senescence, with specific subpopulations linked to IL-6 levels.
- An induction threshold method enhances senescence biomarker quantification, and rapamycin demonstrates targeted efficacy.
