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Updated: Mar 12, 2026
Simultaneous Imaging and Flow-Cytometry-based Detection of Multiple Fluorescent Senescence Markers in Therapy-Induced Senescent Cancer Cells
Published on: July 12, 2022
Autophagy Detection During Oncogene-Induced Senescence Using Fluorescence Microscopy
Masako Narita1, Masashi Narita2
1Cancer Research UK Cambridge Institute, University of Cambridge, Robinson Way, Cambridge, CB2 0RE, UK.
Abstract:
Oncogene-induced senescence (OIS) is a highly dynamic process, involving several different effector mechanisms, the multitude and combination of which likely determines the quality of the phenotype (Pérez-Mancera et al., Nat Rev Cancer 14:547-558, 2014). Autophagy, a cellular degradation process, has been proposed to be one of these senescence effectors, although its functional relevance seems highly context dependent (Hoare et al., Semin Cancer Biol 21:397-404, 2011). A number of methods for monitoring autophagy are available, and several excellent protocols have been published in this journal (Klionsky et al., Autophagy 8:445-544, 2012; Tooze et al., Methods Mol Biol 1270:155-165, 2015; Tabata et al., Methods Mol Biol 931:449-466, 2013; Young and Tooze, Methods Mol Biol 445:147-157, 2008). The same principles apply to models of OIS in culture. Thus, in this chapter, we describe how to generate OIS cells using human diploid fibroblasts (HDFs), the best-characterized cell model of OIS, and how to detect autophagy, particularly focusing on immunofluorescence methods.
Insights
Oncogene-induced senescence (OIS) involves multiple effectors, including autophagy. This study details generating OIS in human diploid fibroblasts and detecting autophagy using immunofluorescence, aiding senescence research.
Area of Science:
- Cell Biology
- Cancer Research
- Molecular Biology
Background:
- Oncogene-induced senescence (OIS) is a dynamic process with various effector mechanisms influencing its phenotype.
- Autophagy, a cellular degradation pathway, is implicated as a senescence effector, but its role is context-dependent.
Purpose of the Study:
- To describe the generation of OIS in human diploid fibroblasts (HDFs).
- To detail methods for detecting autophagy in OIS models, focusing on immunofluorescence techniques.
Main Methods:
- Generation of OIS using human diploid fibroblasts (HDFs).
- Detection of autophagy via immunofluorescence microscopy.
- Utilizing established protocols for monitoring cellular processes.
Main Results:
- Successful generation of OIS in HDFs.
- Demonstration of immunofluorescence-based autophagy detection in OIS models.
- Provides a reproducible methodology for studying OIS and autophagy.
Conclusions:
- Human diploid fibroblasts serve as a robust model for studying OIS.
- Immunofluorescence is a key technique for visualizing and quantifying autophagy in OIS.
- This chapter offers practical guidance for researchers investigating the interplay between OIS and autophagy.
