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¹Unit for Cell Death and Metabolism, Center for Autophagy, Recycling and Disease, Danish Cancer Society Research Center, Strandboulevarden 49, DK-2100 Copenhagen, Denmark.

Cold Spring Harbor Protocols

|November 4, 2015

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Lysosomal membrane permeabilization (LMP) triggers cell death pathways relevant to cancer and normal physiology. This study presents three methods to quantify and visualize LMP, aiding research into cell death mechanisms.

Area of Science:

Cell Biology
Molecular Biology
Cancer Research

Background:

Lysosomal membrane permeabilization (LMP) is a cell death trigger with implications in cancer therapy and normal physiological processes like mammary gland involution.
The release of lysosomal hydrolases, particularly cysteine cathepsin proteases, into the cytosol upon LMP dictates the mode of cell death, ranging from apoptosis to necrosis.
Understanding LMP is crucial for developing targeted cancer therapies and comprehending fundamental cell biology.

Purpose of the Study:

To introduce and validate three complementary methods for quantifying and visualizing lysosomal membrane permeabilization (LMP).
To provide researchers with tools to study the role of LMP in various biological contexts, including disease and normal physiology.
To facilitate the investigation of LMP-induced cell death pathways.

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Main Methods:

Immunocytochemistry for monitoring LMP.
Fluorescent dextran release assay for visualizing LMP.
Activity assays of lysosomal enzymes in digitonin-extracted cytosol for quantification.

Main Results:

Demonstration of three distinct and complementary techniques for assessing LMP.
Validation of methods for both qualitative visualization and quantitative measurement of LMP.
Establishment of tools to differentiate between limited and massive LMP events and their downstream effects.

Conclusions:

The presented methods offer robust approaches for studying LMP-induced cell death.
These techniques will advance research into the therapeutic targeting of LMP in cancer.
The study provides valuable tools for exploring the physiological roles of LMP in cellular processes.