Isolation of functional lysosomes from skeletal muscle

Area of Science:

• Cell Biology
• Muscle Physiology
• Organelle Biology

Background:

• Lysosomes degrade cellular components, including mitochondria, essential for skeletal muscle homeostasis.
• Mitochondrial quality control via mitophagy relies on lysosomal degradation, but isolation from muscle is challenging.
• Understanding skeletal muscle lysosome function is crucial for aging, disease, and exercise adaptation research.

Purpose of the Study:

• To develop a method for isolating functional lysosomes from mouse skeletal muscle.
• To enable assessment of lysosomal integrity and enzymatic activity in skeletal muscle.
• To investigate changes in lysosomal function in aging skeletal muscle.

Main Methods:

• Isolation of intact, functional lysosomes from small amounts of mouse skeletal muscle tissue.
• Development of functional assays to evaluate lysosomal enzymatic activity (e.g., acid phosphatase, cathepsin-B).
• Assessment of calcium release from isolated lysosomes.

Main Results:

• A novel method successfully isolated pure, functional lysosomes from skeletal muscle without density gradients or lysosome-modifying agents.
• Functional assays demonstrated reduced lysosomal degradative activity in lysosomes from aging muscle.
• The method preserves lysosomal membrane integrity and function.

Conclusions:

• This protocol provides a valuable tool for studying lysosomal biology in skeletal muscle.
• The findings highlight impaired lysosomal degradative activity in aging muscle.
• This approach supports investigations into lysosome-related dysfunction in muscle aging, disease, and exercise.