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Extracellular vesicles (EVs) derived from red blood cells (RBCs) have emerged as a promising disease biomarker and drug delivery tool due to their intercellular signaling and cargo transporting properties. RBC-EVs have primarily been induced via calcium ionophore treatment. We recently showed that activation of piezo1, a mechanosensitive stretch-activated Ca2+ ion channel, stimulates RBCs to release EVs. These RBC-EVs may be more physiologically relevant than calcium ionophore-induced RBC-EVs, since RBCs in vivo may be stimulated to release EVs via piezo1 activation as they squeeze through capillaries and experience mechanical forces such as membrane tension and shear stress. This chapter provides a comprehensive overview of how to generate piezo1-stimulated RBC-EVs in vitro. We describe RBC isolation and treatment, along with subsequent RBC-EV isolation and characterization. Through this chapter, readers will learn how to generate piezo1 RBC-EVs for diverse biomedical applications.

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Methods for Isolating and Characterizing Red Blood Cell Extracellular Vesicles

  • 实验方案

  • 发表日期：

    2026年

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  • Gurneet S. Sangha,
  • Marzyeh Kheradmand,
  • Lauren V. Smith,
  • Alisa Morss Clyne

摘要

Extracellular vesicles (EVs) derived from red blood cells (RBCs) have emerged as a promising disease biomarker and drug delivery tool due to their intercellular signaling and cargo transporting properties. RBC-EVs have primarily been induced via calcium ionophore treatment. We recently showed that activation of piezo1, a mechanosensitive stretch-activated Ca2+ ion channel, stimulates RBCs to release EVs. These RBC-EVs may be more physiologically relevant than calcium ionophore-induced RBC-EVs, since RBCs in vivo may be stimulated to release EVs via piezo1 activation as they squeeze through capillaries and experience mechanical forces such as membrane tension and shear stress. This chapter provides a comprehensive overview of how to generate piezo1-stimulated RBC-EVs in vitro. We describe RBC isolation and treatment, along with subsequent RBC-EV isolation and characterization. Through this chapter, readers will learn how to generate piezo1 RBC-EVs for diverse biomedical applications.