Flow Cytometry Reveals the Nature of Oncotic Cells

Area of Science:

• Cell Biology
• Molecular Biology
• Immunology

Background:

• Necrosis is often broadly defined as non-specific cell death following plasma membrane damage.
• Signaling pathways in accidental cell death (ACD), also known as oncosis, remain under-investigated.
• Regulated cell death (RCD) encompasses programmed pathways like apoptosis and programmed necrosis.

Purpose of the Study:

• To characterize the phenotypic nature of cells undergoing oncosis and necrosis using flow cytometry.
• To explore the signaling pathways involved in accidental cell death (ACD) and their relationship with regulated cell death (RCD).
• To investigate the role of DNA damage response (DDR) and parthanatos in oncotic cells.

Main Methods:

• Flow cytometric analysis of intracellular antigens to distinguish cell death phenotypes.
• Induction of oncosis and apoptosis using sodium azide, with apoptosis inhibition by zVAD.
• Assessment of DNA damage response (DDR) markers (pH2AX, PARP) and Receptor-interacting serine/threonine protein kinase 3 (RIP3) expression.
• Treatment with Necrostatin-1 to evaluate its effect on DDR and apoptosis.

Main Results:

• Oncotic cells exhibited a viability+/caspase-3-/RIP3+/- phenotype.
• Both oncotic and apoptotic cells displayed DNA damage response (DDR) phenotypes, including pH2AX+/PARP- and pH2AX+/PARP+ (parthanatos).
• Necrostatin-1 reduced DDR in oncotic cells and enhanced sodium azide-induced apoptosis.

Conclusions:

• Flow cytometry provides a robust approach to differentiate cell death pathways.
• Oncotic cells show distinct phenotypes involving DDR and parthanatos.
• This study highlights a significant link between accidental cell death (ACD) and regulated cell death (RCD) processes.