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Updated: Jun 2, 2026
Induction of Eryptosis in Red Blood Cells Using a Calcium Ionophore
Published on: January 21, 2020
Stimulation of suicidal erythrocyte death by oridonin
Kashif Jilani1, Syed M Qadri, Christine Zelenak
1Department of Physiology, University of Tuebingen, Gmelinstraße 5, D-72076 Tuebingen, Germany.
Abstract:
Oridonin triggers apoptosis of cancer cells and was suggested as anticancer agent. Oridonin is partially effective through mitochondrial depolarization and partially by modifying gene expression. Erythrocytes lack mitochondria and nuclei but may undergo eryptosis, a suicidal cell death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine exposure at the cell surface. Triggers of eryptosis include increase of cytosolic Ca(2+)-activity, ATP depletion and ceramide formation. The present study explored, whether oridonin triggers eryptosis. Cytosolic Ca(2+)-concentration was estimated from Fluo3-fluorescence, cell volume from forward scatter in FACS analysis, phosphatidylserine exposure from binding of fluorescent annexin V, hemolysis from hemoglobin release, ATP concentration utilizing a luciferin-luciferase assay and ceramide abundance utilizing fluorescent anti-ceramide antibodies. A 48 h exposure to oridonin (≥25μM) significantly increased cytosolic Ca(2+)-concentration, increased ceramide formation, decreased forward scatter and triggered annexin V-binding (the latter in >20% of the erythrocytes). Oridonin didn't decrease ATP concentration and hemolysed <5% of erythrocytes. The effects of oridonin on annexin V binding were partially reversed in the nominal absence of Ca(2+) and by the addition of amiloride (1mM). The present observations reveal a completely novel effect of oridonin, i.e. triggering of Ca(2+) entry and ceramide formation as well as suicidal death of erythrocytes.
Insights
Oridonin, an anticancer agent, induces suicidal death in erythrocytes (red blood cells) by increasing calcium levels and ceramide formation. This novel finding reveals a new mechanism of oridonin
Area of Science:
- Biochemistry
- Cell Biology
- Toxicology
Background:
- Oridonin is recognized for its anticancer properties, primarily through inducing apoptosis in cancer cells.
- Eryptosis, a form of suicidal cell death in erythrocytes, is characterized by cell shrinkage and phosphatidylserine exposure.
- Eryptosis can be triggered by increased intracellular calcium, ATP depletion, and ceramide formation.
Purpose of the Study:
- To investigate whether oridonin induces eryptosis in human erythrocytes.
- To elucidate the specific mechanisms by which oridonin affects erythrocytes.
Main Methods:
- Flow cytometry (FACS) was used to measure cell volume and phosphatidylserine exposure (annexin V binding).
- Intracellular calcium levels were assessed using Fluo3-fluorescence.
- Ceramide abundance was quantified using fluorescent antibodies.
- ATP concentration was measured via a luciferin-luciferase assay.
Main Results:
- Oridonin exposure (≥25μM for 48h) significantly increased intracellular calcium concentration and ceramide formation in erythrocytes.
- Oridonin treatment led to cell shrinkage (decreased forward scatter) and phosphatidylserine exposure, indicative of eryptosis.
- Oridonin did not significantly deplete ATP levels or cause substantial hemolysis.
- The eryptotic effects of oridonin were partially reversible by removing extracellular calcium or adding amiloride.
Conclusions:
- Oridonin triggers eryptosis, a novel suicidal death pathway in erythrocytes.
- The mechanism involves calcium influx and ceramide generation, distinct from its known anticancer effects.
- This study uncovers a previously unrecognized biological activity of oridonin.
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