Abstract: TH-PO882
Protein-Bound Uremic Solute 3-Carboxy-4-Methyl-5-Propyl-2-Furanpropanoic Acid (CMPF) Induces Eryptosis through a Piezo1-Dependent Pathway
Session Information
- Anemia and Iron Metabolism
October 24, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Anemia and Iron Metabolism
- 200 Anemia and Iron Metabolism
Authors
- Van Spitzenbergen, Beatriz Akemi Kondo, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
- Dias, Erika S., Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
- Monte-Alegre, Júlia Bacarin, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
- Andrade, Gabriela Bohnen, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
- Brugnolo-Santos, Vitor André, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
- Ozogovski, Yuri Daitschman, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
- Ferreira Dias, Gabriela, Renal Research Institute, New York, New York, United States
- Grobe, Nadja, Renal Research Institute, New York, New York, United States
- Kotanko, Peter, Renal Research Institute, New York, New York, United States
- Moreno-Amaral, Andrea Novais, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
Background
The uremic solute 3-Carboxy-4-methyl-5-propyl-2-furanpropionate (CMPF) exhibits structural resemblances with Jedi1, a known chemical activator of the mechanosensitive ion channel Piezo1. Therefore, we hypothesized that activation of Piezo1 on red blood cells (RBC) may promote eryptosis, i.e., premature RBC death in uremia (Kotanko, FASEB Bioadv. 2022). Our aim was to further probe into that hypothesis.
Methods
Isolated RBC from six healthy individuals were either pre-treated or left untreated with the Piezo1 inhibitor GsMTx-4 (5 min). This step was followed by 30 min incubation with CMPF or Jedi1 (both 87 mM). Subsequently, we stimulated Piezo1 by challenging RBC for 5 min with either hypotonic (NaCl 6g/L) or isotonic (9 g/L) saline. After centrifugation, RBC pellets were stained with Annexin-V to assess phosphatidylserine (PS) exposure on the RBC membrane and with Fluo-4AM to quantify intracellular calcium (icCa2+). Both signals report eryptosis pathways were measured by flow cytometry.
Results
Table 1: When challenged with hypotonic saline, both Jedi1 and CMPF increased PS exposure and icCa2+. In contrast, in the presence of isotonic saline, CMPF, but not Jedi1, increased PS and icCa2+. These effects were prevented by 5-min pre-incubation with GsMTx-4.
Conclusion
Our results support the notion that CMPF, a uremic toxin, activates Piezo1 on RBC and, through that pathway, promotes eryptosis. Further investigation is warranted to determine whether and to what extent this effect aggravates anemia in kidney patients.
| Experimental Condition | NaCl 6 g/L | NaCl 9 g/L | ||
| % PS exposure (MFI) | % icCa2+ (MFI) | % PS exposure (MFI) | % icCa2+ (MFI) | |
| Control (DMSO 0.12% in PBS+4% HSA) | 5.3 ± 1.4 | 21.1 ± 1.8 | 3.8 ± 1.1 | 14.1 ± 3.1 |
| Jedi 1 (87 µM) | 12.4 ± 4.4** | 37.2 ± 8.2** | 6.8 ± 1.2 | 21.8 ± 5.2 |
| GsMTx-4 (2 µM) + Jedi 1 (87 µM) | 5.6 ± 1.2 | 21.4 ± 6.3 | 5 ± 1.1 | 14.1 ± 3.3 |
| CMPF (87 µM) | 17.8 ± 5.7*** | 40.5 ± 8.1*** | 13.2 ± 3.3** | 24.2 ± 4.7** |
| GsMTx-4 (2 µM) + CMPF (87 µM) | 5.5 ± 1.2 | 19.1 ± 6.5 | 4.8 ± 1.3 | 15.4 ± 3.4 |
MFI = Mean Fluorescence Intensity; HAS = Human Albumin Serum, **p<0.01 versus Control. ***p<0.001 versus Control.