Abstract: FR-OR069
Application of a Glutathione (GSH)-Sensitive Biosensor to Monitor Single-Cell Redox Profiles in Immunosuppressed Human Cells and Kidney Transplant Patients: A Feasibility Study
Session Information
- Transplantation: Basic Science Innovations and Advances
November 07, 2025 | Location: Room 370A, Convention Center
Abstract Time: 05:20 PM - 05:30 PM
Category: Transplantation
- 2101 Transplantation: Basic
Authors
- Goerlich, Nina, Charite - Universitatsmedizin Berlin, Berlin, BE, Germany
- Espinar Barranco, Laura, Icahn School of Medicine at Mount Sinai, New York, New York, United States
- Ningoo, Mehek, Icahn School of Medicine at Mount Sinai, New York, New York, United States
- Metzke, Diana, Charite - Universitatsmedizin Berlin, Berlin, BE, Germany
- Gisbert Vilanova, Cayetana, Icahn School of Medicine at Mount Sinai, New York, New York, United States
- Arzig, Joram, Charite - Universitatsmedizin Berlin, Berlin, BE, Germany
- Mirkheshti, Pouneh, Charite - Universitatsmedizin Berlin, Berlin, BE, Germany
- Cravedi, Paolo, Icahn School of Medicine at Mount Sinai, New York, New York, United States
- Fueyo-González, Francisco, Icahn School of Medicine at Mount Sinai, New York, New York, United States
- Enghard, Philipp, Charite - Universitatsmedizin Berlin, Berlin, BE, Germany
- Fribourg, Miguel, Icahn School of Medicine at Mount Sinai, New York, New York, United States
Background
Intracellular redox imbalance is a hallmark of immune dysfunction under immunosuppression (IS), contributing to infection, poor vaccine response, and allograft injury. Yet, clinical tools to monitor immune cell redox states are limited. We developed a GSH-specific, reversible, flow cytometry-compatible biosensor and previously showed it links redox state and capacity with T cell activation. Here, we applied the sensor to assess cell-specific IS effects and tested its feasibility in a clinical cohort of kidney transplant recipients (KTR).
Methods
We analyzed oxidative states in immune cells under 2 conditions: baseline GSH (redox state) and GSH after addition of the oxidative stressor tert-butyl hydroperoxide (TBHP) to assess redox capacity. First, we treated human PBMCs with IS in vitro to validate sensitivity. Second, we applied the sensor to PBMCs from stable KTR (n=34) to compare redox profiles to healthy controls (HC) and assess changes following EPO therapy.
Results
The sensor detected redox changes in immune cells exposed to IS. T cell redox capacity declined dose-dependently under Prednisolone, Tacrolimus, and MPA; monocyte capacity was reduced only by MPA. In patient samples, signals were robust across timepoints. Compared to HC, KTR showed elevated basal GSH (Fig. 1A, redox state, monocytes p < 0.001) and reduced redox capacity (Fig. 1B: CD4+ T cells p = 0.04; CD8+ T cells p = 0.08). EPO treatment altered monocyte redox state (Fig. 1C, p = 0.017), correlating with a marked reduction of TNF-α release (p = 0.0037).
Conclusion
This study demonstrates the feasibility of using our GSH-sensitive biosensor to monitor redox homeostasis in human at the single-cell level, including clinical transplant samples. Our findings lay the groundwork for incorporating redox profiling into personalized immunomonitoring strategies in transplantation and potentially other immunocompromised states.
Funding
- Government Support – Non-U.S.