Membrane-anchored ROS scavenger protects pancreatic islets from hypoxic injury by preserving surface thiol homeostasis

Introduction: Islet graft dysfunction is mainly driven by hypoxia-induced injury. Under hypoxic, excessive reactive oxygen species oxidize membrane thiol groups (-SH) to disulfides (S-S), and disrupt redox homeostasis. Here, we developed a synthetic, high-affinity membrane-anchored ROS trap agent (M-REST) that anchor to islet cell membranes and competitively scavenge hypoxia-induced ROS. M-REST preserves -SH groups, thereby restoring redox homeostasis and mitigating islet hypoxic injury, with efficacy comparable to first-line ROS-scavenging agents (N-acetylcysteine, NAC) while also ensuring prolonged graft retention. Mechanistically, this redox stabilization boosts mitochondrial energy metabolism, supporting β-cell function and insulin secretion for improved transplantation outcomes.
Method: Islet cell viability was evaluated using flow cytometry, intracellular redox balance was measured via DCFH-DA fluorescence, and glucose-stimulated insulin secretion was quantified by ELISA. Ultrastructural morphology of islets was analyzed using transmission electron microscopy. Neonatal porcine islets were transplanted beneath the renal capsule of NCG mice. After 14 days, grafts were harvested for transcriptomic profiling. Graft performance was assessed through insulin immunohistochemical staining, cell viability by TUNEL assay, and redox status by thiol-reactive labeling.
Results: M-REST bound islet membranes with >95% efficiency within 15 min, and showed >90% retention over 14 days, far surpassing the in vivo pharmacokinetic residence time of NAC (~2.3 h). Under hypoxia conditions, islet cells exhibited disrupted redox homeostasis, evidenced by reduced membrane thiol (–SH) levels due to excessive ROS accumulation. M-REST treatment restored membrane thiol expression (45.60 ± 5.51% vs. 59.87 ± 2.53%) and lowered intracellular ROS levels (mean fluorescence intensity: 132.90 ± 3.70 vs. 87.50 ± 6.75), thereby improving islet cell viability (29.33 ± 2.98% vs. 46.20 ± 9.00% vs NAC:40.46±5.34%) and glucose-stimulated insulin secretion (5.15 ± 2.26 vs. 22.58 ± 6.20 vs NAC:20.06±5.62 mIU/L). In subrenal capsule transplantation models, M-REST–treated grafts reduced blood glucose in diabetic mice, maintain surface thiol expression, preserve insulin staining, and show fewer TUNEL-positive cells at days 3, 7 and 14 than controls. Correlation analysis revealed that membrane thiol levels positively associated with graft viability and insulin function. GO/KEGG analysis revealed significant enrichment of differential pathways in redox and mitochondrial metabolic processes. TEM further demonstrated M-REST mediated preservation of mitochondrial ultrastructure under hypoxia, supporting its role in maintaining redox balance and mitochondrial energy metabolism to enhance graft function.
Conclusion: These findings identify the membrane SH/S–S interface as a critical determinant of islet graft outcome and support M-REST as a redox-stabilizing strategy to enhance graft survival and function after islet transplantation.