Sex-differences In Endothelial Dysfunction After Ischemia/Reperfusion Injury: Effect of Postpartum Plasma Supplementation During Storage On Rat Arterial Vessels

Background and Aims: Ischemia/reperfusion injury (IRI) remains a major complication of cardiac bypass surgery, contributing to endothelial dysfunction of vascular grafts. Accumulating evidence supports the role of sex hormones as modulators of cardiovascular risk. Given the established link of female hormonal status and outcomes in ischemic cardiovascular diseases, we hypothesized that supplementing saline with postpartum plasma protects vascular grafts against IRI. This study investigates the effect of storing rat aorta with male, female and postpartum plasma on vascular function following IRI.

Methods: Thoracic aortic rings were obtained from 7-9-month-old male and female Wistar rats and either directly mounted in organ bath chambers (Control_male n=7, 507±25g; Control_female n=8, 297±13g), or subjected to 24h of preservation at 4°C to mimic cold ischemia. Preservation was performed either in saline alone (IR_male n=8, 484±25g; IR_female n=9, 329±9g), or in saline supplemented with 3% plasma. The plasma groups included male plasma (IR+male_male n=10, 495±16g; IR+male_female n=8, 324±10g), female plasma (IR+female_male n=8, 517±31g; IR+female_female n=8, 328±13g) and postpartum plasma (IR+postpartum_male n=9, 507±14g; IR+postpartum_female n=10, 329±12g). To examine vascular function, organ bath experiments were used to evaluate both contraction and relaxation responses ex vivo. Endothelium-dependent relaxation was assessed by measuring the maximal relaxation (R_max) to acetylcholine after pre-constriction with phenylephrine. Additionally, enzyme-linked immunosorbent assay (ELISA) was performed to determine the concentrations of estrogen, progesterone and oxytocin in the male, female and postpartum plasma used for supplementation.

Results: Control groups showed the highest maximum relaxation (R_max) to acetylcholine (Control_male 84±3%; Control_female 78±1%) among all groups. IRI significantly impaired the endothelium function by reducing R_max to 34±3% in males and 30±3% in females. The supplementation of postpartum plasma attenuated this dysfunction in both sexes, increasing R_max to 47±3% in males (p<0.05) and 49±4% in females compared to the IR-groups. Female plasma showed approximately the same tendency in reducing endothelial dysfunction after IRI solely in female aortic rings (IR_female 30±3% vs. IR+female_female 52±3%), whereas male rings derived no benefit (IR_male 34±3% vs. IR+female_male 33±3%). Male plasma had no effect on endothelial function following IRI.

ELISA showed significantly increased progesterone level in postpartum (55±1 ng/ml) and female plasma (55±5 ng/ml) compared to male plasma (28±3 ng/ml; each p<0.0001). The highest level of estrogen was detected in the postpartum plasma (64±8 pg/ml) vs. female plasma (39±5 pg/ml, p<0.05), and vs. male plasma (43±3 pg/ml). The level of oxytocin did not differ among the groups.

Conclusion: Postpartum plasma alleviates endothelial dysfunction following IRI in the arterial vessels of both female and male rats, whereas female plasma exerts a sex-specific protective effect limited to female aortic rings. These effects may be attributed to the presence of progesterone and estrogen. These preclinical data underscore the potential value of tailoring preservation solutions based on sex and hormonal status to improve graft survival and post-transplant outcomes in clinical settings.