Tianyi Li, Lydia Wehrli, Sevastiani Antonouli, Eleftheria M Panagiotou, Spyridon P Deligiannis, Xinchen Liu, Jasmin Hassan, Elisabeth Moussaud-Lamodière, Catarina Arnelo, Karin Pettersson, Kiriaki Papaikonomou, Andres Salumets, Pauliina Damdimopoulou, Valentina Di Nisio
Standard normoxia culture induces substantial transcriptional and metabolic alterations in human ovarian cortical tissue. Reduced oxygen tension partially restores markers of disrupted metabolism in stroma, supports tissue preservation and improves follicle survival. Further optimization of culture conditions should consider additional factors to mitigate the metabolic shift.
RESEARCH QUESTION: How does 6-day in vitro culture of human ovarian cortical tissue affect gene expression and tissue morphology, and can these insights help optimize culture conditions?
DESIGN: Ovarian cortical tissue from donors undergoing gender-affirming surgery or elective caesarean section was analysed. In study 1, fresh and cultured samples (n = 3) were subjected to single-cell transcriptomics to characterize culture-induced changes. In study 2, tissue from 12 donors was cultured under normoxia and hypoxia conditions to test whether altered oxygen tension modulates these effects. End points included metabolic marker gene expression, lactate production, ultrastructural analysis and follicle and stromal assessment.
RESULTS: Six-day culture in normoxia preserved the major somatic cell types in ovarian tissue but also induced broad transcriptional changes suggestive of extracellular matrix (ECM) remodelling and metabolic reprogramming. Increased glycolysis and oxidative phosphorylation signatures were observed across major somatic compartments. Predicted cell‒cell communication based on the transcriptomics data revealed a shift toward ECM-associated signalling after in vitro culture. Hypoxia was tested as a more physiological culture condition to mitigate the normoxia culture-induced changes. Results showed that hypoxia partially restored glycolytic signatures in the stromal compartment whereas it was associated with increased lactate production, reduced glycogen deposition in the tissue fragments, improved follicle survival and preserved stromal features.
CONCLUSIONS: Standard normoxia culture induces substantial transcriptional and metabolic alterations in human ovarian cortical tissue. Reduced oxygen tension partially restores markers of disrupted metabolism in stroma, supports tissue preservation and improves follicle survival. Further optimization of culture conditions should consider additional factors to mitigate the metabolic shift.