Modulating hypoxia and oxidative stress in human xenografts using adipose tissue-derived stem cells

Abstract

Objective: To investigate whether adipose tissue-derived stem cells (ASCs) modulate hypoxia and oxidative stress in human ovarian tissue transplants.

Design: Prospective experimental study SETTING: Gynecological research unit in a university hospital PATIENT(S): Cryopreserved ovarian cortex from 5 adult women.

Intervention(s): Thirty mice were grafted with frozen-thawed human ovarian tissue, with or without ASCs (2-step/ASCs+ovarian tissue [OT] group and OT group). The ovarian grafts were retrieved on days 3 (n = 5), 10 (n = 5), and 21 (n = 5). The 10 animals grafted for 21 days underwent in vivo evaluations using microdialysis. One piece of ovarian tissue per patient was fixed for analysis after thawing (non-grafted controls).

Main outcome measure(s): Direct reactive oxygen species were collected every second day after grafting by means of microdialysis. Analyses of ovarian fragments included immunolabeling for double CD34 (revascularization by host and graft components); immunofluorescence for hypoxia-inducible factor 1α (hypoxia-related response), nuclear factor erythroid 2-related factor 2 (oxidative stress-related response), and 8-hydroxy-deoxyguanosine (oxidative stress-related DNA damage); and gene expression (quantitative reverse transcription polymerase chain reaction) for vascular endothelial growth factor-A (neoangiogenesis), superoxide dismutase 2 (antioxidant activity), and nuclear respiratory factor 1 (mitochondrial biogenesis).

Result(s): Reactive oxygen species peaked earlier in the ASC group (day 2) compared with that in the OT group (day 10) after grafting. Total vascularization was stable in the ASC group at all time points, while it was lower in the OT group 3 days after grafting. Hypoxia-inducible factor 1α expression, also detected in non-grafted controls, was significantly lower in the ASC group than in the OT group on days 3 and 10. The increase in VEGF gene expression lasted significantly longer in the ASC group than in the OT group. There was no significant upturn in the oxidative stress-related response (nuclear factor erythroid 2-related factor 2 pathway) or oocyte DNA damage (8-hydroxy-deoxyguanosine) in any of the grafted groups.

Conclusion(s): Use of ASCs allows faster ovarian graft reperfusion and mitigates the hypoxia-related response through rapid revascularization, sustained by prolonged increase in vascular endothelial growth factor after grafting. No evidence of oxidative stress-related damage was detected irrespective of the transplantation strategy.

Keywords: Adipose tissue-derived stem cells; HIF1α; hypoxia; ovarian tissue transplantation; oxidative stress.