The NADPH oxidase 4 is a major source of hydrogen peroxide in human granulosa-lutein and granulosa tumor cells

Abstract

H 2 O 2 is a reactive oxygen species (ROS), which can diffuse away from its site of generation and may act as a cell-to-cell signaling factor. The mechanisms responsible for the generation of H 2 O 2 in human ovarian follicles and possible signaling role(s) of H 2 O 2 are not well known. We identified a source of H 2 O 2 , the enzyme NADPH oxidase (NOX) 4, in isolated differentiated, in-vitro fertilisation-derived human granulosa-lutein cells (GCs), in proliferating human granulosa tumour cells (KGN), as well as in situ in cells of growing ovarian follicles. H 2 O 2 was readily detected in the supernatant of cultured GCs and KGN cells. H 2 O 2 levels were significantly lowered by the NOX4 blocker GKT137831, indicating a pronounced contribution of NOX4 to overall H 2 O 2 generation by these cells. We provide evidence that extracellular H 2 O 2 is taken up by GCs, which is facilitated by aquaporins (peroxiporins). We thus conclude that GC-derived H 2 O 2 might act as autocrine/paracrine factor. Addition of H 2 O 2 increased MAPK-phosphorylation in GCs. Moreover, reducing H 2 O 2 production with GKT137831 slowed proliferation of KGN cells. Our results pinpoint NOX4 and H 2 O 2 as physiological players in the regulation of GC functions.