Follicle-stimulating hormone stimulates free radical generation without inducing substantial oxidative stress in human granulosa cells
Authors: Lin Nuan , Van Zomeren Koen , Plosch Torsten , Hofsink Naomi , Van Veen Teelkien , Li Hui Ting , Lin Jiazhe , Zhou Xiaoling , Groen Henk , Tietge Uwe J F , Cantineau Astrid , Schirhagl Romana , Hoek Annemieke 
Published: February 2025

Abstract

STUDY QUESTION
Does follicle stimulating hormone (FSH) induce free radical generation with substantial oxidative damage in human cumulus granulosa cells (cGC) and mural granulosa cells (mGC)?

SUMMARY ANSWER
FSH of both physiological and supraphysiological concentrations induced free radical generation on subcellular levels, most notably in the mitochondria, while the elevated free radical load caused neglectable oxidative damage in both cGCs and mGCs.

WHAT IS KNOWN ALREADY
FSH is fundamental for regulation of granulosa cell (GC) function and oocyte maturation, during which a physiological level of reactive oxygen species (ROS) is essential, while excessive amounts lead to oxidative damage. Potential adverse effects of high FSH doses on GCs may be mediated by ROS.

STUDY DESIGN, SIZE, DURATION
This prospective experimental study included patients who attended a reproductive medicine centre in 2023. cGC and mGC were separately isolated and brought into culture on the day of oocyte retrieval, 36 h after ovulation induction with recombinant hCG (250 mg,). Recombinant FSH, at different concentrations mimicking physiological (6 mIU/ml) and supraphysiological (60 mIU/ml and 600 mIU/ml) conditions, was applied (n = 4 in each group).

PARTICIPANTS/MATERIALS, SETTING, METHODS
Women aged 20-35 years, undergoing intracytoplasmic sperm injection (ICSI) with at least 3 follicles were included. Quantum sensing of cGC and mGC free radicals, detected by either cytoplasm-located fluorescent nanodiamonds (FNDs) or mitochondria-targeted FNDs, were tracked for 2 hours following FSH treatment in a magnetometry setup. Mitochondrial function analysis, as well as oxidative damage to DNA/RNA, lipids and proteins, upon FSH exposure were examined.

MAIN RESULTS AND THE ROLE OF CHANCE
FSH induced cytoplasmic and mitochondrial ROS increases in cGC and mGC (p &lt; 0.01 in all concentrations after 2 hours) while showing different patterns along time: cGC showed significantly larger cytoplasmic ROS change compared with mGC to physiological (p &lt; 0.01) and supraphysiological (p &lt; 0.05) concentrations of FSH. Significantly larger free radical changes were observed in mitochondria compared to cytoplasm in response to FSH (all concentrations in cGCs with p &lt; 0.05; supraphysiological concentrations in mGCs with p &lt; 0.05, p &lt; 0.001, respectively) after 2 hours. Mitochondrial basal respiration and ATP production were significantly increased upon FSH exposure to supraphysiological concentrations in both cGCs (p &lt; 0.01) and mGCs (p &lt; 0.05). However, no oxidative damage to GC DNA/RNA, proteins or lipids was found upon FSH exposure at any concentration except elevated lipid peroxidation in the FSH group of 600 mIU/mL (p &lt; 0.05).

LARGE SCALE DATA
N/A.

LIMITATIONS, REASONS FOR CAUTION
The GCs came from females of different biological backgrounds and were stimulated before oocyte and GC retrieval, thereby increasing the risk of variation. In addition, the effects of long-term FSH exposure as well as the effects of the FSH-induced ROS on the oocyte remain to be investigated.

WIDER IMPLICATIONS OF THE FINDINGS
We demonstrate that FSH of both physiological and supraphysiological concentrations induce free radical generation at subcellular levels, most notably in the mitochondria, while the elevated free radical load causes neglectable oxidative damage in both cGCs and mGCs. Our results suggest that the “FSH Ootoxicity” hypothesis would not seem to be mediated by ROS in human GCs.

STUDY FUNDING/COMPETING INTEREST(S)
This study is supported by the Abel Tasman Talent Program (ATTP) of the Graduate School of Medical Sciences of the University Medical Center Groningen/University of Groningen, The Netherlands, as well as an XS grant from NWO. Unrelated to the current work, A.H. is member of an advisory board on the development and application of a lifestyle App for patients with infertility, from Ferring Pharmaceutical Company, The Netherlands. R.S. is the founder of QTsense who commercialize quantum sensing equipment. This article has no direct relation to the work of QTsense. The remaining authors have no conflicts of interest.