Effects of Polyethelene Glycol (PEG)-Fibrinogen Combined in Various Techniques with Nerurotrophin 4/5 (NT4/5) on Development of Human Primordial Follicles in Organ Culture

Michal Perpinial ^1,3,5 Ronit Abir ^1,2,5 יואל שופרו ^1,4,5 Orly Kleiner ^1,2,5 Olga Kossover ⁶ Galia Oron ^1,4 Avi Ben-Haroush ^1,4 Benjamim Fisch ¹ Dror Seliktar ⁶

¹Beilinson Women Hospital, IVF and Infertility Unit, Rabin Medical Center
²Cellular and Developmental Biology, Tel Aviv University, Sackler Faculty of Medicine
³Life Science Faculty, Tel Aviv University
⁴Obstetrics and Gynecology, Tel Aviv University, Sackler Faculty of Medicine
⁵The Felsenstein Medical Research Center, Tel Aviv University, Sackler Faculty of Medicine
⁶Faculty of Biomedical Engineering, Technion-Israel Institute of Technology

Introduction: The signals that induce activation of primordial follicles are unclear. NT4/5 is expressed in human follicles and promoted human follicular assembly. PEG-fibrinogen seems an effective culture matrix for human primordial follicles.

Aim: To investigate if NT4/5 can promote human primordial follicle development and if results can be optimized with medium supplementation or addition to PEG-fibrinogen.

Methods: Human ovarian slices were cultured as follows: (A) embedded in PEG-fibrinogen with culture medium supplemented with NT4/5, (B) embedded in PEG-fibrinogen that contained NT4/5, (C) Culture inserts containing PEG-fibrinogen with NT4/5 were placed above incubated slices. The wells were cultured for six days. Follicular growth was evaluated by follicle counts, Ki67 immunostaining and 17-β estradiol _(E2) and Anti Mullerian Hormone (AMH) measurements in spent medium samples.

Results: In total 509 follicles were counted. There seemed a trend of atretic follicle reduction in parallel with high primordial follicle counts in groups A and C. There was no increase in developing follicles (primary and secondary) in any of the groups and Ki67 staining was positive in all developing follicles. E₂ and AMH measurements are underway.

Conclusions: So far, NT4/5 does not seem to promote growth, but further studies are needed. In addition, the optimal method for growth factor supplementation (in the culture medium or in PEG-fibrinogen) needs investigation.