O-096 Human fallopian tube organoids provide a favorable environment for sperm motility

Abstract

Abstract Study question Does a human fallopian tube (HFT) organoid model offer a favorable apical environment for human sperm survival and motility? Summary answer The apical compartment of a new model of human fallopian tube organoids provides after differentiation a favorable environment for sperm motility. What is known already Human fallopian tubes are the site of major events that are crucial for achieving an ongoing pregnancy, such as gamete survival and competence, fertilization steps, and preimplantation embryo development. In order to better understand the tubal physiology and tubal factors involved in these reproductive functions, and to improve still suboptimal in vitro conditions for gametes preparation and embryo culture during In Vitro Fertilization IVF, we sought to develop a human fallopian tube organoid model from isolated adult stem cells allowing spermatozoa coculture in the apical compartment. Study design, size, duration Over a two-year period, fallopian tube tissues were collected for organoid culture purposes from 10 “donor” patients undergoing bilateral salpingectomy by laparoscopy for definitive sterilization. After tissue digestion, adult stem cells from the isthmus and ampulla regions were separately seeded in 3D Matrigel and cultured with conventional growth factors for organoid culture and specific factors for differentiation of the female genital tract. Participants/materials, setting, methods HFT organoids were characterized by light microscopy, scanning and transmission electron microscopy, immunofluorescence and transcriptomic analysis. Following simultaneous organoid culture on specific inserts, spermatozoa from five donors were placed either in control media or in the apical compartment of colon or HFT organoids (isthmus and ampulla separately) for 96 hours. Vitality and motility were assessed at 0, 48, and 96 hours on 200 spermatozoa in each condition and in duplicate and compared using Wilcoxon test. Main results and the role of chance Specific fallopian tube differentiation of our model was confirmed by immunofluorescence, transcriptome analysis and electron microscopy observations that exhibited ciliary and secretory cells. We succeeded in releasing spermatozoa in the apical compartment of HFT organoids and in recovering them for sperm analysis. Sperm vitality values are similar in HFT organoids and in commercial sperm media. We demonstrated a superiority of HFT organoid apical compartment for sperm motility compared with other controls (colon organoids, organoid culture media, and conventional commercial sperm washing and fertilization media). At 48 hours of incubation, progressive sperm motility was higher in the apical compartment of HFT organoids (ampulla 30.6% ±17.3, isthmus 29.3% ±14.8) than in commercial fertilization media (15.3% ±14.6) (p < 0.05) and compared with all other conditions. At 96 hours progressive sperm motility was almost nil (<1%) in all conditions except for spermatozoa within HFT organoids (p < 0.05): 11.5% ±14.7 and 12.9% ±16.9 in ampulla and isthmus organoids, respectively. Limitations, reasons for caution This was an in vitro study in which conditions of organoid culture could not exactly mimic the in vivo environment of extracellular matrix and the vascularization of fallopian tubes. Wider implications of the findings This work opens up perspectives for better understanding of HFT physiology. For the first time, it highlights the possibility of developing HFT organoids for reproductive purposes. In the future, it could help us to improve gametes fertilizing abilities and culture conditions of embryos during human assisted reproductive technologies. Trial registration number not applicable