Mass Spectrometric Analysis of Clomiphene Citrate-induced Changes in the Secretome Profile of PAX8-positive Human Fallopian Tube Secretory Epithelial Cells and Identification of Biomarkers Relevant to Reproduction and Pregnancy

Clomiphene citrate (CC) is commonly used for infertility treatments, particularly in intrauterine insemination (IUI) and in vitro fertilization (IVF) programs. Despite its effectiveness in inducing ovulation, it is reported to have a low pregnancy rate compared to letrozole. Further, risks of ectopic pregnancy, miscarriage, and fetal malformations have been reported in CC-induced fertility treatment cycles. The present study aimed at understanding whether these effects are associated with CC-induced changes in the secretome profile of the human PAX8-positive fallopian tube secretory epithelial cells (hPFTSECs). Fallopian tube tissues from healthy women who underwent postpartum tubectomy were enzymatically dissociated and cultured with CC, and their secretomes were analyzed by Liquid Chromatography Tandem Mass Spectrometry (LC–MS/MS) to identify CC-induced alterations. Proteomic analysis identified a total of 1549 proteins in the secretome pool of hPFTSECs cultured in vitro and 258 proteins were found to be differentially abundant in response to CC. KEGG and REACTOME analysis revealed that the differentially abundant proteins (DAPs) were mainly associated with pathways such as estrogen signaling, cell cycle and DNA damage. Further, candidate proteins associated with ovulation, embryo development, embryo implantation, pregnancy complications, ectopic pregnancy, sperm functions, apoptosis, senescence, and ovarian cancers were identified. CC exposure induced changes in secretome profile of hPFTSECs may affect the female reproductive health by potentially altering the tubal microenvironment. The study was registered in the Clinical Trials Registry of India (CTRI) under the registration number CTRI/2020/11/028791 on 02/11/2020.