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Objective: To analyze and compare the expression profile of TAC3, TACR3, KISS1, and KISS1R in mural granulosa and cumulus cells from healthy oocyte donors and patients with different infertility etiologies, including advanced maternal age, endometriosis, and low ovarian response. Design: Genetic association study. Setting: Private fertility clinic and public research laboratory. Patient(s): Healthy oocyte donors and infertile women undergoing in vitro fertilization (IVF) treatment. Intervention(s): IVF. Main Outcome Measure(s): Gene expression levels of KISS1, KISS1R, TAC3, and TACR3 in human mural granulosa and cumulus cells. Result(s): Infertile women showed statistically significantly altered expression levels of KISS1 (-2.57 +/- 2.30 vs. -1.37 +/- 2.11), TAC3 (-1.21 +/- 1.40 vs. -1.49 +/- 1.98), and TACR3 (-0.77 +/- 1.36 vs. -0.03 +/- 0.56) when compared with healthy oocyte donors. Advanced maternal age patients, endometriosis patients, and low responders showed specific and altered expression profiles in comparison with oocyte donors. Conclusion(s): Abnormal expression levels of KISS1/KISS1R and TAC3/TACR3 systems in granulosa cells might be involved in the decreased fertility associated to advanced maternal age, endometriosis, and low ovarian response. ((C) 2020 by American Society for Reproductive Medicine.)

[Objective] To explore the relationship between iron and infertility by investigating iron-related gene expression in granulosa and uterine cervical cells., [Design] Case-control study., [Setting] Two tertiary hospitals., [Patient(s)] Two independent cohorts of fertile (n = 18 and n = 17) and infertile (n = 31 and n = 35) women., [Intervention(s)] In vitro fertilization., [Main Outcome Measure(s)] Gene expression levels of ferritin light chain (FTL), ferritin heavy chain (FTH), transferrin receptor (TFRC), and ferroportin (SLC40A1) mRNA were analyzed in granulosa and cervical cells., [Result(s)] In the first cohort, fertile and infertile women were similar in body mass index. Ferroportin mRNA levels were decreased in granulosa cells from infertile women in parallel with increased serum hepcidin levels. A positive association between ferroportin and TFRC mRNA, a gene associated with intracellular iron deficiency, was observed only in granulosa cells from fertile women. The major findings were replicated in a second independent cohort., [Conclusion(s)] Ferroportin mRNAs and circulating hepcidin identify a subset of infertile women and may constitute a target for therapy.

[Objective] To study the expression levels of tachykinins and tachykinin receptors in uterine leiomyomas and matched myometrium., [Design] Laboratory study., [Setting] University research laboratories and academic hospital., [Patient(s)] Women undergoing hysterectomy for symptomatic leiomyomas., [Intervention(s)] Quantitative polymerase chain reaction, immunohistochemistry and Western blot., [Main Outcome Measure(s)] Expression and tissue immunostaining of substance P, neurokinin A, hemokinin-1, neurokinin 1 receptor full-length (NK1R-Fl) and truncated (NK1R-Tr) isoforms, and neurokinin 2 receptor (NK2R) in paired samples of leiomyoma and adjacent normal myometrium., [Result(s)] TAC1 messenger RNA (mRNA) was significantly up-regulated in leiomyomas, whereas intense immunoreaction for the three peptides was particularly abundant in connective tissue cells. Differential regulation of TACR1 mRNA was observed, and at the protein level there was a significant increased expression of NK1R short isoform (NK1R-Tr). TACR2 mRNA was significantly up-regulated in leiomyomas, although levels of NK2R protein were similar in normal and tumor cells., [Conclusion(s)] These and our previous data demonstrate that the whole tachykinin system is differentially regulated in leiomyomas. The increased expression of NK1R-Tr might stimulate leiomyoma growth in a similar way to that observed in other steroid-dependent tumors., Supported in part by research grants from the Agencia Canaria de Innovación y Sociedad de la Información, ACIISI, (PI 2007/001), and from Ministerio de Economía y Competitividad (CTQ2011-25564 and RTC-2014-1431-1), Spain, in both cases with joint financing by FEDER funds from the European Union.

Objective To investigate the presence of neurokinin B (NKB)/NK 3 receptor (NK 3 R) and kisspeptin/KISS1 receptor (KISS1R) messenger RNA (mRNA) and proteins throughout the human female genital tract. Design In vitro study. Setting Academic research laboratories and academic hospitals. Patient(s) Fifteen reproductive-age women and 16 postmenopausal women provided fresh samples of uterus, ovary, or oviduct, and 12 women provided archival samples of endometrium or oviduct. Intervention(s) Fresh and archival samples of uterus, ovary, and oviduct obtained from reproductive-age and postmenopausal women. Main Outcome Measure(s) Results of reverse-transcription polymerase chain reaction (RT-PCR) and immunohistochemistry to investigate the pattern of expression of NKB/NK 3 R and kisspeptin/KISS1R in target tissues. Result(s) Expression of the genes encoding NKB ( TAC3 ) and NK 3 R ( TACR3 ), and kisspeptin ( KISS1 ) and its receptor ( KISS1R ) was found in the uterus, ovary, and oviduct. Both NKB and NK 3 R immunoreactivity was detected in the endometrium, the oviduct, and the ovary, with marked expression in endometrial and oviductal epithelial cells, where intense coexpression of kisspeptin and KISS1R was also detected. Positive staining for NKB and NK 3 R was found in the myometrium where, in contrast, kisspeptin and KISS1R were absent. Conclusion(s) NKB/NK 3 R and kisspeptin/KISS1R are present in female peripheral reproductive tissues with colocalization of both systems in some non-neuronal cell populations of the human female genital tract. Our findings are compatible with a potential modulatory role of NKB and kisspeptin at peripheral reproductive tissues.

Objective To verify the presence of protein precursor pro-enkephalin (PENK) and met-enkephalin in human spermatozoa and to characterize the effects of exogenous and endogenous enkephalins on sperm motility. Design We carried out expression assays for met-enkephalin and its protein precursor PENK by reverse transcription–polymerase chain reaction (RT-PCR), Western blot, and immunofluorescence techniques in sperm cells and motility analysis after incubation of semen samples with met-enkephlin enzyme inhibitors and the opioid receptor antagonist naloxone. Met-enkephalin secretion was analyzed by flow cytometry. Setting Assisted reproduction unit and academic research laboratory. Patient(s) Semen from 50 normozoospermic healthy human donors. Intervention(s) Spermatozoa isolated from semen on discontinuous Percoll gradient (40%–80%) followed by a swim-up was used for all techniques. Main Outcome Measure(s) Immunoblotting blots, indirect immunofluorescence antibody assays, RT-PCR blots, flow cytometry plots, and percentage of motile sperm. Result(s) We found by RT-PCR and immunofluorescence that met-enkephalin and its protein precursor PENK are present in the head of human sperm cells. Endogenous met-enkephalin increased sperm motility, whereas the addition of exogenous met-enkephalin had a biphasic effect on motility, likely due to the activation of distinct receptor subtypes. Conclusion(s) We provide evidence for a new role of met-enkephalin as an endogenous mediator of sperm motility. This autocrine regulation of sperm function by the opioid system represents a new mechanism of regulation of male factor fertility and could be useful as an emerging target for male contraception.