Endometriosis is a disease in which uterine endometrium-like tissue grows outside the uterus, resulting in various symptoms, including dysmenorrhea, pelvic pain, irregular menstruation, and infertility. Although endometriosis affects approximately 10 % of women of reproductive age (Giudice and Kao, 2004), of which 20–70 % are infertile (Counseller and Crenshaw, 1951), the associated etiology remains unclear. Endometriosis can be divided into deep infiltrating endometriosis (DIE) and peritoneal endometriosis (PE); DIE has been implicated in severe menstrual and chronic pelvic pain. However, DIE and PE are diagnosed based on their different disease statuses (Cornillie et al., 1990). PE is an early stage of endometriosis resulting from the implantation of fresh, viable, retroperitoneal endometrium. Local factors may control early ectopic proliferation and superficial invasion into the pelvis. There are various reports on the percentage of NK cells in peritoneal fluid, but in general, the cellular activity of NK cells is reported to be decreased in patients with endometriosis (Osuga et al., 2011). Additionally, the observation that 90 % of women exhibit retrograde menstruation, while only 10–15 % (Halme et al., 1984) exhibit endometriosis suggests that the onset and progression of endometriosis may involve multiple mechanisms, including immune system abnormalities, and epigenetics (Nasu et al., 2011).

Natural killer (NK) cells have been a focal point in research on the etiology of reproductive failure, including recurrent implantation failure (RIF) and recurrent pregnancy loss (RPL) (Mai et al., 2021, Takeyama et al., 2021, Fukui et al., 2017). More specifically, reproductive failure is reportedly caused by increased NK cell cytotoxicity, but NK cell cytotoxicity is decreased in endometriosis (Fukui et al., 2021, Fukui et al., 2017, Oosterlynck et al., 1992, Somigliana et al., 1999, Funamizu et al., 2014).

NK cell receptors are functionally classified into activating and inhibitory types (Lanier, 2005). In humans, natural cytotoxicity receptors (NCRs), comprising NKp46 (NCR1, CD335), NKp44 (NCR2, CD336), and NKp30 (NCR3, CD337), and NKG2D are the primary activating receptors (Moretta and Moretta, 2004, Moretta et al., 2002). NKp46 is primarily activated by cells infected with bacteria or viruses or by tumor cells (Sivori et al., 1997, Zhuang and Long, 2017, Pessino et al., 1998). Furthermore, target cell recognition by NK cells under both pathophysiological and immunotherapeutic conditions is a highly dynamic process, which is controlled by integrated signals from multiple receptors that can promote or inhibit adhesion, granule polarization, and degranulation (Zamai et al., 2020). Currently, researchers have investigated and manipulated NK immune checkpoints, and data on synergistic cooperation induced by different NK surface-activating receptors on distinct NK cell subsets are still being gathered.

We have previously reported that NKp46 expression is reduced in NK cells in the peritoneal fluid of patients with endometriosis (Funamizu et al., 2014). Similarly, Giuliani et al. (2014) reported that NKp46 expression by uterine NK (uNK) cells is decreased in infertile patients with endometriosis compared to that in patients with RIF or RPL. However, NKp46 expression is lower in women with concomitant RPL and endometriosis compared to those without endometriosis (Giuliani et al., 2014). Accordingly, we reported that more precise characterization of the co-expression of activating receptors with NKp46 on NK cells may improve the investigation, diagnosis, and treatment of reproductive failures, while direct modulation of NKp46 expression may improve reproductive outcomes (Mai et al., 2021). NKp46 may also regulate the production of cytokines, including IFN-γ, TNF-α, IL-4, IL-10, and TGF-β, by endometrial NK cells (Yokota et al., 2013). Thus, NKp46, and its co-expressed receptors, may serve as useful markers for endometriosis diagnosis. NK cell expresses not only NKp46 but also various kinds of activating or inhibitory receptors on its surface.

The present study aimed to investigate the immunological relationship between endometriosis and peritoneal fluid immune cells abnormalities. Therefore, we assessed the participation of NK cells in endometriosis by analyzing the co-expression of activating or inhibitory receptors on NK cells and intracellular cytokines production. We investigated the expression of NKp46, and its activating or inhibitory receptors, on NK cells in premenopausal women who underwent laparoscopy for gynecological reasons. We also investigated the relationship between receptor expression and endometriosis severity to determine the involvement of NK cells in the pathogenesis and progression of endometriosis.