Monocyte perturbation modulates the ovarian response to an immune challenge
Introduction
Monocytes and macrophages are key innate immune cells that play critical roles in the body's response to pathogens as well as in the regulation of immune homeostasis. These cells contribute to the initiation, development and resolution of inflammation, as well as to the tissue regeneration and repair processes, promoting angiogenesis and vascular remodelling (Bergmann et al., 2006; Shi and Pamer, 2011; Ginhoux and Jung, 2014). Monocytes and macrophages are also important in the dynamic regulation of organ-specific functions, including in ovulation and the maintenance of pregnancy (Oakley et al., 2011). Within the ovary, macrophages regulate the ovarian immune microenvironment through the release of cytokines, chemokines, and growth factors, as well as through phagocytosis and tissue remodelling (Wu et al., 2004; Cohen et al., 1999; Oakley et al., 2010; Zhang et al., 2021). Disruption of the ovarian macrophage milieu by conditional ablation of these cells in mice has been shown to acutely impair ovarian vascular integrity and steroidogenesis (Turner et al., 2011; Care et al., 2013).
We have recently demonstrated that, in adult rats, the absence of monocytes and macrophages does not overtly disrupt ovarian follicle health or maturation under freely-cycling conditions, suggesting they are not essential for ovarian follicle integrity in the healthy mature animal. As such, conditional depletion of circulating and ovarian monocytes and macrophages using a Cx3Cr1-Dtr knock-in rat model has no acute or sustained effects on ovarian follicle numbers, atresia, or apoptosis, and no effect on corpora lutea regression for up to 3 weeks after disruption to these cells (Sominsky et al., 2021b).
Although monocytes and macrophages may not play a key role in the day-to-day function and maturation of ovarian follicles, these cells are still crucial for maintaining ovarian function in the context of an inflammatory challenge (Herath et al., 2007). Exposure to immune challenge with lipopolysaccharide (LPS) in the early postnatal period results in follicle loss and compromised reproductive capacity later in life (Fuller et al., 2017; Sominsky et al., 2013; Wu et al., 2011; Knox et al., 2009). Exposure to LPS during the prepubertal period influences folliculogenesis and steroidogenesis by reducing ovarian expression of follicle stimulating hormone receptor (Fshr) and disrupting steroid hormone release from both granulosa and theca-interstitial cells, leading to delayed puberty onset (Seibert et al., 2019, DA KYUNG Yoo, 2016; Sominsky et al., 2012). In adults, LPS depletes the ovarian follicle pool (Bromfield and Sheldon, 2013; Besnard et al., 2001). LPS also suppresses hypothalamic pituitary-gonadal (HPG) axis functionality, reducing the pulsatile release of gonadotrophin-releasing hormone (GnRH) and postponing the subsequent surge in follicle stimulating hormone (FSH) and luteinizing hormone (LH) (Bidne et al., 2018a), as well as disrupting ovarian steroidogenesis (Herath et al., 2007).
While the evidence above suggests that immune challenges have negative effects on reproductive function, and specifically the ovary, it is important to note that inflammatory events are commonly experienced throughout life and in most cases will have no consequences for fertility in the longer term (Weiss et al., 2009). Similarly, while prolonged suppression of the immune system with anti-inflammatories, antibiotics or due to chemotherapy treatments can have damaging effects on fertility, temporary suppression of acute inflammation is common and unlikely to affect reproductive capacity in the long-term (Spears et al., 2019; Boots and Jungheim, 2015; Lemeire et al., 2007). Therefore, ovarian immune cells may be necessary for appropriately responding to acute immune and inflammatory challenges, but can be overwhelmed or dysregulated if the challenge is large, prolonged, or occurs during critical programming periods (Weiss et al., 2009; Ye et al., 2016).
In this study we aimed to assess if a transient disruption to the ovarian immune milieu, induced by a reversible depletion of circulating monocytes and ovarian macrophages, would affect subsequent ovarian responses to an acute immune challenge. We hypothesised that an immune challenge with LPS would cause a disruption in the maturation of ovarian follicles, and would increase the presence of macrophages associated with follicle atresia and apoptotic cells (Besnard et al., 2001; Tingen et al., 2011) but that these effects would be exacerbated in those with prior monocyte depletion and repopulation. We have previously established that diphtheria toxin (DT) in the Cx3cr1-Dtr rat depletes ~50–60% circulating and ovarian monocytes and macrophages within 24 h, with repopulation of circulating monocytes by 5 days post-DT and no major disruptions to ovarian follicles at this time (Sominsky et al., 2021b). Administration of DT in this model also leads to depletion of microglia throughout the brain (De Luca et al., 2019). We have previously shown that upon repopulation, microglia display an ameboid morphology, associated with increased activation state (De Luca et al., 2020). Interestingly, changes in microglial morphology upon repopulation are also associated with increased astrocyte density and their phagocytic capacity at that time (De Luca et al., 2020), indicating functional changes of the newly repopulated cells and a coordinated role of the brain immune cells to perturbations in the immune milieu. Here, we explored the possibility that in addition to the previously established effects of systemic DT administration on microglia, circulating monocyte depletion and repopulation in Cx3cr1-Dtr rats would also lead to changes in their functional profile, leading to altered capacity of ovarian monocytes and macrophages to respond to an acute immune challenge. We therefore used this model here and applied DT to deplete monocytes. We then gave LPS at 5 days post-DT to assess the consequences of acute monocyte perturbation on ovarian responses to systemic immune challenge upon restoration of immune homeostasis.
Section snippets
Experimental animals and monocyte depletion
We conducted all experiments at RMIT University, Australia in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes, with approval from the RMIT University Animal Ethics Committee. We used a knock-in rat model, on a Wistar Han background, in which a diphtheria toxin receptor (Dtr) is expressed under the control of the endogenous Cx3cr1 promoter sequence. These rats have been described previously, where we have shown that upon administration of
Transient monocyte depletion does not exacerbate ovarian inflammation in response to an immune challenge
To assess the sustained effects of an acute disruption to the immune milieu on ovarian vulnerability to a subsequent immune challenge, we gave LPS at 5 days after monocyte depletion, a timepoint at which ovarian follicles are not affected by monocyte perturbations (Sominsky et al., 2021b) and culled the rats 24 h later, a timeframe sufficient to induce ovarian inflammation and inflammation-induced disruption of the ovarian follicle pool (Bromfield and Sheldon, 2013). LPS increased the
Discussion
In this study, we aimed to assess whether a transient immune disruption due to circulating and ovarian monocyte depletion and repopulation would influence the susceptibility of the ovary to a subsequent an immune challenge. We have previously shown that circulating and ovarian monocyte and macrophage depletion has no lasting overt effects on ovarian follicle numbers or health at 5 and 21 days post-depletion (Sominsky et al., 2021b). However, here we show that acute monocyte depletion and
CRediT authorship contribution statement
Simin Younesi: Investigation, Methodology, Formal analysis, Writing – original draft, Writing – review & editing. Sarah J. Spencer: Conceptualization, Methodology, Resources, Supervision, Writing – review & editing. Luba Sominsky: Conceptualization, Investigation, Methodology, Formal analysis, Resources, Supervision, Writing – review & editing.
Acknowledgements
This project was supported by RMIT University Ph.D. Scholarship to SY, funding from a National Health and Medical Research Council Career Development Fellowship II (APP1128646) to SJS and by an RMIT Vice-Chancellor's Postdoctoral Fellowship to LS.
References (61)
- et al.
Conditional microglial depletion in rats leads to reversible anorexia and weight loss by disrupting gustatory circuitry
Brain Behav. Immun.
(2019) - et al.
Neuroimmune regulation of female reproduction in health and disease
Current Opinion in Behavioral Sciences
(2019) - et al.
Effects of electroacupuncture on luteal regression and steroidogenesis in ovarian hyperstimulation syndrome model rat
Life Sci.
(2018) - et al.
The antibiotic streptomycin assessed in a battery of in vitro tests for reproductive toxicology
Toxicol. Vitro
(2007) - et al.
Pituitary-ovary-spleen axis in ovulation
Trends Endocrinol. Metabol.
(2011) - et al.
Activation of arcuate nucleus GABA neurons promotes luteinizing hormone secretion and reproductive dysfunction: implications for polycystic ovary syndrome
EBioMedicine
(2019) - et al.
Neonatal immune challenge alters reproductive development in the female rat
Horm. Behav.
(2012) - et al.
Neonatal overfeeding induces early decline of the ovarian reserve: implications for the role of leptin
Mol. Cell. Endocrinol.
(2016) - et al.
Ovarian theca cells in follicular function
Reprod. Biomed. Online
(2007) - et al.
Arteriogenesis depends on circulating monocytes and macrophage accumulation and is severely depressed in op/op mice
J. Leukoc. Biol.
(2006)
Prolactin and lipopolysaccharide treatment increased apoptosis and atresia in rat ovarian follicles
Acta Physiol. Scand.
Disruption of female reproductive function by endotoxins
Reproduction
Disruption of female reproductive function by endotoxins
Reproduction
Inflammation and human ovarian follicular dynamics
Semin. Reprod. Med.
Localization of leukocyte subsets in the rat ovary during the periovulatory period
Biol. Reprod.
Lipopolysaccharide initiates inflammation in bovine granulosa cells via the TLR4 pathway and perturbs oocyte meiotic progression in vitro
Endocrinology
Lipopolysaccharide reduces the primordial follicle pool in the bovine ovarian cortex ex vivo and in the murine ovary in vivo
Biol. Reprod.
Macrophages regulate corpus luteum development during embryo implantation in mice
J. Clin. Invest.
Maternal nutrient restriction impairs young adult offspring ovarian signaling resulting in reproductive dysfunction and follicle loss
Biol. Reprod.
Macrophages: important accessory cells for reproductive function
J. Leukoc. Biol.
Effect of lipopolysaccharide (LPS) exposure on the reproductive organs of immature female rats
Development & reproduction
Glial remodeling enhances short-term memory performance in Wistar rats
J. Neuroinflammation
Chronic predator stress in female mice reduces primordial follicle numbers: implications for the role of ghrelin
J. Endocrinol.
Luteinizing hormone acts directly at granulosa cells to stimulate periovulatory processes
Endocrine
Single-cell reconstruction of follicular remodeling in the human adult ovary
Nat. Commun.
Neonatal immune activation depletes the ovarian follicle reserve and alters ovarian acute inflammatory mediators in neonatal rats
Biol. Reprod.
Monocytes and macrophages: developmental pathways and tissue homeostasis
Nat. Rev. Immunol.
Nuclear exclusion of SMAD2/3 in granulosa cells is associated with primordial follicle activation in the mouse ovary
J. Cell Sci.
Luteinizing hormone facilitates antral follicular maturation and survival via thecal paracrine signaling in cattle
Endocrinology
Ovarian reserve alterations in premenopausal women with chronic inflammatory rheumatic diseases: impact of rheumatoid arthritis, Behcet's disease and spondyloarthritis on anti-Mullerian hormone levels
Rheumatology
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