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OpinionPituitary–ovary–spleen axis inovulationOliver R. Oakley1, Michele L. Frazer2 and CheMyong Ko11  Division of Clinica...
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Relaçòes entre o sistema imunológico e o reprodutor parecem depender de liberação de leucócitos pelo baço, através de comando do LH. Observar as figuras ilustrativas.

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  1. 1. OpinionPituitary–ovary–spleen axis inovulationOliver R. Oakley1, Michele L. Frazer2 and CheMyong Ko11 Division of Clinical and Reproductive Sciences, Department of Clinical Sciences, College of Health Sciences,University of Kentucky, Lexington, Kentucky 40536, USA2 Hagyards Equine Medical Institute, 4250 Iron Works Pike, Lexington, Kentucky 40511, USALeukocytes are rapidly recruited to the preovulatory ovary examine trafficking of leukocytes into the ovary, the re-and play a crucial role as facilitators of ovulation and luteal quirement for leukocytes in ovulation, and consider information. In this article, recent findings on leukocyte depth the spleen as a source of leukocytes.trafficking to the ovary, as well as the physiological roleof leukocytes in the ovary, will be summarized and dis- Trafficking of leukocytes into the ovarycussed. We then explore the novel hypothesis that the The migration of leukocytes in response to chemokines hashypothalamus–pituitary–ovary (HPO) axis might include been implicated in a plethora of normal and pathophysio-the spleen as a reservoir of leukocytes by summarizing logical aspects of reproductive systems [8]. Multiple che-recent reports on this topic, both in the fields of immu- moattractants including interleukin-8 (IL-8) and a varietynology and reproductive biology. of their target populations of leukocytes have been shown to play important roles in ovulation [9–11]. Here we sum-Linking leukocytes with ovulation marize ovarian leukocyte populations, their function andOvulation, a key step in the propagation of life, has always factors that affect their infiltration into the ovary.been a subject of human curiosity. This egg-releasing act ofthe ovary is still a mysterious event and much about the Leukocyte populations and their localization within theprocess has yet to be unveiled. Ovulation is a crucial step in ovaryreproduction and has become a key therapeutic target for Traditionally, immunohistochemical techniques have beentreating female infertility and various ovarian diseases. used to characterize ovarian leukocyte populations. Al-Ovulatory failure is associated with the development of though these methods are effective for identifying theovarian disorders such as polycystic ovarian syndrome localization of leukocytes in ovarian tissues, determining(PCOS), hemorrhagic cyst formation, and hormonal imbal- the precise leukocyte subsets that are present in the tissueance, all of which are major risk factors in women’s health has been challenging. Modern techniques such as flow[1–3]. Furthermore, controlling ovulation has become a cytometry have made it possible to distinguish betweenhallmark for contraception because blockage of ovulation CD4+ T cells, CD8+ T cells, B cells, natural killer (NK) cells,ensures the absence of fertilizable eggs [4]. Understanding regulatory T cells or other cell types, each with differentthe mechanisms that govern this ovulatory process, how- functions. Table 1 summarizes the leukocyte populationsever, is challenging because there is interplay between the that have been identified in the ovary, their localizationreproductive system, the immune system, and possibly and possible functions. As shown in the table, most leuko-other systems. Recently a comprehensive flow cytometry cyte subtypes are found in the ovary and are predominatelyapproach was applied to measure inflammation quantita- localized in the periphery of the follicle, interstitium, andtively during ovulation by determining the spatiotemporal corpora lutea, but not inside follicles.patterns of leukocyte infiltration in the ovaries of imma-ture and adult rats. This effort led to the finding of massive Mechanism of leukocyte infiltration into the ovaryleukocyte infiltration into the ovary induced by the lutei- The initiator of an inflammatory event is often a discretenizing hormone (LH) surge or by human chorionic gonado- signal that is rapidly amplified by chemical signals pro-tropin (hCG) injection during ovulation [5,6]. Surprisingly, duced by responding tissues and infiltrating cells. Unlikeovarian leukocyte infiltration was accompanied by the during infection, where the inflammatory stimuli are ob-release of millions of leukocytes into the bloodstream from vious, the initiating factors for an inflammatory responsethe spleen, indicating that this immune organ might be a that occurs during a normal physiological event such assource of leukocytes that infiltrate the preovulatory ovary. preovulatory inflammation are less clear. Infiltration andSupporting this idea, recent studies showed that spleen distribution of leukocytes in the ovary are correlated withleukocytes are recruited to injured heart tissues following hormonal changes associated with the estrous cyclemyocardial infarction [7]. Both of these studies demon- [12,13], indicating that reproductive hormones such asstrate the importance of the spleen as an immediate source ovarian steroids and gonadotropins can elicit inflammato-of leukocytes for inflammatory events. In this article we ry responses in the ovary. The same adhesion molecules, chemokines and cytokines observed in immune responses Corresponding author: Ko, C. ( to infectious agents are also found in the ovary during1043-2760/$ – see front matter ß 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.tem.2011.04.005 Trends in Endocrinology and Metabolism, September 2011, Vol. 22, No. 9 345
  2. 2. Opinion Trends in Endocrinology and Metabolism September 2011, Vol. 22, No. 9Table 1. Ovarian leukocyte species, localization and functionsCell type Location Possible function(s) ReferencesMonocyte/macrophage Periphery of follicles, tunica Il-8 secretion, luteal regression [5,22,99–111] albugenia, corpora luteaNeutrophils Theca layer, corpora lutea Production of proteolytic enzymes, [5,22,110,112,113] ECM degradation, follicle maturation, ovulation, luteal formationLymphocytes Hilus, stroma, corpora lutea Selection of dominant follicle, [5,101,107,110] luteal formation, luteal regressionNK-cells Follicle, corpora lutea Angiogenesis [114]Mast cells Medulla, cortex, interstitium, ECM degradation, ovulation [109,115,116] corpora luteaEosinophils Theca layer, corpora lutea ECM degradation, neovascularization [111,116,117]estrous [5,14,15], suggesting a similar mechanism of leu- the ovary are low, but production increases in granulosakocyte infiltration in the preovulatory ovary. The cytokines cells and theca cells upon LH stimulation [11]. IL-8 is alsointerleukins (IL)-1, IL-6 and IL-10 could contribute to known to increase vascular permeability [19,20], and thisincreased cell adhesion molecule (CAM) expression during could facilitate the infiltration process. In addition, theovulation [16]. Furthermore, our recent data show in- infiltrating leukocytes can interact with ovarian endothe-creased expression of ICAM-1 and E-selectin corresponds lial cells and other cell types via a multitude of chemokineswith increased infiltration of leukocytes into the ovary [5]. during ovulation [21–23]. Treatment with neutralizingAccordingly, treatment with IL-1 receptor antagonist (IL- antibodies to either IL-8 [24] or neutrophils [1] significant-1Ra) inhibits hCG-induced ovulation rates in rats by 40% ly reduces ovulation rates in animal models.[17]. Two chemokines, monocyte chemotactic protein-1 The classical four step process of tethering and rolling, (MCP-1, also known as chemokine C–C motif ligandactivation, firm adhesion and transmigration into the tis- CCL2) and thymus-expressed cytokine (TECK/CCL25),sues is paramount to any event requiring infiltration of are well characterized in ovarian leukocyte infiltration.leukocytes [18]; we anticipate ovulatory inflammation to be MCP-1 is a potent chemoattractant for monocytes and isthe same. Once the leukocytes are tethered on the endo- also effective in recruiting macrophages and T cells [25,26].thelial cell wall, specific populations of leukocytes, charac- The major producers of MCP-1 are monocytes and macro-terized by their chemokine receptor (CCR) expression, phages, although several other cell types including epithe-migrate towards the source of the corresponding chemo- lial, endothelial, and smooth muscle cells have been shownkine produced by theca cells (TC), granulosa cells (GC) or to produce this protein [27]. In addition to the chemotacticresident ovarian leukocytes (Table 2). For instance, neu- properties, MCP-1 interacts with G-protein-coupled recep-trophils infiltrate the ovary in response to an increased tors to induce multiple intracellular responses includingconcentration gradient of IL-8 [9]. Amounts of basal IL-8 in activation and degranulation (a comprehensive review ofTable 2. List of chemokines, chemokine receptors and responsible cells present in the ovaryChemokine Producing cells Chemokine receptor Responding cells ReferencesCCL-2 (MCP-1) Mo/MF, cDC, BMDC, CCR-2a, b, T, Mo, baso, DC, NK [23,31,118–120] GC, GLC, stromal fibroblast CCR-8, CCR-11CCL-3 (MIP-1a) Mo/MF, cDC, BMDC, CCR-1, CCR-5 Mo/MF, T, baso, eos, neut, [118–120] TE, TM, NK, B, GC, GLC DC, NK, GC, TCCCL-4 (MIP-1b) Mo/MF, cDC, BMDC, CCR-5 Mo/MF, cDC, BMDC, B, T, [118–120] TE, TM, NK, B NK, baso, eos, B, GC, TCCCL-5 (RANTES) Mo/MF, cDC, BMDC, CCR-1, CCR-3, TE, TM, NK, Mo/MF, [118–120] TE, GC, TM, NK CCR-5 DC, eos, baso, GC, TCCCL-20(MIP-3a/LARC) Epithelial, GC CCR-6 PBMC, TE, TM, NKT, DC, B [118,121]CCL25 (TECK) Mo/MF, cDC, BMDC, TC CCR-9 MF, thymocyte, DC, B [37,118]CXCL-1 (Groa) Mo/MF, DC, BMDC, GC CXCR1, CXCR2 Neut, fibroblast [20,118,120]CXCL-5 (ENA-78) BMDC CXCR2 Neut, endo, BMDC [120]CXCL-8 (IL-8) Mast, GC, GLC, TC, CXCR1, CXCR2 Neut, baso, T, endo [20] stromal fibroblastCXCL-10 (IP-10) Osteoblasts, BM endo, GC CXCR3A CXCR3B T, NK, B, endo, Mo/MF, DC, GC [119,120]CXCL-12 (SDF-1) BM reticular, GC, endo, CXCR4, CXCR7 CD34+ BM, thymocytes, [122,123] stromal, fibroblast Mo/MF, TE, B, plasma, neut, cDC, BMDCCX3CL1 (Fractalkine) Mast cells, GC, TC and CX3CR1 NK, Mo, neut, mast, astrocytes, [118] stromal fibroblasts, neurons, activated T cells endotheliumBaso, basophil; BMDC, bone marrow-derived dendritic cell; cDC, conventional dendritic cell; DC, dendritic cell; endo, endothelial cell; GC, granulosa cell; GLC, granulosa-lutein cell; NK, natural killer cell; neut, neutrophil; mast, mast cell; Mo, monocyte; MF, macrophage; PBMC, polymorphonuclear cell; TC, theca cell; TE, effector T cell; TM,memory T cell.346
  3. 3. Opinion Trends in Endocrinology and Metabolism September 2011, Vol. 22, No. 9alternative functions for MCP-1 is given in [28]). Interest- Many immune mediators, such as cytokines, chemokines,ingly, studies have demonstrated in humans and rats that cell surface receptors and adhesion molecules, are alsoMCP-1 is involved in all aspects of ovarian function includ- substrates for MMP action [46]. In this regard, leukocyte-ing follicular development [29,30], ovulation and luteolysis secreted MMPs might act both to break down the ECM and[23,31]. Several studies have demonstrated that inhibiting to regulate the breakdown of chemotactic proteins, therebythe production of monocytes/macrophages [32,33] or the limiting leukocyte infiltrationdirect neutralization of ovarian macrophages [34] results Angiogenesis and neovascularization take place recur-in reduced or inhibited ovulation rates in mice, further rently within the ovary, and vascular endothelial growthsupporting the role of monocytes/macrophages in ovula- factor (VEGF) expression closely correlates to the dynamiction. TECK was first described in the development of T changes that take place in the ovary. In particular, a dra-cells in the thymus, but now has a well-accepted role as a matic increase in angiogenesis occurs before ovulation and achemokine that recruits cells to sites of inflammation fine network of capillaries develops and infiltrates the theca[35,36]. Neutralization of TECK with specific antibodies layer. An increase also occurs immediately after ovulation toinhibited leukocyte infiltration into the ovary by 85%, generate an extensive capillary network in the developingresulting in a lack of ovulation. Interestingly, ovulatory corpus luteum (CL) [47]. The source of VEGF that drives thisfailure has been attributed to the lack of infiltration of a active angiogenesis has not yet been clearly determined.rare CD8a+ T cell population [37,38]. Consistent with this Although a body of literature suggests VEGF is expressed byfinding are reports that ovarian TECK expression is tightly TC, GC and the CL [48,49], monocytes, macrophages andregulated by gonadotropins [39]. neutrophils also produce VEGF in many scenarios [50,51]. Two fascinating aspects of ovarian leukocyte infiltration Macrophages isolated from human follicular aspirates upre-are the speed and extent to which it takes place. In rats, gulate VEGF production more than fivefold upon stimula-this increased expression of CAMs and corresponding tion with hCG or LH [52]. Although the function of MMPsinfiltration commences in as little as 1–3 hours after and VEGF appear mutually exclusive, several investigatorsovulatory gonadotropin stimulation. This is an extremely have shown a mutual regulation between MMPs and VEGFfast event, particularly in contrast to inflammation caused [53]. It is expected that more detailed information on theby infection in which infiltration of leukocytes occurs over leukocyte function in the ovary will emerge as new assayseveral days [40]. By contrast, preovulatory leukocyte methods and animal models are developed.infiltration is not an occasional event but a frequent onebecause leukocyte infiltration takes place each time ovula- HPO axis in regulating ovulationtion occurs, which is every 4–5 days in rodents and ap- The hypothalamus, pituitary and ovary have long beenproximately once per month in women. considered to constitute the axis of a regulatory loop that controls ovulation. These three key organs, the HPO axis,Roles of ovarian leukocytes in ovulation communicate between one another via hormonal signals.Leukocytes are involved in three main aspects of ovarian The result is cyclic hormonal changes that result in thefunction: (i) loosening of the follicular wall to facilitate periodic expulsion of eggs in the process known as ovula-follicle growth and ovulation, (ii) tissue repair following tion. Gonadotropin releasing hormone (GnRH) secretedfollicle rupture, and (iii) luteal formation and regression. from the hypothalamus stimulates pituitary gonadotrophsOvulation, as well as the events that follow, requires major to synthesize and release the gonadotropins, follicle-stim-modifications in the extracellular matrix (ECM), and these ulating hormone (FSH) and LH. These two hormones thenrearrangements of the ECM involve tightly controlled exert their effects on the ovary, leading to the growth andproduction of tissue proteases. Matrix metalloproteinases maturation of follicles and the expulsion of the oocyte [54–(MMPs) are a family of soluble and membrane type (MT- 57]. The ovary is a complex organ, comprised of folliclesMMPs) zinc-dependent endopeptidases [41]. Both follicu- that are at different stages of development including thelar cells (GC and TC) and leukocytes are known to produce quiescent primordial, primary, small pre-antral, antral,MMPs [42–44]. However, further studies are needed for and large antral (or preovulatory) follicles. During follicu-better characterization of the cells types responsible for lar growth, GCs surround the oocyte, a basement mem-production of each MMP subtype. brane forms, and a TC layer develops and surrounds the Leukocytes were recently described as major producers of follicle. The two cell layers act cooperatively in the produc-MMP-9, the most abundant MMP found in the preovulatory tion of steroid hormones. TCs produce androgens thatovary [45]. MMP-producing cells were identified as mono- traverse the basement membrane to the GCs, where aro-cytes/macrophages and granulosa cells are the major pro- matase converts these androgens to estrogens that regu-ducers of inhibitors of MMPs (TIMPs). In inflamed tissues, late FSH and LH release from the pituitary [54].as monocytes move through tissue they secrete MMPs thatdigest matrix proteins, facilitating migration through tis- Inflammation and ovulationsues to the sites of inflammation. It is feasible that ovarian Many hallmarks of inflammation are also observed in themonocytes produce one class of MMPs for migration pur- ovary at the time of ovulation. Similar vascular changesposes and, upon tissue specific differentiation, produce more include increased blood flow and vascular permeability,potent MMPs that could further facilitate matrix break- and cellular events such as increased leukocyte extravasa-down. Interestingly, although the major role of MMPs in the tion and activation occur at the site of inflammation and inovary is related to their function in ECM breakdown, the the ovulating ovary. In addition, chemical mediators in-substrates for MMPs are not restricted to matrix proteins. cluding prostaglandins, vasoactive amines (histamine and 347
  4. 4. Opinion Trends in Endocrinology and Metabolism September 2011, Vol. 22, No. 9serotonin), cytokines and chemokines are produced both in In particular, the ovary utilizes the same molecular signalsinflammatory responses and ovulation. The characteristic that attract leukocytes via the mechanism that governstissue damage, repair and remodeling that result from their infiltration at the site of infections or injuriesinflammation in non-ovarian tissues also occur in the [11,15,74,75]. Thus, does the spleen serve as a source ofovary during ovulation [58–64]. Therefore, ovulation is infiltrating leukocytes during this period of ovulatory in-now considered an outcome of acute inflammatory reac- flammation? To answer this question, a study recentlytions in the ovary. measured sequential changes of leukocyte content in the Leukocytes as the main mediators of ovarian inflamma- ovary and spleen after inducing superovulation, by inject-tory responses are a major target of investigation. Leuko- ing gonadotropins [a bolus injection with pregnant marecytes circulate in the blood, become attracted by serum gonadotropin (PMSG) to stimulate follicular growthchemokines and adhesion molecules in inflamed tissues, followed 48 h later by hCG injection to induce ovulation]and traverse the blood vessel wall infiltrating interstitial [5]. Flow cytometry was employed to count the actualtissues to reach their sites of action. Cytokines that are numbers of leukocytes in these two distal organs usinginitially released by the inflamed tissue play a crucial role CD45-specific fluorescent antibodies. This approachin increasing adhesion molecule expression on endothelial revealed that as intraovarian leukocyte numbers in-cells and in upregulating the corresponding receptor ex- creased, leukocyte numbers in the spleen decreased sharp-pression on leukocytes, both of which greatly enhance ly. The same inverse relationship was observed in adultleukocyte migration to the target tissues [65,66]. At sites rats during the period of proestrus to estrus, when ovula-of inflammation, leukocytes and vascular endothelial cells tory inflammation occurs. Lower numbers of leukocytesrelease chemokines and cytokines that accelerate leuko- infiltrated the ovary upon superovulation induction incyte recruitment and modulate leukocyte function. Even- splenectomized rats [5]. Together, these findings stronglytually, however, the main function of leukocytes in the indicate that the spleen supplies leukocytes to the preovu-ovary is exerted through release of proteases [67,68]. A latory ovary.number of molecules, including diverse cytokines, chemo- These findings raise the interesting question, should thekines, and proteases that are commonly associated with spleen be considered a key component of the reproductiveimmunological responses, are present in the preovulatory axis in regulating ovulation? Are spleen leukocytes underhuman ovary as well as in ovaries in animal models [60– the regulation of LH, progesterone and/or prostaglandins63]. Therefore, the past 30 years of research have clearly whose ovarian functions are crucial for successful ovula-demonstrated the significance of infiltrating leukocytes in tion? How does the HPO axis communicate with the spleenovarian function. Much of this work implies that the to trigger the preovulatory leukocyte release? Unfortu-infiltration of leukocytes in periovulatory period is a key nately, none of these questions can be clearly answeredevent in ovulation [69]. However, the origin of these infil- at present because very little research on the spleen hastrating leukocytes, their phenotype, and the mechanisms been done in relation to its reproductive function. However,that govern their trafficking to the ovary are elusive. a glimpse of the role the spleen might play in ovulation can be gathered from past and current literature.Adding the spleen to the HPO axisLeukocytes are hematopoietic in origin and are produced in The impact of splenectomy on female fertilitythe bone marrow. Upon release into the bloodstream they The removal of the entire spleen, splenectomy, has been acirculate and infiltrate inflamed tissues or are stored in successful surgical procedure for many hematological, im-lymphoid organs, such as the spleen, for future activation munological, and traumatic conditions. However, althoughand release. The spleen releases leukocytes following in- the removal is advantageous in treating the specific dis-duction of acute inflammation in the heart by ischemic orders, it leaves the patient with a significant defect in bothmyocardial injury [7,70,71]. Using a sophisticated ap- innate and adaptive immune responses. As a consequence,proach that involved the transplantation of spleens from splenectomized patients have a 60–100-fold increased riskGFP mice into wild-type mice, it was demonstrated that of sepsis [76]. The ovulatory consequences of splenectomyspleen leukocytes infiltrate heart tissues during acute in humans, however, are difficult to assess because otherinflammation. This finding indicates that at least one role treatment regimens such as chemotherapy or radiotherapyof the spleen is to act as an immediate supplier of leuko- that often accompany the splenectomy procedure alsocytes for tissues that experience acute inflammation. This result in severe damage to ovulatory function. However,concept is supported by the fact that, upon stimulation by there are several animal studies that indicate a role of thean inflammatory signal, the bone marrow takes days to spleen in ovarian function. In particular, studies in rodentsproduce leukocytes whereas spleen leukocytes reach sites and rabbits show that splenectomy results in either a delayof inflammation within minutes to hours [7,72,73]. in ovulation [77], aberrant corpus luteal function [78], or an Upon ovulatory gonadotropin stimulation the ovary absence of leuteolysis [79,80].experiences an acute inflammatory response. This inflam-mation is different from responses to infectious insults Proposed method of communication between the HPObecause ovulatory inflammation is in response to a normal axis and the spleenphysiological event, LH stimulation. However, the nature How would the HPO axis communicate with spleen toand sequence of the inflammatory events occurring in the induce spleen leukocyte release? Because leukocytes arepreovulatory ovary are essentially identical to those reac- released as early as one hour after ovulatory LH surge, LHtions taking place at the sites of infectious inflammation. could directly stimulate spleen (Figure 1). However, this is348
  5. 5. [()TD$FIG] Opinion Trends in Endocrinology and Metabolism September 2011, Vol. 22, No. 9 Hypothalamus Hypothalamus Anterior pituitary GnRH ? Pituitary Spleen E2 LH ? Leukocytes Ovary TRENDS in Endocrinology & MetabolismFigure 1. Proposed interplay between the HPO axis and spleen. Preovulatory rise of E2 (estradiol) stimulates release of LH into the bloodstream. LH then triggers leukocyterelease from the spleen either by acting directly on leukocytes or through indirect effects on spleen tissues. Spleen leukocytes released into the bloodstream migrate to theovary in response to cytokines and chemokines that act as leukocyte attractants; the cells then enter the tissue through interactions between leukocyte receptors andadhesion molecules on the endothelial cells.[()TD$FIG] White pulp Spleen MZ Red pulp Theca cells Ovary Granulosa cells Key: Chemokine receptors Chemokines Leukocyte expressed CAM Endothelial cell expressed CAM TRENDS in Endocrinology & MetabolismFigure 2. Spleen leukocyte trafficking to the ovary. Spleen reservoir leukocytes are tethered in the open blood system of the red pulp through either specific chemokine–chemokine receptor or adhesion-molecule interactions. Upon LH surge, changes in leukocyte CCR expression or the reduced chemokine production by reticular fibroblast withinthe red pulp results in the mobilization of spleen reservoir leukocytes into the bloodstream. Upon arriving at the ovary, LH-mediated upregulation of adhesion moleculeexpression on ovarian endothelial cells, together with LH-mediated vasodilation, increases leukocyte adherence to the endothelial cells. Once tethered to the ovarian endothelialcells, leukocytes respond to follicle-produced chemokines. The migration of leukocytes through the interstitial space towards the source (GC, TC or ovarian leukocytes) results inthe production of MMPs to facilitate movement through the ECM. At the mature follicle, leukocytes become activated by locally produced cytokines and produce reactive oxygenspecies (ROS), MMPs and proteolytic granules that weaken the basement membrane of the follicle, enabling the release of the oocyte. MZ, marginal zone. 349
  6. 6. Opinion Trends in Endocrinology and Metabolism September 2011, Vol. 22, No. 9unlikely because the presence of LH receptors in the spleen Concluding remarksis reported only in poultry [81], and the spleen does not The immune system is not only important for battlingexpress LH receptors in mammals [82]. Instead, an endo- foreign invaders but is also essential for female reproduc-crine molecule(s) produced by the ovary in response to LH tion. We propose here that the spleen can bridge thestimulation could travel to the spleen via the circulation immune and reproductive systems by serving as a reser-and stimulate leukocyte release (Figure 1). In the case of voir for the leukocytes required for the inflammatorythe myocardial injury model, angiotensin (Ang) II was events that regulate ovulation. Rigorous collaborativeshown to have such activity [7]. It will be interesting to studies between the fields of immunology and reproductivedetermine if Ang II has this role in the periovulatory biology will be required to validate this hypothesis andrelease of spleen leukocytes. In support of the potential elucidate the interaction between the reproductive organsrole of angiotensin II, studies indicate that ovarian Ang II and the spleen.synthesis and secretion increases immediately after ovu-latory LH/hCG stimulation and ovulation is inhibited if Acknowledgments We thank Dr. Thomas E. Curry for critical comments and Mr. Tom DolanPD123319 or Saralasin, Ang II receptor antagonists, are for the artwork. This work was supported by National Institutes ofinjected into superovulation-induced rabbits and rats, re- Health grant RO1HD052694 (to C.K.).spectively [83–85]. Other candidate molecules that couldoriginate from the ovary and induce spleen leukocyte Referencesrelease include chemokines and cytokines that are pro- 1 Urman, B. and Yakin, K. (2006) Ovulatory disorders and infertility. J.duced by the ovary upon LH stimulation. It will be inter- Reprod. Med. 51, 267–282esting to determine whether receptors for these ovary- 2 Gibson, M. (1995) Reproductive health and polycystic ovary syndrome. Am. J. Med. 98, 67S–75Sborne molecules are present in the spleen and if the 3 Mitwally, M.F. and Casper, R.F. (2006) Potential of aromatasereceptors are localized in leukocytes or spleen cells. It is inhibitors for ovulation and superovulation induction in infertilehowever considered that the spleen leukocyte release women. Drugs 66, 2149–2160could be a homeostatic response to the decrease in circu- 4 Cremer, M. et al. (2010) Recent innovations in oral contraception.lating leukocytes because many of them infiltrate the Semin. Reprod. Med. 28, 140–146 5 Oakley, O.R. et al. (2010) Periovulatory leukocyte infiltration in theovary. Taken together, we propose the following path of rat ovary. Endocrinology 151, 4551–4559spleen leukocyte trafficking to the ovary (Figure 2). The 6 Hedin, L. (2010) Invaders from the spleen: an unexpected origin ofleukocytes reside in steady-state equilibrium within the the leukocytes participating in ovulation. Endocrinology 151,red pulp of the spleen where they are held via chemokine 4096–4099 7 Swirski, F.K. et al. (2009) Identification of splenic reservoir monocytesreceptors (CCRs). The LH surge could alter CCR expres- and their deployment to inflammatory sites. Science 325, 612–616sion on the leukocytes and/or induce chemokine expres- 8 Kitaya, K. and Yamada, H. (2010) Pathophysiological roles ofsion in the spleen endothelial cells, thereby stimulating chemokines in human reproduction: an overview. Am. J. Reprod.mobilization of the leukocytes through the endothelial cell Immunol. 65, 449–459layer of the blood vessel and into the circulation. Leuko- 9 Runesson, E. et al. (1996) The human preovulatory follicle is a source of the chemotactic cytokine interleukin-8. Mol. Hum. Reprod. 2,cytes circulate through the periphery until they reach the 245–250ovary where an interaction with appropriate adhesion 10 Arici, A. et al. (1996) Interleukin-8 expression and modulation inmolecules on the endothelial cell wall occurs. Simulta- human preovulatory follicles and ovarian cells. Endocrinology 137,neously, in the ovary the LH surge decreases blood flow by 3762–3769dilating vessels and increasing the expression of leukocyte 11 Bukulmez, O. and Arici, A. (2000) Leukocytes in ovarian function. Hum. Reprod. Update 6, 1–15receptors such as adhesion molecules to promote the 12 Karaca, T. et al. (2008) Distribution and heterogeneity of mast cells ininteraction of leukocytes with receptors on ovarian endo- female reproductive tract and ovary on different days of the oestrusthelial cells. cycle in Angora goats. Reprod. Domest. Anim. 43, 451–456 13 Smith, M.P. et al. (2005) Leukocyte origin and profile in follicularLeukocytes in other reproductive organs aspirates at oocyte retrieval. Hum. Reprod. 20, 3526–3531 14 Bonello, N. et al. (2004) Periovulatory expression of intercellularLeukocytes also play crucial roles in reproductive tissues adhesion molecule-1 in the rat ovary. Biol. Reprod. 71, 1384–1390other than the ovary, including the uterus [86–88], pitui- 15 Rohm, F. et al. (2002) Correlation between expression of selectins andtary [89,90], oviduct [91–93], testis [94], and vagina [95– migration of eosinophils into the bovine ovary during the97]. Thus, implantation, pregnancy maintenance, embryo periovulatory period. Cell Tissue Res. 309, 313–322 16 Robker, R.L. et al. (2010) The inflammatory response at ovulation isdevelopment, menstrual tissue shedding and many other altered in ovaries of progesterone receptor null (PRKO) mice., In Inreproductive functions are regulated by leukocytes. With 43rd Annual Meeting of the Society for the Study of Reproduction,the recent finding that the spleen is a reservoir for leuko- abstract #95cytes that rapidly respond to perturbation, it is likely that 17 Simon, C. et al. (1994) Interleukin-1 receptor antagonist suppressesthe spleen could also serve as the leukocyte reservoir for human chorionic gonadotropin-induced ovulation in the rat. Biol. Reprod. 51, 662–667these reproductive tissues. In fact, we found a discrepancy 18 Langer, H.F. and Chavakis, T. (2009) Leukocyte–endothelialbetween the numbers of leukocytes that leave the spleen interactions in inflammation. J. Cell. Mol. Med. 13, 1211–1220upon LH stimulation and the combined increase of leuko- 19 Murayama, C. et al. (2010) Effect of VEGF (vascular endothelialcyte numbers in the bloodstream and ovary [5], indicating growth factor) on expression of IL-8 (interleukin-8) IL-1beta andthat large numbers of spleen leukocytes migrate to other their receptors in bovine theca cells. 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