Treg Cell Pool Research Articles

Peripheral regulatory T lymphocytes recirculating to the thymus suppress the development of their precursors.

Most T lymphocytes, including regulatory T cells (Treg cells), differentiate in the thymus. The age-dependent involution of this organ leads to decreasing production of T cells. Here we found that the output of new Treg cells from the thymus decreased substantially more than that of conventional T cells. Peripheral mouse and human Treg cells recirculated back to the thymus, where they constituted a large proportion of the pool of Treg cells and displayed an activated and differentiated phenotype. In the thymus, the recirculating cells exerted their regulatory function by inhibiting interleukin 2 (IL-2)-dependent de novo differentiation of Treg cells. Thus, Treg cell development is controlled by a negative feedback loop in which mature progeny cells return to the thymus and restrain development of precursors of Treg cells.

A subpopulation of CD103(pos) ICOS(pos) Treg cells occurs at high frequency in lymphopenic mice and represents a lymph node specific differentiation stage.

Regulatory T (Treg) cells are pivotal for the maintenance of peripheral tolerance by controlling self-reactive, chronic, and homeostatic T-cell responses. Here, we report that the increase in Treg-cell suppressive function observed in lymphopenic mice correlates with the degree of lymphopenia and is caused by a higher frequency of a novel subpopulation of CD103(pos) ICOS(pos) Treg cells. Though present in the thymus, CD103(pos) ICOS(pos) Treg cells are not generated there but recirculate from the periphery to that site. The acquisition and maintenance of this distinctive phenotype requires the LN microenvironment and the in situ availability of antigen. Contrary to conventional effector and other Treg cells, the cellularity of CD103(pos) ICOS(pos) Treg cells is not affected by the absence of IL-7 and thymic stroma lymphopoetin. Given their increased frequency in lymphopenia, the absolute number of CD103(pos) ICOS(pos) Treg cells remains unchanged in the periphery irrespective of a paucity of total Treg cells. We furthermore demonstrate, with cell transfers in mice, that the CD103(pos) ICOS(pos) phenotype represents a LN-specific differentiation stage arrived at by several other Treg-cell subsets. Thus, tissue-specific cues determine the overall potency of the peripheral Treg-cell pool by shaping its subset composition.

Membrane progesterone receptors in human regulatory T cells: a reality in pregnancy.

To provide evidence of the existence of membrane progesterone receptor alpha (mPRα) on regulatory T cells (Treg) in peripheral blood during pregnancy, postulating a possible explanation for the effect of progesterone on preterm birth. Cross-sectional study. Tertiary Obstetric Department in a University Hospital. Healthy pregnant women. Treg cells from peripheral blood samples were studied by flow cytometry using multiple monoclonal antibody expression. Evaluate the number and percentage of CD4(+) CD25(high) CD127(low) , the number and percentage of Treg cells among the total CD4(+) T cells, and the percentage and mean fluorescence intensity (MFI) of mPRα in that population, using several gating strategies. 43 peripheral blood samples were collected from healthy women during pregnancy, whose median gestational age was 28.7±7.1 (16-40) weeks. The percentage of CD4(+) in the total lymphocytes was 43% (32-51) and the percentage of CD4(+) CD25(high) CD127(low) was 4.8% (1.6-5.9), with only 45% (16-72) of those cells expressing the intracellular marker FoxP3 (Treg cell pool). We confirmed the existence of mPRα in that specific population because 8.0% (2.02-33) of the Treg cells were marked with the specific monoclonal antibody, with an mPRα(+) MFI of 719 (590-1471). This research shows that Treg cells express mPRα during pregnancy, which might play an important role in immune modulation by progesterone.

Both retention and recirculation contribute to long-lived regulatory T-cell accumulation in the thymus.

Natural Treg cells acquire their lineage-determining transcription factor Foxp3 during development in the thymus and are important in maintaining immunologic tolerance. Here, we analyzed the composition of the thymic Treg-cell pool using RAG2-GFP/FoxP3-RFP dual reporter mice and found that a population of long-lived GFP(-) Treg cells exists in the thymus. These long-lived Treg cells substantially increased with age, to a point where they represent >90% of the total thymic Treg-cell pool at 6 months of age. In contrast, long-lived conventional T cells remained at ∼ 15% of the total thymic pool at 6 months of age. Consistent with these studies, we noticed that host-derived Treg cells represented a large fraction (∼ 10%) of the total thymic Treg-cell pool in bone marrow chimeras, suggesting that long-lived Treg cells also reside in the thymus of these mice. The pool of long-lived Treg cells in the thymus was sustained by retention of Treg cells in the thymus and by recirculation of peripheral Treg cells back into the thymus. These long-lived thymic Treg cells suppressed T-cell proliferation to an equivalent extent to splenic Treg cells. Together, these data demonstrate that long-lived Treg cells accumulate in the thymus by both retention and recirculation.

Regulatory T‐cell homeostasis: steady‐state maintenance and modulation during inflammation

Regulatory T (Treg) cells play a vital role in the prevention of autoimmunity and the maintenance of self-tolerance, but these cells also have an active role in inhibiting immune responses during viral, bacterial, and parasitic infections. Although excessive Treg activity can lead to immunodeficiency, chronic infection, and cancer, too little Treg activity results in autoimmunity and immunopathology and impairs the quality of pathogen-specific responses. Recent studies have helped define the homeostatic mechanisms that support the diverse pool of peripheral Treg cells under steady-state conditions and delineate how the abundance and function of Treg cells changes during inflammation. These findings are highly relevant for developing effective strategies to manipulate Treg cell activity to promote allograft tolerance and treat autoimmunity, chronic infection, and cancer.

Extrathymic induction of Foxp3+ regulatory T cells declines with age in a T‐cell intrinsic manner

Extrathymically induced Foxp3⁺ regulatory T (Treg) cells contribute to the pool of Treg cells and are implicated in the maintenance of immune tolerance at environmental interfaces. The impact of T-cell senescence on their generation and function is, however, poorly characterized. We report here that steady-state induction of Foxp3 is impaired in aged T cells in vivo. In vitro assays further revealed that this defective generation of Treg cells was independent from the strength of TCR stimulation and arose before T-cell proliferation. Importantly, they also revealed that this impairment of Foxp3 induction is unrelated to known age-related T-cell defects, such as IL-2 secretion impairment, accumulation of activated T-cell populations, or narrowing of the T-cell repertoire. Finally, a loss of extrathymic induction of Foxp3 and tolerance to minor-mismatched skin graft were observed in aged mice treated by nondepleting anti-CD4 antibody. The T-cell intrinsic impairment of Treg-cell generation revealed here highlights age as a key factor to be considered in immune tolerance induction.

Antiapoptotic Mcl-1 is critical for the survival and niche-filling capacity of Foxp3+ regulatory T cells

Foxp3⁺ regulatory T (Treg) cells are a crucial immunosuppressive population of CD4⁺ T cells, yet the homeostatic processes and survival programs that maintain the Treg cell pool are poorly understood. Here we report that peripheral Treg cells markedly alter their proliferative and apoptotic rates to rapidly restore numerical deficit through an interleukin 2-dependent and costimulation-dependent process. By contrast, excess Treg cells are removed by attrition, dependent on the Bim-initiated Bak- and Bax-dependent intrinsic apoptotic pathway. The antiapoptotic proteins Bcl-xL and Bcl-2 were dispensable for survival of Treg cells, whereas Mcl-1 was critical for survival of Treg cells, and the loss of this antiapoptotic protein caused fatal autoimmunity. Together, these data define the active processes by which Treg cells maintain homeostasis via critical survival pathways.

457 REGULATORY T CELLS IN CLINICALLY LOCALIZED RENAL CELL CARCINOMA: COMPARISON WITH AN AGE-MATCHED HEALTHY CONTROL

You have accessJournal of UrologyKidney Cancer: Basic Research (III)1 Apr 2013457 REGULATORY T CELLS IN CLINICALLY LOCALIZED RENAL CELL CARCINOMA: COMPARISON WITH AN AGE-MATCHED HEALTHY CONTROL Taekmin Kwon, In Gab Jeong, Yunlim Kim, Jong Hyun Yoon, Kwang Hyun Kim, Dalsan You, Jun Hyuk Hong, Hanjong Ahn, and Choung-Soo Kim Taekmin KwonTaekmin Kwon Seoul, Korea, Republic of More articles by this author , In Gab JeongIn Gab Jeong Seoul, Korea, Republic of More articles by this author , Yunlim KimYunlim Kim Seoul, Korea, Republic of More articles by this author , Jong Hyun YoonJong Hyun Yoon Seoul, Korea, Republic of More articles by this author , Kwang Hyun KimKwang Hyun Kim Seoul, Korea, Republic of More articles by this author , Dalsan YouDalsan You Seoul, Korea, Republic of More articles by this author , Jun Hyuk HongJun Hyuk Hong Seoul, Korea, Republic of More articles by this author , Hanjong AhnHanjong Ahn Seoul, Korea, Republic of More articles by this author , and Choung-Soo KimChoung-Soo Kim Seoul, Korea, Republic of More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1848AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES There has been increasing evidence for focusing on the role of regulatory T (Treg) cells in cancer immunity. However, the literature regarding the frequency of regulatory T cells (Treg) in renal cell carcinoma (RCC) compared with normal control is controversial. Also, there is increasing evidence for age-dependent development of Treg cells. We investigated the frequency of Tregs in the peripheral blood of RCC patients compared with age-matched healthy control and compared the prevalence of Treg cells in the tumor infiltrating lymphocytes (TIL) with lymphocyte in normal kidney tissue. METHODS The frequency of Treg cells in the peripheral blood of 62 patients with clinically localized RCC and from 65 healthy controls was evaluated by flow cytometry using specific monoclonal antibodies recognizing CD4(+), CD25 (+) and Foxp3 (+) markers. The prevalence of Treg cells in the TIL and lymphocyte in normal kidney tissue from 54 patients with RCC were also evaluated. RESULTS The median age of the RCC and control group was 55 (<40 years: n=8, 40-49: n=13, 50-59: n=16, ≥60: n=25) and 53 (<40 years: n=9, 40-49: n=18, 50-59: n=16, ≥60: n=22) years, respectively. The median tumor size was 6.0cm (1.7-13.7cm) and 49 patients (79%) had clear cell histology on pathologic examination. There was no difference in the frequency of Treg cells in the peripheral blood in entire RCC patients compared with healthy donors (2.8 ± 4.0% vs 1.9 ± 2.6%, p=0.150). Treg cells frequency was found to significantly higher in RCC patients with large tumor size (≥7cm, 4.2 ± 5.5%) than those from normal healthy control or small to medium sized RCC patients (<7cm, 2.0 ± 2.6%, p=0.016). Furhman nuclear grade did not affect the frequency of Treg cells among RCC patients (Gr 1-2: 2.5 ± 3.6% vs Gr 3-4: 3.0 ± 4.4%, p=0.675). There was no statistically significant difference in the prevalence of Treg cells between TIL and lymphocyte in normal kidney tissue (5.5 ± 9.5% vs 4.8 ± 5.8%, p=0.629). CONCLUSIONS We provide evidence of an increased pool of Treg cells in the peripheral blood of clinically localized RCC patients with substantial tumor burden compared with age-matched healthy control. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e187-e188 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Taekmin Kwon Seoul, Korea, Republic of More articles by this author In Gab Jeong Seoul, Korea, Republic of More articles by this author Yunlim Kim Seoul, Korea, Republic of More articles by this author Jong Hyun Yoon Seoul, Korea, Republic of More articles by this author Kwang Hyun Kim Seoul, Korea, Republic of More articles by this author Dalsan You Seoul, Korea, Republic of More articles by this author Jun Hyuk Hong Seoul, Korea, Republic of More articles by this author Hanjong Ahn Seoul, Korea, Republic of More articles by this author Choung-Soo Kim Seoul, Korea, Republic of More articles by this author Expand All Advertisement Advertisement PDF DownloadLoading ...

Seminal Fluid and the Generation of Regulatory T Cells for Embryo Implantation

T regulatory (Treg) cells are essential mediators of the maternal immune adaptation necessary for embryo implantation. In mice, insufficient Treg cell activity results in implantation failure, or constrains placental function and fetal growth. In women, Treg cell deficiency is linked with unexplained infertility, miscarriage, and pre-eclampsia. To devise strategies to improve Treg cell function, it is essential to define the origin of the Treg cells in gestational tissues, and the regulators that control their functional competence and recruitment. Male seminal fluid is a potent source of the Treg cell-inducing agents TGFβ and prostaglandin E, and coitus is one key factor involved in expanding the pool of inducible Treg cells that react with paternal alloantigens shared by conceptus tissues. In mice, coitus initiates a sequence of events whereby female dendritic cells cross-present seminal fluid antigens and activate T cells, which in turn circulate via the blood to be sequestered into the endometrium. Similar events may occur in the human genital tract, where seminal fluid induces immune cell changes that appear competent to prime Treg cells. Improved understanding of how seminal fluid influences Treg cells in women should ultimately assist in the development of new therapies for immune-mediated pathologies of pregnancy.

The extent of HLA-DR expression on HLA-DR+Tregs allows the identification of patients with clinically relevant borderline rejection

Regulatory T cells (Tregs) were shown to be involved into the pathogenesis of acute rejection after transplantation. The suppressive activity of the total regulatory T cell pool depends on its percentage of highly suppressive HLA-DR(+) -Treg cells. Therefore, both the suppressive activity of the total Treg pool and the extent of HLA-DR expression of HLA-DR(+) -Tregs (MFI HLA-DR) were estimated in non transplanted volunteers, patients with end-stage renal failure (ESRF), healthy renal transplant patients with suspicion on rejection, due to sole histological Bord-R or sole acute renal failure (ARF), and patients with clinically relevant borderline rejection (Bord-R and ARF). Compared to patients with only Bord-R or only ARF, the suppressive activity of the total Treg cell pool was exclusively reduced in patients with clinically relevant Bord-R. In parallel, the HLA-DR MFI of the DR(+) -Treg subset was significantly decreased in these patients, due to a significantly lower proportion of DR(high+) -Tregs, which were shown to have the highest suppressive capacity within the total Treg pool. Our findings clearly demonstrate that the determination of the HLA-DR MFI of the HLA-DR(+) -Treg subset allows a highly sensitive, specific and non-invasive discrimination between patients with clinically relevant Bord-R (Bord and ARF) and patients with subclinical rejection or other causes of transplant failure.

Trafficking of regulatory T cells in the intestinal immune system

Intestinal tolerance toward dietary antigens and the microbiota critically depends on regulatory T cells (T(reg) cells). To exert their function, T(reg) cells need to get into the proximity of their target cells foremost within lymphoid organs and the lamina propria in the small and large intestines. Thus, T(reg)-cell-mediated immunosuppression is intrinsically linked to cell migration and localization. However, the gut lamina propria not only constitutes an effector site but also actively influences T(reg)-cell function. In particular, gut-resident macrophages take an active role in shaping the T(reg)-cell pool. Here, the author summarizes recent observations to suggest that T(reg)-cell migration not only directs cells to sites of immunosuppression but also allows for the coordinated shaping of the T(reg)-cell population.

The Extent of HLA-DR Expression on HLA-DR+ Tregs Allows the Identification of Patients with Clinically Relevant Borderline Rejection

Introduction: Regulatory T cells (Tregs) were shown to be involved into the pathogenesis of acute rejection after transplantation. The suppressive activity of the total regulatory T cell pool depends on its percentage of highly suppressive HLA-DR+-Treg cells. Methods: Therefore, both the suppressive activity of the total Treg pool and the extent of HLA-DR expression of HLA-DR+-Tregs (MFI HLA-DR) were estimated in non transplanted volunteers, patients with end-stage renal failure (ESRF), healthy renal transplant patients with suspicion on rejection, due to sole histologic Bord-R or sole acute renal failure (ARF), and patients with clinically relevant borderline rejection (Bord-R and ARF). Results: Compared to healthy non-transplanted controls, the suppressive activity of the total Treg cell pool was exclusively reduced in patients with clinically relevant Bord-R. In parallel, the HLA-DR MFI of the DR+-Treg subset was significantly decreased in these patients, due to a significantly lower proportion of DRhigh+-Tregs, which were shown to have the highest suppressive capacity within the total Treg pool.Figure: [HLA-DR MFI of the DR+-Tregs]Conclusion: Our findings clearly demonstrate that the determination of the HLA-DR MFI of the HLA-DR+-Treg subset allows a highly sensitive, specific and non-invasive discrimination between patients with clinically relevant Bord-R (Bord and ARF) and patients with subclinical rejection or other causes of transplant failure.

Methylprednisolone treatment increases the proportion of the highly suppressive HLA-DR+-Treg-cells in transplanted patients

Methylprednisolone is widely used to improve immune suppression in transplanted patients threatened by acute rejection. Recently, we showed that the suppressive activity of a Treg cell population depends decisively on their percentage of highly suppressive HLA-DRhigh+-Treg cells, which are strongly reduced in rejecting transplant patients.In order to examine whether the composition of the total CD4+CD127low+/−FoxP3+‐Treg cell pool with different Treg-subsets (DRhigh+CD45RA−-Tregs, DRlow+CD45RA−-Tregs, DR−CD45RA−-Tregs, DR−CD45RA+-Tregs) is affected by methylprednisolone bolus therapy we compared the percentages of these four different Treg cell subsets in transplant patients with biopsy proven rejection before and after steroid bolus therapy (n=23).In patients treated with steroid bolus therapy, the percentage of the naïve DR−CD45RA+-Tregs was significantly decreased, whereas the percentage of the DR+CD45RA−-Tregs was significantly increased. By that, the strongest increase was detected for the most suppressive DRhigh+CD45RA−-Tregs. However, these effects were only temporarily and closely associated to the duration of the bolus therapy.Our results suggest that besides various anti-inflammatory effects on cells of the adaptive and innate immune system, methylprednisolone also has the capacity to enhance the suppressive activity of the total Treg cell pool by increasing its percentage of highly differentiated and highly suppressive DRhigh+CD45RA−-Tregs.

Term and preterm labor: decreased suppressive activity and changes in composition of the regulatory T‐cell pool

Regulatory T cells (Tregs) exert a key role in tolerance induction to the semi-allogeneic fetus. Currently, it is not known whether immunological rejection processes are involved in the induction of normal term or irresistible preterm labor. In this study, we examined whether there were differences in the percentage of the total CD4(+)CD127(low+/-)CD25(+)FoxP3(+)-Treg-cell pool, its suppressive activity and its composition with distinct Treg subsets (HLA-DR(low+)-, HLA-DR(high+)-, HLA-DR(-)- and naive CD45RA(+)-Tregs) between preterm and term laboring women. We found that its percentage was decreased neither in term nor in preterm laboring women. Its suppressive activity was strongly diminished in preterm laboring women and to a lesser extent in spontaneously term laboring women. During the normal course of pregnancy, its composition changed in such a way that the percentage of naive CD45RA(+)-Tregs increased while the percentage of the highly suppressive HLA-DR(low+)- and HLA-DR(high+)-Tregs decreased significantly until term. With the onset of spontaneous term labor this phenomenon was reversed and reached significant values postpartum. In addition, we confirmed that both the decreased percentage of HLA-DR(+)-Tregs within the total Treg-cell pool and their decreased level of HLA-DR expression (depending on the percentage of HLA-DR(low+)- and HLA-DR(high+)-Tregs) had a reducing effect on the suppressive activity of the total Treg cell pool in preterm laboring women. However, spontaneous term delivery was associated with increasing percentages of HLA-DR(+)-Tregs and increasing HLA-DR expression of this Treg subset. Therefore, it becomes apparent that the mechanisms inducing term or preterm labor may be completely different.

Enhanced GITR/GITRL interactions augment IL‐27 expression and induce IL‐10‐producing Tr‐1 like cells

The glucocorticoid-induced TNFR-related (GITR) protein is a coactivating receptor that is constitutively expressed on Treg cells and induced on activated T cells. To better under-stand the role of long-term GITR signaling, we generated a mouse that constitutively expresses GITR ligand (GITRL) on APCs that mimics the physiological distribution of GITRL in vivo. Despite a five-fold expansion of the Treg-cell pool, there is increased activation and depletion of naive T cells in the transgenic (Tg) mice, suggesting that the increased number of Treg cells cannot fully suppress T-cell activation. Interestingly, GITRL Tg mice have multiorgan lymphocytic infiltrates yet display no overt autoimmunity, indicating the existence of a compensatory immunoregulatory mechanism(s). In the spleens and tissue infiltrates ofGITRL Tg mice, we found increased numbers of Foxp3(-) IL-10-producing type 1 regulatory T (Tr-1)-like cells that suppress naïve T-cell proliferation in an IL-10-dependent fashion. Increased IL-27 production from Tg APCs and activation of c-Maf in the Tr1-like cells suggest a possible mechanism for their induction. Our results demonstrate that enhanced GITR/GITRL interactions have a pleiotropic role on the regulation of T-cell responses, which includes promoting the differentiation of Tr-1-like cells, which contribute to the maintenance of peripheral T-cell tolerance.

Interleukin-7 Influences FOXP3+CD4+ Regulatory T Cells Peripheral Homeostasis

Mechanisms governing peripheral CD4+ FOXP3+ regulatory T cells (Treg) survival and homeostasis are multiple suggesting tight and complex regulation of regulatory T cells homeostasis. Some specific factors, such as TGF-β, interleukin-2 (IL-2) and B7 costimulatory molecules have been identified as essentials for maintenance of the peripheral Treg compartment. Conversely, Treg dependency upon classical T cell homeostatic factors such as IL-7 is still unclear. In this work, we formally investigated the role of IL-7 in Treg homeostasis in vivo in murine models. We demonstrated that IL-7 availability regulated the size of peripheral Treg cell pool and thus paralleled the impact of IL-7 on conventional T cell pool. Moreover, we showed that IL-7 administration increased Treg cell numbers by inducing thymic-independent Treg peripheral expansion. Importantly the impact of IL-7 on Treg expansion was detected whether conventional T cells were present or absent as IL-7 directly participates to the peripheral expansion of Treg after adoptive transfer into lymphopenic hosts. Our results definitively identify IL-7 as a central factor contributing to Treg peripheral homeostasis, thus reassembling Treg to other T cell subsets in respect of their need for IL-7 for their peripheral maintenance.

DRhigh+CD45RA−-Tregs Potentially Affect the Suppressive Activity of the Total Treg Pool in Renal Transplant Patients

Recent studies show that regulatory T cells (Tregs) play an essential role in tolerance induction after organ transplantation. In order to examine whether there are differences in the composition of the total CD4+CD127low+/−FoxP3+- Treg cell pool between stable transplant patients and patients with biopsy proven rejection (BPR), we compared the percentages and the functional activity of the different Treg cell subsets (DRhigh+CD45RA−-Tregs, DRlow+CD45RA−-Tregs, DR−CD45RA−-Tregs, DR−CD45RA+-Tregs). All parameters were determined during the three different periods of time after transplantation (0–30 days, 31–1,000 days, >1,000 days). Among 156 transplant patients, 37 patients suffered from BPR. The most prominent differences between rejecting and non-rejecting patients were observed regarding the DRhigh+CD45RA−-Treg cell subset. Our data demonstrate that the suppressive activity of the total Treg pool strongly depends on the presence of these Treg cells. Their percentage within the total Treg pool strongly decreased after transplantation and remained relatively low during the first year after transplantation in all patients. Subsequently, the proportion of this Treg subset increased again in patients who accepted the transplant and reached a value of healthy non-transplanted subjects. By contrast, in patients with acute kidney rejection, the DRhigh+CD45RA−-Treg subset disappeared excessively, causing a reduction in the suppressive activity of the total Treg pool. Therefore, both the monitoring of its percentage within the total Treg pool and the monitoring of the HLA-DR MFI of the DR+CD45RA−-Treg subset may be useful tools for the prediction of graft rejection.

Pregnancy-associated diseases are characterized by the composition of the systemic regulatory T cell (Treg) pool with distinct subsets of Tregs

Dysregulations concerning the composition and function of regulatory T cells (T(regs)) are assumed to be involved in the pathophysiology of complicated pregnancies. We used six-colour flow cytometric analysis to demonstrate that the total CD4(+) CD127(low+/-) CD25(+) forkhead box protein 3 (FoxP3)(+) T(reg) cell pool contains four distinct T(reg) subsets: DR(high+) CD45RA(-), DR(low+) CD45RA(-), DR(-) CD45RA(-) T(regs) and naive DR(-) CD45RA(+) T(regs). During the normal course of pregnancy, the most prominent changes in the composition of the total T(reg) cell pool were observed between the 10th and 20th weeks of gestation, with a clear decrease in the percentage of DR(high+) CD45RA(-) and DR(low+) CD45RA(-) T(regs) and a clear increase in the percentage of naive DR(-) CD45RA(+) T(regs). After that time, the composition of the total T(reg) cell pool did not change significantly. Its suppressive activity remained stable during normally progressing pregnancy, but decreased significantly at term. Compared to healthy pregnancies the composition of the total T(reg) cell pool changed in the way that its percentage of naive DR(-) CD45RA(+) T(regs) was reduced significantly in the presence of pre-eclampsia and in the presence of preterm labour necessitating preterm delivery (PL). Interestingly, its percentage of DR(high+) CD45RA(-) and DR(low+) CD45RA(-) T(regs) was increased significantly in pregnancies affected by pre-eclampsia, while PL was accompanied by a significantly increased percentage of DR(-) CD45RA(-) and DR(low+) CD45RA(-) T(regs). The suppressive activity of the total T(reg) cell pool was diminished in both patient collectives. Hence, our findings propose that pre-eclampsia and PL are characterized by homeostatic changes in the composition of the total T(reg) pool with distinct T(reg) subsets that were accompanied by a significant decrease of its suppressive activity.

Seminal Fluid Regulates Accumulation of FOXP3+ Regulatory T Cells in the Preimplantation Mouse Uterus Through Expanding the FOXP3+ Cell Pool and CCL19-Mediated Recruitment1

Regulatory T (Treg) cells facilitate maternal immune tolerance of the semiallogeneic conceptus in early pregnancy, but the origin and regulation of these cells at embryo implantation is unclear. During the preimplantation period, factors in the seminal fluid delivered at coitus cause expansion of a CD4(+)CD25(+) putative Treg cell population in the para-aortic lymph nodes draining the uterus. Using flow cytometry, immunohistochemistry, and real-time quantitative PCR (qPCR) for the signature Treg cell transcription factor FOXP3, we confirmed the identity of the expanded lymph node population as FOXP3(+) Treg cells and showed that this is accompanied by a comparable increase in the uterus of FOXP3(+) Treg cells and expression of Foxp3 mRNA by Day 3.5 postcoitum. Seminal plasma was necessary for uterine Treg cell accumulation, as mating with seminal vesicle-deficient males failed to elicit an increase in uterine Treg cells. Furthermore seminal fluid induced expression of mRNA encoding the Treg chemokine CCL19 (MIP3beta), which acts through the CCR7 receptor to regulate Treg cell recruitment and retention in peripheral tissues. Glandular and luminal epithelial cells were identified as the major cellular origins of uterine CCL19, and exposure to both seminal plasma and sperm was required for maximum expression. Together, these results indicate that Treg cells accumulate in the uterus prior to embryo implantation and that seminal fluid is a key regulator of the uterine Treg cell population, operating by both increasing the pool of available Treg cells and promoting their CCL19-mediated recruitment from the circulation into the implantation site.

Repeated Seminal Fluid Exposure Expands Regulatory T Cell Populations in Mice.

Regulatory T cells (Treg cells) are required to achieve successful allogeneic pregnancy through their actions in mediating maternal immune tolerance towards conceptus antigens at the time of embryo implantation. In mice, we have shown that seminal fluid exposure is a key factor in maximising the Treg cell pool available systemically and present in the implantation site. Both the seminal plasma and sperm components of seminal fluid contribute to Treg cell activation and proliferation through provision of alloantigens and the immune deviating cytokine transforming growth factor-beta (TGFB), as well as potentially other signalling agents. Additionally, seminal fluid induces uterine expression of chemokines such as CCL19 that facilitate Treg cell recruitment into the uterine endometrium in the peri-implantation period. In women, clinical studies have shown that regular coitus in the absence of barrier methods of contraception over at least a three month period can diminish the incidence of preeclampsia and improve reproductive outcomes. We thus hypothesised that in mice, repeated mating events expand the Treg cell pool beyond that present after a single mating event. To examine the effect of repeated seminal fluid exposure on the maternal Treg cell population, C57BL/6 (B6) female mice were mated once or four times (over a 4-6 week period) to vasectomised (VAS) BALB/c males, then killed on day 3.5 post coitum (pc). Treg cells were quantified by flow cytometry as CD3+CD4+CD25+FoxP3+ cells in the uterus-draining para-aortic lymph nodes, peripheral lymph nodes, spleen and blood. Four matings resulted in an approximately 2-fold increase in the size of the Treg cell population in para-aortic lymph nodes compared with no mating or one mating event. Increases were also seen in the cervical lymph nodes but not the spleen or blood. Surgical removal of the seminal vesicle gland to generate SVX Balb/c males abolished their capacity to elicit expansion of the Treg cell pool. Repeated syngeneic matings of B6 females with VAS B6 males also failed to elicit any further expansion compared with one mating. In conclusion, this study demonstrates that repeated exposure to seminal fluid can expand the Treg cell pool, and that seminal plasma and male alloantigens are involved in eliciting this response. Our findings begin to provide a mechanistic explanation for observations in humans that pregnancy complications are more prevalent after a short duration of sexual relationship or use of barrier methods of contraception. This research was supported by NHMRC Program ID453556 and Fellowship ID627017 awarded to SAR. (platform)