Reproductive biology
Abstract
Materno-fetal immunity possesses specialized characteristics to ensure pathogen clearance while maintaining tolerance to the semiallogeneic fetus. Most of our understanding on human materno-fetal immunity is based on conventional rodent models that may not precisely represent human immunological processes owing to the huge evolutionary divergence. Herein, we developed a pregnant human immune system (HIS) mouse model through busulfan preconditioning, which hosts multilineage human immune subset reconstitution at the materno-fetal interface. Human materno-fetal immunity exhibits a tolerogenic feature at the midgestation stage (embryonic day [E] 14.5), and human immune regulatory subsets were detected in the decidua. However, the immune system switches to an inflammatory profile at the late gestation stage (E19). A cell–cell interaction network contributing to the alternations in the human materno-fetal immune atmosphere was revealed based on single-cell RNA-Seq analysis, wherein human macrophages played crucial roles by secreting several immune regulatory mediators. Furthermore, depletion of Treg cells at E2.5 and E5.5 resulted in severe inflammation and fetus rejection. Collectively, these results demonstrate that the pregnant HIS mouse model permits the development of functional human materno-fetal immunity and offers a tool for human materno-fetal immunity investigation to facilitate drug discovery for reproductive disorders.
Authors
Shuai Dong, Cong Fu, Chang Shu, Min Xie, Yan Li, Jun Zou, Yi-Zi Meng, Peng Xu, Yan-Hong Shan, Hui-Min Tian, Jin He, Yong-Guang Yang, Zheng Hu
Abstract
Embryo implantation is crucial for ensuring a successful pregnancy outcome and subsequent child health. The intrauterine environment during the peri-implantation period shows drastic changes in gene expression and cellular metabolism in response to hormonal stimuli and reciprocal communication with embryos. Here, we performed spatial transcriptomic analysis to elucidate the mechanisms underlying embryo implantation. Transcriptome data revealed that lipid metabolism pathways, especially arachidonic acid–related (AA-related) ones, were enriched in the embryo-receptive luminal epithelia. Cyclooxygenases (COXs), rate-limiting enzymes involved in prostaglandin production by AA, were spatiotemporally regulated in the vicinity of embryos during implantation, but the role of each COX isozyme in the uterus for successful pregnancy was unclear. We established uterine-specific COX2-knockout (uKO) and COX1/uterine COX2-double-KO (COX1/COX2-DKO) mice. COX2 uKO caused deferred implantation with failed trophoblast invasion, resulting in subfertility with reduced pregnancy rates and litter sizes. COX1/COX2 DKO induced complete infertility, owing to abrogated embryo attachment. These results demonstrate that both isozymes have distinct roles during embryo implantation. Spatial transcriptome and lipidome analyses revealed unique profiles of prostaglandin synthesis by each COX isozyme and spatiotemporal expression patterns of downstream receptors throughout the endometrium. Our findings reveal previously unappreciated roles of COXs at the fetomaternal interface to establish early pregnancy.
Authors
Shizu Aikawa, Mitsunori Matsuo, Shun Akaeda, Yukihiko Sugimoto, Makoto Arita, Yosuke Isobe, Yuki Sugiura, Shu Taira, Rae Maeda, Ryoko Shimizu-Hirota, Norihiko Takeda, Daiki Hiratsuka, Xueting He, Chihiro Ishizawa, Rei Iida, Yamato Fukui, Takehiro Hiraoka, Miyuki Harada, Osamu Wada-Hiraike, Yutaka Osuga, Yasushi Hirota
IL-27/Blimp-1 axis regulates the differentiation and function of Tim-3+ Tregs during early pregnancy
Abstract
Decidual regulatory T cells (Tregs) are essential for successful pregnancy outcome. A subset of Tregs, T cell immunoglobulin and mucin domain-containing protein 3–positive regulatory T cells (TregsTim-3+), plays a central role in the acceptance of the fetus during early stages of normal pregnancy. The molecular mechanism regulating the differentiation and function of TregsTim-3+ is unknown. Here, we investigated the role of the transcription factor B lymphocyte-induced maturation protein 1 (Blimp-1) on decidual TregTim-3+ differentiation. We demonstrated that Blimp-1 enhanced the coexpression of negative costimulatory molecules (Tim-3, T cell immunoreceptor with Ig and ITIM domains, and programmed cell death protein 1) on Tregs and improved their immunosuppressive functions, including increased IL-10 secretion, suppression of effector T cell proliferation, and promotion of macrophage polarization toward the M2 phenotype. Furthermore, we showed that IL-27 regulated the expression of Tim-3 and Blimp-1 through the STAT1 signaling pathway and that transfer of TregsBlimp-1+ into an abortion-prone mouse model effectively reduced embryo absorption rate. We postulated that abnormalities in the IL-27/Blimp-1 axis might be associated with recurrent pregnancy loss (RPL). These findings provided insights for developing more efficient immunotherapies for women with RPL.
Authors
Si-Jia Zhao, Xiao-Hui Hu, Xin-Xiu Lin, Yu-Jing Zhang, Jing Wang, Huan Wang, Guang-Shun Gong, Gil Mor, Ai-Hua Liao
Abstract
Spermatogenesis requires precise posttranslational control in the endoplasmic reticulum (ER), but the mechanism remains largely unknown. The protein disulfide isomerase (PDI) family is a group of thiol oxidoreductases responsible for catalyzing the disulfide bond formation of nascent proteins. In this study, we generated 14 strains of KO mice lacking the PDI family enzymes and found that only PDI deficiency caused spermatogenesis defects. Both inducible whole-body PDI-KO (UBC-Cre/Pdifl/fl) mice and premeiotic PDI-KO (Stra8-Cre/Pdifl/fl) mice experienced a significant decrease in germ cells, testicular atrophy, oligospermia, and complete male infertility. Stra8-Cre/Pdifl/fl spermatocytes had significantly upregulated ER stress–related proteins (GRP78 and XBP1) and apoptosis-related proteins (Cleaved caspase-3 and BAX), together with cell apoptosis. PDI deletion led to delayed DNA double-strand break repair and improper crossover at the pachytene spermatocytes. Quantitative mass spectrometry indicated that PDI deficiency downregulated vital proteins in spermatogenesis such as HSPA4L, SHCBP1L, and DDX4, consistent with the proteins’ physical association with PDI in normal testes tissue. Furthermore, PDI served as a thiol oxidase for disulfide bond formation of SHCBP1L. Thus, PDI plays an essential role in protein quality control for spermatogenesis in mice.
Authors
Yaqiong Zhang, Aizhen Yang, Zhenzhen Zhao, Fengwu Chen, Xiaofeng Yan, Yue Han, Depei Wu, Yi Wu
Abstract
MAPK activating death domain (MADD) is a multifunctional protein regulating small GTPases RAB3 and RAB27, MAPK signaling, and cell survival. Polymorphisms in the MADD locus are associated with glycemic traits, but patients with biallelic variants in MADD manifest a complex syndrome affecting nervous, endocrine, exocrine, and hematological systems. We identified a homozygous splice site variant in MADD in 2 siblings with developmental delay, diabetes, congenital hypogonadotropic hypogonadism, and growth hormone deficiency. This variant led to skipping of exon 30 and in-frame deletion of 36 amino acids. To elucidate how this mutation causes pleiotropic endocrine phenotypes, we generated relevant cellular models with deletion of MADD exon 30 (dex30). We observed reduced numbers of β cells, decreased insulin content, and increased proinsulin-to-insulin ratio in dex30 human embryonic stem cell–derived pancreatic islets. Concordantly, dex30 led to decreased insulin expression in human β cell line EndoC-βH1. Furthermore, dex30 resulted in decreased luteinizing hormone expression in mouse pituitary gonadotrope cell line LβT2 but did not affect ontogeny of stem cell–derived GnRH neurons. Protein-protein interactions of wild-type and dex30 MADD revealed changes affecting multiple signaling pathways, while the GDP/GTP exchange activity of dex30 MADD remained intact. Our results suggest MADD-specific processes regulate hormone expression in pancreatic β cells and pituitary gonadotropes.
Authors
Kristiina Pulli, Jonna Saarimäki-Vire, Pekka Ahonen, Xiaonan Liu, Hazem Ibrahim, Vikash Chandra, Alice Santambrogio, Yafei Wang, Kirsi Vaaralahti, Anna-Pauliina Iivonen, Johanna Känsäkoski, Johanna Tommiska, Yasmine Kemkem, Markku Varjosalo, Sanna Vuoristo, Cynthia L. Andoniadou, Timo Otonkoski, Taneli Raivio
Abstract
Fetuses with growth restriction (FGR) have an early activation of hepatic glucose production (HGP), a hallmark of type 2 diabetes (T2D). Here we used fetal hepatic catheterization to directly measure HGP and substrate flux in an FGR sheep model. We hypothesized that FGR fetuses would have increased hepatic lactate and amino acid uptake to support increased HGP. Indeed, FGR compared to normal (CON) fetuses had increased HGP and activation of gluconeogenic genes. Unexpectedly, hepatic pyruvate output was increased while hepatic lactate and gluconeogenic amino acid uptake rates were decreased in FGR fetal liver. Hepatic oxygen consumption and total substrate uptake rates were lower. In FGR liver tissue, metabolite abundance, 13C-metabolite labeling, enzyme activity, and gene expression support decreased pyruvate oxidation and increased lactate production. Isolated hepatocytes from FGR fetuses had greater intrinsic capacity for lactate-fueled glucose production. FGR livers also had lower energy (ATP) and redox state (NADH:NAD+). Thus, reduced hepatic oxidative metabolism may make carbons available for increased HGP but also produces nutrient and energetic stress in FGR fetal liver. Intrinsic programming of these pathways regulating HGP in the FGR fetus may underlie increased HGP and T2D risk postnatally.
Authors
Laura D. Brown, Paul J. Rozance, Dong Wang, Evren C. Eroglu, Randall B. Wilkening, Ashley Solmonson, Stephanie R. Wesolowski
Abstract
Acute bacterial orchitis (AO) is a prevalent cause of intra-scrotal inflammation, often resulting in sub- or infertility. A frequent cause eliciting AO is uropathogenic Escherichia coli (UPEC), a gram negative pathovar, characterized by the expression of various iron acquisition systems to survive in a low-iron environment. On the host side, iron is tightly regulated by iron regulatory proteins (IRP) 1 and 2 and these factors are reported to play a role in testicular and immune cell function, however, their precise role remains unclear. Here, we showed in a mouse model of UPEC-induced orchitis that the absence of IRP1 results in reduced immune response and testicular damage. Compared to infected wild-type (WT)-mice, testis of UPEC-infected Irp1–/– mice showed impaired ERK signaling. Conversely, IRP2 deletion led to a stronger inflammatory response. Notably, differences in immune cell infiltrations were observed among the different genotypes. In contrast to WT and Irp2–/– mice, no increase in monocytes and neutrophils was detected in testis of Irp1–/– mice upon UPEC-infection. Interestingly, in Irp1–/– UPEC-infected testis, we observed an increase in a subpopulation of macrophages (F4/80+ CD206+) associated with anti-inflammatory and wound-healing activities compared to WT. These findings suggest that IRP1 deletion may protect against UPEC-induced inflammation by modulating ERK signaling and dampening the immune response.
Authors
Niraj Ghatpande, Aileen Harrer, Bar Azoulay-Botzer, Noga Guttmann-Raviv, Sudhanshu Bhushan, Andreas Meinhardt, Esther G. Meyron-Holtz
Abstract
Benign prostatic hyperplasia (BPH) is the nodular proliferation of the prostate transition zone in older men, leading to urinary storage and voiding problems that can be recalcitrant to therapy. Decades ago, John McNeal proposed that BPH originates with the “reawakening” of embryonic inductive activity by adult prostate stroma, which spurs new ductal proliferation and branching morphogenesis. Here, by laser microdissection and transcriptional profiling of the BPH stroma adjacent to hyperplastic branching ducts, we identified secreted factors likely mediating stromal induction of prostate glandular epithelium and coinciding processes. The top stromal factors were Insulin Like Growth Factor 1 (IGF1) and C-X-C Motif Chemokine Ligand 13 (CXCL13), which we confirmed by RNA in situ hybridization to be co-expressed in BPH fibroblasts, along with their cognate receptors (IGF1R and CXCR5) on adjacent epithelium. In contrast, IGF1 but not CXCL13 was expressed in human embryonic prostate stroma. Finally, we demonstrated that IGF1 is necessary for the generation of BPH-1 cell spheroids and patient-derived BPH cell organoids in three-dimensional culture. Our findings partially support historic speculations on the etiology of BPH, and provide what we believe to be new molecular targets for rational therapies directed against the underlying processes driving BPH.
Authors
Anna S. Pollack, Christian A. Kunder, Noah Brazer, Zhewei Shen, Sushama Varma, Robert B. West, Gerald R. Cunha, Laurence S. Baskin, James D. Brooks, Jonathan R. Pollack
Abstract
CXCR4 is a key regulator of the development of NK cells and dendritic cells, both of which play an important role in early placental development and immune tolerance at the maternal-fetal interface. However, the role of CXCR4 in pregnancy is not well understood. Our study demonstrates that adult-induced global genetic CXCR4 deletion, but not uterine-specific CXCR4 deletion, was associated with increased pregnancy resorptions and decreased litter size. CXCR4-deficient mice had decreased NK cells and increased granulocytes in the decidua, and increased leukocyte numbers in peripheral blood. We found that CXCR4-deficient mice had abnormal decidual NK cell aggregates and NK cell infiltration into trophoblast areas beyond the giant cell layer. This was associated with low NK cell expression of granzyme B, a NK cell granule effector, indicative of NK cell dysfunction. Pregnancy failure in these mice was associated with abnormalities in placental vascular development and increased placental expression of inflammatory genes. Importantly, adoptive bone marrow transfer of wild type CXCR4+ bone marrow cells into CXCR4-deficient mice rescued the reproductive deficits by normalizing NK cell function and mediating normal placental vascular development. Collectively, our study found an important role for maternal CXCR4 expression in immune cell function, placental development and pregnancy maintenance.
Authors
Fang Lyu, Chase A. Burzynski, Yuan yuan Fang, Aya Tal, Alice Y. Chen, Jacqueline Kisa, Kriti Agrawal, Yuval Kluger, Hugh S. Taylor, Reshef Tal
Abstract
Maternal decidual CD8+ T cells must integrate the antithetical demands of providing immunity to infection while maintaining immune tolerance for fetal and placental antigens. Human decidual CD8+ T cells were shown to be highly differentiated memory T cells with mixed signatures of dysfunction, activation, and effector function. However, no information is present on how specificity for microbial or fetal antigens relates to their function or dysfunction. In addition, a key question, whether decidual CD8+ T cells include unique tissue-resident memory T cells (Trm) or also effector memory T cell (Tem) types shared with peripheral blood populations, is unknown. Here, high-dimensional flow cytometry of decidual and blood CD8+ T cells identified 2 Tem populations shared in blood and decidua and 9 functionally distinct Trm clusters uniquely found in decidua. Interestingly, fetus- and virus-specific decidual CD8+ Trm cells had similar features of inhibition and cytotoxicity, with no significant differences in their expression of activation, inhibitory, and cytotoxic molecules, suggesting that not all fetus-specific CD8+ T cell responses are suppressed at the maternal-fetal interface. Understanding how decidual CD8+ T cell specificity relates to their function and tissue residency is crucial in advancing understanding of their contribution to placental inflammation and control of congenital infections.
Authors
Shweta Mahajan, Aria Alexander, Zachary Koenig, Nicholas Saba, Nina Prasanphanich, David A. Hildeman, Claire A. Chougnet, Emily DeFranco, Sandra Andorf, Tamara Tilburgs
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