Karen Racicot

Inhibitor of DNA Binding 4 Is Expressed Selectively by Single Spermatogonia in the Male Germline and Regulates the Self-Renewal of Spermatogonial Stem Cells in Mice

Continual spermatogenesis at a quantitatively normal level is required to sustain male fertility. The foundation of this process relies on maintenance of an undifferentiated spermatogonial population consisting of spermatogonial stem cells (SSCs) that self-renew as well as transient amplifying progenitors produced by differentiation. In mammals, type Asingle spermatogonia form the SSC population, but molecular markers distinguishing these from differentiating progenitors are undefined and knowledge of mechanisms regulating their functions is limited. We show that in the mouse male germline the transcriptional repressor ID4 is expressed by a subpopulation of undifferentiated spermatogonia and selectively marks Asingle spermatogonia. In addition, we found that ID4 expression is up-regulated in isolated SSC-enriched fractions by stimulation from GDNF, a key growth factor driving self-renewal. In mice lacking ID4 expression, quantitatively normal spermatogenesis was found to be impaired due to progressive loss of the undifferentiated spermatogonial population during adulthood. Moreover, reduction of ID4 expression by small interfering RNA treatment abolished the ability of wild-type SSCs to expand in vitro during long-term culture without affecting their survival. Collectively, these results indicate that ID4 is a distinguishing marker of SSCs in the mammalian germline and plays an important role in the regulation of self-renewal.

Understanding the Complexity of the Immune System during Pregnancy

Progress in our understanding of the role of the maternal immune system during healthy pregnancy will help us better understand the role of the immune system in adverse pregnancy outcomes. In this review, we discuss our present understanding of the ‘immunity of pregnancy’ in the context of the response to cervical and placental infections and how these responses affect both the mother and the fetus. We discuss novel and challenging concepts that help explain the immunological aspects of pregnancy and how the mother and fetus respond to infection.

MicroRNAs 221 and 222 regulate the undifferentiated state in mammalian male germ cells.

Continuity of cycling cell lineages relies on the activities of undifferentiated stem cell-containing subpopulations. Transition to a differentiating state must occur periodically in a fraction of the population to supply mature cells, coincident with maintenance of the undifferentiated state in others to sustain a foundational stem cell pool. At present, molecular mechanisms regulating these activities are poorly defined for most cell lineages. Spermatogenesis is a model process that is supported by an undifferentiated spermatogonial population and transition to a differentiating state involves attained expression of the KIT receptor. We found that impaired function of the X chromosome-clustered microRNAs 221 and 222 (miR-221/222) in mouse undifferentiated spermatogonia induces transition from a KIT– to a KIT+ state and loss of stem cell capacity to regenerate spermatogenesis. Both Kit mRNA and KIT protein abundance are influenced by miR-221/222 function in spermatogonia. Growth factors that promote maintenance of undifferentiated spermatogonia upregulate miR-221/222 expression; whereas exposure to retinoic acid, an inducer of spermatogonial differentiation, downregulates miR-221/222 abundance. Furthermore, undifferentiated spermatogonia overexpressing miR-221/222 are resistant to retinoic acid-induced transition to a KIT+ state and are incapable of differentiation in vivo. These findings indicate that miR-221/222 plays a crucial role in maintaining the undifferentiated state of mammalian spermatogonia through repression of KIT expression.

Sertoli Cells Dictate Spermatogonial Stem Cell Niches in the Mouse Testis

Sustained spermatogenesis in adult males relies on the activity of spermatogonial stem cells (SSCs). In general, tissue-specific stem cell populations such as SSCs are influenced by contributions of support cells that form niche microenvironments. Previous studies have provided indirect evidence that several somatic cell populations and the interstitial vasculature influence SSC functions, but an individual orchestrator of niches has not been described. In this study, functional transplantation of SSCs, in combination with experimental alteration of Sertoli cell content by polythiouracil (PTU)-induced transient hypothyroidism, was used to explore the relationship of Sertoli cells with SSCs in testes of adult mice. Transplantation of SSCs from PTU-treated donor mice into seminiferous tubules of normal recipient mice revealed a greater than 3-fold increase in SSCs compared to those from testes of non-PTU-treated donors. In addition, use of PTU-treated mice as recipients for transplantation of SSCs from normal donors revealed a greater than 3-fold increase of accessible niches compared to those of testes of non-PTU treated recipient mice with normal numbers of Sertoli cells. Importantly, the area of seminiferous tubules bordered by interstitial tissue and percentage of seminiferous tubules associated with blood vessels was found to be no different in testes of PTU-treated mice compared to controls, indicating that neither the vasculature nor interstitial support cell populations influenced the alteration of niche number. Collectively, these results provide direct evidence that Sertoli cells are the key somatic cell population dictating the number of SSCs and niches in mammalian testes.

Viral Infection of the Pregnant Cervix Predisposes to Ascending Bacterial Infection

Preterm birth is the major cause of neonatal mortality and morbidity, and bacterial infections that ascend from the lower female reproductive tract are the most common route of uterine infection leading to preterm birth. The uterus and growing fetus are protected from ascending infection by the cervix, which controls and limits microbial access by the production of mucus, cytokines, and antimicrobial peptides. If this barrier is compromised, bacteria may enter the uterine cavity, leading to preterm birth. Using a mouse model, we demonstrate, to our knowledge for the first time, that viral infection of the cervix during pregnancy reduces the capacity of the female reproductive tract to prevent bacterial infection of the uterus. This is due to differences in susceptibility of the cervix to infection by virus during pregnancy and the associated changes in TLR and antimicrobial peptide expression and function. We suggest that preterm labor is a polymicrobial disease, which requires a multifactorial approach for its prevention and treatment.

Viral infections during pregnancy.

Viral infections during pregnancy have long been considered benign conditions with a few notable exceptions, such as herpes virus. The recent Ebola outbreak and other viral epidemics and pandemics show how pregnant women suffer worse outcomes (such as preterm labor and adverse fetal outcomes) than the general population and non‐pregnant women. New knowledge about the ways the maternal–fetal interface and placenta interact with the maternal immune system may explain these findings. Once thought to be ‘immunosuppressed’, the pregnant woman actually undergoes an immunological transformation, where the immune system is necessary to promote and support the pregnancy and growing fetus. When this protection is breached, as in a viral infection, this security is weakened and infection with other microorganisms can then propagate and lead to outcomes, such as preterm labor. In this manuscript, we review the major viral infections relevant to pregnancy and offer potential mechanisms for the associated adverse pregnancy outcomes.

The role of inflammation for a successful implantation.

Approximately half of all human embryo implantations result in failed pregnancy. Multiple factors may contribute to this failure, including genetic or metabolic abnormalities of the embryo. However, many of these spontaneous early abortion cases are attributed to poor uterine receptivity. Furthermore, although many fertility disorders have been overcome by a variety of assisted reproductive techniques, implantation remains the rate‐limiting step for the success of the in vitro fertilization (IVF) treatments. We, as well as others, have demonstrated that endometrial biopsies performed either during the spontaneous, preceding cycle, or during the IVF cycle itself, significantly improve the rate of implantation, clinical pregnancies, and live births. These observations suggest that mechanical injury of the endometrium may enhance uterine receptivity by provoking the immune system to generate an inflammatory reaction. In strong support of this idea, we recently found that dendritic cells (DCs), an important cellular component of the innate immune system, play a critical role in successful implantation in a mouse model. In this review, we discuss the hypothesis that the injury‐derived inflammation in the biopsy‐treated patients generates a focus for uterine DCs and Mac accumulation that, in turn, enhance the endometrial expression of essential molecules that facilitate the interaction between the embryo and the uterine epithelium.

Type I Interferon Regulates the Placental Inflammatory Response to Bacteria and is Targeted by Virus: Mechanism of Polymicrobial Infection-Induced Preterm Birth

Preterm birth (PTB) affects approximately 12% of pregnancies and at least 50% of cases have no known risk factors. We hypothesize that subclinical viral infections of the placenta are a factor sensitizing women to intrauterine bacterial infection. Specifically, we propose that viral‐induced placental IFN‐β inhibition results in a robust inflammatory response to low concentrations of bacteria.

Trophoblast Induces Monocyte Differentiation Into CD14+/ CD16+ Macrophages

During early pregnancy, macrophages and trophoblast come into close contact during placenta development, and regulated cross talk between these cellular compartments is crucial for maintaining a healthy pregnancy. As trophoblast cells constitutively secrete many chemokines and cytokines, we hypothesize that trophoblast‐secreted factors can differentiate monocytes into a decidual phenotype. In this study, we describe a unique macrophage phenotype, following monocytes’ exposure to trophoblast‐soluble factors.

Human Chorionic Gonadotropin Enhances Trophoblast–Epithelial Interaction in an In Vitro Model of Human Implantation

Embryo implantation, which is an absolute requirement for reproduction, starts with blastocyst apposition to the uterine endometrium, followed by attachment to the endometrial surface epithelium. Recent clinical studies reported an increase in implantation and pregnancy rates among women receiving intrauterine human chorionic gonadotropin (hCG) prior to embryo transfer suggesting that, at least in some cases, female infertility is a result of inadequate secretion of hCG. In this study, we characterized the effect of hCG on trophoblast–epithelial interaction by further developing our recently described in vitro model of implantation. Here, we confirmed hCG increased attachment of trophoblast to epithelial cells, using a single-cell trophoblast–epithelial coculture system in addition to a blastocyst-like spheroid–epithelial coculture system. Furthermore, we discovered that the source and concentration was pivotal; the first preparation of hCG affected 2 molecules related to implantation, MUC16 and osteopontin, while the second preparation required additional cytokines to mimic the effects. Using this system, we can develop a comprehensive knowledge of the cellular and gene targets of hCG and other factors involved in embryo apposition and implantation and potentially increase the number of therapeutic targets for subfertile patients.