Telomeres are repeated DNA sequences whose main function is to preserve genome stability, protecting chromosomes ends from shortening caused by progressive loss during each cell replication or DNA damage. Telomere length regulation is normally achieved by telomerase enzyme, whose activity is progressively shut off during embryonic differentiation in somatic tissues, whereas it is maintained in germ cells, activated lymphocytes, and certain types of stem cell populations. The maintenance of telomerase activity for a longer time is necessary for germ cells to delay telomere erosion, thus avoiding chromosome segregation defects that could contribute to aneuploid or unbalanced gametes. Over the last few years, telomere biology has become an important topic in the field of human reproduction, encouraging several studies to focus on the relation between telomere length and spermatogenesis and male fertility, embryo development and quality during assisted reproductive treatment, and female pathologies as polycystic ovary, premature ovarian insufficiency, and endometriosis. This review analyzes whether telomere length in germ cells is related to reproduction fitness, whether telomere length is related to pathologies associated with male and female fertility, and whether measurement of telomere length could represent a biomarker of germ cell and embryo quality. Telomere length could be considered a molecular marker of spermatogenesis and sperm quality and is somewhat related to male fertility potential. Fewer evidence, although promising, is available for oocytes, female (in)fertility, and embryo quality. The increasing evidence for a role of telomeres and telomere length in human reproduction, indeed, has expanded the historical view of considering them just a marker of aging. Telomere length might have in the future a prognostic potential in couple infertility, especially useful to select best germ cells with the greatest potential of fertilization.

Summary Sentence

Telomere length could be considered a molecular marker of spermatogenesis and sperm quality and is somewhat related tomale fertility potential. Fewer evidence, although promising, is available for oocytes, female (in)fertility, and embryo quality.

Sperm telomere length (STL) is a promising new parameter for sperm quality analysis that may elucidate the molecular mechanisms underlying the idiopathic cases of male factor infertility, which represent almost half of all the male factor infertility cases worldwide. Telomeres consist of nucleoprotein structures present at the ends of eukaryotic chromosomes, whose protective functions maintain the genomic stability. Their role in reproduction includes an active intervention during gametogenesis, fertilization, and preimplantation embryo development. In consonance, studies have shown that compromised telomere homeostasis is associated with male infertility. Since critically short telomeres have their function affected, assessing STL may be a fast and economic method for sperm quality analysis and expectantly contribute to improve the success of fertility treatments. This hypothesis is supported by several reports associating STL with seminal parameters, sperm genome integrity, and clinical outcomes. However, there are other studies in the literature that do not demonstrate these associations. Additionally, it is still not clear whether the lengthening mechanisms of telomeres occurring during early embryo development resume the inherited telomere length. Further research is essential to clarify the suitability of STL as a biomarker for male infertility, before it could be routinely implemented in medically assisted reproduction centers. Understanding the molecular mechanisms underlying STL function and dynamics will provide us new insights into the origins of male infertility and a possible new useful tool as an outcome predictor for assisted reproduction.

Summary Sentence

The importance of telomeres in human reproduction is increasingly evident and sperm telomere length has been suggested as a new biomarker for male infertility, possibly contributing to improve the success of fertility treatments.

In mammals, small non-coding RNAs (sncRNAs) have been reported to be important during early embryo development. However, a comprehensive assessment of the inventory of sncRNAs during the maternal-to-zygotic transition (MZT) has not been performed in an animal model that better represents the sncRNA biogenesis pathway in human oocytes and embryos. The objective of this study was to examine dynamic changes in expression of sncRNAs during the MZT in bovine embryos produced by in vitro fertilization (IVF), which occurs at the 8-cell stage. An unbiased, discovery-based approach was employed using small RNAseq to profile sncRNAs in bovine oocytes, 8-cell stage embryos and blastocyst stage embryos followed by network and ontology analyses to explore the functional relevance of differentially expressed micro-RNAS (miRNAs). The relative abundance of miRNAs was markedly higher in 8-cell stage embryos compared to oocytes or blastocyst stage embryos. This shift in miRNA population was largely associated with upregulation of miRNAs predicted to target genes involved in the biological processes of cell development, cell division, Wnt signaling, and pluripotency, among others. Distinct populations of piwi-interacting-like RNAs (pilRNAs) were identified in bovine oocytes and blastocyst stage embryos, though pilRNAs were nearly absent in 8-cell stage embryos. Also, small nucleolar RNAs were highly expressed in 8-cell stage embryos. Overall, these data reveal a strong dynamic shift in the relative abundance of sncRNAs associated with the MZT in bovine oocytes and embryos, suggesting that these molecules may play important roles in the shift from maternal to zygotic control of gene expression.

Summary Sentence

In bovine oocytes and IVF embryos, the maternal-to-zygotic transition is associated with a marked shift in abundance of specific sncRNA classes, including miRNAs, pilRNAs, and snoRNAs.

To investigate possible involvement of glucocorticoid receptor (GR) in mediating effects of maternal stress or therapeutically administered glucocorticoids on early embryo, we analyzed the expression of GR subtypes in ovulated mouse oocytes and preimplantation embryos. RT-PCR analysis results showed that GRα and GRγ transcripts are relatively highly expressed in mouse oocytes, and both transcripts are present at lower amounts in preimplantation embryos. We also detected low expression of two other splice variants, GRβ and a transcript orthologous to the human GR-P subtype, mainly at the blastocyst stage. Using western blot analysis, we detected several GR protein bands that differed in size between oocytes and preimplantation embryos. To compare the effects of corticosterone (a major endogenous glucocorticoid in rodents) and dexamethasone (a synthetic glucocorticoid) on early embryos, we cultured mouse preimplantation embryos in the presence of these glucocorticoids. Corticosterone showed a strong inhibitory effect on embryo development (starting from a 50 µM concentration), without a significant influence on apoptosis incidence. On the other hand, dexamethasone induced apoptosis in early embryo cells (starting from a 1.5 µM concentration), and its effect on embryo development was less detrimental than that found with the same dose of corticosterone. In summary, our results showed that different GR subtypes are expressed in ovulated mouse oocytes and preimplantation embryos and that the composition of GR subtypes changes during early embryo development. Moreover, we found significant differences in the effects of the two glucocorticoids on early embryo development, which might be associated with activation of different GR subtypes.

Summary Sentence

Different glucocorticoid receptor subtypes are expressed in ovulated mouse oocytes and preimplantation embryos, and two glucocorticoid molecule types differ in their effects on preimplantation embryo development.

This study investigated bovine conceptus-induced modifications to the endometrial transcriptome related to effects of interferon tau (IFNT), conceptus origin (in vivo vs. in vitro), and conceptus sex. In vitro (IVF) or in vivo (superovulation and artificial insemination, AI) produced blastocysts were transferred into recipient heifers on day 7 of the estrous cycle. On day 15, IVF- or AI-derived conceptuses were obtained by uterine flushing and individually placed on endometrial explants in media for 6 h. Explants were also cultured with media alone as a control or media containing 100 ng/mL IFNT. Total explant RNA was analyzed by RNA-Seq. Incubation of endometrium with IFNT or IVF- or AI-derived conceptuses changed (P ≤ 0.001) expression of 491, 498, and 576 transcripts, respectively, compared to the control. Further, 369 differentially expressed genes (DEGs) were common between explants exposed to IFNT or a conceptus. A total of 240 DEGs were uniquely altered by conceptuses (IVF- and AI-derived) but not IFNT. Of these transcripts, 46 were shared between the IVF and AI groups, while 61 and 133 were specific to IVF and AI conceptuses, respectively. Five genes [melanophilin (MLPH), prominin-2 (PROM2), myeloid associated differentiation marker (MYADM), vomeronasal 1 receptor 4 like (VN1R4L) and 5-hydroxytryptamine receptor 1A (HTR1A)] were more abundant in endometrium exposed to female compared to male conceptuses (P < 0.001). A single gene [ADP-ribosylation factor like GTPase 4C (ARL4C)] was more abundant in response to male conceptuses (P < 0.001) than female conceptuses. These data support the hypothesis that conceptus regulation of gene expression in the endometrium is complex and involves factors other than IFNT that may have a biological role in pregnancy establishment.

Summary Sentence

Elongating bovine conceptuses modify the adjacent endometrial transcriptome in a manner dependent and independent of interferon tau and related to conceptus origin and sex.

Embryo implantation rates have been found to be enhanced by precedent endometrial injuries, but the underlying mechanism is not fully investigated. Endometrial inflammation occurs both at peri-implantation period and after endometrial injury, in which vascular reaction is a distinctive feature of inflammation. In this study, intentional endometrial injury was done with a 0.7-mmdiameter brush inserted into the left uterine horn of female ICR mice, then turned around 720° (group 2), and the right uterine horn served as the controls without endometrial injuries (group 1). Intraperitoneal equine chorionic gonadotropin 2.5 IU was injected, followed by human chorionic gonadotropin 10 IU injection, and the uterus was dissected 5 days later, roughly at the periimplantation period. The peri-implantation endometrium was obtained, and angiogenesis protein array revealed that matrix metalloproteinase-3 (MMP-3), plasminogen activator inhibitor-1 (PAI-1), insulin-like growth factor binding protein 1 (IGFBP-1), and IL-1α were more strongly expressed in injured endometrium (group 2) than in the controls (group 1). Immunohistochemical CD34 staining was more prominently expressed in group 2 uterus, and the treatment with LY294002, a phosphoinositide 3-kinase (PI3K) inhibitor, significantly decreased CD34 immunopositive cells. The capabilities of permeability, proliferation, tube formation, and migration of mouse endometrial endothelial cells were significantly enhanced in group 2 than in group 1. Our results demonstrate that enhanced endometrial angiogenesis is a possible mechanism accounting for the increased endometrial receptivity after endometrial injury.

Summary Sentence

Increased endometrial receptivity after intentional injury is probably due to enhanced endometrial angiogenesis

Cell fusion is involved in the development of some adult organs, is implicated in the pathogenesis of specific types of cancer, and is known to participate in repair/regeneration processes mediated by bone-marrow-derived cells (BMDCs). Endometriosis is a disease characterized by growth of functional endometrial tissue outside of the uterine cavity. Endometriosis shares some molecular properties with cancer and BMDCs home to endometriosis lesions in a mouse model. Our objective was to determine if cell fusion can occur in endometriosis and establish whether bone-marrow-derived cells participate in cell fusion events in lesions. We employed a Cre-Lox system to identify cell fusion events in a mouse model of endometriosis. Fused cells were detected in endometriotic lesions, albeit at a low frequency (∼⃒1 in 400 cells), localized to the stromal compartment, and displayed restricted proliferation. Using 5-fluorouracil-based nongonadotoxic bone marrow transplantation model, we demonstrate that bone marrow cells represent a principal cell source for fusion events in lesions. Cell fusion progeny uniformly lacked expression of selected markers of hematopoietic, endothelial, and epithelial markers, though they expressed the mesenchymal/stromal markers Sca-1 and CD29. This study is the first to describe the phenomenon of cell fusion in endometriosis and points to a mesenchymal population derived from cell fusion events with limited proliferative activity, properties previously attributed to endometrial stem cells. Their putative role in the pathogenesis of the disease remains to be elucidated.

Summary Sentence

Cells of the endometriotic lesion fuse with cells of the host animal; bone-marrow-derived cells recruited to the endometriosis lesion contribute to this process.

Endometriosis causes severe chronic pelvic pain and infertility. We have recently reported that niclosamide treatment reduces growth and progression of endometriosis-like lesions and inflammatory signaling (NFKB and STAT3) in a mouse model. In the present study, we examined further inhibitory mechanisms by which niclosamide affects endometriotic lesions using an endometriotic epithelial cell line, 12Z, and macrophages differentiated from a monocytic THP-1 cell line. Niclosamide dose dependently reduced 12Z viability, reduced STAT3 and NFKB activity, and increased both cleaved caspase-3 and cleaved PARP. To model the inflammatory microenvironment in endometriotic lesions, we exposed 12Z cells to macrophage conditioned media (CM). Macrophages were differentiated from THP-1 cells using 12-O-tetradecanoylphorbol-13-acetate as M0, and then M0 macrophages were polarized into M1 or M2 using LPS/IFNγ or IL4/IL13, respectively. Conditioned media from M0, M1, or M2 cultures increased 12Z viability. This effect was blocked by niclosamide, and cell viability returned to that of CM from cells treated with niclosamide alone. To assess proteins targeted by niclosamide in 12Z cells, CM from 12Z cells cultured with M0, M1, or M2 with/without niclosamide were analyzed by cytokine/chemokine protein array kits. Conditioned media from M0, M1, and/or M2 stimulated the secretion of cytokines/chemokines from 12Z cells. Production of most of these secreted cytokines/chemokines in 12Z cells was inhibited by niclosamide. Knockdown of each gene in 12Z cells using siRNA resulted in reduced cell viability. These results indicate that niclosamide can inhibit the inflammatory factors in endometriotic epithelial cells stimulated by macrophages by targeting STAT3 and/or NFKB signaling.

Summary Sentence

Niclosamide can inhibit the inflammatory microenvironment established between endometriotic epithelial cells and macrophages by targeting STAT3 and NFκB signaling.

Genes involved in sexual reproduction diverge rapidly as a result of reproductive fitness. Here, we identify a novel protein domain in the germline-specific Polycomb protein SCML2 that is required for the establishment of unique gene expression programs after the mitosis-to-meiosis transition in spermatogenesis. We term this novel domain, which is comprised of rapidly evolved, DNAbinding repeat units of 28 amino acids, the SCML2 DNA-binding (SDB) repeats. These repeats are acquired in a specific subgroup of the rodent lineage, having been subjected to positive selection in the course of evolution. Mouse SCML2 has two DNA-binding domains: one is the SDB repeats and the other is an RNA-binding region, which is conserved in human SCML2. For the recruitment of SCML2 to target loci, the SDB repeats cooperate with the other functional domains of SCML2 to bind chromatin. The cooperative action of these domains enables SCML2 to sense DNA hypomethylation in an in vivo chromatin environment, thereby enabling SCML2 to bind to hypomethylated chromatin. We propose that the rapid evolution of SCML2 is due to reproductive adaptation, which has promoted species-specific gene expression programs in spermatogenesis.

Summary Sentence

Here, we identify a novel protein domain in the germline-specific Polycomb protein SCML2; because the domain is comprised of rapidly evolved DNA-binding repeats, it has been termed the SCML2 DNA-binding (SDB) repeats.

ATP supply is essential for sperm performance and increases in ATP content coevolve with enhanced sperm swimming velocity as a response to sperm competition in rodents. ATP content is the balance between production and consumption but, although ATP production has received much attention, little is known about ATP consumption. The rate of ATP consumption is crucial for the propagation of the flagellar wave, becoming a main determinant of the time and distance sperm could move before exhausting their reserves. A high yield in distance per unit of ATP consumed (efficiency) could provide advantages in sperm competition. We characterized sperm ATP consumption rate in a group of mouse species with different sperm competition levels to understand its impact on swimming velocity, duration, and yield of sperm ATP reserves. Interspecific comparisons revealed that sperm of species with higher sperm competition levels had high ATP consumption rates and faster swimming velocity. Moreover, sperm that consumed ATP at a faster rate swam more efficiently, since they were able to cover more distance per unit of ATP consumed. Our results suggest that by coupling the advantages of higher ATP turnover rates to increased efficiency of ATP expenditure, sperm would respond to increasingly competitive environments while maintaining a positive ATP balance.

Summary Sentence

Sperm of mouse species with higher sperm competition level swim faster because (a) they consume more ATP, and (b) swim greater distance per unit of ATP consumed.

The self-renewal, proliferation, and differentiation of the spermatogonial populations must be finely coordinated in the mammalian testis, as dysregulation of these processes can lead to subfertility, infertility, or the formation of tumors. There are wide gaps in our understanding of how these spermatogonial populations are formed and maintained, and our laboratory has focused on identifying the molecular and cellular pathways that direct their development. Others and we have shown, using a combination of pharmacologic inhibitors and genetic models, that activation of mTOR complex 1 (mTORC1) is important for spermatogonial differentiation in vivo. Here, we extend those studies to directly test the germ cell-autonomous requirement for mTORC1 in spermatogonial differentiation. We created germ cell conditional knockout mice for “regulatory associated protein of MTOR, complex 1″ (Rptor), which encodes an essential component of mTORC1. While germ cell KO mice were viable and healthy, they had smaller testes than littermate controls, and no sperm were present in their cauda epididymides. We found that an initial cohort of Rptor KO spermatogonia proliferated, differentiated, and entered meiosis (which they were unable to complete). However, no self-renewing spermatogonia were formed, and thus the entire germline was lost by adulthood, resulting in Sertoli cell-only testes. These results reveal the cell autonomous requirement for RPTOR in the formation or maintenance of the foundational self-renewing spermatogonial stem cell pool in the mouse testis and underscore complex roles for mTORC1 and its constituent proteins in male germ cell development.

Summary Sentence

SSC maintenance requires RPTOR

Mammalian sperm undergo a series of biochemical and physiological changes collectively known as capacitation in order to acquire the ability to fertilize. Although the increase in phosphorylation associated with mouse sperm capacitation is well established, the identity of the proteins involved in this signaling cascade remains largely unknown. Tandem mass spectrometry (MS/MS) has been used to identify the exact sites of phosphorylation and to compare the relative extent of phosphorylation at these sites. In the present work, we find that a novel site of phosphorylation on a peptide derived from the radial spoke protein Rsph6a is more phosphorylated in capacitated mouse sperm. The Rsph6a gene has six exons, five of which are conserved during evolution in flagellated cells. The exon containing the capacitation-induced phosphorylation site was found exclusively in eutherian mammals. Transcript analyses revealed at least two different testis-specific splicing variants for Rsph6a.Rsph6a mRNA expression was restricted to spermatocytes. Using antibodies generated against the Rsph6a N-terminal domain, western blotting and immunofluorescence analyses indicated that the protein remains in mature sperm and localizes to the sperm flagellum. Consistent with its role in the axoneme, solubility analyses revealed that Rsph6 is attached to cytoskeletal structures. Based on previous studies in Chlamydomonas reinhardtii, we predict that Rsph6 participates in the interaction between the central pair of microtubules and the surrounding pairs. The findings that Rsph6a is more phosphorylated during capacitation and is predicted to function in axonemal localization make Rsph6a a candidate protein mediating signaling processes in the sperm flagellum.

Summary Sentence

A sequence corresponding to the N-terminal domain of the radial spoke protein Rsph6 was found phosphorylated in capacitated sperm. Rsph6 expression is a testis-specific gene. Rsph6 protein remains present in mature sperm, and is localized to the sperm flagellum.

The direct role of melatonin in mammary glands of dairy goats has remained obscure. This study aimed to evaluate the expression of melatonin membrane receptors (MT1 and MT2) in the pituitary and mammary glands of dairy goats during lactation, and to investigate the role of melatonin in mammary function. Both MT1 and MT2 were consistently expressed in the pituitary and mammary eight glands throughout the lactation period, and their levels were lower in 9March (group I), June (group III), and September (group V) than in May (group II) and August (group IV). The expression patterns of pituitary and mammary MT1 and MT2 were consistent with those of blood melatonin during lactation. Furthermore, the mammary prolactin (PRL), and pituitary growth hormone (GH) and PRL mRNA expression showed an inverse trend in relation to blood melatonin levels. In mammary tissues, MT1 and MT2 immunoreactivity was predominantly located in the mammary epithelial cells (MECs). In addition, a dose- and time-dependent inhibition on cell viability was observed in cultured MECs. At the dose of 10 and 100 pg/ml, melatonin decreased mammary β-casein and PRL expression. Furthermore, the inhibitory effects of melatonin were blocked by luzindole, a nonselective MT1 and MT2 receptor antagonist. In addition, melatonin promoted MT1 and MT2 expression in cultured MECs. In conclusion, the presence of MT1 and MT2 in the pituitary and mammary glands and the inhibitory effects of melatonin on cell viability, β-casein, and PRL expression in MECs suggest the potential regulation by melatonin in goat mammary function.

Summary Sentence

Melatonin directly decreased cell viability, and β-casein and prolactin expression in cultured mammary epithelial cells of lactating dairy goats.

The role of androgenic steroids on ovarian development has attracted much attention in recent years, but the molecular mechanism is still largely unknown. In this study, using zebrafish as a model, we found that the trace metal zinc mediates the action of androgen on oocyte maturation. The ovarian and serum testosterone is transiently stimulated by LH during oocyte maturation. Testosterone could mimic the action of LH on oocyte maturation, and its action appears to be independent of the classical nuclear androgen receptor. Consistent with a recent finding that a zinc transporter (Zip9) has been suggested as a novel androgen receptor, we found the labile zinc concentration could be induced by testosterone in the ovarian follicular cells, and zinc couldmimic the action of testosterone on oocytematuration and signaling. Moreover, the action of testosterone on oocyte maturation could be abolished by the chelation of zinc. Thus, the evidence supports the notion that zinc could mediate the action of androgen on oocyte maturation in zebrafish. This finding would shed light on understanding the role of androgen in ovary development and the molecular mechanism of oocyte maturation in fish.

Summary Sentence

Zinc could mediate the action of androgen on oocyte maturation in zebrafish.

Insufficient perfusion of the trophoblast by maternal blood is associated with an increased generation of reactive oxygen species and complications of the placenta. In this study, we first examined whether rosiglitazone, an agonist of the peroxisome proliferator-activated receptor-γ (PPARγ ), protects the human trophoblast from oxidative injury by regulating key antioxidant proteins, catalase (CAT) and the superoxide dismutases (SOD1 and SOD2). In first trimester placental explants, localization of CAT was limited to cytotrophoblasts, whereas SOD1 was expressed in both the cytoand syncytiotrophoblasts. In first trimester placental explants, hypoxia decreased the expression of both SOD1 and SOD2, and increased apoptosis. Treatment with rosiglitazone dose-dependently upregulated anti-oxidative CAT and SOD2, and rescued hypoxic injury in first trimester villous explants and JEG-3 cells, strongly suggesting the involvement of the PPARγ in regulating their expressions. Rosiglitazone facilitated transcription activity of PPARγ , and enhanced promotor binding, increased transcriptional activity at the CAT promoter, and elevated protein expression/activity. Treatment of hypoxic JEG-3 cells with rosiglitazone resulted in mitochondrial membrane potential increase and a reduction of caspase 9 and caspase 3 activity which is consistent with improved cell survival. To complement PPARγ activation data, we also utilized the antagonist (SR-202) and siRNA to suppress PPARγ expression and demonstrate the specific role of PPARγ in reducing ROS and oxidative stress. Ex vivo examination of term human placenta revealed lower expression of antioxidant proteins in pathologic compared to healthy placental tissues, which could be rescued by rosiglitazone, indicating that rosiglitazone can improve survival of the trophoblast under pathological conditions. These findings provide evidence that the PPARγ pathway directly influences cellular antioxidants production and the pathophysiology of placental oxidative stress.

Summary Sentence

Rosiglitazone (a PPARγ agonist) improves survival of the trophoblast under pathological conditions by regulation of antioxidants.

Pregnancy is a physiologic state of substantially elevated estrogen biosynthesis that maintains vasodilator production by uterine artery endothelial cells (P-UAECs) and thus uterine perfusion. Estrogen receptors (ER-α and ER-β; ESR1 and ESR2) stimulate nongenomic rapid vasodilatory responses partly through activation of endothelial nitric oxide synthase (eNOS). Rapid estrogenic responses are initiated by the ∼⃒4% ESRs localized to the plasmalemma of endothelial cells. Caveolin-1 (Cav-1) interactions within the caveolae are theorized to influence estrogenic effects mediated by both ESRs. Hypothesis: Both ESR1 and ESR2 display similar spatial partitioning between the plasmalemma and nucleus of UAECs and have similar interactions with Cav-1 at the plasmalemma. Using transmission electron microscopy, we observed numerous caveolae structures in UAECs, while immunogold labeling and subcellular fractionations identified ESR1 and ESR2 in three subcellular locations: membrane, cytosol, and nucleus. Bioinformatics approaches to analyze ESR1 and ESR2 transmembrane domains identified no regions that facilitate ESR interaction with plasmalemma. However, sucrose density centrifugation and Cav-1 immunoisolation columns uniquely demonstrated very high protein–protein association only between ESR1, but not ESR2, with Cav-1. These data demonstrate (1) both ESRs localize to the plasmalemma, cytosol and nucleus; (2) neither ESR1 nor ESR2 contain a classic region that crosses the plasmalemma to facilitate attachment; and (3) ESR1, but not ESR2, can be detected in the caveolar subcellular domain demonstrating ESR1 is the only ESR bound in close proximity to Cav-1 and eNOS within this microdomain. Lack of protein–protein interaction between Cav-1 and ESR2 demonstrates a novel independent association of these proteins at the plasmalemma.

Summary Sentence

Since ESR1 and ESR2 plasma membrane location are respectively affiliated with Cav-1-dependent and Cav-1-indepenent mechanisms, a novel difference of ER regulation in endothelial cells derived from the uterine vasculature was identified.

Aims: Globally, sepsis is a major cause of mortality through the combination of cardiovascular collapse and multiorgan dysfunction. Pregnancy appears to increase the risk of death in sepsis, but the exact reason for the greater severity is unclear. In this study, we used polymicrobial sepsis induced by cecal ligation and puncture (CLP) and high-dose intraperitoneal lipopolysaccharide (LPS; 10 or 40 mg, serotype 0111: B4) to test the hypotheses that pregnant mice are more susceptible to sepsis and that this susceptibility was mediated through an excessive innate response causing a more severe cardiovascular collapse rather than a reduction in microbe killing.

Methods and Results: Initial studies found that mortality rates were greater, and that death occurred sooner in pregnant mice exposed to CLP and LPS. In pregnant and nonpregnant CD1 mice monitored with radiotelemetry probes, cardiovascular collapse occurred sooner in pregnant mice, but once initiated, occurred over a similar timescale. In a separate study, tissue, serum, and peritoneal fluid (for protein, flow cytometry, nitric oxide, and bacterial load studies) were collected. At baseline, there was no apparent Th1/Th2 bias in pregnant mice. Post CLP, the circulating cytokine response was the same, but leukocyte infiltration in the lung was greater in pregnant mice, but only TNFα levels were greater in lung tissue. The bacterial load in blood and peritoneal fluid was similar in both groups.

Conclusion: Sepsis-related mortality was markedly greater in pregnant mice. Cardiovascular collapse and organ dysfunction occurred sooner in pregnancy, but bacterial killing was similar. Circulating and tissue cytokine levels were similar, but immune cell extravasation into other organs was greater in pregnant mice. These data suggest that an excessive innate immune system response as shown by the exaggerated lung infiltration of leukocytes may be responsible for the greater mortality. Approaches that reduce off-site trafficking may improve the prognosis of sepsis in pregnancy.

Summary Sentence

Overall, our data suggests that severe septic shock in pregnancy occurred earlier and resulted in greater mortality. Markedly greater inflammatory cell extravasation in the lungs suggests that this off-target effect may mediate the high rates of mortality.

Endogenous hydrogen sulfide (H₂S) is a potent vasodilator and proangiogenic second messenger synthesized from L-cysteine by cystathionine β-synthase (CBS) and cystathionine γ -lyase (CTH). Estrogens are potent vasodilators that stimulate H₂S biosynthesis in uterine arteries (UA) in vivo; however, the underlying mechanisms are unknown. We hypothesized that estrogens stimulate H₂S biosynthesis in UA endothelial cells (UAEC) via specific estrogen receptor (ER)-dependent mechanisms. In cultured primary UAEC, treatment with estradiol-17β (E₂β) stimulated CBS and CTH mRNAs and proteins in a time- and concentration-dependent fashion. As little as 0.1 nM E₂β was effective in increasing CBS and CTH expressions and these stimulatory effects maximized with 10–100 nM E₂β at 48–72 h. E₂β also activated CBS and CTH promoters in UAEC, leading to CBS and CTH expression. Treatment with E₂β stimulated H₂S production, which was blocked by specific inhibitors of either CBS or CTH and their combination and the ER antagonist ICI 182780. Treatment with either specific agonist of ERα or ERβ stimulated both CBS and CTH mRNA and protein expressions and H₂S production to levels similar to that of E₂β. Specific antagonist of either ERα or ERβ blocked E₂β-stimulated CBS and CTH mRNA and protein expressions and H₂S production. Combinations of either ERα or ERβ agonists or their antagonists had no additive effects. Thus, E₂β stimulates H₂S production by upregulating CBS and CTH mRNA and protein expressions through specific ERα or ERβ-dependent CBS and CTH transcription in UAEC in vitro.

Summary Sentence

Estradiol-17β stimulates uterine artery endothelial cell hydrogen sulfide biosynthesis.

Spermatogonial stem cells (SSCs) provide the foundation of spermatogenesis. However, because of their small number and slow self-renewal, transfection of SSCs has met with limited success. Although several viral vectors can infect SSCs, genome integration and an inability to maintain long-term gene expression have hampered studies on SSCs. Here we report successful SSC infection by Sendai virus (SV), an RNA virus in the Paramyxoviridae. The SV efficiently transduced germline stem (GS) cells, cultured spermatogonia with enriched SSC activity, and maintained gene expression for at least 5 months. It also infected freshly isolated SSCs from adult testes. The transfected GS cells reinitiated spermatogenesis following spermatogonial transplantation into seminiferous tubules of infertile mice, suggesting that SV transfection does not interfere with spermatogenesis progression. On the other hand, microinjection of SV into the seminiferous tubules of immature mice transduced SSCs and Sertoli cells, but did not transduce Leydig or peritubular cells by interstitial virus injection. SV-infected hamster GS cells, and freshly isolated rabbit or monkey SSC-like cells were identified following xenogeneic spermatogonial transplantation, suggesting that SV transduces SSCs from several mammalian species. Thus, SV is a useful vector that can transduce both SSCs and Sertoli cells and overcome problems associated with other viral vectors.

Summary Sentence

Sendai virus can transduce spermatogonia without genome integration.

A stable system for producing sterile domesticated fish is required to prevent genetic contamination to native populations caused by aquaculture escapees. The objective of this study was to develop a system to mass produce stock for aquaculture that is genetically sterile by surrogate broodstock via spermatogonial transplantation (SGTP).We previously discovered that female medaka carrying mutations on the follicle-stimulating hormone receptor (fshr) gene become sterile. In this study, we demonstrated that sterile hybrid recipient females that received spermatogonia isolated from sex-reversed XX males (fshr (–/–)) recovered their fertility and produced only donorderived fshr (–) X eggs. Natural mating between these females and fshr (–/–) sex-reversed XX males successfully produced large numbers of sterile fshr (–/–) female offspring. In conclusion, we established a new strategy for efficient mass production of sterile fish. This system can be applied to any aquaculture species for which SGTP and methods for producing sterile recipients can be established.

Summary Sentence

Novel method for mass producing genetically sterile fish from surrogate broodstock was established via transplantation of spermatogonia isolated from follicle-stimulating hormone receptor mutants

Despite the essential role of the active metabolite of vitamin A, all-trans retinoic acid (atRA) in spermatogenesis, the enzymes, and cellular populations responsible for its synthesis in the postnatal testis remain largely unknown. The aldehyde dehydrogenase 1A (ALDH1A) family of enzymes residing within Sertoli cells is responsible for the synthesis of atRA, driving the first round of spermatogenesis. Those studies also revealed that the atRA required to drive subsequent rounds of spermatogenesis is possibly derived from the ALDH1A enzymes residing within the meiotic and post-meiotic germ cells. Three ALDH1A isozymes (ALDH1A1, ALDH1A2, and ALDH1A3) are present in the testis. Although, ALDH1A1 is expressed in adult Sertoli cells and is suggested to contribute to the atRA required for the pre-meiotic transitions, ALDH1A2 is proposed to be the essential isomer involved in testicular atRA biosynthesis. In this report, we first examine the requirement for ALDH1A2 via the generation and analysis of a conditional Aldh1a2 germ cell knockout and a tamoxifen-induced Aldh1a2 knockout model. We then utilized the pan-ALDH1A inhibitor (WIN 18446) to test the collective contribution of the ALDH1A enzymes to atRA biosynthesis following the first round of spermatogenesis. Collectively, our data provide the first in vivo evidence demonstrating that animals severely deficient in ALDH1A2 postnatally proceed normally through spermatogenesis. Our studies with a pan-ALDH1A inhibitor (WIN 18446) also suggest that an alternative source of atRA biosynthesis independent of the ALDH1A enzymes becomes available to maintain atRA levels for several spermatogenic cycles following an initial atRA injection.

Summary Sentence

Elimination of ALDH1A enzymatic activity following a single pulse of retinoic acid does not immediately ablate spermatogenesis due to the presence of an additional source of atRetinal oxidation.

The expression of many genes during the postmeiotic stages of spermatogenesis is largely regulated by germ cell-specific RNA-binding proteins at the level of posttranscription. One of these RNA-binding proteins, YBX2, participates in mRNA storage and regulation of translation in haploid spermatids. How YBX2-stored mRNAs become translationally competent during spermiogenesis remains unknown. In the present study, we report for the first time that YBX2 interacts with PAIP1, a protein translation enhancer, in vitro and in vivo. In murine testes, PAIP1 is highly expressed and colocalizes with YBX2 in round spermatids. Using sequential RNA immunoprecipitation and sequence analysis, we identified a group of spermiogenic mRNAs indirectly bound by PAIP1 through YBX2. Translation of mRNAs bearing the YBX2 target sequence was significantly blocked by YBX2 protein, but was reinitiated when YBX2 was coexpressed with PAIP1 in vitro. Taken together, these results indicate that PAIP1 regulates the translation of YBX2-masked mRNAs during spermiogenesis, and we propose this YBX2–PAIP1 interaction plays an important role in male germ cell development.

Summary Sentence

PAIP1 interacts with YBX2 in the testis and stimulates translation of spermiogenic mRNAs stored by YBX2.