Regulation of the innate immune cells during pregnancy: An immune checkpoint perspective.

The foetus can be regarded as a half-allograft implanted into the maternal body. In a successful pregnancy, the mother does not reject the foetus because of the immune tolerance mechanism at the maternal-foetal interface. The innate immune cells are a large part of the decidual leukocytes contributing significantly to a successful pregnancy. Although the contributions have been recognized, their role in human pregnancy has not been completely elucidated. Additionally, the accumulated evidence demonstrates that the immune checkpoint molecules expressed on the immune cells are co-inhibitory receptors regulating their activation and biological function. Therefore, it is critical to understand the immune microenvironment and explore the function of the innate immune cells during pregnancy. This review summarizes the classic immune checkpoints such as PD-1, CTLA-4 and some novel molecules recently identified, including TIM-3, CD200, TIGIT and the Siglecs family on the decidual and peripheral innate immune cells during pregnancy. Furthermore, it emphasizes the role of the immune checkpoint molecules in pregnancy-associated complications and reproductive immunotherapy.

Decreased nitric oxide content mediated by asymmetrical dimethylarginine and protein l-arginine methyltransferase 3 in macrophages induces trophoblast apoptosis: a potential cause of recurrent miscarriage.

STUDY QUESTION: Is the protein l-arginine methyltransferase 3 (PRMT3)/asymmetrical dimethylarginine (ADMA)/nitric oxide (NO) pathway involved in the development of recurrent miscarriage (RM), and what is the potential mechanism? SUMMARY ANSWER: Elevated levels of PRMT3 and ADMA inhibit NO formation in the decidua, thereby impairing the functions of trophoblast cells at the maternal-foetal interface. WHAT IS KNOWN ALREADY: Decreased NO bioavailability is associated with RM. ADMA, an endogenous inhibitor of nitric oxide synthase (NOS), is derived from the methylation of protein arginine residues by PRMTs and serves as a predictor of mortality in critical illness. STUDY DESIGN, SIZE, DURATION: A total of 145 women with RM and 149 healthy women undergoing elective termination of an early normal pregnancy were enrolled. Ninety-six female CBA/J, 24 male DBA/2 and 24 male BALB/c mice were included. CBA/J × DBA/2 matings represent the abortion group, while CBA/J × BALB/c matings represent the normal control group. The CBA/J pregnant mice were then categorised into four groups: (i) normal + vehicle group (n = 28), (ii) abortion + vehicle group (n = 28), (iii) normal + SGC707 (a PRMT3 inhibitor) group (n = 20) and (iv) abortion + SGC707 group (n = 20). All injections were made intraperitoneally on Days 0.5, 3.5 and 6.5 of pregnancy. Decidual tissues were collected on Days 8.5, 9.5 and 10.5 of gestation. The embryo resorption rates were calculated on Day 9.5 and Day 10.5 of gestation. PARTICIPANTS/MATERIALS, SETTING, METHODS: NO concentration, ADMA content, NOS activity, expression levels of NOS and PRMTs in decidual tissues were determined using conventional assay kits or western blotting. PRMT3 expression was further analysed in decidual stromal cells, macrophages and natural killer cells. A co-culture system between decidual macrophages (DMs) and HTR-8/SVneo trophoblasts was constructed to study the roles of the PRMT3/ADMA/NO signalling pathway. Trophoblast apoptosis was analysed via Annexin V-fluorescein isothiocyanate/propidium iodide staining. CBA/J × DBA/2 mouse models were used to investigate the effects of SGC707 on embryo resorption rates. MAIN RESULTS AND THE ROLE OF CHANCE: Our results show that NO concentration and NOS activity were decreased, but ADMA content and PRMT3 expression were increased in the decidua of RM patients. Moreover, compared with the normal control subjects, PRMT3 expression was significantly up-regulated in the macrophages but not in the natural killer cells or stromal cells of the decidua from RM patients. The inhibition of PRMT3 results in a significant decrease in ADMA accumulation and an increase in NO concentration in macrophages. When co-cultured with DMs, which were treated with SGC707 and ADMA, trophoblast apoptosis was suppressed and induced, respectively. In vivo experiments revealed that the administration of SGC707 reduced the embryo resorption rate of CBA/J × DBA/2 mice. LIMITATIONS, REASONS FOR CAUTION: All sets of experiments were not performed with the same samples. The main reason is that each tissue needs to be reserved for clinical diagnosis and only a small piece of each tissue can be cut and collected for this study. WIDER IMPLICATIONS OF THE FINDINGS: Our results indicate that the PRMT3/ADMA/NO pathway is a potential marker and target for the clinical diagnosis and therapy of RM. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Key Research and Development Program of China (2017YFC1001401), National Natural Science Foundation of China (81730039, 82071653, 81671460, 81971384 and 82171657) and Shanghai Municipal Medical and Health Discipline Construction Projects (2017ZZ02015). The authors have declared no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.

Design, synthesis, and biological activity of a novel series of benzofuran derivatives against oestrogen receptor-dependent breast cancer cell lines.

A docking study of a novel series of benzofuran derivatives with ERα was conducted. In this study, we report the synthesis of a novel series of benzofuran derivatives and evaluation of their anticancer activity in vitro against MCF-7 human breast cancer cells, as well as their potential toxicity to ER-independent MDA-MB-231 breast cancer cells, human renal epithelial HEK-293 cells, and human immortal keratinocytes (HaCaT cells) by using the MTT colorimetric assay. The screening results indicated that the target compounds exhibited anti-breast cancer activity. The target compound 2-benzoyl-3-methyl-6-[2-(morpholin-4-yl)ethoxy]benzofuran hydrochloride (4e) exhibited excellent activity against anti-oestrogen receptor-dependent breast cancer cells and low toxicity. The preliminary structure-activity relationships of the target benzofuran derivatives have been summarised. In conclusion, the novel benzofuran scaffold may be a promising lead for the development of potential oestrogen receptor inhibitors.

Phytotoxic, Antifungal and Immunosuppressive Metabolites from Aspergillus terreus QT122 Isolated from the Gut of Dragonfly.

Insect gut microbes have been considered as a resource for bioactive metabolites. The aim of this study was to characterize the compounds of a fungus Aspergillus terreus QT122 associated with the gut of dragonfly. Five main phytotoxic, antifungal, and immunosuppressive substances were isolated from the fungus QT122. The structures of such compounds were identified as emodin (1), 1-methyl emodin (2), terrein (3), methyl 6-acetyl-4-methoxy-7,8-dihydroxynaphthalene-2-carboxylate (4), and dihydrogeodin (5) on the basis of spectroscopic analysis and by comparison of the corresponding data to those reported in the literature previously. The compound 3 exhibited the best phytotoxic activity against the radicle growth of A. retroflexus L. and E. crusgalli L. with their IC50 values of 11.2 and 3.1 µg/mL, which were comparable to that of the positive control of 2,4-dichlorophenoxyacetic acid (2,4-D) with the IC50 values of 8.1 and 1.6 µg/mL, respectively. The compounds 2-3 showed potent antifungal activity in the growth of Alternaria solani with the IC50 value of less than 0.1 µg/mL and the compound 2 also had great inhibitory effect against the growth of Fusarium oxysporum f. sp. cucumerinum (IC50 < 0.1 µg/mL), which was comparable to that of referenced cycloheximide with IC50 value of below 0.1 µg/mL. The compounds 3-5 exhibited strong immunosuppressive activities against the T cell viability with the inhibition rates of more than 99%, which were comparable to positive cyclosporin A under the concentration of 20 µM. These results suggest that the compounds 2-5 have the potential to be used as bio-control agents in agriculture or immunosuppressive agents.

Trade-offs and evolution of thermal adaptation in the Irish potato famine pathogen Phytophthora infestans.

Temperature is one of the most important environmental parameters with crucial impacts on nearly all biological processes. Due to anthropogenic activity, average air temperatures are expected to increase by a few degrees in coming decades, accompanied by an increased occurrence of extreme temperature events. Such global trends are likely to have various major impacts on human society through their influence on natural ecosystems, food production and biotic interactions, including diseases. In this study, we used a combination of statistical genetics, experimental evolution and common garden experiments to investigate the evolutionary potential for thermal adaptation in the potato late blight pathogen, Phytophthora infestans, and infer its likely response to changing temperatures. We found a trade-off associated with thermal adaptation to heterogeneous environments in P. infestans, with the degree of the trade-off peaking approximately at the pathogen's optimum growth temperature. A genetic trade-off in thermal adaptation was also evidenced by the negative association between a strain's growth rate and its thermal range for growth, and warm climates selecting for a low pathogen growth rate. We also found a mirror effect of phenotypic plasticity and genetic adaptation on growth rate. At below the optimum, phenotypic plasticity enhances pathogen's growth rate but nature selects for slower growing genotypes when temperature increases. At above the optimum, phenotypic plasticity reduces pathogen's growth rate but natural selection favours for faster growing genotypes when temperature increases further. We conclude from these findings that the growth rate of P. infestans will only be marginally affected by global warming.

Macrophages promote the growth and invasion of endometrial stromal cells by downregulating IL-24 in endometriosis.

Macrophages play an important role in the origin and development of endometriosis. Estrogen promoted the growth of decidual stromal cells (DSCs) by downregulating the level of interleukin (IL)-24. The aim of this study was to clarify the role and mechanism of IL-24 and its receptors in the regulation of biological functions of endometrial stromal cells (ESCs) during endometriosis. The level of IL-24 and its receptors in endometrium was measured by immunohistochemistry. In vitro analysis was used to measure the level of IL-24 and receptors and the biological behaviors of ESCs. Here, we found that the expression of IL-24 and its receptors (IL-20R1 and IL-20R2) in control endometrium was significantly higher than that in eutopic and ectopic endometrium of women with endometriosis. Recombinant human IL-24 (rhIL-24) significantly inhibited the viability of ESCs in a dosage-dependent manner. Conversely, blocking IL-24 with anti-IL-24 neutralizing antibody promoted ESCs viability. In addition, rhIL-24 could downregulate the invasiveness of ESCs in vitro After co-culture, macrophages markedly reduced the expression of IL-24 and IL-20R1 in ESCs, but not IL-22R1. Moreover, macrophages significantly restricted the inhibitory effect of IL-24 on the viability, invasion, the proliferation relative gene Ki-67, proliferating cell nuclear antigen (PCNA) and cyclooxygenase2 (COX-2), and the stimulatory effect on the tumor metastasis suppressor gene CD82 in ESCs. These results indicate that the abnormally low level of IL-24 in ESCs possibly induced by macrophages may lead to the enhancement of ESCs' proliferation and invasiveness and contribute to the development of endometriosis.

Embryonic trophoblasts induce decidual regulatory T cell differentiation and maternal-fetal tolerance through thymic stromal lymphopoietin instructing dendritic cells.

Physiological pregnancy requires the maternal immune system to recognize and tolerate embryonic Ags. Although multiple mechanisms have been proposed, it is not yet clear how the fetus evades the maternal immune system. In this article, we demonstrate that trophoblast-derived thymic stromal lymphopoietin (TSLP) instructs decidual CD11c(+) dendritic cells (dDCs)with increased costimulatory molecules; MHC class II; and Th2/3-type, but not Th1-type, cytokines. TSLP-activated dDCs induce proliferation and differentiation of decidual CD4(+)CD25(-) T cells into CD4(+)CD25(+)FOXP3(+) regulatory T cells (Tregs) through TGF-ß1. TSLP-activated dDC-induced Tregs display immunosuppressive features and express Th2-type cytokines. In addition, decidual CD4(+)CD25(+)FOXP3(+) Tregs promote invasiveness and HLA-G expression of trophoblasts, resulting in preferential production of Th2 cytokines and reduced cytotoxicity in decidual CD56(bright)CD16(-) NK cells. Of interest, decreased TSLP expression and reduced numbers of Tregs were observed at the maternal-fetal interface during miscarriage. Our study identifies a novel feedback loop between embryo-derived trophoblasts and maternal decidual leukocytes, which induces a tolerogenic immune response to ensure a successful pregnancy.

Chemokine CCL24 promotes the growth and invasiveness of trophoblasts through ERK1/2 and PI3K signaling pathways in human early pregnancy.

Chemokine CCL24, acting through receptor CCR3, is a potent chemoattractant for eosinophil in allergic diseases and parasitic infections. We recently reported that CCL24 and CCR3 are co-expressed by trophoblasts in human early pregnant uterus. Here we prove with evidence that steroid hormones estradiol (E), progesterone (P), and human chorionic gonadotropin (hCG), as well as decidual stromal cells (DSCs) could regulate the expression of CCL24 and CCR3 of trophoblasts. We further investigate how trophoblast-derived CCL24 mediates the function of trophoblasts in vitro, and conclude that CCL24/CCR3 promotes the proliferation, viability and invasiveness of trophoblasts. In addition, analysis of the downstream signaling pathways of CCL24/CCR3 show that extracellular signal-regulated kinases (ERK1/2) and phosphoinositide 3-kinase (PI3K) pathways may contribute to the proliferation, viability and invasiveness of trophoblasts by activating intracellular molecules Ki67 and matrix metallopeptidase 9 (MMP9). However, we did not observe any inhibitory effect on trophoblasts when blocking c-Jun N-terminal kinase (JNK) or p38 pathways. In conclusion, our data suggests that trophoblast-derived CCL24 at the maternal-fetal interface promotes trophoblasts cell growth and invasiveness by ERK1/2 and PI3K pathways. Meanwhile, pregnancy-related hormones (P and hCG), as well as DSCs could up-regulate CCL24/CCR3 expression in trophoblasts, which may indirectly influence the biological functions of trophoblasts. Thus, our results provide a possible explanation for the growth and invasion of trophoblasts in human embryo implantation.

IL-33 enhances proliferation and invasiveness of decidual stromal cells by up-regulation of CCL2/CCR2 via NF-κB and ERK1/2 signaling.

Interleukin (IL)-33, a newly described member of the IL-1 family, has been reported to facilitate primary tumor progression and metastatic dissemination. However, its biological function on decidual stromal cells (DSCs) remains unclear. In this study, we tested the hypothesis whether IL-33 promotes proliferation and invasion of DSCs, and the possible mechanism. IL-33 and its orphan receptor ST2 was found to be co-expressed by DSCs in human first-trimester pregnancy. Addition of IL-33, enhanced the proliferation and invasion of DSCs in a dosage-dependent manner, concomitantly with increasing expression of proliferation relative gene (PCNA, survivin) and invasion relative gene (titin, MMP2). Blocking IL-33/ST2 signaling by soluble sST2 apparently abolished the stimulatory effect on the proliferation, invasiveness and related gene expression in DSCs. We also demonstrated that chemokines CCL2/CCR2 was significantly increased with IL-33 administration. Moreover, inhibition of CCL2/CCR2 activation using CCL2 neutralizing antibody or CCR2 blocker prevented IL-33-stimulated proliferation and invasiveness capacity of DSCs. Increasing phosphorylation of nuclear factor NF-κB p65 and extracellular signal-regulated kinases ERK1/2 after treatment with IL-33 was confirmed by western blotting. And the IL-33-induced CCL2/CCR2 expression was abrogated by treatment with the NF-κB inhibitor BAY 11-7082 or ERK1/2 inhibitor U0126. Finally, we showed that decreased IL-33/ST2 expression was observed in DSCs from spontaneous abortion compared with normal pregnancy at both gene and protein levels. This study provides evidence for the molecular mechanism of IL-33 in promoting proliferation and invasiveness of DSCs by up-regulation of CCL2/CCR2 via NF-κB and ERK1/2 signal pathways and thus contributes insight to the potential of IL-33 involved in successful pregnancy via inducing DSCs mitosis and invasion.

Thymic stromal lymphopoietin downregulates NME1 expression and promotes invasion in human trophoblasts via the activation of STAT3 signaling pathway.

Our previous study has demonstrated that TSLP induces the invasion of human trophoblasts and plays a role in embryo implantation. The present study is undertaken to explore the mechanism by which TSLP modulates trophoblast invasion. It was revealed that TSLP treatment significantly downregulated NME1 and TIMP1 expression in both human trophoblasts and JEG-3 cells. The stimulatory effect of TSLP on trophoblast invasion was partially inhibited by either STAT3 inhibitor or STAT5 inhibitor. The downregulation of NME1 expression by TSLP was completely abrogated by STAT3 inhibitor. TIMP1 expression was attenuated by NME1 siRNA interference and enhanced by NME1 overexpression in JEG-3 cells. TSLP-stimulated increase in trophoblast invasion was partially attenuated by NME1 overexpression. Our data suggest that TSLP might downregulate NME1 expression via STAT3 signaling pathway, affecting TIMP1 expression in influencing trophoblast invasion. Our studies suggest that further studies on TSLP as a potential therapeutic for recurrent spontaneous abortion are warranted.

Inhibition of IDO1 suppresses cyclooxygenase-2 and matrix metalloproteinase-9 expression and decreases proliferation, adhesion and invasion of endometrial stromal cells.

Indoleamine 2,3-dioxygenase-1 (IDO1) is an intracellular enzyme that catalyses essential amino acid tryptophan along the kynurenine pathway. The aim of this study was to determine the impact of IDO1 expression on the biological characteristic of the endometrial stromal cells (ESCs). IDO1, cyclooxygenase-2 (COX-2) and matrix metalloproteinases (MMPs) in endometriotic ectopic stromal cells, endometriosis-derived eutopic stromal cells and normal ESCs (control) were detected by the in-cell Western analysis. After being treated with lipopolysaccharide, levo-1-methyl-tryptophan (L-1-MT) alone or a combination, a comparative analysis of the above protein expression was evaluated. The effects of IDO1 on ESCs proliferation, adhesion and invasion were detected through ELISA, adhesion assay and Matrigel invasion assay, respectively. The results showed that, contrary to healthy ESCs from control women, the expression of IDO1 was significantly higher in eutopic and ectopic ESCs obtained from women with endometriosis. Inhibition of IDO1 by L-1-MT suppressed the expression of COX-2 and MMP-9 in ESCs. It could also decrease the ESCs proliferation, adhesion and invasion, while stimulating ESCs decidualization. Thus, IDO1 is possibly involved in endometriosis pathogenesis via promoting COX-2 and MMP-9 expression and regulation of ESCs biological characteristics. The information may be useful for developing a new therapeutic strategy for endometriosis.

Thymic stromal lymphopoietin from trophoblasts induces dendritic cell-mediated regulatory TH2 bias in the decidua during early gestation in humans.

Thymic stromal lymphopoietins (TSLPs) play critical roles in dendritic cell-mediated immune responses. In this study, we found that human trophoblasts and decidual epithelial cells in maternal-fetal interface of early placentas express TSLP mRNA and protein, but only trophoblast cells secret soluble TSLP. Human decidual CD1c(+) DCs (dDCs) highly express the functional TSLP receptor complex TSLP receptor and interleukin-7 receptor-α. Recombinant human TSLP activates CD1C(+) decidual DCs and peripheral monocyte-derived DCs with increased costimulatory molecules, major histocompatibility complex class II, and OX-40L. Human TSLP or supernatants from human trophoblasts specifically stimulate dDCs to highly produce interleukin-10 and T(H)2-attracting chemokine CCL-17. The TSLP-activated dDCs prime decidual CD4(+) T cells for T(H)2 cell differentiation, involved in maternal-fetal immunotolerance. Interestingly, the protein expression of TSLP in normal pregnancy with significant T(H)2 bias is much higher than that of miscarriage showing T(H)1 bias at the maternal-fetal interface. Therefore, human trophoblasts may contribute to maternal-fetal tolerance by instructing dDCs to induce regulatory T(H)2 bias in human early pregnancy via TSLP.

CXCL8 enhances proliferation and growth and reduces apoptosis in endometrial stromal cells in an autocrine manner via a CXCR1-triggered PTEN/AKT signal pathway.

BACKGROUND: Chemokine CXCL8 (also known as IL-8) has been identified as a potential regulator of endometrial stromal cells (ESCs), but it is unclear how CXCL8 regulates the survival of ESCs in the pathogenesis of endometriosis. METHODS: We assessed the secretion of CXCL8 by enzyme-linked immunosorbent assays and the expression of its receptors, CXCR1 and CXCR2, by in-cell Western assay and immunohistochemistry. The effects of CXCL8 on the activation or expression of various cell mediators were also investigated by in-cell Western assay. The effects of CXCL8 on the proliferation, growth and apoptosis of ESCs in vitro were assessed by BrdU assays, cell counts and annexin V labeling, respectively. RESULTS: Secretion of CXCL8 and expression of CXCR1 in the eutopic ESCs from women with endometriosis were significantly higher than that in control ESCs, but the expression of CXCR2 showed no significant difference between these two cell types. CXCL8 stimulated proliferation and growth and reduced apoptosis of ESCs in an autocrine manner, and these effects were abolished by anti-human CXCL8 and CXCR1 neutralizing antibodies and by a PI3K/Akt inhibitor. Moreover, CXCL8 up-regulated the expression of the anti-apoptotic proteins, survivin and Bcl-2, inhibited the expression of the Phosphatase and tensin homolog (PTEN) and activated the phosphorylation of Akt. CONCLUSIONS: This study suggests that CXCL8 and CXCR1 are involved in the pathogenesis of endometriosis by up-regulating proliferation and growth and restricting apoptosis in ESCs by activating the PTEN/Akt pathway and mediating the expression of survivin and Bcl-2.

Fructose-1,6-bisphosphate prevents pregnancy loss by inducing decidual COX-2+ macrophage differentiation.

Decidualization is an intricate biological process in which extensive remodeling of the endometrium occurs to support the development of an implanting blastocyst. However, the immunometabolic mechanisms underlying this process are still largely unknown. We found that the decidualization process is accompanied by the accumulation of fructose-1,6-bisphosphate (FBP). The combination of FBP with pyruvate kinase M stimulated IL-27 secretion by endometrial stromal cells in an ERK/c-FOS-dependent manner. IL-27 induced decidual COX-2+ M2-like macrophage differentiation, which promotes decidualization, trophoblast invasion, and maternal-fetal tolerance. Transfer of Ptgs2+/COX-2+ macrophages prevented fetal loss in Il27ra-deleted pregnant mice. FBP levels were low in plasma and decidual tissues of patients with unexplained recurrent spontaneous abortion. In therapeutic studies, FBP supplementation significantly improved embryo loss by up-regulation of IL-27-induced COX-2+ macrophage differentiation in a mouse model of spontaneous abortion. These findings collectively provide a scientific basis for a potential therapeutic strategy to prevent pregnancy loss.

The decidual gamma-delta T cells up-regulate the biological functions of trophoblasts via IL-10 secretion in early human pregnancy.

The functions of decidual γδT cells in early human pregnancy are not fully understood. The present study was undertaken to characterize the action of decidual γδT cells, and analyze their regulatory roles in proliferation and invasion of trophoblasts in early human pregnancy. It was revealed that decidual γδT cells were a CD4(-) and CD8(-) subset whose numbers increased significantly in either the decidua or peripheral blood. The main cytokines synthesized in decidual γδT cells in decreasing concentration were; IL-10>TGF-ß>TNF-α≥IFN-γ, with little IL-2 or IL-4 being produced. Decidual γδT cells promoted trophoblast proliferation and invasion, and suppressed trophoblast apoptosis through IL-10 secretion in co-culture. These results suggest that γδT cells in human decidua might play an important role in the Th2 bias at maternal-fetal interface and in the development and progression of the placenta, which is beneficial to maternal immunotolerance toward the fetus in early human pregnancy.

Upregulation of miR-29a suppressed the migration and invasion of trophoblasts by directly targeting LOXL2 in preeclampsia.

OBJECTIVE: Preeclampsia is a pregnancy-specific disorder that is a major cause of maternal and foetal morbidity and mortality, with a prevalence of 6-8% of pregnancies. Although the downregulation of lysyl oxidase (LOX) and LOX-like protein 2 (LOXL2), which leads to reduced trophoblast cell migration and invasion through activation of the TGF-ß1/Smad3/collagen pathway, is relevant to preeclampsia, the mechanisms regulating differences in the gene expression of LOX and LOXL2 in placentas are not yet understood. This study aimed to investigate the mechanisms regulating differences in the gene expression of LOX and LOXL2 in placentas. METHODS: The expression of miRNAs, LOX and LOXL2 in preeclamptic placentas and control placentas was analysed by qPCR. Localisation of miR29a and LOXL2 in preeclamptic placentas was performed by RNA-Fluorescence in-situ hybridization assay. The direct regulation of LOXL2 by miR-29a was assessed by dual-luciferase reporter assays in human extravillous trophoblast cells (HTR8/SVneo). Cell migration and invasion were evaluated by Transwell assays in HTR8/SVneo cells. RESULTS: miR-29a expression was upregulated in preeclamptic placentas and negatively correlated with LOXL2 mRNA expression levels. RNA-Fluorescence in-situ hybridization assay revealed a clear overlap between miR-29a and LOXL2 in the placentas of preeclampic women. LOXL2 was a direct target gene of miR-29a, as confirmed by a dual-luciferase reporter assay in HTR8/SVneo trophoblast cells. miR-29a suppressed HTR8/SVneo trophoblast cell migration and invasion. LOXL2 overexpression reversed the inhibitory effects of miR-29a on HTR8/SVneo trophoblast cell migration and invasion. CONCLUSION: Our results suggest that the upregulation of miR-29a suppresses the migration and invasion of HTR8/SVneo trophoblast cells by directly targeting LOXL2 in preeclampsia.

Antibacterial Potential of Termite-Associated Streptomyces spp.

Twenty-one strains of termite-associated actinomycetes were tested for their activities against three bacteria. The results showed that nine strains showed bacteriostatic activities against at least one tested bacterium, and the actinomycete YH01, which was isolated from the body surface of the queen of Odontotermes formosanus, had potent antibacterial activity. The YH01 was further identified as Streptomyces davaonensis. Two metabolites roseoflavin (1) and 8-methylamino-8-demethyl-d-riboflavin (2) were isolated and purified from S. davaonensis YH01. Their structures were determined by NMR, MS, and the related literature. The metabolite 1 showed strong inhibition activities against Bacillus subtilis (MIC = 1.56 µg/mL) and Staphylococcus aureus (MIC = 3.125 µg/mL), which were comparable to referenced gentamycin sulfate, with MIC values of 1.56 and 1.56 µg/mL, respectively. Furthermore, the anti-MRSA potential of compound 1 was determined against nine kinds of MRSA strains, with inhibition zones in the ranges of 12.7-19.7 mm under a concentration of 15 µg/6 mm discs and 18.3-22.7 mm under a concentration of 30 µg/6 mm discs. However, metabolite 1 had no inhibitory effect on Gram-negative bacteria. These results suggested that roseoflavin produced by YH01 holds promise for use against Gram-positive bacteria, especially to MRSA.

Low chorionic villous succinate accumulation associates with recurrent spontaneous abortion risk.

Dysregulated extravillous trophoblast invasion and proliferation are known to increase the risk of recurrent spontaneous abortion (RSA); however, the underlying mechanism remains unclear. Herein, in our retrospective observational case-control study we show that villous samples from RSA patients, compared to healthy controls, display reduced succinate dehydrogenase complex iron sulfur subunit (SDHB) DNA methylation, elevated SDHB expression, and reduced succinate levels, indicating that low succinate levels correlate with RSA. Moreover, we find high succinate levels in early pregnant women are correlated with successful embryo implantation. SDHB promoter methylation recruited MBD1 and excluded c-Fos, inactivating SDHB expression and causing intracellular succinate accumulation which mimicked hypoxia in extravillous trophoblasts cell lines JEG3 and HTR8 via the PHD2-VHL-HIF-1α pathway; however, low succinate levels reversed this effect and increased the risk of abortion in mouse model. This study reveals that abnormal metabolite levels inhibit extravillous trophoblast function and highlights an approach for RSA intervention.

Functional regulation of thymic stromal lymphopoietin on proliferation and invasion of trophoblasts in human first-trimester pregnancy.

BACKGROUND: Thymic stromal lymphopoietin (TSLP) is a novel cytokine that triggers the dendritic cell-mediated T(H)2 response and regulatory T cell expansion. The aim of this study is to evaluate whether TSLP and TSLP receptor (TSLPR) are expressed in primary human extravillous trophoblast (EVT), how proimflammatory cytokines (tumor necrosis factor (TNF)-alpha, IL-1beta), T(H)2 and T(H)3 cytokines (IL-4, TGF-beta) and pregnancy-associated hormones regulate TSLP production by EVT and whether the SLP-TSLPR interaction affects the biological behavior of trophoblsts. METHODS: We assessed TSLP mRNA and protein expression by real-time RT-PCR, ELISA and immunochemistry, respectively. We further investigated effects of TSLP on the proliferation and invasion of trophoblast cells in vitro. RESULTS: The primary EVTs constitutively expressed TSLP and TSLPR. IL-4 and TNF-alpha or pregnancy-associated hormones result in a significant increase in TSLP mRNA expression and protein release from EVT, and TSLP promotes primary EVT proliferation and invasion in vitro. CONCLUSIONS: This study has demonstrated that the first-trimester human trophoblast cells express TSLP and TSLPR, that cytokine milieu which mimics the maternal-fetal interface modulates expression of TSLP in trophoblast and that TSLP stimulates trophoblast proliferation and invasion. This suggests that TSLP plays an important role in human EVT invasion and placentation in human early pregnancy.

Molecular characteristics and possible functions of innate lymphoid cells in the uterus and gut.

With the discovery of innate lymphoid cells (ILCs), which are especially enriched in barrier surfaces, the family of innate lymphocytes has grown. A unique characterization of these cells can provide a phenotypical definition of ILCs and their specific functions in different tissue environments. Although ILCs are part of the innate immune system, they are derived from lymphoid lineages lacking rearranged antigen-specific and pattern-recognition receptors. The International Union of Immunological Societies (IUIS) favors the notion that ILCs can be generally divided into five main groups, namely, NK cells, ILC1s, ILC2s, ILC3s and LTi cells. These cells can be specifically stimulated by environmental and pathogen-derived signals. Upon stimulation, ILCs can rapidly secrete a wide range of soluble cytokines that can modulate the functions of effector cells. Over the last decade, ILCs, especially helper ILCs, which do not include NK cells, have been recognized to be a crucial cell type involved in integrating diverse host immune responses. Recently, emerging research has shown that helper ILCs also play a critical role in promoting tissue restoration and immune responses at barrier surfaces. Notably, helper ILCs act as a double-edged sword, being involved in the inflammatory and reparative responses during homeostasis and disease. Therefore, in this review, we summarize the current findings regarding the molecular characteristics and tissue-specific effector functions of helper ILCs in the uterus during physiological and pathological pregnancy and in the intestine during homeostasis and inflammation.