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Uterine leiomyosarcoma (LMS) is a rare but deadly disease. Due to poor understanding of the molecular and genetic causes of the disease, the diagnosis of LMS has been based primarily on histology. Nuclear atypia is one of hallmarks in LMS, however, it also occurs in 2 clinically benign variants, including smooth muscle tumors with fumarate hydratase alteration (SMT‐FH) and leiomyoma with bizarre nuclei (LM‐BN). In addition to nuclear atypia, many well recognized biomarkers used for LMS are also frequently overexpressed in LM‐BN, and the histogenesis and molecular natures for LM‐BN and LMS remain largely unknown. To characterize the molecular profiling of LMS, SMT‐FH, and LM‐BN, we performed integrated comprehensive genomic profiling including whole‐genome sequencing (WGS) and RNA sequencing and genomic microarray analyses to assess genome‐wide copy number alterations (CNAs) and immunohistochemistry (IHC) in all 3 tumor types. We found that both LM‐BN and LMS showed genomic instability and harbored extensive CNAs throughout the whole genome. By contrast, the SMT‐FH presented its characteristic 1q43‐44 deletions in all cases tested, with minimal CNAs in the rest of genomic regions. Further analyses revealed that LMS and LM‐BN groups showed similar patterns of CNAs that are tended to cluster together and separated from the SMT‐FH group. The integrated molecular profiling enabled the detection of novel and traditional biomarkers and showed excellent discrimination between LM‐BN and LMS. Our study suggests that LM‐BN, despite having similar nuclear atypia to SMT‐FH, showed similar genomic instability but distinct genomic alterations with its malignant counterpart of LMS. The integrated molecular profiling is of clinical importance in characterizing these rare uterine smooth muscle tumors., Leiomyosarcoma and 2 benign mimics (leiomyoma with FH alteration [SMT‐FH] and leiomyoma with bizarre nuclei [LM‐BN]) have characteristic nuclear atypia and often cause diagnostic challenge. LM‐BN and LMS are highly genomically unstable and harbor extensive CNAs in the genome, whereas SMT‐FH shows simple 1q43‐44 deletions. Integrated molecular profiling enabled the detection of novel and traditional biomarkers and showed excellent discrimination between LM‐BN and LMS.

Does the aggregation of M2 macrophages affect the expression of gene associated with retinoid-interferon-induced mortality 19 (GRIM-19) in adenomyosis?Endometrial tissues were collected from patients with (n = 15) and without (n = 15) adenomyosis. Tissues were analysed for GRIM-19 and toll-like receptor 4 (TLR4) expression by immunohistochemistry and western blotting. Apoptosis was analysed by TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling (TUNEL) assay. Human endometrial stromal cells (HESC) were transfected with GRIM-19 small interfering RNA (SiRNA) to knockdown GRIM-19 expression. The HESC were co-cultured with M2 macrophages to detect the influence of M2 macrophages in HESC cells. Analyses included GRIM-19, caspase-3 and TLR4 expression by western blotting, and GRIM-19 and TLR4 by quantitative real-time polymerase chain reaction. Apoptosis was measured by flow cytometry and TUNEL assay. Cell proliferation (Cell Counting Kit-8 assay) and migration assays were carried out.The expression of GRIM-19 was significantly lower in adenomyosis lesions compared with controls (P0.001). Deficiency of GRIM-19 induced by siRNA decreased apoptosis and increased proliferation and migration in HESC. A significant decrease in GRIM-19 expression occurred in HESC after co-culture with M2 macrophages (P = 0.018). After co-culture with M2 macrophage, apoptosis decreased and proliferation and cell invasion in HESC increased. Protein (P = 0.006) and mRNA (P = 0.013) expression of TLR4 in HESC also reduced after this co-culture. Up-regulation of GRIM-19 occurred in HESC treated with the activator TLR4 (P = 0.016). Up-regulation of GRIM-19 was significantly reversed in cells treated with the TLR4 inhibitor (P = 0.011).M2 macrophages may be involved in regulating the expression of GRIM-19 partly through the TLR4 signalling axis in adenomyosis.

Background The aim of this study was to evaluate the effectiveness and safety of different treatment strategies for endogenic caesarean scar pregnancy (CSP) patients. Methods According to Vial’s standard, we defined endogenic-type CSP as (1) the gestational sac growing towards the uterine cavity and (2) a greater than 0.3 cm thickness of myometrial tissue at the caesarean scar. A total of 447 endogenic CSP patients out of 527 patients from 4 medical centres in China were enrolled in this study. A total of 120 patients were treated with methotrexate (MTX) followed by surgery, 106 received ultrasound-guided curettage directly and 221 received curettage combined with hysteroscopy. The clinical information and clinical outcomes of these patients were reviewed. Successful treatment was defined as (1) no additional treatment needed, (2) no retained mass of conception and (3) serum β subunit of human chorionic gonadotropin (β-hCG) level returning to a normal level within 4 weeks. The success rate was analysed based on these factors. Result Among 447 patients, no significant difference was observed in baseline characteristics between groups except for foetal heartbeat. The success rate was significantly different (p Conclusion The combination of curettage and hysteroscopy represents the most effective strategy. Pretreatment with MTX did not result in better clinical outcomes. Ultrasound-guided curettage directly should not be considered a first-line treatment choice for endogenic CSP patients.

Activin A promotes human trophoblast invasion during the first trimester of pregnancy and is associated with preeclampsia and pregnancy-induced hypertension (PE/PIH) in naturally conceived pregnancies. However, whether integrin β1 mediates activin A-increased trophoblast invasion remains unknown and the evidence is limited regarding the predictive value of activin A for PE/PIH in women receiving in vitro fertilization (IVF) treatment. Here, we studied the role and underlying molecular mechanisms of integrin β1 in activin A-promoted invasion in immortalized (HTR8/SVneo) and primary human extravillous trophoblast (EVT) cells. A nest case-control study was designed to investigate the predictive/diagnostic value of activin A in IVF pregnancies. Results showed that integrin β1 expression increased after activin A treatment and knockdown of integrin β1 significantly decreased both basal and activin A-increased HTR8/SVneo cell invasion. SB431542 (TGF-β type I receptors inhibitor) abolished activin A-induced SMAD2/SMAD3 phosphorylation and integrin β1 overexpression. Activin A-upregulated integrin β1 expression was attenuated after the depletion of ALK4 or SMAD4 in both HTR8/SVneo and primary EVT cells. Furthermore, we found similar first-trimester activin A levels in IVF patients with or without subsequent PE/PIH. These results reveal that integrin β1 mediates activin A-promoted trophoblast invasion through ALK4-activated SMAD2/3-SMAD4 pathway, and the predictive/diagnostic value of first-trimester maternal serum activin A for hypertensive disorders of pregnancy might be different in IVF population.

As a commonly used phthalate compound, di(n-butyl) phthalate (DBP) is an emerging group of polyvinyl chloride plasticizers. The acute toxicity of DBP has been extensively studied using the aquatic indicator organism, Daphnia magna. However, little is known about chronic and transgenerational toxicity of DBP. In this study, acute LC50 values were 3.04 mg/L (24 h) and 2.55 mg/L (48 h). Chronic toxicity tests in the case of maternal exposure to DBP revealed that DBP had negligible effects on growth and reproduction of F3 generation of D. magna, although the growth rate of body length and the intrinsic rate of increase were prominently reduced, to a pretty small extent. At specific concentrations, DBP generated beneficial effects on the parental generation of D. magna and no obvious impacts on the F1 generation. This study showed that maternal exposure to DBP did not cause any transgenerational effects on D. magna.

AIM To explore whether neutrophil-to-lymphocyte ratio (NLR) and monocyte-to-lymphocyte ratio (MLR) could predict missed abortion (MA) at 7-13 week's gestation. METHODS A total of 363 women with a diagnosis of MA and 232 women with normal pregnancy at 7-13 week's gestation, who visited our hospital from June 2012 to May 2018 were retrospectively analyzed. At 7 week's gestation, total and differential leukocyte counts, NLR and MLR were compared between women with MA (n = 69) and normal controls (n = 53). The receiver operating characteristic curve was used to select the optimal indicator and its cut-off point. The role of the optimal indicator was further assessed at 8-13 week's gestation. RESULTS The mean white blood cell counts, the mean neutrophil counts, the median monocyte counts, the mean NLR and the median MLR in women with MA were significantly lower than those in normal controls (P

The aim of the study was to compare the efficacy of laparoscopy and hysteroscopy for the treatment of cesarean scar pregnancy (CSP) and analyze the clinical factors associated with successful selection for hysteroscopic or laparoscopic treatment of CSP. We retrospectively studied 112 cases of CSP that were treated by laparoscopy and/or hysteroscopy in our hospital from December 2014 to December 2017. In total, 72 of these patients underwent ultrasound-guided curettage and hysteroscopic resection without uterine scar defect repair. Fourty of these patients underwent laparoscopic resection and repair of the uterine scar defect. We analyzed the different clinical variables between the 2 groups and identified the clinical factors which could predict the need for the laparoscopic repair of uterine scar defect. Results showed that laparoscopy and hysteroscopy were safe ways to treat CSP, and no patient underwent hysterectomy. The β-hCG level in both of the 2 groups decreased to normal 4 to 8 weeks after surgery. There were significant differences between the hysteroscopy group and laparoscopy uterine scar repair group in terms of days of amenorrhea, gestational sac diameter, myometrial thickness, operation time, intraoperative blood loss, and hospitalization duration (P

Summary Hydropic leiomyoma (HLM) is a variant of uterine leiomyoma with characteristic features of zonal distributions of edema, increased vascularity, and tumor cells arranged in nodules or cords. Diagnostic difficulty and patient management are further complicated by a lack of studies and unknown cause of the disease. To study this tumor's nature, 24 HLM cases were selected for analysis of cytohistologic features, immunohistochemical profile (HMGA2, FH, CD34, pAKT, p16, ER, SMA, and Ki-67), and molecular alterations of HMGA2 by fluorescence in situ hybridization and MED12 mutations. HLM showed large tumor size (average 14.4 cm) and unique histology, characterized by edematous areas of tumor cells with mostly round-oval nuclei, arranged in cords and/or with perinodular growth around vessels, and increased thick-walled vessels (average 17 vessels/10× medium-power field). Immunohistochemistry revealed that 76% (18/24) of HLMs had HMGA2 overexpression, 32% (6/19) of which harbored HMGA2 rearrangement detected by fluorescence in situ hybridization. Thick-walled vessels in HLM were composed of mostly HMGA2-positive tumor cells, and HLM with HMGA2 overexpression also showed CD34-positive tumor vessel–supporting pericytes. In contrast to usual-type leiomyoma with a high frequency of MED12 mutations, no MED12 mutations were found in any HLM. HLM showed increased pAKT activity, indicating a strong contribution of AKT pathway signaling in HLM promoting tumor growth. Our findings suggest that HLM is a distinct variant of uterine smooth muscle tumor likely driven by HMGA2 overexpression.

To study the effect of indoleamine 2,3-dioxygenase (IDO) expressed in HTR-8/SVneo cells on NKG2D and NKp46 expression and cytotoxicity of decidual NK (dNK) and peripheral NK (pNK) cells.CD56(+) dNK and pNK cells purified were cultured with HTR-8/SVneo cell conditioned medium (CM), 1-MT+HTR-8/SVneo cell CM, and complete RPMI 1640 medium (negative control) in vitro. The mRNA and protein expression of NKG2D and NKp46 in NK cells were then assessed by qRT-PCR and flow cytometry, respectively. Their cytotoxicity was evaluated with LDH assays, and TNF-α secretion was analyzed by ELISA.For dNK cells, the mRNA and protein expression of NKp46 as well as NKG2D did not differ significantly among the three groups (P0.05), whereas for pNK cells, the expression level was significantly decreased in HTR-8/SVneo cell CM group than the other two groups (P0.01). Peripheral NK cells cultured with HTR-8/SVneo cell CM showed reduced cytotoxicity and TNF-α secretion than the other two groups (P0.01), although there were no significant differences among three groups for dNK cells (P0.05).IDO expressed by HTR-8/SVneo cells can down-regulate NKp46 and NKG2D expression and reduce cytotoxicity in pNK cells, and may contribute to keep dNK cytotoxicity at a low level, suggesting an important role for IDO in the maintenance of normal pregnancy.

Uterine leiomyomas (ULM) are histologically and molecularly heterogeneous and clinically they grow at vastly different rates. Several driver gene mutations have been identified in ULM, including MED12 mutations, HMGA2 overexpression, and biallelic FH inactivation. ULM with different driver mutant genes may use different molecular pathways, but currently no clear correlation between gene mutations and growth related pathways has been established. To better define this relationship, we collected ULM with MED12 (n = 25), HMGA2 (n = 15), and FH (n = 27) mutations and examined the sex steroid hormone, cell cycle, and AKT pathway genes by immunohistochemistry. While ER and PR were highly expressed in all types of ULM, FH ULM showed lower ER expression and higher PR expression. HMGA2 tumors had significantly higher levels of AKT signaling and mitogenic activity than other ULM types. HMGA2 activated AKT signaling through upregulation of IGF2BP2. Silencing HMGA2 in ULM cells resulted in downregulation of AKT and upregulation of p16 and p21, which eventually led to cell senescence. HMGA2 overexpression in ULM is not only related to tumor development but also plays a role in controlling cellular proliferation through the AKT pathway.

To evaluate uterine artery chemoembolization (UAEC) followed by vacuum aspiration as a conservative treatment for complicated cesarean scar pregnancy.A retrospective review was performed of women presenting with cesarean scar pregnancy between January 2002 and December 2008. The medical record was evaluated to determine the method of treatment.During the time period studied, 13 women were identified who underwent UAEC followed by vacuum aspiration. 12 women successfully had bilateral UAEC followed by vacuum aspiration alone, one woman had unilateral UAEC followed by vacuum aspiration but subsequently required laparotomy. All 13 women were successfully cured and retained uterus, there was no case with severe complicating disease. With the follow-up period, two women who were planning future pregnancy conceived, and spontaneous abortion occurred in one of them during the first trimester, another had an elective cesarean delivery at term.UAEC combined with vacuum aspiration is technically feasible and may help avoid laparotomy in women with cesarean scar pregnancy.

Summary Dendritic cells (DCs) can acquire unique features or phenotypes in different tissue microenvironments and decide whether immunity or tolerance develops. DCs observed within the decidua have been implicated in pregnancy maintenance. However, the precise distribution of decidual DC subsets and their phenotypic characteristics are largely unknown. Using flow cytometry, we identified three DC subsets in normal human first-trimester decidua: BDCA-1+ CD19- CD14- myeloid DC type 1 (MDC1), BDCA-3+ CD14- myeloid DC type 2 (MDC2) and BDCA-2+ CD123+ plasmacytoid DC (PDC). The percentage of MDC1 to mononuclear cells in the decidua was similar to that in the peripheral blood controls. The percentage of MDC2 in the decidua was significantly higher than that in the peripheral blood controls, whereas the percentage of PDC was significantly lower. Both MDC1 and MDC2 subsets expressed human leucocyte antigen D-related, CD86 and CD80 at low levels, suggesting a characteristic of immature myeloid DCs. Immunoglobulin-like transcript 3, suggested to be involved in immune tolerance induction, was also expressed on decidual MDC1 and MDC2 subsets. In addition, as gestational age increased from 6 to 9 weeks, the numbers of MDC1 decreased but MDC2 increased significantly. This is the first study to demonstrate the presence of three previously unidentified BDCA-1+, BDCA-3+ and BDCA-2+ DC subsets in human decidua, these decidual DCs might play important role in the maintenance of pregnancy.

NK cells accumulate at the maternal-fetal interface (MFI) and play essential roles in maintaining immune tolerance during pregnancy. The mechanisms that facilitate NK cells tolerance to fetal tissue are largely unknown. T cell Ig and mucin domain-containing protein 3 (Tim-3) is a newly defined molecule with essential immunological function in many physiological and pathological processes. Recent study showed that Tim-3 was involved in the regulation of immune tolerance at MFI. However, whether Tim-3 regulates NK cells cytotoxicity toward trophoblasts is unclear. Here, we showed Tim-3 was mainly expressed by decidual NK cells (dNK) and Tim-3 level in dNK was higher than peripheral NK cells (pNK). Tim-3(+) dNK expressed more levels of mature markers CD94 and CD69 than Tim-3- dNK cells and blocking Tim-3 significantly inhibited dNK IFN-γ and TNF-α secretion. Furthermore, we found TGF-β1 may contribute to such up-regulation of Tim-3 in NK cells. Interestingly, blocking Tim-3 enhanced NK cytotoxicity toward trophoblast cell line HTR-8 but not K562. We found HTR-8 expressed Tim-3 ligand Galectin-9, in contrast K562 did not. Small interfering RNA-mediated silencing of Galectin-9 expression enhanced NK cytotoxicity toward HTR-8. We further showed Tim-3/Galecin-9 inhibited NK cytotoxicity toward trophoblast partially via impairing the degranulation process. In addition, clinical data showed that abnormal Tim-3 level on pNK might be associated with recurrent spontaneous abortion (RSA). Thus, our data demonstrate Tim-3/Galectin-9 pathway maintains local tolerance by suppressing NK cytotoxicity toward trophoblasts which may represent a new immunologic tolerance mechanism at MFI.

Objectives To localize indoleamine 2,3-dioxygenase (IDO) mRNA and protein and to undertake a functional study at the first trimester fetal–maternal interface in order to determine whether the distribution and function of IDO are related to recurrent spontaneous abortion (RSA). Methods Women undergoing legal pregnancy termination and women with RSA participated in this prospective study. Immunohistochemistry and real-time reverse transcription–polymerase chain reaction were used to analyse levels of IDO protein and mRNA in placenta, decidua and HTR-8/SVneo cells. Culture medium collected from trophoblast villous explant or HTR-8/SVneo cell cultures was used to measure IDO activity in response to interferon (IFN)-γ treatment. Results A total of 40 healthy women and 26 women with RSA provided samples of placenta and decidua. For normal pregnancies, IDO protein and mRNA was identified in placental trophoblasts, invasive extravillous trophoblasts and decidual glandular epithelium. IFN-γ significantly increased IDO activity in trophoblast villous explants and HTR-8/SVneo cells. Levels of IDO protein and mRNA in the placenta and decidua from normal pregnancies were significantly higher than in those from RSA. Conclusions Decreased levels of IDO protein and mRNA in the placenta and decidua from RSA suggest an important role for IDO in the maintenance of normal pregnancy.