MANF Promotes Unexplained Recurrent Miscarriages by Interacting with NPM1 and Downregulating Trophoblast Cell Migration and Invasion.

Dysplasia and invasive defects in early trophoblasts contribute to unexplained recurrent miscarriages (URMs). Mesencephalic astrocyte-derived neurotrophic factor (MANF) inhibits migration and invasion in some cancer cells, but its role in pregnancy-related diseases remains unresolved. Here, we found that MANF levels in the peripheral blood and aborted tissue of URM women were higher than in normal controls, irrespective of pregnancy or miscarriage. We confirm the interaction between MANF and nucleophosmin 1 (NPM1) in trophoblasts of URM patients, which increases the ubiquitination degradation of NPM1, leading to upregulation of the p53 signaling pathway and inhibition of cell proliferation, migration, and invasion ability. Using a URM mouse model, we found that MANF downregulation resulted in reduced fetal resorption; however, concomitant NPM1 downregulation led to increased abortion rates. These data indicate that MANF triggers miscarriage via NPM1 downregulation and p53 activation. Thus, MANF downregulation or disruption of the MANF-NPM1 interaction could be targets for URM therapeutics.

HMGB1: a double-edged sword and therapeutic target in the female reproductive system.

HMGB1 that belongs to the High Mobility Group-box superfamily, is a nonhistone chromatin associated transcription factor. It is present in the nucleus of eukaryotes and can be actively secreted or passively released by kinds of cells. HMGB1 is important for maintaining DNA structure by binding to DNA and histones, protecting it from damage. It also regulates the interaction between histones and DNA, affecting chromatin packaging, and can influence gene expression by promoting nucleosome sliding. And as a DAMP, HMGB1 binding to RAGE and TLRs activates NF-κB, which triggers the expression of downstream genes like IL-18, IL-1ß, and TNF-α. HMGB1 is known to be involved in numerous physiological and pathological processes. Recent studies have demonstrated the significance of HMGB1 as DAMPs in the female reproductive system. These findings have shed light on the potential role of HMGB1 in the pathogenesis of diseases in female reproductive system and the possibilities of HMGB1-targeted therapies for treating them. Such therapies can help reduce inflammation and metabolic dysfunction and alleviate the symptoms of reproductive system diseases. Overall, the identification of HMGB1 as a key player in disease of the female reproductive system represents a significant breakthrough in our understanding of these conditions and presents exciting opportunities for the development of novel therapies.

Investigation of bedaquiline resistance and genetic mutations in multi-drug resistant Mycobacterium tuberculosis clinical isolates in Chongqing, China.

BACKGROUND: To investigate the prevalence and molecular characterization of bedaquiline resistance among MDR-TB isolates collected from Chongqing, China. METHODS: A total of 205 MDR-TB isolates were collected from Chongqing Tuberculosis Control Institute between March 2019 and June 2020. The MICs of BDQ were determined by microplate alamarblue assay. All strains were genotyped by melting curve spoligotyping, and were subjected to WGS. RESULTS: Among the 205 MDR isolates, the resistance rate of BDQ was 4.4% (9/205). The 55 (26.8%) were from male patients and 50 (24.4%) were new cases. Furthermore, 81 (39.5%) of these patients exhibited lung cavitation, 13 (6.3%) patients afflicted with diabetes mellitus, and 170 (82.9%) isolates belonged to Beijing family. However, the distribution of BDQ resistant isolates showed no significant difference among these characteristics. Of the 86 OFX resistant isolates, 8 isolates were XDR (9.3%, 8/86). Six BDQ resistant isolates (66.7%, 6/9) and two BDQ susceptible isolates (1.0%, 2/196) carried mutations in Rv0678. A total of 4 mutations types were identified in BDQ resistant isolates, including mutation in A152G (50%, 3/6), T56C (16.7%, 1/6), GA492 insertion (16.7%, 1/6), and A274 insertion (16.7%, 1/6). BDQ showed excellent activity against MDR-TB in Chongqing. CONCLUSIONS: BDQ showed excellent activity against MDR-TB in Chongqing. The resistance rate of BDQ was not related to demographic and clinical characteristics. Mutations in Rv0678 gene were the major mechanism to BDQ resistance, with A152G as the most common mutation type. WGS has a good popularize value and application prospect in the rapid detection of BDQ resistance.

Exposure to acrylamide inhibits testosterone production in mice testes and Leydig cells by activating ERK1/2 phosphorylation.

Acrylamide (ACR) is formed during the cooking of starchy foods at high temperatures. Accumulating evidence has shown that ACR has toxic effects, but the mechanism of its potential reproductive toxicity remains unclear. In this study, we observed that ACR caused weight loss in mice. There was no significant difference in the weight of testis and epididymis between the low/medium-dose ACR group and the control group. And the number of epididymal sperms, testicular Leydig cells, serum testosterone level, testicular steroidogenic genes and enzymes, including cytochrome P450 family 11 subfamily A member 1 (CYP11A1) and cytochrome P450 family 17 subfamily A member 1 (CYP17A1), were decreased in the medium/high-dose ACR group. Additional cell experiments showed that the apoptosis rate and the level of reactive oxygen species (ROS) were increased, and testosterone levels and CYP17A1 protein expression were reduced in Leydig cells with treated ACR. Furthermore, the phosphorylation levels of extracellular signal-regulated kinases (ERK1/2) increased significantly; however, there was no significant difference in the levels of serine-threonine protein kinase (AKT) phosphorylation in the testis of mice and Leydig cells treated with ACR. These results suggest that ACR exposure leads to the damage of testicular structure and function and a decline in testosterone synthesis in Leydig cells and mouse testis, which may be related to the activated phosphorylation of ERK1/2.

Frozen embryo transfer in the menstrual cycle after moderate-severe ovarian hyperstimulation syndrome: a retrospective analysis.

BACKGROUND: Ovarian hyperstimulation syndrome (OHSS) is a rare but serious complication of controlled ovarian stimulation. Frozen-embryo transfer (ET) is prompted to be performed in the next menstrual cycles after cancellation of fresh-ET after occurrence of OHSS. However, effects of frozen-ET in the second menstrual cycle have never been investigated. Therefore, this study aimed to assess this in the menstrual cycle after OHSS. METHODS: The OHSS group included 342 women with moderate-severe OHSS who underwent the first frozen-ET in the second menstrual cycle in the First Affiliated Hospital of Anhui Medical University from June 2018 to September 2019. A total of 342 women without OHSS who received frozen-ET in the second menstrual cycle were selected as control group matched by age, body mass index, fertility history, ovulation induction scheme. Uni- and multi-variable conditional logistic regression was used to estimate the association between moderate-severe OHSS and pregnancy outcomes. RESULTS: There were no significant differences in maternal outcomes (miscarriage, preterm birth and pregnancy complications including gestational diabetes mellitus, pregnancy-induced hypertension, placenta previa, premature rupture of membranes and postpartum hemorrhage) and in neonatal outcome (birth-weight and body length, neonatal congenital diseases and other complications) between the two groups in either uni- or multi-variable models. CONCLUSIONS: Frozen-ET in the menstrual cycle after OHSS has similar maternal and neonatal outcomes as in women without OHSS. This study indicates that frozen-ET could be performed in the second menstrual cycle in women who recovered from moderate-severe OHSS.

Low-dose aspirin protects unexplained recurrent spontaneous abortion via downregulation of HMGB1 inflammation activation.

Background: High mobility group box protein 1 (HMGB1) is considered as a kind of sterile inflammatory mediators, which is an overexpression in patients with unexplained recurrent spontaneous abortion (URSA). Specific targeting effect of aspirin on HMGB1 has been revealed. Our previous studies have explored the application of HMGB1 as a therapeutic target of aspirin in URSA disease of mice model and human, but the dynamic process of aspirin downregulating HMGB1 concentration has not been demonstrated. Methods: From December 2018 to November 2020, women with URSA (n = 91) and control women (n = 90) with no history of recurrent abortion or adverse pregnancy were included in the Reproductive Medicine Center of the First Affiliated Hospital of Anhui Medical University. ELISA was applied to detect the concentrations of HMGB1 and IFN-γ in the peripheral blood. Thirty-one URSA patients were monitored for low-dose aspirin treatment (2 and 4 weeks), the changes of HMGB1 and IFN-γ concentrations in peripheral blood of URSA patients before and after using aspirin were compared, and pregnancy outcomes after aspirin treatment were followed up. Results: The levels of HMGB1 in peripheral blood were significantly higher in URSA patients compared with controls, decreasing trends of HMGB1 and IFN-γ concentrations in plasma of URSA patients were observed after treatment with low-dose aspirin continuously, and the expression of HMGB1 was positively correlated with IFN-γ. There were no birth abnormalities in the babies of the URSA patients treated with aspirin. Conclusions: High levels of HMGB1 may be one of the pathogenesis of URSA. Low-dose aspirin may provide protective effect on the HMGB1-triggered URSA.

The associations of serum metals concentrations with the intermediate and pregnancy outcomes in women undergoing in vitro fertilization (IVF).

BACKGROUND: Toxic and essential trace elements are reported to have impact on female fertility. However, studies on the potential synergistic or antagonistic effects of metal mixtures on IVF outcomes remain limited. OBJECTIVE: To evaluate whether serum concentrations of metals, individually and as mixtures, are associated with pregnancy outcomes in women undergoing IVF. METHODS: In a prospective birth cohort study about IVF from the First Affiliated Hospital of Anhui Medical University (n = 1184), we measured the concentrations of serum metals by ICP-MS according to a previously established method. Oocyte/embryo development indicators and follow-up results were also collected. The individual and joint effects of metals were estimated using logistic regressions and Bayesian kernel machine regressions (BKMR). RESULTS: At embryonic stage, we found negative associations between the serum lead (Pb) (ß = -0.14, 95%CI: -0.32, -0.04) and cadmium (Cd) (ß = -0.24, 95%CI: -0.39, -0.09) concentrations and the high-quality embryos rate; and positive associations between the serum cobalt (Co) (ß = 0.18, 95%CI: 0.05, 0.31) and selenium (Se) (ß = 0.17, 95%CI: 0.06, 0.41) concentrations and the MII rate. Regarding to the pregnancy outcomes, the serum Pb was negatively related with successful implantation (OR=0.85, 95%CI: 0.77, 0.94) and clinical pregnancy (OR=0.95, 95%CI: 0.91, 0.99); and positively associated with spontaneous abortion (OR=1.39, 95% CI: 1.02, 1.91). The BKMR analysis showed linear or parabolic associations between the metal mixtures and pregnancy outcomes, with Pb showing the highest posterior inclusion probabilities. CONCLUSIONS: The toxic (Pb, Cd) and essential (Co, Se) metals could be incorporated as simultaneous predictors of IVF outcomes including potential antagonistic effects, in which Pb exhibits major contributions.

Upregulated HMGB1 levels in maternal-fetal interface of patients with unexplained recurrent spontaneous abortion from different sources.

OBJECTIVE: To investigate the expression and sources of high mobility group box 1 (HMGB1) protein in the maternal-fetal interface of patients with unexplained recurrent spontaneous abortion (URSA), and further to verify the role of HMGB1 in the etiology of URSA. METHODS: 55 women at early pregnancy with URSA and 55 women undergoing selective termination of normal early pregnancy as control were included. The abortion tissues including villi and decidua were collected. The expression of HMGB1, CD45, CK7, and vimentin in abortion tissues was detected, and the localization and sources of HMGB1 were analyzed. RESULTS: Infiltrating immune cells and expression of HMGB1 were significantly increased in villi and decidua in URSA group compared with those in the control group. In the URSA group, HMGB1 was colocalized with the CD45-labeled immune cells, and it was more obvious in decidua than in villi; in addition, HMGB1 was colocalized with the vimentin-labeled decidual stromal cells, but not with the CK7- labeled villous epithelial cells. CONCLUSION: High expression of HMGB1 in the maternal-fetal interface in URSA patients was actively secreted by the infiltrating immune cells, and decidual stromal cells may passively release HMGB1 during necrosis.

Bi-allelic mutations in MCIDAS and CCNO cause human infertility associated with abnormal gamete transport.

Reduced generation of multiple motile cilia (RGMC) and the consequent primary ciliary dyskinesia (PCD) cause infertility due to a substantial reduction in the number of multiciliated cells (MCCs) in the efferent ducts (EDs)/oviducts. MCIDAS acts upstream of CCNO to regulate the biogenesis of basal bodies (BBs); therefore, both genes play a vital role in the multiciliogenesis of the reproductive tract epithelium. In this study, whole-exome sequencing was performed to identify the causative genes in 10 unrelated infertile patients with PCD: seven males and three females. Notably, homozygous frameshift mutations in MCIDAS (c.186dupT, p.Pro63Serfs*22) and CCNO (c.262_263insGGCCC, p.Gln88Argfs*8) were identified in one male and one female participant from two unrelated consanguineous families. Haematoxylin-eosin staining/scanning electron microscopy revealed abnormal MCCs in the mutated EDs/oviducts. Furthermore, transmission electron microscopy revealed significantly reduced BBs. Immunofluorescence staining showed the absence of MCIDAS and CCNO signals in the affected tissues and confirmed that MCIDAS acts upstream of CCNO in the context of multiciliogenesis in the reproductive tract epithelium. In vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) was successful, with a positive pregnancy outcome in both MCIDAS- and CCNO-mutated patients. Our results support the use of IVF/ICSI interventions to treat infertility due to RGMC in couples.

Inhibition of HMGB1 Ameliorates the Maternal-Fetal Interface Destruction in Unexplained Recurrent Spontaneous Abortion by Suppressing Pyroptosis Activation.

Recurrent spontaneous abortion (RSA) is a common complication of pregnancy that affects the physical and mental health of pregnant women, and approximately 50% of the mechanisms are unclear. Our previous studies have found that high mobility group box 1 (HMGB1) molecules are highly expressed at the maternal-fetal interface of unexplained recurrent spontaneous abortion (URSA) patients. The purpose of this study was to further detect the expression of HMGB1 and pyroptosis in decidual tissue of URSA patients, and explore the potential mechanism of the protective role of HMGB1 in URSA patients and mouse model. The decidua tissues of 75 URSA patients and 75 women who actively terminated pregnancy were collected, and URSA mouse models were established and treated with HMGB1 inhibitor-aspirin. The expression of HMGB1, and their receptors (RAGE, TLR2, TLR4), pyroptosis-associated proteins (NLRP-3, caspase-1, GSDMD) and NF-κB was examined at the maternal-fetal interface of human and mouse. Our study found that HMGB1, NLRP-3, Caspase-1, GSDMD, RAGE, TLR2 and TLR4 were highly expressed and NF-κB signaling pathway were activated in the decidua tissue of URSA group. Moreover, immune cell disorder and co-localization of HMGB1 and macrophages were found at the maternal-fetal interface of URSA mice. However, HMGB1, TLR2, TLR4, NF-κB, and pyroptosis-associated proteins can be down-regulated by administering low-dose aspirin. These data may indicate that highly expressed HMGB1 was actively secreted by macrophages and then activated pyroptosis through the TLR2/TLR4-NF-κB pathway to cause aseptic inflammation, leading to the occurrence and development of URSA. Moreover, low-dose aspirin can reduce HMGB1 protein levels of serum and decidual in URSA.

Destruction in maternal-fetal interface of URSA patients via the increase of the HMGB1-RAGE/TLR2/TLR4-NF-κB signaling pathway.

AIMS: HMGB1 has been reported to play a crucial role in the physiological and pathophysiological responses during pregnancy. However, it is still unknown whether excessively expressed HMGB1 at the maternal-fetal interface related to Unexplained Recurrent Spontaneous Abortion (URSA). This study was designed to investigate the local capability of HMGB1 in the pathology of URSA, determined the distributions and characteristics of HMGB1, its receptors (RAGE/TLR2/TLR4) and important signaling molecule NF-κB p65 expression at the maternal-fetal interface，as well as compared the differences of HMGB1 expression between the URSA group, control group and aspirin treatment group. MATERIAL AND METHODS: H&E staining, Western blot analysis, immunofluorescence assay and immunohistochemical staining were applied to determine the effect of HMGB1 and its receptors at the maternal-fetal interface. ELISA was utilized to detect the concentration of HMGB1 in plasma. KEY FINDINGS: Our study demonstrated that the activation of the HMGB1-RAGE/TLR2/TLR4-NF-κB pathway at the maternal-fetal interface may have occurred in the URSA group. HMGB1 concentration in plasma was higher in the URSA group than the control group. Furthermore, the levels of HMGB1 of subjects with URSA could be reduced by administrating low doses of aspirin (ASPL). SIGNIFICANCE: This is the first report indicating the roles of HMGB1 at the maternal-fetal interface of URSA patients and broadening the horizons for clinical treatment of URSA. HMGB1-RAGE/TLR2/TLR4-NF-κB signaling pathway may be activated at the maternal-fetal interface in URSA and account for its pathogenesis. HMGB1 have the potential to be promising biomarkers in prevention and therapy of URSA.