hsa_circ_0005991 promotes epithelial-mesenchymal transition by regulating miR-30b-3p/Cdc42EP1 axis in ovary endometriosis.

Endometriosis is a common gynecological disease with an enigmatic pathogenesis. This work explored the function of hsa_circ_0005991 in ovarian endometriosis. High-throughput RNA-Seq was conducted in five matched ectopic (EC) and eutopic (EU) samples. Further, several types of cell function experiments were conducted. According to bioinformatics analysis, a competing endogenous RNA network was established. It included 5 circRNAs, 13 miRNAs, and 551 mRNAs. The expression levels of hsa_circ_0005991 and Cdc42EP1 were significantly elevated, while miR-30b-3p was reduced in the EC group. Upregulation of hsa_circ_0005991 raised Cdc42EP1 levels, induced EMT, and boosted Ishikawa cell proliferation, migration, and invasion. hsa_circ_0005991 knockdown indicated the opposite effects. When co-transfected with miR-30b-3p mimics or inhibitors, these effects could be reversed, respectively. Western blot assays showed alterations of EMT markers in EC samples. hsa_circ_0005991/miR-30b-3p/Cdc42EP1 axis promotes the EMT process in endometriosis, which may offer a theoretical foundation for the mechanism exploration and therapy of this disease.

Deferasirox protects against hydrogen peroxide-induced cell apoptosis by inhibiting ubiquitination and degradation of p21WAF1/CIP1.

Deferasirox (DFX) is an iron chelator approved for the treatment of iron overload diseases. However, the role of DFX in oxidative stress-induced cell apoptosis and the exact molecular mechanisms underlying these processes remain poorly understood and require further investigation. In this study, we found that DFX rendered resistant to H2O2-induced apoptosis in HEK293T cells, reduced the intracellular levels of the labile iron pool (LIP) and oxidative stress induced by H2O2. Furthermore, DFX inhibited the ubiquitination and degradation of the cyclin-dependent kinase inhibitor p21WAF1/CIP1 (p21) via modulation of the interaction of p21 with SCF-Skp2. DFX also showed the inhibition effect on the activation of c-Jun N-terminal kinase (JNK), pro-caspase-3 and related mitochondrial apoptosis pathway induced by H2O2. These results provide novel insights into the molecular mechanism underpinning iron-mediated oxidative stress and apoptosis, and they may represent a promising target for therapeutic interventions in related pathological conditions.

Deferoxamine attenuates lipopolysaccharide-induced inflammatory responses and protects against endotoxic shock in mice.

To examine the role of the intracellular labile iron pool (LIP) in the induction of inflammatory responses, we investigated the anti-inflammatory effect of the iron chelator deferoxamine (DFO) on lipopolysaccharide (LPS)-induced inflammatory responses in RAW264.7 macrophage cells and endotoxic shock in mice in the present study. Our data showed that DFO significantly decreased LPS-induced LIP and ROS upregulation. We then found that DFO inhibited phosphorylation of MAP kinases such as ERK and p38 and also inhibited the activation of NF-κB induced by LPS. Furthermore, the production of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), nitric oxide (NO) and prostaglandin E2 (PGE2) induced by LPS was inhibited by DFO in RAW264.7 macrophages. Administration of DFO significantly decreased the mortality and improved the survival of septic mice with lethal endotoxemia in LPS-injected mice. These results demonstrate that iron plays a pivotal role in the induction of inflammatory responses and against septic shock. DFO has effective inhibitory effect on the production of inflammatory mediators via suppressing activation of MAPKs and NF-κB signaling pathways; it also has a protective effect on LPS-induced endotoxic shock in mice. Our findings open doors to further studies directed at exploring a new class of drugs against septic shock or other inflammatory diseases by modulating cellular chelatable iron.

Regulation of histone demethylase KDM6B by hypoxia-inducible factor-2α.

Lysine (K)-specific demethylase 6B (KDM6B) is a histone H3K27 demethylase, which specifically catalyzes the demethylation of H3 lysine-27 tri/dimethylation (H3K27me3/2). KDM6B can activate gene transcription by promoting transcriptional elongation which is associated with RNA polymerase II and related elongation factors. So KDM6B is important for the regulation of gene expression. Previous studies have indicated that several histone demethylases such as KDM3A, KDM4B, and KDM4C are regulated by hypoxia-inducible factor (HIF). But, the effect of hypoxia on KDM6B is not fully understood. In this study, we found that the expression levels of KDM6B mRNA and protein are modestly up-regulated under hypoxia (1% O2) or mimic hypoxia (desferrioxamine mesylate or CoCl2 treatment) (P<0.05). The result of RNAi shows that the up-regulation of KDM6B is dependent on HIF-2α, but not on HIF-1α. The result of chromatin immunoprecipitation assay indicates that there is a hypoxia response element in KDM6B promoter (-4041 to -4037). The result of Co-IP assay indicates that KDM6B can form complex with HIF-2α or HIF-1α. The knockdown experiment implies that KDM6B is a potential regulator for HIF-2α target genes. These data demonstrate that KDM6B is a new hypoxia response gene regulated by HIF-2α. Our results also show that KDM6B is a potential co-activator of HIF-α, which is important for the activation of hypoxia response genes.

Human umbilical cord mesenchymal stem cell-derived exosomes inhibit ovarian granulosa cells inflammatory response through inhibition of NF-κB signaling in polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a common endocrine disorder and the leading cause of anovulatory infertility in women of reproductive age. Chronic low-grade inflammation is considered to be the major contributors to ovarian dysfunction. Human umbilical mesenchymal stem cell-derived exosomes (hUC-MSC-exos) have been proven to exhibit highly effective anti-inflammatory ability. However, its ability to protect against inflammatory response in PCOS remains unclear. Therefore, the aim of the present study was to explore the effect and underlying mechanisms of hUC-MSC-exos on inflammatory response of ovarian granulosa cells (GCs) in PCOS. In this study, we demonstrated that hUC-MSC-exos enhanced the expression of anti-inflammatory factor IL-10 and inhibited the expression of pro-inflammatory factors TNF-α and IFN-γ. Moreover, hUC-MSC-exos had the effects of inhibiting apoptosis and promoting progesterone production. Mechanistically, exposure to hUC-MSC-exos inhibited NF-κB signaling through significantly decreased the levels of the phosphorylation IκB and p65 in GCs. Moreover, the nuclear translocation of p65 was inhibited. Taken together, these results indicate that hUC-MSC-exos ameliorates the inflammation of granulosa cells in patients with PCOS by inhibiting NF-κB signaling pathway.

Exosomal miR-143-3p derived from follicular fluid promotes granulosa cell apoptosis by targeting BMPR1A in polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is an endocrine disorder that occurs in women of reproductive age. Anovulation caused by abnormal follicular development is still the main characteristic of PCOS patients with infertile. Granulosa cell (GC) is an important part of the follicular microenvironment, the dysfunction of which can affect follicular development. Increasing evidence indicates that exosomal miRNAs derived from the follicular fluid (FF) of patients play critical roles during PCOS. However, which follicular fluid-derived exosomal miRNAs play a pivotal role in controlling granulosa cell function and consequently follicular development remain largely unknown, as does the underlying mechanism. Herein, we showed that miR-143-3p is highly expressed in the follicular fluid exosomes of patients with PCOS and can be delivered into granulosa cells. Furthermore, functional experiments showed that translocated miR-143-3p promoted granulosa cell apoptosis, which is important in follicle development. Mechanistically, BMPR1A was identified as a direct target of miR-143-3p. Overexpression of BMPR1A reversed the effects of exosomal miR-143-3p on GC apoptosis and proliferation by activating the Smad1/5/8 signaling pathway. These results demonstrate that miR-143-3p-containing exosomes derived from PCOS follicular fluid promoted granulosa cell apoptosis by targeting BMPR1A and blocking the Smad1/5/8 signaling pathway. Our findings provide a novel mechanism underlying the roles of exosomal-miRNAs in the follicular fluid of PCOS patients and facilitate the development of therapeutic strategies for PCOS.

miR­519d­3p released by human blastocysts negatively regulates endometrial epithelial cell adhesion by targeting HIF1α.

Successful embryo implantation requires a competent embryo, a receptive endometrium and synchronized communication between them. The selection of embryos with the highest implantation potential remains a challenge in the field of assisted reproductive technology. Moreover, little is known about the precise molecular mechanisms underlying embryo­endometrium crosstalk. MicroRNAs (miRNAs/miRs) have been detected in the spent embryo culture medium (SCM); however, their functions at the preimplantation stage remain unclear. In the present study, human SCM samples were collected during in vitro fertilization/intracytoplasmic sperm injection­embryo transfer and divided into implanted and not­implanted groups according to the clinical pregnancy outcomes. Total RNA was extracted and six miRNAs (miR­372­3p, miR­373­3p, miR­516b­5p, miR­517a­3p, miR­519d­3p and miR­520a­3p) were selected for reverse transcription­quantitative PCR (RT­qPCR) analysis. The results revealed that miR­372­3p and miR­519d­3p were markedly increased in SCM from blastocysts that failed to implant compared with in blastocysts that implanted. The receiver operating characteristic curve analysis revealed that miR­519d­3p was superior to miR­372­3p in predicting pregnancy outcomes. In vitro miRNA uptake and cell adhesion assays were performed to determine whether miR­519d­3p could be taken up by endometrial epithelial cells and to examine the biological roles of miR­519d­3p after internalization. Potential targets of miR­519d­3p were verified using a dual­luciferase reporter system. The results demonstrated that miR­519d­3p was taken up by human endometrial epithelial cells and that it may inhibit embryo adhesion by targeting HIF1α. Using RT­qPCR, western blot analysis and flow cytometry assay, HIF1α was shown to inhibit the biosynthesis of fucosyltransferase 7 and sialyl­Lewis X (sLex), a cell­surface oligosaccharide that serves an important role in embryonic apposition and adhesion. In addition, a mouse model was established and the results suggested that miR­519d­3p overexpression hampered embryo implantation in vivo. Taken together, miRNAs in SCM may serve as novel biomarkers for embryo quality. Furthermore, miR­519d­3p was shown to mediate embryo­endometrium crosstalk and to negatively regulate embryo implantation by targeting HIF1α/FUT7/sLex pathway.

Case report: Analysis of novel compound heterozygous TPP1 variants in a Chinese patient with neuronal ceroid lipofuscinosis type 2.

Neuronal ceroid lipofuscinosis type 2 (CLN2) is an autosomal recessive neurodegenerative disease caused by variants in the TPP1 gene that lead to the deficiency of the lysosomal enzyme tripeptidyl peptidase I (TPP1) activity. Herein, we report a rare case of CLN2 caused by two novel variants of TPP1. The patient presented with seizures at onset, followed by progressive cognitive impairment, motor decline, and vision loss. Novel compound heterozygous variants, c.544_545del and c.230-3C>G, in TPP1 were identified by whole-exome sequencing. The variant assessment showed that the c.544_545del is a frameshift variant mediating mRNA decay and that c.230-3C>G is a splice variant generating aberrantly spliced TPP1 mRNA, as confirmed by a Splicing Reporter Minigene assay. In conclusion, clinical history, variant assessment, and molecular analyses demonstrate that the novel compound heterozygous variants are responsible for CLN2 disease in this patient. This study expands the mutation spectrum of TPP1.

Facial expression recognition in patients with type 2 diabetes mellitus.

BACKGROUND: Facial expression recognition is an important social cognitive skill. Type 2 diabetes mellitus (T2DM) affects cognitive function. Whether facial expression recognition deficits and attention bias exist in T2DM is unknown. Facial expression search task is a commonly used paradigm to measure emotional processing. In this study, facial expression recognition features of T2DM patients were studied by facial expression search task. METHODS: Thirty outpatients with T2DM and 30 normal controls matched by sex, age and education etc. were selected. Standardized stick drawings with happy, neutral and sad emotion expressions were selected as stimulus materials, and facial expression search task was used to Search for expression targets in neutral interferers to compare the response time between the two groups. RESULTS: The reaction time of identifying the positive expression (happy) in the diabetic group and the control group was greater than that of the negative expression (sad). The response time of the diabetic group to identify positive expressions and negative expressions was greater than that of the control group. The slope of the search for positive expressions in the diabetic group was 419.14 ms, and the search slope for negative expressions in the diabetic group was 237.97 ms. The slope of the search for positive expressions in the control group was 300.4 ms, and that of the control group for negative expressions was 119.07 ms. CONCLUSIONS: In the diabetic group and the control group, the reaction time of identifying the positive expression was positively delayed compared with the negative expression, which showed a negative attention bias; Patients with type 2 diabetes significantly prolonged the response time of recognizing positive expression and negative expression without obvious clinical cognitive impairment.