Estrogen Regulates Scribble Localization in Endometrial Epithelial Cells Through Acyl Protein Thioesterase (APT)-Mediated S-Palmitoylation in Adenomyosis.

Despite its prevalence and the severity of symptoms, little is known about the pathogenesis and etiology of adenomyosis. In our previous study, Scribble localization has been found to be partially translocated to cytoplasm; however, its regulatory mechanism is known. In consideration of the important role of supraphysiologic estrogen production in the endometrium in the development of adenomyosis, we analyzed the effect and mechanism of estrogen on Scribble localization in vivo and in vitro. Firstly, we found Scribble translocation from the basolateral membrane to the cytoplasm was easily to be seen in women and mice with adenomyosis (68% vs 27%, 60% vs 10% separately). After treatment with the S-palmitoylation inhibitor 2-bromopalmitate for 48H, cytoplasmic enrichment of Scribble and the reduced level of palm-Scribble was observed by immunofluorescence, Western blot, and acyl-biotin exchange palmitoylation assay. High estrogen exposure could not only induce partially cytoplasmic translocation of Scribble but also decrease the expression level of palm-Scribble, which can be recovered by estrogen receptor inhibitor ICI182,780. Based on following experiments, we found that estrogen regulated Scribble localization by APT through S-palmitoylation of Scribble protein. At last, IHC was performed to verify the expression of APT1 and APT2 in human clinical tissue specimens and found that they were all increased dramatically. Furthermore, positive correlations were found between APT1 or APT2 and aromatase P450. Therefore, our research may provide a new understanding of the pathogenesis of adenomyosis.

Desmethoxycurcumin aids IFNα's anti-HBV activity by antagonising CRYAB reduction and stabilising IFNAR1 protein.

The eradication of chronic hepatitis B (CHB) caused by hepatitis B virus (HBV) infection is a crucial goal in clinical practice. Enhancing the anti-HBV activity of interferon type I (IFNI) is a key strategy for achieving a functional cure for CHB. In this study, we investigated the effect of combined treatment with IFNα and Desmethoxycurcumin (DMC) on HBV replication in HepG2 cells and explored the underlying mechanism. Our results indicated IFNα alone was ineffective in completely inhibiting HBV replication, which was attributed to the virus-induced down-regulation of IFNI receptor 1 (IFNAR1) protein. However, the addition of a low dose of DMC significantly synergized with IFNα, leading to notable enhancement of IFNα anti-HBV activity. This effect was achieved by stabilising the IFNAR1 protein. Further investigation revealed that low dose DMC effectively blocked the ubiquitination-mediated degradation of IFNAR1, which was accomplished by rescuing the protein levels of alphaB-crystallin (CRYAB) and orchestrating the interaction between CRYAB and the E3 ubiquitin ligase, ß-Trcp. Importantly, over-expression of CRYAB was found to favour the antiviral activity of IFNα against HBV replication. In conclusion, our study demonstrates that low-dose DMC enhanced the anti-HBV activity of IFNα by counteracting the reduction of CRYAB and stabilising the IFNAR1 protein.

Metformin in the Treatment of Amisulpride-Induced Hyperprolactinemia: A Clinical Trial.

Objective: To evaluate the efficacy and safety of metformin in the treatment of amisulpride-induced hyperprolactinemia. Methods: A total of 86 schizophrenic patients who developed hyperprolactinemia after taking amisulpride were screened and randomly assigned to the metformin group (42 patients) and placebo group (44 patients) and followed up for eight weeks. The patients' serum prolactin levels, blood glucose and lipids were measured at the baseline and the end of the intervention. The treatment emergent symptom scale (TESS) was also assessed. Results: After eight weeks of intervention, serum prolactin levels in the metformin group decreased from (1737.360 ± 626.918) mIU/L at baseline to (1618.625 ± 640.865) mIU/L, whereas serum prolactin levels in the placebo group increased from (2676.470 ± 1269.234) mIU/L at baseline to (2860.933 ± 1317.376) mIU/L. There was a significant difference in prolactin changes (Fcovariance = 9.982, P = 0.002) between the two groups. There was no significant difference in the incidence of adverse drug reactions (P > 0.05) between the two groups. Conclusion: Metformin is able to improve amisulpride-induced hyperprolactinemia with its safety.

Scribble downregulation in adenomyosis compromises endometrial stromal decidualization by decreasing FOXO1 expression.

STUDY QUESTION: Does Scribble (SCRIB) contribute to aberrant decidualization of endometrial stromal cells (ESC) in adenomyosis? SUMMARY ANSWER: SCRIB knockdown impairs decidualization of ESC by decreasing Fork-head box O1A (FOXO1) expression through the protein kinase B (AKT) and atypical protein kinase C (aPKC) activated pathways. WHAT IS KNOWN ALREADY: Stromal SCRIB is required for primary decidual zone formation and pregnancy success in mice. In our previous studies, decidualization was dampened in ESC isolated from adenomyosis patients, yet the underlying molecular mechanisms remain elusive. STUDY DESIGN, SIZE, DURATION: Eutopic endometrium tissue samples from diffuse adenomyosis and non-adenomyosis patients in proliferative, early-secretory and mid-secretory phase (n = 10 per phase for each group) were explored. In parallel, in vitro decidualization studies were carried out in ESC isolated from non-adenomyosis women (n = 8). PARTICIPANTS/MATERIALS, SETTING, METHODS: The endometrial SCRIB expression was analyzed using immunohistochemistry staining and western blot. Quantitative RT-PCR (qRT-PCR), western blot and immunofluorescence staining were used to explore the expression of SCRIB in ESC during in vitro decidualization. siRNA-mediated SCRIB knockdown followed by decidual markers expression analysis, flow cytometry for cell cycle analysis and phalloidin staining for morphological analysis were performed to examine the function of SCRIB in ESC decidualization. RNA-sequencing was performed to examine the SCRIB-mediated transcriptional changes in decidualized ESC (DSC). Rescue experiments using an AKT inhibitor MK2206 and aPKC inhibitor NSC37044 were used to investigate the signaling pathways through which could mediate SCRIB-regulated FOXO1 protein expression and ESC decidualization. MAIN RESULTS AND THE ROLE OF CHANCE: We found that the expression of SCRIB in the mid-secretory phase eutopic endometrial stroma of adenomyosis patients was significantly lower than that of non-adenomyosis. SCRIB knockdown reduced the expression of decidual markers, abrogated the epithelioid-like morphological changes, inhibited the mesenchymal-to-epithelial transitions process and promoted the cell cycle progression of ESC during in vitro decidualization. SCRIB knockdown-induced decidualization defects were attributed to a decrease in expression of transcription factor FOXO1, known to regulate decidualization. Furthermore, we found that SCRIB knockdown induced the aberrant activation of AKT and aPKC, which led to FOXO1 phosphorylation and degradation. Rescue assay confirmed that restoring the expression of FOXO1 effectively reversed the decidualization defects and cell cycle progression caused by SCRIB knockdown. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: In this study, it was demonstrated that SCRIB knockdown mediated the activation of AKT and aPKC, contributing to FOXO1 degradation and aberrant decidualization, however, the molecular link between AKT and aPKC signaling was not determined, and still requires further exploration. WIDER IMPLICATIONS OF THE FINDINGS: Our findings support the hypothesis that adenomyosis interferes with embryo implantation due to insufficient endometrial receptivity. Abnormal decidualization of the endometrial stroma may clarify the possible association between adenomyosis and infertility. Our findings may be clinically useful for counseling and treatment of infertile adenomyosis patients. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Natural Science Foundation of China (82001523 and 82171639). The authors have no conflicts of interest to disclose. TRIAL REGISTRATION NUMBER: N/A.

Impaired decidualization of human endometrial stromal cells from women with adenomyosis†.

Differentiation of endometrial stromal cells (ESCs) into secretory decidualized cells (dESCs) is essential for embryo implantation. Adenomyosis is a common benign gynecological disease that causes infertility. However, whether adenomyosis affects decidualization of human ESCs is elusive. Primary eutopic ESCs were obtained from patients with adenomyosis (n = 9) and women with nonendometrial diseases (n = 12). We determined the capacity of decidualization of human ESCs by qRT-PCR, Edu proliferation assay, cytokine array, and ELISA assay. We found that the expression of decidualization markers (IGFBP1 and PRL) in ESCs of adenomyosis was reduced, concomitant with increased cell proliferation. Differential secretion of cytokines in dESCs, including CXCL1/2/3, IL-6, IL-8, MCP-1, VEGF-A, MIP-3α, OPN, SDF-1α, HGF, and MMP-9, was observed between adenomyosis and nonadenomyosis. Moreover, the expression of decidualization regulators (HOXA10 at both mRNA and protein levels, FOXO1, KLF5, CEBPB, and HAND2 at mRNA levels) in the eutopic endometrium of adenomyosis was lower than that of nonadenomyosis. We propose that ESCs from adenomyosis have defected ability to full decidualization, which may lead to a nonreceptive endometrium.

Celecoxib, a selective COX-2 inhibitor, markedly reduced the severity of tamoxifen-induced adenomyosis in a murine model.

The aim of the present study was to evaluate the effects of the selective cyclooxygenase (COX)-2 inhibitor celecoxib on the development of uterine adenomyosis in mice. ICR neonatal mice were first exposed to tamoxifen to establish a mouse model of adenomyosis. Following 60 days of celecoxib treatment, pathological formation of adenomyosis lesions and the depth of myometrial infiltration were evaluated using hematoxylin and eosin staining. To examine thermal pain modulation in mice, a hotplate test was conducted every 15 days from postnatal day 30 onwards. Immunohistochemistry was performed to assess the expression of aromatase P450, N-cadherin, E-cadherin, COX-2 and cluster of differentiation 31, whereas the levels of estrogen were analyzed in uterine tissue homogenates using ELISA. Masson trichrome staining was performed to assess the extent of fibrosis in the uterus. Celecoxib treatment significantly inhibited the depth of infiltration into the myometrium, resulting in significantly reduced disease severity. Treatment with high doses of celecoxib significantly prolonged thermal response latency. Following celecoxib treatment, the expression of E-cadherin was significantly increased whereas the expression of N-cadherin was significantly decreased. Concomitantly, the extent of fibrosis was also reduced following celecoxib treatment. Uterine tissue homogenates isolated from mice treated with both high and low doses of celecoxib exhibited lower concentrations of estrogen and decreased expression of aromatase P450. These observations suggest that celecoxib reduces adenomyosis severity by suppressing estrogen production in the uterus, reversing epithelial-mesenchymal transition and relieving fibrosis. Taken together, the results of the present study support the potential use of celecoxib, a selective COX-2 inhibitor, for the treatment of adenomyosis.

Estrogen degrades Scribble in endometrial epithelial cells through E3 ubiquitin ligase HECW1 in the development of diffuse adenomyosis†.

Despite its prevalence and the severity of symptoms, little is known about the pathogenesis and etiology of adenomyosis. In previous studies, the protein expression level of the polarity protein Scribble in the eutopic endometrium of patients with adenomyosis was found to be significantly decreased; however, little is known about its regulatory mechanism. In consideration of the important role of supraphysiologic estrogen production in the endometrium in the development of adenomyosis, we analyzed the effect and mechanism of estrogen on the expression of Scribble in vivo and in vitro. Firstly, we found Scribble was downregulated in eutopic endometrium and negatively related with aromatase P450 in tamoxifen-induced adenomyosis. Then, we established a 3D culture of primary endometrial epithelial cells and found that estrogen could disrupt apical-basal polarity of endometrial glandular epithelial cells. Based on the following experiments and GEO dataset screening, we found that estrogen regulates the expression level of Scribble by HECW1 through ubiquitination of Scribble protein. At last, we verified the expression of Scribble, HECW1, and aromatase P450 in eutopic endometrium of human and mouse specimens and found that the expression of HECW1 and aromatase P450 was significantly increased, while the expression of Scribble was significantly downregulated. Furthermore, a positive correlation was found between HECW1 and aromatase P450, while a negative correlation was found between HECW1 and Scribble in human clinical tissue specimens. Therefore, our research may provide a new understanding of the pathogenesis of adenomyosis.

Baicalein reduces endometriosis by suppressing the viability of human endometrial stromal cells through the nuclear factor-κB pathway in vitro.

The aim of the present study was to evaluate the effects of baicalein on human endometrial stromal cells in vitro. Ectopic endometrium samples were obtained from 6 female patients with ovarian endometriosis who underwent laparoscopic surgical procedures from July to September 2015. After culturing the cells, immunocytochemistry was performed to verify the purity and homogeneity of the endometrial stromal cells, and a Cell Counting Kit-8 assay was used to evaluate cell viability. In addition, cell cycle progression was analyzed using flow cytometry, and the effects of baicalein on the expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), proliferating cell nuclear antigen (PCNA) and cyclin D1 in endometrial stromal cells were evaluated using western blot analysis. The related signaling pathways were also investigated by incubating cells with inhibitors of signaling pathways, prior to adding 40 µM baicalein for 48 h, followed by analysis of cell viability using a Cell Counting Kit-8 assay. The results indicated that treatment with baicalein significantly induced a dose-dependent decrease (P<0.05) in the viability of human endometrial stromal cells, which was abolished by inhibition of the nuclear factor (NF)-κB signaling pathway. However, baicalein treatment did not induce a time-dependent decrease in viability, as cell viabilities between the 24, 48 and 72 h treatment groups did not differ significantly. The number of cells in the G0/G1 phase significantly increased following treatment with baicalein (P<0.05), while the number of cells in the S and G2/M phases significantly decreased (P<0.05). Baicalein-treated cells also exhibited significantly reduced expression of Bcl-2, PCNA and cyclin D1 compared with control cells (P<0.05). These results suggested that baicalein may suppress the viability of human endometrial stromal cells through the NF-κB signaling pathway in vitro, and may induce apoptosis and promote cell cycle arrest at the G0/G1 phase. Thus, baicalein may provide a novel treatment option for endometriosis.

Vascular endothelial growth factor receptor-2 inhibitor cediranib causes regression of endometriotic lesions in a rat model.

Vascular endothelial growth factor (VEGF) receptor-2 plays an essential role in angiogenesis, and it also expressed in the glandular epithelium and stromal cells of ectopic endometrium. Cediranib is a protein tyrosine kinase inhibitor that potently inhibits VEGF receptor-2, but there is no study about its effects on the endometriosis. We induced endometriosis on both sides of the abdominal wall in 20 female Sprague-Dawley rats and randomly divided them into 2 groups. They were administered: cediranib 4 mg/kg/day (group 1), equal saline (group 2) for 12 days. Then, the lesion volumes were calculated, and Masson trichrome was used to detect fibrosis. Angiogenesis was evaluated by CD-31 immunohistochemistry and serum VEGF levels. Proliferation was indicated by proliferating cell nuclear antigen immunohistochemistry. Apoptosis was measured by a TUNEL assay and cleaved caspase-3 immunohistochemistry. In the treatment group, the lesion volumes were smaller (P < 0.05), and the degree of fibrosis was greater. The microvessel density was lower (P < 0.05) than control, however, serum VEGF was up-regulated by a negative feedback mechanism (P < 0.01). In addition, proliferation was significantly suppressed (P < 0.01), and apoptosis in the lesions was more obvious in the treatment group. These data indicated that cediranib can inhibit development of endometriotic lesions in rats.

Effect of GuiXiong Xiaoyi Wan in Treatment of Endometriosis on Rats.

Objective. To evaluate the effect of GuiXiong Xiaoyi Wan (GXXYW) on the development of endometriosis in a rat model. Methods. Sprague-Dawley rats with surgically induced endometriosis were randomly treated with low-dose GXXYW, high-dose GXXYW, or vehicle (negative control) for 28 days. Immunohistochemistry was used to assess cell proliferation in the lesions. The terminal deoxynucleotidyl transferase- (TdT-) mediated dUTP biotin nick end labelling (TUNEL) method was performed to analyse the apoptosis induced by GuiXiong Xiaoyi Wan. The percentages of CD3+ lymphocytes, CD4+ lymphocytes, and CD8+ lymphocytes in the spleens of the rats were evaluated using flow cytometric analysis. Results. Treatment with GXXYW significantly decreased the lesion size, inhibited cell proliferation, and induced apoptosis in endometriotic tissue. The spleens of GXXYW-treated rats also demonstrated a significant increase in the percentage of CD4+ lymphocytes and a significant decrease in the percentage of CD8+ lymphocytes. Conclusions. These results suggest that, in a rat model, GXXYW may be effective in the suppression of the growth of endometriosis, possibly through the inhibition of cell proliferation, the induction of apoptosis of endometriotic cells, and the regulation of the immune system.

An ERP study on whether the P600 can reflect the presence of unexpected phonology.

In this study, we conducted a new approach to determine whether the P600 could reflect the presence of unexpected phonology. In the experiment, the critical Chinese characters in the poems are either original or substituted by homophones or synonyms. The N400 effect is observed only under the homophonic condition. Late positive shifts are revealed under both homophonic and synonymous conditions, but the amplitude under the homophonic condition is smaller than that under the synonymous condition. The observation of the N400 effect under the homophonic condition indicates that semantic processing occurs in Chinese poem comprehension, while the P600 effect elicited under the homophonic condition is a reflection of unfulfilled semantic expectations. The existence of the late positivity under the synonymous condition reveals that P600 can reflect the presence of unexpected phonology. Interestingly, late positivities under the two conditions both consist of two parts. A discussion ensues wherein these two parts of the late positive shift relate, respectively, to monitoring and resolution processes in language comprehension.

The processing of phonological, orthographical, and lexical information of Chinese characters in sentence contexts: an ERP study.

In the current work, we aimed to study the processing of phonological, orthographical, and lexical information of Chinese characters in sentence contexts, as well as to provide further evidence for psychological models. In the experiment, we designed sentences with expected, homophonic, orthographically similar, synonymous, and control characters as endings, respectively. The results indicated that P200 might be related to the early extraction of phonological information. Moreover, it might also represent immediate semantic and orthographic lexical access. This suggested that there might be a dual-route in cognitive processing, where the direct access route and the phonologically mediated access route both exist and interact with each other. The increased N400 under the control condition suggested that both phonological and orthographical information would influence semantic integration in Chinese sentence comprehension. The two positive peaks of the late positive shift might represent the semantic monitoring, and orthographical retrieval and reanalysis processing, respectively. Under the orthographically similar condition, orthographical retrieval and reanalysis processing was more difficult in comparison with the other conditions, which suggested that there might be direct access from orthography to semantic representation in cognitive processing. In conclusion, it was shown that the direct access hypothesis or the dual-route hypothesis could better explain cognitive processing in the brain.

The influence on cognitive processing from the switches of shooting angles in videos of real-world events: an ERP study.

This work mainly focuses on the influence from switches of shooting angles in videos during the cognitive processing in the human brain. In the experiment we used the videos with switches of shooting angles as materials and compared the ERPs elicited by the switch frames and the non-switch frames in the videos, it was found that when subjects were asked to pay attention to the video contents, the switch frames would trigger P3a-RON waveforms, but no N400 waveform was found in the ERP results. This showed that when subjects were concerned with the video contents, the switches of shooting angles would distract their attention from the video contents, but as long as the semantic meaning of the videos were coherent, the switches of shooting angles would not lead to significant difficulties in semantic comprehension. At the same time, the experimental results also further proved that the P3a and RON generally reflect the processing of task-irrelevant visual stimuli.

The interaction between pictures and words: evidence from positivity offset and negativity bias.

In order to study the interaction between pictures and words, we investigated the variations of their actual arousal levels through the observation of the positivity offset and negativity bias under different conditions. We used emotional pictures and emotional Chinese words to construct stimuli under four conditions: (1) only a word was presented (Word Only condition), (2) only a picture was presented (Picture Only condition), (3) a word was presented before a picture (Word Before Picture condition) and (4) a picture was presented before a word (Picture Before Word condition). The picture and word in each target pair under the conditions (3) and (4) were congruous in content and emotion. Significant negativity bias is noticed under the Picture Only and the Word Only conditions. Effects analogous to a positivity offset are observed under the other two conditions. It is suggested that the actual arousal levels of multiple stimuli, such as those presented under the conditions (3) and (4) are different from those of individual pictures and individual words, while the actual arousal levels of individual pictures do not differ significantly from those of individual words. The results indicate that content consistency can lead to a reduction in emotional attention, and also that the influence of pictures on words will differ from the opposite condition.

Emotional facilitation effect in the picture-word interference task: an ERP study.

In this paper, we aimed to verify the emotional facilitation effect in the picture-word interference task using event-related potentials. Twenty-one healthy subjects were asked to categorize the emotional valences of pictures accompanied by emotionally congruent, either centrally or laterally positioned Chinese words. For both the foveal and lateral word presentations, the reaction times were faster compared to the picture-only presentation. Compared to the picture-only presentation, P200 amplitude increased under the foveal word presentation condition and decreased under the lateral word presentation condition, indicating that more attentional resources were required when an accompanying word was in the center of a picture than when the word was in the lateral position. Latency of P300 was shorter in response to picture-word stimuli irrespective of word position, indicating that an emotionally congruent word facilitated the emotional processing of the target picture, which verified the emotional facilitation effect and was consistent with the results in psychology. The late positive component was more positive when the picture-word stimuli were more positive, which reflected the feature of positivity offset. The findings suggest that the time window for an emotional facilitation effect may be limited to processing aspects associated with P300 (i.e., affective evaluation).

The integration processing of the visual and auditory information in videos of real-world events: an ERP study.

In real life, the human brain usually receives information through visual and auditory channels and processes the multisensory information, but studies on the integration processing of the dynamic visual and auditory information are relatively few. In this paper, we have designed an experiment, where through the presentation of common scenario, real-world videos, with matched and mismatched actions (images) and sounds as stimuli, we aimed to study the integration processing of synchronized visual and auditory information in videos of real-world events in the human brain, through the use event-related potentials (ERPs) methods. Experimental results showed that videos of mismatched actions (images) and sounds would elicit a larger P400 as compared to videos of matched actions (images) and sounds. We believe that the P400 waveform might be related to the cognitive integration processing of mismatched multisensory information in the human brain. The results also indicated that synchronized multisensory information would interfere with each other, which would influence the results of the cognitive integration processing.