Inhibition of Foxp3 expression in the placenta of mice infected intraperitoneally by toxoplasma gondii tachyzoites: insights into the PPARγ/miR-7b-5p/Sp1 signaling pathway.

BACKGROUND: Toxoplasma gondii, an obligate intracellular parasitic protozoa, infects approximately 30% of the global population. Contracting T. gondii at the primary infection of the mother can result in neonatal microcephaly, chorioretinitis, hydrocephalus, or mortality. Our previous study indicated that pregnant mice infected with T. gondii displayed a decrease in both the number and the suppressive ability of regulatory T cells, accompanied by the reduced Forkhead box P3 (Foxp3). Numerous studies have proved that microRNAs (miRNAs) are implicated in T. gondii infection, but there is meager evidence on the relationship between alterations of miRNAs and downregulation of Foxp3 induced by T. gondii. METHODS: Quantitative reverse transcription polymerase chain reaction was utilized to detect the transcriptions of miRNAs and Foxp3. Protein blotting and immunofluorescence were used to detect the expressions of Foxp3 and related transcription factors. The structure of mouse placenta was observed by hematoxylin and eosin (HE) staining. To examine the activity of miR-7b promoter and whether miR-7b-5p targets Sp1 to suppress Foxp3 expression, we constructed recombinant plasmids containing the full-length/truncated/mutant miR-7b promoter sequence or wildtype/mutant of Sp1 3' untranslated region (3' UTR) to detect the fluorescence activity in EL4 cells. RESULTS: In T. gondii-infected mice, miR-7b transcription was significantly elevated, while Foxp3 expression was decreased in the placenta. In vitro, miR-7b mimics downregulated Foxp3 expression, whereas its inhibitors significantly upregulated Foxp3 expression. miR-7b promoter activity was elevated upon the stimulation of T. gondii antigens, which was mitigated by co-transfection of mutant miR-7b promoter lacking peroxisome proliferator-activated receptor γ (PPARγ) target sites. Additionally, miR-7b mimics diminished Sp1 expression, while miR-7b inhibitors elevated its expression. miR-7b mimics deceased the fluorescence activity of Sp1 3' untranslated region (3' UTR), but it failed to impact the fluorescence activity upon the co-transfection of mutant Sp1 3' UTR lacking miR-7b target site. CONCLUSIONS: T. gondii infection and antigens promote miR-7b transcription but inhibit Foxp3 protein and gene levels. T. gondii antigens promote miR-7b promoter activity by a PPARγ-dependent mechanism. miR-7b directly binds to Sp1 3' UTR to repress Sp1 expression. Understanding the regulatory functions by which T. gondii-induced miR-7b suppresses Foxp3 expression can provide new perspectives for the possible therapeutic avenue of T. gondii-induced adverse pregnancy outcomes.

Construction and analysis of a joint diagnostic model of machine learning for cryptorchidism based on single-cell sequencing.

BACKGROUND: Cryptorchidism is a condition in which one or both of a baby's testicles do not fully descend into the bottom of the scrotum. Newborns with cryptorchidism are at increased risk of developing infertility later in life. The aim of this study was to develop a novel diagnostic model for cryptorchidism and to identify new biomarkers associated with cryptorchidism. METHODS: The study data were obtained from RNA sequencing data of cryptorchid patients from Nantong University Hospital and the Gene Expression Omnibus (GEO) database. Differential expression analysis was used to obtain differentially expressed genes (DEGs) between the control and cryptorchid groups. These DEGs were analyzed for their functions by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment using GSEA software. Random Forest algorithm was used to screen central genes based on these DEGs. Neuralnet software package was used to develop artificial neural network models. Based on clinical data, receiver operating characteristic (ROC) was used to validate the models. Single-cell sequencing analysis was used for the pathogenesis of cryptorchidism. RESULTS: We obtained a total of 525 important DEGs related to cryptorchidism, which are mainly associated with biological functions such as supramolecular complexes and microtubule cytoskeleton. Random forest approach screening obtained eight hub genes. A neural network based on the hub genes showed a 100% success rate of the model. Finally, single-cell sequencing analysis validated the hub genes. CONCLUSION: We developed a novel diagnostic model for cryptorchidism using artificial neural networks and validated its utility as an effective diagnostic tool.

miR-942-5p Regulates Proliferation, Invasion and EMT of Trophoblast Cells in Gestational Diabetes by Targeting the CEBPA.

Objective: Gestational diabetes mellitus (GDM) is a prevalent metabolic disorder caused by abnormal glucose metabolism during pregnancy. Trophoblast dysfunction induced by hyperglycemia during pregnancy is the main factor leading to the development of GDM. In this study, we evaluated the expression of miR-942-5p in the placenta of patients with GDM and its regulation of trophoblast cell biological function. Methods: HTR-8/SVneo trophoblast cells were incubated with glucose to establish in vitro models, and miR-942-5p mimics transfected cells were added. The expression levels of miR-942-5p, CCAAT-enhancer-binding protein alpha (CEBPA) and N-cadherin in tissues and cells were detected by reverse-transcription quantitative polymerase chain reaction (RT-qPCR), protein blotting and immunohistochemistry (IHC). MTT, flow cytometry and Transwell assays were used to determine changes in cell proliferation, apoptosis and invasion. Dual-luciferase reporter assay and Pearson analysis were used to confirm the association between miR-942-5p and CEBPA. Results: miR-942-5p and N-cadherin were decreased in placental tissue and in human placental trophoblast cells (HTR-8/SVneo) exposed to high glucose (HG) conditions, while CEBPA was increased in placental tissue and HTR-8/SVneo exposed to HG conditions. Elevated levels of miR-942-5p suppressed apoptosis induced by HG and facilitated the proliferative and invasive capacities of HTR-8/SVneo. Mechanistically, we confirmed that miR-942-5p overexpression directly targeted CEBPA and suppressed CEBPA expression, while upregulating N-cadherin expression, which is involved in the EMT process of trophoblast cells and alleviated the dysfunction of trophoblast cells induced by HG in GDM. Conclusion: Overexpression of miR-942-5p promotes proliferation, invasion and EMT of trophoblast cells by targeting and negatively regulating CEBPA. These findings offer novel understanding regarding the treatment of GDM.

A comprehensive study of oxidative stress-related effects on the prognosis and drug therapy of cervical cancer.

BACKGROUND: Cervical cancer (CC) is a serious global disease with poor prognoses and a significant recurrence rate in patients with advanced disease. Oxidative stress (OS) greatly influences many types of human cancers, making it crucial to understand the functional mechanisms of OS-related genes in CC. METHODS: The transcriptome and clinical data of three normal samples and 306 patients with CC were obtained from The Cancer Genome Atlas dataset. The GSE44001 dataset was acquired from the Gene Expression Omnibus database. OS-related subtypes in the cohort with CC were identified using unsupervised hierarchical clustering, univariate Cox analysis, gene set enrichment analysis (GSEA), and least absolute shrinkage and selection operator regression analysis. Additionally, molecular pathways that differ across subtypes were determined and OS-related genes linked to the prognosis of patients of CC were determined. Finally, a clinical prognostic gene signature was developed and validated. The relative infiltration level of immune cell subpopulations in different risk groups and subtypes was evaluated using the cell-type identification by estimating relative subsets of RNA transcripts (CIBERPORT) algorithm and single-sample GSEA (ssGSEA) techniques. RESULTS: The present study established two distinct OS subtypes (OS clusters A and B). Analysis using ssGSEA and CIBERSPORT revealed that OS cluster B exhibited a significant level of immune infiltration. A clinical prognostic gene signature was established using OS-related characteristic genes identified by examining the differentially expressed genes across both subtypes. Furthermore, patients with CC were grouped into high- and low-risk groups, with the low-risk group showing higher survival rates. Additionally, these individuals exhibited significant advantages in terms of survival and immunotherapy. Receiver operating characteristic curve analysis demonstrated the higher predictive value of the clinical prognostic gene signature. The outcomes of the validation group depicted congruence with those recorded in the training group. CONCLUSIONS: A new model was constructed based on eight OS-related characteristic genes to aid the prediction of the survival rates of individuals with CC. The present study contributes to the existing literature on the mechanisms of OS genes in CC and offers a fresh perspective for future advancements in immunotherapy for such individuals.

Single-cell sequencing of brain tissues reveal the central nervous system's susceptibility to SARS-CoV-2 and the drug.

Background: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the current COVID-19 pandemic, resulting in a public health crisis that required immediate action. The SARS-CoV-2 virus enters human cells via three receptors, namely cathepsin, angiotensin-converting enzyme 2 (ACE2) and SARS-CoV receptors. Cathepsin destroys the spike protein (S protein), thereby allowing the entry of viral nucleic acid into human host cells. Methods: Utilizing single-cell transcriptome analysis of brain tissues, the vulnerability of the central nervous system to infection with SARS-CoV-2 in humans was investigated. Results: ACE2 is mainly expressed in endothelial cells, with the highest levels found in ageing endothelial cells. Drug prediction suggests that (-)-catechin reduces the effects of COVID-19 on the nervous system. Immunohistochemistry analysis showed that ACE2 was mainly expressed in cerebral vessels. Immunofluroscenceresults showed the co-expression of CD31 and ACE2 in human tissues. Western blot further showed that ACE2 expression was higher in old rats than in young rats. Conclusion: This study provides insight into the mechanism of SARS-CoV-2 brain invasion. Accordingly, patients with neurological symptoms who are infected with SARS-CoV-2 should be given individualised care.

Toxoplasma gondii promotes microRNA-34a to inhibit Foxp3 expression in adverse outcomes of pregnancy in mice.

Infection with Toxoplasma gondii (T. gondii) during the pregnant period causes microcephaly, mental and psychomotor retardation or death. Miserable outcomes are mainly linked with regulatory T cells (Tregs) dysfunction. Forkhead box P3 (Foxp3), a vital regulator of establishment and maintenance of Tregs, can be modulated by microRNAs (miRNAs). Previously, our study revealed that T. gondii infection in pregnant mice induced Tregs dysfunction, accompanied with reduced Foxp3 expression. The role of miRNAs in the inhibition of Foxp3 triggered by T. gondii remains unclear. Herein, T. gondii infection promotes miR-34a expression in the placenta of mice. miR-34a mimic inhibits Foxp3 expression via targeting 3' untranslated region (3'UTR) whereas its inhibitor promotes Foxp3 expression in vitro. T. gondii antigens could enhance the activity of miR-34a promoter via a Smad4-dependent mechanism. Collectively, our data reveal a new avenue through which T. gondii inhibits Foxp3 expression necessary to drive adverse outcomes of pregnancy in mice.

Waste-converted nitrogen and fluorine co-doped porous carbon nanosheets for high performance supercapacitor with ionic liquid electrolyte.

In this work, nitrogen and fluorine co-doped porous carbon nanosheets (NFPCNS) were fabricated from pharmaceutical drug residues derived from the fermentation synthesis of lincomycin hydrochloride via high-temperature pyrolysis and subsequent KOH activation without adding any nitrogen and fluorine reagents. The obtained NFPCNS exhibits an optimized integration of three dimensional interconnected nanosheet structure, large specific surface area of 2912 m2 g-1, hierarchical porous structure with large mesopore proportion (Smeso/Smicro = 151.5%, Vmeso/Vmicro = 248.2%) and high level heteroatom content (13.2 at.% O, 4.3 at.% N and 1.0 at.% F). Therefore, NFPCNS based supercapacitors using 1-ethyl-3-methylimidazolium tetrafluoroborate electrolyte exhibit an excellent gravimetric capacitance of 296F g-1 at 1 A g-1, good rate capability of 65% at 20 A g-1 and high energy density of 125 Wh kg-1. Furthermore, an ultra-high energy density of 173 Wh kg-1 and a long cycling life with 93% capacitance retention after 2000 cycles has been achieved by NFPCNS based supercapacitors with 1-ethyl-3-methylimidazolium bis[(trifluoromethyl)sulfonyl]imide electrolyte. NFPCNS should be a green and efficient electrode materials for next-generation high-energy supercapacitors.

PD-L1 Expression and Intra-Tumoral CD8+ T Lymphocytes in Esophageal Carcinosarcoma.

We detected PD-L1 and intra-tumoral CD8+ T lymphocytes (CD8+ TIL) in 19 patients with esophageal carcinosarcoma (ECS). The median follow-up period of these patients was 43 months, and the three- and five-year survival rates were 78.9 and 63.2%, respectively. No statistically significant correlation was observed between PD-L1 and CD8+ TIL in sarcomatous components(SC) (r = -0.262, p = 0.279) and epithelial carcinomatous (EC) (r = 0.055, p = 0.824). This study examined the immunological markers in ECS for the first time. PD-L1 is highly expressed in the SC and is associated with a poorer prognosis.

High GNG13 expression is associated with poor survival in epithelial ovarian cancer and breast cancer.

Recently, change in the GNG13 expression has been shown to result in multiple congenital malformations and sexual reversal, and it was also found in the brain. The aim of this study was to measure the expression levels in epithelial ovarian cancer (EOC) and breast cancer (BC) and assess their value as a potential prognostic marker. The correlation of GNG13 protein expression was detected by immunohistochemistry (IHC) in 119 EOC and 125 BC tissues. Assessment of the associations between GNG13 levels and various clinicopathological features was identified, the relationship between GNG13 and prognosis in BC and EOC patients was analyzed using online resources of Oncomine and Kaplan-Meier plotter. Protein expression levels of GNG13 were both significantly lower in BC and EOC compared with normal tissues (p<0.0001 and p<0.001, respectively). Among the clinicopathological characteristics of BC, tumor grade (p=0.001) and TNM stage (p=0.001) were significantly associated with low expression of GNG13. While in EOC, low expression of GNG13 was significantly related to FIGO stage (p=0.001), presence of metastasis (p=0.001), and CA125 (p=0.001). Our data suggest that GNG13 expression maybe as a new inhibitor, which can strongly inhibit metastasis and partially attenuates tumor growth in EOC and BC.

Postpartum septic pelvic thrombophlebitis and ovarian vein thrombosis after caesarean section: a rare case report.

BACKGROUND: Septic pelvic thrombophlebitis (SPT) is a well-recognized but rare puerperal complication that has two types: ovarian vein thrombophlebitis (OVT) and deep septic pelvic thrombophlebitis (DSPT). The present case report describes the clinical and imaging findings of a female patient diagnosed with right ovarian vein infectious thrombophlebitis after caesarean section (C-section). CASE PRESENTATION: A 35-year-old G3P2 female who presented with a foetal vein Galen malformation at 41 weeks of gestation underwent C-section. The patient had high fever after C-section, and anti-inflammatory treatment was not effective within 1 week. An abdominal wall incision haematoma was found, and a second surgery for the removal of the abdominal wall haematoma was performed. The patient was ultimately diagnosed with abdominal incision haematoma and right ovarian vein infectious thrombophlebitis after C-section. We used imipenem and tigecycline to strengthen the anti-inflammatory effects, simultaneously administrating low-molecular-weight heparin and warfarin as anticoagulant therapy. On the 30th day after C-section, the right ovarian vein thrombus disappeared. CONCLUSION: This case illustrates the need to consider the potential relationship between abdominal incision haematoma and ovarian vein thrombophlebitis. Despite advances in the management of venous thromboembolism globally, more data on epidemiology in terms of first incidence, prevalence, recurrence and risk factors, management of bleeding complications, and increased awareness in Asian populations are necessary.

Development of a Prognostic Model for Ovarian Cancer Patients Based on Novel Immune Microenvironment Related Genes.

Ovarian cancer (OV) has become the most lethal gynecological cancer. However, its treatment methods and staging system are far from ideal. In the present study, taking the advantage of large-scale public cohorts, we extracted a list of immune-related prognostic genes that differentially expressed in tumor and normal ovarian tissues. Importantly, an individualized immune-related gene based prognostic model (IPM) for OV patients were developed. Furthermore, we validated our IPM in Gene Expression Omnibus (GEO) repository and compared the immune landscape and pathways between high-risk and low-risk groups. The results of our study can serve as an important model to identify the immune subset of patients and has potential for use in immune therapeutic selection and patient management.

Global 3'UTR shortening and down-regulation of repeated element related piRNA play crucial roles in boys with cryptorchidism.

BACKGROUND: Cryptorchidism is the most common congenital defect in children's genitourinary system. Decades of research have identified both environmental and genetic factors contribute to the etiology. METHODS: Small-RNA/mRNA-seq were performed on testicular tissues from cryptorchidism patients. Downstream analysis included mRNA expression, piRNA expression and miRNA expression. RESULTS: We find a global downregulation of repeated element related piRNA expression as well as a global 3'UTR shortening of mRNAs in patients with cryptorchidism. We also find that genes with shortened 3'UTR which are highly enriched in vascular endothelial growth and protein ubiquitination, tend to be up-regulated in cryptorchidism. These results indicate that boys with cryptorchidism may not have normal piRNA functions to protect developmental tissues from transposon invasion. Dysregulated shortened 3'UTR genes may affect normal testicular tissue development. CONCLUSION: In summary, our findings also provided the first landscape of gene regulation in cryptorchidism, especially in terms of post-transcriptional regulations.

High neuropilin and tolloid-like 1 expression associated with metastasis and poor survival in epithelial ovarian cancer via regulation of actin cytoskeleton.

Abnormal expression of neuropilin and tolloid-like 1 (NETO1) has been detected in some human carcinomas. However, the expression of NETO1 and the underlying mechanism in epithelial ovarian cancer (EOC) remain unknown. In this study, we found that a higher NETO1 expression in EOC tissue samples compared to normal ovarian tissue samples was significantly correlated with worse overall survival. Additionally, Cox regression analysis suggested that NETO 1 was independently associated with overall survival. NETO1 overexpression enhanced the EOC cells' migration and invasion capability in vitro via regulation of actin cytoskeleton. Mechanistically, silencing NETO1 reduced the expression of ß-tubulin, F-actin and KIF2A. In conclusion, our results demonstrated the critical role of NETO1 in EOC invasion, and therapies aimed at inhibiting its expression or activity might significantly control EOC growth, invasion and metastatic dissemination.

Dysfunction of Tregs contributes to FGR pathogenesis via regulating Smads signalling pathway.

Fetal growth restriction (FGR) is ranked number two of most common complication of abnormal pregnancy worldwide. The pathogenesis of FGR is complicated due to multiple aetiologies and the exact mechanism for FGR development is currently unknown. T regulatory cells (Tregs) are proven to play central roles in the maintenance of normal pregnancy. Peripheral blood samples of 102 pregnant human were collected analysed using flow cytometry to identify Tregs. We found that reduced Tregs and down-regulation of Foxp3 were observed in peripheral blood of FGR patients. In FGR mouse model, we have found that Tregs were not only reduced in spleen but also in placenta. In vitro, Foxp3 and its transcription regulatory signalling molecules, including P-Smad2, P-Smad3 and Smad4, were diminished as well. Inhibition on Foxp3 expression was partially reversed by overexpression of Smad2 and Smad4. In FGR patients, Western blot results revealed that Foxp3, P-Smad2, P-Smad3 and Smad4 expression was inhibited in placenta. Our preliminary result suggests that maternal-foetal immune tolerance mediated by Tregs plays an essential role in the development of FGR. The inhibited expression of Foxp3 and down-regulated Smad2/Smad3/Smad4 signalling pathway were involved in the FGR pathogenesis. Targeting maternal-foetal immune tolerance through Tregs might represent a novel therapeutic option for FGR.

miR-203a-3p regulates the cellular processes of esophageal cancer cells via targeting CtBP2.

BACKGROUND: MicroRNAs (miRNA) (small noncoding RNAs) are vital modulators of gene expression by mRNA degradation and translational silencing. However, the definite mechanism and character of miR-203a-3p in regulating esophageal carcinoma cells remain unexplained. Here we further investigate the effect and the latent target gene of miR-203a-3p on the progression of esophageal squamous cell cancer (ESCC) tissues and cells. METHODS: The expressions of miR-203a-3p in ESCC tissues and peri-neoplastic tissues were further measured by RT-quantitative-PCR (RT-qPCR). Luciferase assay was applied to confirm that C-terminal-binding protein 2 (CtBP2) was the potential target gene of miR-203a-3p. miRNA mimic was transfected into ECA109 cells to up-regulate the miR-203a-3p expression, and its and CtBP2 expression were tested using RT-qPCR and Western blot. In vitro, MTT, transwell, wound healing, TUNEL and flow cytometry (FCM) assay were used to explore the role of miR-203a-3p on the cellular processes of ECA109 cells via targeting CtBP2. Furthermore, we designed rescue experiments by using CtBP2 stable over-expression ECA109 cells. RESULTS: We found the miR-203a-3p expressions in ESCC tissues and cells were significantly raised. miR-203a-3p negatively regulated the CtBP2 expression, and caused to inhibiting proliferation, migration and invasion, and promoting apoptosis in ECA109 cell. In addition, proteins involved in epithelial-mesenchymal transition (EMT) were measured by Western blot in ECA109 cells. miR-203a-3p enhanced the E-cadherin and ß-catenin expression, while reduced vimentin expression in ECA109 cells. In vivo, Xenograft tumor model demonstrated that tumor volume in miR-203a-3p agomir group was remarkably decreased. CONCLUSIONS: miR-203a-3p plays a vital role in the metastasis of ESCC cell by targeting CtBP2, and offers a promising therapeutic target for ESCC treatment.

Therapeutic effects of modified Devine surgery for concealed penis in children.

OBJECTIVE: Considerable controversy exists regarding the surgical indications for a concealed penis. We herein describe a modified technique for correction of a concealed penis. The superfluous inner plate is resected to accelerate the disappearance of the postoperative lymphedema, and the skin between the penis and scrotum is trimmed to recover the penoscrotal angle. METHODS: From January 2014 to October 2017, 79 patients with a concealed penis underwent our modified Devine penoplasty procedure. We measured the penile length preoperatively and postoperatively to confirm the improvement. A questionnaire was administered to the patients' parents to assess satisfaction regarding penile size, morphology, voiding status, and hygiene. RESULTS: The perpendicular penile length was 1.88 ± 0.76 cm preoperatively and 4.42 ± 0.48 cm postoperatively, representing a significant improvement(p < 0.05). The parents' satisfaction grades for penile size, morphology, voiding status, and hygiene were significantly improved postoperatively (p < 0.05). Almost every patient had postoperative penile lymphedema; however, this symptom had spontaneously resolved by 6 weeks. No other complications occurred, such as skin necrosis, tissue contracture, or wound infection. CONCLUSION: The herein-described modified repair technique for a concealed penis was technically feasible and safe, and excellent postoperative satisfaction was achieved. Additionally, the postoperative penis exhibited a good cosmetic appearance. Because of the successful outcomes with few complications, we believe that this surgical method for selected patients with a concealed penis is more effective than the traditional method. LEVEL OF EVIDENCE: Level IV, therapeutic study.

Overexpression of KIF2A is Suppressed by miR-206 and Associated with Poor Prognosis in Ovarian Cancer.

BACKGROUND/AIMS: This study aimed to investigate the expression and prognostic value of kinesin family member 2A (KIF2A) and the suppression effects of microRNA-206 (miR-206) on KIF2A in ovarian cancer. METHODS: Ovarian cancer tissues from patients and ovarian cancer cell lines (A2780 and SKOV3) were used in this study. miR-206 mimics and control were transiently transfected into cells. RT-qPCR was performed to detect KIF2A mRNA and miR-206 expression levels, Western blot was performed to detect KIF2A protein levels, Dual-Luciferase Reporter Assay was used to examine the inhibition effects of miR-206 on KIF2A mRNA, immunohistochemical staining was used to examine the expression of KIF2A in tissue sections. CCK-8, transwell and Annexin-V-FITC/Propidium Iodide staining with flow cytometry were used to detect the cell proliferation, migration/invasion, and apoptosis respectively. RESULTS: Our study explored the expression profiles of KIF2A and miR-206 in the patients with ovarian cancer. We found that overexpression of KIF2A was associated with a poor prognosis in ovarian cancer. We also found that KIF2A mRNA contains two target sites for miR-206 binding and confirmed that miR-206 directly suppresses KIF2A; inhibits ovarian cancer cell proliferation, migration, and invasion; and induces apoptosis. CONCLUSION: The results suggest KIF2A could serve a valuable prognostic indicator in ovarian cancer and provide a rationale for treatment of ovarian cancer by targeting KIF2A via miR-206.

Differentiation of human umbilical cord Wharton's jelly-derived mesenchymal stem cells into endometrial cells.

BACKGROUND: Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) are a novel and promising strategy for tissue engineering because of their ability to differentiate into many cell types. We characterized the differentiation of WJ-MSCs into endometrial epithelial cell (EEC)-like and endometrial stromal cell (ESC)-like cells and assessed the effect of 17ß-estradiol and 8-Br-cAMP on the differentiation system. METHODS: WJ-MSCs were treated in two ways to differentiate into EEC-like and ESC-like cells respectively: cocultured with ESCs in control/differentiation medium (17ß-estradiol, growth factors); and cultured in control/differentiation medium (8-Br-cAMP alone or 8-Br-cAMP plus 17ß-estrogen and growth factors). Three signaling pathway inhibitors (SB203580, PD98059, H89) were used to investigate the mechanism of WJ-MSC differentiation into ESC-like cells. Immunofluorescence, western blot and flow cytometry analyses were used to analyze expression of epithelial markers and stromal cell markers. Enzyme-linked immunosorbent assays were used to test the production of secretory proteins associated with the differentiation of ESC-like cells. RESULTS: 17ß-estradiol at 1 µM downregulated vimentin and CD13 and upregulated cytokeratin and CD9 proteins, promoting the differentiation of WJ-MSCs into EEC-like cells in the coculture system. 8-Br-cAMP at 0.5 mM upregulated vimentin and CD13 and downregulated CK and CD9, promoting the differentiation of WJ-MSCs into ESC-like cells. Prolactin (PRL) and insulin-like growth factor-binding protein 1 (IGFBP1) were upregulated and the protein kinase A (PKA) signaling pathway was activated, whereas extracellular signal-regulated (ERK)1/2 and p38 mitogen-activated protein kinase (MAPK) were not affected. CONCLUSIONS: 17ß-estradiol at 1 µM is a good inducer for facilitating the differentiation of WJ-MSCs into EEC-like cells. 8-Br-cAMP plus estrogen and growth factors can induce the differentiation of WJ-MSCs into ESC-like cells. During the differentiation of WJ-MSCs into ESC-like cells, PRL and IGFBP1 were upregulated by the treatment and the PKA signaling pathway was activated, whereas ERK1/2 and p38 MAPK were not affected. These findings suggest a promising approach to the treatment of endometrial damage and other endometrial diseases and suggest new applications for WJ-MSCs in clinical practice.

CD200Fc reduces LPS-induced IL-1ß activation in human cervical cancer cells by modulating TLR4-NF-κB and NLRP3 inflammasome pathway.

Chronic inflammation plays an important role in tumorigenesis of cervical cancer. CD200Fc, a CD200R1 agonist, has been found to have anti-inflammatory effects in autoimmune diseases and neuro-degeneration. However, the anti-inflammatory effect of CD200Fc on cervical cancer has not yet to be completely understood. This study investigated the anti-inflammatory effects and mechanisms of CD200Fc in LPS-induced human SiHa cells and Caski cells. SiHa cells and Caski cells were stimulated with 40 µg/ml LPS under different concentrations of CD200Fc for 90 min or 12 hours. The mRNA and protein levels of pro-IL-1ß, cleaved-IL-1ß and NLRP3, as well as the protein level of cleaved caspase-1, were significantly increased in LPS-induced SiHa cells and Caski cells. LPS stimulation did not change ASC and pro-caspase-1 expression. CD200Fc down-regulated protein expression of cleaved caspase-1 and mRNA and protein expression of pro-IL-1ß, cleaved-IL-1ß and NLRP3. In addition, the protein levels of TLR4, p-P65 and p-IκB, as well as the translocation of P65 to nucleus, were significantly increased in LPS-induced SiHa cells and Caski cells. LPS stimulation did not change t-P65 and t-IκB on protein levels, which were components of TLR-NF-κB pathway. CD200Fc down-regulated protein expression of TLR4, p-P65 and p-IκB and inhibited the translocation of P65 to nucleus in LPS-induced SiHa cells and Caski cells. These results indicated that CD200Fc appeared to suppress the inflammatory activity of TLR4-NF-κB and NLRP3 inflammasome pathway in LPS-induced SiHa cells and Caski cells. It provided novel mechanistic insights into the potential therapeutic uses of CD200Fc for cervical cancer.

LAMP3 expression correlated with poor clinical outcome in human ovarian cancer.

Lysosome-associated membrane protein 3 belongs to the lysosome-associated membrane glycoprotein family, which is associated with lymph node, metastasis, poor overall survival, and resistance to chemotherapy and radiotherapy. Epithelial ovarian cancer is one of the most deadly global female gynecologic malignant tumors. Its clinical outcome is poor and most epithelial ovarian cancer patients tend to relapse because of drug resistance. However, lysosome-associated membrane protein 3 expression in epithelial ovarian cancer and its relationship between clinicopathologic factors remain poorly understood. To clarify the prognostic implications of lysosome-associated membrane protein 3 in epithelial ovarian cancer, we analyzed both messenger RNA and protein levels of lysosome-associated membrane protein 3 in ovarian carcinomas. Polymerase chain reaction results showed higher expression of lysosome-associated membrane protein 3 messenger RNA in epithelial ovarian cancer than in noncancerous tissues. Immunohistochemical results showed that high lysosome-associated membrane protein 3 cytoplasmic expression was significantly related to tumor grade ( p = 0.038), lymph node metastasis ( p = 0.049), metastasis ( p < 0.001), level of CA125 ( p = 0.030), and International Federation of Gynecology and Obstetrics (FIGO) ( p < 0.001). High lysosome-associated membrane protein 3 nuclear expression was significantly associated with tumor grade ( p = 0.046), tumor single or double (representative whether the tumor involving one or both ovaries) ( p = 0.016), metastasis ( p < 0.001), and FIGO stage ( p < 0.001). Survival analysis indicated that high lysosome-associated membrane protein 3 cytoplasmic expression (hazard ratio: 4.632, 95% confidence interval: 2.421-8.864; p < 0.001), patients' age (hazard ratio: 1.729, 95% confidence interval: 1.027-2.911; p = 0.039), and FIGO stage (hazard ratio: 2.049, 95% confidence interval: 1.113-3.774; p = 0.021) were significantly correlated with poor survival outcome of epithelial ovarian cancer patients.