H3K4me3-Mediated FOXJ2/SLAMF8 Axis Aggravates Thrombosis and Inflammation in ß2GPI/Anti-ß2GPI-Treated Monocytes.

Antiphospholipid syndrome (APS) is characterized by thrombus formation, poor pregnancy outcomes, and a proinflammatory response. H3K4me3-related monocytes activation are key regulators of APS pathogenesis. Therefore, H3K4me3 CUT&Tag and ATAC-seq are performed to examine the epigenetic profiles. The results indicate that the H3K4me3 signal and chromatin accessibility at the FOXJ2 promoter are enhanced in an in vitro monocyte model by stimulation with ß2GPI/anti-ß2GPI, which mimics APS, and decreases after OICR-9429 administration. Furthermore, FOXJ2 is highly expressed in patients with primary APS (PAPS) and is the highest in patients with triple-positive antiphospholipid antibodies (aPLs). Mechanistically, FOXJ2 directly binds to the SLAMF8 promoter and activates SLAMF8 transcription. SLAMF8 further interacts with TREM1 to stimulate TLR4/NF-κB signaling and prohibit autophagy. Knockdown of FOXJ2, SLAMF8, or TREM1 blocks TLR4/NF-κB and provokes autophagy, subsequently inhibiting the release of inflammatory and thrombotic indicators. A mouse model of vascular APS is established via ß2GPI intraperitoneal injection, and the results suggest that OICR-9429 administration attenuates the inflammatory response and thrombus formation by inactivating FOXJ2/SLAMF8/TREM1 signaling. These findings highlight the overexpression of H3K4me3-mediated FOXJ2 in APS, which consequently accelerates APS pathogenesis by triggering inflammation and thrombosis via boosting the SLAMF8/TREM1 axis. Therefore, OICR-9429 is a promising candidate drug for APS therapy.

Depletion of circ_0006459 protects human brain microvascular endothelial cells from oxygen-glucose deprivation-induced damage through the miR-940/FOXJ2 pathway.

BACKGROUND: Multiple circular RNAs (circRNAs) play important roles in ischemic stroke. The present study aims to reveal the role and the mechanism of circ_0006459 in ischemic stroke. METHODS: Human brain microvascular endothelial cells (HBMECs) were treated with oxygen-glucose deprivation (OGD) to mimic an in vitro ischemic stroke model. RNA expression of circ_0006459, microRNA-940 (miR-940), and forkhead box J2 (FOXJ2) was detected by quantitative real-time polymerase chain reaction. Cell proliferation was analyzed by cell counting kit-8 (CCK-8) and 5-Ethynyl-29-deoxyuridine (EdU) assays. Cell apoptotic rate was quantified by flow cytometry analysis. The protein expression of proliferating cell nuclear antigen (PCNA), clusters of differentiation 6 (CDK6), BCL2-associated x protein (Bax), B-cell lymphoma 2 (Bcl2), interleukin-1ß (IL-1ß), IL-8, IL-18 and tumor necrosis factor-α (TNF-α) was analyzed by Western blotting. The regulatory relationships among circ_0006459, miR-940, and F 《》 OXJ2 were identified by dual-luciferase reporter assay, RNA immunoprecipitation assay, and RNA pull-down assay. RESULTS: Circ_0006459 and FOXJ2 expression were significantly upregulated, whereas miR-940 expression was downregulated in HBMECs after OGD. Circ_0006459 depletion assuaged OGD-induced inhibition in cell proliferation and promotion in cell apoptosis and inflammation in HBMECs. Circ_0006459 acted as a sponge for miR-940, and miR-940 targeted FOXJ2 in HBMECs. Besides, miR-940 silencing or FOXJ2 overexpression relieved circ_0006459 knockdown-induced promotion in cell proliferation and inhibition in cell apoptosis and inflammation in OGD-induced HBMECs. Further, circ_0006459 depletion decreased FOXJ2 protein expression by interacting with miR-940. CONCLUSION: Depletion of circ_0006459 protected human brain microvascular endothelial cells from oxygen-glucose deprivation-induced damage through miR-940/FOXJ2 pathway, providing a promising therapeutic target for ischemic stroke.

FOXJ2 Inhibits the Proliferation, Migration and Epithelial-Mesenchymal Transition of Prostate Carcinoma Cells.

OBJECTIVE: Forkhead box J2 (FOXJ2) which belongs to FOX transcription factors family has been regarded as diagnostic, prognostic biomarker and therapeutic target of various cancers. The aim of this study is to investigate the role of FOXJ2 in prostate carcinoma. METHODS: Western blot and qRT-PCR were applied to detect expression of FOXJ2 in prostate carcinoma cells. MTT and colony formation assays were used to detect cell proliferation. Cell migration and invasion were assessed by wound healing and transwell assays. RESULTS: FOXJ2 was down-regulated in primary prostate carcinoma tissues compared to the normal tissues based on the TCGA database. Prostate carcinoma cells also showed lower expression of FOXJ2 than normal prostate cell line (RWPE-1). pcDNA-mediated ectopical expression of FOXJ2 increased cell viability of prostate carcinoma cells, and promoted the proliferation, migration and invasion. Over-expression of FOXJ2 enhanced protein expression of E-cadherin, reduced N-cadherin, vimentin and fibronectin in the prostate carcinoma cells. Protein expression of Notch 1, Jagged-1, and Hes 1 were up-regulated in prostate carcinoma cells by over-expression of FOXJ2. CONCLUSIONS: FOXJ2 inhibited the proliferation, migration and epithelial-mesenchymal transition (EMT) of prostate carcinoma cells through inactivation of Jagged-1/Notch-1/Hes-1 pathway.

[Impact of overexpressed FOXJ2 on mouse spermatogenesis and its action mechanism].

OBJECTIVE: To analyze the phenotype of the male reproductive system in the germline-specific conditional Foxj2 knock-in mouse model (Stra8-cre; Foxj2tg/+), identify a target gene of the transcription factor FOXJ2, and investigate the effect of the overexpression of Foxj2 on mouse spermatogenesis and its action mechanism. METHODS: Based on the Cre-loxP recombination system, we generated a germline-specific conditional Foxj2 knock-in mouse model (Stra8-cre; Foxj2tg/+). We determined male fertility by counting the number of pups per litter and the fertilization rate after intracytoplasmic sperm injection (ICSI), observed the morphology of the testes and epididymides by HE staining, examined the sperm quality by computer assisted sperm analysis (CASA), detected the expression and localization of Cx43 in the testis by RT-qPCR, Western blot and immunohistochemistry, and verified the binding site of FOXJ2 to the Cx43 promoter using ChIP-PCR and dual luciferase reporter assay. RESULTS: The number of pups per litter and fertilization rate after ICSI were lower in the Stra8-cre; Foxj2tg/+ male mice than in the controls, and so were the size and weight of the testis. HE staining exhibited obvious exfoliation of germ cells and dramatically decreased spermatocytes and spermatids in the seminiferous tubules of the Stra8-cre; Foxj2tg/+ mice. Moreover, sperm concentration in the cauda epididymides was reduced, and the transcription and expression levels of Cx43 in the testis were increased. ChIP-PCR and dual luciferase reporter assay showed direct binding of FOXJ2 to the Cx43 promoter in the testis. CONCLUSIONS: Overexpressed FOXJ2 may lead to spermatogenic failure and subfertility in Stra8-cre; Foxj2tg/+ male mice by upregulating the expression of Cx43.

Foxj2 overexpression is associated with poor prognosis, progression, and metastasis in nasopharyngeal carcinoma.

Foxj2, a novel member of Forkhead box family, has been reported to play an important role in tumorigenesis, progression, and metastasis of certain cancers. However, the expression status and effects of Foxj2 on nasopharyngeal carcinoma (NPC) progression and metastasis remain debated. In this study, we first examined the expression of Foxj2 in NPC by immunohistochemistry and Western blotting analysis. We confirmed significantly elevated expression of Foxj2 in NPC tissues and cell lines. Next, the relationships between Foxj2 expression levels and the clinicopathological factors were investigated. Its expression level correlated with T-classification (P=0.026), distant metastasis (P=0.004), and clinical stage (P=0.029). In addition, high expression of Foxj2 was associated with poor prognosis in NPC patients. The effects of Foxj2 on cell proliferation and migration were explored by RNA interference (RNAi) with CCK-8 assay, cell cycle analyses, wound healing, and transwell assay. In conclusion, our data indicate that Foxj2 upregulation promotes the progression and migration of NPC. It makes Foxj2 serve as a potential therapeutic target for the treatment of NPC.

Effects of FOXJ2 on TGF-ß1-induced epithelial-mesenchymal transition through Notch signaling pathway in non-small lung cancer.

As one member of Forkhead box transcription factors, Forkhead box J2 (FOXJ2) has been found to be involved in epithelial-mesenchymal transition (EMT) process. However, the role and mechanism of FOXJ2 in non-small cell lung cancer (NSCLC) and EMT regulation have not been fully revealed. In this paper, it was revealed that the expression of FOXJ2 was lower in NSCLC samples compared with matched peritumoral lung tissue. We demonstrated that FOXJ2 expression was down-regulated by transforming growth factor-ß1 (TGF-ß1) treated, and overexpression of FOXJ2 inhibited TGF-ß1-induced EMT. Mechanistically, knocking out the expression of FOXJ2 promoted EMT by increasing the expression of Notch1 and NICD. This study implicates the potential value of FOXJ2 as a molecular marker for NSCLC.

Expression and Its Clinical Significance of Forkhead Box J2 in Non-small Cell Lung Cancer Tissues and Paracancerous Tissues / 现代检验医学杂志

Objective To evaluate the expressions and its clinical significance of Foxj2 in non-small cell lung cancer (NSCLC) patients.Methods Fifty cancer tissues from patients who underwentsurgery were collected.Western blot and immunohisto-chemical methods were used to detect the expressions of Foxj2.Results Results of western blot showed that Foxj2 expres-sion in lung cancer tissue was significantly lower than adjacent tissues.The difference between the two groups was statisti-cally significant(P<0.05).Immunohistoehemical results showed that the number of Foxj2-positive cells in adjacent tissues was significantly higher than that in cancer tissues,and the expression of Foxj2 in non-lymphatic node metastasis group was higher than that in lymphatic node metastasis.The difference between the two groups was statistically significant (P<0.01).Conclusion The expressions and location of Foxj2 was related to the lymphatic node metastasis of lung cancer.This phenomenon indicates that Foxj2 may play an important role in the development of lung cancer.

The role of FoxJ2 in the migration of human glioma cells.

Previous studies have demonstrated that FoxJ2 (forkhead box J2) is a member of Forkhead Box transcription factors and acts as an important prognostic indicator in human breast cancer. Our study aimed to assess the expression and function in human glioma. Western blot analysis and immunohistochemistry were performed in human glioma tissues. Low FoxJ2 expression was observed in 80 samples and its level was correlated with the grade of malignancy. A strongly positive correlation was observed between FoxJ2 and E-cadherin. Overexpression of FoxJ2 increased E-cadherin expression and decreased vimentin expression. The wound healing and transwell assays showed that overexpression of FoxJ2 significantly inhibited their migration in U87 cells. Consistent with this, knockdown of FoxJ2 promoted cellular motility. In a word, FoxJ2 suppressed cell migration and invasion in glioma, which might be a potential novel molecular targeted therapy for surgery and immune treatment.

STAT6 silencing up-regulates cholesterol synthesis via miR-197/FOXJ2 axis and induces ER stress-mediated apoptosis in lung cancer cells.

MiRNAs and transcription factors have emerged as important regulators for gene expression and are known to regulate various biological processes, including cell proliferation, differentiation and apoptosis. Previously, using genome-wide expression profiling studies, we have shown an inverse relationship of STAT6 and cholesterol biosynthesis and also identified FOXJ2 binding sites in the upstream region of 3 key genes (HMGCR, HMGCS1 and IDI1) of the cholesterol synthesis pathway. Our previous study also provided clues toward the anti-apoptotic role played by STAT6. For better understanding of the cellular response and underlying signaling pathways activated by STAT6 silencing, we examined the changes in miRNome profile after the siRNA-mediated silencing of STAT6 gene in NCI-H460 cells using LNA-based miRNA microarray. Our analysis showed significant downregulation of miRNAs, let-7b and miR-197, out of which miR-197 was predicted to target FOXJ2. We here show that miR-197 not only negatively regulates FOXJ2 expression through direct binding to its respective binding site in its 3'UTR but also alters total cholesterol levels by regulating genes associated with cholesterol biosynthesis pathway. We further demonstrated that STAT6 silencing elicited ER stress-mediated apoptosis in NCI-H460 cells through C/EBP homologous protein (CHOP) induction, alteration of BH3 only proteins expression and ROS production. The apoptosis induced by STAT6 downregulation was partially reversed by NAC, the ROS scavenger. Based on the above findings, we suggest that ER stress plays a major role in STAT6-induced apoptosis.