Structure-guided mutagenesis of Henipavirus receptor-binding proteins reveals molecular determinants of receptor usage and antibody-binding epitopes.

Nipah virus (NiV) is a highly lethal, zoonotic Henipavirus (HNV) that causes respiratory and neurological signs and symptoms in humans. Similar to other paramyxoviruses, HNVs mediate entry into host cells through the concerted actions of two surface glycoproteins: a receptor-binding protein (RBP) that mediates attachment and a fusion glycoprotein (F) that triggers fusion in an RBP-dependent manner. NiV uses ephrin-B2 (EFNB2) and ephrin-B3 (EFNB3) as entry receptors. Ghana virus (GhV), a novel HNV identified in a Ghanaian bat, uses EFNB2 but not EFNB3. In this study, we employ a structure-informed approach to identify receptor-interfacing residues and systematically introduce GhV-RBP residues into a NiV-RBP backbone to uncover the molecular determinants of EFNB3 usage. We reveal two regions that severely impair EFNB3 binding by NiV-RBP and EFNB3-mediated entry by NiV pseudotyped viral particles. Further analyses uncovered two-point mutations (NiVN557SGhV and NiVY581TGhV) pivotal for this phenotype. Moreover, we identify NiV interaction with Y120 of EFNB3 as important for the usage of this receptor. Beyond these EFNB3-related findings, we reveal two domains that restrict GhV binding of EFNB2, confirm the HNV-head as an immunodominant target for polyclonal and monoclonal antibodies, and describe putative epitopes for GhV- and NiV-specific monoclonal antibodies. Cumulatively, the work presented here generates useful reagents and tools that shed insight to residues important for NiV usage of EFNB3, reveal regions critical for GhV binding of EFNB2, and describe putative HNV antibody-binding epitopes. IMPORTANCE: Hendra virus and Nipah virus (NiV) are lethal, zoonotic Henipaviruses (HNVs) that cause respiratory and neurological clinical features in humans. Since their initial outbreaks in the 1990s, several novel HNVs have been discovered worldwide, including Ghana virus. Additionally, there is serological evidence of zoonotic transmission, lending way to concerns about future outbreaks. HNV infection of cells is mediated by the receptor-binding protein (RBP) and the Fusion protein (F). The work presented here identifies NiV RBP amino acids important for the usage of ephrin-B3 (EFNB3), a receptor highly expressed in neurons and predicted to be important for neurological clinical features caused by NiV. This study also characterizes epitopes recognized by antibodies against divergent HNV RBPs. Together, this sheds insight to amino acids critical for HNV receptor usage and antibody binding, which is valuable for future studies investigating determinants of viral pathogenesis and developing antibody therapies.

Sequence basis for selectivity of ephrin-B2 ligand for Eph receptors and pathogenic henipavirus G glycoproteins.

IMPORTANCE: Ephrin-B2 (EFNB2) is a ligand for six Eph receptors in humans and regulates multiple cell developmental and signaling processes. It also functions as the cell entry receptor for Nipah virus and Hendra virus, zoonotic viruses that can cause respiratory and/or neurological symptoms in humans with high mortality. Here, we investigate the sequence basis of EFNB2 specificity for binding the Nipah virus attachment G glycoprotein over Eph receptors. We then use this information to engineer EFNB2 as a soluble decoy receptor that specifically binds the attachment glycoproteins of the Nipah virus and other related henipaviruses to neutralize infection. These findings further mechanistic understanding of protein selectivity and may facilitate the development of diagnostics or therapeutics against henipavirus infection.

Ephrin B2 (EFNB2) potentially protects against intervertebral disc degeneration through inhibiting nucleus pulposus cell apoptosis.

Nucleus pulposus (NP) cell apoptosis is a significant indication of accelerated intervertebral disc degeneration; however, the precise mechanism is unelucidated as of yet. Ephrin B2 (EFNB2), the only gene down-regulated in the three degraded intervertebral disc tissue microarray groups (GSE70362, GSE147383 and GSE56081), was screened for examination in this study. Subsequently, EFNB2 was verified to be down-regulated in degraded NP tissue samples. Interleukin-1 (IL-1ß) treatment of NP cells to simulate the IDD environment indicated that IL-1ß treatment decreased EFNB2 expression. In degenerative NP cells stimulated by IL-1ß, EFNB2 knockdown significantly increased the rate of apoptosis as well as the apoptosis-related molecules cleaved-caspase-3 and the Bax to Bcl-2 ratio. EFNB2 was found to promote AKT, PI3K, and mTOR phosphorylation; the PI3K/AKT signaling role was investigated using the PI3K inhibitor LY294002. EFNB2 overexpression significantly increased PI3K/AKT pathway activity in IL-1ß-stimulated NP cells than the normal control. Moreover, EFNB2 partially alleviated NP cell apoptosis induced by IL-1ß, reduced the cleaved-cas3 level, and decreased the Bax/Bcl-2 ratio after the addition of the inhibitor LY294002. Additionally, EFNB2 overexpression inhibited the ERK1/2 phosphorylation; the effects of EFNB2 overexpression on ERK1/2 phosphorylation, degenerative NP cell viability, and cell apoptosis were partially reversed by ERK signaling activator Ceramide C6. EFNB2 comprehensively inhibited the apoptosis of NP cells by activating the PI3K/AKT signaling and inhibiting the ERK signaling, obviating the exacerbation of IDD. EFNB2 could be a potential target to protect against degenerative disc changes.

Unveiling EFNB2 as a Key Player in Sorafenib Resistance: Insights from Bioinformatics Analysis and Functional Validation in Hepatocellular Carcinoma.

Sorafenib resistance has become a big hurdle for treating advanced HCC; thus, identifying novel targets to overcome sorafenib resistance is of great importance. Thanks to the massive progress in the sequencing and data analysis, high-throughput screening of novel targets in HCC development has been extensively used in recent years. In present study, we harnessed the public dataset and aimed to identify novel targets related to sorafenib resistance in HCC via bioinformatics analysis and in vitro validation. This study examined three GEO datasets (GSE140202, GSE143233, GSE182593) and identified 20 common DEGs. Functional enrichment analysis suggested these DEGs might play a role in regulating drug resistance pathways. PPI network analysis pinpointed 14 hub genes, with EFNB2 showing high connectivity to other genes. Subsequent in vitro experiments demonstrated that EFNB2 was up-regulated in sorafenib-resistant HCC cells. EFNB2 suppression sensitized HepG2 and Huh7 sorafenib-resistant cells. Furthermore, EFNB2 knockdown increased caspase-3/-7 activities and hindered EMT in sorafenib-resistant HCC cells. Conversely, EFNB2 overexpression promoted sorafenib resistance, decreased caspase-3/-7 activity, and enhanced EMT in HCC cells. Overall, this study identified 14 promising genes potentially linked to sorafenib resistance in HCC, with EFNB2 emerging as a potential contributor to this resistance mechanism.

LncRNA BBOX1-AS1 Contributes to Laryngeal Carcinoma Progression by Recruiting SRSF1 to Maintain EFNB2 mRNA Stability.

Laryngeal cancer is a common malignancy of the larynx with a generally poor prognosis. This study systematically assessed the functional role of lncRNA BBOX1-AS1 in laryngeal carcinoma progression and associated molecular regulatory mechanisms. The proliferation, migration, and invasion of laryngeal carcinoma cells were detected by Cell Counting Kit-8, wound healing, clonal formation, and transwell assays. In addition, the interaction between BBOX1-AS1, Serine/Arginine Splicing Factor 1 (SRSF1), and Ephrin-B2 (EFNB2) mRNA was examined employing RNA immunoprecipitation and RNA pull-down experiments. Furthermore, western blotting, and RT-qPCR assays were adopted to detect the expression levels of BBOX1-AS1, SRSF1, and EFNB2. The impact of BBOX1-AS1 and SRSF1 on EFNB2 mRNA stability was examined using the RNA stability assay. BBOX1-AS1 was highly expressed in human laryngeal carcinoma tissues and cell lines. BBOX1-AS1 knockdown suppressed the growth, proliferation, migration, and invasion of laryngeal carcinoma cells. BBOX1-AS1 maintained the stability of EFNB2 mRNA in laryngeal carcinoma cells by recruiting SRSF1. EFNB2 knockdown inhibited the growth and metastatic function of laryngeal carcinoma cells in vitro. EFNB2 overexpression reversed the influence of BBOX1-AS1 knockdown on laryngeal cancer tumorigenesis. BBOX1-AS1 maintained EFNB2 mRNA stability by recruiting SRSF1, thereby aggravating laryngeal carcinoma malignant phenotypes. BBOX1-AS1 might be a new theoretical target for the treatment of laryngeal carcinoma.

EFNB2 haploinsufficiency causes a syndromic neurodevelopmental disorder.

Ephrin B2, one of the ligand of the EphB receptors, is involved in a complex signaling pathway regulating the development of the nervous system, neuronal migration, erythropoiesis and vasculogenesis. We report a patient with a de novo variant in EFNB2 and a family in which segregates a 610-kb deletion at chromosome 13q33 encompassing only ARGLU1 and EFNB2 genes. The de novo variant was observed in a patient with anal stenosis, hypoplastic left ventricle and mild developmental delay. The deletion was identified in 2 sibs with congenital heart defect and mild developmental delay. One of the affected sibs further had myoclonic epilepsy and bilateral sensorineural hearing loss. The carrier mother was apparently asymptomatic. Because EFNB2 is located in the subtelomeric region of 13q chromosome, we reviewed the previous reports of terminal 13q deletion. We suggest that haploinsufficiency of the EFNB2 could be at the origin of several clinical features reported in 13qter deletions, including intellectual disability, seizures, congenital heart defects, anorectal malformation and hearing loss.

Conserved role of Sonic Hedgehog in tonotopic organization of the avian basilar papilla and mammalian cochlea.

Sound frequency discrimination begins at the organ of Corti in mammals and the basilar papilla in birds. Both of these hearing organs are tonotopically organized such that sensory hair cells at the basal (proximal) end respond to high frequency sound, whereas their counterparts at the apex (distal) respond to low frequencies. Sonic hedgehog (Shh) secreted by the developing notochord and floor plate is required for cochlear formation in both species. In mice, the apical region of the developing cochlea, closer to the ventral midline source of Shh, requires higher levels of Shh signaling than the basal cochlea farther away from the midline. Here, gain-of-function experiments using Shh-soaked beads in ovo or a mouse model expressing constitutively activated Smoothened (transducer of Shh signaling) show up-regulation of apical genes in the basal cochlea, even though these regionally expressed genes are not necessarily conserved between the two species. In chicken, these altered gene expression patterns precede morphological and physiological changes in sensory hair cells that are typically associated with tonotopy such as the total number of stereocilia per hair cell and gene expression of an inward rectifier potassium channel, IRK1, which is a bona fide feature of apical hair cells in the basilar papilla. Furthermore, our results suggest that this conserved role of Shh in establishing cochlear tonotopy is initiated early in development by Shh emanating from the notochord and floor plate.

Up-regulation of miR-582-5p regulates cellular proliferation of prostate cancer cells under androgen-deprived conditions.

BACKGROUND: MicroRNAs are noncoding small RNA that negatively regulate target gene expression by binding to the 3'-UTR of mRNA. Previous studies have shown that several microRNAs play a pivotal role in prostate cancer by acting as oncogenes or tumor suppressors. This study was aimed at identifying microRNAs that contribute to the progression to castration resistant prostate cancer. METHODS: MicroRNAs expression profiles of a xenograft model and cell lines were examined by microarray analysis and real-time PCR. Functional analysis of miR-582-5p in cellular proliferation was examined by cell counting. Furthermore, in order to investigate a candidate target of miR-582-5p, microarray analysis and analysis in silico were utilized. RESULTS: MiR-582-5p was identified to be up-regulated at the castration resistant stage of a xenograft model, KUCaP2 and in castration resistant cell line, AILNCaP#1. Overexpression of miR-582-5p increased the number and the percentage of S phase of LNCaP cells under androgen deprived condition. Moreover, suppression of miR-582-5p decreased the number and the percentage of S phase of AILNCaP#1 cells. Furthermore, we identified that miR-582-5p down-regulates EFNB2 expression, which is down-regulated at the castration resistant stage of a xenograft model, KUCaP2 and in castration resistant cell line, AILNCaP#1. CONCLUSIONS: Our results suggest that up-regulation of miR-582-5p contributes to an increase in the proliferation of prostate cancer cells under androgen deprived conditions.

Tetrasomy 13q32.2qter due to an apparent inverted duplicated neocentric marker chromosome in an infant with hemangiomas, failure to thrive, laryngomalacia, and tethered cord.

BACKGROUND: Approximately 100 small supernumerary marker chromosomes (sSMCs) with a non-α-satellite neocentromere structure have been reported in the literature. Of the few derived from chromosome 13, five have consisted of inverted duplicated segment 13q32qter. CASE REPORT: We herein describe the sixth case, characterized by genome wide SNP array, conventional cytogenetics and FISH studies. The de novo occurrence of the marker, the poor prognosis and the presence of hemangiomas are consistent with previous cases. CONCLUSION: We hereby expand the clinical spectrum of this rare cytogenetic disorder and suggest a possible mechanism for the pathogenesis of associated congenital vascular malformations.

Theranostic Potential of EFNB2 for Cetuximab Resistance in Head and Neck Cancer.

Only 13% of head and neck cancer (HNC) patients respond to cetuximab therapy despite its target (EGFR) is expressed in about 80-90% of HNC patients. However, this problem remained unresolved till date despite of numerous efforts. Thus, the current study aimed to establish hub genes involved in cetuximab resistance via series of bioinformatics approach. The GSE21483 dataset was analysed for differentially expressed genes (DEGs) using GEO2R and enrichment analysis was carried out using DAVID. STRING 11.5 and Cytoscape 3.7.2 were used for protein-protein interactions and hub genes respectively. The significant hub genes (p < 0.05) were validated using ULCAN and Human protein atlas. Validated genes were further queried for tumor infiltration using TIMER2.0. Out of total 307 DEGs, 38 hub genes were identified of which IL1A, EFNB2, SPRR1A, ROBO1 and SOCS3 were the significant hub genes associated with both mRNA expression and overall survival. IL1A, ROBO1, and SOCS3 were found to be downregulated whereas EFNB2 and SPRR1A were found to be upregulated in our study. However, using UALCAN, we found that high expression of IL1A, EFNB2, SOCS3 negatively affects overall survival whereas high expression of SPRR1A and ROBO1 positively affects overall survival. Protein level for EFNB2 and SPRR1A expression was significant in tumor HNC tissue as compared to normal HNC tissue. EFNB2 was found to be a key regulator of CTX resistance among HNC patients. Targeting EFNB2 and associated PPI circuits might improve the response rate to CTX. Thus, EFNB2 has potential to be theranostic marker for CTX resistance. Supplementary Information: The online version contains supplementary material available at 10.1007/s12070-023-03739-9.

Elevated ITGA5 facilitates hyperactivated mTORC1-mediated progression of laryngeal squamous cell carcinoma via upregulation of EFNB2.

Background: Laryngeal squamous cell carcinoma (LSCC) is one of the most common malignant tumors of the head and neck, and it has shown increasing incidence and mortality. The mechanistic target of rapamycin complex 1 (mTORC1) is frequently dysregulated in LSCC, but its underlying mechanisms remain unclear. Methods: Establishment of a novel LSCC cell line using primary LSCC tumor tissues with dysregulated mTORC1 activity and then stable knockdown of Raptor (an mTORC1 specific component) in this cell line. Transcriptomic sequencing, quantitative real-time PCR, western blot analysis, and immunofluorescence assays were used to identify the crucial downstream effector of mTORC1. A series of experiments were conducted to investigate the functions and underlying mechanisms of the mTORC1 target gene in LSCC progression. Clinical LSCC samples were used to evaluate the association of mTORC1 and its downstream targets with clinicopathological features and patient prognosis. Finally, the influence on cisplatin (CDDP) sensitivity upon depletion of the mTORC1 target gene was assessed using a cell culture system, a cell line-derived xenograft (CDX) model, and a patient-derived xenograft (PDX) model. Results: We successfully established a novel LSCC cell line with hyperactivated mTORC1 activity and then identified integrin subunit alpha 5 (ITGA5) as a novel functional downstream effector of mTORC1 in the progression of LSCC. Elevated ITGA5 promotes LSCC progression through augmentation of ephrin-B2 (EFNB2). Clinical data analysis indicated that the activation of the mTORC1-ITGA5-EFNB2 signaling pathway is associated with malignant progression and poor prognosis of LSCC patients. Inhibition of ITGA5 significantly sensitized LSCC cells to CDDP. Conclusions: Our findings highlight a novel molecular mechanism for the tumorigenesis driven by deregulated mTORC1 signaling in LSCC, suggesting that the ITGA5-EFNB2 axis may be a therapeutic target for the treatment of mTORC1-related LSCC.

Corrigendum: miR-193a-3p Mediates Placenta Accreta Spectrum Development by Targeting EFNB2 via Epithelial-Mesenchymal Transition Pathway Under Decidua Defect Conditions.

[This corrects the article DOI: 10.3389/fmolb.2020.613802.].

Glycoprotein attachment with host cell surface receptor ephrin B2 and B3 in mediating entry of nipah and hendra virus: a computational investigation.

Nipah virus (NiV) and Hendra virus (HeV) are highly pathogenic paramyxovirus which belongs to Henipavirus family, causes severe respiratory disease, and may lead to fatal encephalitis infections in humans. NiV and HeV glycoproteins (G) bind to the highly conserved human ephrin-B2 and B3 (EFNB2 & EFNB3) cell surface proteins to mediate the viral entry. In this study, various molecular modelling approaches were employed to understand protein-protein interaction (PPI) of NiV and HeV glycoprotein (84% sequence similarity) with Human EFN (B2 and B3) to investigate the molecular mechanism of interaction at atomic level. Our computational study emphasized the PPI profile of both the viral glycoproteins with EFN (B2 and B3) in terms of non-bonded contacts, H-bonds, salt bridges, and identification of interface hotspot residues which play a critical role in the formation of complexes that mediate viral fusion and entry into the host cell. According to the reports, EFNB2 is considered to be more actively involved in the attachment with the NiV and HeV glycoprotein; interestingly the current computational study has displayed more conformational stability in HeV/NiV glycoprotein with EFNB2 complex with relatively high binding energy as compared to EFNB3. During the MD simulation, the number of H-bond formations was observed to be less in the case of EFNB3 complexes, which may be the possible reason for less conformational stability in the EFNB3 complexes. The current detailed interaction study on the PPI may put a path forward in designing peptide inhibitors to obstruct the interaction of viral glycoproteins with host proteins, thereby inhibiting viral entry. Graphical abstract: The viral attachment and fusion of Nipah and Hendra virus was explored through the interaction between viral glycoprotein and the host cell surface ephrin protein. The MD simulation results displayed more stability in Nipah and Hendra glycoprotein with EFNB2 as compared to EFNB3. The residue Glu533 in the central cavity of HeV/NiV glycoprotein protein identified as the potential hotspot in binding with the G-H loop of EFNB2. Supplementary Information: The online version contains supplementary material available at 10.1007/s12039-022-02110-9.

Ephrin B Activate Src Family Kinases in Fibroblasts Inducing Stromal Remodeling in Prostate Cancer.

Through stromal-epithelial interactions, carcinoma associated fibroblasts (CAF) play a critical role in tumor growth and progression. Activation of erythrophoyetin-producing human hepatocellular (Eph) receptors has been implicated in cancer. Eph receptor interactions with Ephrin ligands lead to bidirectional signals in the recipient and effector cells. The consequences of continuous reverse Ephrin signaling activation in fibroblasts on prostate cancer (PCa) is unknown. When compared to benign prostate fibroblast, CAF displayed higher expression of Ephrin B1, B2, and B3 ligands (EFNB1, EFNB2, and EFNB3). In this study, we found that continuous activation of EFNB1 and EFNB3 in a benign human prostate stromal cell line (BHPrS1) increased the expression of CAF markers and induced a CAF phenotype. BHPrS1EFNB1 and BHPrS1EFNB3 displayed a pro-tumorigenic secretome with multiple effects on neovascularization, collagen deposition, and cancer cell proliferation, overall increasing tumorigenicity of a premalignant prostate epithelial cell line BPH1 and PCa cell line LNCaP, both in vitro and in vivo. Inhibition of Src family kinases (SFK) in BHPrS1EFNB1 and BHPrS1EFNB3 suppressed EFNB-induced ɑ-SMA (Alpha-smooth muscle actin) and TN-C (Tenascin-C) in vitro. Our study suggests that acquisition of CAF characteristics via SFK activation in response to increased EFNB ligands could promote carcinogenesis via modulation of TME in PCa.

RNA-seq co-expression network analysis reveals anxiolytic behavior of mice with Efnb2 knockout in parvalbumin+ neurons.

Anxiety disorders are the most common psychiatric disorders, and the change in the activity of the prefrontal cortex (PFC) is considered as the underlying pathological mechanism. Parvalbumin-expressing (PV+) inhibition contributes to the overall activity of the PFC. However, the molecular mechanism underlying the excitation-inhibition imbalance of PV+ neurons in the PFC is unknown. Efnb2 is a membrane-bound molecule that plays an important role in the nervous system through binding the Eph receptor. To investigate whether the loss of Efnb2 in PV+ affects anxiety, we examined the behavior of wild type and Efnb2 in PV+ neurons knockout (KO) mice. We monitored the defensive responses to aversive stimuli of elevated plus maze (EPM) and found that KO mice exhibited obvious fearless and anxiolytic behaviors. To further investigate the underlying regulatory mechanism, we performed RNA sequencing, analyzed the differentially expressed genes (DEGs), and constructed the weighted gene co-expression network analysis (WGCNA). The WGCNA identified 12 characteristic modules. Among them, the MEgreen module showed the most significant correlation with KO mice of EPM stimuli. The Gene Ontology enrichment and the Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that this was related to the distal axon, Ras signaling pathway and insulin signaling pathway. Furthermore, the whole-cell voltage clamp recordings also proved that Efnb2 gene knock-out could affect synaptic function. Together with the transcriptomic analysis of mice with Efnb2 knockout on PV+ neurons, our findings suggest that Efnb2 gene in the PV+ neuron of PFC may be a crucial factor for fear and anxiety, which provide an insight into anxiety pathophysiology.

Efnb2 haploinsufficiency induces early gap junction plaque disassembly and endocytosis in the cochlea.

Chromosome 13q deletions encompassing EFNB2, which encodes the transmembrane protein ephrin-B2, are likely to cause syndromic forms of sensorineural hearing loss of unclear origin. Thus, unravelling the pathogenic mechanisms could help to improve therapeutic strategies. In the cochlea, adjacent non-sensory epithelial cells are connected via gap junction channels, the activity of which is critical to maintain cochlear homeostasis. Here we show that ephrin-B2 promotes the assembly of connexin 30 (Cx30) gap junction plaques (GJPs) between adjacent non-sensory Deiters' cells. An in situ proximity ligation assay revealed that ephrin-B2 preferentially interacts with Cx30 in the periphery of the GJPs, i.e. where newly synthesized connexin hemichannels accrue to the GJP. Moreover, we observed that heterozygous mice encoding an Efnb2 null allele display excessive clathrin-mediated internalization of Cx30 GJPs in early postnatal stages. Finally, an in vitro organotypic assay revealed that ectopic activation of ephrin-B2 reverse signalling promotes the internalization of Cx30 GJPs. These data argue in favor of a cell-autonomous, Eph receptor-independent role of ephrin-B2 in the assembly of Cx30 GJPs. According to recent observations, early GJP degradation could certainly play a role in the pathogenic process leading to progressive sensorineural hearing loss due to Efnb2/EFNB2 haploinsufficiency.

Expression levels of EPHB4, EFNB2 and caspase-8 are associated with clinicopathological features and progression of esophageal squamous cell cancer.

The upregulation of EPH receptor B4 (EPHB4) results in a survival advantage for tumor cells via the inhibition of the casapse-8-mediated apoptotic pathway, which begins from the cell membrane. The present study investigated the expression patterns of EPHB4, ephrin B2 (EFNB2) and caspase-8 in patients with esophageal squamous cell carcinoma (ESCC). The association between the expression patterns and certain clinicopathological characteristics of the patients was also determined. mRNA levels of EPHB4, EFNB2 and caspase-8 in paired primary ESCC samples and adjacent esophageal tissues collected from 96 patients with ESCC were quantified using quantitative PCR. Upregulation of EPHB4 and EFNB2 mRNA expression, and downregulation of caspase-8 mRNA were detected in ESCC samples compared with that in the adjacent esophageal tissues. The expression levels of EPHB4 and EFNB2 were positively correlated with each other, whereas the mRNA levels of both EPHB4 and EFNB2 exhibited a negative correlation with that of caspase-8. The mRNA levels of both EPHB4 and EFNB2 demonstrated a significant positive association with certain clinicopathological features of patients with ESCC, including family history, tumor size, metastasis and stage. Conversely, a negative association was revealed between the expression level of caspase-8 and clinicopathological features of patients with ESCC. Moreover, mRNA expression levels of EPHB4 and EFNB2 were negatively associated with survival times of patients with ESCC, whereas the level of caspase-8 was positively associated with patient outcome. The results from the present study suggested that EPHB4, EFNB2 and caspase-8 may be implicated in the tumorigenesis and progression of ESCC, and that consequently, they may serve as useful prognostic markers, as well as potential therapeutic targets.

miR-193a-3p Mediates Placenta Accreta Spectrum Development by Targeting EFNB2 via Epithelial-Mesenchymal Transition Pathway Under Decidua Defect Conditions.

Objective: To clarify the role of microRNA-193a-3p (miR-193a-3p) in the pathogenesis of placenta accreta spectrum. Methods: The placental tissue expression levels of miR-193a-3p and Ephrin-B2 (EFNB2) were compared between a placenta accreta spectrum group and a control group. Transwell migration and invasion assays were used to verify the effect of miR-193a-3p and EFNB2 on HTR-8/SVneo cells cultured in human endometrial stromal cell (hESC)-conditioned medium. Epithelial-mesenchymal transition (EMT)-related proteins were examined by western blotting to establish whether the EMT pathway was altered in placenta accreta spectrum. To determine whether EFNB2 is a target gene of miR-193a-3p, luciferase activity assays were performed. Results: miR-193a-3p was upregulated but EFNB2 downregulated in the placenta accreta spectrum group and EFNB2 was a direct target of miR-193a-3p. Overexpression or inhibition of miR-193a-3p revealed that miR-193a-3p promoted the migration and invasion of HTR-8/SVneo cells cultured in hESC-conditioned medium. Furthermore, EMT was induced, as shown by increased N-cadherin, vimentin, MMP2, and MMP9 and decreased E-cadherin in the placenta accreta spectrum group and in HTR-8/SVneo cells transfected with miR-193a-3p mimics or si-EFNB2. The negative effect of miR-193a-3p inhibitor was reversed by co-transfection with si-EFNB2 in function studies and in analyses of EMT-related proteins in vitro. Conclusion: miR-193a-3p which upregulated in placenta accreta spectrum group increases HTR-8/SVneo cell migration and invasion by targeting EFNB2 via the EMT pathway under decidua defect conditions to lead to placenta accreta spectrum.

Ephrin-B2 inhibits Aß25-35-induced apoptosis by alleviating endoplasmic reticulum stress and promoting autophagy in HT22 cells.

Previous studies have shown that Erythropoietin-producing hepatocellular carcinoma receptor B2 (EphB2) inhibited Aß-induced neuron apoptosis and improved cognition in AD mice, but the role of which ligand Ephrin B2 (Ephrin-B2, efnB2) is not clear. The aim of this study was to investigate the effect of the efnB2-activated Eph/efn forward signaling pathway on Aß-induced HT22 hippocampal cell apoptosis using recombination mouse Ephrin B2-Fc chimera protein (efnB2-Fc). We found that non-toxic concentrations of efnB2-Fc decreased the release of lactate dehydrogenase (LDH) in HT22 cells and reduced cell apoptosis in a dose-dependent manner. Further studies revealed that efnB2-Fc alleviated Aß-induced ER stress and decreased the expression of ER-stress-related transcriptional factor C/EBP homologous protein (CHOP), binding immunoglobulin protein (Bip) and phosphorylated eukaryotic translation initiation factor 2 subunit α (p-eIF2α) in HT22 cells. These effects of efnB2-Fc were related to the inhibition of Akt/mTOR signal transduction, an increase in the level of microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ), and the activation of the autophagy pathway. Our results indicate that the efnB2-mediated forward signaling pathway may activate the autophagy pathway to alleviate the Aß-induced ER stress and apoptosis in the HT22 cell.

EFNB2 acts as the target of miR-557 to facilitate cell proliferation, migration and invasion in pancreatic ductal adenocarcinoma by bioinformatics analysis and verification.

This study aims to identify the pivotal microRNAs (miRNAs) and genes, and their potential regulatory mechanisms in pancreatic ductal adenocarcinoma (PDAC) through bioinformatics analysis and experimental verification. We comprehensively analyzed two miRNA microarray datasets (GSE32678 and GSE43796) and three gene microarray datasets (GSE28735, GSE41368 and GSE71989), which were downloaded from the Gene Expression Omnibus (GEO) database, and identified the total of 8 differentially expressed miRNAs (DEMs) and 257 differentially expressed genes (DEGs) in common. Next, a new miRNA-mRNA regulatory network was constructed by bioinformatics methods, including 7 miRNAs, 58 putative target genes and 80 interaction pairs of miRNA-mRNA. Scrutinized by OncoLnc and GEPIA, it was found that 3 of 7 miRNAs (miR-21, miR-196b and miR-203) and 20 of 58 genes (MXRA5, EPYC, ECT2, COL12A1, SLC6A14, SLC7A2, BTG2, PDK4, CTNND2, NRP2, PXDN, CD109, TGFBI, LRRN1, ITGA2, DKK1, GREM1, EFNB2, SEMA3C and NT5E) were notably associated with prognosis in patients with PDAC. Furthermore, EFNB2 was significantly upregulated in PDAC compared with normal controls from different public databases. Cellular function experiments demonstrated that EFNB2 knockdown inhibited cell proliferation, migration and invasion in SW1990 cells. Western blot and luciferase reporter assays revealed that miR-557 negatively regulated the expression of EFNB2 by directly binging its 3' UTR. In conclusion, we performed integrated analysis for multiple expression profiles, and provided novel candidate miRNAs and genes to be exploited for functional studies. In addition, our findings suggested that EFNB2 contributes to PDAC progression by acting as the target gene of miR-557. It is useful for uncovering miRNA-based treatments in PDAC.