Knockdown of LPCAT1 Repressed Hepatocellular Carcinoma Growth and Invasion by Targeting S100A11.

OBJECTIVE: To explore the function of LPCAT1 in hepatocellular carcinoma progression. METHODS: Bioinformatics analysis was utilized to the data from TCGA to explore the level of LPCAT1 between normal and tumor tissues, as well as the relationship between LPCAT1 level and tumor grade and prognosis of HCC. Subsequently, we used siRNA to silence LPCAT1 in HCC cells to detect cell proliferation, migration, and invasion ability. RESULTS: The expression of LPCAT1 was significantly increased in HCC tissues. High LPCAT1 expression was correlated with high histologic grade and poor prognosis of HCC. In addition, silencing of LPCAT1 inhibited the proliferation, migration and invasion of liver cancer cells. Moreover, LPCAT1 knockdown suppressed S100A11 and Snail both at mRNA and protein level. CONCLUSION: LPCAT1 promoted the growth, invasion and migration of HCC cells by regulating S100A11 and Snail. Therefore, LPCAT1 may serve as a potential molecular target for the diagnosis and treatment of HCC.

Leaky Gut Plays a Critical Role in the Pathophysiology of Autism in Mice by Activating the Lipopolysaccharide-Mediated Toll-Like Receptor 4-Myeloid Differentiation Factor 88-Nuclear Factor Kappa B Signaling Pathway.

Increased intestinal barrier permeability, leaky gut, has been reported in patients with autism. However, its contribution to the development of autism has not been determined. We selected dextran sulfate sodium (DSS) to disrupt and metformin to repair the intestinal barrier in BTBR T+tf/J autistic mice to test this hypothesis. DSS treatment resulted in a decreased affinity for social proximity; however, autistic behaviors in mice were improved after the administration of metformin. We found an increased affinity for social proximity/social memory and decreased repetitive and anxiety-related behaviors. The concentration of lipopolysaccharides in blood decreased after the administration of metformin. The expression levels of the key molecules in the toll-like receptor 4 (TLR4)-myeloid differentiation factor 88 (MyD88)-nuclear factor kappa B (NF-κB) pathway and their downstream inflammatory cytokines in the cerebral cortex were both repressed. Thus, "leaky gut" could be a trigger for the development of autism via activation of the lipopolysaccharide-mediated TLR4-MyD88-NF-κB pathway.

Study on the Antioxidant Effect of Tanshinone IIA on Diabetic Retinopathy and Its Mechanism Based on Integrated Pharmacology.

Aim: To explore the effect of tanshinone IIA on diabetic retinopathy (DR) and its mechanism. Methods: GeneCards and OMM databases were used to mine DR-related genes. The chemical structure of tanshinone IIA was searched by PubChem, and the potential target was predicted by PharmMapper. Cystape 3.8.2 was used to visualize and analyze the tanshinone IIA-DR protein interaction network. DAVID ver 6.8 data were used to perform enrichment analysis of the tanshinone IIA-DR protein interaction network. Then animal experiments were carried out to further explore the mechanism of tanshinone IIA in the treatment of DR. Male SD rats were intraperitoneally injected with streptozotocin to establish a diabetes model and were randomly divided into a model group, a low-dose tanshinone IIA group and a high-dose group. Normal rats served as the control group. Hematoxylin-eosin (HE) staining was used to observe the structural changes of the retina; the SOD, GSH-Px, and MDA levels in the retina were detected by the xanthine oxidase method; the expression of VEGF, IL-1ß, IL-6, TNF-α, and caspase-3 mRNA were detected by qRT-PCR; and the Bcl-2, Bax, and VEGFA proteins were determined by the western blot. Results: A total of 213 tanshinone IIA potential targets and 223 DR-related genes were obtained. The enrichment analysis showed that tanshinone IIA may regulate hypoxia, oxidative stress, positive regulation of ERK1 and ERK2 cascade, steroid hormone-mediated signaling pathway, inflammatory response, angiogenesis, VEGF signaling pathway, apoptosis, PI3K-Akt signaling pathway, TNF signaling pathway, and biological processes and signaling pathways. The structure of the retina in the normal control group was clear, the retina in the model group was not clear, the nerve fiber layer was edema, the retinal cell layers of the tanshinone IIA low-dose group are arranged neatly, the inner and outer nuclear layers are slightly disordered, and the tanshinone IIA low-dose group was large. The structure of the mouse retina was further improved compared with the low-dose tanshinone IIA group. Compared with the model group, the retinal tissue SOD and GSH-PX of rats in the tanshinone IIA group increased, and the MDA level decreased (P < 0.05). Compared with the model group, the expression of VEGF, IL-1ß, IL-6, TNF-α, and caspase-3 mRNA in the retina of tanshinone IIA groups was significantly reduced (P < 0.01). Compared with the model group, the Bcl-2 protein in the tanshinone IIA groups increased, while the Bax and VEGFA proteins decreased (P < 0.05). Conclusion: Tanshinone IIA may improve the morphological performance of the retina of diabetic rats and inhibit DR, the mechanism of which may be anti-inflammatory, antiangiogenesis, etc.

Narciclasine ameliorated T cell mediated acute liver injury through activating AMPK pathway.

Hepatitis is closely related to cirrhosis and liver cancer, and it is vital that we develop new drugs and identify new drug targets. Traditional Chinese medicine has demonstrated excellent curative effects on liver diseases. The ingredients from Chinese herbals are important source for drug development in the treatment of hepatitis. Here, we found that narciclasine (NCS), a major component extracted from narcissus bulbs, showed hepatoprotective effect against concanavalin A (Con A) induced hepatitis. NCS treatment significantly reduced hepatocyte death, hepatic inflammatory cells infiltration, and serum cytokine levels in Con A challenged mice. We further observed that NCS directly inhibited Con A induced splenocytes proliferation and cytokine production in vitro. RNA-seq results showed that genes related to immune response were upregulated in Con A treated CD4+ T cells, which were down-regulated in the presence of NCS. Moreover, the AMPK pathway had been found activated in response to NCS treatment, suggesting a potential target for NCS targets. In conclusion, our results reveal that NCS is a powerful immunosuppressor against T cell activation, thus leading to protection against Con A induced liver injury in mice. These findings provide new insights into the use of natural products in the treatment of autoimmune hepatitis.

EBV infection-induced GPX4 promotes chemoresistance and tumor progression in nasopharyngeal carcinoma.

Epstein-Barr virus (EBV) was the first oncogenic virus identified in humans. It is primarily associated with multiple lymphoid and epithelial cancers, including nasopharyngeal carcinoma (NPC). However, its association with ferroptosis and its role in cancer therapy resistance have not been fully elucidated. Here, we show that EBV infection reduces the sensitivity of NPC cells to ferroptosis by activating the p62-Keap1-NRF2 signaling pathway in conjunction with upregulation of SLC7A11 and GPX4 expression. Knockdown of endogenous GPX4 or blockade of GPX4 using a specific inhibitor enhanced the chemosensitivity of EBV-infected NPC cells. Functional studies revealed that GPX4 knockdown suppresses the proliferation and colony formation of NPC cells. Mechanistically, GPX4 interacts with the TAK1-TAB1/TAB3 complex, regulates TAK1 kinase activity, and further activates downstream MAPK-JNK and NFκB pathways. High GPX4 expression is correlated with poor clinical outcomes in patients with NPC and other cancer types. Taken together, our findings suggest that EBV infection has important effects on redox homeostasis, revealing a previously unappreciated role for GPX4 in tumor progression. This novel mechanism provides a potential new target for the treatment of EBV-related tumors.

Nogo-B promotes invasion and metastasis of nasopharyngeal carcinoma via RhoA-SRF-MRTFA pathway.

Distant metastasis remains the major cause for treatment failure in patients with nasopharyngeal carcinoma (NPC). Thus, it is necessary to investigate the underlying regulation mechanisms and potential biomarkers for NPC metastasis. Nogo-B (neurite outgrowth inhibitor B), encoded by reticulon-4, has been shown to be associated with the progression and advanced stage of several cancer types. However, the relationship between Nogo-B and NPC remains unknown. In this study, we found that higher expression of Nogo-B was detected in NPC cells and tissues. Higher expression of Nogo-B was statistically relevant to N stage, M stage, and poor prognosis in NPC patients. Further functional investigations indicated that Nogo-B overexpression could increase the migration, invasion, and metastasis ability of NPC cells in vitro and in vivo. Mechanistically, Nogo-B promoted epithelial-mesenchymal transition (EMT) and enhanced the invasive potency by interacting directly with its receptor NgR3 in NPC. Additionally, overexpression of Nogo-B could upregulate the protein levels of p-RhoA, SRF, and MRTFA. A positive relationship was found between the expression of Nogo-B and the p-RhoA in NPC patients as well as in mouse lung xenografts. Nogo-Bhigh p-RhoAhigh expression was significantly associated with N stage, M stage, and poor prognosis in NPC patients. Notably, CCG-1423, an inhibitor of the RhoA-SRF-MRTFA pathway, could reverse the invasive potency of Nogo-B and NgR3 in NPC cell lines, and decrease the expression of N-Cadherin, indicating that CCG-1423 may be a potential target drug of NPC. Taken together, our findings reveal that Nogo-B enhances the migration and invasion potency of NPC cells via EMT by binding to its receptor NgR3 to regulate the RhoA-SRF-MRTFA pathway. These findings could provide a novel insight into understanding the metastasis mechanism and targeted therapy of advanced NPC.

Staged transverse preputial island flap urethroplasty for some proximal hypospadias with moderate-to-severe chordee.

BACKGROUND: We aimed to assess the outcome of staged transverse preputial island flap (TPIF) urethroplasty for repairing certain cases of primary proximal hypospadias with moderate-to-severe chordee in children. METHODS: Nighty-two consecutive boys who underwent either one-stage or staged TPIF urethroplasty for the repair of proximal hypospadias with moderate-to-severe chordee between August 2015 and December 2019 were evaluated retrospectively. Patients were divided into two groups: one-stage TPIF urethroplasty group (n = 44) and staged TPIF urethroplasty group (n = 48). We noted and compared the postoperative complications, including urethrocutaneous fistula, urethral diverticula, residual penile curvature, and urethral stricture in both groups. RESULTS: Both groups were followed up for 1-5 years, with an average of 3 years. No cases of residual or recurrence of penile chordee were reported in either group. In Group A, 9 patients (9/44, 20.4%) had postoperative urethrocutaneous fistula, and all patients underwent urinary fistula repair or urethroplasty. In Group B, postoperative urethrocutaneous fistula occurred in 2 cases (2/48, 4.1%), and one patient developed a urethrocutaneous fistula after the first operation, which was successfully repaired during the second operation. A urethrocutaneous fistula occurred in 1 case after completion of the second-stage operation; urethral fistula repair was performed successfully 6 months later. There were 2 cases of urethral stricture in Group A (2/44, 4.5%) and none in Group B. There were 6 cases of urethral diverticulum in Group A (6/44, 13.6%) and no cases of urethral diverticulum in Group B. The operative success rates were 61.3% and 95.8% in Group A and Group B, respectively (P < 0.001). CONCLUSIONS: Compared with one-stage TPIF urethroplasty, staged TPIF urethroplasty in the treatment of certain cases of primary proximal hypospadias with moderate-to-severe chordee resulted in fewer postoperative fistulas, urethral strictures and urethral diverticula. The staged TPIF urethroplasty procedure was effective in reducing the operation difficulty and complication rate of hypospadias, improving the curative effect of complex hypospadias and having good clinical application value.

Comparative analysis of key immune protection factors in H9N2 avian influenza viruses infected and immunized specific pathogen-free chicken.

H9N2 avian influenza viruses (AIV) continue to circulate in vaccinated chicken flocks in China, which prompted us to investigate the differential immune protection factors induced by H9N2 AIV infection and immunization for analyzing the reason of protection deficiency of H9N2 AIV inactivated vaccine. In this study, we firstly explored virus-induced optimal immune responses in chicken after H9N2 AIV infection. And, we found that H9N2 hemagglutination inhibition (HI) antibody level, antiviral interferon-stimulated genes including 2',5'-oligoadenylate synthetase-like and myxovirus resistance 1, CD8+ T cell response in peripheral blood lymphocytes (PBL) accompanied by the cytotoxicity-associated genes, including poly (ADP-ribose) polymerase and IFN-r play important roles in defending against H9N2 infection. Besides, we observed that vaccine immunization triggered the similar H9N2 HI antibody level as viral infection, the increase of CD4+ T cell percentage instead of CD8+ T cell percentage in PBL. Moreover, we further made a comparative analysis of immune-related gene expression profile in PBL and lung after H9N2 AIV infection and immunization, respectively. The results showed that vaccine immunization contributed to the up-regulation of Th2 cytokine. But the deficiency of cytotoxicity-associated genes induced by H9N2 AIV inactivated vaccine may be the potential key reason of protection deficiency. These findings provide evidence and direction for developing effective H9N2 AIV vaccines.

Autocrine INSL5 promotes tumor progression and glycolysis via activation of STAT5 signaling.

Metabolic reprogramming plays important roles in development and progression of nasopharyngeal carcinoma (NPC), but the underlying mechanism has not been completely defined. In this work, we found INSL5 was elevated in NPC tumor tissue and the plasma of NPC patients. Plasma INSL5 could serve as a novel diagnostic marker for NPC, especially for serum VCA-IgA-negative patients. Moreover, higher plasma INSL5 level was associated with poor disease outcome. Functionally, INSL5 overexpression increased, whereas knockdown of its receptor GPCR142 or inhibition of INSL5 reduced cell proliferation, colony formation, and cell invasion in vitro and tumorigenicity in vivo. Mechanistically, INSL5 enhanced phosphorylation and nuclear translocation of STAT5 and promoted glycolytic gene expression, leading to induced glycolysis in cancer cells. Pharmaceutical inhibition of glycolysis by 2-DG or blockade of INSL5 by a neutralizing antibody reversed INSL5-induced proliferation and invasion, indicating that INSL5 can be a potential therapeutic target in NPC. In conclusion, INSL5 enhances NPC progression by regulating cancer cell metabolic reprogramming and is a potential diagnostic and prognostic marker as well as a therapeutic target for NPC.

EphA2 phosphorylates NLRP3 and inhibits inflammasomes in airway epithelial cells.

Inflammasomes are intracellular complexes that form in the cytosol of inflammatory cells. NLRP3 is one of the sensor proteins in the complex that can recognize a wide variety of stimuli ranging from microbial components to environmental particulates. Here, we report that in mouse airway epithelial cells (AECs), inflammasome activation is inhibited by EphA2, a member of the transmembrane tyrosine kinase receptor family, via tyrosine phosphorylation of NLRP3 in a model of reovirus infection. We find that EphA2 depletion markedly enhances interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) production in response to the virus. EphA2-/- mice show stronger inflammatory infiltration and enhanced inflammasome activation upon viral infection, and aggravated asthma symptoms upon ovalbumin (ova) induction. Mechanistically, EphA2 binds to NLRP3 and induces its phosphorylation at Tyr132, thereby interfering with ASC speck formation and blocking the activation of the NLRP3-inflammasome. These data demonstrate that reovirus employs EphA2 to suppress inflammasome activation in AECs and that EphA2 deficiency causes a pathological exacerbation of asthma in an ova-induced asthma model.

Systematic Identification of Host Immune Key Factors Influencing Viral Infection in PBL of ALV-J Infected SPF Chicken.

Although research related to avian leukosis virus subgroup J (ALV-J) has lasted for more than a century, the systematic identification of host immune key factors against ALV-J infection has not been reported. In this study, we establish an infection model in which four-week-old SPF chickens are infected with ALV-J strain CHN06, after which the host immune response is detected. We found that the expression of two antiviral interferon-stimulated genes (ISGs) (Mx1 and IFIT5) were increased in ALV-J infected peripheral blood lymphocytes (PBL). A significant CD8+ T cell response induced by ALV-J appeared as early as seven days post-infection (DPI), and humoral immunity starting from 21 DPI differed greatly in the time scale of induction level. Meanwhile, the ALV-J viremia was significantly decreased before antibody production at 14 DPI, and eliminated at 21 DPI under a very low antibody level. The up-regulated CD8+ T cell in the thymus (14DPI) and PBL (7 DPI and 21 DPI) was detected, indicating that the thymus may provide the output of CD8+ T cell to PBL, which was related to virus clearance. Besides, up-regulated chemokine CXCLi1 at 7 DPI in PBL was observed, which may be related to the migration of the CD8+ T cell from the thymus to PBL. More importantly, the CD8 high+ T cell response of the CD8αß phenotype may produce granzyme K, NK lysin, or IFN-γ for clearing viruses. These findings provide novel insights and direction for developing effective ALV-J vaccines.

Knockout of LATS1 induces neoplastic phenotype in hepatic oval cells.

BACKGROUND: Primary liver cancer (PLC) is the second leading cause of cancer-related death worldwide. It has been reported that PLC can be originated from malignant transformed adult hepatic progenitor cells. Mammalian large tumor suppressor kinase 1 (LATS1) is one of the core components of the Hippo pathway and it has been implicated in regulating invasion and metastasis of different cancer cell. However, the underlying connections between hepatic progenitor cells and LATS1 in the pathogenesis of PLC are still elusive. METHODS: LATS1 gene knockout (LATS1-KO) hepatic oval cells (HOCs) were constructed by the CRISPR/Cas9 system. Cell viability was evaluated by the CCK-8 assay. Cell migration was measured by scrape assay. Cell invasion was examined by Transwell assay. Cell apoptosis was evaluated by flow cytometry. The expression of LATS1 and Yes-associated protein (YAP) in HOCs was determined by Q-PCR and Western blot analysis. RESULTS: Here, we found that knockout of LATS1 significantly induced the migration and invasion of WB-F344 cells. Knockdown of YAP suppressed the neoplastic phenotype of LATS1-KO WB-F344 cells. Furthermore, overexpression of YAP promoted the migration and invasion of LATS1-KO WB-F344 cells. CONCLUSIONS: In summary, the current study demonstrated that LATS1 is required for inhibiting the neoplastic phenotype of normal hepatic progenitor cell via downregulating YAP.

Correction to: Elevated expression of CST1 promotes breast cancer progression and predicts a poor prognosis.

In Figure 7f the panel for c-myc of MDA-MB-468 was erroneously duplicated. The corrected version of the figure is shown in this paper. This correction does not influence the conclusion of the study and we sincerely apologize for this oversight.

Plasma Macrophage Migration Inhibitory Factor and CCL3 as Potential Biomarkers for Distinguishing Patients with Nasopharyngeal Carcinoma from High-Risk Individuals Who Have Positive Epstein-Barr Virus Capsid Antigen-Specific IgA.

PURPOSE: The purpose of this study was to identify novel plasma biomarkers for distinguishing nasopharyngeal carcinoma (NPC) patients from healthy individuals who have positive Epstein-Barr virus (EBV) viral capsid antigen (VCA-IgA). MATERIALS AND METHODS: One hundred seventy-four plasma cytokines were analyzed by a Cytokine Array in eight healthy individuals with positive EBV VCA-IgA and eight patients with NPC. Real-time polymerase chain reaction, Western blotting, enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry were employed to detect the expression levels of macrophage migration inhibitory factor (MIF) and CC chemokine ligand 3 (CCL3) in NPC cell lines and tumor tissues. Plasma MIF and CCL3 were measured by ELISA in 138 NPC patients, 127 EBV VCA-IgA negative (VN) and 100 EBV VCA-IgA positive healthy donors (VP). Plasma EBV VCA-IgA was determined by immunoenzymatic techniques. RESULTS: Thirty-four of the 174 cytokines varied significantly between the VP and NPC group. Plasma MIF and CCL3 were significantly elevated in NPC patients compared with VN and VP. Combination of MIF and CCL3 could be used for the differential diagnosis of NPC from VN cohort (area under the curve [AUC], 0.913; sensitivity, 90.00%; specificity, 80.30%), and combination of MIF, CCL3, and VCA-IgA could be used for the differential diagnosis of NPC from VP cohort (AUC, 0.920; sensitivity, 90.00%; specificity, 84.00%), from (VN+VP) cohort (AUC, 0.961; sensitivity, 90.00%; specificity, 92.00%). Overexpressions of MIF and CCL3 were observed in NPC plasma, NPC cell lines and NPC tissues. CONCLUSION: Plasma MIF, CCL3, and VCA-IgA combination significantly improves the diagnostic specificity of NPC in high-risk individuals.

Identification of a sodium pump Na+/K+ ATPase α1-targeted peptide for PET imaging of breast cancer.

The sodium pump Na+/K+ ATPase a1 subunit(NKA a1), an attractive cancer-related biomarker and therapeutic target, is closely related to the development and progression of several cancers including breast cancer. Currently, a NKA a1 inhibitor, UNBS1450, has already evidenced its great therapeutic potential in personalized cancer treatment. The ability of non-invasive imaging of NKA a1 expression would be useful for selecting cancer patients who may benefit from this drug. Here, we identified an S3 peptide that is specifically homed to breast cancer by phage display. All data of in vitro and in vivo experiments suggested the excellent targeting character of the S3 peptide. As the binding activity of the S3 phage was positively correlated to the level of NKA α1 expression in various breast cancer cells, NKA α1 was validated as the primary target of the S3 peptide. Based on immunohistochemistry staining result of 107 breast cancer patients, NKA α1 was verified to be a novel tracking marker and a prognostic predictor for breast cancer. Importantly, we proposed and validated an S3 peptide-based radiotracer 18F-ALF-NOTA-S3 for PET (Positron Emission Tomography) imaging of breast cancer and other NKA α1-overexpressing cancers, including hepatocellular carcinoma and non-small cell lung cancer, in mouse models. Our findings demonstrated the potential application of 18F-ALF-NOTA-S3 for visualization of NKA α1-positive lesions, which provide a new approach to character tumor phenotypic imaging.

Author Correction: Ephrin receptor A2 is an epithelial cell receptor for Epstein-Barr virus entry.

In the version of this Letter originally published, the authors reported on the use of 2,5-dimethylpyrrolyl benzoic acid to block Ephrin receptors. In 2011, it was reported that newly synthesized 2,5-dimethylpyrrolyl benzoic acid lacked the previously reported EphA2 antagonizing activity1. However, the purchased compound did in fact have the activity initially reported, suggesting that an uncharacterized alteration occurred during storage. The authors therefore wish to clarify that the compound used in their study should be more accurately referred to as a 2,5-dimethylpyrrolyl benzoic acid derivative. All references to 2,5-dimethylpyrrolyl benzoic acid in the Letter have now been changed to reflect this.Although 2,5-dimethylpyrrolyl benzoic acid derivatives have been reported to have off-target effects2, as do most small-molecule inhibitors, the multiple complementary methods and techniques used demonstrate that EphA2 is a key Epstein-Barr virus epithelial cell receptor. The conclusions of the study are therefore unchanged.

CRLF1 promotes malignant phenotypes of papillary thyroid carcinoma by activating the MAPK/ERK and PI3K/AKT pathways.

Papillary thyroid carcinoma (PTC) is the one of the most common types of endocrine cancer and has a heterogeneous prognosis. Tumors from patients with poor prognosis may differentially express specific genes. Therefore, an analysis of The Cancer Genome Atlas (TCGA) database was performed and revealed that cytokine receptor-like factor 1 (CRLF1) may be a potential novel target for PTC treatment. The objective of the current study was to explore the expression of CRLF1 in PTC and to investigate the main functions and mechanisms of CRLF1 in PTC. PTC tissues exhibited higher CRLF1 expression at both the mRNA and protein levels than it did with normal thyroid tissues. High CRLF1 levels were associated with aggressive clinicopathological features and poor disease-free survival rates. By using loss-of-function and gain-of-function assays, we found that CRLF1 not only increased cell migration and invasion in vitro but also promoted tumor growth both in vitro and in vivo. In addition, CRLF1 induced epithelial-mesenchymal transitions. Overexpression of CRLF1 activated the ERK1/2 and AKT pathways. The oncogenic effects induced by CRLF1 were suppressed by treating the cells with the MEK inhibitor U0126 or the AKT inhibitor MK-2206. These results suggest that CRLF1 enhances cell proliferation and metastasis in PTC and thus may therefore be a potential therapeutic target for PTC.

Ephrin receptor A2 is an epithelial cell receptor for Epstein-Barr virus entry.

Epstein-Barr virus (EBV) is causally associated with nasopharyngeal carcinoma, 10% of gastric carcinoma and various B cell lymphomas 1 . EBV infects both B cells and epithelial cells 2 . Recently, we reported that epidermal growth factor and Neuropilin 1 markedly enhanced EBV entry into nasopharyngeal epithelial cells 3 . However, knowledge of how EBV infects epithelial cells remains incomplete. To understand the mechanisms through which EBV infects epithelial cells, we integrated microarray and RNA interference screen analyses and found that Ephrin receptor A2 (EphA2) is important for EBV entry into the epithelial cells. EphA2 short interfering RNA knockdown or CRISPR-Cas9 knockout markedly reduced EBV epithelial cell infection, which was mostly restored by EphA2 complementary DNA rescue. EphA2 overexpression increased epithelial cell EBV infection. Soluble EphA2 protein, antibodies against EphA2, soluble EphA2 ligand EphrinA1, or the EphA2 inhibitor 2,5-dimethylpyrrolyl benzoic acid efficiently blocked EBV epithelial cell infection. Mechanistically, EphA2 interacted with EBV entry proteins gH/gL and gB to facilitate EBV internalization and fusion. The EphA2 Ephrin-binding domain and fibronectin type III repeats domain were essential for EphA2-mediated EBV infection, while the intracellular domain was dispensable. This is distinct from Kaposi's sarcoma-associated herpesvirus infection through EphA2 4 . Taken together, our results identify EphA2 as a critical player for EBV epithelial cell entry.

Elevated expression of CST1 promotes breast cancer progression and predicts a poor prognosis.

Cystatin SN (CST1) belongs to the type 2 cystatin (CST) superfamily, which restricts the proteolytic activities of cysteine proteases. CST1 has been recently considered to be involved in the development of several human cancers. However, the prognostic significance and function of CST1 in breast cancer remains unknown. In the current study, we found that CST1 was generally upregulated in breast cancer at both mRNA and protein level. Furthermore, overall survival (OS) and disease-free survival (DFS) in the low CST1 expression subgroup were significantly superior to the high CST1 expression subgroup (OS, p < 0.001; DFS, p < 0.001), which indicated that CST1 expression level was closely correlated to the survival risk of these patients. Univariate and multivariate analyses demonstrated that CST1 expression was an independent prognostic factor, the same as ER status and nodal status. Next, CST1 overexpression promoted breast cancer cell proliferation, clonogenicity, migration, and invasion abilities. By contrast, knockdown of CST1 attenuated these malignant characteristics in breast cancer cells. Collectively, our study indicates that CST1 cannot only serve as a significant prognostic indicator but also as a potential therapeutic target for breast cancer. KEY MESSAGES: High CST1 expression is negatively correlated with survival of breast cancer patients. CST1 promotes cell proliferation, clone formation, and metastasis in breast cancer cells. CST1 is a novel potential prognostic biomarker and therapeutic target for breast cancer.