HOXB-AS1 accelerates the tumorigenesis of glioblastoma via modulation of HOBX2 and HOBX3 at transcriptional and posttranscriptional levels.

Glioblastoma (GBM) is the most universal and invasive brain tumor among adults. Increasing studies have reported that long noncoding RNAs play vital roles in regulating downstream molecules at the transcriptional or posttranscriptional level in tumor progression. The purpose of the current research was to inquire the modulation mechanism by which homeobox B cluster antisense RNA 1 (HOXB-AS1) functioned in GBM. Our study first discovered the lifted expression of HOXB-AS1 and its nearby genes HOXB2 and HOXB3 in GBM and the positive relationship between HOXB-AS1 and HOXB2 or HOXB3. Loss-of-function assays and in vivo study detected that silencing of HOXB-AS1, HOXB2, or HOXB3 restrained the proliferation and induced the apoptosis in GBM. In addition, mechanism experiments demonstrated that HOXB-AS1 recruited interleukin enhancer-binding factor 3 (ILF3) to regulate HOXB2 and HOXB3 expression at the transcriptional level, and HOXB-AS1 sponged miR-186-5p to modulate HOXB2 and HOXB3 expression at posttranscriptional level. Finally, the regulatory mechanism of HOXB-AS1 in GBM was certified through rescue experiments. Our results indicated that HOXB-AS1 boost the HOXB2 or HOXB3 expression at the transcriptional and posttranscriptional levels. We detected the HOXB-AS1-ILF3-HOXB2/HOXB3 axis and HOXB-AS1-miR-186-5p-HOXB2/HOXB3 axis driving the GBM progression, which might generate more effective diagnostic biomarkers and therapeutic targets for patients with GBM.

Long noncoding RNA HNF1A-AS1 regulates proliferation and apoptosis of glioma through activation of the JNK signaling pathway via miR-363-3p/MAP2K4.

Long noncoding RNAs (lncRNAs) have been proven to exert important functions in the various biological processes of human cancers. It has been reported that lncRNA HNF1 homeobox A antisense RNA 1 (HNF1A-AS1) was abnormally expressed and played a role in the initiation and development of various human cancers. In this study, we confirmed that the expression level of HNF1A-AS1 was increased in glioma tissues and cells. Knockdown of HNF1A-AS1 inhibited cell proliferation and promoted cell apoptosis in glioma. Then, we disclosed the downregulation of miR-363-3p in glioma tissues and cell lines. The interaction between HNF1A-AS1 and miR-363-3p was identified in glioma cells. Furthermore, an inverse correlation between HNF1A-AS1 and miR-363-3p was observed in glioma tissues. Afterwards, we recognized that MAP2K4 was a direct target of miR-363-3p. The expression of MAP2K4 was negatively correlated with miR-363-3p while positively related to HNF1A-AS1 in glioma tissues. We also found the regulatory effect of HNF1A-AS1 on the MAP2K4-dependent JNK signaling pathway. All findings indicated that HNF1A-AS1 induces the upregulation of MAP2K4 to activate the JNK signaling pathway to promote glioma cell growth by acting as a miR-363-3p sponge.

Long noncoding RNA FAM83H-AS1 exerts an oncogenic role in glioma through epigenetically silencing CDKN1A (p21).

Gliomas are the commonest and most aggressive primary malignant tumor in the central nervous system. Long noncoding RNAs (lncRNAs) have been identified to act as crucial regulators in multiple biological processes, including tumorigenesis. FAM83H antisense RNA1 (FAM83H-AS1) has been uncovered to be dysregulated in several cancers. However, the biological role of FAM83H-AS1 in glioma still needs to be investigated. Currently, our findings indicated that FAM83H-AS1 was upregulated in glioma tissues and cell lines and high level of FAM83H-AS1 was associated with poor prognosis of glioma. Loss-of-function assays demonstrated that silenced FAM83H-AS1 obviously suppressed cell proliferation via regulating the cell-cycle distribution and cell apoptosis rate, and mechanistic experiments revealed that FAM83H-AS1 could epidemically silence CDKN1A expression through recruiting EZH2 to the promoter of CDKN1A, thereby influencing the cell cycle and proliferation. Collectively, our findings suggested that FAM83H-AS1 participated in the progression of glioma and might act as a potential therapeutic target and prognosis biomarker for human glioma.

N-Glycosylation at Asn 402 Stabilizes N-Cadherin and Promotes Cell-Cell Adhesion of Glioma Cells.

Cadherin is crucial for cell-cell adhesion and N-glycosylation of N-cadherin has been implicated in the process of mammary, renal, and ovarian carcinogenesis. However, whether N-glycosylation of N-cadherin plays a role in glioma remains unknown. Previous studies had indicated that N-glycosylation could occur at three asparagine residues of N-cadherin. By generating and over-expressing N-glycosylation-deficient N-cadherin mutants in the human glioma cell lines SHG66 and U87, we found that mutation of N402 but not of the other potentially N-glycosylated residues destabilized N-cadherin and led to its ubiquitylation and subsequent proteasomal degradation. Furthermore, destabilized N-cadherin inhibited cadherin-mediated cell-cell adhesion and promoted cell migration. Our findings reveal that N-glycosylation controls N-cadherin stability and plays a role in glioma migration. J. Cell. Biochem. 118: 1423-1431, 2017. © 2016 Wiley Periodicals, Inc.

Overexpression of Wnt3a facilitates the proliferation and neural differentiation of neural stem cells in vitro and after transplantation into an injured rat retina.

Neural stem cell-based therapy is a promising option for repair after injury. However, poor stem cell proliferation and insufficient differentiation of the stem cells into neurons are still difficult problems. The present study investigated whether transplantation of neural stem cells (NSCs) genetically modified to express Wnt3a is a promising approach to overcome these difficulties. We explored the possibility that Wnt3a might contribute to the therapeutic effect of NSC transplantation in retinal repair. The relative promotion of proliferation and neural differentiation by modified NSCs was investigated in a rat model of optic nerve crush. A recombinant lentivirus (Lenti-Wnt3a) was engineered to express Wnt3a. NSCs infected with control lentivirus (Lenti-GFP) or Lenti-Wnt3a were transplanted into the subretinal space immediately after the optic nerve crush. The proliferation and neural differentiation activity of the NSCs were assessed in vitro and in vivo. Overexpression of Wnt3a in NSCs induced activation of Wnt signaling, promoted proliferation, and directed the differentiation of the NSCs into neurons both in vitro and in vivo. Our study suggests that Wnt3a can potentiate the therapeutic benefits of NSC-based therapy in the injured retina.

Glial cells activation potentially contributes to the upregulation of stromal cell-derived factor-1α after optic nerve crush in rats.

Stromal cell-derived factor-1α (SDF-1α) plays an important role after injury. However, little is known regarding its temporal and spatial expression patterns or how it interacts with glial cells after optic nerve crush injury. We characterized the temporal and spatial expression pattern of SDF-1α in the retina and optic nerve following optic nerve crush and demonstrated that SDF-1α is localized to the glial cells that are distributed in the retina and optic nerve. CXCR4, the receptor for SDF-1α, is expressed along the ganglion cell layer (GCL). The relative expression levels of Sdf-1α mRNA and SDF-1α protein in the retina and optic nerve 1, 2, 3, 5, 7, 10 and 14 days after injury were determined using real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay, respectively, and the Cxcr4 mRNA expression was determined using real-time PCR. Immunofluorescence and immunohistochemical approaches were used to detect the localization of SDF-1α and CXCR4 after injury. The upregulation of Sdf-1α and Cxcr4 mRNA was detected as early as day one after injury in the retina and day two in the optic nerve, the expression peaks 5-7 days after injury. The expression of Sdf-1α and Cxcr4 mRNA was maintained for at least 14 days after the optic nerve crush injury. Furthermore, SDF-1α-positive zones were distributed locally in the reactive glial cells, which suggested potential autocrine stimulation. CXCR4 was mainly expressed in the GCL, which was also adjacent to the the glial cells. These findings suggest that following optic nerve crush, the levels of endogenous SDF-1α and CXCR4 increase in the retina and optic nerve, where activated glial cells may act as a source of increased SDF-1α protein.

LncRNA-PVT1 indicates a poor prognosis and promotes angiogenesis via activating the HNF1B/EMT axis in glioma.

Recent studies identified that long non-coding RNAs (lncRNAs) exhibited critical roles in tumor migration and invasion. However, the roles of lncRNAs in glioma remain unclear. The aim of this study was to uncover the underlying mechanisms of glioma progression and provide potential therapeutic targets for its treatment in clinic. Our microarray study showed that lncRNA-PVT1 was significantly upregulated in glioma tissues and played an important role in cell proliferation, migration, invasion and angiogenesis. Our data showed that the expression of lncRNA-PVT1 was increased obviously and associated with advanced tumor stage, metastasis, invasion ability, and poor prognosis in glioma patients. Up-regulation of lncRNA-PVT1 was observed to promote glioma cells proliferation, and invasion abilities in vitro as well as tumor growth in vivo by regulating miR-1207-3p expression. Online software (TargetScan, miRDB and miR TarBase) were used to predict the regulating mechanisms of lncRNA-PVT1, miR-1207-3p and HNF1B, which were validated by dual-luciferase reporter gene system. In vivo tumor-bearing mice models were established to validate the cellular results. Therefore, we suggested that lncRNA-PVT1/miR-1207-3p/HNF1B axis might play critical roles in glioma progression, indicating that lncRNA-PVT1/miR-1207-3p/HNF1B signaling axis may serve as novel molecular targets for glioma prevention and treatment.

Blocking Wnt/ß-catenin Signal Amplifies Anti-PD-1 Therapeutic Efficacy by Inhibiting Tumor Growth, Migration, and Promoting Immune Infiltration in Glioblastomas.

Glioblastoma (GBM), as the immunologically cold tumor, respond poorly to programmed cell death 1 (PD-1) immune checkpoint inhibitors because of insufficient immune infiltration. Herein, through the analysis of The Cancer Genome Atlas data and clinical glioma samples, we found Wnt/ß-catenin signal was activated in GBM and inversely related to the degree of immune cell (CD8+) infiltration and programmed cell death ligand 1 (PD-L1) expression. Blockade of Wnt/ß-catenin signal could inhibit GBM U118 cells' growth and migration, and upregulate their PD-L1 expression which indicated the possible better response to anti-PD-1 immunotherapy. Besides, in a co-culture system comprising U118 cells and Jurkat cells, Wnt inhibition alleviated Jurkat cell's apoptosis and enhanced its cytotoxic function as evidenced by obviously increased effector cytokine IFNγ secretion and lactate dehydrogenase release. Moreover, the enhanced anti-GBM effect of PD-1 antibody triggered by Wnt inhibition was observed in GL261 homograft mouse model, and the upregulation of immune cell (CD4+/CD8+) infiltration and IFNγ secretion in tumor tissues suggested that Wnt/ß-catenin inhibition could inflame cold tumor and then sensitize GBM to PD-1 blockade therapy. Taken together, our study verified the blockade of Wnt/ß-catenin signal could augment the efficacy of PD-1 blockade therapy on GBM through directly inhibiting tumor proliferation and migration, as well as facilitating T-cell infiltration and PD-L1 expression in tumor microenvironment.

A novel hybrid of 3-benzyl coumarin seco-B-ring derivative and phenylsulfonylfuroxan induces apoptosis and autophagy in non-small-cell lung cancer.

BACKGROUND: Compound 6, as a novel hybrid of 3-benzyl coumarin seco-B-ring derivative and nitric oxide (NO) donor phenylsulfonylfuroxan, has the potential to develop into an anticancer drug because it displays significant antiproliferation activitity for various solid cancer cell lines including non-small-cell lung cancer (NSCLC) cells. PURPOSE: We attempt to uncover the capacities of compound 6 to induce apoptosis and autophagy in NSCLC cells, as well as the underlying mechanism involved in this process. METHODS: The effect of compound 6 on cell viability was evaluated in A549 cells by MTT assay. Apoptosis was mainly detected by flow cytometry. The induction of autophagy was observed by transmission electron microscopy (TEM), confocal microscopy as well as western-blotting technique. The expression of all related-proteins including PI3K/Akt/mTOR signaling pathway were also examined by western-blotting technique. RESULTS: Above all, distinct growth inhibition and caspase-dependent apoptosis were detected in A549 cells administered with compound 6. Then, we confirmed the induction of autophagy triggered by compound 6 in A549 cells. Noticeably, blocking autophagy using a series of inhibitors and ATG5 siRNA had little effect on the cytotoxicity of compound 6, elucidating nonprotective autophagy triggered in NSCLC cells. Further research illustrated that PI3K/Akt/mTOR signaling pathway was involved in compound 6-induced apoptosis, and 3-MA as well as LY294002 had synergistic inhibiting effect on proliferation of A549 cells through the pathway mentioned above. CONCLUSION: These findings raise a rationale that this 3-benzyl coumarin seco-B-ring derivative and phenylsulfonylfuroxan hybrid could be a promising candidate for developing as a therapeutic agent toward NSCLC, and the combination therapy through PI3K/Akt/mTOR signaling pathway may result in optimized treatment outcomes.

Surface-Enhanced Resonance Raman Scattering-Guided Brain Tumor Surgery Showing Prognostic Benefit in Rat Models.

Glioma is the most frequent form of malignant brain tumors. Surgical debulking is a major strategy for glioma treatment. However, there is a great challenge for the neurosurgeons to intraoperatively identify the true margins of glioma because of its infiltrative nature. Tumor residues or microscopic satellite foci left in the resection bed are the main reasons leading to early recurrence as well as poor prognosis. In this study, a surface-enhanced resonance Raman scattering (SERRS) probe was developed to intraoperatively guide glioma resection. In this probe, molecular reporters with absorptive maxima at the near-infrared wavelength range were covalently functionalized on the surface of gold nanostars. This SERRS probe demonstrated an ultrahigh sensitivity with a detection limit of 5.0 pM in aqueous solution. By the development of glioma xenografts in a mouse dorsal skin window chamber, extravasation of this probe from leaky tumor vasculature as functions of time and distance to tumor boundary was investigated. Importantly, the invasive margin of the tumor xenograft was demarcated by this probe with a high signal-to-background ratio. Preoperative magnetic resonance imaging (MRI) first defined the position of orthotopic glioma xenografts in the brain of rat models, and the craniotomy plan was designed. The brain tumor was then excised intraoperatively step-by-step with the assistance of a handheld Raman scanner till the Raman signals of the probe completely disappeared in the resection bed. Notably, longitudinal MRI showed that SERRS-guided surgery significantly reduced the tumor recurrence rate and improved the overall survival of rat models compared with the white light-guided surgery. Overall, this work demonstrates the prognostic benefit of SERRS-guided glioma surgery in animal models. Because delineation of tumor-invasive margins is a common challenge faced by the surgeons, this SERRS probe with a picomolar detection limit holds the promise in improving the surgical outcome of different types of infiltrated tumors.

PirB inhibits axonal outgrowth via the PI3K/Akt/mTOR signaling pathway.

Accumulating data strongly suggests that leukocyte immunoglobulin like receptor B1 (PirB) inhibits axonal outgrowth. However, the underlying mechanisms remain unclear. In the present study, cortical neurons of newborn mice were cultured with Nogo­66 (Nogo­p4; 4 µmol/l; a PirB ligand) together with NEP1­40 (Nogo inhibitory peptide) and/or anti­PirB body (50 mg/ml). PirB mRNA and protein was higher in cultured neurons induced by Nogo­66 compared with untreated cells. Neurite outgrowth assays demonstrated that the inhibitory effects of Nogo­66 on axonal outgrowth were reversed by anti­PirB body. Reverse transcription­quantitative polymerase chain reaction and western blot assays demonstrated that anti­PirB treatment led to reduced mRNA and protein expression of phosphoinositide 3­kinase (PI3K), Akt serine/threonine kinase (Akt), mechanistic target of rapamycin kinase (mTOR), myosin IIA and cofilin, which are involved in axonal outgrowth. Furthermore, blockade of the PI3K/Akt/mTOR pathway using a PI3K inhibitor or an mTOR inhibitor diminished the stimulatory effect of anti­PirB on axonal outgrowth, and the reduced effect of anti­PirB on factors that were activation by anti­PirB. In addition, blockade of PI3K/Akt/mTOR enhanced anti­PirB­induced gene and protein expression. These results revealed that PirB functions as a potential suppressor in axonal outgrowth via repressing PI3K/Akt/mTOR signaling pathway, and PirB/PI3K/Akt/mTOR may be a novel target for enhancing axonal outgrowth for developing rational therapeutic strategies.

From the vascular microenvironment to neurogenesis.

In response to injury, the function of the adult mammalian central nervous system (CNS) is repaired by angiogenesis and neurogenesis. The proliferation, differentiation and migration of the neural progenitor cells (NPCs) are regulated by several components of the vascular microenvironment within the neural stem cell niche. Vascular endothelial cells (VECs), which make up blood vessels, as well as extracellular components, affect neurogenesis directly via contacting with NPCs. Additionally, soluble factors from the vascular system that are released into the CNS enhance neurogenesis in the form of paracrine signaling. The purpose of this mini-review is to highlight the roles of the vascular microenvironment as a mediator in promoting neurogenesis.

Stem/progenitor cells: a potential source of retina-specific cells for retinal repair.

Retinal injury generally results in permanent visual disturbance or even blindness. Any effort to restore vision in such condition would require replacement of the highly specialized retinal cells. Stem/progenitor cells have been proposed as a potential source of new retina-specific cells to replace those lost due to retina injury. Evidence to date suggests that continued development of stem cell therapies may ultimately lead to viable treatment options for retina injury. A wide range of stem/progenitor cells from various sources is currently being investigated for the treatment of retinal injury. This article reviews the recent achievements about stem/progenitor cell source for retinal repair.