HMGB1 is an architectural chromatin-binding protein that can be released actively by activated cells or passively by dying cells and can serve as a DAMP molecule to drive the pathogenesis of inflammatory and angiogenic diseases. Through TLR4 and RAGE signaling pathways, HMGB1 could regulate vascular growth in vivo and in vitro through diverse mechanisms, including induction of proangiogenic cytokine release and activation of ECs, macrophages, EPCs, and mesoangioblasts, all of which could contribute to vessel formation. Accordingly, HMGB1 plays a significant role in many angiogenesis-related conditions, such as tumors, PDR, wound-healing, and ischemia-induced angiogenesis. In this review, we focus on the regulatory role of HMGB1 in angiogenesis and recent progress in therapeutic strategies targeting HMGB1.

PurposeTo study the role of long non-coding RNA (lncRNA) MALAT1 in transforming growth factor beta 1 (TGF-β1)-induced epithelial-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells.MethodsARPE-19 cells were cultured and exposed to TGF-β1. The EMT of APRE-19 cells is confirmed by morphological change, as well as the increased expression of alpha-smooth muscle actin (αSMA) and fibronectin, and the down-regulation of E-cadherin and Zona occludin-1(ZO-1) at both mRNA and protein levels. The expression of lncRNA MALAT1 in RPE cells were detected by quantitative real-time PCR. Knockdown of MALAT1 was achieved by transfecting a small interfering RNA (SiRNA). The effect of inhibition of MALAT1 on EMT, migration, proliferation, and TGFβ signalings were observed. MALAT1 expression was also detected in primary RPE cells incubated with proliferative vitreoretinopathy (PVR) vitreous samples.ResultsThe expression of MALAT1 is significantly increased in RPE cells incubated with TGFβ1. MALAT1 silencing attenuates TGFβ1-induced EMT, migration, and proliferation of RPE cells, at least partially through activating Smad2/3 signaling. MALAT1 is also significantly increased in primary RPE cells incubated with PVR vitreous samples.ConclusionLncRNA MALAT1 is involved in TGFβ1-induced EMT of human RPE cells and provides new understandings for the pathogenesis of PVR.

Complement is known to play a role in alcoholic fatty liver disease (AFLD), but the underlying mechanisms are poorly understood, thereby constraining the development of a rational approach for therapeutic intervention in the complement system. C3 deficiency has been shown to impart protective effects against ethanol-induced hepatic steatosis and inflammation. Here we demonstrate a protection effect in wild-type mice by treatment with CR2-Crry, a specific inhibitor of C3 activation. The expression of glycine transfer (t) RNA-derived fragments (Gly-tRFs) is upregulated in ethanol-fed mice and inhibition of Gly-tRFs in vivo decreases chronic ethanol feeding-induced hepatosteatosis without affecting inflammation. The expression of Gly-tRF was downregulated in C3-deficient or CR2-Crry-treated mice, but not in C5-deficient mice; Gly-tRF expression was restored by the C3 activation products C3a or Asp (C3a-des-Arg) via the regulation of CYP2E1. Transcriptome profiling of hepatic tissues showed that Gly-tRF inhibitors upregulate the expression of sirtuin1 (Sirt1) and subsequently affect downstream lipogenesis and β-oxidation pathways. Mechanistically, Gly-tRF interacts with AGO3 to downregulate Sirt1 expression via sequence complementarity in the 3′ UTR. Notably, the expression levels of C3d, CYP2E1 and Gly-tRF are upregulated, whereas Sirt1 is decreased in AFLD patients compared to healthy controls. Collectively, our findings suggest that C3 activation products contribute to hepatosteatosis by regulating the expression of Gly-tRF. Complement inhibition at the C3 activation step and treatment with Gly-tRF inhibitors may be potential and precise therapeutic approaches for AFLD.

BackgroundTemozolomide (TMZ) is an oral DNA-alkylating agent used for treating patients with glioblastoma. However, therapeutic benefits of TMZ can be compromised by the expression of O6-methylguanine methyltransferase (MGMT) in tumor tissue. Here we used MGMT-expressing glioblastoma stem cells (GSC) lines as a model for investigating the molecular mechanism underlying TMZ resistance, while aiming to explore a new treatment strategy designed to possibly overcome resistance to the clinically relevant dose of TMZ (35 μM).MethodsMGMT-expressing GSC cultures are resistant to TMZ, and IC50 (half maximal inhibitory concentration) is estimated at around 500 μM. Clonogenic GSC surviving 500 μM TMZ (GSC-500 μM TMZ), were isolated. Molecular signatures were identified via comparative analysis of expression microarray against parental GSC (GSC-parental). The recombinant protein of top downregulated signature was used as a single agent or in combination with TMZ, for evaluating therapeutic effects of treatment of GSC.ResultsThe molecular signatures characterized an activation of protective stress responses in GSC-500 μM TMZ, mainly including biotransformation/detoxification of xenobiotics, blocked endoplasmic reticulum stress-mediated apoptosis, epithelial-to-mesenchymal transition (EMT), and inhibited growth/differentiation. Bone morphogenetic protein 7 (BMP7) was identified as the top down-regulated gene in GSC-500 μM TMZ. Although augmenting BMP7 signaling in GSC by exogenous BMP7 treatment did not effectively stop GSC growth, it markedly sensitized both GSC-500 μM TMZ and GSC-parental to 35 μM TMZ treatment, leading to loss of self-renewal and migration capacity. BMP7 treatment induced senescence of GSC cultures and suppressed mRNA expression of CD133, MGMT, and ATP-binding cassette drug efflux transporters (ABCB1, ABCG2), as well as reconfigured transcriptional profiles in GSC by downregulating genes associated with EMT/migration/invasion, stemness, inflammation/immune response, and cell proliferation/tumorigenesis. BMP7 treatment significantly prolonged survival time of animals intracranially inoculated with GSC when compared to those untreated or treated with TMZ alone (p = 0.0017), whereas combination of two agents further extended animal survival compared to BMP7 alone (p = 0.0489).ConclusionsThese data support the view that reduced endogenous BMP7 expression/signaling in GSC may contribute to maintained stemness, EMT, and chemoresistant phenotype, suggesting that BMP7 treatment may provide a novel strategy in combination with TMZ for an effective treatment of glioblastoma exhibiting unmethylated MGMT.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0459-1) contains supplementary material, which is available to authorized users.

Glioblastoma stem cells (GSC) co-exhibiting a tumor-initiating capacity and a radio-chemoresistant phenotype, are a compelling cell model for explaining tumor recurrence. We have previously characterized patient-derived, treatment-resistant GSC clones (TRGC) that survived radiochemotherapy. Compared to glucose-dependent, treatment-sensitive GSC clones (TSGC), TRGC exhibited reduced glucose dependence that favor the fatty acid oxidation pathway as their energy source. Using comparative genome-wide transcriptome analysis, a series of defense signatures associated with TRGC survival were identified and verified by siRNA-based gene knockdown experiments that led to loss of cell integrity. In this study, we investigate the prognostic value of defense signatures in glioblastoma (GBM) patients using gene expression analysis with Probeset Analyzer (131 GBM) and The Cancer Genome Atlas (TCGA) data, and protein expression with a tissue microarray (50 GBM), yielding the first TRGC-derived prognostic biomarkers for GBM patients. Ribosomal protein S11 (RPS11), RPS20, individually and together, consistently predicted poor survival of newly diagnosed primary GBM tumors when overexpressed at the RNA or protein level [RPS11: Hazard Ratio (HR) = 11.5, p<0.001; RPS20: HR = 4.5, p = 0.03; RPS11+RPS20: HR = 17.99, p = 0.001]. The prognostic significance of RPS11 and RPS20 was further supported by whole tissue section RPS11 immunostaining (27 GBM; HR = 4.05, p = 0.01) and TCGA gene expression data (578 primary GBM; RPS11: HR = 1.19, p = 0.06; RPS20: HR = 1.25, p = 0.02; RPS11+RPS20: HR = 1.43, p = 0.01). Moreover, tumors that exhibited unmethylated O-6-methylguanine-DNA methyltransferase (MGMT) or wild-type isocitrate dehydrogenase 1 (IDH1) were associated with higher RPS11 expression levels [corr (IDH1, RPS11) = 0.64, p = 0.03); [corr (MGMT, RPS11) = 0.52, p = 0.04]. These data indicate that increased expression of RPS11 and RPS20 predicts shorter patient survival. The study also suggests that TRGC are clinically relevant cells that represent resistant tumorigenic clones from patient tumors and that their properties, at least in part, are reflected in poor-prognosis GBM. The screening of TRGC signatures may represent a novel alternative strategy for identifying new prognostic biomarkers.

Exosomes have recently emerged as a pivotal mediator of many physiological and pathological processes. However, the role of exosomes in proliferative vitreoretinopathy (PVR) has not been reported. In this study, we aimed to investigate the role of exosomes in PVR. Transforming growth factor beta 2 (TGFß‐2) was used to induce epithelial‐mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells, as an in vitro model of PVR. Exosomes from normal and EMTed RPE cells were extracted and identified. We incubated extracted exosomes with recipient RPE cells, and co‐cultured EMTed RPE cells and recipient RPE cells in the presence of the exosome inhibitor GW4869. Both experiments suggested that there are further EMT‐promoting effects of exosomes from EMTed RPE cells. MicroRNA sequencing was also performed to identify the miRNA profiles in exosomes from both groups. We identified 34 differentially expressed exosomal miRNAs (P <. 05). Importantly, miR‐543 was found in exosomes from EMTed RPE cells, and miR‐543‐enriched exosomes significantly induced the EMT of recipient RPE cells. Our study demonstrates that exosomal miRNA is differentially expressed in RPE cells during EMT and that these exosomal miRNAs may play pivotal roles in EMT induction. Our results highlight the importance of exosomes as cellular communicators within the microenvironment of PVR.

Abstract. Drusen are considered a hallmark characteristic of age-related macular degeneration (AMD). In our previous study, we found that amyloid-β (Aβ) peptide, a component of drusen, induced the cells of the retinal pigment epithelium (RPE; RPE cells) to enter senescence; however, its effects in vivo remain unknown. Thus, the present study was carried out to explore the in vivo effects of Aβ peptide on RPE cell senescence and senescence-associated inflammation in C57BL/6 mice. C57BL/6 mice received a subretinal injection of Aβ(1-42) peptide; on day 7 post-injection, the mice were anesthetized and subjected to whole-body perfusion with 4% paraformaldehyde (PFA) in PBS and the whole eyes were then enucleated. Retinal function was assessed by electroretinography (ERG), and the morphological characteristics of the retina were examined by light and electron microscopy. Fundus autofluorescence (FAF) was examined by confocal scanning laser ophthalmoscopy (cSLO). The expression of p16INK4a , a marker of cellular senescence, was examined by immunofluorescence staining and western blot analysis. The RPE-choroid was analyzed for cytokine expression by RT-PCR. In Aβ(1-42)-injected mice, scotopic ERG responses declined. Degenerative alterations, including the disruption of the inner segment (IS)/outer segment (OS) junction and extensive vacuolation and thickness of Bruch's membrane (BrM) were observed under a a light microscope. The accumulation of vacuoles and the loss of basal infoldings in the RPE were identified using an electron microscope. FAF and p16INK4a expression increased in Aβ(1-42)-injected mice. In addition, Aβ(1-42) upregulated interleukin (IL)-6 and IL-8 gene expression in the RPE-choroid. In conclusion, our results confirm the effects of Aβ(1-42) peptide on RPE senescence in vivo. The Aβ-injected mice developed AMD-like ocular pathology. It is thus suggested that RPE cell senescence is a potential mechanistic link between inflammation and retinal degeneration.

Tumor‐associated macrophages (TAMs) are vital constituents in mediating cell‐to‐cell communication within the tumor microenvironment. However, the molecular mechanisms underlying the interplay between TAMs and tumor cells that guide cell fate are largely undetermined. Extracellular vesicles, also known as exosomes, which are derived from TAMs, are the components exerting regulatory effects. Thus, understanding the underlying mechanism of “onco‐vesicles” is of crucial importance for prostate cancer (PCa) therapy. In this study, we analyzed micro RNA sequences in exosomes released by THP‐1 and M2 macrophages and found a significant increase in miR‐95 levels in TAM‐derived exosomes, demonstrating the direct uptake of miR‐95 by recipient PCa cells. In vitro and in vivo loss‐of‐function assays suggested that miR‐95 could function as a tumor promoter by directly binding to its downstream target gene, JunB, to promote PCa cell proliferation, invasion, and epithelial–mesenchymal transition. The clinical data analyses further revealed that higher miR‐95 expression results in worse clinicopathological features. Collectively, our results demonstrated that TAM‐mediated PCa progression is partially attributed to the aberrant expression of miR‐95 in TAM‐derived exosomes, and the miR‐95/JunB axis provides the groundwork for research on TAMs to further develop more‐personalized therapeutic approaches for patients with PCa.