The Long-Noncoding RNA TUG1 Regulates Oxygen-Induced Retinal Neovascularization in Mice via MiR-299.

Purpose: The oxygen-induced retinal neovascularization mouse model closely approximates pathological changes associated with human retinal neovascularization-associated diseases, including retinopathies. We used this model and human retinal endothelial cells (HRECs) under hypoxia to explore the relationship between taurine upregulated gene-1 (TUG1), vascular endothelial growth factor (VEGF), and miR-299-3p on retinopathy of prematurity (ROP). Methods: An oxygen-induced retinopathy (OIR) mouse model was established; the mice were divided into a normal control group, OIR group, TUG1 control group (lentivirus control), and TUG1-knockdown group. The apoptosis of retinal cells was evaluated using a TUNEL assay. Angiogenic, apoptotic, and inflammatory factors were detected by Western blot, immunohistochemistry, and immunofluorescence analyses. HRECs were cultured under hypoxia and assessed for VEGF expression, apoptosis, tubule formation, and migration ability. The relationship between TUG1, VEGF, and miR-299-3p was detected via a dual luciferase reporter gene assay. Results: Intravitreal injection of TUG1 lentivirus reduced the inflammatory response in the mouse retinal tissue and markedly reduced pathological changes in the retina. Overexpression of miR-299 in HRECs reduced the apoptosis rate, tube formation, and migration ability of hypoxia-treated cells, thereby inhibiting the formation of new blood vessels. The dual luciferase reporter gene assay suggested that miR-299 has binding sites for TUG1 and VEGF. Conclusions: TUG1 reduces the expression of VEGFA by competitively adsorbing miR-299-3p and facilitates the regulation of retinal neovascularization, suggesting that it may serve as a new therapeutic target for retinal neovascular diseases.

Maternally expressed gene 3 regulates retinal neovascularization in retinopathy of prematurity.

The mouse model of oxygen induced retinopathy is suitable for the study of various retinal neovascularization diseases, including retinopathy of prematurity. The maternally expressed gene 3 (MEG3) has been demonstrated to have an inhibitory effect on diabetic retinopathy. In this study, we investigated the role of MEG3 overexpression in oxygen-induced retinopathy in mice. The results showed that MEG3 overexpression effectively inhibited the production of retinal neovascularization in oxygen-induced retinopathy mice. It acts by down-regulating the expression of phosphoinositide 3-kinase, serine/threonine kinase, and vascular endothelial growth factor and pro-inflammatory factors. MEG3 overexpression lentivirus has a future as a new method for the clinical treatment of retinopathy of prematurity. The animal experiments were approved by the Animal Ethics Committee of Shengjing Hospital of China Medical University, China (approval No. 2016PS074K) on February 25, 2016.

Effect and mechanism of the long noncoding RNA MALAT1 on retinal neovascularization in retinopathy of prematurity.

AIMS: The most typical pathological manifestation of retinopathy of prematurity (ROP) is Retinal neovascularization (RNV). Long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has been reported to mediate angiogenesis. Our experiment aimed to research the effect and mechanism of the MALAT1 on RNV in ROP. MAIN METHODS: C57 mice was used to establish oxygen-introduced retinopathy (OIR), and divided into control, hyperoxia, hyperoxia control siRNA, and hyperoxia MALAT1 siRNA groups. KEY FINDINGS: It was shown that MALAT1 mRNA was high expressed in the retinas of OIR mice. Further studies revealed that after intravitreal injection of MALAT1 siRNA, the degree of retinopathy was significantly reduced compared with OIR group. In addition, the protein and mRNA expression levels of CCN1, AKT and VEGF were significantly decreased. This was accompanied by a decrease in inflammatory genes including IL-1ß, IL-6, and TNF-α compared with the hyperoxia control siRNA mice. SIGNIFICANCE: The result suggested that MALAT1 may be involved in the process of RNV in ROP and MALAT1 siRNA may be a promising agent for the treatment of ROP by inhibiting RNV.

Expression profiles of long noncoding RNAs in retinopathy of prematurity.

Long noncoding RNA (lncRNA) regulates the proliferation and migration of human retinal endothelial cells, as well as retinal neovascularization in diabetic retinopathy. Based on similarities between the pathogenesis of retinopathy of prematurity (ROP) and diabetic retinopathy, lncRNA may also play a role in ROP. Seven-day-old mice were administered 75 ± 2% oxygen for 5 days and normoxic air for another 5 days to establish a ROP model. Expression of lncRNA and mRNA in the retinal tissue of mice was detected by high-throughput sequencing technology, and biological functions of the resulted differentially expressed RNAs were evaluated by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses. The results showed that compared with the control group, 57 lncRNAs were differentially expressed, including 43 upregulated and 14 downregulated, in the retinal tissue of ROP mice. Compared with control mice, 42 mRNAs were differentially expressed in the retinal tissue of ROP mice, including 24 upregulated and 18 downregulated mRNAs. Differentially expressed genes were involved in ocular development and related metabolic pathways. The differentially expressed lncRNAs may regulate ROP in mice via microRNAs and multiple signaling pathways. Our results revealed that these differentially expressed lncRNAs may be therapeutic targets for ROP treatment. This study was approved by the Medical Ethics Committee of Shengjing Hospital of China Medical University on February 25, 2016 (approval No. 2016PS074K).

Citreoviridin induces ROS-dependent autophagic cell death in human liver HepG2 cells.

Citreoviridin (CIT) is one of toxic mycotoxins derived from fungal species in moldy cereals. Whether CIT exerts hepatotoxicity and the precise molecular mechanisms of CIT hepatotoxicity are not completely elucidated. In this study, the inhibitor of autophagosome formation, 3-methyladenine, protected the cells against CIT cytotoxicity, and the autophagy stimulator rapamycin further decreased the cell viability of CIT-treated HepG2 cells. Knockdown of Atg5 with Atg5 siRNA alleviated CIT-induced cell death. These finding suggested the hypothesis that autophagic cell death contributed to CIT-induced cytotoxicity in HepG2 cells. CIT increased the autophagosome number in HepG2 cells observed under a transmission electron microscope, and this effect was confirmed by the elevated LC3-II levels detected through Western blot. Reduction of P62 protein levels and the result of LC3 turnover assay indicated that the accumulation of autophagosomes in the CIT-treated HepG2 cells was due to increased formation rather than impaired degradation. The pretreatment of HepG2 cells with the ROS inhibitor NAC reduced autophagosome formation and reversed the CIT cytotoxicity, indicating that CIT-induced autophagic cell death was ROS-dependent. In summary, ROS-dependent autophagic cell death of HpeG2 cells described in this study may help to elucidate the underlying mechanism of CIT cytotoxicity.