Regulatory mechanisms of retinal ganglion cell death in normal tension glaucoma and potential therapies.

Normal tension glaucoma (NTG) is a multifactorial optic neuropathy characterized by normal intraocular pressure, progressive retinal ganglion cell (RGC) death, and glaucomatous visual field loss. Recent studies have described the mechanisms underlying the pathogenesis of NTG. In addition to controlling intraocular pressure, neuroprotection and reduction of RGC degeneration may be beneficial therapies for NTG. In this review, we summarized the main regulatory mechanisms of RGC death in NTG, including autophagy, glutamate neurotoxicity, oxidative stress, neuroinflammation, immunity, and vasoconstriction. Autophagy can be induced by retinal hypoxia and axonal damage. In this process, ischemia can cause mutations of optineurin and activate the nuclear factor-kappa B pathway. Glutamate neurotoxicity is induced by the over-stimulation of N-methyl-D-aspartate membrane receptors by glutamate, which occurs in RGCs and induces progressive glaucomatous optic neuropathy. Oxidative stress also participates in NTG-related glaucomatous optic neuropathy. It impairs the mitochondrial and DNA function of RGCs through the apoptosis signal-regulating kinase-JUN N-terminal kinase pathway. Moreover, it increases inflammation and the immune response of RGCs. Endothelin 1 causes endothelial dysfunction and impairment of ocular blood flow, promoting vasospasm and glaucomatous optic neuropathy, as a result of NTG. In conclusion, we discussed research progress on potential options for the protection of RGCs, including TANK binding kinase 1 inhibitors regulating autophagy, N-methyl-D-aspartate receptor antagonists inhibiting glutamate toxicity, ASK1 inhibitors regulating mitochondrial function, and antioxidants inhibiting oxidative stress. In NTG, RGC death is regulated by a network of mechanisms, while various potential targets protect RGCs. Collectively, these findings provide insight into the pathogenesis of NTG and potential therapeutic strategies.

[Effect of MiR-142-3p Targeting HOXA5 on Proliferation, Cycle Arrest and Apoptosis of Acute B Lymphocytic Leukemia Cells].

OBJECTIVE: To investigate the effect and molecular mechanism of miR-142-3p to the proliferation, cycle and apoptosis of acute B lymphocytic leukemia (B-ALL) cells by regulating the homeobox gene 5 (HOXA5) expression. METHODS: Real-time fluorescence quantitative PCR was used to detect the expression levels of miR-142-3p and HOXA5 in human B-ALL cell Nalm6 cell line and human B lymphoblast Hmy2-cir cells. Nalm6 was transfected by using liposome transfection technology, miR-142-3p mimic, pcDNA-HOXA5 overexpression plasmid, miR-142-3p mimic+pcDNA-HOXA5 overexpression plasmid, and control. The binding site of HOXA5 and miR-142-3p was predicted according to microRNA.org, and the targeting relationship between miR-142-3p and HOXA5 gene was detected by double luciferase reporter gene experiment. The effect of miR-142-3p to the proliferation of Nalm6 cells was detected using the Cell Counting Box-8 (CCK-8) method and cell clone formation experiments. Flow cytometry was used to detect the effects of miR-142-3p to cell cycle distribution and apoptosis of Nalm6 cells. The expression levels of cell cycle-related proteins, including G1 /S-specific cyclin-D1 (CyclinD1), Cyclin-dependent kinase 4 (CDK4) and B-cell lymphoma/ leukemia-2 protein (BCL-2), BCL-2 related X protein (Bax), cysteine-aspartate-specific protease (Caspase-3) were detected by Western blot. RESULTS: Compared with Hmy2-cir cells, miR-142-3p showed low expression in Nalm6 cells and HOXA5 showed high expression (P<0.05). MiR-142-3p and HOXA5 3'-UTR showed complementary binding regions, the luciferase activity of miR-142-3p mimic and wild-type HOXA5 3'-UTR was significantly lower than that of miR-142-3p negative control and wild-type HOXA5 3'-UTR (P<0.05). The proliferation of Nalm6 cells and the number of cell clones could be inhibited by miR-142-3p mimic after 48 and 72 hours of transfection (P<0.05), which causing G1 phase arrest of Nalm6 cells and inhibiting the expression of CyclinD1 and CDK4 protein (P<0.01), promoting the apoptosis of Nalm6 cells, and inhibiting the expression of BCL-2 protein, moreover, promoting the expression of Bax and Caspase-3 protein (P<0.05). CONCLUSION: MiR-142-3p can inhibit the proliferation of Nalm6 cells by targeting down-regulation the expression of HOXA5, arrest the G1 phase of cells, and promote apoptosis of the cells.

Oxidative Stress-Induced circHBEGF Promotes Extracellular Matrix Production via Regulating miR-646/EGFR in Human Trabecular Meshwork Cells.

Primary open-angle glaucoma (POAG), a leading cause of irreversible vision loss, presents with increased prevalence and a higher degree of clinical severity in the world. Growing evidence has shown that ncRNAs are involved in the fibrotic process, which is thought to be the proegumenal cause of POAG. Here, we screened out a differentially expressed circRNA (named circHBEGF) in human trabecular meshwork cells (HTMCs) under oxidative stress, which is spliced from pre-HBEGF. circHBEGF promotes the expression of extracellular matrix (ECM) genes (fibronectin and collagen I). Further studies revealed that circHBEGF could competitively bind to miR-646 as a miRNA sponge to regulate EGFR expression in HTMCs. Importantly, HBEGF can also activate EGF signaling pathways, through which can transcriptionally activate ECM genes in HTMCs. In summary, this study investigates the functions and molecular mechanisms of oxidative stress-induced circHBEGF in the regulation of ECM production in HTMCs through the miR646/EGFR pathway. These findings further elucidate the pathogenic mechanism and may identify novel targets for the molecular therapy of POAG.

Long non-coding RNA RP11-820 promotes extracellular matrix production via regulating miR-3178/MYOD1 in human trabecular meshwork cells.

Primary open angle glaucoma (POAG) is the most common type of glaucoma. At the mechanistic level, POAG is thought to be caused by extracellular matrix (ECM) deposition in the trabecular meshwork (TM). Growing evidence has shown that long noncoding RNAs (lncRNAs) are involved in the fibrotic process underlying many diseases. This study was undertaken to explore the role of lncRNA-RP11-820 in ECM production of human TM cells (HTMCs). Our results revealed that lncRNA-RP11-820 was significantly upregulated under oxidative stress in HTMCs. Further investigation revealed that lncRNA-RP11-820 directly binds miR-3178, through which the expression of MYOD1 is regulated. Importantly, MYOD1 can transcriptionally activate ECM genes in HTMCs, in complex with STAT3. Taken together, our data established that oxidative stress-induced lncRNA-RP11-820 plays a key role in regulating the miR-3178/MYOD1/ECM axis in HTMCs. These findings further elucidate the pathogenic mechanism and provide a novel therapeutic target relevant to POAG.

Ocular Surface Changes in Prostaglandin Analogue-Treated Patients.

Glaucoma is the second leading cause of blindness globally. Reducing intraocular pressure (IOP) has been acknowledged to be the main therapy for glaucoma. Prostaglandin analogues (PGAs) have become the first-line therapy for patients with glaucoma due to their powerful efficacy for lowering (IOP). However, usage of PGAs can also cause several notable side effects, including the changes in ocular surface. The relationship between PGAs and ocular surface changes is complicated and still remains unclear. In the present review, we summarize the recent studies of the effects of PGAs on ocular changes as well as the possible mechanisms that might provide new considerations during clinical medication.

tRF/miR-1280 Suppresses Stem Cell-like Cells and Metastasis in Colorectal Cancer.

Several studies have shown that tRNAs can be enzymatically cleaved to generate distinct classes of tRNA-derived fragments (tRF). Here, we report that tRF/miR-1280, a 17-bp fragment derived from tRNALeu and pre-miRNA, influences Notch signaling pathways that support the function of cancer stem-like cells (CSC) in colorectal cancer progression. tRF/miR-1280 expression was decreased in human specimens of colorectal cancer. Ectopic expression of tRF/miR-1280 reduced cell proliferation and colony formation, whereas its suppression reversed these effects. Mechanistic investigations implicated the Notch ligand JAG2 as a direct target of tRF/miR-1280 binding through which it reduced tumor formation and metastasis. Notably, tRF/miR-1280-mediated inactivation of Notch signaling suppressed CSC phenotypes, including by direct transcriptional repression of the Gata1/3 and miR-200b genes. These results were consistent with findings of decreased levels of miR-200b and elevated levels of JAG2, Gata1, Gata3, Zeb1, and Suz12 in colorectal cancer tissue specimens. Taken together, our results established that tRF/miR-1280 suppresses colorectal cancer growth and metastasis by repressing Notch signaling pathways that support CSC phenotypes. Furthermore, they provide evidence that functionally active miRNA can be derived from tRNA, offering potential biomarker and therapeutic uses. Cancer Res; 77(12); 3194-206. ©2017 AACR.

HES1 promotes extracellular matrix protein expression and inhibits proliferation and migration in human trabecular meshwork cells under oxidative stress.

Glaucoma is the leading cause of irreversible blindness. The most prevalent form of glaucoma is primary open-angle glaucoma (POAG). Oxidative stress is one of the major pathogenic factors of the POAG, and can elicit molecular and functional changes in trabecular meshwork cells, causing increased aqueous humor outflow resistance and elevated intraocular pressure. However, the regulatory mechanisms underlying oxidative stress-induced cell phenotypic changes remain elusive. Herein, we exposed primary human trabecular meshwork cells to the oxidative stress induced by 300 µM H2O2 for 2 h, and found significantly up-regulated expression of extracellular matrix proteins and a transcription factor, hairy and enhancer of split-1 (HES1). The cell functions, including migration and proliferation, were impaired by the oxidative stress. Furthermore, HES1 shRNA abrogated the extracellular matrix protein up-regulation and rescued the functional defects caused by the oxidative stress; conversely, HES1 overexpression resulted in the molecular and functional changes similar to those induced by H2O2. These results suggest that HES1 promotes extracellular matrix protein expression and inhibits proliferative and migratory functions in the trabecular meshwork cells under oxidative stress, thereby providing a novel pathogenic mechanism underlying and a potential therapeutic target to the POAG.

Relationship between the Pathogenesis of Glaucoma and miRNA.

Small RNA (microRNA or miRNA) is a kind of small noncoding single-stranded RNA that regulates complementary mRNA at the posttranscriptional level in eukaryotic organisms. As important regulatory factors, miRNAs play an important role in the occurrence and development of glaucoma and widely participate in regulating biological processes of glaucoma-related genes. This article reviews the connection between the aqueous humor, trabecular meshwork, the apoptosis of retinal ganglion cells, and miRNA.

MicroRNA-483-3p Inhibits Extracellular Matrix Production by Targeting Smad4 in Human Trabecular Meshwork Cells.

PURPOSE: This study investigated the effects of microRNA-483-3p (miR-483-3p) on extracellular matrix (ECM) production, and clarified the regulatory mechanism of microRNA-483-3p in human trabecular meshwork cells (HTMCs) under oxidative stress. METHODS: The expression levels of ECM (fibronectin, laminin, collagen I) in HTMCs under oxidative stress were measured by Western blot. Changes of miR-483-3p expression in HTMCs were evaluated by quantitative polymerase chain reaction (qPCR). After using lentivirus stably expressing pri-miR-483, the effects of miR-483-3p on the ECM were assessed by qPCR and Western blot. Smad4, the potential target of miR-483-3p according to mRNA target-predicting algorithms, was confirmed by luciferase assay and Western blot. Furthermore, the effects of Smad4 knockdown on ECM expression were investigated by qPCR and Western blot. RESULTS: The mRNA and protein levels of ECM (fibronectin, laminin, collagen I) were upregulated in HTMCs induced by oxidative stress. The expression level of miR-483-3p decreased in HTMCs under oxidative stress, and the ectopic expression of miR-483-3p decreased the levels of ECM. In addition, miR-483-3p targeted Smad4 through two binding sites, resulting in a decrease of Smad4 expression. Furthermore, knockdown of Smad4 reduced the levels of ECM in HTMCs. CONCLUSIONS: MicroRNA-483-3p has an inhibitory effect on ECM production in HTMCs through downregulating Smad4, which indicates that miR-483-3p may serve as a potential therapeutic target in glaucoma.