Background/Aims: Oxidized low-density lipoprotein (oxLDL) promotes unregulated platelet activation in patients with dyslipidemic disorders. Although oxLDL stimulates activating signaling, researchers have not clearly determined how these events drive accelerated thrombosis. Here, we describe the mechanism by which ROS regulate autophagy during ox-LDL-induced platelet activation by modulating the PI3K/AKT/mTOR signaling pathway. Methods: For in vitro experiments, ox-LDL, the ROS scavenger N-acetylcysteine (NAC), the mTOR inhibitor rapamycin and the autophagy inhibitor 3-MA were used alone or in combination with other compounds to treat platelets. Then, platelet aggregation was evaluated on an aggregometer and platelet adhesion was measured under shear stress. The levels of a platelet activation marker (CD62p) were measured by flow cytometry, reactive oxygen species (ROS) levels were then quantified by measuring DCFH-DA fluorescence intensity via flow cytometry. Nitric oxide (NO) and superoxide (O2·-) levels were determined by the nitric acid deoxidize enzyme method and lucigenin-enhanced chemiluminescence (CL), respectively. Transmission electron microscopy was used to observe the autophagosome formation, immunofluorescence staining was employed to detect LC3 expression and western blotting was used to measure the levels of PI3K/AKT/mTOR pathway- and autophagy-related proteins. Results: Ox-LDL-induced platelets showed a significant increase in platelet aggregation and adhesion, CD62p expression, ROS level and O2·- content, with an elevated LC3II/LC3I ratio and Beclin1 expression, but a dramatic reduction in the levels of p62 and pathway-related proteins (all P < 0.05). However, platelet activation and autophagy were aggravated by the Rapamycin treatment, and decreased following treatment with NAC, 3-MA, or NAC and 3-MA, together with increased activity of the PI3K/AKT/mTOR pathway. Additionally, decreased platelet activation and autophagy were observed in platelets treated with NAC and Rapamycin or Rapamycin and 3-MA compared with platelets treated with Rapamycin alone, suggesting that both NAC and 3-MA reversed the effects of Rapamycin. Conclusion: Inhibition of ROS production may reduce autophagy to suppress ox-LDL-induced platelet activation by activating PI3K/AKT/mTOR pathway.
This research planned to grab the expression and impact of lncRNA HAGLROS in the biology and progression of mantle cell lymphoma. HAGLROS level in mantle cell lymphoma cell lines was detected, followed by investigation of the influences of HAGLROS silencing on Mino cell biological performances. Afterwards, the express patterns of HAGLROS vs. miR-100, as well as miR-100 vs. ATG5, were investigated. Furthermore, whether HAGLROS could regulate the signals of PI3K/AKT/mTOR was analyzed. HAGLROS level was high in mantle cell lymphoma cell lines. Silencing of HAGLROS inhibited Mino cell viability, increased apoptosis and decreased autophagy by sponging miR-100. Moreover, miR-100 targeted ATG5 fixed. Furthermore, HAGLROS suppression resulted in inhibition on the briskness of PI3K/ AKT/mTOR signals. Concurrently HAGLROS suppression and miR-100 inhibitor markedly changed the impacts of HAGLROS down-regulation alone on activating PI3K/AKT/mTOR signals, which could further change after co-transfection of si-HAGLROS þ miR-100 inhibitor þ siATG5. Our findings point out that expression of HAGLROS is increased in mantle cell lymphoma cells and may function as an oncogene in mantle cell lymphoma. HAGLROS may promote tumour development by regulating miR-100/ATG5/ PI3K/AKT/mTOR axis.
Introduction: AML1-ETO produced by t(8;21) abnomality has multiple effects on the leukemogenesis of acute myeloid leukemia (AML). SET domain, bifurcated 2 (SETDB2) can mediate gene silencing by trimethylation of the ninth lysine residue of histone H3 protein (H3K9) of the promoter and has been confirmed as an oncogene in many cancers. The role of SETDB2 in AML1-ETO positive AML is not clear. Methods: Quantitative reverse transcription PCR was performed to measure SETDB2 expression in bone marrow from AML patients and healthy people. Kaplan-Meier analysis was performed to investigate the effect of SETDB2 on prognosis of AML patients. Dual luciferase reporter gene assay, chromatin immunoprecipitation were performed to investigate the regulatory mechanism of AML1-ETO on SETDB2. CCK-8 and colony formation assay were performed to measure the effect of SETDB2 on leukemic cells. Results: SETDB2 is highly expressed in AML1-ETO positive AML. The overall survival, event-free and relapse-free survival rate of patients with high SETDB2 expression was lower than those of patients with low SETDB2 expression. SETDB2 is epigenetically upregulated by AML1-ETO fusion protein. Downregulation of SETDB2 expression significantly inhibits the proliferation and clonality of leukemic cells and promotes the sensitivity of leukemic cells to an epigenetic inhibitor JQ1. Conclusion: AML1-ETO/SETDB2 is a novel epigenetic pathway of leukemogenesis and SETDB2 is a potential therapeutic target of t(8;21) AML.
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