MiR-568 mitigated cardiomyocytes apoptosis, oxidative stress response and cardiac dysfunction via targeting SMURF2 in heart failure rats.

Chronic heart failure (CHF), a conventional, complex, and severe syndrome, is generally defined by myocardial output inadequate to satisfy the metabolic requirements of body tissues. Recently, miR-568 was identified to be down-regulated in CHF patients' sera and negatively correlated with left ventricular mass index in symptomatic CHF patients with systolic dysfunction. Nevertheless, the role of miR-568 during CHF development remains obscure. The current study is aimed to investigate the role of miR-568 in CHF. The MTT assay, flow cytometry analysis, RT-qPCR analysis, western blot analysis and luciferase reporter assays were conducted to figure out the function and potential mechanism of miR-568 in vitro. Rats were operated with aortic coarctation to establish CHF animal model. The effects of miR-568 and SMURF2 on CHF rats were evaluated by hematoxylin-eosin staining, Masson's staining, serum index testing, cardiac ultrasound detection, and TUNEL staining assays. We discovered that miR-568 level was downregulated by H2O2 treatment in cardiomyocytes. In mechanism, miR-568 directly targeted and negatively regulated SMURF2. In function, SMURF2 overexpression reversed the effects of miR-568 on cardiac function and histological changes in vivo. Additionally, SMURF2 overexpression reversed the effects of miR-568 on the content of LDH, AST, CK and CK-MB in vivo. Moreover, SMURF2 overexpression reversed the effects of miR-568 on oxidative stress response in vivo. MiR-568 mitigated cardiomyocytes apoptosis, oxidative stress response and cardiac dysfunction via targeting SMURF2 in CHF rats. This discovery may serve as a potential biomarker for CHF treatment.

MiR-568 inhibits the activation and function of CD4⁺ T cells and Treg cells by targeting NFAT5.

CD4(+) T cells play critical roles in orchestrating adaptive immune responses. Their activation and proliferation are critical steps that occur before they execute their biological functions. Despite the important role of this process, the underlying molecular events are not fully understood. MicroRNAs (miRNAs) have been shown to play important roles in lymphocyte development and function. However, the miRNAs that regulate T-cell differentiation, activation and proliferation are still largely unknown. In our previous study, using a miRNA array, we found that several miRNAs (including miR-202, 33b, 181c, 568 and 576) are differentially expressed between resting and activated CD4(+) T cells. In this study, we focused on the function of miR-568 during CD4(+) T-cell activation. We showed that the expression level of miR-568 decreased during the activation of T cells, including Jurkat cells and human peripheral blood CD4(+) T cells. When Jurkat or human peripheral blood CD4(+) T cells were transfected with miR-568 mimics, cell activation was significantly inhibited, as shown by the inhibited expression of activation markers such as CD25, CD69 and CD154; decreased IL-2 production; and inhibited cell proliferation. Using software predictions and confirmatory experiments, we demonstrated that nuclear factor of activated T cells 5 (NFAT5) is a target of miR-568. Treg cells are an important CD4(+) T-cell subpopulation, so we also evaluated the function of miR-568 in Treg-cell activation and differentiation. We showed that the miR-568 level decreased, while the NFAT5 protein level increased during CD4(+)CD25(+) Treg-cell activation, and the transfection of miR-568 mimics inhibited the NFAT5 expression, inhibited the production of both TGF-ß and IL-10 and also inhibited the proliferation of Treg cells. Our further study showed that over-expression of miR-568 can inhibit Treg-cell differentiation and can inhibit the suppressive effect of these cells on effector cells. In addition, inhibition of NFAT5 by siRNA-mediated knockdown can inhibit the activation and differentiation of Treg cells. These findings reveal that miR-568 can inhibit the activation and function of both CD4(+) T cells and Treg cells by targeting NFAT5. Since miR-568 plays an important role in both CD4(+) T cells and Treg cells, these findings may provide leads for the development of novel treatments for human inflammatory and autoimmune diseases.

LINC00680 enhances hepatocellular carcinoma stemness behavior and chemoresistance by sponging miR-568 to upregulate AKT3.

BACKGROUND: Hepatocellular carcinoma (HCC) has an extremely poor prognosis due to the development of chemoresistance, coupled with inherently increased stemness properties. Long non-coding RNAs (LncRNAs) are key regulators for tumor cell stemness and chemosensitivity. Currently the relevance between LINC00680 and tumor progression was still largely unknown, with only one study showing its significance in glioblastoma. The study herein was aimed at identifying the role of LINC00680 in the regulation HCC stemness and chemosensitivity. METHODS: QRT-PCR was used to detect the expression of LINC00680, miR-568 and AKT3 in tissue specimen and cell lines. Gain- or loss-of function assays were applied to access the function of LINC00680 in HCC cells, including cell proliferation and stemness properties. HCC stemness and chemosensitivity were determined by sphere formation, cell viability and colony formation. Luciferase reporter, RNA immunoprecipitation (RIP), and RNA pull-down assays were performed to examine the interaction between LINC00680 and miR-568 as well as that between miR-568 and AKT3. A nude mouse xenograft model was established for the in vivo study. RESULTS: We found that LINC00680 was remarkably upregulated in HCC tissues. Patients with high level of LINC00680 had poorer prognosis. LINC00680 overexpression significantly enhanced HCC cell stemness and decreased in vitro and in vivo chemosensitivity to 5-fluorouracil (5-Fu), whereas LINC00680 knockdown led to opposite results. Mechanism study revealed that LINC00680 regulated HCC stemness and chemosensitivity through sponging miR-568, thereby expediting the expression of AKT3, which further activated its downstream signaling molecules, including mTOR, elF4EBP1, and p70S6K. CONCLUSION: LINC00680 promotes HCC stemness properties and decreases chemosensitivity through sponging miR-568 to activate AKT3, suggesting that LINC00680 might be a potentially important HCC diagnosis marker and therapeutic target.