Retracted: LncRNA-ROR alleviates hypoxia-triggered damages by downregulating miR-145 in rat cardiomyocytes H9c2 cells

Chronic hypoxic heart disease (CHD) is a common clinical type of congenital heart disease. Long noncoding RNA regulator of reprogramming (lncRNA-ROR) exerts an important regulating effect in cardiovascular diseases. In our study, we explored the effect of lncRNA-ROR and the possible mechanisms against hypoxia-caused apoptosis in H9c2 cells. H9c2 cells were exposed to hypoxia (1% O₂) to construct the in vitro model of CHD. The level of lncRNA-ROR and microRNA (miRNA/miR)-145 was detected. To upregulate the level of lncRNA-ROR and miR-145, transfection was carried out. Western blot assay was performed to quantified protein expression. The interaction of lncRNA-ROR with miR-145 was verified by RIP and Dual-luciferase reporter assays. The expression of p53 and Bax was largely elevated and Bcl-2 was suppressed by hypoxia induction. We found that lncRNA-ROR was elevated by hypoxia. LncRNA-ROR overexpression was able to relieve the damages of H9c2 cells induced by hypoxia through rescuing viability, suppressing apoptosis, and blocking Cytochrome c release. miR-145 was suppressed by overexpressed lncRNA-ROR and the combination of miR-145 mimic was able to abolish the protective effect of lncRNA-ROR. Moreover, we found that lncRNA-ROR activated Ras/Raf/MEK/ERK and PI3K/AKT transduction cascades by suppressing miR-145. Besides, lncRNA-ROR directly targeted miR-145 and negatively modulated the level of miR-145. Our present study revealed that lncRNA-ROR protected H9c2 cells against hypoxia-caused damages by regulation of miR-145 through activating Ras/Raf/MEK/ERK and PI3K/AKT.     

Estrogen inhibits osteoclasts formation and bone resorption via microRNA-27a targeting PPARγ and APC

Inhibition of osteoclasts formation and bone resorption by estrogen is very important in the etiology of postmenopausal osteoporosis. The mechanisms of this process are still not fully understood. Recent studies implicated an important role of microRNAs in estrogen-mediated responses in various cellular processes, including cell differentiation and proliferation. Thus, we hypothesized that these regulatory molecules might be implicated in the process of estrogen-decreased osteoclasts formation and bone resorption. Western blot, quantitative real-time polymerase chain reaction, tartrate-resistant acid phosphatase staining, pit formation assay and luciferase assay were used to investigate the role of microRNAs in estrogen-inhibited osteoclast differentiation and bone resorption. We found that estrogen could directly suppress receptor activator of nuclear factor B ligand/macrophage colony-stimulating factor-induced differentiation of bone marrow-derived macrophages into osteoclasts in the absence of stromal cell. MicroRNA-27a was significantly increased during the process of estrogen-decreased osteoclast differentiation. Overexpressing of microRNA-27a remarkably enhanced the inhibitory effect of estrogen on osteoclast differentiation and bone resorption, whereas which were alleviated by microRNA-27a depletion. Mechanistic studies showed that microRNA-27a inhibited peroxisome proliferator-activated receptor gamma (PPARγ) and adenomatous polyposis coli (APC) expression in osteoclasts through a microRNA-27a binding site within the 3′-untranslational region of PPARγ and APC. PPARγ and APC respectively contributed to microRNA-27a-decreased osteoclast differentiation and bone resorption. Taken together, these results showed that microRNA-27a may play a significant role in the process of estrogen-inhibited osteoclast differentiation and function.     

MiR-26b-3p regulates osteoblast differentiation via targeting estrogen receptor α

BackgroundUnderstanding of the molecular mechanisms of miRNAs involved in osteoblast differentiation is important for the treatment of bone-related diseases.MethodsMC3T3-E1 cells were induced to osteogenic differentiation by culturing with bone morphogenetic protein 2 (BMP2). After transfected with miR-26b-3p mimics or inhibitors, the osteogenic differentiation of MC3T3-E1 cells was detected by ALP and ARS staining. Cell viability was analyzed by MTT. The expressions of miR-26b-3p and osteogenic related markers and signaling were examined by qPCR and western blot. Direct binding of miR-26b-3p and ER-α were determined by dual luciferase assay.ResultsmiR-26b-3p was significantly down-regulated during osteoblast differentiation. Overexpression of miR-26b-3p inhibited osteoblast differentiation, while inhibition of miR-26b-3p enhanced osteoblast differentiation. Further studies demonstrated miR-26b-3p inhibited the expression of estrogen receptor α (ER-α) by directly targeting to the CDS region of ER-α mRNA. Overexpression of ER-α rescued the suppression effects of miR-26b-3p on osteoblast differentiation, while knockdown of ER-α reversed the upregulation of osteoblast differentiation induced by knockdown of miR-26b-3p.ConclusionOur study demonstrates that miR-26b-3p suppresses osteoblast differentiation of MC3T3-E1 cells via directly targeting ER-α.     

Low expression of miR-186-5p regulates cell apoptosis by targeting toll-like receptor 3 in high glucose–induced cardiomyocytes

To investigate the effect and mechanism of microRNA-186-5p (miR-186-5p) on the apoptosis in high glucose (HG)–treated cardiomyocytes. Diabetic cardiomyopathy model was established in cardiomyocytes by stimulating with HG. The expressions of miR-186-5p and toll-like receptor 3 (TLR3) were detected by quantitative polymerase chain reaction or Western blot analysis, respectively. Apoptosis was detected in HG-treated cardiomyocytes by flow cytometry and Western blot analysis. The interaction between miR-186-5p and TLR3 was explored by bioinformatics analysis and luciferase activity assay. Results showed that miR-186-5p expression was downregulated in HG-treated cardiomyocytes and its overexpression reversed HG-induced apoptosis and cleaved caspase-3 protein expression. Moreover, TLR3 was indicated as a target of miR-186-5p and regulated by miR-186-5p. Knockdown of TLR3 suppressed HG-induced apoptosis and cleaved caspase-3 protein expression. Besides, restoration of TLR3 ablated the effect of miR-186-5p on cell apoptosis. Collectively, miR-186-5p attenuated HG-induced apoptosis by regulating TLR3 in cardiomyocytes, providing novel biomarker for treatment of diabetic cardiomyopathy.