Silencing of Long Noncoding RNA TUG1 Ameliorates Atherosclerosis-Induced Myocardial Injury by Upregulating microRNA-30b-3p and Downregulating Brd4.

ABSTRACT: Long noncoding RNAs and microRNAs (miRNAs) are emerging biomarkers involved in human diseases, and we focused on the roles of long noncoding RNA taurine upregulated gene 1 (TUG1) and miR-30b-3p in the related mechanisms of atherosclerosis-induced myocardial injury. ApoE-deficient mice were fed with high-fat diet to establish atherosclerotic models and then were subjected to either TUG1 downregulation or miR-30b-3p upregulation treatment. The serum myocardial enzymes, inflammatory biomarkers, pathological changes, intramyocardial macrophage infiltration, and apoptosis of cardiomyocytes in atherosclerotic mice were determined. The expression of TUG1, miR-30b-3p, and bromodomain protein 4 (Brd4) in atherosclerotic models was evaluated. Moreover, the correlations of TUG1, miR-30b-3p, and Brd4 were verified. TUG1 and Brd4 were increased while miR-30b-3p was decreased in atherosclerotic mice. The silenced TUG1 or elevated miR-30b-3p attenuated atherosclerosis-induced myocardial injury mainly by reducing serum myocardial enzyme content and inflammatory response, improving pathological changes, and preventing macrophage infiltration and cardiomyocyte apoptosis in atherosclerotic mice. Mechanistically, TUG1 could competitively bind with miR-30b-3p to prevent the degradation of its target gene Brd4. This study reveals that the silencing of TUG1 ameliorates atherosclerosis-induced myocardial injury by upregulating miR-30b-3p and downregulating Brd4, which may provide novel targets for atherosclerosis treatment.

Role of miRNA-99a-5p in Modulating the Function of Hepatocellular Carcinoma Cells: Bioinformatics Analysis and In Vitro Assay.

AIM: This study aimed to investigate the biological functions of miRNAs in hepatobiliary tumors as the focus of targeted therapy research. BACKGROUND: Hepatobiliary tumors are among the leading causes of cancer-related deaths worldwide. Many microRNAs (miRNAs) play an important regulatory role in tumor progression. Our study aims to explore some biologically functional miRNAs from different datasets of hepatobiliary tumors for disease diagnosis or treatment. OBJECTIVE: In this study, we tried to filter out differentially expressed miRNAs in different tumor datasets from the GEO database. METHODS: In this study, we first perform analyses in different GEO data sets. After taking the intersection, the initial scope is limited to several differential RNAs. Then, combined with the existing research results from Kaplan-Meier survival analysis and literature, the candidate molecule was finally identified to be studied. Furthermore, the biological characteristics analysis of the candidate molecule was performed on the basis of Cancermirnome online tool, including expression levels in tumors, KEGG and GO analysis, ROC analysis, and target gene prediction. Furthermore, the effect of the candidate molecule on the biological functions of liver cancer was verified by In Vitro assay. RESULTS: The preliminary analysis of bioinformatics shows that 16 differentially expressed miRNAs may play an important role in HCC or ICC. Ultimately, we identified miRNA-99a-5p as the only molecule to study. The results showed that miRNA-99a-5p is abnormally expressed in many tumors, and in liver cancer, its level of expression in tumor tissue is significantly lower than that in normal tissue. Then, the KEGG and GO analysis found that it functions in multiple pathways. At the same time, the ROC analysis found that it showed great potential for prognostic prediction in HCC and we also predicted that RUNDC3B is the most likely target to which it binds. Finally, the experimental results of overexpression and knockdown confirmed that miRNA-99a-5p could inhibit cell proliferation in HCC, which also suggested that it may be an important tumor suppressor in HCC. CONCLUSION: MiRNA-99a-5p was negatively correlated with HCC progression and could act as a novel therapeutic target for HCC.

Let-7a regulates EV secretion and mitochondrial oxidative phosphorylation by targeting SNAP23 in colorectal cancer.

BACKGROUND: MicroRNAs (miRNAs) are abundant in tumor-derived extracellular vesicles (EVs) and the functions of extracellular miRNA to recipient cells have been extensively studied with tumorigenesis. However, the role of miRNA in EV secretion from cancer cells remains unknown. METHODS: qPCR and bioinformatics analysis were applied for determining extracellular let-7a expression from CRC patient serum and cells. Nanosight particle tracking analysis was performed for investigating the effect of let-7a on EV secretion. Luciferase reporter assays was used for identifying targeted genes synaptosome-associated protein 23 (SNAP23). In vitro and in vivo assays were used for exploring the function of let-7a/SNAP23 axis in CRC progression. Bioenergetic assays were performed for investigating the role of let-7a/SNAP23 in cellular metabolic reprogramming. RESULTS: let-7a miRNA was elevated in serum EVs from CRC patients and was enriched in CRC cell-derived EVs. We determined that let-7a could suppress EV secretion directly targeting SNAP23. In turn, SNAP23 promotes EV secretion of let-7a to downregulate the intracellular let-7a expression. In addition, we found a novel mechanism of let-7a/SNAP23 axis by regulating mitochondrial oxidative phosphorylation (OXPHOS) through Lin28a/SDHA signaling pathway. CONCLUSIONS: Let-7a plays an essential role in not only inhibiting EV secretion, but also suppressing OXPHOS through SNAP23, resulting in the suppression of CRC progression, suggesting that let-7a/SNAP23 axis could provide not only effective tumor biomarkers but also novel targets for tumor therapeutic strategies.

Metformin Improves Functional Recovery After Spinal Cord Injury via Autophagy Flux Stimulation.

Spinal cord injury (SCI) is a severe neurological disease with few efficacious drugs. Autophagy is a cellular process to confront with stress after SCI and considered to be a therapeutic target of SCI. In this study, we investigated the therapeutic effect of metformin on functional recovery after SCI and its underlying mechanism of autophagy regulation. Using a rat model of traumatic SCI, we found improved function recovery which was paralleled by a reduction of apoptosis after metformin treatment. We further examined autophagy via detecting autophagosomes by transmission electron microscopy and immunofluorescence, as well as autophagy markers by western blot in each groups. The results showed that the number of autophagosomes and expression of autophagy markers such as LC3 and beclin1 were increased in SCI group, while autophagy substrate protein p62 as well as ubiquitinated proteins were found to accumulate in SCI group, indicating an impaired autophagy flux in SCI. But, metformin treatment attenuated the accumulation of p62 and ubiquitinated proteins, suggesting a stimulative effect of autophagy flux by metformin. Blockage of autophagy flux by chloroquine partially abolished the apoptosis inhibition and functional recovery effect of metformin on SCI, which suggested that the protective effect of metformin on SCI was through autophagy flux stimulation. Activation of AMPK as well as inhibition of its downstream mTOR signaling were detected under metformin treatment in vivo and in vitro; inhibition of AMPK signaling by compound C suppressed autophagy flux induced by metformin in vitro, indicating that AMPK signaling was involved in the effect of metformin on autophagy flux regulation. Together, these results illustrated that metformin improved functional recovery effect through autophagy flux stimulation and implied metformin to be a potential drug for SCI therapy.

Regulation of Caveolin-1 and Junction Proteins by bFGF Contributes to the Integrity of Blood-Spinal Cord Barrier and Functional Recovery.

The blood-spinal cord barrier (BSCB) plays important roles in the recovery of spinal cord injury (SCI), and caveolin-1 is essential for the integrity and permeability of barriers. Basic fibroblast growth factor (bFGF) is an important neuroprotective protein and contributes to the survival of neuronal cells. This study was designed to investigate whether bFGF is beneficial for the maintenance of junction proteins and the integrity of the BSCB to identify the relations with caveolin-1 regulation. We examined the integrity of the BSCB with Evans blue dye and fluorescein isothiocyanate-dextran extravasation, measured the junction proteins and matrix metalloproteinases, and evaluated the locomotor function recovery. Our data indicated that bFGF treatment improved the recovery of BSCB and functional locomotion in contusive SCI model rats, reduced the expression and activation of matrix metalloproteinase-9, and increased the expressions of caveolin-1 and junction proteins, including occludin, claudin-5, p120-catenin, and ß-catenin. In the brain, in microvascular endothelial cells, bFGF treatment increased the levels of junction proteins, caveolin-1 small interfering RNA abolished the protective effect of bFGF under oxygen-glucose deprivation conditions, and the expression of fibroblast growth factor receptor 1 and co-localization with caveolin-1 decreased significantly, which could not be reversed by bFGF treatment. These findings provide a novel mechanism underlying the beneficial effects of bFGF on the BSCB and recovery of SCI, especially the regulation of caveolin-1.