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A schematic overview of this study showing corneal stromal stem cell (CSSC) isolation from anterior limbal stromal tissue, ex vivo cell culture, extraction of extracellular vesicles (EV) to identify the expression of miR-29a and 381-5p by Nanostring assay. The anti-inflammatory and anti-fibrotic effects were determined by in vitro and in vivo assays. Screening of miR-29a expression in CSSC-EV distinguished CSSC with good anti-scarring quality for cell-based therapy of corneal scarring after injury. [Display omitted] • Corneal stromal stem cell (CSSC) therapy rectifies corneal scarring, a cause of global blindness. • Cell potency and quality control are needed to assure cell product safety for patient use. • Transcriptomic assay found microRNAs enriched in extracellular vesicles (EV) of healing CSSC. • EV express miR-29a/381 to reduce inflammation and fibrosis, indicating anti-scarring potency. • High miR-29a levels in EV distinguished CSSC with good anti-scarring quality for clinical use. Corneal blindness due to scarring is treated with corneal transplantation. However, a global problem is the donor material shortage. Preclinical and clinical studies have shown that cell-based therapy using corneal stromal stem cells (CSSCs) suppresses corneal scarring, potentially mediated by specific microRNAs transported in extracellular vesicles (EVs). However, not every CSSC batch from donors achieves similar anti-scarring effects. To examine miRNA profiles in EVs from human CSSCs showing "healing" versus "non-healing" effects on corneal scarring and to design a tool to select CSSCs with strong healing potency for clinical applications. Small RNAs from CSSC-EVs were extracted for Nanostring nCounter Human miRNA v3 assay. MicroRNAs expressed > 20 folds in "healing" EVs (P < 0.05) were subject to enriched gene ontology (GO) term analysis. MiRNA groups with predictive regulation on inflammatory and fibrotic signalling were studied by mimic transfection to (1) mouse macrophages (RAW264.7) for M1 phenotype assay; (2) human corneal keratocytes for cytokine-induced fibrosis, and (3) human CSSCs for corneal scar prevention in vivo. The expression of miR-29a was screened in additional CSSC batches and the anti-scarring effect of cells was validated in mouse corneal wounds. Twenty-one miRNAs were significantly expressed in "healing" CSSC-EVs and 9 miRNA groups were predicted to associate with inflammatory and fibrotic responses, and tissue regeneration (P <10−6). Overexpression of miR-29a and 381-5p significantly prevented M1 phenotype transition in RAW264.7 cells after lipopolysaccharide treatment, suppressed transforming growth factor β1-induced fibrosis marker expression in keratocytes, and reduced scarring after corneal injury. High miR-29a expression in EV fractions distinguished human CSSCs with strong healing potency, which inhibited corneal scarring in vivo. We characterized the anti-inflammatory and fibrotic roles of miR-29a and 381-5p in CSSCs, contributing to scar prevention. MiR-29a expression in EVs distinguished CSSCs with anti-scarring quality, identifying good quality cells for a scarless corneal healing. [ABSTRACT FROM AUTHOR]

Circulating miRNAs have been proposed as the effective diagnostic biomarkers for muscular fibrosis-associated diseases. However, circulating biomarkers for early diagnosis of contracture muscles are limited in gluteal muscle contracture (GMC) patients. Here we sought to explore the abnormally expressed miRNAs in plasma and contraction bands of GMC patients. The results showed miR-29a-3p expression in plasma and contraction bands tissue was significantly reduced in GMC patients compared with normal control. Cell viability and levels of proliferation-associated protein cyclin D1 and cyclindependent- kinase 2 (CDK2) were powerfully inhibited by miR-29a mimics and enhanced by miR-29a inhibitor compared with negative control. Furthermore, miR-29a mimics effectively impeded, while miR-29a inhibitor enhanced the expression of collagen I and collagen III, followed by the secretion of transforming growth factor β1 (TGF-β1), TGF-β3 and connective tissue growth factor (CTGF) in primary human contraction bands (CB) fibroblasts. The miR-29a-3p negatively regulated the expression of TGF-β1 through binding to the 3' UTR region of SERPINH1 (encoding heat shock protein HSP47), but had no effect on Smad2 activity. The miR-29a-3p was inversely correlated with HSP47 in contraction bands tissue from GMC patients. Collectively, miR-29a was notably depressed and regulated cell viability and fibrosis by directly targeting HSP47 in GMC, which suggest that circulating miR-29a might be a potential biomarker for early diagnosis and provides a novel therapeutic target for GMC. [ABSTRACT FROM AUTHOR]

OBJECTIVE: Oral squamous cell carcinoma (OSCC) comprises approximately ~90% of all oral malignancies and exhibits a significant mortality rate worldwide. Although the dysregulation of small nucleolar RNA host gene 20 (SNHG20) participates in the development of multiple malignancies, the molecular mechanisms underlying its regulation of OSCC progression remain to be fully elucidated. PATIENTS AND METHODS: The expression levels of SNHG20, microRNA-29a (miR-29a), and Disheveled-Axin Domain Containing 1 (DIXDC1) were detected by Real Time-quantitative Polymerase Chain Reaction (RT-qPCR). The protein expression levels of DIXDC1 and β-catenin were measured by Western blotting. In addition, MTT assay was performed to measure the cell proliferation ability in SCC9 and SCC15 cells. Cell migration and invasion abilities were measured by wound healing assay and transwell assay, respectively. The cell apoptosis was assessed by flow cytometry assay. Besides, Luciferase reporter assay was employed to examine the interrelation between miR-29a and SNHG20 or DIXDC1. RESULTS: It was demonstrated that SNHG20 and DIXDC1 were significantly upregulated in OSCC tissues and cell lines, while miR-29a was markedly downregulated. Moreover, the high expression of SNHG20 was found to predict a lower survival rate in OSCC patients. In addition, loss-of-function experiments demonstrated that SNHG20 knockdown inhibited the development and progression of OSCC, whereas the miR-29a inhibitor significantly abolished the effect of SNHG20 depletion on OSCC progression by directly binding to SNHG20. DIXDC1 was shown to enhance si-SNHG20 and miR-29a mimic-attenuated cell viability, migration, and invasion by directly binding to miR-29a. Furthermore, it was also found that DIXDC1 activated Wnt signaling in OSCC cells. CONCLUSIONS: Our study demonstrated that SNHG20 promoted OSCC progression via the miR-29a/DIXDC1/Wnt signaling pathway, which might provide a novel theoretical basis for the treatment of OSCC. [ABSTRACT FROM AUTHOR]

OBJECTIVE: The long non-coding ribonucleic acid (lncRNA)-myocardial infarction associated transcript (MIAT) has been demonstrated to serve as a key regulator in various physiological and pathological processes. This study aims to explore whether lncRNA-MIAT regulates the expression of micro RNA (miR)-29a to affect kidney’s injury in sepsis rats. MATERIALS AND METHODS: A total of 30 healthy male Sprague-Dawley (SD) rats were randomly divided into experimental group and control group. Rats in the experimental group were injected with lipopolysaccharide (LPS) through the tail vein to prepare the model of sepsis-induced kidney injury, while those in the control group with the equal volume of normal saline. After the levels of serum creatinine (SCr) and blood urea nitrogen (BUN) in rats were determined to ascertain successful modeling, fluorescence quantitative Real- time polymerase chain reaction (qRT-PCR) was performed to measure the expression levels of the lncRNA-MIAT and miR-29a messenger RNAs (mRNAs) in renal tissues. The normal rat kidney epithelial (NRK-52E) cell line was cultured in vitro, and the model was established in vitro via LPS to study the influences of lncRNA-MIAT and miR-29a on the kidney injury in sepsis rats. Moreover, cell apoptosis was detected using Western blotting. RESULTS: According to the results of the rat in vivo experiment and NRK-52E cell line in vitro experiment, the model of kidney injury was established successfully, and compared with the control group, experiment group had significantly raised SCr and BUN levels (p<0.01) and a remarkably increased lincRNA-MIAT gene expression level (p<0.01), but a substantially decreased miR-29a gene expression level (p<0.01). Additionally, when the expression of lncRNA-MIAT was up-regulated, the expression of miR-29a was prominently decreased (p<0.01), but that of the cell apoptosis gene cysteine-aspartic proteases (Caspase)-8 protein was remarkably increased (p<0.01). However, the expression of Caspase-8 protein was significantly lowered (p<0.01) once the expression of miR-29a was up-regulated. CONCLUSIONS: LncRNA-MIAT may bind to miR-29a to participate in sepsis-related kidney injury. [ABSTRACT FROM AUTHOR]

OBJECTIVE: Abnormal expression of circular RNAs (circRNAs) has been observed in various biological processes and cancer pathogenesis. However, the expression of circRNAs in pediatric acute myeloid leukemia (AML) remains largely unknown so far. PATIENTS AND METHODS: Twelve bone marrow samples from pediatric AML patients and healthy controls were analyzed using Agilent circRNA microarray (n = 6, respectively). The circRNAs profiles and regulatory networks were analyzed by integrated bioinformatics methods. Functional analysis (Gene Ontology and KEGG) was performed by KOBAS. The expression of circRNA in patient samples was validated via qRT-PCR assay (n > 30). Luciferase reporter assay was performed to validate the binding of miRNAs. CCK8 and colony formation assay were conducted to measure cell proliferation. RESULTS: A total of 273 circRNAs were upregulated in AML and 296 were downregulated (Fold change > 2, p-value < 0.05), the majority of these circRNAs were distributed among chr1, chr6, and chr16, while few in chr13 and chr21. Top 20 differentially expressed circRNAs were chosen to build circRNAs-miRNAs regulatory relationships. Bioinformatics algorithms indicated that circ-0004136 acts as a sponge for several pediatric AML-related miRNAs. Target genes involved in the circ0004136-miRNA-mRNA network were enriched in leukemia-related functions and signaling pathways. Circ-0004136 was found to be significantly upregulated in pediatric AML and could sponge AML-related miRNAs, including miR-29a and miR-142. Furthermore, circ-0004136 was demonstrated to promote the proliferation of AML by sponging miR-142. CONCLUSIONS: Taken together, this study revealed the circRNAs expression profile and regulatory networks of circRNAs-miRNAs-mRNAs in pediatric AML for the first time. Circ-0004136 was significantly upregulated in pediatric AML and could promote cell proliferation by sponging miR-142. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To evaluate the effects of miR-29a on the high glucose (HG)-induced apoptosis and the correlation between miR-29a and heme oxygenase-1 (HO-1) and the underlying molecular mechanism. MATERIALS AND METHODS: The cell apoptosis was analyzed by the flow cytometry, and the cells autophagy was evaluated using the transmission electron microscopy. Luciferase reporter assay was carried out to detect the correlation between miR-29a and HO-1. Besides, reverse transcription-PCR and Western blot were applied to detect the mRNA and protein levels. RESULTS: The expression of miR-29a was significantly decreased in the HG-treated podocytes. Besides, miR-29a overexpression could promote cellular autophagy and significantly reduce HG-induced podocytes apoptosis. Moreover, HO-1 was a direct target of miR-29a and the pre-autophagy and the anti-apoptotic effects of miR-29a on HG-treated podocytes could be significantly reversed by the HO-1 siRNA administration. CONCLUSIONS: MiR-29a functionally promoted podocytes autophagy and inhibited apoptosis through the HO-1dependent pathway in the HG condition. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To investigate the biological role of micro-ribonucleic acid (miR)-29a in non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS: 55 cases of NSCLC tissue specimens and paired normal lung tissue specimens collected in the Department II of Oncology, Ruikang Hospital Affiliated to Guangxi University of Chinese Medicine from July 2012 to April 2015 were randomly included. The fluorescent quantitative polymerase chain reaction, Western blotting, and immunohistochemistry were performed to detect the expression levels of miR-29a and metastasis suppressor 1 (MTSS1). Pearson correlation analysis was utilized to investigate the relationship between miR-29a expression and MTSS1 expression in NSCLC tissues, and the Kaplan-Meier survival curves were constructed to analyze the association of miR-29a expression with the survival time of NSCLC patients. A54 proliferation and invasion abilities were measured by means of plate clone formation assay, and transwell assay after the miR-29a was suppressed by miRNA inhibitor. Luciferase assay was used to detect the target gene of miR-29a. RESULTS: In NSCLC tissues, the miR-29a expression level was higher than that in normal lung tissues (p<0.05), while the expression level of MTSS1 protein was remarkably lower than that in normal lung tissues (p<0.05). The median survival time of the patients was 15.1 months in high miR-29a expression group and 18.3 months in low miR-29a expression group (p<0.05). The miR-29a expression was negatively correlated with the expression level of MTSS1 protein in NSCLC tissues (r=-0.762, p<0.05). Luciferase results suggest that miR-29a binds to the promoter region of MTSS1 and inhibits its transcription level. The expression of MTSS1 protein was up-regulated notably after miR-29a knockdown by an inhibitor. It was revealed in the results of transwell assay and plate clone formation assay that the proliferative and invasive capacity of A549 cells was significantly decreased after knockdown of miR-29a. CONCLUSIONS: The transcribed miR-29a down-regulates the protein level of MTSS1, suppressor of tumor proliferation and invasion, thereby promoting the proliferative and invasive capacity of NSCLC cells. Both miR-29a and MTSS1 are expected to become potential therapeutic targets for NSCLC. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To investigate the role of miR-29a in regulating the differentiation mesenchymal stem cells into osteoblasts. MATERIALS AND METHODS: For the first step, the changes of expression of miR-29a during the process of mesenchymal stem cells (MSCs) differentiation into osteoblast were detected. Then, we infected the MSCs with mimics or inhibitors of miR-29a to explore the roles of miR-29a in the differentiation. Further, the prediction and verification of the possible target genes of miR-29a were achieved by bioinformatics analysis and luciferase reporter assay. RESULTS: MiR-29a was up-regulated during the process of MSCs differentiation into osteoblasts. Overexpression or inhibition of miR-29a using mimics or inhibitors had no significant effect on cell proliferation. Furthermore, the differentiation was enhanced when miR-29a was artificially overexpressed in vitro, whereas silencing of miR-29a attenuated this process. It was evidenced by alkaline phosphatase (ALP) staining, matrix mineralization, and increased expression of osteoblast-specific genes. Furthermore, we determined that the gene HDAC4 might be a direct target of miR-29a. CONCLUSIONS: In the current study, miR- 29a promotes osteogenesis via suppressing HDAC4, indicating that targeting miR-29a may be feasible in the management of osteoporosis. [ABSTRACT FROM AUTHOR]

OBJECTIVE: The fractional laser and topical retinoic acid treatment have been applied for skin rejuvenation; however, the possible molecular mechanism of promoting remodeling of dermis is not clearly. Here we aimed to compare the effects of 10600 nm CO2 fractional laser and topical retinoic acid formulation on the skin collagen proliferation of Wistar rats, and to further explore the possible molecular mechanism of promoting remodeling of dermis. MATERIALS AND METHODS: The hair on the back of Wistar rats was removed, and the back was divided equally into four regions with the cross-streaking method: A (the control group), B (the retinoic acid group), C (retinoic acid and fractional laser combination treatment group), and D (the fractional laser group). Specimens were collected at 3rd day and in 1-8 weeks after CO2 fractional laser irradiation; then they were used for detection of the changes of dermis thickness and content of hydroxyproline in the four regions of the rats; back. Real-time PCR method was used to detect the dynamic changes of the expression level of type III procollagen mRNA and the expression levels of miR-29a, Akt and transforming growth factor-β (TGF-β) mRNA at 3rd week in the skin tissue of Wistar rats. RESULTS: The thickness of dermis, content of hydroxyproline and expression level of type III procollagen mRNA in the treatment groups (B, C, and D) were found all significantly increased compared with those in the control group (A) (p<0.05); at 3rd week, up-regulation of Akt and TGF-β mRNA expression and down-regulation of miR-29a mRNA expression were observed in the treatment groups (B, C, and D). The difference in the combination treatment group (C) was the most significant (p<0.05). CONCLUSIONS: These results demonstrate that retinoic acid formulation and CO2 fractional laser both can promote collagen proliferation and reconstruction, with the skin rejuvenation efficacy in group C > group D > group B. miR-29a/Akt/TGF-β signal pathways may play a certain role in the promotion of collagen synthesis and proliferation. [ABSTRACT FROM AUTHOR]

Atherosclerotic cardiovascular diseases, such as coronary heart disease, have become a major public health problem all over the world. MicroRNA-29a (miR-29a) modulates expression levels of collagen, inflammatory reaction and other extracellular matrix mRNAs, while adiponectin (APN), a circulating protein secreted by adipocytes, has anti-inflammatory properties. Both play multifaceted roles in angiogenesis or vascular remodelling. However, little is known about plasma miR-29a and APN levels in patients with atherosclerosis. We therefore investigated the relationship between the plasma levels of miR-29a or APN and carotid intima-media thickness (cIMT) in atherosclerosis patients (n = 85, cIMT ≥ 1.2 mm) and the controls (n = 85, cIMT < 1.2 mm). We found that the atherosclerosis group showed higher miR-29a levels (31.15 ± 3.99 vs. 26.39 ± 1.05 Ct, P < 0.001) and lower APN levels (15.93 ± 4.61 vs. 21.80 ± 7.74 ng/ml, P < 0.001), compared with control group. Thus, increased cIMT was associated with higher plasma miR-29a levels (r = 0.688, P < 0.001) and with lower plasma APN levels (r = -0.494, P < 0.001). Furthermore, multiple logistic regression analysis indicated that higher miR-29a levels (OR: 1.136, 95% CI: 1.042-1.240, P = 0.004) increased the risk for atherosclerosis, whereas higher APN levels appeared to be protective (OR: 0.122, 95% CI: 0.055-0.271, P < 0.001). The present study indicates that elevated miR-29a levels and reduced APN levels are associated with atherosclerosis. [ABSTRACT FROM AUTHOR]

The article presents a study which aims to determine the effect of microRNA expression in the neuronal dysfunction caused by rabies virus (RABV) infection using the bioinformatics and gene analysis method.

We previously reported that aluminum (Al) can cause a range of neurotoxic injuries including progressive irreversible synaptic structural damage and synaptic dysfunction, and eventually neuronal deaths. Mechanism of Al-induced electrophysiological and neuronal connectivity changes in neurons may indicate damage to the neuronal network. Here, mouse primary hippocampal neurons were cultured on micro-electrode array (MEA)- and high-content analysis (HCA)-related plates, showing that Al exposure significantly inhibited hippocampal neuronal electrical spike activity and neurite outgrowth characterized by a reduction in neurite branching and a decrease in the average total neurite length in relation to both Al dose and time of incubation. In recent years, miR-29a/ phosphatase and tensin homolog (PTEN) have been found to play pivotal roles in the morphogenesis of neurons, it has been confirmed in vitro and in vivo that the PTEN–Glycogen synthase kinase-3β (GSK-3β) axis regulates neurite outgrowth. The present study demonstrated that increases in Al exposure and dose gradually reduce miR-29a expression. Up-regulation of miR-29a in the hippocampal neurons by lentivirus transfection reversed the decrease in electrical spike activity and the reduction in both neurite branching and length induced by Al. Moreover, miR-29a suppressed the expression of PTEN and increased the level of phosphorylated Protein Kinase B (p-AKT) and p-GSK-3β which were inhibited by the Al treatment. This suggests that miR-29a is critically involved in the functional and structural neuronal damage induced by Al and is a potential target for Al neurotoxicity. Moreover, the reduction of neurite length and branching induced by Al exposure was regulated by miR-29a and its target neuronal PTEN–GSK3β signaling pathway, which also represents a possible mechanism of Al-induced the inhibition of the electrical activity. Collectively, Al-induced damage to the neuronal network occurred through miR-29a-mediated alterations of the PTEN–GSK3β signaling pathway. [Display omitted] • Al exposure significantly inhibited hippocampal neuronal electrical spike activity and neurite outgrowth in relation to both Al dose and time of incubation. • Al suppressed miR-29a expression dose and time dependently. • Al-induced damage to the neuronal network occurred through miR-29a-mediated alterations of PTEN–GSK3β signaling pathway. [ABSTRACT FROM AUTHOR]