Objective To detect traces of microRNAs (miRNAs) in plasma and assess the expression stability of two common reference genes by stem-loop and poly A polymerase (PAP) real-time quantitative polymerase chain reaction (PCR) method, as miRNAs are the new bio-markers of tumor diagnosis and molecular targeted therapy, and its quantitative research is very important. Methods We extracted miRNAs from plasma of adult Sprague Dawley (SD) rats’ plasma, and detected the expressions of rno-miR-200b-3p and rno-miR-126-3p with stem-loop and PAP real-time PCR quantitative method, with rno-miR-103a-3p and U6 as internal controls. All the results were evaluated by 2△Ct method. Results Compared with PAP method, the stem-loop method reduced Ct value by 2-4 cycles and improved sensitivity by 10 times. In PAP method, the melting curve showed two peaks, a main peak and a small non-specific peak. Yet the melting curve of stem-loop method demonstrated a single specific peak. Furthermore, we validated the stability of internal references in the two real time PCR methods. U6 presented a more stable Ct value than rno-miR-103 in adult SD rats’ plasma samples. Conclusions Stem-loop real-time PCR is recommended as a major way to detect some samples with a low concentration of miRNAs, owing to its high accuracy and sensitivity. However, if a large number of tissue samples is going to be detected, PAP real-time PCR is more suitable and convenient than stem-loop method. U6 is more stable and repeatable than rno-miR-103a-3p as the reference gene to evaluate the semi-quantitative consequence of miRNAs.
ObjectiveTo investigate the value of plasma microRNA-216 (miR-216) in patients with acute pancreatitis as a clinical biomarker to early identify severe acute pancreatitis (SAP).MethodsPatients with acute pancreatitis who admitted to the hospital within 48 hours after the onset of disease between September and November 2014 were enrolled in this study. Plasam and clinical data of all the patients were collected. MiR-216 in the plasma was detected using quantitative real time-polymerase chain reaction.ResultsA total of 25 patients were enrolled. The Ct value of plasma miR-216 in SAP patients (32.40±1.43) was significantly upregulated than mild acute pancreatitis (MAP) (35.85±1.91, P<0.05) and moderately severe acute pancreatitis (MSAP) patients (35.90±2.44,P<0.05), respectively. The area under receiver operating characteristic curve for plasmamiR-216 in predicting SAP was 0.792 (P<0.05), which did not differ much from other conventional parameters such as C-reactive protein, urinary nitrogen, and cytokines (P>0.05).ConclusionPlasma miR-216 is significantly upregulated in SAP patients compared with MAP and MSAP, but it shows no inferior efficiency than the investigated conventional predictors in predicting SAP.
Recent advances in epigenetics indicate that several epigenetic modifications, including acetylation, methylatio, and microRNA (miRNA), play an important role in the pathogenesis of acute kidney injury (AKI). Our study reveales that enhancement of protein acetylation by pharmacological inhibition of class I histone deacetylases leads to more severe tubular injury, and delays the restoration of renal structure and function. The changes in promoter DNA methylation occurs in the kidney with ischemia/reperfusion. MiRNA expression is associated with the regulation of both renal injury and regeneration after AKI. Targeting the epigenetic process may provide a therapeutic treatment for patients with AKI. The purpose of this review is to summarize recent advances in epigenetic regulation of AKI and provide mechanistic insight into the role of acetylation, methylation, and miRNA expression in the pathological processes of AKI.
Circular RNAs (circRNAs) are a novel class of non-coding RNAs, which are more stable than linear RNAs for their closed circular structure by covalent bond. CircRNAs exist in a large variety of cells and regulate the expressions of target genes. Moreover, circRNAs are closely related to various diseases and have a potential value as biomarkers and prognostic markers clinically. In this article, the classification and biological functions of circRNA molecules (including being as microRNA sponges, regulating gene transcription, regulating RNA binding protein and the potential translation function) are summarized, and the latest research progress of circRNAs in rheumatoid arthritis is reviewed.
ObjectiveTo investigate the effect of curcumin on lipopolysaccharide (LPS)-induced inflammation and apoptosis in alveolar macrophage via microRNA-132 (miR-132)/high mobility group protein B1 (HMGB1).MethodsThe cultured mouse alveolar macrophage line (RAW264.7 cells) were divided into the control group, the LPS group, the LPS+50 μmol/L curcumin group, and the LPS+100 μmol/L curcumin group. Forty-eight hours after drug treatment, the levels of miR-132/HMGB1, inflammatory mediator and apoptotic were detected. Secondly, the empty vector, synthetic miR-132 mimics and inhibitors were transfected into another cultured mouse alveolar macrophage line (RAW264.7 cells) to detect the inflammation and apoptosis of alveolar macrophage after transfection.ResultsCompared with the control group, in the LPS group, the apoptosis of alveolar macrophage, the levels of interleukin (IL)-6, IL-8 and tumor necrosis factor (TNF)-α, and the expression of miR-132 increased, while the expression of HMGB1 decreased (P<0.05); compared with the LPS group, in the two curcumin groups, the apoptosis of alveolar macrophage, the levels of IL-6, IL-8 and TNF-α, and the expression of miR-132 decreased, while the expression of HMGB1 increased (P<0.05); and the greater the drug concentration, the more obvious the effect (P<0.05). In addition, up-regulation of miR-132 reduced the expression of HMGB1 in alveolar macrophage, increased inflammatory factor, and induced apoptosis in alveolar macrophage; however, down-regulation of miR-132 increased the expression of HMGB1 in alveolar macrophage, reduced inflammatory factor, and inhibited apoptosis in alveolar macrophage (P<0.05).ConclusionCurcumin could decrease LPS-induced inflammation and apoptosis in alveolar macrophage via decreasing miR-132 and increasing HMGB1.
ObjectiveTo investigate the regulatory effect of miRNA-21-5p (miR-21) on spinal fibroblasts, and to explore the mechanism of miR-21 related pathological process of spinal cord injury.MethodsSpinal cord fibroblasts were identified by immunofluorescence. Spinal fibroblasts damage model was established by scratch method. Quantitative real-time polymerase chain reaction (RT-PCR) was used to determine the relative expression of miR-21 and fibrosis-related genes in spinal cord fibroblasts after injury. The expression of miR-21 in spinal cord fibroblasts was up-regulated and down-regulated by using miR-21 mimics/inhibitor, and the expression levels of apoptosis and proliferation-related proteins were detected by Western Blot (WB).ResultsThe expression of miR-21 and fibrosis-related genes were increased after spinal cord fibroblast scratch (P<0.05). Up-regulation of the miR-21 can increase the expression of apoptosis-related genes in fibroblasts (P<0.05), and vice versa. The proliferation of fibroblasts was consistent with the expression of miR-21, while the apoptosis of fibroblasts was contrary to the expression of miR-21.ConclusionsmiR-21 enhanced the fibrosis and proliferation, inhibited the apoptosis of spinal cord fibroblasts after mechanical injury. This indicates that miR-21 is closely related with the formation of fibrotic scar after spinal cord injury, which also providesa potential therapeutic target for spinal cord injury.
At present, there are relatively many clinical gene studies. microRNA -215 (miR-215) is a miRNA induced by p53. It exists in animals and humans. miR-215 can exist not only in tumor tissues, but also in blood, urine and feces. miR-215 is abnormally expressed in a variety of tumors and plays a role in promoting and inhibiting cancer. Therefore, miR-215 may provide a new research direction for tumor diagnosis, treatment and prognosis. This paper reviews the expression of miR-215 as a oncogene and tumor suppressor gene in tumors and in non tumors.
Objective To explore the differential expression of circular RNAs (circRNAs) in polycystic ovary syndrome (PCOS) by bioinformatics, and predict the microRNAs (miRNAs) associated with them. Methods The expression profile of cumulus cells gene chip in PCOS was searched in the Gene Expression Omnibus database, and differential circRNAs were screened by GEO2R tool of the database. The Gene Ontology and Kyoto Encyclopedia of Genes and Genomes signaling pathways of different circRNA genes were analyzed using the DAVID 6.8 database. Circular RNA interactome was used to predict the potential regulated miRNAs. Cytoscape software was used to establish circRNA-miRNA network map. The potential regulatory miRNAs were predicted by the 10 circRNAs with the most significant differences in up-regulation and down-regulation. Results A total of 247 circRNAs were obtained in PCOS, and 277 miRNAs binding to up-regulated circRNA genes and 125 miRNAs binding to down-regulated circRNA genes were predicted. The top 10 miRNAs that could bind to multiple differential circRNAs were hsa-miR-557, hsa-miR-507, hsa-miR-224, hsa-miR-136, hsa-miR-127-5p, hsa-miR-579, hsa-miR-502-5p, hsa-miR-186, hsa-miR-1253, and hsa-miR-432. Conclusion The differential expression analysis of circRNAs is helpful to understand the main role of circRNAs in PCOS, and the prediction of potential regulated miRNAs can help to understand the pathogenesis of the disease.
Intervertebral disc degeneration is a multifactorial pathological process which is one of the leading causes of disability worldwide. The main pathological changes of intervertebral disc degeneration are the degradation of extracellular matrix, apoptosis, autophagy, senescence and inflammation. Dysregulation of microRNAs has been implicated in various pathologies, including various degenerative diseases such as disc degeneration. This article reviews the research status of microRNA in degenerative disc pathology, with emphasis on the biological mechanisms and potential therapeutic prospects of microRNA in extracellular matrix degradation, apoptosis, inflammation, and cartilage endplate degeneration.