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Objective:To explore the role of microRNA-133a (miR-133a) and silent mating information regulation 2 homolog 1 (SIRT1) in the effects of electroacupuncture on persons with disuse muscular atrophy.Methods:Thirty C57BL/6 mice were randomly divided into a control group, an experimental control group and an experimental group, each of 10. Disuse muscular atrophy was induced in the mice of the experimental and experimental control groups using tail suspension. The mice in the electroacupuncture group were given 15 minutes of electroacupuncture over the Yanglingquan and Zusanli points every day for 14 days. Wet weight ratio and the cross-sectional area of the gastrocnemius and soleus were tracked, and the structure of the mitochondria in the skeletal muscles was observed using a transmission electron microscope. The protein expressions of SIRT1, peroxisome proliferator-activated receptor γ coactivator-1a (PGC-1a), nicotinamide phosphoribosyl transferase (NAMPT), adenosine 5-monophosphate-activated protein kinase-a (AMPK-a) and phospho-AMPK-a (P-AMPK-a) were detected using western blotting. The expressions of the muscle atrophy F-box (Atrogin-1), muscle ring finger1 (MuRF1), miR-133a, SIRT1, paired box gene 7 (Pax7), myogenic determination (MyoD) and myogenin (MyoG) genes were detected through polymerase chain reactions. The concentration of Niacylamide adenine dinucleotide (NAD+ ) and the ratio of NAD+ to reduced nicotinamide adenine dinucleotide were also measured.Results:Compared with the experimental control group, the average wet weight increased by 21% and the cross-sectional area of the soleus increased by 30% in the experimental group. The average wet weight of the gastrocnemius increased by 5% and the area by 17%. The average expressions of Atrogin-1, MuRF1, SIRT1, PGC-1a and NAMPT, the concentration of NAD+ , as well as the average value of P-AMPK-a/AMPK-a and NAD+ /NADH were significantly lower in the experimental group than in the experimental control group, while the average expression of miR-133a in the experimental group was significantly (163%) higher. The average expressions of Pax7 and MyoD were significantly up-regulated in the experimental control group compared with the other two groups, while MyoG was highly expressed in the experimental group compared with the other 2 groups.Conclusions:The SIRT1 pathway is one of the reflexive protective mechanisms that mediate in the natural recovery of skeletal muscles. Electroacupuncture enhances myoblast differentiation, improves energy metabolism in the mitochondria, and thus promotes recovery from disuse muscular atrophy.
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Objective To explore expression level of circulating microRNA (miR)-133a and Galectin-3 and their potential clinical application in differential diagnosis between patients with chronic Keshan disease and dilated cardiomyopathy.Methods Twenty-eight patients with chronic Keshan disease and 28 cases of age-and sex-matched healthy people as control from the same severe historical endemic areas of Keshan disease in Heilongjiang Province,and another 28 patients with dilated cardiomyopathy from non-affected areas were chosen for the study.All the subjects were asked for disease history and did physical examination,examined by Doppler echocardiography for left ventricular ejection fraction (LVEF) and left ventricular end-diastolic diameter (LVEDD),and collected fasting venous blood specimen (elbow vein).The plasma miR-133a and the serum Galectin-3 were determined by Real-time PCR and enzyme-linked immunosorbent method,respectively.Meanwhile,the correlation was analyzed between miR-133a,galectin-3,LVEF and LVEDD.Results The miR-133a and Galectin-3 levels in different groups were statistically different (F =48.789,9.485,P ＜ 0.01).The plasma miR-133a level in chronic Keshan disease group and dilated cardiomyopathy group [median (quartile):0.394 (0.271,0.770),1.665 (0.943,2.713)] were both significantly lower than those in control group [2.382 (1.502,3.302],P ＜ 0.01 or ＜ 0.05],and the plasma miR-133a level in chronic Keshan disease group was lower than that in dilated cardiomyopathy group (P ＜ 0.01).There was no significant difference of serum Galectin-3 level between chronic Keshan disease group and dilated cardiomyopathy group [17.710 (9.624,27.799),12.692 (9.376,26.290) μg/L,P ＞ 0.05],but both were significantly higher than those in control group [8.070 (7.135,9.308) μg/L,P ＜ 0.01].The miR-133a was positively correlated with LVEF (rs =0.297,P ＜ 0.01),while negatively correlated with LVEDD,and Galectin-3 (rs =-0.271,-0.318,P ＜ 0.05 or ＜ 0.01);the serum Galectin-3 was negatively correlated with LVEF (rs =-0.392,P ＜ 0.01),and positively correlated with LVEDD (rs =0.385,P ＜ 0.01).Conclusion The combined application of miR-133a,Galectin-3,LVEF and LVEDD may provide assistance in clinical differential diagnosis of chronic Keshan disease and dilated cardiomyopathy.
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AIM:To investigate the effect of up-regulated expression of microRNA-133a (miR-133a) on myo-cardial fibrosis in spontaneously hypertensive rats (SHR). METHODS:Wistar-Kyoto (WKY) rats with homologous nor-mal blood pressure served as the normal control group. SHR were divided into SHR group, SHR+ adeno-associated virus (AAV) group and SHR+miR-133a-AAV group randomly. miR-133a carried by miR-133a-AAV was transfected into SHR heart by coronary perfusion. The rat tail artery pressure was monitored. The myocardial collagen deposition was observed by Masson staining. The expression of miR-133a in myocardial tissue was detected by real-time PCR. The protein levels of transforming growth factor-β1 (TGF-β1) and connective tissue growth factor (CTGF) were determined by immunohisto-chemistry and Western blot. RESULTS:Compared with the WKY rats, the tail artery pressure of the SHR increased sig-nificantly. The expression of miR-133a in heart decreased, and the expression levels of TGF-β1 and CTGF increased (P<0.05), and myocardial fibrosis occurred. After up-regulating the expression level of miR-133a in the heart of SHR, the myocardial fibrosis was significantly reduced, and the expression levels of TGF-β1 and CTGF decreased (P<0.05). CONCLUSION:Up-regulation of the miR-133a expression improves myocardial fibrosis induced by hypertension, which may be related to inhibiting the protein expression of TGF-β1 and CTGF in myocardium.
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Osteosarcoma (OS) has a high incidence, malignity, and frequency of recurrence and metastasis. In this study, we aimed to explore the potential anti-cancer effects of Astragalus polysaccharides (APS) on human OS MG63 cells as well as underlying mechanisms. Viability of MG63 cells was assessed by CCK-8 assay to determine the adequate concentration of APS. Then, effects of APS on MG63 cell proliferation, cell cycle distribution, apoptosis, and migration and invasion were analyzed by BrdU incorporation, PI staining, flow cytometry, and transwell assays, respectively. The expression levels of proteins involved in these physiological processes were assessed by western blot analysis. Afterwards, miR-133a level in APS-treated cells was determined by qRT-PCR, and whether APS affected MG63 cells through regulation of miR-133a was determined. Finally, the activation of c-Jun N-terminal protein kinase (JNK) pathway was detected. We found that APS treatment suppressed the viability, proliferation, migration, and invasion of MG63 cells, as well as induced cell apoptosis. Moreover, APS enhanced the expression of miR-133a in MG63 cells. Knockdown of miR-133a reversed the APS treatment-induced MG63 cell proliferation, migration and invasion inhibition, as well as cell apoptosis. Furthermore, APS inactivated JNK pathway in MG63 cells. Knockdown of miR-133a reversed the APS treatment-induced inactivation of JNK pathway in MG63 cells. To conclude, APS repressed proliferation, migration, and invasion while induced apoptosis of OS MG63 cells by up-regulating miR-133a and then inactivating JNK pathway.
Subject(s)
Humans , Bone Neoplasms/pathology , Cell Movement/drug effects , Apoptosis/drug effects , Astragalus Plant/chemistry , Cell Proliferation/drug effects , Bone Neoplasms/drug therapy , Cell Cycle/drug effects , Up-Regulation/drug effects , Cell Survival/drug effects , Blotting, Western , Reproducibility of Results , Analysis of Variance , MicroRNAs/analysis , Cell Line, Tumor , JNK Mitogen-Activated Protein Kinases/analysis , Antineoplastic Agents/pharmacologyABSTRACT
Objective To observe the influence of high-salt intake on the blood pressure,weight of left ventricle,myocardial microRNA133a and fibrosis levels by implementing the high-salt diet intervention trial in Dahl salt-sensitive rats and SD rats so as to explore the role of microRNA133a in salt-sensitive hypertensive cardiac fibrosis with high-salt intervention.Methods Sixteen 4-week male Dahl salt-sensitive rats are divided into two groups,8 in the SLS group and 8 in the SHS group.Similarly,sixteen 4-week male SD rats were divided into two groups. Systolic blood pressure of the rats was measured by the indirect tail-cuff method.Left ventricular mass index was measured after the rats were sacrificed. The expression levels of left ventricular myocardial tissue collagen I (LVMI),connective tissue growth factor (CTGF)expression and microRNA133a were observed.Results Blood pressure in high-salt groups both increased (P0.05).After the intervention,microRNA133a expression was significantly decreased in all high-salt groups compared with that in low-salt groups (P<0 .0 1 ),and the expression decreased more significantly in high-salt group of Dahl salt-sensitive rats than in high-salt group of SD rats (P<0.05).Conclusion High-salt diet is probably involved in salt-sensitive hypertension myocardial fibrosis by downregulating the expression of myocardial microRNA133a.
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AIM:To investigate the levels and clinical significance of microRNA-133a/b (miR-133a/b) and multidrug resistance-associated protein 1 ( MRP1 ) in peripheral blood of the patients with drug-refractory epilepsy. METHODS:Prediction of the miRNAs targeting transcriptional regulation of MRP1 was conducted by bioinformatics analy-sis.The plasmids containing wild type and mutant 3’UTR of MRP1 reporter gene were constructed.Dual luciferase report-er gene assay was used to verify this prediction.In addition, the peripheral blood samples of the epilepsy patients (37 cases were drug-refractory, the other 58 cases were nonresistant) were collected.The levels of miR-133a/b and MRP1 were measured by real-time PCR and ELISA.RESULTS:Through TargetScan database, it was predicted that miR-133a/b tran-scriptionally regulated MRP1.The results of dual luciferase report gene assay suggested that luciferase activity in experi-mental group with miR-133a/b mimics, pMIR-MRP1 and pRLTK plasmids were down-regulated by 76.9% and 64.1%compared with that in control group with scramble mimic, pMIR-MRP1 and pRLTK plasmids.The luciferase activity was up-regulated by 3.62 times and 2.04 times in mutation group with miR-133a/b mimics, pMIR-mut-MRP1 and pRLTK plasmids compared with experimental group.Before administration, the serum levels of miR-133a/b in the epilepsy patients without drug resistance was 2.18 times and 1.74 times higher than than in the epilepsy patients with drug resistance ( P<0.05), respectively, while MRP1 expression level was 3.72 times higher in the epilepsy patients with drug resistance than those in the epilepsy patients without drug resistance.After administration, the levels of miR-133a/b in the epilepsy pa-tients without drug resistance were 2.76 times and 2.95 times higher than those in the epilepsy patients with drug resistance (P<0.05), respectively, while the serum level of MRP1 in the epilepsy patients with drug resistance was 4.99 times higher than that in the epileptic patients without drug resistance (P<0.01).CONCLUSION:miR-133a/b transcriptio-nally regulates MRP1.There are lower expression levels of miR-133a/b and higher expression level of MRP1 in the epilep-sy patients with drug resistance compared with those in the epilepsy patients without drug resistance.miR-133a/b and MRP1 may be a diagnostic indicator for determining refractory epilepsy.
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Objective To explore the inhibitory effect of calcitonin gene-related peptide (CGRP) on cardiomyocyte apoptosis and the underlying mechanism.Methods In cultured rat cardiomyocytes,apoptosis was induced by the incubation of isoprenaline ( 10ˉ5 mol/L) for 48 h.CGRP ( 10ˉ8 or 10ˉ7 mol/L) was administrated for 1 h before incubating isoprenaline to evaluate its effect on cardiomyocyte apoptosis.At the end of the drug treatment,the rate of apoptotic cells and intracellular reactive oxygen species (ROS) were determined,and RNA was extracted to examine the expression of microRNA-1 and microRNA-133a.Results Isoprenaline significantly increased the rate of apoptotic cells and intracellular ROS production concomitantly with the increased microRNA-1 expression and the decreased microRNA-133a expression,which were inhibited by pretreatment with CGRP.The effects of CGRP were reversed by CGRP receptor antagonist.Conclusion CGRP can inhibit the isoprenaline-induced cardiomyocyte apoptosis.The beneficial effect of CGRP is related to regulating microRNA-1 and microRNA-133a expression through the prevention of isoprenaline-induced ROS production.