Muscle-specific microRNAs Research Articles

Influence of statins on distinct circulating microRNAs during prolonged aerobic exercise.

Statins exacerbate exercise-induced skeletal muscle injury. Muscle-specific microRNAs (myomiRs) increase in plasma after prolonged exercise, but the patterns of myomiRs release after statin-associated muscle injury have not been examined. We examined the relationships between statin exposure, in vitro and in vivo muscle contraction, and expression of candidate circulating myomiRs. We measured plasma levels of myomiRs, circulating microRNA-1 (c-miR-1), c-miR-133a, c-miR-206, and c-miR-499-5p from 28 statin-using and 28 nonstatin-using runners before (PRE), immediately after (FINISH), and 24 h after they ran a 42-km footrace (the 2011 Boston marathon) (POST-24). To examine these cellular-regulation myomiRs, we used contracting mouse C2C12 myotubes in culture with and without statin exposure to compare intracellular and extracellular expression of these molecules. In marathoners, c-miR-1, c-miR-133a, and c-miR-206 increased at FINISH, returned to baseline at POST-24, and were unaffected by statin use. In contrast, c-miR-499-5p was unchanged at FINISH but increased at POST-24 among statin users compared with PRE and runners who did not take statins. In cultured C2C12 myotubes, extracellular c-miR-1, c-miR-133a, and c-miR-206 were significantly increased by muscle contraction regardless of statin use. In contrast, extracellular miR-499-5p was unaffected by either isolated statin exposure or isolated carbachol exposure but it was increased when muscle contraction was combined with statin exposure. In summary, we found that statin-potentiated muscle injury during exercise is accompanied by augmented extracellular release of miR-499-5p. Thus c-miR-499-5p may serve as a biomarker of statin-potentiated muscle damage.

MicroRNA expression and protein acetylation pattern in respiratory and limb muscles of Parp-1−/− and Parp-2−/− mice with lung cancer cachexia

BackgroundCurrent treatment options for cachexia, which impairs disease prognosis, are limited. Muscle-enriched microRNAs and protein acetylation are involved in muscle wasting including lung cancer (LC) cachexia. Poly(ADP-ribose) polymerases (PARP) are involved in muscle metabolism. We hypothesized that muscle-enriched microRNA, protein hyperacetylation, and expression levels of myogenic transcription factors (MTFs) and downstream targets, muscle loss and function improve in LC cachectic Parp-1−/− and Parp-2−/− mice. MethodsBody and muscle weights, grip strength, muscle phenotype, muscle-enriched microRNAs (miR-1, -133, -206, and -486), protein acetylation, acetylated levels of FoxO1, FoxO3, and PGC-1α, histone deacetylases (HDACs) including SIRT1, MTFs, and downstream targets (α-actin, PGC-1α, and creatine kinase) were evaluated in diaphragm and gastrocnemius of LC (LP07 adenocarcinoma) wild type (WT), Parp-1−/− and Parp-2−/− mice. ResultsCompared to WT cachectic animals, in both respiratory and limb muscles of Parp-1−/− and Parp-2−/− cachectic mice: downregulation of muscle-specific microRNAs was counterbalanced especially in gastrocnemius of Parp-1−/− mice; increased protein acetylation was attenuated (improvement in HDAC3, SIRT-1, and acetylated FoxO3 levels in both muscles, acetylated FoxO1 levels in the diaphragm); reduced MTFs and creatine kinase levels were mitigated; body and muscle weights, strength, and muscle fiber sizes improved, while tumor weight and growth decreased. ConclusionsThese molecular findings may explain the improvements seen in body and muscle weights, limb muscle force and fiber sizes in both Parp-1−/− and Parp-2−/− cachectic mice. General significancePARP-1 and -2 play a role in cancer-induced cachexia, thus selective pharmacological inhibition of PARP-1 and -2 may be of interest in clinical settings.

MicroRNA and mRNA Expression Changes in Steroid Naïve and Steroid Treated DMD Patients.

Background:Duchenne Muscular Dystrophy (DMD) is a recessive X-linked form of muscular dystrophy. Steroid therapy has clinical benefits for DMD patients, but the mechanism remains unclear.Objective:This study was designed to identify mRNAs and microRNAs regulated in Duchenne Muscular Dystrophy patients prior to and after steroid therapy.Methods:Genome wide transcriptome profiling of whole blood was performed to identify mRNAs and microRNAs regulated in DMD patients.Results:The data show many regulated mRNAs and some microRNAs, including some muscle-specific microRNAs (e.g., miR-206), that were significantly altered in blood of young (age 3–10) DMD patients compared to young controls. A total of 95 microRNAs, but no mRNAs, were differentially expressed in older DMD patients compared to matched controls (age 11–20). Steroid treatment reversed expression patterns of several microRNAs (miR-206, miR-181a, miR-4538, miR-4539, miR-606, and miR-454) that were altered in the young DMD patients. As an example, the over-expression of miR-206 in young DMD patients is predicted to down-regulate a set of target genes (e.g., RHGAP31, KHSRP, CORO1B, PTBP1, C7orf58, DLG4, and KLF4) that would worsen motor function. Since steroids decreased miR-206 expression to control levels, this could provide one mechanism by which steroids improve motor function.Conclusions:These identified microRNA-mRNA alterations will help better understand the pathophysiology of DMD and the response to steroid treatment.

Myogenic Differentiation of Mouse Embryonic Stem Cells That Lack a Functional Pax7 Gene.

The transcription factor Pax7 plays a key role during embryonic myogenesis and sustains the proper function of satellite cells, which serve as adult skeletal muscle stem cells. Overexpression of Pax7 has been shown to promote the myogenic differentiation of pluripotent stem cells. However, the effects of the absence of functional Pax7 in differentiating embryonic stem cells (ESCs) have not yet been directly tested. Herein, we studied mouse stem cells that lacked a functional Pax7 gene and characterized the differentiation of these stem cells under conditions that promoted the derivation of myoblasts in vitro. We analyzed the expression of myogenic factors, such as myogenic regulatory factors and muscle-specific microRNAs, in wild-type and mutant cells. Finally, we compared the transcriptome of both types of cells and did not find substantial differences in the expression of genes related to the regulation of myogenesis. As a result, we showed that the absence of functional Pax7 does not prevent the in vitro myogenic differentiation of ESCs.

Identification and expression of SRF targeted by miR-133a during early development of Paralichthys olivaceus.

Serum response factor (SRF) is a MADS-box transcription factor that regulates the expression of genes involved in development, metabolism, cell proliferation, and differentiation. In the present study, we cloned the full-length SRF cDNA which includes the coding region of 1503bp, a 573-bp 5'untranslated region (UTR) and a 400-bp 3'-UTR. The deduced 501 amino acid sequence of the SRF protein contained a MADS domain and NLS at the N terminus, similar to other organisms, and it also is highly phylogenetically conserved. SRF mRNA is ubiquitously expressed in various tissues, with the highest level in the kidneys, and it is also highly expressed during the embryonic and metamorphic stages. During metamorphosis, the SRF mRNA levels are down-regulated by exogenous thyroid hormone (TH) at 17dph and by thiourea (TU) at 29, 36, and 41dph, whereas SRF mRNA levels were significantly up-regulated by the added exogenous TH to the TU-treated larvae at 41dph, which indicates that thyroid hormone is essential for expression of SRF mRNA, so, higher levels of TH did not result in changes of SRF mRNA levels, while TH deficiency or inhibited by the non-specific TU toxicity cause down-regulation of SRF mRNA, which indicated that TH can indirectly affect the SRF mRNA levels. Meanwhile, using a luciferase reporter assay, we verified that SRF is a common target gene of miR-133a which is a muscle-specific microRNA (miRNA), which indicated that SRF may be involved in the signaling pathway of miRNA that regulates muscle development.

Epigenetic changes as a common trigger of muscle weakness in congenital myopathies.

Congenital myopathies are genetically and clinically heterogeneous conditions causing severe muscle weakness, and mutations in the ryanodine receptor gene (RYR1) represent the most frequent cause of these conditions. A common feature of diseases caused by recessive RYR1 mutations is a decrease of ryanodine receptor 1 protein content in muscle. The aim of the present investigation was to gain mechanistic insight into the causes of this reduced ryanodine receptor 1. We found that muscle biopsies of patients with recessive RYR1 mutations exhibit decreased expression of muscle-specific microRNAs, increased DNA methylation and increased expression of class II histone deacetylases. Transgenic mouse muscle fibres over-expressing HDAC-4/HDAC-5 exhibited decreased expression of RYR1 and of muscle-specific miRNAs, whereas acute knock-down of RYR1 in mouse muscle fibres by siRNA caused up-regulation of HDAC-4/HDAC-5. Intriguingly, increased class II HDAC expression and decreased ryanodine receptor protein and miRNAs expression were also observed in muscles of patients with nemaline myopathy, another congenital neuromuscular disorder. Our results indicate that a common pathophysiological pathway caused by epigenetic changes is activated in some forms of congenital neuromuscular disorders.

Diagnostic and prognostic role of circulating miR-206 in rhabdomyosarcoma patients.

10038 Background: miR-206 is a muscle-specific microRNA and is much more strongly expressed in rhabdomyosarcoma (RMS) tumors than in non-RMS tumors. Here, we analyzed the diagnostic and prognostic ...

MiR-133a Enhances the Protective Capacity of Cardiac Progenitors Cells after Myocardial Infarction

SummarymiR-133a and miR-1 are known as muscle-specific microRNAs that are involved in cardiac development and pathophysiology. We have shown that both miR-1 and miR-133a are early and progressively upregulated during in vitro cardiac differentiation of adult cardiac progenitor cells (CPCs), but only miR-133a expression was enhanced under in vitro oxidative stress. miR-1 was demonstrated to favor differentiation of CPCs, whereas miR-133a overexpression protected CPCs against cell death, targeting, among others, the proapoptotic genes Bim and Bmf. miR-133a-CPCs clearly improved cardiac function in a rat myocardial infarction model by reducing fibrosis and hypertrophy and increasing vascularization and cardiomyocyte proliferation. The beneficial effects of miR-133a-CPCs seem to correlate with the upregulated expression of several relevant paracrine factors and the plausible cooperative secretion of miR-133a via exosomal transport. Finally, an in vitro heart muscle model confirmed the antiapoptotic effects of miR-133a-CPCs, favoring the structuration and contractile functionality of the artificial tissue.

Epigenetic mechanisms in respiratory muscle dysfunction of patients with chronic obstructive pulmonary disease.

Epigenetic events are differentially expressed in the lungs and airways of patients with chronic obstructive pulmonary disease (COPD). Moreover, epigenetic mechanisms are involved in the skeletal (peripheral) muscle dysfunction of COPD patients. Whether epigenetic events may also regulate respiratory muscle dysfunction in COPD remains unknown. We hypothesized that epigenetic mechanisms would be differentially expressed in the main inspiratory muscle (diaphragm) of patients with COPD of a wide range of disease severity compared to healthy controls. In diaphragm muscle specimens (thoracotomy due to lung localized neoplasms) of sedentary patients with mild-to-moderate and severe COPD, with preserved body composition, and sedentary healthy controls, expression of muscle-enriched microRNAs, histone acetyltransferases (HATs) and deacetylases (HDACs), total DNA methylation and protein acetylation, small ubiquitin-related modifier (SUMO) ligases, muscle-specific transcription factors, and muscle structure were explored. All subjects were also clinically evaluated: lung and muscle functions and exercise capacity. Compared to healthy controls, patients exhibited moderate airflow limitation and diffusion capacity, and reduced exercise tolerance and transdiaphragmatic strength. Moreover, in the diaphragm of the COPD patients, muscle-specific microRNA expression was downregulated, while HDAC4 and myocyte enhancer factor (MEF)2C protein levels were higher, and DNA methylation levels, muscle fiber types and sizes did not differ between patients and controls. In the main respiratory muscle of COPD patients with a wide range of disease severity and normal body composition, muscle-specific microRNAs were downregulated, while HDAC4 and MEF2C levels were upregulated. It is likely that these epigenetic events act as biological adaptive mechanisms to better overcome the continuous inspiratory loads of the respiratory system in COPD. These findings may offer novel therapeutic strategies to specifically target respiratory muscle dysfunction in patients with COPD.

Coxsackievirus B3 engineered to contain microRNA targets for muscle-specific microRNAs displays attenuated cardiotropic virulence in mice.

Coxsackievirus B3 (CVB3) is trophic for cardiac tissue and is a major causative agent for viral myocarditis, where local viral replication in the heart may lead to heart failure or even death. Recent studies show that inserting microRNA target sequences into the genomes of certain viruses can eradicate these viruses within local host tissues that specifically express the cognate microRNA. Here, we demonstrated both in vitro and in vivo that incorporating target sequences for miRNA-133 and -206 into the 5' untranslated region of the CVB3 genome ameliorated CVB3 virulence in skeletal muscle and myocardial cells that specifically expressed the cognate cellular microRNAs. Compared to wild-type CVB3, viral replication of the engineered CVB3 was attenuated in human TE671 (rhabdomyosarcoma) and L6 (skeletal muscle) cell lines in vitro that expressed high levels of miRNA-206. In the in vivo murine CVB3-infection model, viral replication of the engineered CVB3 was attenuated specifically in the heart that expressed high levels of both miRNAs, but not in certain tissues, which allowed the host to retain the ability to induce a strong and protective humoral immune response against CVB3. The results of this study suggest that a microRNA-targeting strategy to control CVB3 tissue tropism and pathogenesis may be useful for viral attenuation and vaccine development. Coxsackievirus B3 (CVB3) is a major causative agent for viral myocarditis, and viral replication in the heart may lead to heart failure or even death. Limiting CVB3 replication within the heart may be a promising strategy to decrease CVB3 pathogenicity. miRNAs are ∼21-nucleotide-long, tissue-specific endogenous small RNA molecules that posttranscriptionally regulate gene expression by imperfectly binding to the 3' untranslated region (UTR), the 5' UTR, or the coding region within a gene. In our study, muscle-specific miRNA targets (miRT) were incorporated into the CVB3 genome. Replication of the engineered viruses was restricted in the important heart tissue of infected mice, which reduced cardiac pathology and increased mouse survival. Meanwhile, replication ability was retained in other tissues, thus inducing a strong humoral immune response and providing long-term protection against CVB3 rechallenge. This study suggests that a microRNA-targeting strategy can potentially control CVB3 tissue tropism and pathogenesis and may be useful for viral attenuation and vaccine development.

MyomiR-dependent switching of BAF60 variant incorporation into Brg1 chromatin remodeling complexes during embryo myogenesis.

Myogenesis involves the stable commitment of progenitor cells followed by the execution of myogenic differentiation, processes that are coordinated by myogenic regulatory factors, microRNAs and BAF chromatin remodeling complexes. BAF60a, BAF60b and BAF60c are structural subunits of the BAF complex that bind to the core ATPase Brg1 to provide functional specificity. BAF60c is essential for myogenesis; however, the mechanisms regulating the subunit composition of BAF/Brg1 complexes, in particular the incorporation of different BAF60 variants, are not understood. Here we reveal their dynamic expression during embryo myogenesis and uncover the concerted negative regulation of BAF60a and BAF60b by the muscle-specific microRNAs (myomiRs) miR-133 and miR-1/206 during somite differentiation. MicroRNA inhibition in chick embryos leads to increased BAF60a or BAF60b levels, a concomitant switch in BAF/Brg1 subunit composition and delayed myogenesis. The phenotypes are mimicked by sustained BAF60a or BAF60b expression and are rescued by morpholino knockdown of BAF60a or BAF60b. This suggests that myomiRs contribute to select BAF60c for incorporation into the Brg1 complex by specifically targeting the alternative variants BAF60a and BAF60b during embryo myogenesis, and reveals that interactions between tissue-specific non-coding RNAs and chromatin remodeling factors confer robustness to mesodermal lineage determination.

MiR-133b regulates bladder cancer cell proliferation and apoptosis by targeting Bcl-w and Akt1.

BackgroundMiR-133b is a muscle-specific microRNA; it has a role in the formation of cardiocytes and the expression of myocardium ion channels by regulating target genes. Many human malignant tumors demonstrate a low expression of miR-133b, as noted in colorectal, lung, esophagus and bladder cancers, but the role of miR-133b in bladder cancer is unknown.MethodsThe expression of miR-133b in clinical bladder cancer specimens and adjacent normal tissues was confirmed by stem-loop RT-PCR. We also analyzed the relationship between miR-133b expression and clinicopathological factors of bladder cancer. Bcl-w and Akt1 protein expression in 41 bladder cancer specimens and adjacent normal tissues was detected by Western blot. After transfection of miR-133b mimics or inhibitor into a T24 human bladder cancer cell line, Bcl-w and Akt1 protein and mRNA expression were examined by Western blot and RT-PCR, respectively. The effect of miR-133b on T24 cell proliferation and apoptosis was measured by CCK-8 tests and flow cytometry, respectively.ResultsThe expression of miR-133b in bladder cancer tissues from 41 patients was significantly down-regulated (P < 0.01); low expression of miR-133b was strongly associated with high-grade bladder cancer (P < 0.01). Bcl-w and Akt1 proteins were significantly overexpressed in bladder cancer tissues versus adjacent normal tissues (P < 0.01 for both). The expression of Akt1 and Bcl-w proteins and Akt1 mRNA, in T24 cells was significantly down-regulated or up-regulated after transfection of miR-133b mimics or inhibitor, respectively; however, there was no significant difference in Bcl-w mRNA expression. Transfection of HEK-293 T cells with miR-133b significantly suppressed a luciferase-reporter containing the Bcl-w or Akt 1 3′-untranslated regions. MiR-133b mimics significantly inhibited T24 cell proliferation, as well as increased T24 cell apoptosis (P < 0.05 and P < 0.01, respectively) while the miR-133b inhibitor increased and decreased these, respectively (P < 0.05 for both).ConclusionsMiR-133b may play a very important role in the proliferation and apoptosis of T24 cells by regulating the expression of Bcl-w and Akt1.

Sevoflurane anesthesia persistently downregulates muscle-specific microRNAs in rat plasma

The volatile anesthetic, sevoflurane, is widely used in surgery. Over the years, there has been a growing interest in the biological effects of sevoflurane on tissue and organ systems and the molecular mechanisms involved. MicroRNAs (miRNAs or miRs) acting as pivotal post‑transcriptional regulators for fine-tuning gene networks are not only expressed intracellularly, but are also secreted into the plasma. However, the sevoflurane‑associated dynamics of circulating miRNAs and the effects of sevoflurane on tissues remain unknown. Thus, the aim of this study was to perform a comprehensive analysis of circulating miRNA levels and compositions in sevoflurane‑anesthetized rats. The rats were allowed to breathe spontaneously under 2% sevoflurane anesthesia for 6 h, and we performed a quantitative polymerase chain reaction (PCR)‑based array analysis of the time-dependent changes in plasma miRNA levels and compositions. Subsequently, we validated the levels of muscle‑specific miRNAs (also known as myomiRNAs; miR-1, miR‑133a, miR-133b and miR-206) of the plasma, heart and skeletal muscle by quantitative PCR following 3 and 6 h of anesthesia, as well as at 1, 3, 7 and 14 days post-anesthesia. Of the 210 miRNAs detected in the rat plasma from the control group (no anesthesia), 161 plasma miRNAs (77%) were transiently downregulated as a result of sevoflurane anesthesia. Although the downregulation of the plasma miRNAs (148 out of the 161 plasma mRNAs; 92%) recovered immediately after anesthesia, the plasma levels of 4 muscle-specific miRNAs were persistently downregulated until 14 days post-anesthesia. In the cardiac and skeletal muscles, the expression levels of the muscle-specific miRNAs were upregulated within 2 weeks post-anesthesia, indicating that the expression levels of the muscle-specific miRNAs in the cardiac and skeletal muscles and their plasma levels are substantially inversely correlated following anesthesia. Our data suggest that sevoflurane predominantly affects cardiac and skeletal muscles and suppresses the release of miRNA from these tissues into the circulation. This new information provides novel insight into the molecular mechanisms of action of the anesthetic, sevoflurane.

Serum miR‐206 and other muscle‐specific microRNAs as non‐invasive biomarkers for Duchenne muscular dystrophy

Creatine kinase has been utilized as a diagnostic marker for Duchenne muscular dystrophy (DMD), but it correlates less well with the DMD pathological progression. In this study, we hypothesized that muscle-specific microRNAs (miR-1, -133, and -206) in serum may be useful for monitoring the DMD pathological progression, and explored the possibility of these miRNAs as potential non-invasive biomarkers for the disease. By using real-time quantitative reverse transcription-polymerase chain reaction in a randomized and controlled trial, we detected that miR-1, -133, and -206 were significantly over-expressed in the serum of 39 children with DMD (up to 3.20 ± 1.20, 2(-ΔΔCt) ): almost 2- to 4-fold enriched in comparison to samples from the healthy controls (less than 1.15 ± 0.34, 2(-ΔΔCt) ). To determine whether these miRNAs were related to the clinical features of children with DMD, we analyzed the associations compared to creatine kinase. There were very good inverse correlations between the levels of these miRNAs, especially miR-206, and functional performances: high levels corresponded to low muscle strength, muscle function, and quality of life. Moreover, by receiver operating characteristic curves analyses, we revealed that these miRNAs, especially miR-206, were able to discriminate DMD from controls. Thus, miR-206 and other muscle-specific miRNAs in serum are useful for monitoring the DMD pathological progression, and hence as potential non-invasive biomarkers for the disease. There has been a long-standing need for reliable, non-invasive biomarkers for Duchenne muscular dystrophy (DMD). We found that the levels of muscle-specific microRNAs, especially miR-206, in the serum of DMD were 2- to 4-fold higher than in the controls. High levels corresponded to low muscle strength, muscle function, and quality of life (QoL). These miRNAs were able to discriminate DMD from controls by receiver operating characteristic (ROC) curves analyses. Thus, miR-206 and other muscle-specific miRNAs are useful as non-invasive biomarkers for DMD.

Transcriptomic dissection of myogenic differentiation signature in caprine by RNA-Seq

Muscle growth and development from the embryonic to the adult stage of an organism consists of a series of exquisitely regulated and orchestrated changes in expression of genes leading to muscle maturation. In this study, we performed whole transcriptome profiling of adult caprine skeletal muscle derived myoblast and fused myotubes. Using Ion Torrent PGM sequencing platform, a total of 948,776 and 799,976 reads were generated in myoblasts and fused myotubes, respectively. The sequence reads were analyzed on CLC Genomics Workbench using Bos taurus RNA database to study the gene expression in both stages to study different genes responsible for muscle development and regeneration. The up and down-regulated genes were analyzed for gene ontology (GO) and KEGG pathways by Database for Annotation, Visualization and Integrated Discovery (DAVID) database. We found many genes exclusive to multinuclear fused myotubes and contractile nature of skeletal muscle, whereas up-regulated genes in myoblast stage were related to cell division and transcriptional regulation. Out of 27 genes selected for expression validation by RT-qPCR (reverse transcriptase-quantitative polymerase chain reaction), 19 genes showed the expression pattern comparable with CLC Genomics Workbench findings. Further, mRNA originated muscle specific microRNAs (miRNA-1 and miRNA-133b) were also observed in the fused myotubes along with other miRNAs with possible importance in muscle development. This study highlights important genes responsible for muscle development and differentiation in adult skeletal muscle system.

Muscle-specific microRNAs as biomarkers of Duchenne Muscular Dystrophy progression and response to therapies.

Recent studies have revealed the contribution of fibro-adipogenic progenitors (FAPs) to the pathogenesis and progression of Duchenne Muscular Dystrophy (DMD). While FAPs direct compensatory regeneration at early stages of disease, as the disease progresses they contribute to the progressive replacement of contractile myofibers with fibrotic scars and fatty infiltration. Using the mouse model of DMD – the mdx mice - we have recently reported that FAPs mediate the ability of HDAC inhibitors (HDACi) to promote muscle regeneration and prevent fibro-adipogenic degeneration at early stages of disease. This effect is mediated by the induction of myomiRs that, in turn, target the SWI/SNF components BAF60A and B, thereby favoring the formation of BAF60C-based SWI/SNF complex, which directs the switch from the fibro-adipogenic to the myogenic lineage. Here we show direct evidence of induction of miR-206 and BAF60C, and reduction of BAF60A, in FAPs isolated from mdx muscles exposed to the HDACi Trichostatin A (TSA). We also discuss how increased expression of myomiRs in dystrophic muscles can be integrated with circulating myomiRs to provide accurate biomarkers of disease progression and response to treatment.

MiR-133b, a muscle-specific microRNA, is a novel prognostic marker that participates in the progression of human colorectal cancer via regulation of CXCR4 expression

BackgroundMicroRNA-133b (miR-133b), which is a muscle-specific microRNA, has been reported to be downregulated in human colorectal carcinoma (CRC) when compared to adjacent non-tumor tissue. However, its diagnostic value and role in CRC have yet to be described. CXC chemokine receptor-4 (CXCR4), which participates in multiple cell processes such as cell invasion-related signaling pathways, was predicted to be a potential target of miR-133b. The aim of this study was to investigate the associations and functions of miR-133b and CXCR4 in CRC initiation and invasion.MethodsMature miR-133b and CXCR4 expression levels were detected in 31 tumor samples and their adjacent, non-tumor tissues from patients with CRC, as well as in 6 CRC cell lines, using real-time quantitative RT-PCR (qRT-PCR). Luciferase reporter assays and Western blots were used to validate CXCR4 as a putative target gene of miR-133b. Regulation of CXCR4 expression by miR-133b was assessed using qRT-PCR and Western blot analysis, and the effects of exogenous miR-133b and CXCR4 on cell invasion and migration were evaluated in vitro using the SW-480 and SW-620 CRC cell lines.ResultsA significant downregulation of miR-133b was observed in 93.55% of CRC tissues, and the expression of miR-133b was much lower in metastatic tumors (stage C and D, stratified by the Modified Dukes Staging System) than in primary tumors (stage A and B). In contrast, CXCR4 protein expression significantly increased in 52.63% of CRC samples, and increased CXCR4 expression in CRC was associated with advanced tumor stage. CXCR4 was shown to be a direct target of miR-133b by luciferase reporter assays, and transfection of miR-133b mimics inhibited invasion and stimulated apoptosis of SW-480 and SW-620 CRC cells.ConclusionsOur study demonstrated that downregulated miR-133b contributed to increased cell invasion and migration in CRC by negatively regulating CXCR4. These findings may be significant for the development of therapy target for CRC.

Downregulation of miR-133 via MAPK/ERK signaling pathway involved in nicotine-induced cardiomyocyte apoptosis

Tobacco smoking is a risk factor for many diseases, and nicotine is a major component of tobacco. Our previous work revealed that nicotine can induce myocardial fibrosis. This study aimed to investigate whether nicotine can induce cardiomyocyte apoptosis and to explore the mechanisms involved. Cardiomyocytes were exposed to different nicotine concentrations for 48h. MTT assay showed that the viability of cardiomyocytes was significantly inhibited by nicotine in a dose- and time-dependent manner. Loss of mitochondrial membrane potential, nuclear and DNA defragmentation determined by TUNEL and ELISA assays, and morphological alterations all revealed the pro-apoptotic property of nicotine. Meanwhile, miR-133, a muscle-specific microRNA, was markedly downregulated by nicotine. Consistently, caspase-9, a target gene for miR-133, was significantly upregulated, leading to an increase in caspase-3, in nicotine-treated cardiomyocytes compared to non-treated cells. Furthermore, ERK1/2 protein levels were considerably downregulated, along with reduction of serum response factor (SRF), which is a downstream target protein of ERK1/2 and an upstream transactivator of miR-133 as well. Our findings therefore revealed that inhibition of the ERK1/2-SRF-miR-133 signaling pathway to increase caspases-9 and -3 is a novel mechanism for nicotine to induce cardiomyocyte apoptosis and these tobacco smokers.

Increased skeletal muscle-specific microRNA in the blood of patients with COPD

BackgroundSkeletal muscle weakness in chronic obstructive pulmonary disease (COPD) carries a poor prognosis, therefore a non-invasive marker of this process could be useful. Reduced expression of muscle-specific microRNA (myomiRs) in...

Impaired regeneration in LGMD2A supported by increased PAX7‐positive satellite cell content and muscle‐specific microrna dysregulation

Recent in vitro studies suggest that CAPN3 deficiency leads initially to accelerated myofiber formation followed by depletion of satellite cells (SC). In normal muscle, up-regulation of miR-1 and miR-206 facilitates transition from proliferating SCs to differentiating myogenic progenitors. We examined the histopathological stages, Pax7 SC content, and muscle-specific microRNA expression in biopsy specimens from well-characterized LGMD 2A patients to gain insight into disease pathogenesis. Three distinct stages of pathological changes were identified that represented the continuum of the dystrophic process from prominent inflammation with necrosis and regeneration to prominent fibrosis, which correlated with age and disease duration. Pax7-positive SCs were highest in the fibrotic group and correlated with down-regulation of miR-1, miR-133a, and miR-206. These observations, and other published reports, are consistent with microRNA dysregulation leading to inability of Pax7-positive SCs to transit from proliferation to differentiation. This results in impaired regeneration and fibrosis.