SMARCB1 expression in epithelioid sarcoma is regulated by miR‐206, miR‐381, and miR‐671‐5p on Both mRNA and protein levels

Proximal type epithelioid sarcoma shares similarities with malignant rhabdoid tumor, including the lack of nuclear immunoreactivity of SMARCB1. Biallelic mutation of SMARCB1 has been convincingly established as the cause of loss of protein expression in rhabdoid tumor, but the cause in epithelioid sarcoma remains unknown. In our previous work, we demonstrated that DNA hypermethylation and post‐translational modification mechanisms were not involved. In this current work, we explored the hypothesis that miRNAs regulate SMARCB1 gene expression in epithelioid sarcomas. In silico target prediction analysis revealed eight candidate miRNAs, and quantitative PCR—in 32 formalin‐fixed, paraffin‐embedded tumor samples comprising 30 epithelioid sarcomas and two malignant rhabdoid tumors—demonstrated significant (P < 0.001) overexpression of four miRNAs in epithelioid sarcomas: miR‐206, miR‐381, miR‐671‐5p, and miR‐765. Two human tumors (fibrosarcoma and colon adenocarcinoma) and a normal cell line (human dermal fibroblast) with retained SMARCB1 expression were cultured for miRNA transient transfection (electroporation) experiments. SMARCB1 mRNA expression was analyzed by quantitative real‐time PCR and immunostaining of SMARCB1 was performed to examine the effect of miRNAs transfections on both RNA and protein levels. Only three of the overexpressed miRNAs (miR‐206, miR‐381, and miR‐671‐5p) could silence the SMARCB1 mRNA expression in cell cultures; most effectively miR‐206. Transfection of miR‐206, miR‐381, miR‐671‐5p, and some combination of them also eliminated SMARCB1 nuclear staining, demonstrating a strong effect on not only mRNA but also protein levels. Our results suggest loss of SMARCB1 protein expression in epithelioid sarcoma is due to the epigenetic mechanism of gene silencing by oncomiRs. © 2013 Wiley Periodicals, Inc.     

Altered miR‐155 Expression in Allergic Asthmatic Airways

We and others have previously identified microRNAs (miRNAs) with pathological roles in animal models of asthma, where miR‐146a and miR‐155 have been described to play important roles in inflammatory responses. To date, few studies have investigated miRNA expression in human asthmatics. In the current study, significantly lower levels of miR‐155 were detected in cell‐free sputum from allergic asthmatics compared to healthy controls. Induced sputum isolated from allergic asthmatics in and out of pollen season revealed that miR‐155 expression, but not miR‐146a, is reduced in lymphocytes in season compared to post‐season. In contrast, miR‐155 was found to increase, whereas miR‐146a decreased in PBMCs and cell‐free PBMC culture media upon T cell receptor stimulation via αCD3/CD28 in both allergic asthmatics and healthy controls. Our findings suggest that miR‐155 is differentially expressed ex vivo in airways of allergic asthmatics compared to healthy controls, which may have implications in the local immune response in allergic asthma.     

miR‐1, regulated by LMP1, suppresses tumour growth and metastasis by targeting K‐ras in nasopharyngeal carcinoma

There is evidence to show that downregulation of miR‐1 expression is closely related to cancer progression, including in nasopharyngeal carcinoma (NPC). However, the molecular mechanisms underlying miR‐1 downregulation in NPC remain largely unknown, especially its association with Epstein–Barr virus (EBV). In this study we found that restoration of miR‐1 dramatically inhibited cell invasion in vitro, together with tumour growth and metastasis in vivo. Importantly, we found that LMP1, an Epstein–Barr virus (EBV)‐associated protein, suppressed miR‐1 expression. Furthermore, we identified K‐ras as a novel direct target of miR‐1. Our results demonstrated for the first time that miR‐1 was suppressed by LMP1 and its tumour‐suppressive effects were mediated chiefly by repressing K‐ras expression. We propose that miR‐1 could serve as an independent biomarker to identify patients with different clinical characteristics.     

Analysis of Circulating miR‐1, miR‐23a,and miR‐26a in Atrial Fibrillation Patients Undergoing Coronary Bypass Artery Grafting Surgery

Atrial fibrillation (AF) is the most common arrhythmia after cardiac surgery. From a pathophysiological point of view, a myriad of factors such as trauma, atrial dilation, ischemia, mechanical myopericarditis, autonomic imbalance, loss of connexins, AF nest remodeling, inflammation, sutures, and dysfunction caused by postextracorporeal circulation can contribute to postoperative atrial fibrillation (POAF) resulting in a longer hospital stay and consequently higher cost. Recent studies showed that short fragments of RNA, called microRNA (miRNA), can contribute to the development of several cardiovascular diseases, including AF. The aim of this study was to evaluate the levels of circulating miRNAs (miR‐1, ‐23a, and ‐26a) that can be involved in POAF. Patients submitted to coronary artery bypass graft surgery were grouped in POAF (24 patients) and without POAF (24 patients). Results showed older age, longer clamp‐time, and more days in the intensive care unit as well as a longer total hospital stay in the POAF group. Preoperative levels of circulating miRNAs were similar. Analysis of miRNAs revealed significantly lower circulating levels of miRNA‐23a (P = 0.02) and ‐26a (P = 0.01) in the POAF group during the postoperative period. Receiver operating characteristic (ROC) analysis showed the area under the ROC curve of miR‐23a and miR‐26a for predicting FA was 0.63 (95% confidence interval [CI]: 0.51–0.74; P = 0.02) and 0.66 (95% CI: 0.55–0.77; P = 0.01), respectively. Our data suggests that circulating miRNA‐23a and ‐26a may be involved in the underlying biology of postoperative AF development.     

Increased miR‐223 expression in T cells from patients with rheumatoid arthritis leads to decreased insulin‐like growth factor‐1‐mediated interleukin‐10 production

We hypothesized that the aberrant expression of microRNAs (miRNAs) in rheumatoid arthritis (RA) T cells was involved in the pathogenesis of RA. The expression profile of 270 human miRNAs in T cells from the first five RA patients and five controls were analysed by real‐time polymerase chain reaction. Twelve miRNAs exhibited potentially aberrant expression in RA T cells compared to normal T cells. After validation with another 22 RA patients and 19 controls, miR‐223 and miR‐34b were over‐expressed in RA T cells. The expression levels of miR‐223 were correlated positively with the titre of rheumatoid factor (RF) in RA patients. Transfection of Jurkat cells with miR‐223 mimic suppressed insulin‐like growth factor‐1 receptor (IGF‐1R) and transfection with miR‐34b mimic suppressed cAMP response element binding protein (CREB) protein expression by Western blotting. The protein expression of IGF‐1R but not CREB was decreased in RA T cells. The addition of recombinant IGF‐1‐stimulated interleukin (IL)‐10 production by activated normal T cells, but not RA T cells. The transfection of miR‐223 mimic impaired IGF‐1‐mediated IL‐10 production in activated normal T cells. The expression levels of SCD5, targeted by miR‐34b, were decreased in RA T cells after microarray analysis. In conclusion, both miR‐223 and miR‐34b were over‐expressed in RA T cells, but only the miR‐223 expression levels were correlated positively with RF titre in RA patients. Functionally, the increased miR‐223 expression could impair the IGF‐1‐mediated IL‐10 production in activated RA T cells in vivo, which might contribute to the imbalance between proinflammatory and anti‐inflammatory cytokines.     

Up‐regulation of miR‐455‐5p by the TGF‐β–SMAD signalling axis promotes the proliferation of oral squamous cancer cells by targeting UBE2B

MicroRNAs (miRNAs) are involved in the tumourigenesis of various cancers by regulating their downstream targets. To identify the changes of miRNAs in oral squamous cell carcinoma (OSCC), we investigated the expression profiles of miRNAs in 40 pairs of OSCC specimens and their matched non‐tumour epithelial tissues. Our data revealed higher miR‐455‐5p expression in the tumour tissues than in the normal tissues; the expression was also higher in oral cancer cell lines than in normal keratinocyte cell lines. MiR‐455‐5p knockdown reduced both the anchorage‐independent growth and the proliferative ability of oral cancer cells, and these factors increased in miR‐455‐5p‐overexpressing cells. Furthermore, by analysing the array data of patients with cancer and cell lines, we identified ubiquitin‐conjugating enzyme E2B (UBE2B) as a target of miR‐455‐5p, and further validated this using 3′‐untranslated region luciferase reporter assays and western blot analysis. We also demonstrated that UBE2B suppression rescued the impaired growth ability of miR‐455‐5p‐knockdown cells. Furthermore, we observed that miR‐455‐5p expression was regulated, at least in part, by the transforming growth factor‐β (TGF‐β) pathway through the binding of SMAD3 to specific promoter regions. Notably, miR‐455‐5p expression was associated with the nodal status, stage, and overall survival in our patients, suggesting that miR‐455‐5p is a potential marker for predicting the prognosis of patients with oral cancer. In conclusion, we reveal that miR‐455‐5p expression is regulated by the TGF‐β‐dependent pathway, which subsequently leads to UBE2B down‐regulation and contributes to oral cancer tumourigenesis. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Down‐Regulation of miR‐19a as a Biomarker for Early Detection of Silicosis

The accumulated data indicate that there is significant genetic heterogeneity underlying the etiology of silicosis. Recent reports have revealed that microRNAs (miRNAs) play an important role in regulating pulmonary fibrosis. This study, therefore, aimed to identify some miRNAs as biomarkers for silicosis, and to explore the early diagnostic value of biomarkers for silicosis. Total RNAs were collected from the peripheral blood leukocytes of 23 silicosis patients and 23 healthy controls, the different miRNAs were screened using microarrays. The potential biomarker miRNAs were identified by quantitative real‐time polymerase chain reaction (qPCR) and receiver operating characteristic (ROC) curves. Eighteen differential miRNAs in leukocytes were up‐regulated and twenty differential miRNAs were down‐regulated in the silicosis group, compared with the control group. The expression levels of miR‐181a and miR‐19a were 0.8854 ± 0.1037 and 0.2929 ± 0.0342 by the relative quantitation method 2^?△△CT of qPCR, respectively. The sensitivity and specificity for miR‐181a at a cut‐off value of 1.8917 were 70% and 75%, respectively, whereas, those for miR‐19a at a cut‐off value of 3.6828 were 95% and 95%, respectively. Thus, miR‐19a in peripheral blood leukocyte could be used as an effective biomarker for silicosis. Anat Rec, 299:1300–1307, 2016. © 2016 Wiley Periodicals, Inc.     

Increased levels of serum miR‐148a‐3p are associated with prostate cancer

Prostate cancer (PCa) is one of the most common types of cancer and the fifth leading cause of death among men worldwide. The tools for diagnosing PCa have limited value, and to improve correct diagnosis there is a need for markers that can contribute to a more precise diagnosis, which would lead to proper treatment of only those patients who need it. Micro RNA (miRNA) plays a key role in the development of cancer and is therefore a potential marker for PCa. Next‐generation sequencing was used to discover differences in miRNA expression between serum samples from PCa patients and healthy controls, and the results were validated by quantitative real‐time polymerase chain reaction. Detection of the miRNA of interest was attempted in prostate tissue by in situ hybridization. All samples were collected in collaboration with Biobank1^®. By miRNA sequencing of serum samples, significant expression of some miRNAs in patients with PCa and healthy controls was detected. This study showed that miR‐148a‐3p is upregulated in men with PCa, and the miRNA is differentially expressed in PCa patients compared to healthy controls. The results also showed that miR‐148a‐3p is located in prostate tissue.     

Antibody against the Epstein-Barr virus BHRF1 protein,a homologue of Bcl-2, in patients with nasopharyngeal carcinoma

The Epstein-Barr virus (EBV) open reading frame BHRF1, a homologue of the oncogene bcl-2, was cloned from a patient with nasopharyngeal carcinoma (NPC) and overexpressed in Escherichia coli. The resulting recombinant BHRF1 fusion protein, with an apparent molecular weight of 35 KD, was used as antigen in an immunoblotting assay for IgG antibody in human sera. Anti-BHRF1 antibody was detected in 57 (61.3%) of 93 patients with NPC, 5 (5.7%) of 87 patients with nonmalignant diseases of the nasopharynx, and in 1 (1.3%) of 78 healthy blood donors. The positivity rate in these nonmalignant patients was 4.4 times that of the normal controls. Negative results were observed in four patients with infectious mononucleosis and patients with other cancers, including 4 with esophageal cancer, 11 with lung cancer, 10 with lymphoma, 13 with gastric carcinoma, 10 with cervical carcinoma, and 10 with other head and neck cancers. Antibody neutralizing EBV DNase and IgA antibody to viral capsid antigen (VCA) were assayed in parallel. The results showed that 7.5% of the NPC patients were negative for anti-DNase and anti-VCA antibodies and EBV infection could be detected by the anti-BHRF1 antibody alone. The demonstration of anti-BHRF1 antibody in most NPC sera strongly supports the hypothesis that the EBV BHRF1 protein is expressed in most NPC patients and its specific antibody can be a useful marker for the diagnosis of NPC. J. Med. Virol. 56:179–185, 1998. © 1998 Wiley-Liss, Inc.