Identification of differential expressed PE exosomal miRNA in lung adenocarcinoma, tuberculosis, and other benign lesions.
ABSTRACT: Pleural effusion (PE) is a common clinical complication of many pulmonary and systemic diseases, including lung cancer and tuberculosis. Nevertheless, there is no clinical effective biomarker to identify the cause of PE. We attempted to investigate differential expressed exosomal miRNAs in PEs of lung adenocarcinoma (APE), tuberculous (TPE), and other benign lesions (NPE) by using deep sequencing and quantitative polymerase chain reaction (qRT-PCR). As a result, 171 differentiated miRNAs were observed in 3 groups of PEs, and 11 significantly differentiated exosomal miRNAs were validated by qRT-PCR. We identified 9 miRNAs, including miR-205-5p, miR-483-5p, miR-375, miR-200c-3p, miR-429, miR-200b-3p, miR-200a-3p, miR-203a-3p, and miR-141-3p which were preferentially represented in exosomes derived from APE when compared with TPE or NPE, while 3 miRNAs, including miR-148a-3p, miR-451a, and miR-150-5p, were differentially expressed between TPE and NPE. These different miRNAs profiles may hold promise as biomarkers for differential diagnosis of PEs with more validation based on larger cohorts.
Project description:Exosomal microRNAs (miRNAs) are promising candidate biomarkers for diagnosis or prognosis for breast cancer. We investigated the prognostic role of exosomal miRNAs in serum samples derived from patients with breast cancer and compared miRNA expression between serum and tumor tissues.The miRNA profile derived from exosome between breast cancer patients with recurrence (n = 16) and without recurrence (n = 16) were compared by miRNA PCR array. Further, we examined the expression of miRNAs derived from tissues in the patients with breast cancer with (n = 35) and without recurrence (n = 39) by qRT-PCR.Of 384 miRNAs, three miRNAs (miR-338-3p, miR-340-5p, and miR-124-3p) were significantly upregulated and eight (miR-29b-3p, miR-20b-5p, miR-17-5p, miR-130a-3p, miR-18a-5p, miR-195-5p, miR-486-5p, and miR-93-5p) were significantly downregulated in the patients with recurrence. We evaluated the expression of the miRNAs in tumor tissues. The patients with recurrence had higher levels of miR-340 at their primary site as well as in the serum. In contrast, miR-195-5p, miR-17-5p, miR-93-5p, and miR-130a-3p, derived from tumor tissues that were downregulated in the serum from patients with recurrence, were higher in the patients with recurrence than in those with no recurrence. In logistic regression analysis, miR-340-5p, miR-17-5p, miR-130a-3p, and miR-93-5p were significantly associated with recurrence.Several exosomal miRNAs may be useful biomarkers to predict breast cancer recurrence. We show the different expression patterns of miRNAs between tumor tissues and serum. These findings may suggest selective mechanism of release of exosomal miRNAs by cancer cells to regulate their progression.
Project description:Gastric cancer (GC) remains a leading cause of cancer-related mortality in the United States and China, there is an urgent need to discover novel non-invasive biomarkers for the early diagnosis of GC to improve the prognosis of GC patients. Exosomal miRNAs are considered promising biomarkers for cancer diagnosis. Using next-generation sequencing (NGS), bioinformatics and further validation, we identified and evaluated exosomal miRNAs in serum as early diagnostic markers for GC. NGS revealed that the average mappable reads in the RNA libraries were about 6.5 million per patient including miRNAs (73.38%), rRNAs (17.10%), snRNAs (8.83%), snoRNAs (0.65%), and tRNAs (0.04%). A total of 66 up and 13 down-regulated exosomal miRNAs were found in the screened cohort. In the validation cohort, by comparing with healthy individuals, higher levels of serum exosomal miR-92b-3p, let-7g-5p, miR-146b-5p, and miR-9-5p were found to be significantly associated with early-stage GC (p < 0.05). Diagnostic power of the combined panels of the exosomal miRNAs or the combination of exosomal miRNAs and CEA outperformed that of single exosomal miRNA marker for establishing a diagnosis of early-stage GC. The combined diagnosis of exosomal miR-92b-3p + let-7g-5p + miR-146b-5p + miR-9-5p with CEA had the most powerful efficiency with an AUC up to 0.786. In addition, serum levels of exosomal miR-92b-3p were significantly associated with poor cohesiveness (p = 0.0021), let-7g-5p and miR-146b-5p were significantly correlated with nerve infiltration (p = 0.0234 and p = 0.0126, respectively), and miR146b-5p was statistically correlated with tumor invasion depth in early-stage GC (p = 0.0089). In conclusion, serum exosomal miR-92b-3p, -146b-5p, -9-5p, and let-7g-5p may serve as potential non-invasive biomarkers for early diagnosis of GC.
Project description:Background: A liquid biopsy using circulating exosomal genetic materials provides new insights for thyroid cancer diagnosis. This study aimed to identify plasma-derived exosomal biomarkers that could be used for early detection of papillary thyroid carcinoma (PTC). Method: Exosomal miRNAs in plasma were isolated from patients with benign thyroid nodules and patients with PTC. Profiling of exosomal miRNA was performed using RNA sequencing (RNA-seq) to identify miRNA candidates and differentiate the benign from malignant. The validation cohort consisted of 30 patients with benign thyroid nodules, 35 PTC patients, and 31 healthy individuals. Real-time PCR was used to quantify the expression of miRNA candidates. The diagnostic potential of the candidates was evaluated by receiver operating characteristic (ROC) curves. Results: After RNA-seq, eight plasma exosomal miRNAs were selected as candidates. Further validation indicated that the levels of exosomal miR-16-2-3p, miR-223-5p, miR-34c-5p, miR-182-5p, miR-223-3p, and miR-146b-5p were significantly lower in nodules compared to healthy controls (p < 0.0001), whereas miR-16-2-3p and miR-223-5p were significantly higher in the PTC cases than in those with benign nodules (p < 0.05). ROC analyses revealed that the above six miRNAs were potent indicators for detection of thyroid nodules. Meanwhile, miR-16-2-3p and miR-223-5p can be utilized for detecting PTC from benign nodules. Additionally, combined miRNA panels showed increased diagnostic sensitivities and specificities compared to single miRNA markers. Conclusion: Six aberrantly expressed plasma exosomal miRNAs may be used as diagnostic biomarkers to differentiate thyroid nodules from healthy individuals. The panel consisting of miR-16-2-3p, miR-223-5p, miR-101-3p, and miR-34c-5p are eligible for discriminating benign from malignant thyroid nodules.
Project description:Specific types of human papillomaviruses (HPVs) cause cervical cancer. Cervical cancers exhibit aberrant cellular microRNA (miRNA) expression patterns. By genome-wide analyses, we investigate whether the intracellular and exosomal miRNA compositions of HPV-positive cancer cells are dependent on endogenous E6/E7 oncogene expression. Deep sequencing studies combined with qRT-PCR analyses show that E6/E7 silencing significantly affects ten of the 52 most abundant intracellular miRNAs in HPV18-positive HeLa cells, downregulating miR-17-5p, miR-186-5p, miR-378a-3p, miR-378f, miR-629-5p and miR-7-5p, and upregulating miR-143-3p, miR-23a-3p, miR-23b-3p and miR-27b-3p. The effects of E6/E7 silencing on miRNA levels are mainly not dependent on p53 and similarly observed in HPV16-positive SiHa cells. The E6/E7-regulated miRNAs are enriched for species involved in the control of cell proliferation, senescence and apoptosis, suggesting that they contribute to the growth of HPV-positive cancer cells. Consistently, we show that sustained E6/E7 expression is required to maintain the intracellular levels of members of the miR-17~92 cluster, which reduce expression of the anti-proliferative p21 gene in HPV-positive cancer cells. In exosomes secreted by HeLa cells, a distinct seven-miRNA-signature was identified among the most abundant miRNAs, with significant downregulation of let-7d-5p, miR-20a-5p, miR-378a-3p, miR-423-3p, miR-7-5p, miR-92a-3p and upregulation of miR-21-5p, upon E6/E7 silencing. Several of the E6/E7-dependent exosomal miRNAs have also been linked to the control of cell proliferation and apoptosis. This study represents the first global analysis of intracellular and exosomal miRNAs and shows that viral oncogene expression affects the abundance of multiple miRNAs likely contributing to the E6/E7-dependent growth of HPV-positive cancer cells.
Project description:Background:A reliable noninvasive biomarker is not yet available for endometriosis diagnosis. Novel biomarkers for the diagnosis of endometriosis are urgently needed. The molecular constituents of exosomes, especially exosomal microRNAs (miRNAs), have considerable potential as novel biomarkers for clinical diagnosis. This study is aimed at exploring aberrant exosomal miRNA profiles by using miRNA microarray and at providing more accurate molecular biomarkers of endometriosis. Methods:Exosomes were isolated from the serum of patients with endometriosis and negative controls and identified by electron microscopy, nanoparticle tracking analysis, and Western blot. Exosomal miRNAs were profiled by miRNA microarrays. The expression of selective serum exosomal miRNA was validated by qRT-PCR. Receiver operating characteristic (ROC) curves were established to explore the diagnostic value of selective miRNAs. Finally, GO annotation and KEGG pathway enrichment analyses were used to display possible functions associated with the two miRNAs. Results:A total of 24 miRNAs showed differential levels of enrichment with P < 0.05 and |log2?fold?change| > 1 by miRNA microarrays. Among the six selective miRNAs (i.e., miR-134-5p, miR-197-5p, miR-22-3p, miR-320a, miR-494-3p, and miR-939-5p), qRT-PCR analysis revealed that miR-22-3p and miR-320a were significantly upregulated in serum exosomes from patients with endometriosis compared with negative individuals. ROC curve revealed that the serum exosomal miR-22-3p and miR-320a yielded the area under the curve values of 0.855 and 0.827, respectively. Conclusion:Our results demonstrated that exosomal miR-22-3p and miR-320a were significantly increased in the sera of patients with endometriosis. The two miRNAs may be useful potential biomarkers for endometriosis diagnosis.
Project description:Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS). There is currently no single definitive test for MS. Circulating exosomes represent promising candidate biomarkers for a host of human diseases. Exosomes contain RNA, DNA, and proteins, can cross the blood-brain barrier, and are secreted from almost all cell types including cells of the CNS. We hypothesized that serum exosomal miRNAs could present a useful blood-based assay for MS disease detection and monitoring. Exosome-associated microRNAs in serum samples from MS patients (n?=?25) and matched healthy controls (n?=?11) were profiled using small RNA next generation sequencing. We identified differentially expressed exosomal miRNAs in both relapsing-remitting MS (RRMS) (miR-15b-5p, miR-451a, miR-30b-5p, miR-342-3p) and progressive MS patient sera (miR-127-3p, miR-370-3p, miR-409-3p, miR-432-5p) in relation to controls. Critically, we identified a group of nine miRNAs (miR-15b-5p, miR-23a-3p, miR-223-3p, miR-374a-5p, miR-30b-5p, miR-433-3p, miR-485-3p, miR-342-3p, miR-432-5p) that distinguished relapsing-remitting from progressive disease. Eight out of nine miRNAs were validated in an independent group (n?=?11) of progressive MS cases. This is the first demonstration that microRNAs associated with circulating exosomes are informative biomarkers not only for the diagnosis of MS, but in predicting disease subtype with a high degree of accuracy.
Project description:MicroRNAs (miRNAs) serve important roles in drug?resistance; however, exosomal miRNAs associated with drug?resistance in ovarian cancer (OC) have not been reported to date. The current study aimed to analyze the drug resistance?associated exosomal miRNAs in original OC cells and their derived exosomes using microarray data downloaded from the Gene Expression Omnibus database (series GSE76449). The chemosensitive OC cell lines SKOV3_ip1, A2780_PAR and HEYA8, as well as the chemoresistant cell lines SKOV3_TR, A2780_CP20 and HEYA8_MDR, were investigated. Differentially expressed miRNAs (DE?miRNAs) were identified using the limma method, and their mRNA targets were predicted using the miRWalk and LinkedOmics database. Functions of target genes were analyzed with DAVID tool, while TCGA data were used to explore the survival association of identified miRNAs. According to the results, 28 DE?miRNAs were found to be common in exosomal and original samples of A2780_CP20 cells, among which the functions of 5 miRNAs were predicted (including miR?146b?5p, miR?509?5p, miR?574?3p, miR?574?5p and miR?760). In addition, 16 and 35 DE?miRNAs were detected for HEYA8_MDR and SKOV3_TR, respectively, with the functions of 4 of these miRNAs predicted for each cell line (HEYA8_MDR: miR?30a?3p, miR?30a?5p, miR?612 and miR?617; SKOV3_TR: miR?193a?5p, miR?423?3p, miR?769?5p and miR?922). It was also reported that miR?183?5p was the only one common miRNA among the three cell lines. Furthermore, miR?574?3p, miR?30a?5p and miR?922 may regulate CUL2 to mediate HIF?1 cancer signaling pathway, while miR?183?5p may modulate MECP2, similar to miR?760, miR?30a?5p and miR?922, to influence cell proliferation. Finally, the downregulated miR?612 may promote the expression of TEAD3 via the Hippo signaling pathway, and this miRNA was associated with poor prognosis. In conclusion, the findings of the present study suggested several underlying miRNA targets for improving the chemotherapy sensitivity of OC.
Project description:Exosomal miRNAs are proposed as excellent candidate biomarkers for clinical applications. However, little is known about their potential roles as prognostic biomarkers in lung cancer. In this study, we explored the prognostic value of plasma exosomal microRNAs (miRNAs) for non-small-cell lung cancer (NSCLC). Using a quantitative polymerase chain reaction (qPCR) array panel, we analyzed 84 plasma exosomal miRNAs in 10 lung adenocarcinoma patients and 10 matched healthy controls. The qPCR array showed 30 aberrantly expressed exosomal miRNAs. Nine candidate miRNAs were selected based on differential expression and previous reports for further evaluating their prognostic roles in 196 NSCLC patients. Elevated levels of exosomal miR-23b-3p, miR-10b-5p and miR-21-5p were independently associated with poor overall survival (with hazard ratio [95% confidence interval]: 2.42 (1.45 - 4.04), P = 0.001; 2.22 (1.18 - 4.16), P = 0.013; 2.12 (1.28 - 3.49), P = 0.003, respectively). When compared to the clinical prognostic variables only model, adding the three exosomal miRNA signatures significantly improved survival predictive accuracy with an increase of time-dependent area under the receiver operating characteristic curve from 0.88 to 0.91 (P=0.015). Our results indicated that plasma exosomal miR-23b-3p, miR-10b-5p and miR-21-5p are promising non-invasive prognostic biomarkers of NSCLC.
Project description:Exosomal miRNAs are currently being explored as a novel class of biomarkers in cardiovascular diseases. However, few reports have focused on the value of circulating exosomal miRNAs as biomarkers for stable coronary artery disease (SCAD). Here, we aimed to investigate whether miRNAs involved in cardiovascular diseases in circulating exosomes could serve as novel diagnostic biomarkers for SCAD. Firstly, the serum exosomes were isolated and purified by the ExoQuick reagent and identified by transmission electron microscopy, western blot, and nanoparticle tracking analysis. Then, the purified exosomes were quantified by measuring the exosome protein concentration and calculating the total protein amount. Next, eight miRNAs involved in cardiovascular diseases, miR-192-5p, miR-148b-3p, miR-125a-3p, miR-942-5p, miR-149-5p, miR-32-5p, miR-144-3p, and miR-142-5p, were quantified in circulating exosomes from the control group (<i>n</i> = 20) and the SCAD group (<i>n</i> = 20) by quantitative real-time polymerase chain reaction (qPCR). Finally, the gene targets of the differentially expressed miRNAs were predicted, and the functions and signaling pathways of these targets were analyzed using an online database. The isolated exosomes had a bilayer membrane with a diameter of about 100?nm and expressed exosomal markers including CD63, Tsg101, and Flotillin but negatively expressed Calnexin. Both the exosome protein concentration and total protein amount exhibited no significant differences between the two groups. The qPCR assay demonstrated that among the eight miRNAs, the expression levels of miR-942-5p, miR-149-5p, and miR-32-5p in the serum exosomes from the SCAD group were significantly higher than that from the control group. And the three miRNAs for SCAD diagnosis exhibited AUC values of 0.693, 0.702, and 0.691, respectively. GO categories and signaling pathways analysis showed that some of the predictive targets of these miRNAs were involved in the pathophysiology processes of SCAD. In conclusion, our findings suggest that serum exosomal miR-942-5p, miR-149-5p, and miR-32-5p may serve as potential diagnostic biomarkers for SCAD.
Project description:Oral lichen planus (OLP) is a common chronic inflammatory autoimmune disease with unclear etiology. The aim of the present study was to identify the expression profiles of circulating exosomal miRNAs, which have been shown to be potent stimulators of inflammatory and immune responses, in OLP patients. Plasma exosomes were isolated from the patients and healthy individuals, and RAE scoring system was used to evaluate the severity of OLP. Differentially deregulated exosomal miRNAs associated with inflammatory response and autoimmunity in OLP were identified by miScript® miRNA PCR Array, and the results were confirmed by RT-PCR. The relationship between exosomal miRNAs and RAE scores was then analyzed, and bioinformatics analysis was used to predict the target genes and pathways of the differentially expressed exosomal miRNAs. Expression profiling showed that circulating exosomal miR-34a-5p and miR-130b-3p were upregulated, while miR-301b-3p was downregulated in OLP patients. Exosomal miR-34a-5p was positively correlated with the severity of OLP. Bioinformatics analysis revealed that the target genes of miR-34a-5p were mainly involved in regulation of gene expression, cell communication, signaling, and metabolic process, and modulated OLP progression through the PI3K/Akt signaling pathway. In conclusion, circulating exosomal miR-34a-5p could be a potential biomarker for evaluating the severity of OLP.