Exosomal mRNAs Research Articles

Characteristic changes in the mRNA expression profile of plasma exosomes from patients with MPO-ANCA-associated vasculitis and its possible correlations with pathogenesis

To explore the expression patterns and potential roles of mRNAs in exosomes from patients with myeloperoxidase-specific anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (MPO-AAV). Plasma exosomes were isolated from MPO-AAV patients and healthy controls (HCs) to screen for differential mRNA expression via exosomal mRNA sequencing. The differentially expressed mRNAs in exosomes from the 2 groups were comparatively explored by bioinformatics analysis. The six most differentially expressed mRNAs were selected and validated in larger groups of MPO-AAV patients and HCs by real-time quantitative polymerase chain reaction (RT‒qPCR). The relationships between these selected mRNAs and patient characteristics were statistically analyzed. Compared with HCs, a total of 1077 mRNAs in exosomes from MPO-AAV patients were found to be significantly upregulated, including DEPDC1B and TPST1, while NSUN4 and AK4 were significantly downregulated. Statistical analysis did not reveal any correlation between the six selected mRNAs and clinical indicators, including disease activity. GO enrichment analysis revealed that these differentially expressed genes participate in various enzyme activities, protein synthesis, etc. KEGG pathway analysis revealed that metabolic pathways, cell adhesion molecules, epithelial signaling, and mitogen-activated protein kinase (MAPK) signaling pathways were significantly enriched in the exosomal mRNAs. There were significant differences in the expression of exosomal mRNAs between MPO-AAV patients and HCs, which may be related to the occurrence and development of MPO-AAV. These findings provide clues for further investigations of MPO-AAV pathogenesis and the identification of new potential therapeutic targets.

Unveiling potential: urinary exosomal mRNAs as non-invasive biomarkers for early prostate cancer diagnosis

BackgroundThis study investigated the use of urinary exosomal mRNA as a potential biomarker for the early detection of prostate cancer (PCa).MethodsNext-generation sequencing was utilized to analyze exosomal RNA from 10 individuals with confirmed PCa and 10 individuals without cancer. Subsequent validation through qRT-PCR in a larger sample of 43 PCa patients and 92 healthy controls revealed distinct mRNA signatures associated with PCa.ResultsNotably, mRNAs for RAB5B, WWP1, HIST2H2BF, ZFY, MARK2, PASK, RBM10, and NRSN2 showed promise as diagnostic markers, with AUC values between 0.799 and 0.906 and significance p values. Combining RAB5B and WWP1 in an exoRNA diagnostic model outperformed traditional PSA tests, achieving an AUC of 0.923, 81.4% sensitivity, and 89.1% specificity.ConclusionsThese findings highlight the potential of urinary exosomal mRNA profiling, particularly focusing on RAB5B and WWP1, as a valuable strategy for improving the early detection of PCa.

Unraveling the Prefrontal Cortex-Basolateral Amygdala Pathway's Role on Schizophrenia's Cognitive Impairments: A Multimodal Study in Patients and Mouse Models.

This study investigated the role of the medial prefrontal cortex (mPFC)-basolateral amygdala (BLA) pathway in schizophrenia (SCZ)-related cognitive impairments using various techniques. This study utilized clinical scales, magnetic resonance imaging, single-cell RNA sequencing, and optogenetics to investigate the mPFC-BLA pathway in SCZ patients. In the mouse model, 6-week-old methylazoxymethanol acetate-induced mice demonstrated significant cognitive deficits, which were addressed through stereotaxic injections of an adeno-associated viral vector to unveil the neural connection between the mPFC and BLA. Significant disparities in brain volume and neural activity, particularly in the dorsolateral prefrontal cortex (DLPFC) and BLA regions, were found between SCZ patients and healthy controls. Additionally, we observed correlations indicating that reduced volumes of the DLPFC and BLA were associated with lower cognitive function scores. Activation of the mPFC-BLA pathway notably improved cognitive performance in the SCZ model mice, with the targeting of excitatory or inhibitory neurons alone failing to replicate this effect. Single-cell transcriptomic profiling revealed gene expression differences in excitatory and inhibitory neurons in the BLA of SCZ model mice. Notably, genes differentially expressed in the BLA of these model mice were also found in the blood exosomes of SCZ patients. Our research provides a comprehensive understanding of the role of the PFC-BLA pathway in SCZ, underscoring its significance in cognitive impairment and offering novel diagnostic and therapeutic avenues. Additionally, our research highlights the potential of blood exosomal mRNAs as noninvasive biomarkers for SCZ diagnosis, underscoring the clinical feasibility and utility of this method.

Urinary exosomal mRNAs as biomarkers for predicting the therapeutic effect of renin-angiotensin system inhibitors in IgA nephropathy patients

BackgroundRenin-angiotensin system inhibitors (RASi) treatment is the basic therapy for IgA nephropathy (IgAN) patients. However, there is few of biomarker that can predict the efficacy of RASi. This study aimed to find urinary exosomal mRNAs related to the therapeutic effect of RASi in the treatment of proteinuria in IgAN patients. Methods: We divided IgAN patients in screening cohort into A1 (proteinuria increase at 3 months), B1 (proteinuria decrease less than 50 % at 3 months), C1 (proteinuria decrease more than 50 % at 3 months) groups according to changes of proteinuria after treatment. The urinary exosomes were collected before biopsy, RNAs were extracted and analyzed with the microarray assay. The candidate genes were screened by differentially expressed genes (DEGs) analysis and then validated by quantitative real-time polymerase chain reaction (qPCR) in a validation cohort. A receiver operating characteristic (ROC) curve was used to evaluate gene performance in predicting therapeutic effect on RASi reducing proteinuria in IgAN patients. Results: ECE1 and PDE1A mRNAs were significantly different among the three groups, and were gradually decreased among A1, B1 and C1 groups. In the validation cohort, the level of urinary exosomal ECE1 and PDE1A mRNAs were also significantly lower in A2 group compared with C2 group(ECE1, P < 0.001;PDE1A, P < 0.01). Besides, the level of ECE1 mRNA was also lower in B2 group compared with C2 group (P < 0.01). The ROC curve verified that urinary exosomal ECE1 and PDE1A gene level predicted RASi efficacy in IgAN patients with area under curve (AUC) 0.68 and 0.63 respectively. Conclusion: Urinary exosomal ECE1 and PDE1A mRNAs expression can serve as potential biomarkers for predicting the RASi efficacy to reduce proteinuria in IgAN patients.

Modulating the mesenchymal stromal cell microenvironment alters exosome RNA content and ligament healing capacity.

Although mesenchymal stromal cell (MSC) based therapies hold promise in regenerative medicine, their clinical application remains challenging due to issues such as immunocompatibility. MSC-derived exosomes are a promising off-the-shelf therapy for promoting wound healing in a cell-free manner. However, the potential to customize the content of MSC-exosomes, and understanding how such modifications influence exosome effects on tissue regeneration remain underexplored. In this study, we used an in vitro system to compare the priming of human MSCs by 2 inflammatory inducers TNF-α and CRX-527 (a highly potent synthetic TLR4 agonist that can be used as a vaccine adjuvant or to induce anti-tumor immunity) on exosome molecular cargo, as well as on an in vivo rat ligament injury model to validate exosome potency. Different microenvironmental stimuli used to prime MSCs in vitro affected their exosomal microRNAs and mRNAs, influencing ligament healing. Exosomes derived from untreated MSCs significantly enhance the mechanical properties of healing ligaments, in contrast to those obtained from MSCs primed with inflammation-inducers, which not only fail to provide any improvement but also potentially deteriorate the mechanical properties. Additionally, a link was identified between altered exosomal microRNA levels and expression changes in microRNA targets in ligaments. These findings elucidate the nuanced interplay between MSCs, their exosomes, and tissue regeneration.

Abstract 2431: Identification of highly expressed exosomal RNAs derived from pancreatic cancer cell lines and exploration of their potential as biomarkers

Abstract Background: Not all exosomes in the blood of cancer patients originate from cancer cells. This study evaluated the relationships between mRNAs and miRNAs in pancreatic cancer (PC) cell lines and those in exosomes secreted into the supernatant. We aimed to identify candidates for PC biomarkers from cancer-specific exosomal mRNAs and miRNAs. Materials and Methods: Three PC cell lines were utilized. Exosomes were isolated from the supernatant, and cellular and exosomal mRNAs and miRNAs were extracted. Subsequently, RNA sequencing (RNA-seq) was performed, and the data were analyzed using R software. Results: In the MIAPaCa-2 cell line, the expression of cellular mRNAs and corresponding exosomal mRNAs showed a strong correlation. In the other two cell lines, a moderate correlation was observed. Across all three cell lines, cellular miRNAs and corresponding exosomal miRNAs exhibited moderate correlation. We focused on RNAs that were lowly expressed in cells but highly expressed in exosomes and conducted differential expression analysis using R. Compared with healthy volunteers in the GEO database, some RNAs showed high expression in the supernatant and low expression in the plasma of healthy volunteers. Conclusion: We identified three mRNAs and three miRNAs that may serve as promising biomarker candidates for PC. Citation Format: Yuhan Rong, Naoki Okubo, Noritoshi Kobayashi, Eriko Katsuta, Yasushi Ichikawa. Identification of highly expressed exosomal RNAs derived from pancreatic cancer cell lines and exploration of their potential as biomarkers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2431.

Exosomal IGFBP2 derived from highly metastatic promotes hepatocellular carcinoma metastasis by inducing epithelial mesenchymal transition

Liver cancer metastasis is the main cause of death in liver cancer patients. Exosomes, which are small vesicles released by cancer cells, play a crucial role in the metastasis of cancer. The aim of this study was to investigate the effect of exosomes derived from high metastatic potential liver cancer cells acting as cell to cell communication on liver cancer metastasis. Bioinformatics analysis was used to obtain the differential expression of exosomal mRNAs from the plasma of both liver cancer patients and healthy volunteers. Transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and protein blot were employed to characterize the exosomes. The molecular mechanisms and were explored by conducting CCK8, Transwell, Tunel, RTqPCR, western blot, and immunofluorescence staining. We examined IGFBP2 special expression in the plasma exosomes of both liver cancer patients and healthy volunteers, and its presence was associated with a poor prognosis in liver cancer patients. Furthermore, we observed that exosomes from highly metastatic liver cancer cells (MHCC97H) contained high levels of IGFBP2 and could enhance the metastatic potential of less aggressive liver cancer cells (Hep3B). Additionally, we discovered that IGFBP2 in MHCC97H-derived exosomes activated ERK signaling pathway, which triggered epithelial-mesenchymal transition (EMT) in Hep3B cells. Our study underscores the significance of exosomal IGFBP2 from highly metastatic liver cancer cells as a driver of metastasis in less invasive liver cancer cells. This suggests that targeting IGFBP2 in exosomes could be a promising strategy for the treatment and prognosis of liver cancer patients.

B-383 Exosomal PRPSAP1 in Plasma Predicts Microvascular Invasion in Hepatocellular Carcinoma

Abstract Background Microvascular invasion (MVI), a critical predictor of postoperative tumor recurrence in hepatocellular carcinoma (HCC), is only detectable by histopathological examination of the surgical specimen. Prediction of MVI before surgery is of great significance to prognosis, especially by non-invasive means. Here, we report that exosome-derived mRNA in plasma can be promising biomarkers for MVI preoperatively. Methods Patients with initially suspected HCC who were undergoing hepatectomy were prospectively recruited with preoperative peripheral blood collection and followed up. Exosomal mRNA profiling was performed using RNA sequencing in the discovery cohort, and the edgeR package in R software was used to screen out significantly differential expression exosomal mRNAs between MVI and non-MVI groups. Multiplex droplet digital PCR (ddPCR) technology was utilized to verify filtered exosomal mRNAs in the validation cohort. Results A total of 131 HCC patients were enrolled with 37 in the discovery cohort and 94 in the validation cohort. RNA-sequencing data identified six candidate exosomal mRNAs (ZC3H3, RSAD2, PRPSAP1, HOXA2, CHPM4A and KCTD10) as potential predictors for MVI. In the validation cohort, expression levels of PRPSAP1 and HOXA2 were slightly higher and the expression level of CHMP4A was slightly lower in the MVI group than the non-MVI one, which were consistent with the discovery cohort though without statistically significant difference (P &amp;gt; 0.05). Further analysis between MVI number&amp;gt;5 and non-MVI patients validated that the expression level of exosomal PRPSAP1 was significantly higher in MVI number&amp;gt;5 patients (0.147 vs 0.070, P = 0.035). Altogether, exosomal PRPSAP1 in plasma was a promising biomarker predicting MVI for HCC patients. Conclusion We validated for the first time that high level expression of exosome-derived PRPSAP1 can preoperatively predict MVI, especially with the number &amp;gt;5, to improve prognosis.

RNA-Sequencing and Bioinformatics Analysis of Exosomal Long Noncoding RNAs Revealed a Novel ceRNA Network in Stable COPD.

Exosomes are able to exchange their bioactive RNA cargo to recipient cells. In COPD, exosomes can be controlled and engineered for its use as targeted diagnostic and therapeutic tool.Our study explored novel lncRNAs and mRNAs in plasma exosomes that could be involved in the pathogenesis of COPD. High-throughput sequencing was conducted to detect the alterations in the expression of exosomal lncRNAs and mRNAs. Gene ontology (GO) functional analyses and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were used to determine the significant functions and pathways associated with differentially expressed (DE) lncRNAs. The mRNA expression profile dataset, GSE76925, and microRNA expression profile dataset, GSE70080, were obtained from the GEO database. Venn diagrams were used to find common DE mRNAs between my mRNAs dataset and GSE76925. These common DEGs were subjected to PPI analyses to identify Hub genes. Subsequently, Venn diagrams were used to identify common genes between the target genes of DE-miRNAs and Hub genes as well as DE-miRNAs and my lncRNAs dataset. Finally, a lncRNA-miRNA-mRNA co-expression network was constructed byprediction using proprietary software. The lncRNA and mRNA expressions were then validated by quantitative reverse-transcription polymerase chain reaction (qRT-PCR). We identified 1578 differentially regulated lncRNAs and 3071 differentially regulated mRNAs. GO and KEGG pathway analyses suggested that the DE lncRNAs are involved in the pathogenesis of COPD. A lncRNA-miRNA-mRNA meshwork was established to predict the potential interactions among these RNAs. RP3-329A5.8 and MRPS11 expression was then subjected to qRT-PCR for validation. Correlations between MRPS11 and clinic-pathological features were explored. Our study provided a set of lncRNAs and mRNAs that may be involved in the pathogenesis of COPD, thereby highlighting the need for further research on both diagnostic biomarkers and molecular mechanisms.

Abstract 879: Exosomal mRNAs secreted from pancreatic cancer (PC) cell lines might include good candidates of biomarkers of PC patients

Abstract Background: Exosomal mRNAs secreted from cancer cells are regarded as important messengers in communication system between cells. In PC, some exosomal RNAs from patients’ plasma were expected as biomarkers of PC. However, exosomes in the blood of cancer patients are not derived only from cancer cells. Relationship of RNAs produced in cancer cells and those secreted into exosomes is still unclear. In this study, we evaluated relationships between mRNAs in PC cell lines and those of exosomes secreted into supernatant and determined candidates for PC biomarkers from cancer specific exosomal mRNAs. Material and Method: Three PC cell lines (PANC-1, MIAPaCa-2 and Capan-1) were used. Three studies were performed for each cell line. After 48h serum-free culture, the supernatants were collected, and exosomes were isolated by ultracentrifugation. The remaining cells in the culture dish were also collected and mRNAs were extracted. RNA-seq was performed for mRNAs extracted from cell and exosomal RNAs extracted from supernatant. Data was analyzed by R (version 4.2.1). Results: In MIAPaCa-2, expression of cellular mRNAs and corresponding exosomal mRNAs showed strong correlation (R was higher than 0.7). In the other 2 cell lines, they showed moderate correlation (R was higher than 0.5). Then, expression of mRNA in the cells was approximately reflected in those in the exosomes. However, some mRNAs showed high expression in exosome and low expression in the cell. We focused on these mRNAs, and then, performed a differential analysis of the whole raw RNA data by using TCC package of R and found differentially expressed mRNAs. Conclusion: Comparing with data of GEO database showing exosomal mRNA extracted from healthy volunteers, we finally find candidates of cancer specific exosomal mRNAs of PC. Citation Format: Yuhan Rong, Naoki Okubo, Risa Fukui, Akihiro Suzuki, Motokazu Sato, Motohiko Tokuhisa, Yuma Takeda, Noritoshi Kobayashi, Itaru Endo, Yasushi Ichikawa. Exosomal mRNAs secreted from pancreatic cancer (PC) cell lines might include good candidates of biomarkers of PC patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 879.

The predictive value of plasma exosomal lncRNAs/mRNAs in NSCLC patients receiving immunotherapy

PurposeThere is an urgent need to explore the use of plasma-derived exosomal long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs) as potential biomarkers to select the most suitable patient population to receive immunotherapy for advanced NSCLC with no actionable molecular markers. Patients and methodsIn the present study, 7 patients with advanced NSCLC who received nivolumab were enrolled for molecular studies. Plasma-derived exosomal lncRNAs/mRNAs expression profiles differed between patients exhibiting differential immunotherapy efficacy. ResultsIn the non-responders, 299 differentially expressed exosomal mRNAs and 154 lncRNAs were significantly upregulated. In GEPIA2, 10 mRNAs were upregulated in the NSCLC patients compared to that of the normal population. The up-regulation of CCNB1 related to the cis-regulation of lnc-CENPH-1 and lnc-CENPH-2. KPNA2, MRPL3, NET1 and CCNB1 were trans-regulated by lnc-ZFP3-3. In addition, IL6R exhibited a trend of increased expression in the non-responders at baseline, and this expression was further downregulated after treatment in responders. The association between CCNB1 with lnc-CENPH-1 and lnc-CENPH-2, as well as the lnc-ZFP3-3-TAF1 pair, may represent potential biomarkers of poor immunotherapy efficacy. Patients may obtain increased effector T cell function when IL6R is suppressed by immunotherapy. ConclusionsOur study suggests that plasma-derived exosomal lncRNA and mRNA expression profiles differ between responders and non-responders to nivolumab immunotherapy. Lnc-ZFP3-3-TAF1-CCNB1 pair and IL6R might be key factors predicting efficiency of immunotherapy. Large scale clinical studies seem warranted to further validate the potential of plasma-derived exosomal lncRNAs and mRNAs as a biomarker to aid the selection of NSCLC patients for nivolumab immunotherapy.

Uterine Flushing Fluid-Derived Let-7b Targets CXCL10 to Regulate Uterine Receptivity in Goats during Embryo Implantation.

Exosomes have the ability to carry a wide range of chemicals, convey them to target cells or target regions, and act as "messengers." For the purpose of investigating embryo attachment, it is helpful to comprehend the range of exosomal mRNAs and miRNAs derived from the uterine flushing fluid before and after embryo attachment. In this study, we recovered exosomes from goat uterine rinsing fluid at 5, 15, and 18 days of gestation and used RNA-Seq to identify the mRNA and miRNA profiles of exosomes obtained from uterine rinsing fluid before and after embryo implantation. In total, 91 differently expressed miRNAs and 27,487 differentially expressed mRNAs were found. The target genes predicted by the differentially expressed miRNAs and the differentially expressed mRNAs were mainly membrane-related organelles with catalytic activity, binding activity, transcriptional regulation activity, and involved in metabolism, biological regulation, development, and other processes. This was revealed by GO analysis. Furthermore, KEGG analysis revealed that they were abundant in signaling pathways associated with embryo implantation, including the "PI3K-Akt signaling pathway," "Toll-like receptor signaling pathway," "TGF-beta signaling route," "Notch signaling pathway," and others. Moreover, our research has demonstrated, for the first time, that chi-let-7b-5p specifically targets the 3'UTR of CXCL10. Our research offers a fresh viewpoint on the mechanics of embryo attachment.

Dynamic immune and exosome transcriptomic responses in patients undergoing psychostimulant methamphetamine withdrawal.

Methamphetamine (METH) addiction and withdrawal cause serious harm to both the immune system and nervous system. However, the pathogenesis remains largely unknown. Herein, we investigated the peripheral cytokines and exosomal transcriptome regulatory networks in the patients with METH use disorders (MUDs) undergoing withdrawal. Twenty-seven cytokines were simultaneously assessed in 51 subjects, including 22 at the acute withdrawal (AW) stage and 29 at the protracted withdrawal (PW) stage, and 31 age and gender-matched healthy controls (HCs). Compared to the HCs, significantly decreased levels of interleukin (IL)-1β, IL-9, IL-15, Basic FGF, and MIP1a, increased levels of IL-1rα, IL-6, Eotaxin IP-10, VEGF, and RANTES were identified in AW. These disturbances were mostly or partly restored to the baseline in PW. However, the cytokines IL-6, IL-7, and IL-12p70 were consistently increased even after one year of withdrawal. Besides, a significant decrease in CD3+T and CD4+T cell numbers was observed in AW, and the diminishment was restored to baseline in PW. Comparatively, there were no statistically significant changes in CD8+T, NK, and B cells. Furthermore, the exosomal mRNAs and long non-coding RNAs (lncRNA) were profiled, and the lncRNA-miRNA-mRNA networks were constructed and associated with METH AW and PW stages. Notably, the chemokine signaling was remarkably upregulated during AW. By contrast, the differentially expressed mRNAs/lincRNAs were significantly enriched in neurodegeneration-related diseases. Taken together, a group of METH withdrawal-related cytokines and exosomal mRNA/lncRNA regulatory networks were obtained, which provides a useful experimental and theoretical basis for further understanding of the pathogenesis of the withdrawal symptoms in MUDs.

Identification of Diagnostic Exosomal LncRNA-miRNA-mRNA Biomarkers in Colorectal Cancer Based on the ceRNA Network.

Background: Colorectal cancer (CRC) is currently the fourth most common cancer worldwide. The roles of exosomal competing endogenous RNAs (ceRNAs) in CRC remain unclear. In this study, we constructed an exosomal ceRNA network to identify the core ceRNAs and investigate the diagnostic biomarkers in CRC. Methods and Patients: Serum exosomes were isolated from four CRC patients and two healthy donors by ultracentrifugation, and then subjected to RNA isolation, sequencing and microarray. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) analyses were performed to identify functional enrichment implications of differentially expressed exosomal mRNAs. TargetScan and miRanda were used for identifying the miRNA-mRNA and miRNA-LncRNA interactions. The predicted lncRNAs and mRNAs were intersected with the differentially expressed genes, for which the screening criterion was fold change >1.5 in the microarray. Differentially expressed exosomal miRNAs were identified in the GSE71008 dataset, and differentially expressed mRNAs (DEmRNAs) were further summarized from The Cancer Genome Atlas (TCGA) database. Results: A total of 1186 exosomal DEmRNAs, 2088 exosomal DElncRNAs and 29 exosomal miRNAs were detected in CRC patients compared to the healthy donors. Functional enrichment analysis suggested that exosomal DEmRNAs might participate in pathways related to carcinogenesis and development of cancer. An exosomal ceRNA regulatory network of CRC was constructed based on 40 lncRNAs, two miRNAs, and five mRNAs. Exosomal miR-150-5p and miR-10b-5p expression levels were increased in healthy donors compared with CRC patients in the GSE71008 dataset, and five DEmRNAs (TOMM70A, RBM48, BEND3, RHOBTB1, and ADAMTS2) were significantly upregulated in TCGA database. Two potential exosomal regulatory axes of lncRNA G016261-miR-150-5p-RBM48 and lncRNA XLOC_011677-miR-10b-5p-BEND3 were identified from the network. Conclusion: The current study revealed potential molecular biological regulation pathways and diagnostic biomarkers through the exosomal ceRNA regulatory network.

Plasma-derived exosomal mRNA profiles associated with type 1 diabetes mellitus.

BackgroundExosomes carry various types of transcripts, such as messenger RNAs (mRNAs), and play an important role in mediating cell-to-cell communication, thus influencing multiple physiological and pathological processes. However, the role of exosomal mRNAs in T1DM is largely unknown. Here, we aimed to identify the plasma-derived exosomal mRNA expression profiles in T1DM and to explore their potential biological functions in T1DM.Materials and MethodsPlasma-derived exosomes were isolated from 10 patients with T1DM and 10 age- and sex-matched control subjects by size exclusion chromatography methods. Transmission electron microscopy, nanoparticle tracking analysis, and western blot analysis confirmed the presence of exosomes. The exosomal mRNAs were analyzed using the Illumina HiSeq platform. Six differentially expressed mRNAs (DEMs) were randomly selected to determine the expression level by quantitative real-time PCR (qRT−PCR) in a larger cohort (T1DM subjects N=40; control subjects N=40). The biological functions of DEMs were predicted by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Protein−protein interaction networks were constructed to explore the potential associations among DEMs.ResultsIn total, 112 DEMs were identified in T1DM, among which 66 mRNAs were upregulated and 46 mRNAs were downregulated. Four of six candidate exosomal mRNAs were successfully validated by qRT−PCR. Bioinformatics analysis indicated that these mRNAs were most significantly involved in positive regulation by host viral transcription (GO enrichment analysis) and oxidative phosphorylation (KEGG pathway analysis).ConclusionsOur study reported the plasma-derived exosomal mRNA expression profiles of T1DM for the first time. The identified DEMs might be associated with the pathogenesis of T1DM, and some DEMs have the potential to serve as biomarkers and therapeutic targets for T1DM.

Reorganization of the Landscape of Translated mRNAs in NSUN2-Deficient Cells and Specific Features of NSUN2 Target mRNAs.

The RNA cytosine C5 methyltransferase NSUN2 has a variety of RNA substrates and plays an important role in mRNA metabolism. NSUN2 binds to specific sequences enriched in exosomal mRNAs, suggesting its possible involvement in the sorting of mRNAs into exosomes. We applied the photoactivatable.4-thiouridine-enhanced cross-linking and immunoprecipitation assay involving high-throughput RNA sequencing (RNA-seq) to HEK293T cells to determine NSUN2 mRNA targets. NSUN2 cross-linking sites were found in more than one hundred relatively abundant mRNAs with a high GC content and a pronounced secondary structure. Then, utilizing RNA-seq for the total and polysome-associated mRNA from HEK293T cells with and without the knockdown of NSUN2, we identified differentially expressed genes, as well as genes with altered translational efficiency (GATEs). It turned out that the up-regulated GATE mRNAs were much shorter on average than the down-regulated ones, and their GC content was higher; moreover, they contained motifs with C residues located in GC-rich environments. Our findings reveal the specific features of mRNAs that make them potential targets for NSUN2 and expand our understanding of the role of NSUN2 in controlling translation and, possibly, in mRNA sorting into exosomes implemented through the methylation of cytosine residues.

Bladder cancer-derived exosomal KRT6B promotes invasion and metastasis by inducing EMT and regulating the immune microenvironment

BackgroundTumour-derived exosomes have recently been shown to participate in the formation and progression of different cancer processes, including tumour microenvironment remodelling, angiogenesis, invasion, metastasis, and drug resistance. However, the function and mechanism of exosome-encapsulated nucleic acids and proteins in bladder cancer remain unclear. This study aimed to investigate the effects of tumour-derived exosomes on the tumorigenesis and development of bladder cancer.MethodsIn this study, gene expression profiles and clinical information were collected from The Cancer Genome Atlas (TCGA) database and two independent Gene Expression Omnibus (GEO) datasets. The nucleic acids and proteins encapsulated in bladder cancer-derived exosomes were obtained from the ExoCarta database. Based on these databases, the expression patterns of exosomal mRNAs and proteins and the matched clinicopathological characteristics were analysed. Furthermore, we carried out a series of experiments to verify the relevant findings.ResultsA total of 7280 differentially expressed mRNAs were found in TCGA data, of which 52 mRNAs were shown to be encapsulated in bladder cancer-derived exosomes. Survival analysis based on the UALCAN database showed that among the top 10 upregulated and the top 10 downregulated exosomal genes, only the expression of KRT6B had a statistically significant effect on the survival of bladder cancer patients. Additionally, clinical correlation analysis showed that the elevated level of KRT6B was highly associated with bladder cancer stage, grade, and metastasis status. GSEA revealed that KRT6B was involved not only in epithelial–mesenchymal transition-related pathways but also in the immune response in bladder cancer. Ultimately, our experimental results were also consistent with the bioinformatic analysis.ConclusionKRT6B, which can be detected in bladder cancer-derived exosomes, plays an important role in the epithelial–mesenchymal transition and immune responses in bladder cancer. Further research will enable its potentially prognostic marker and therapeutic target for bladder cancer.

CircRNA, lncRNA, and mRNA profiles of umbilical cord blood exosomes from preterm newborns showing bronchopulmonary dysplasia

Bronchopulmonary dysplasia (BPD) represents a multifactorial chronic pulmonary pathology and a major factor causing premature illness and death. The therapeutic role of exosomes in BPD has been feverishly investigated. Meanwhile, the potential roles of exosomal circRNAs, lncRNAs, and mRNAs in umbilical cord blood (UCB) serum have not been studied. This study aimed to detect the expression profiles of circRNAs, lncRNAs, and mRNAs in UCB-derived exosomes of infants with BPD. Microarray analysis was performed to compare the RNA profiles of UCB-derived exosomes of a preterm newborn with (BPD group) and without (non-BPD, NBPD group) BPD. Then, circRNA/lncRNA–miRNA–mRNA co-expression networks were built to determine their association with BPD. In addition, cell counting kit-8 (CCK-8) assay was used to evaluate the proliferation of lipopolysaccharide (LPS)-induced human bronchial epithelial cells (BEAS-2B cells) and human umbilical vein endothelial cells (HUVECs). The levels of tumor necrosis factor (TNF)-α and interleukin (IL)-1β in LPS-induced BEAS-2B cells and HUVECs were assessed through Western blot analysis. Then, quantitative reverse transcription–polymerase chain reaction assay was used to evaluate the expression levels of four differentially expressed circRNAs (hsa_circ_0086913, hsa_circ_0049170, hsa_circ_0087059, and hsa_circ_0065188) and two lncRNAs (small nucleolar RNA host gene 20 (SNHG20) and LINC00582) detected in LPS-induced BEAS-2B cells or HUVECs. A total of 317 circRNAs, 104 lncRNAs, and 135 mRNAs showed significant differential expression in UCB-derived exosomes of preterm infants with BPD compared with those with NBPD. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted to examine differentially expressed exosomal circRNAs, lncRNAs, and mRNAs. The results showed that the GO terms and KEGG pathways mostly involving differentially expressed exosomal RNAs were closely associated with endothelial or epithelial cell development. In vitro, CCK-8 and Western blot assays revealed that LPS remarkably inhibited the viability and promoted inflammatory responses (TNF-α and IL-1β) of BEAS-2B cells or HUVECs. The expression levels of circRNAs hsa_circ_0049170 and hsa_circ_0087059 were upregulated in LPS-induced BEAS-2B cells; the expression level of hsa_circ_0086913 was upregulated and that of hsa_circ_0065188 was downregulated in LPS-induced HUVECs. Moreover, the expression level of lncRNA SNHG20 was upregulated and that of LINC00582 was downregulated in LPS-induced BEAS-2B cells. Further, 455 circRNA/lncRNA–miRNA–mRNA interaction networks were predicted, including hsa_circ_0086913/hsa-miR-103a-3p/transmembrane 4 L six family member 1 (TM4SF1) and lncRNA-SNHG20/hsa-miR-6720-5p/spermine synthase (SMS) networks, which may take part in BPD.Conclusion: This study provided a systematic perspective on UCB-derived exosomal circRNAs and lncRNAs and laid an important foundation for further investigating the potential biological functions of exosomal circRNAs and lncRNAs in BPD.What is Known:• BPD represents a multifactorial chronic pulmonary pathology and a major factor causing premature illness and death.• The therapeutic role of exosomes in BPD has been feverishly investigated, and exosomal RNAs were ignored.What is New:• The profiles of UCB-derived exosomal circRNAs, lncRNAs, and mRNAs were performed.• Several differentially expressed circRNAs and lncRNAs were identified in LPS-induced BEAS-2B cells and HUVECs.

Association Between Mitochondrial Function and Rehabilitation of Parkinson's Disease: Revealed by Exosomal mRNA and lncRNA Expression Profiles.

Rehabilitation has been proposed as a valid measure complementary to the management of Parkinson's disease (PD). However, the mechanism underlying is not clear yet. The differential expressions of exosomal messenger RNA (mRNA) and long noncoding RNAs (lncRNAs) may play a critical role in PD progression and rehabilitation. To compare the differential expressions of exosomal mRNAs and lncRNAs, patients with PD (PWPs, Hoehn and Yahr stages 1.5-2.5, n = 6) and age- and sex-matched healthy controls (HCs, n = 6) were included in this study. All PWPs received a 2-week rehabilitation treatment in the hospital, which seemingly led to improvement in both the motor and non-motor functions. A set of differentially expressed mRNAs (DEmRNAs) and differentially expressed lncRNAs (DElncRNAs) extracted from exosomes in blood samples via next-generation sequencing (NGS) was screened out. Compared to HCs, 2,337 vs. 701 mRNAs and 1,278 vs. 445 lncRNAs were significantly upregulated and significantly downregulated, respectively, in pre-rehabilitation (pre-rehab) PWPs; 2,490 vs. 629 mRNAs and 1,561 vs. 370 lncRNAs were significantly upregulated and significantly downregulated, respectively, in post-rehabilitation (post-rehab) PWPs. Compared to pre-rehab PWPs, 606 vs. 1,056 mRNAs and 593 vs. 1,136 lncRNAs were significantly upregulated and significantly downregulated, respectively, in post-rehab PWPs. Overall, 14 differentially expressed mRNAs (DEmRNAs) and 73 differentially expressed lncRNAs (DElncRNAs) were expressed in the blood exosomes of HCs, pre- and post-rehab PWPs, simultaneously. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses identified 243 significantly co-expressed lncRNA-mRNA pairs. One DEmRNA of interest (ENSG00000099795, NDUFB7) and three corresponding DElncRNAs (ENST00000564683, ENST00000570408, and ENST00000628340) were positively related. Quantitative real-time polymerase chain reaction (qRT-PCR) validated that the expression levels of NDUFB7 mRNA and the 3 DElncRNAs increased significantly in pre-rehab PWPs, but decreased significantly in post-rehab PWPs compared to HCs. NDUFB7 mRNA is a marker related to mitochondrial respiration. It is reasonably believed that mitochondrial function is associated with PD rehabilitation, and the mitochondrial pathway may involve in the pathogenesis of PD.

Exosomal RNAs: Novel Potential Biomarkers for Diseases-A Review.

Exosomes are a subset of nano-sized extracellular vesicles originating from endosomes. Exosomes mediate cell-to-cell communication with their cargos, which includes mRNAs, miRNAs, lncRNAs, and circRNAs. Exosomal RNAs have cell specificity and reflect the conditions of their donor cells. Notably, their detection in biofluids can be used as a diagnostic marker for various diseases. Exosomal RNAs are ideal biomarkers because their surrounding membranes confer stability and they are detectable in almost all biofluids, which helps to reduce trauma and avoid invasive examinations. However, knowledge of exosomal biomarkers remains scarce. The present review summarizes the biogenesis, secretion, and uptake of exosomes, the current researches exploring exosomal mRNAs, miRNAs, lncRNAs, and circRNAs as potential biomarkers for the diagnosis of human diseases, as well as recent techniques of exosome isolation.