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MicroRNAs (miRNAs) and isoforms of their archetype, called isomiRs, regulate gene expression via complementary base-pair binding to messenger RNAs (mRNAs). The partially evolutionarily conserved isomiR sequence variations are differentially expressed among tissues, populations, and genders, and between healthy and diseased states. Aiming towards the clinical use of isomiRs as diagnostic biomarkers and for therapeutic purposes, several challenges need to be addressed, including (i) clarification of isomiR definition, (ii) improved annotation in databases with new standardization (such as the mirGFF3 format), and (iii) improved methods of isomiR detection, functional verification, and in silico analysis. In this review we discuss the respective challenges, and highlight the opportunities for clinical use of isomiRs, especially in the light of increasing amounts of next-generation sequencing (NGS) data.
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Non-thermal plasma, a partially ionized gas, holds significant potential for clinical applications, including wound-healing support, oral therapies, and anti-tumour treatments. While its applications showed promising outcomes, the underlying molecular mechanisms remain incompletely understood. We thus apply non-thermal plasma to mouse auricular skin and conducted non-coding RNA sequencing, as well as single-cell blood sequencing. In a time-series analysis (five timepoints spanning 2 hours), we compare the expression of microRNAs in the plasma-treated left ears to the unexposed right ears of the same mice as well as to the ears of unexposed control mice. Our findings indicate specific effects in the treated ears for a set of five miRNAs: mmu-miR-144-5p, mmu-miR-144-3p, mmu-miR-142a-5p, mmu-miR-223-3p, and mmu-miR-451a. Interestingly, mmu-miR-223-3p also exhibits an increase over time in the right non-treated ear of the exposed mice, suggesting systemic effects. Notably, this miRNA, along with mmu-miR-142a-5p and mmu-miR-144-3p, regulates genes and pathways associated with wound healing and tissue regeneration (namely ErbB, FoxO, Hippo, and PI3K-Akt signalling). This co-regulation is particularly remarkable considering the significant seed dissimilarities among the miRNAs. Finally, single-cell sequencing of PBMCs reveals the downregulation of 12 from 15 target genes in B-cells, Cd4+ and Cd8+ T-cells. Collectively, our data provide evidence for a systemic effect of non-thermal plasma.
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Regulación de la Expresión Génica , MicroARNs , Gases em Plasma , Piel , MicroARNs/genética , Animales , Ratones , Piel/metabolismo , Gases em Plasma/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Perfilación de la Expresión Génica , Cicatrización de Heridas/efectos de los fármacos , Transducción de Señal , Sistema Inmunológico/metabolismoRESUMEN
The massive amount of human biological, imaging, and clinical data produced by multiple and diverse sources necessitates integrative modeling approaches able to summarize all this information into answers to specific clinical questions. In this paper, we present a hypermodeling scheme able to combine models of diverse cancer aspects regardless of their underlying method or scale. Describing tissue-scale cancer cell proliferation, biomechanical tumor growth, nutrient transport, genomic-scale aberrant cancer cell metabolism, and cell-signaling pathways that regulate the cellular response to therapy, the hypermodel integrates mutation, miRNA expression, imaging, and clinical data. The constituting hypomodels, as well as their orchestration and links, are described. Two specific cancer types, Wilms tumor (nephroblastoma) and non-small cell lung cancer, are addressed as proof-of-concept study cases. Personalized simulations of the actual anatomy of a patient have been conducted. The hypermodel has also been applied to predict tumor control after radiotherapy and the relationship between tumor proliferative activity and response to neoadjuvant chemotherapy. Our innovative hypermodel holds promise as a digital twin-based clinical decision support system and as the core of future in silico trial platforms, although additional retrospective adaptation and validation are necessary.
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Osteolyses are common findings in elderly patients and most frequently represent malignant or locally aggressive bone tumors, infection, inflammatory and endocrine disorders, histiocytoses, and rare diseases such as Gorham-Stout syndrome. We here report on a novel entity of massive multifocal osteolyses in both shoulders, the right hip and left knee joint and the dens of an 83-year-old patient not relatable to any previously known etiopathology of bone disorders. The soft tissue mass is of myxoid stroma with an unspecific granulomatous inflammatory process, aggressively destroying extensive cortical and cancellous bone segments and encroaching on articulating bones in diarthrodial large joints. Radiological, nuclear medical, serological, histological, and immunohistochemical analyses were incapable of further classifying the disease pattern within the existing scheme of pathology. Quantitative polymerase chain reaction and next generation sequencing revealed that mutations are not suggestive of any known hereditary or acquired bone disease. Possible treatment options include radionuclide therapy for pain palliation and percutaneous radiation to arrest bone resorption while surgical treatment is inevitable for pathological fractures. This case study shall increase the awareness of the musculoskeletal community and motivate to collect further information on this rare but mutilating disorder.
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The identification of targetomes remains a challenge given the pleiotropic effect of miRNAs, the limited effects of miRNAs on individual targets, and the sheer number of estimated miRNA-target gene interactions (MTIs), which is around 44,571,700. Currently, targetome identification for single miRNAs relies on computational evidence and functional studies covering smaller numbers of targets. To ensure that the targetome analysis could be experimentally verified by functional assays, we employed a systematic approach and explored the targetomes of four miRNAs (miR-129-5p, miR-129-1-3p, miR-133b, and miR-873-5p) by analyzing 410 predicted target genes, both of which were previously associated with Parkinson's disease (PD). After performing 13,536 transfections, we validated 442 of the 705 putative MTIs (62,7%) through dual luciferase reporter assays. These analyses increased the number of validated MTIs by at least 2.1-fold for miR-133b and by a maximum of 24.3-fold for miR-873-5p. Our study contributes to the experimental capture of miRNA targetomes by addressing i) the ratio of experimentally verified MTIs to predicted MTIs, ii) the sizes of disease-related miRNA targetomes, and iii) the density of MTI networks. A web service to support the analyses on the MTI level is available online ( https://ccb-web.cs.uni-saarland.de/utr-seremato ), and all the data have been added to the miRATBase database ( https://ccb-web.cs.uni-saarland.de/miratbase ).
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Regiones no Traducidas 3' , MicroARNs , Enfermedad de Parkinson , MicroARNs/genética , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismo , Humanos , Regulación de la Expresión Génica , Biología Computacional/métodos , Redes Reguladoras de Genes , Biblioteca de GenesRESUMEN
Molecular mechanisms of organismal and cell aging remain incompletely understood. We, therefore, generated a body-wide map of noncoding RNA (ncRNA) expression in aging (16 organs at ten timepoints from 1 to 27 months) and rejuvenated mice. We found molecular aging trajectories are largely tissue-specific except for eight broadly deregulated microRNAs (miRNAs). Their individual abundance mirrors their presence in circulating plasma and extracellular vesicles (EVs) whereas tissue-specific ncRNAs were less present. For miR-29c-3p, we observe the largest correlation with aging in solid organs, plasma and EVs. In mice rejuvenated by heterochronic parabiosis, miR-29c-3p was the most prominent miRNA restored to similar levels found in young liver. miR-29c-3p targets the extracellular matrix and secretion pathways, known to be implicated in aging. We provide a map of organism-wide expression of ncRNAs with aging and rejuvenation and identify a set of broadly deregulated miRNAs, which may function as systemic regulators of aging via plasma and EVs.
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MicroARNs , Ratones , Animales , MicroARNs/metabolismo , Envejecimiento/genética , Hígado/metabolismo , ParabiosisRESUMEN
MicroRNAs (miRNAs) play indispensable roles in posttranscriptional gene regulation. Their cellular regulatory impact is determined not solely by their sheer number, which likely amounts to >2000 individual miRNAs in human, than by the regulatory effectiveness of single miRNAs. Although, one begins to develop an understanding of the complex mechanisms underlying miRNA-target interactions (MTIs), the overall knowledge of MTI functionality is still rather patchy. In this critical review, we summarize key features of mammalian MTIs. We especially highlight latest insights on (i) the dynamic make-up of miRNA binding sites including non-canonical binding sites, (ii) the cooperativity between miRNA binding sites, (iii) the adaptivity of MTIs through sequence modifications, (iv) the bearing of intra-cellular miRNA localization changes and (v) the role of cell type and cell status specific miRNA interaction partners. The MTI biology is discussed against the background of state-of-the-art approaches with particular emphasis on experimental strategies for evaluating miRNA functionality.
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Regulación de la Expresión Génica , MicroARNs , Animales , Humanos , Sitios de Unión , Mamíferos/genética , MicroARNs/metabolismoRESUMEN
During cervical carcinogenesis, T-helper (Th)-17 cells accumulate in the peripheral blood and tumor tissues of cancer patients. We previously demonstrated that Th17 cells are associated with therapy resistance as well as cervical cancer metastases and relapse; however, the underlying Th17-driven mechanisms are not fully understood. Here, using microarrays, we found that Th17 cells induced an epithelial-to-mesenchymal transition (EMT) phenotype of cervical cancer cells and promoted migration and invasion of 2D cultures and 3D spheroids via induction of microRNA miR-142-5p. As the responsible mechanism, we identified the subunits C and D of the succinate dehydrogenase (SDH) complex as new targets of miR-142-5p and provided evidence that Th17-miR-142-5p-dependent reduced expression of SDHC and SDHD mediated enhanced migration and invasion of cancer cells using small interfering RNAs (siRNAs) for SDHC and SDHD, and miR-142-5p inhibitors. Consistently, patients exhibited high levels of succinate in their serum associated with lymph node metastases and diminished expression of SDHD in patient biopsies correlated with increased numbers of Th17 cells. Correspondingly, a combination of weak or negative SDHD expression and a ratio of Th17/CD4+ T cells > 43.90% in situ was associated with reduced recurrence-free survival. In summary, we unraveled a previously unknown molecular mechanism by which Th17 cells promote cervical cancer progression and suggest evaluation of Th17 cells as a potential target for immunotherapy in cervical cancer.
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Previous work on murine models and humans demonstrated global as well as tissue-specific molecular ageing trajectories of RNAs. Extracellular vesicles (EVs) are membrane vesicles mediating the horizontal transfer of genetic information between different tissues. We sequenced small regulatory RNAs (sncRNAs) in two mouse plasma fractions at five time points across the lifespan from 2-18 months: (1) sncRNAs that are free-circulating (fc-RNA) and (2) sncRNAs bound outside or inside EVs (EV-RNA). Different sncRNA classes exhibit unique ageing patterns that vary between the fcRNA and EV-RNA fractions. While tRNAs showed the highest correlation with ageing in both fractions, rRNAs exhibited inverse correlation trajectories between the EV- and fc-fractions. For miRNAs, the EV-RNA fraction was exceptionally strongly associated with ageing, especially the miR-29 family in adipose tissues. Sequencing of sncRNAs and coding genes in fat tissue of an independent cohort of aged mice up to 27 months highlighted the pivotal role of miR-29a-3p and miR-29b-3p in ageing-related gene regulation that we validated in a third cohort by RT-qPCR.
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Vesículas Extracelulares , MicroARNs , ARN Pequeño no Traducido , Humanos , Ratones , Animales , MicroARNs/genética , MicroARNs/metabolismo , Vesículas Extracelulares/metabolismo , ARN Pequeño no Traducido/genética , ARN Pequeño no Traducido/metabolismo , ARN de Transferencia/metabolismo , Envejecimiento/genéticaRESUMEN
The expression levels of various genes involved in human spermatogenesis are influenced by microRNAs (miRNAs), specifically microRNA-23a/b-3p. While certain genes are essential for spermatogenesis and male germ cell function, the regulation of their expression remains unclear. This study aimed to investigate whether microRNA-23a/b-3p targets genes involved in spermatogenesis and the impact of this targeting on the expression levels of these genes in males with impaired fertility. In-silico prediction and dual-luciferase assays were used to determine the potential connections between microRNA-23a/b-3p overexpression and reduced expression levels of 16 target genes. Reverse transcription-quantitative PCR (RT-qPCR) was conducted on 41 oligoasthenozoospermic men receiving infertility treatment and 41 age-matched normozoospermic individuals to verify the lower expression level of target genes. By employing dual-luciferase assays, microRNA-23a-3p was found to directly target eight genes, namely NOL4, SOX6, GOLGA6C, PCDHA9, G2E3, ZNF695, CEP41, and RGPD1, while microRNA-23b-3p directly targeted three genes, namely SOX6, GOLGA6C, and ZNF695. The intentional alteration of the microRNA-23a/b binding site within the 3' untranslated regions (3'UTRs) of the eight genes resulted in the loss of responsiveness to microRNA-23a/b-3p. This confirmed that NOL4, SOX6, GOLGA6C, PCDHA9, and CEP41 are direct targets for microRNA-23a-3p, while NOL4, SOX6, and PCDHA9 are direct targets for microRNA-23b-3p. The sperm samples of oligoasthenozoospermic men had lower expression levels of target genes than age-matched normozoospermic men. Correlation analysis indicated a positive correlation between basic semen parameters and lower expression levels of target genes. The study suggests that microRNA-23a/b-3p plays a significant role in spermatogenesis by controlling the expression of target genes linked to males with impaired fertility and has an impact on basic semen parameters.
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Sperm motility is a prerequisite for achieving pregnancy, and alterations in sperm motility, along with sperm count and morphology, are commonly observed in subfertile men. The aim of the study was to determine whether the expression level of genes annotated with the Gene Ontology (GO) term 'sperm motility' differed in sperm collected from healthy men and men diagnosed with oligoasthenozoospermia. Reverse transcription quantitative real-time PCR (RT-qPCR), quantitative mass spectrometry (LC-MS/MS), and enrichment analyses were used to validate a set of 132 genes in 198 men present at an infertility clinic. Out of the 132 studied sperm-motility-associated genes, 114 showed differentially expressed levels in oligoasthenozoospermic men compared to those of normozoospermic controls using an RT-qPCR analysis. Of these, 94 genes showed a significantly lower expression level, and 20 genes showed a significantly higher expression level. An MS analysis of sperm from an independent cohort of healthy and subfertile men identified 692 differentially expressed proteins, of which 512 were significantly lower and 180 were significantly higher in oligoasthenozoospermic men compared to those of the normozoospermic controls. Of the 58 gene products quantified with both techniques, 48 (82.75%) showed concordant regulation. Besides the sperm-motility-associated proteins, the unbiased proteomics approach uncovered several novel proteins whose expression levels were specifically altered in abnormal sperm samples. Among these deregulated proteins, there was a clear overrepresentation of annotation terms related to sperm integrity, the cytoskeleton, and energy-related metabolism, as well as human phenotypes related to spermatogenesis and sperm-related abnormalities. These findings suggest that many of these proteins may serve as diagnostic markers of male infertility. Our study reveals an extended number of sperm-motility-associated genes with altered expression levels in the sperm of men with oligoasthenozoospermia. These genes and/or proteins can be used in the future for better assessments of male factor infertility.
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Infertilidad Masculina , Semen , Embarazo , Femenino , Humanos , Masculino , Semen/metabolismo , Cromatografía Liquida , Espectrometría de Masas en Tándem , Espermatozoides/metabolismo , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Fertilidad/genéticaRESUMEN
Although the proteome of sperm has been characterized, there is still a lack of high-throughput studies on dysregulated proteins in sperm from subfertile men, with only a few studies on the sperm proteome in asthenozoospermic and oligoasthenozoospermic men. Using liquid chromatography-mass spectrometry (LC-MS/MS) along with bioinformatics analyses, we investigated the proteomic landscape of sperm collected from subfertile men (n = 22), i.e., asthenozoospermic men (n = 13), oligoasthenozoospermic men (n = 9) and normozoospermic controls (n = 31). We identified 4412 proteins in human sperm. Out of these, 1336 differentially abundant proteins were identified in 70% of the samples. In subfertile men, 32 proteins showed a lower abundance level and 34 showed a higher abundance level when compared with normozoospermic men. Compared to normozoospermic controls, 95 and 8 proteins showed a lower abundance level, and 86 and 1 proteins showed a higher abundance level in asthenozoospermic and oligoasthenozoospermic men, respectively. Sperm motility and count were negatively correlated with 13 and 35 and positively correlated with 37 and 20 differentially abundant proteins in asthenozoospermic and oligoasthenozoospermic men, respectively. The combination of the proteins APCS, APOE, and FLOT1 discriminates subfertile males from normozoospermic controls with an AUC value of 0.95. Combined APOE and FN1 proteins discriminate asthenozoospermic men form controls with an AUC of 1, and combined RUVBL1 and TFKC oligoasthenozoospermic men with an AUC of 0.93. Using a proteomic approach, we revealed the proteomic landscape of sperm collected from asthenozoospermic or oligoasthenozoospermic men. Identified abundance changes of several specific proteins are likely to impact sperm function leading to subfertility. The data also provide evidence for the usefulness of specific proteins or protein combinations to support future diagnosis of male subfertility.
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Astenozoospermia , Proteoma , Humanos , Masculino , Proteoma/metabolismo , Proteómica , Cromatografía Liquida , Semen/metabolismo , Motilidad Espermática , Espectrometría de Masas en Tándem , Espermatozoides/metabolismo , Astenozoospermia/diagnóstico , Apolipoproteínas E , ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Proteínas Portadoras/metabolismo , ADN Helicasas/metabolismoRESUMEN
BACKGROUND: Although a pivotal role of microRNA (miRNA, miR) in the pathogenesis of Huntington's disease (HD) is increasingly recognized, the molecular functions of miRNAs in the pathomechanisms of HD await further elucidation. One of the miRNAs that have been associated with HD is miR-34a-5p, which was deregulated in the mouse R6/2 model and in human HD brain tissues. METHODS: The aim of our study was to demonstrate interactions between miR-34a-5p and HD associated genes. By computational means we predicted 12 801 potential target genes of miR-34a-5p. An in-silico pathway analysis revealed 22 potential miR-34a-5p target genes in the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway "Huntington's disease". RESULTS: Using our high-throughput miRNA interaction reporter assay (HiTmIR) we identified NDUFA9, TAF4B, NRF1, POLR2J2, DNALI1, HIP1, TGM2 and POLR2G as direct miR-34a-5p target genes. Direct binding of miR-34a-5p to target sites in the 3'UTRs of TAF4B, NDUFA9, HIP1 and NRF1 was verified by a mutagenesis HiTmIR assay and by determining endogenous protein levels for HIP1 and NDUFA9. STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) analysis identified protein-protein interaction networks associated with HD like "Glutamine Receptor Signaling Pathway" and "Calcium Ion Transmembrane Import Into Cytosol". CONCLUSION: Our study demonstrates multiple interactions between miR-34a-5p and HD associated target genes and thereby lays the ground for future therapeutic interventions using this miRNA.
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Enfermedad de Huntington , MicroARNs , Ratones , Animales , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Modelos Animales de Enfermedad , Mapas de Interacción de Proteínas , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Perfilación de la Expresión GénicaRESUMEN
Cytotoxic CD8+ T lymphocytes (CTL) eliminate infected cells or transformed tumor cells by releasing perforin-containing cytotoxic granules at the immunological synapse. The secretion of such granules depends on Ca2+-influx through store operated Ca2+ channels, formed by STIM (stromal interaction molecule)-activated Orai proteins. Whereas molecular mechanisms of the secretion machinery are well understood, much less is known about the molecular machinery that regulates the efficiency of Ca2+-dependent target cell killing. CTL killing efficiency is of high interest considering the number of studies on CD8+ T lymphocytes modified for clinical use. Here, we isolated total RNA from primary human cells: natural killer (NK) cells, non-stimulated CD8+ T-cells, and from Staphylococcus aureus enterotoxin A (SEA) stimulated CD8+ T-cells (SEA-CTL) and conducted whole genome expression profiling by microarray experiments. Based on differential expression analysis of the transcriptome data and analysis of master regulator genes, we identified 31 candidates which potentially regulate Ca2+-homeostasis in CTL. To investigate a putative function of these candidates in CTL cytotoxicity, we transfected either SEA-stimulated CTL (SEA-CTL) or antigen specific CD8+ T-cell clones (CTL-MART-1) with siRNAs specific against the identified candidates and analyzed the killing capacity using a real-time killing assay. In addition, we complemented the analysis by studying the effect of inhibitory substances acting on the candidate proteins if available. Finally, to unmask their involvement in Ca2+ dependent cytotoxicity, candidates were also analyzed under Ca2+-limiting conditions. Overall, we identified four hits, CCR5 (C-C chemokine receptor type five), KCNN4 (potassium calcium-activated channel subfamily N), RCAN3 (regulator of calcineurin) and BCL (B-cell lymphoma) 2 which clearly affect the efficiency of Ca2+ dependent cytotoxicity in CTL-MART-1 cells, CCR5, BCL2, and KCNN4 in a positive manner, and RCAN3 in a negative way.
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Antineoplásicos , Linfocitos T CD8-positivos , Humanos , Calcio , Citotoxicidad Inmunológica , Linfocitos T Citotóxicos , Células Asesinas NaturalesRESUMEN
Seminal plasma contains a variety of extracellular vesicles (EVs) that deliver RNAs including microRNAs (miRNAs) molecules. However, the roles of these EVs along with their delivered RNAs and their interactions with male infertility are not clear. Sperm-associated antigen 7 (SPAG 7) is expressed in male germ cells and plays a crucial role in several biological functions associated with sperm production and maturation. In this study, we aimed to identify the post-transcriptional regulation of SPAG7 in seminal plasma (SF-Native) and seminal plasma-derived extracellular vesicles (SF-EVs) collected from 87 men undergoing infertility treatment. Among the multiple binding sites for miRNAs within its 3'UTR of SPAG7, we identified the binding of four miRNAs (miR-15b-5p, miR-195-5p, miR-424-5p, and miR-497-5p) to the 3'UTR of SPAG7 by the dual luciferase assays. Analyzing sperm, we found reduced mRNA expression levels of SPAG7 in SF-EVs and SF-Native samples from oligoasthenozoospermic men. By contrast, two miRNAs (miR-424-5p and miR-497-5p) form the SF-Native samples, and four miRNAs (miR-195-5p, miR-424-5p, miR-497-5p, and miR-6838-5p) from the SF-EVs samples showed significantly higher expression levels in oligoasthenozoospermic men. The expression levels of miRNAs and SPAG7 were significantly correlated with basic semen parameters. These findings contribute significantly to our understanding of regulatory pathways in male fertility by showing a direct link between upregulated miRNA, notably miR-424, and downregulated SPAG7 both in seminal plasma and in plasma-derived EVs likely contributing to oligoasthenozoospermia.
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Vesículas Extracelulares , Infertilidad Masculina , MicroARNs , Humanos , Masculino , Semen , Regiones no Traducidas 3' , Antígenos de SuperficieRESUMEN
The tumor microenvironment stimulates the angiogenic activity of endothelial cells (ECs) to facilitate tumor vascularization, growth, and metastasis. The involvement of microRNA-186-5p (miR-186) in regulating the aberrant activity of tumor-associated ECs has so far not been clarified. In the present study, we demonstrated that miR-186 is significantly downregulated in ECs microdissected from human non-small cell lung cancer (NSCLC) tissues compared with matched non-malignant lung tissues. In vitro analyses of primary human dermal microvascular ECs (HDMECs) exposed to different stimuli indicated that this miR-186 downregulation is triggered by hypoxia via activation of hypoxia-inducible factor 1 alpha (HIF1α). Transfection of HDMECs with miR-186 mimic (miR-186m) significantly inhibited their proliferation, migration, tube formation, and spheroid sprouting. In contrast, miR-186 inhibitor (miR-186i) exerted pro-angiogenic effects. In vivo, endothelial miR-186 overexpression inhibited the vascularization of Matrigel plugs and the initial growth of tumors composed of NSCLC cells (NCI-H460) and HDMECs. Mechanistic analyses revealed that the gene encoding for protein kinase C alpha (PKCα) is a bona fide target of miR-186. Activation of this kinase significantly reversed the miR-186m-repressed angiogenic activity of HDMECs. These findings indicate that downregulation of miR-186 in ECs mediates hypoxia-stimulated NSCLC angiogenesis by upregulating PKCα.
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Among the concepts in biology that are widely taken granted is a potentiated cooperative effect of multiple miRNAs on the same target. This strong hypothesis contrasts insufficient experimental evidence. The quantity as well as the quality of required side constraints of cooperative binding remain largely hidden. For miR-21-5p and miR-155-5p, two commonly investigated regulators across diseases, we selected 15 joint target genes. These were chosen to represent various neighboring 3'UTR binding site constellations, partially exceeding the distance rules that have been established for over a decade. We identified different cooperative scenarios with the binding of one miRNA enhancing the binding effects of the other miRNA and vice versa. Using both, reporter assays and whole proteome analyses, we observed these cooperative miRNA effects for genes that bear 3'UTR binding sites at distances greater than the previously defined limits. Astonishingly, the experiments provide even stronger evidence for cooperative miRNA effects than originally postulated. In the light of these findings the definition of targetomes specified for single miRNAs need to be refined by a concept that acknowledges the cooperative effects of miRNAs.
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MicroARNs , MicroARNs/genética , MicroARNs/metabolismo , Regiones no Traducidas 3' , Sitios de UniónRESUMEN
Parkinson's disease (PD) emerges as a complex, multifactorial disease. While there is increasing evidence that dysregulated T cells play a central role in PD pathogenesis, elucidation of the pathomechanical changes in related signaling is still in its beginnings. We employed time-resolved RNA expression upon the activation of peripheral CD4+ T cells to track and functionally relate changes on cellular signaling in representative cases of patients at different stages of PD. While only few miRNAs showed time-course related expression changes in PD, we identified groups of genes with significantly altered expression for each different time window. Towards a further understanding of the functional consequences, we highlighted pathways with decreased or increased activity in PD, including the most prominent altered IL-17 pathway. Flow cytometric analyses showed not only an increased prevalence of Th17 cells but also a specific subtype of IL-17 producing γδ-T cells, indicating a previously unknown role in PD pathogenesis.
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For cancers and other pathologies, early diagnosis remains the most promising path to survival. Profiling of longitudinal cohorts facilitates insights into trajectories of biomarkers. We measured microRNA expression in 240 serum samples from patients with colon, lung, and breast cancer and from cancer-free controls. Each patient provided at least two serum samples, one prior to diagnosis and one following diagnosis. The median time interval between the samples was 11.6 years. Using computational models, we evaluated the circulating profiles of 21 microRNAs. The analysis yielded two sets of biomarkers, static ones that show an absolute difference between certain cancer types and controls and dynamic ones where the level over time provided higher diagnostic information content. In the first group, miR-99a-5p stands out for all three cancer types. In the second group, miR-155-5p allows to predict lung cancers and colon cancers. Classification in samples from cancer and non-cancer patients using gradient boosted trees reached an average accuracy of 79.9%. The results suggest that individual change over time or an absolute value at one time point may predict a disease with high specificity and sensitivity.
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MicroARN Circulante , MicroARNs , Neoplasias , Humanos , Biomarcadores , Biomarcadores de Tumor/genética , Detección Precoz del Cáncer , Perfilación de la Expresión Génica , MicroARNs/genética , Neoplasias/diagnóstico , Neoplasias/genéticaRESUMEN
The unique physicochemical properties make inorganic nanoparticles (INPs) an exciting tool in diagnosis and disease management. However, as INPs are relatively difficult to fully degrade and excrete, their unintended accumulation in the tissue might result in adverse health effects. Herein, we provide a methylome-transcriptome framework for chronic effects of INPs, commonly used in biomedical applications, in human kidney TH-1 cells. Renal clearance is one of the most important routes of nanoparticle excretion; therefore, a detailed evaluation of nanoparticle-mediated nephrotoxicity is an important task. Integrated analysis of methylome and transcriptome changes induced by INPs (PEG-AuNPs, Fe3O4NPs, SiO2NPs, and TiO2NPs) revealed significantly deregulated genes with functional classification in immune response, DNA damage, and cancer-related pathways. Although most deregulated genes were unique to individual INPs, a relatively high proportion of them encoded the transcription factors. Interestingly, FOS hypermethylation inversely correlating with gene expression was associated with all INPs exposures. Our study emphasizes the need for a more comprehensive investigation of INPs' biological safety, especially after chronic exposure.