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1.
Aging (Albany NY) ; 15(1): 53-69, 2023 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-36629518

RESUMO

BACKGROUND: microRNAs (miRNAs) are involved in the progression of Lung adenocarcinoma (LUAD), however, the functions of miR-6742-5p in LUAD remains unknown, thereby this study was carried on. METHODS: The mRNA and miRNA expression data from the LUAD and normal control were obtained from Gene Expression Omnibus (GEO) database, TargetScan and mirDIP were applied to predict the relationship between miR-6742-5p and FGF8.Q-PCR, western blot, dual-luciferase, wound Healing and transwell assays were performed to test the functions of miR-6742-5p in LUAD. RESULTS: Bioinformatics analysis and dual-luciferase identified FGF8 is the target-gene of miR-6742-5p, which is declined in LUAD of human tissues and cell lines, and miR-6742-5P OE suppressed the progression of LUAD in nude mice. MiR-6742-5p OE and KD suppressed or increased the abilities of LUAD' metastasis tested by wound healing and transwell assays H522 and PC-9 cells, these effects about miR-6742-5p OE were reversed by FGF8; miR-6742-5p OE, KD inhibited and increased the expression of FGF8 as its downstream p-ERK1/2, MMP-2/-9, these results were corrected by ERK1/2 inhibitor: Ro 67-7476; the miR-6742-5p KD increased the migrated and invaded cells and suppressed by MMPs inhibitor: S3304. These results identified the negative correlation of miR-6742-5p with FGF8-ERK1/2 signal pathway in LUAD progression. CONCLUSIONS: We conclude that miR-6742-5p might be a regulator of LUAD progression by targeting FGF8/ERK1/2/MMPs signaling pathway, which provides a novel therapeutic target for LUAD.


Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , MicroRNAs , Animais , Camundongos , Humanos , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Camundongos Nus , Linhagem Celular Tumoral , Adenocarcinoma de Pulmão/metabolismo , Neoplasias Pulmonares/patologia , MicroRNAs/genética , MicroRNAs/metabolismo , Transdução de Sinais , Movimento Celular/genética , Regulação Neoplásica da Expressão Gênica , Proliferação de Células/genética , Fator 8 de Crescimento de Fibroblasto/genética , Fator 8 de Crescimento de Fibroblasto/metabolismo
2.
Int J Mol Sci ; 24(2)2023 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-36674732

RESUMO

MiRNAs are critical regulators of numerous physiological and pathological processes. Ascosphaera apis exclusively infects bee larvae and causes chalkbrood disease. However, the function and mechanism of miRNAs in the bee larval response to A. apis infection is poorly understood. Here, ame-miR-34, a previously predicted miRNA involved in the response of Apis mellifera larvae to A. apis invasion, was subjected to molecular validation, and overexpression and knockdown were then conducted to explore the regulatory functions of ame-miR-34 in larval body weight and immune response. Stem-loop RT-PCR and Sanger sequencing confirmed the authenticity of ame-miR-34 in the larval gut of A. mellifera. RT-qPCR results demonstrated that compared with that in the uninfected larval guts, the expression level of ame-miR-34 was significantly downregulated (p < 0.001) in the guts of A. apis-infected 4-, 5-, and 6-day-old larvae, indicative of the remarkable suppression of host ame-miR-34 due to A. apis infection. In comparison with the corresponding negative control (NC) groups, the expression level of ame-miR-34 in the larval guts in the mimic-miR-34 group was significantly upregulated (p < 0.001), while that in the inhibitor-miR-34 group was significantly downregulated (p < 0.01). Similarly, effective overexpression and knockdown of ame-miR-34 were achieved. In addition, the body weights of 5- and 6-day-old larvae were significantly increased compared with those in the mimic-NC group; the weights of 5-day-old larvae in the inhibitor-miR-34 group were significantly decreased in comparison with those in the inhibitor-NC group, while the weights of 4- and 6-day-old larvae in the inhibitor-miR-34 group were significantly increased, indicating the involvement of ame-miR-34 in modulating larval body weight. Furthermore, the expression levels of both hsp and abct in the guts of A. apis-infected 4-, 5-, and 6-day-old larvae were significantly upregulated after ame-miR-34 overexpression. In contrast, after ame-miR-34 knockdown, the expression levels of the aforementioned two key genes in the A. apis-infected 4-, 5-, and 6-day-old larval guts were significantly downregulated. Together, the results demonstrated that effective overexpression and knockdown of ame-miR-34 in both noninfected and A. apis-infected A. mellifera larval guts could be achieved by the feeding method, and ame-miR-34 exerted a regulatory function in the host immune response to A. apis invasion through positive regulation of the expression of hsp and abct. Our findings not only provide a valuable reference for the functional investigation of bee larval miRNAs but also reveal the regulatory role of ame-miR-34 in A. mellifera larval weight and immune response. Additionally, the results of this study may provide a promising molecular target for the treatment of chalkbrood disease.


Assuntos
Arthrodermataceae , Abelhas , MicroRNAs , Animais , Abelhas/genética , Abelhas/imunologia , Abelhas/microbiologia , Peso Corporal , Imunidade , Larva/imunologia , MicroRNAs/genética , MicroRNAs/metabolismo , Arthrodermataceae/fisiologia
3.
Int J Mol Sci ; 24(2)2023 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-36675068

RESUMO

Stress is a key risk factor in the onset of neuropsychiatric disorders. The study of the mechanisms underlying stress response is important to understand the etiopathogenetic mechanisms and identify new putative therapeutic targets. In this context, microRNAs (miRNAs) have emerged as key regulators of the complex patterns of gene/protein expression changes in the brain, where they have a crucial role in the regulation of neuroplasticity, neurogenesis, and neuronal differentiation. Among them, miR-135a-5p has been associated with stress response, synaptic plasticity, and the antidepressant effect in different brain areas. Here, we used acute unavoidable foot-shock stress (FS) and chronic mild stress (CMS) on male rats to study whether miR-135a-5p was involved in stress-induced changes in the prefrontal cortex (PFC). Both acute and chronic stress decreased miR-135a-5p levels in the PFC, although after CMS the reduction was induced only in animals vulnerable to CMS, according to a sucrose preference test. MiR-135a-5p downregulation in the primary neurons reduced dendritic spine density, while its overexpression exerted the opposite effect. Two bioinformatically predicted target genes, Kif5c and Cplx1/2, were increased in FS rats 24 h after stress. Altogether, we found that miR-135a-5p might play a role in stress response in PFC involving synaptic mechanisms.


Assuntos
MicroRNAs , Córtex Pré-Frontal , Estresse Fisiológico , Estresse Psicológico , Animais , Masculino , Ratos , Regulação para Baixo/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/patologia , Córtex Pré-Frontal/fisiologia , Doença Aguda/psicologia , Doença Crônica/psicologia , Estresse Fisiológico/genética , Estresse Psicológico/genética , Estresse Psicológico/psicologia , Sinapses/genética , Sinapses/metabolismo , Sinapses/patologia , Espinhas Dendríticas/genética , Espinhas Dendríticas/metabolismo , Espinhas Dendríticas/patologia
4.
Int J Mol Sci ; 24(2)2023 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-36675088

RESUMO

The aim of this study was to assess the interrelation between vascular ultrasonography (US) findings, histopathological data, and the expression of selected dysregulated microRNAs (miRNAs) in giant cell arteritis (GCA). The study included data on the clinical parameters, US measurements, and temporal artery biopsies (TABs) of 46 treatment-naïve patients diagnosed with GCA and 22 age-matched non-GCA patient controls. We performed a comprehensive comparative and correlation analysis along with generation of receiver operating characteristic (ROC) curves to ascertain the diagnostic performance of US examination parameters and selected miRNAs for GCA diagnosis. We showed significant differences in the US-measured intima-media thickness of the temporal arteries, the presence of a halo sign, and the presence of luminal stenosis between GCA-positive/TAB-positive, GCA-positive/TAB-negative, and non-GCA patients. Correlation analysis revealed significant associations between several histopathological parameters, US-measured intima-media thickness, and the halo sign. We found that the significant overexpression of miR-146b-5p, miR-155-5p, miR-511-5p, and miR-21-5p, and the under-expression of the miR-143/145 cluster, miR-30a-5p, and miR-125a-5p, coincides and is associated with the presence of a halo sign in patients with GCA. Notably, we determined a high diagnostic performance of miR-146b-5p, miR-21-3p, and miR-21-5p expression profiles in discriminating GCA patients from non-GCA controls, suggesting their potential utilization as putative biomarkers of GCA. Taken together, our study provides an insight into the US-based diagnostic evaluation of GCA by revealing the complex interrelation of clearly defined image findings with underlying vascular immunopathology and altered arterial tissue-specific miRNA profiles.


Assuntos
Arterite de Células Gigantes , MicroRNAs , Artérias Temporais , Humanos , Biópsia , Espessura Intima-Media Carotídea , Arterite de Células Gigantes/diagnóstico por imagem , Arterite de Células Gigantes/genética , Arterite de Células Gigantes/patologia , MicroRNAs/genética , MicroRNAs/metabolismo , Sensibilidade e Especificidade , Artérias Temporais/diagnóstico por imagem , Artérias Temporais/metabolismo , Artérias Temporais/patologia , Ultrassonografia
5.
Nat Commun ; 14(1): 351, 2023 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-36681689

RESUMO

Pancreatic ß-cell compensation is a major mechanism in delaying T2DM progression. Here we report the abnormal high expression of circGlis3 in islets of male mice with obesity and serum of people with obesity. Increasing circGlis3 is regulated by Quaking (QKI)-mediated splicing circularization. circGlis3 overexpression enhances insulin secretion and inhibits obesity-induced apoptosis in vitro and in vivo. Mechanistically, circGlis3 promotes insulin secretion by up-regulating NeuroD1 and Creb1 via sponging miR-124-3p and decreases apoptosis via interacting with the pro-apoptotic factor SCOTIN. The RNA binding protein FUS recruits circGlis3 and collectively assemble abnormal stable cytoplasmic stress granules (SG) in response to cellular stress. These findings highlight a physiological role for circRNAs in ß-cell compensation and indicate that modulation of circGlis3 expression may represent a potential strategy to prevent ß-cell dysfunction and apoptosis after obesity.


Assuntos
Células Secretoras de Insulina , MicroRNAs , Camundongos , Masculino , Animais , RNA Circular/genética , RNA Circular/metabolismo , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Apoptose/genética , Obesidade/genética , Obesidade/metabolismo
6.
Arch Biochem Biophys ; 734: 109501, 2023 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-36592647

RESUMO

A well-tuned inflammatory response is crucial for an effective immune process. Nuclear factor-kappa B (NF-κB) is a key mediator of inflammatory and innate immunity responses, and its dysregulation is closely associated with immune-related diseases. MicroRNAs (miRNAs) are important inflammation modulators. However, miRNA-regulated mechanisms that implicate NF-κB activity are not fully understood. This study aimed to identify a potential miRNA that could modulate the dysregulated NF-κB signaling during inflammation. We identified miR-582-5p that was significantly downregulated in inflamed murine adipose tissues and RAW264.7 cells. S-phase kinase-associated protein 1 (SKP1), a core component of an E3 ubiquitin ligase that regulates the NF-κB pathway, was proposed as a biological target of miR-582-5p by using TargetScan. The binding of miR-582-5p to a 3'-untranslated region site on Skp1 was confirmed using a dual-luciferase reporter assay; in addition, transfection with a miR-582-5p mimic suppressed SKP1 expression in RAW264.7 cells. Importantly, exogenous miR-582-5p attenuated the production of pro-inflammatory cytokines such as tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6 through suppressing the degradation of the NF-κB inhibitor alpha, followed by the nuclear translocation of NF-κB. Therefore, exogenously applied miR-582-5p can attenuate the NF-κB signaling pathway via targeting Skp1; this provides a prospective therapeutic strategy for treating inflammatory and immune diseases.


Assuntos
MicroRNAs , NF-kappa B , Animais , Camundongos , Inflamação/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Proteínas Quinases Associadas a Fase S/genética , Proteínas Quinases Associadas a Fase S/metabolismo , Transdução de Sinais
7.
Cell Mol Biol Lett ; 28(1): 5, 2023 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-36658478

RESUMO

BACKGROUND: Secondary spinal cord injury (SCI) often causes the aggravation of inflammatory reaction and nerve injury, which affects the recovery of motor function. Bone-marrow-derived macrophages (BMDMs) were recruited to the injured area after SCI, and the M1 polarization is the key process for inducing inflammatory response and neuronal apoptosis. We previously showed that photobiomodulation (PBM) can inhibit the polarization of M1 phenotype of BMDMs and reduce inflammation, but the underlying mechanisms are unclear. The purpose of this study is to explore the potential target and mechanism of PBM in treating SCI. METHODS: Transcriptome sequencing and bioinformatics analysis showed that long noncoding RNA taurine upregulated gene 1 (lncRNA TUG1) was a potential target of PBM. The expression and specific mechanism of lncRNA TUG1 were detected by qPCR, immunofluorescence, flow cytometry, western blotting, fluorescence in situ hybridization, and luciferase assay. The Basso mouse scale (BMS) and gait analysis were used to evaluate the recovery of motor function in mice. RESULTS: Results showed that lncRNA TUG1 may be a potential target of PBM, regulating the polarization of BMDMs, inflammatory response, and the axial growth of DRG. Mechanistically, TUG1 competed with TLR3 for binding to miR-1192 and attenuated the inhibitory effect of miR-1192 on TLR3. This effect protected TLR3 from degradation, enabling the high expression of TLR3, which promoted the activation of downstream NF-κB signal and the release of inflammatory cytokines. In vivo, PBM treatment could reduce the expression of TUG1, TLR3, and inflammatory cytokines and promoted nerve survival and motor function recovery in SCI mice. CONCLUSIONS: Our study clarified that the lncRNA TUG1/miR-1192/TLR3 axis is an important pathway for PBM to inhibit M1 macrophage polarization and inflammation, which provides theoretical support for its clinical application in patients with SCI.


Assuntos
MicroRNAs , RNA Longo não Codificante , Traumatismos da Medula Espinal , Receptor 3 Toll-Like , Animais , Camundongos , Citocinas/genética , Hibridização in Situ Fluorescente , Inflamação/genética , Inflamação/metabolismo , Macrófagos/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Traumatismos da Medula Espinal/genética , Receptor 3 Toll-Like/genética
8.
Mediators Inflamm ; 2023: 9380398, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36660176

RESUMO

Myocardial ischemia-reperfusion (IR) injury is the restoration of blood flow post ischemia, which threatens the human life. Long noncoding RNA distal-less homeobox 6 antisense 1 (DLX6-AS1) has been found to take part in the IR-induced cerebral injury. Here, we determined the functional role of DLX6-AS1 in IR-induced myocardial injury. We ligated the left anterior descending coronary artery of rats to induce IR injury. IR injury rats exhibited severe tissue damage and increase of infraction size. The levels of lactate dehydrogenase (LDH), creatine kinase (CK), proinflammatory factors including MCP-1, IL-6, and IL-1ß, and cell apoptosis were also enhanced in IR rats, indicating that IR induced significant myocardial injury in rats. DLX6-AS1 expression was elevated in the myocardial tissues of IR injury rats, while DLX6-AS1 deficiency alleviated IR-induced myocardial injury in rats by reducing inflammatory response and cell apoptosis. Moreover, rat embryonic cardiomyocyte cell line H9c2 was subjected to hypoxia reoxygenation (HR). DLX6-AS1 was upregulated in the HR-treated H9c2 cells, and DLX6-AS1 enhanced the expression of F-box and WD40 repeat domain-containing 7 (FBXW7) by sponging miR-204-5p. Inhibition of DLX6-AS1 inhibited inflammatory response and cell apoptosis in H9c2 cells via miR-204-5p/FBXW7 axis. In conclusion, this work demonstrates that DLX6-AS1 accelerates myocardial IR injury through regulating miR-204-5p/FBXW7 axis. Thus, this work provides a novel ceRNA DLX6-AS1/miR-204-5p/FBXW7 axis in myocardial IR injury, and DLX6-AS1 may be a potential target for the treatment of myocardial IR injury.


Assuntos
MicroRNAs , Traumatismo por Reperfusão Miocárdica , RNA Longo não Codificante , Animais , Ratos , Apoptose/genética , Linhagem Celular , Proteína 7 com Repetições F-Box-WD/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , MicroRNAs/metabolismo , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo
9.
Methods Mol Biol ; 2619: 257-271, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36662476

RESUMO

Circulating microRNAs (miRNAs) possess significant roles in normal homeostasis and disease conditions, including cardiovascular diseases, fibrosis, inflammatory response, and cancer. Secreted miRNAs, via the membrane vesicles, actively communicate with extracellular matrix (ECM) components to affect cell-cell and cell-matrix interactions, thereby affecting matrix remodeling and metabolic pathways in the recipient cells. Matrix macromolecules regulate the expression of miRNAs, and in turn miRNAs have been identified as emerging mediators of matrix constituents, serving as appealing biomarkers for many pathophysiological processes. Therefore, the expression profile of certain miRNAs highlights the importance of their targeting in several aspects of human pathologies. In this chapter, we report molecular biology protocols to determine the effects of selected miRNAs on the expression and activity of matrix biomolecules.


Assuntos
MicroRNA Circulante , MicroRNAs , Neoplasias , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Proteoglicanas/metabolismo , Matriz Extracelular/metabolismo , Neoplasias/patologia
10.
Methods Mol Biol ; 2619: 273-292, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36662477

RESUMO

MicroRNAs are small noncoding RNAs that regulate gene expression at the posttranscriptional level. Proteoglycans are glycoproteins characterized by covalent attachment of a glycosaminoglycan chain, which have been identified as regulatory targets of microRNAs in a physiological and pathophysiological context. We present a strategy and detailed methods for the functional analysis of microRNA regulation of proteoglycans using human cancer cells as an application example. The experimental setup includes in silico microRNA target prediction, transfection of cancer cells with microRNA precursors, validation of target regulation by qPCR, flow cytometry and luciferase reporter assays, and an example for functional analysis and phenotype confirmation by complementation analysis.


Assuntos
MicroRNAs , Neoplasias , Humanos , MicroRNAs/metabolismo , Proteoglicanas/genética , Proteoglicanas/metabolismo , Transfecção , Luciferases/metabolismo
11.
PLoS One ; 18(1): e0280425, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36662897

RESUMO

microRNAs (miRNAs) are extracellularly released by cells for intercellular communication, while intracellularly, they inhibit the expression of specific genes. An increasing number of studies suggest that extracellular miRNAs have great potential as both therapeutic targets and disease-specific biomarkers in a variety of diseases, including pain disorders. However, little is known about miRNA release from dorsal root ganglion (DRG) neurons in neuropathic pain caused by peripheral nerve injury. In this study, we investigated the changes in the extracellular release of miRNAs from DRG neurons in a rat model of neuropathic pain induced by chronic constriction injury of the sciatic nerve. We found increased release of six miRNAs (let-7d, miR-21, miR-142-3p, miR-146b, miR-203-3p and miR-221) from primary cultured DRG neurons prepared from rats 7 days after nerve injury. Among these, miR-221 was also increased in serum from days 7 to 28 after nerve injury. In contrast, serum miR-221 levels and its release from DRG neurons were unchanged in an inflammatory pain model produced by intraplantar injection of complete Freund's adjuvant. These results suggest that the increased release of specific miRNAs by DRG neurons may be involved in the pathophysiology of neuropathic pain through extracellular as well as intracellular mechanisms. Furthermore, serum miR-221 may be useful as a biomarker of neuropathic pain caused by peripheral nerve injury.


Assuntos
MicroRNAs , Neuralgia , Traumatismos dos Nervos Periféricos , Ratos , Animais , Gânglios Espinais/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Neuralgia/etiologia , Nervo Isquiático/lesões
12.
Biomolecules ; 13(1)2023 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-36671508

RESUMO

Osteoarthritis (OA), the most prevalent joint disease and the leading cause of disability, remains an incurable disease largely because the etiology and pathogenesis underlying this degenerative process are poorly understood. Low-grade inflammation within joints is a well-established factor that disturbs joint homeostasis and leads to an imbalance between anabolic and catabolic processes in articular cartilage; however, the complexity of the network between inflammatory factors that often involves positive and negative feedback loops makes current anti-cytokine therapy ineffective. MicroRNAs (miRNAs) have emerged as key regulators to control inflammation, and aberrant miRNAs expression has recently been linked to OA pathophysiology. In the present study, we characterized transcriptomic profiles of miRNAs in primary murine articular chondrocytes in response to a proinflammatory cytokine, IL-1ß, and identified miR-146a-5p as the most responsive miRNA to IL-1ß. miR-146a-5p was also found to be upregulated in human OA cartilage. We further demonstrated that knockdown of miR-146a-5p antagonized IL-1ß-mediated inflammatory responses and IL-1ß-induced catabolism in vitro, and silencing of miR-146a in chondrocytes ameliorated articular cartilage destruction and reduced OA-evoked pain in an injury-induced murine OA model. Moreover, parallel RNA sequencing revealed that differentially expressed genes in response to IL-1ß were enriched in pathways related to inflammatory processes, cartilage matrix homeostasis, and cell metabolism. Bioinformatic analyses of putative miR-146a-5p gene targets and following prediction of protein-protein interactions suggest a functional role of miR-146a-5p in mediating inflammatory processes and regulation of cartilage homeostasis. Our genetic and transcriptomic data define a crucial role of miR-146a-5p in OA pathogenesis and implicate modulation of miR-146a-5p in articular chondrocytes as a potential therapeutic strategy to alleviate OA.


Assuntos
Cartilagem Articular , MicroRNAs , Osteoartrite , Humanos , Camundongos , Animais , Osteoartrite/patologia , MicroRNAs/genética , MicroRNAs/metabolismo , Condrócitos , Inflamação/metabolismo , Cartilagem Articular/patologia , Apoptose
13.
Biosensors (Basel) ; 13(1)2023 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-36671913

RESUMO

miRNA is considered a novel biomarker for cancer diagnosis and due to its low level in vivo, the development of new detection methods for it has become a research hotspot in recent years. Here, we firstly found that miR-625-5p was significantly upregulated in colorectal cancer tissues by means of differential expression analysis of the dbDEMC database and clinical validation. Subsequently, it was found that miR-625-5p promoted cell proliferation and migration but inhibited apoptosis through phenotypic experiments; thus, we initially identified miR-625-5p as a potential biomarker for colorectal cancer. Moreover, in order to monitor slight changes in the miR-625-5p level, we developed a novel detection method for it based on strand displacement amplification (SDA). In this system, a hairpin was designed to recognize and pair with miR-625-5p, which was used as a primer to initiate SDA, and a large number of complementary DNAs were generated via cyclic amplification, followed by the addition of SYBR Gold to achieve quantitative analysis of miR-625-5p. Moreover, this method showed a good response to miR-625-5p with a detection limit of 8.6 pM and a dynamic range of 0.01 to 200 nM, and the specificity of it was verified using a set of other miRNAs as an interference. Finally, we set up different concentrations of biologic samples for detection to verify the practicability of the method. The results of this study indicate that this detection method has great potential in clinical diagnosis.


Assuntos
Neoplasias Colorretais , MicroRNAs , Humanos , Biomarcadores Tumorais , MicroRNAs/metabolismo , DNA Complementar , Apoptose , Neoplasias Colorretais/diagnóstico
14.
Cells ; 12(2)2023 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-36672162

RESUMO

Colorectal cancer has proven to be difficult to treat as it is the second leading cause of cancer death for both men and women worldwide. Recent work has shown the importance of microRNA (miRNA) in the progression and metastasis of colorectal cancer. Here, we develop a metric based on miRNA-gene target interactions, previously validated to be associated with colorectal cancer. We use this metric with a regularized Cox model to produce a small set of top-performing genes related to colon cancer. We show that using the miRNA metric and a Cox model led to a meaningful improvement in colon cancer survival prediction and correct patient risk stratification. We show that our approach outperforms existing methods and that the top genes identified by our process are implicated in NOTCH3 signaling and general metabolism pathways, which are essential to colon cancer progression.


Assuntos
Neoplasias do Colo , MicroRNAs , Masculino , Humanos , Feminino , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias do Colo/patologia , Transdução de Sinais/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Análise de Sequência de RNA
15.
Cells ; 12(2)2023 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-36672174

RESUMO

Rapid responses to changes within subcellular compartments of highly polarized cells, such as neuron axons, depend on local translation and post-transcriptional regulation. The mechanism by which microRNAs (miRNAs) regulate this process is not fully understood. Here, using live cell imaging and RNA sequencing analysis, we demonstrated how miRNAs can differentially control hundreds of transcripts at the subcellular level. We demonstrated that the seed match length of the miRNA target-sequence regulates both mRNA stability and protein translation rates. While longer seed matches have an increased inhibitory effect, transcriptome analysis did not reveal differences in seed match length between axonal and somata mRNAs of motor neurons. However, mRNA variants with longer 3'UTR are enriched in axons and contain multiple repeats of specific miRNA target sequences. Finally, we demonstrated that the long 3'UTR mRNA variant of the motor protein Kif5b is enriched explicitly in motor neuron axons and contains multiple sequence repeats for binding miR-129-5p. This subsequently results in the differential post-transcriptional regulation of kif5b and its synthesis in axons. Thus, we suggest that the number of miRNA binding sites at the 3'UTR of the mRNA, rather than the miRNA seed match length, regulates the axonal transcriptome.


Assuntos
MicroRNAs , Regiões 3' não Traduzidas/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Axônios/metabolismo , Sítios de Ligação , Proteínas de Transporte/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
16.
Cells ; 12(2)2023 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-36672241

RESUMO

MicroRNAs (miRNAs) are versatile, post-transcriptional regulators of gene expression. Canonical miRNAs are generated through the two-step DROSHA- and DICER-mediated processing of primary miRNA (pri-miRNA) transcripts with optimal or suboptimal features for DROSHA and DICER cleavage and loading into Argonaute (AGO) proteins, whereas multiple hairpin-structured RNAs are encoded in the genome and could be a source of non-canonical miRNAs. Recent advances in miRNA biogenesis research have revealed details of the structural basis of miRNA processing and cluster assistance mechanisms that facilitate the processing of suboptimal hairpins encoded together with optimal hairpins in polycistronic pri-miRNAs. In addition, a deeper investigation of miRNA-target interaction has provided insights into the complexity of target recognition with distinct outcomes, including target-mediated miRNA degradation (TDMD) and cooperation in target regulation by multiple miRNAs. Therefore, the coordinated or network regulation of both miRNA biogenesis and miRNA-target interaction is prevalent in miRNA biology. Alongside recent advances in the mechanistic investigation of miRNA functions, this review summarizes recent findings regarding the ordered regulation of miRNA biogenesis and miRNA-target interaction.


Assuntos
MicroRNAs , MicroRNAs/metabolismo , Processamento Pós-Transcricional do RNA
17.
Cells ; 12(2)2023 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-36672257

RESUMO

The existence of circular RNA (circRNA) research in mainstream science can be attributed to the contemporary synergism of big data and keen attention to detail by several research groups worldwide. Since the re-emergence of these non-canonical RNA transcripts, seminal advances have been made in understanding their biogenesis, interactome, and functions in diverse fields and a myriad of human diseases. However, most research outputs to date have focused on the ability of highly stable circRNAs to interact with, and impact signalling through, microRNAs. This is likely to be the result of seminal papers in the field ascribing a few remarkable circRNAs as "miRNA sponges". However, the stoichiometric ratio between the (often-lowly-expressed) circRNA and their (commonly-more-abundant) target is rarely in favour of a biologically relevant and functional consequence of these interactions. It is time for yet another revolution in circRNA research to uncover functions beyond their documented ability to bind miRNAs. This Special Issue aims to highlight non-canonical functions for this non-canonical family of RNA molecules.


Assuntos
MicroRNAs , RNA Circular , Humanos , RNA Circular/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Transdução de Sinais
18.
Clinics (Sao Paulo) ; 78: 100155, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36681070

RESUMO

FOXO3a dysregulation is frequently implicated in tumorigenesis, and its inhibition can occur by several molecular mechanisms. Among these, post-transcriptional suppression by miRNAs has been associated with various cancers initiation. Here, we assessed the expression profiles of the most relevant miRNAs for breast tumorigenesis, using Luminal A (LA) and Triple-Negative (TN) breast cancer from Brazilian patients, by the quantitative real time-PCR method. Their potential prognostic role for the patients was also evaluated. We identified the miRNAs miR-96-5p and miR-182-5p, de-scribed as negative regulators of FOXO3A, with differential expression both in LA and TN tumors when compared to normal tissue. The miR-96-5p and miR-182-5p miRNAs were upregulated in LA (7.82 times, p < 0.005; 6.12 times, p < 0.005, respectively) and TN breast cancer samples (9.42 times, p < 0.0001; 8.51 times, p < 0.0001) compared to normal tissues. The samples with higher miR-96-5p and miR-182-5p expression (FR ≥ 4) were submitted for FOXO3a immunostaining. Reduced protein detection was observed in all of the tumors compared to normal tissues. The most prominent miRNA expression and FOXO3a protein suppression were observed in TN samples (p < 0.001), indicating the relevant role of these molecules in this tumor biology and clinical behavior. Our results corroborate the literature regarding to the relevance of FOXO3a in the breast cancer, and they open new perspectives for alternative target therapy options for Brazilian patients expressing both FOXO3a and its regulatory miRNAs.


Assuntos
Neoplasias da Mama , MicroRNAs , Neoplasias de Mama Triplo Negativas , Humanos , Feminino , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Brasil , Biomarcadores Tumorais/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Prognóstico , Carcinogênese , Regulação Neoplásica da Expressão Gênica , Perfilação da Expressão Gênica
19.
Environ Pollut ; 319: 120981, 2023 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-36587786

RESUMO

Numerous studies have revealed that ambient long-term exposure to fine particulate matter (PM2.5) is significantly related to the development of lung cancer, but the molecular mechanisms of PM2.5 exposure-induced lung cancer remains unknown. As an important epigenetic regulator, microRNAs (miRNAs) play vital roles in responding to environment exposure and various diseases including lung cancer development. Here we constructed a PM2.5-induced malignant transformed cell model and found that miR-200 family, especially miR-200a-3p, was involved in the process of PM2.5 induced lung cancer. Further investigation of the function of miR-200 family (miR-200a-3p as a representative revealed that miR-200a-3p promoted cell migration by directly suppressing TNS3 expression. These results suggested that ambient PM2.5 exposure may increase the expression of miR-200 family and then promote the proliferation and migration of lung cancer cells. Our study provided novel model and insights into the molecular mechanism of ambient PM2.5 exposure-induced lung cancer.


Assuntos
Neoplasias Pulmonares , MicroRNAs , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias Pulmonares/metabolismo , Material Particulado/toxicidade , Material Particulado/metabolismo , Células Epiteliais/patologia , Transformação Celular Neoplásica/metabolismo
20.
J Ethnopharmacol ; 305: 116032, 2023 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-36587882

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Kabasura Kudineer (KK), the traditional Indian medicine of Siddha, effectively manages common respiratory symptoms such as flu, cold, and fever. However, there is no evidence of the immunomodulatory capacity of KK in the cultured Jurkat T-lymphocytes under the LPS insult studied. AIM OF THE STUDY: Assess the effect of the traditional Indian medicine of Siddha, Kabasura Kudineer (KK) on immunomodulation by suppressing oxidative damage in cultured Jurkat T cells in vitro. The miRNA activity on anti-inflammatory gene receptors and cellular nitric oxide levels also was studied. MATERIALS AND METHODS: Jurkat T cells were exposed to LPS treatment in the presence or absence of KK. Cell viability and nitric oxide (NO) were measured with MTT and Griess assay. Cellular antioxidant systems (glutathione and SOD) were determined using glutathione and SOD assay. Lipid peroxidation was measured using an MDA assay. MiRNA-15a-5p expression was performed using microRNA qPCR Assays. Both inflammatory and anti-inflammatory genes (IL-6, IL-1, IL-10, IL-13) were performed using a qPCR and ELISA assay. RESULTS: The data showed that reduced cell proliferation and exaggerated NO production was observed in LPS treated condition compared to the control condition. Further, LPS treatment increased lipid peroxidation and reduced antioxidant enzyme activities (SOD and glutathione) in cultured Jurkat T cells. However, treatment with KK or N-acetyl cysteine (NAC; antioxidant) treatment mitigates the above effect. Mechanistically, LPS-induced oxidative stress upregulated miR- 15-5p expression and suppressed IL-10 Receptor alpha (IL-10Rα) by binding to its 3'-UTR region. The deregulated expression of IL-10Rα expression leads to increased IL-6 and IL-1ß expression in LPS-induced Jurkat T cells; however, treatment with KK or NAC reversed the above effects. CONCLUSION: Collectively, our study revealed the previously undefined mechanistic role of Kabasura Kudineer (KK) that alleviates the LPS-induced oxidative damage associated with inflammation by inhibiting the miRNA-15-5p/IL-10Rα axis.


Assuntos
MicroRNAs , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos/farmacologia , Óxido Nítrico/metabolismo , Inflamação , Estresse Oxidativo , Anti-Inflamatórios/farmacologia , Glutationa/metabolismo , Superóxido Dismutase/metabolismo
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