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AIM: To examine the regulatory role of microRNA-204 (miR-204) on silent information regulator 1 (SIRT1) and vascular endothelial growth factor (VEGF) under high-glucose-induced metabolic memory in human retinal pigment epithelial (hRPE) cells. METHODS: Cells were cultured with either normal (5 mmol/L) or high D-glucose (25 mmol/L) concentrations for 8d to establish control and high-glucose groups, respectively. To induce metabolic memory, cells were cultured with 25 mmol/L D-glucose for 4d followed by culture with 5 mmol/L D-glucose for 4d. In addition, exposed in 25 mmol/L D-glucose for 4d and then transfected with 100 nmol/L miR-204 control, miR-204 inhibitor or miR-204 mimic in 5 mmol/L D-glucose for 4d. Quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect miR-204 mRNA levels. SIRT1 and VEGF protein levels were assessed by immunohistochemical and Western blot. Flow cytometry was used to investigate apoptosis rate. RESULTS: It was found that high glucose promoted miR-204 and VEGF expression, and inhibited SIRT1 activity, even after the return to normal glucose culture conditions. Upregulation of miR-204 promoted apoptosis inhibiting SIRT1 and increasing VEGF expression. However, downregulation of miR-204 produced the opposite effects. CONCLUSION: The study identifies that miR-204 is the upstream target of SIRT1 and VEGF, and that miR-204 can protect hRPE cells from the damage caused by metabolic memory through increasing SIRT1 and inhibiting VEGF expression.
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OBJECTIVE: The exosomal cargo mainly comprises proteins, lipids, and microRNAs (miRNAs). Among these, miRNAs undertake multiple biological effects of exosomes (Exos). Some stem cell-derived exosomal miRNAs have shown the potential to treat diabetic nephropathy (DN). However, there is little research into the therapeutic effects of adipose-derived stem cell (ADSC)-derived exosomal miRNAs on DN. We aimed to explore the potential of miR-204-modified ADSC-derived Exos to mitigate DN. METHODS: Exos were extracted and identified from ADSCs. Histopathological injury, oxidative stress (OS), mitochondrial function, cell viability, and apoptosis were assessed to explore the effects of ADSC-derived Exos on DN. For mechanism exploration, quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to measure miR-204, methyltransferase (METTL3, METTL14, and METTL7A), and CIDEC. Also, CIDEC m6A methylation and miR-204-METTL7A, and METTL7A-CIDEC interactions were determined. RESULTS: Initially, OS-induced mitochondrial dysfunction was observed in DN rats. ADSC-derived Exos inhibited histopathological injury, cell apoptosis, OS, and mitochondrial dysfunction in DN rats. The similar therapeutic effects of ADSC-derived Exos were detected in the in vitro model. Intriguingly, miR-204 was released by ADSC-derived Exos and its upregulation enhanced the anti-DN effects of Exos. Mechanically, miR-204 reduced METTL7A expression to CIDEC m6A methylation, thus suppressing OS and mitochondrial dysfunction. CONCLUSIONS: ADSC-derived exosomal miR-204 rescued OS-induced mitochondrial dysfunction by inhibiting METTL7A-mediated CIDEC m6A methylation. This study first revealed the significant role of ADSC-derived exosomal miR-204 in DN, paving the way for the development of novel therapeutic strategies to improve the clinical outcomes of DN patients.
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Diabetes Mellitus , Nefropatías Diabéticas , Exosomas , MicroARNs , Enfermedades Mitocondriales , Nanoestructuras , Humanos , Ratas , Animales , Exosomas/metabolismo , Nefropatías Diabéticas/genética , Nefropatías Diabéticas/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Metiltransferasas/genética , Metiltransferasas/metabolismo , Metilación , Enfermedades Mitocondriales/metabolismo , Diabetes Mellitus/metabolismoRESUMEN
Tumors have high requirements in terms of nutrients and oxygen. Angiogenesis is the classical mechanism for vessel formation. Tumoral vascularization has the function of nourishing the cancer cells to support tumor growth. Vasculogenic mimicry, a novel intratumoral microcirculation system, alludes to the ability of cancer cells to organize in three-dimensional (3D) channel-like architectures. It also supplies the tumors with nutrients and oxygen. Both mechanisms operate in a coordinated way; however, their functions in breast cancer stem-like cells and their regulation by microRNAs remain elusive. In the present study, we investigated the functional role of microRNA-204 (miR-204) on angiogenesis and vasculogenic mimicry in breast cancer stem-like cells. Using flow cytometry assays, we found that 86.1% of MDA-MB-231 and 92% of Hs-578t breast cancer cells showed the CD44+/CD24- immunophenotype representative of cancer stem-like cells (CSCs). The MDA-MB-231 subpopulation of CSCs exhibited the ability to form mammospheres, as expected. Interestingly, we found that the restoration of miR-204 expression in CSCs significantly inhibited the number and size of the mammospheres. Moreover, we found that MDA-MB-231 and Hs-578t CSCs efficiently undergo angiogenesis and hypoxia-induced vasculogenic mimicry in vitro. The transfection of precursor miR-204 in both CSCs was able to impair the angiogenesis in the HUVEC cell model, which was observed as a diminution in the number of polygons and sprouting cells. Remarkably, miR-204 mimics also resulted in the inhibition of vasculogenic mimicry formation in MDA-MB-231 and Hs-578t CSCs, with a significant reduction in the number of channel-like structures and branch points. Mechanistically, the effects of miR-204 were associated with a diminution of pro-angiogenic VEGFA and ß-catenin protein levels. In conclusion, our findings indicated that miR-204 abrogates the angiogenesis and vasculogenic mimicry development in breast cancer stem-like cells, suggesting that it could be a potential tool for breast cancer intervention based on microRNA replacement therapies.
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Exosomes are a type of cell-derived vesicles that range in size from 30 to 100 nm. They are widely present in various organisms and participate in diverse biological processes, playing crucial roles in tumorigenesis and progression. This study aimed to investigate whether LINC01480 in tumor-derived exosomes is involved in the molecular mechanism of gastric cancer by competitively upregulating the VCAM1 expression through binding miR-204-5p. The study analyzed transcriptome data related to gastric cancer from the cancer genome atlas database and constructed a risk-scoring model for epithelial-mesenchymal transition (EMT)-related lncRNAs to identify eight EMT-related lncRNAs associated with prognosis. EMT-related mRNAs positively correlated with LINC01480 were screened in the ExoRBase database. In vitro cell experiments showed that exosomal LINC01480 can promote the proliferation, migration, invasion, and EMT of gastric cancer cells by upregulating VCAM1 expression through competitive binding with miR-204-5p. In vivo experiments on nude mice showed that exosomal LINC01480 promotes the development of gastric cancer. These results suggest that exosomal LINC01480 could be a potential therapeutic target for gastric cancer.
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MicroARNs , ARN Largo no Codificante , Neoplasias Gástricas , Animales , Ratones , MicroARNs/genética , Neoplasias Gástricas/genética , ARN Endógeno Competitivo , Ratones Desnudos , ARN Largo no Codificante/genética , Proliferación Celular/genética , Línea Celular TumoralRESUMEN
Imatinib resistance in chronic myelogenous leukemia (CML) is a clinical problem. The present study examined the role of NMyc downstream regulatory gene 3 (NDRG3) in imatinib resistance in CML. Quantitative PCR demonstrated that NDRG3 was highly expressed in patients with CML. Cell Counting Kit (CCK)8 experiments proved that NDRG3 promoted the proliferation of K562 CML cells and enhanced imatinib resistance. Dualluciferase assay showed that microRNA (miR)2045p inhibited expression of NDRG3 and immunofluorescence experiments showed that NDRG3 promoted accumulation of ßcatenin in the nucleus, thereby increasing the expression of downstream drug resistance and cell cycleassociated factors (cMyc and MDR1). At the same time, cell proliferation experiments showed that ßcatenin played a role in cell proliferation and drug resistance. Cotransfection with small interfering (si)ßcatenin partially reversed the effect of NDRG3. This finding indicated that NDRG3 plays an important role in imatinib resistance and miR2045p and ßcatenin are involved in the biological behavior of NDRG3. The present results provide theoretical support for overcoming drug resistance in CML.
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Leucemia Mielógena Crónica BCR-ABL Positiva , MicroARNs , Humanos , Mesilato de Imatinib/farmacología , Mesilato de Imatinib/uso terapéutico , beta Catenina/genética , MicroARNs/genética , MicroARNs/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Células K562 , Péptidos y Proteínas de Señalización IntracelularRESUMEN
Ischemic stroke has extremely high mortality and disability rates worldwide. miR-204-5p has been reported to be associated with neurological diseases. However, the relationship linking miR-204-5p to ischemic stroke and its molecular mechanism remain unclear. Herein, we found that expression of miR-204-5p was significantly decreased while EphA4 increased in vivo and vitro, which reached the peak at 24 h after cerebral ischemia/reperfusion. Then, we altered miR-204-5p expression in rats by cerebroventricular injection. Our study showed that miR-204-5p overexpression obviously reduced the brain infarction area and neurological score. We successfully cultured neurons to investigate the downstream mechanism. Upregulation of miR-204-5p increased cell viability and suppressed the release of LDH. Moreover, the proportion of apoptotic cells tested by TUNEL and flow cytometry and protein expression of Cleaved Caspase3 and Bax were inhibited. The relative expression of IL-6, TNF-α, and IL-1ß was repressed. In contrary, knockdown of miR-204-5p showed the opposite results. Bioinformatics and a dual luciferase assay illustrated that EphA4 was a target gene. Further research studies demonstrated that the neuroprotective effects of miR-204-5p could be partially mitigated by upregulating EphA4. Next, we proved that the miR-204-5p/EphA4 axis furtherly activated the PI3K/AKT pathway. We thoroughly illustrated the role of neuroinflammation and apoptosis. However, whether there are other mechanisms associated with the EphA4/PI3K/AKT pathway needs further investigation. Altogether, the miR-204-5p axis ameliorates neurological injury via the EphA4/PI3K/AKT pathway, which is expected to serve as an effective treatment for ischemic stroke.
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Accidente Cerebrovascular Isquémico , MicroARNs , Ratas , Animales , Proteínas Proto-Oncogénicas c-akt/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Accidente Cerebrovascular Isquémico/genética , MicroARNs/genética , MicroARNs/metabolismo , Transducción de Señal , ApoptosisRESUMEN
Objective To explore the role and mechanism of microRNA-204(miR-204) carried by the exosomes of human umbilical cord-derived mesenchymal stem cells(hUC-MSC) in regulating the polarization of macrophages in a mouse model of myocardial ischemia-reperfusion(I/R) injury. Methods After the hUC-MSCs were isolated,cultured,and identified,their adipogenic and osteogenic differentiation capabilities were determined.The exosomes of hUC-MSCs were separated by ultracentrifugation,and the expression of CD81,CD63,tumor susceptibility gene 101(Tsg101),and calnexin in the exosomes was determined by Nanoparticle Tracking Analysis software,transmission electron microscopy,and Western blotting.Three groups(hUC-MSC,miR-204 mimic,and negative control) were designed for the determination of the expression of miR-204 in the cells and their exosomes by qRT-PCR.The C57BL/6J mice were randomly assigned into a sham operation group,an I/R group,a hUC-MSC exosomes group,a negative control group,and a miR-204 mimic group.Except the sham operation group,the I/R model was established by ligating the left anterior descending artery.The echocardiography system was employed to detect the heart function of mice.HE staining was employed to observe the pathological changes of mouse myocardium.ELISA was employed to determine the levels of interleukin-1ß(IL-1ß),tumor necrosis factor-α(TNF-α),arginase 1(Arg-1),and IL-10 in the myocardial tissue.After the macrophages of mouse myocardial tissue were isolated,flow cytometry was employed to determine the expression of CD11c and CD206,and ELISA to measure the levels of IL-1ß,TNF-α,Arg-1,and IL-10 in the macrophages. Results hUC-MSCs had adipogenic and osteogenic differentiation capabilities,and the exosomes were successfully identified.Compared with the negative control group,the miR-204 mimic group showed up-regulated expression of miR-204 in hUC-MSCs and their exosomes(P<0.001,P<0.001).Compared with the sham operation group,the modeling of I/R increased the left ventricular end-diastolic diameter(LVEDD)(P<0.001),left ventricular end-systolic diameter(LVESD)(P<0.001),myocardial injury score(P<0.001),and the levels of IL-1ß(P<0.001),TNF-α(P<0.001),and CD11c(P<0.001).Meanwhile,it lowered the left ventricular ejection fraction(LVEF)(P<0.001),left ventricular fractional shortening(LVFS)(P<0.001),Arg-1(P<0.001),IL-10(P<0.001),and CD206(P<0.001).Compared with those in the I/R group,the LVEDD(P<0.001),LVESD(P<0.001),myocardial injury score(P<0.001),and the levels of IL-1ß(P<0.001),TNF-α(P=0.010),and CD11c(P<0.001) reduced,while LVEF(P<0.001),LVFS(P<0.001),and the levels of Arg-1(P<0.001),IL-10(P=0.028),and CD206(P=0.022) increased in the hUC-MSC exosomes group.Compared with those in the negative control group,the LVEDD(P<0.001),LVESD(P<0.001),myocardial injury score(P=0.001),and the levels of IL-1ß(P=0.048),TNF-α(P<0.001),and CD11c(P=0.007) reduced,while the LVEF(P<0.001),LVFS(P<0.001),and the levels of Arg-1(P<0.001),IL-10(P=0.001),and CD206(P=0.001) increased in the miR-204 mimic group. Conclusion The hUC-MSC exosomes overexpressing miR-204 can inhibit the polarization of macrophages in the I/R mouse model to M1-type and promote the polarization to M2-type.
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Exosomas , Células Madre Mesenquimatosas , MicroARNs , Daño por Reperfusión Miocárdica , Animales , Humanos , Ratones , Modelos Animales de Enfermedad , Exosomas/metabolismo , Exosomas/patología , Interleucina-10/metabolismo , Macrófagos , Ratones Endogámicos C57BL , MicroARNs/genética , Osteogénesis , Volumen Sistólico , Factor de Necrosis Tumoral alfa/metabolismo , Cordón Umbilical/metabolismo , Cordón Umbilical/patología , Función Ventricular IzquierdaRESUMEN
INTRODUCTION: At present, the mortality rate of clear cell renal cell carcinoma (ccRCC) remains high. The development of biomarkers is conducive to the early diagnosis and treatment of cancer. This research aimed to explore the role of microRNA-204-5p and the downstream gene in ccRCC. METHODS: We used bioinformatics analysis and qRT-PCR for measurement of microRNA-204-5p, qRT-PCR, and Western blot for GXYLT2 mRNA and protein levels, respectively. We conducted in vitro experiments like CCK-8, colony formation, Transwell, wound healing, and cell cycle assays to assess the role of microRNA-204-5p and GXYLT2 in ccRCC. Besides, the target relationship of microRNA-204-5p and GXYLT2 was confirmed through dual-luciferase assay. RESULTS: This study disclosed that microRNA-204-5p was underexpressed in ccRCC cells and tissues, which was closely associated with prognosis of patients with ccRCC. Stable forced expression of microRNA-204-5p hindered malignant phenotypes of ccRCC cells. Further detection unfolded that micro-RNA-204-5p bound the 3'-UTR of GXYLT2 to repress its expression. Besides, forced expression of microRNA-204-5p restored the promoting impact of overexpression of GXYLT2 on malignant progression of ccRCC cells. CONCLUSION: These findings revealed the vital role of microRNA-204-5p and GXYLT2 in ccRCC progression, as well as the possibility of microRNA-204-5p in improving ccRCC prognosis and treatment.
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Carcinoma de Células Renales , Neoplasias Renales , MicroARNs , Humanos , Regiones no Traducidas 3' , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/patología , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Regulación hacia Abajo , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales/genética , Neoplasias Renales/patología , MicroARNs/genética , MicroARNs/metabolismoRESUMEN
BACKGROUND: Vasculogenic mimicry (VM) is characterized by formation of three-dimensional (3D) channels-like structures by tumor cells, supplying the nutrients needed for tumor growth. VM is stimulated by hypoxic tumor microenvironment, and it has been associated with increased metastasis and clinical poor outcome in cancer patients. cAMP responsive element (CRE)-binding protein 5 (CREB5) is a hypoxia-activated transcription factor involved in tumorigenesis. However, CREB5 functions in VM and if its regulated by microRNAs remains unknown in breast cancer. OBJECTIVE: We aim to study the functional relationships between VM, CREB5 and microRNA-204-5p (miR-204) in breast cancer cells. METHODS: CREB5 expression was evaluated by mining the public databases, and using RT-qPCR and Western blot assays. CREB5 expression was silenced using short-hairpin RNAs in MDA-MB-231 and MCF-7 breast cancer cells. VM formation was analyzed using matrigel-based cultures in hypoxic conditions. MiR-204 expression was restored in cancer cells by transfection of RNA mimics. Luciferase reporter assays were performed to evaluate the binding of miR-204 to 3'UTR of CREB5. RESULTS: Our data showed that CREB5 mRNA expression was upregulated in a set of breast cancer cell lines and clinical tumors, and it was positively associated with poor prognosis in lymph nodes positive and grade 3 basal breast cancer patients. Silencing of CREB5 impaired the hypoxia-induced formation of 3D channels-like structures representative of the early stages of VM in MDA-MB-231 cells. In contrast, VM formation was not observed in MCF-7 cells. Interestingly, we found that CREB5 expression was negatively regulated by miR-204 mimics in breast cancer cells. Functional analysis confirmed that miR-204 binds to CREB5 3'-UTR indicating that it's an ulterior effector. CONCLUSIONS: Our findings suggested that CREB5 could be a potential biomarker of disease progression in basal subtype of breast cancer, and that perturbations of the miR-204/CREB5 axis plays an important role in VM development in breast cancer cells.
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Neoplasias de la Mama , MicroARNs , Regiones no Traducidas 3' , Neoplasias de la Mama/patología , Línea Celular Tumoral , Proteína de Unión al Elemento de Respuesta al AMP Cíclico/genética , Proteína de Unión al Elemento de Respuesta al AMP Cíclico/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Hipoxia/genética , MicroARNs/genética , MicroARNs/metabolismo , Neovascularización Patológica/genética , Factores de Transcripción/genética , Microambiente TumoralRESUMEN
BACKGROUND: MicroRNAs regulate cardiac hypertrophy development, which precedes and predicts the risk of heart failure. microRNA-204-5p (miR-204) is well expressed in cardiomyocytes, but its role in developing cardiac hypertrophy and cardiac dysfunction (CH/CD) remains poorly understood. METHODS: We performed RNA-sequencing, echocardiographic, and molecular/morphometric analysis of the heart of mice lacking or overexpressing miR-204 five weeks after trans-aortic constriction (TAC). The neonatal rat cardiomyocytes, H9C2, and HEK293 cells were used to determine the mechanistic role of miR-204. RESULTS: The stretch induces miR-204 expression, and miR-204 inhibits the stretch-induced hypertrophic response of H9C2 cells. The mice lacking miR-204 displayed a higher susceptibility to CH/CD during pressure overload, which was reversed by the adeno-associated virus serotype-9-mediated cardioselective miR-204 overexpression. Bioinformatic analysis of the cardiac transcriptomics of miR-204 knockout mice following pressure overload suggested deregulation of apelin-receptor (APJ) signalling. We found that the stretch-induced extracellular signal-regulated kinase 1/2 (ERK1/2) activation and hypertrophy-related genes expression depend on the APJ, and both of these effects are subject to miR-204 levels. The dynamin inhibitor dynasore inhibited both stretch-induced APJ endocytosis and ERK1/2 activation. In contrast, the miR-204-induced APJ endocytosis was neither inhibited by dynamin inhibitors (dynasore and dyngo) nor associated with ERK1/2 activation. We find that the miR-204 increases the expression of ras-associated binding proteins (e.g., Rab5a, Rab7) that regulate cellular endocytosis. CONCLUSIONS: Our results show that miR-204 regulates trafficking of APJ and confers resistance to pressure overload-induced CH/CD, and boosting miR-204 can inhibit the development of CH/CD.
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Receptores de Apelina/antagonistas & inhibidores , Cardiomegalia/prevención & control , MicroARNs/farmacología , Animales , Receptores de Apelina/metabolismo , Cardiomegalia/tratamiento farmacológico , Modelos Animales de Enfermedad , Cardiopatías/tratamiento farmacológico , Cardiopatías/prevención & control , MicroARNs/metabolismo , Ratas , Transducción de Señal/efectos de los fármacosRESUMEN
Cardiovascular ischemia/reperfusion (I/R) injury is primarily caused by oxygen recovery after prolonged hypoxia. Previous studies found that the long non coding RNA (lncRNA) nuclear enriched abundant transcript 1 (NEAT1) was involved in cardiovascular pathology, and that NODlike receptor protein 3 (NLRP3) inflammasome activationdependent pyroptosis played a key role in cardiovascular I/R injury. The present study aimed to explore the molecular mechanism of I/R pathogenesis in order to provide novel insights for potential future therapies. Cell viability and lactate dehydrogenase enzyme activity assays were used to detect cell injury after human umbilical vein endothelial cells (HUVECs) were subjected to hypoxia/reoxygenation (H/R). The expression of the NEAT1/microRNA (miR)204/BRCA1/BRCA2containing complex subunit 3 (BRCC3) axis was examined by reverse transcriptionquantitative PCR, and the associations among genes were confirmed by luciferase reporter assays. Western blotting and ELISA were used to measure the level of NLRP3 inflammasome activationdependent pyroptosis. The results demonstrated that NEAT1, BRCC3 expression and NLRP3 inflammasome activationdependent pyroptosis were significantly increased in H/Rinjured HUVECs, whereas silencing BRCC3 or NEAT1 attenuated H/Rinduced injury and pyroptosis. NEAT1 positively regulated BRCC3 expression via competitively binding with miR204. Moreover, NEAT1 overexpression counteracted miR204 mimicinduced injury, BRCC3 expression and NLRP3 inflammasome activationdependent pyroptosis. Taken together, these findings demonstrated that inhibition of lncRNA NEAT1 protects HUVECs against H/Rinduced NLRP3 inflammasome activation by targeting the miR204/BRCC3 axis.
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Enzimas Desubicuitinizantes/antagonistas & inhibidores , Endotelio Vascular/efectos de los fármacos , Hipoxia/fisiopatología , Inflamación/prevención & control , MicroARNs/antagonistas & inhibidores , Proteína con Dominio Pirina 3 de la Familia NLR/antagonistas & inhibidores , ARN Largo no Codificante/antagonistas & inhibidores , Supervivencia Celular , Enzimas Desubicuitinizantes/genética , Enzimas Desubicuitinizantes/metabolismo , Células Endoteliales de la Vena Umbilical Humana , Humanos , Inflamasomas/efectos de los fármacos , Inflamasomas/metabolismo , Inflamación/etiología , Inflamación/metabolismo , Inflamación/patología , MicroARNs/genética , Sustancias Protectoras/farmacología , Piroptosis , Daño por ReperfusiónRESUMEN
Oxalate-induced oxidative stress causes damage to cells, accompanied with renal deposition of calcium oxalate crystals. Recent studies have highlighted the extensive functions of microRNAs (miRNAs) in various processes, including cellular responses to oxidative stress. Hence, this study was intended to analyze the role of miR-204 in the calcium oxalate kidney-stone formation and the underlying mechanism. In silico analysis was performed to determine the miRNA/mRNA interaction involved in calculus, while dual-luciferase reporter assay was conducted for validation. A calcium oxalate kidney-stone model was established by H2O2 induction in RTEC HK-2 cells, in which the expression of miR-204 was examined. Gain- and loss-of-function approaches were employed to alter the expression of miR-204/MUC4 so as to assess the detailed role of miR-204 in oxidative stress injury in renal tubular epithelial cells (RTECs) and calcium oxalate kidney-stone formation. MUC4, an up-regulated gene in H2O2-induced HK-2 cells, was a target of MUC4. miR-204 functionally targeted MUC4 and blocked the ERK pathway activation. Furthermore, up-regulated miR-204 contributed to promotion of RTEC proliferation and suppression of ROS levels, RTEC apoptosis as well as formation of calcium oxalate crystal. Taken together, miR-204 impairs MUC4-dependent activation of the ERK signaling pathway and consequently ameliorates oxidative stress damage to RTECs and prevents calcium oxalate kidney-stone formation.
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Oxalato de Calcio , MicroARNs , Oxalato de Calcio/metabolismo , Células Epiteliales/metabolismo , Humanos , Peróxido de Hidrógeno/metabolismo , Peróxido de Hidrógeno/toxicidad , Riñón/metabolismo , Sistema de Señalización de MAP Quinasas , MicroARNs/genética , MicroARNs/metabolismo , Estrés Oxidativo , Transducción de SeñalRESUMEN
Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the cell Transwell assay data in the article (featured in Figs. 3B and 6B) were strikingly similar to data that appearing in different form in another article by different authors at different research institutions, which had already been published elsewhere at the time of the present article's submission. Owing to the fact that the contentious data in the above article had already appeared in different form in another article prior to its submission to Oncology Reports, the Editor has decided that this paper should be retracted from the Journal. The authors did not reply to indicate whether or not they agreed with the retraction of the paper. The Editor apologizes to the readership for any inconvenience caused. [the original article was published on Oncology Reports 34: 399406, 2015; DOI: 10.3892/or.2015.3986].
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Mitochondrial dysfunction is an early feature of Alzheimer's disease (AD), whereby accumulation of damaged mitochondria in conjunction with impaired mitophagy contributes to neurodegeneration. Various non-transcribed microRNAs (miRNAs) are involved in this process. In the present study, we aimed to decipher the participation of miR-204 in a murine AD model. Primary hippocampal neurons were isolated from mice and treated with ß-amyloid 1-42 (Aß1-42) to establish a cell model of AD. Dichloro-dihydro-fluorescein diacetate and dihydrorhodamine 123 staining assays were performed to measure total reactive oxygen species (ROS) and mitochondrial ROS production in neurons, and MitoSOX staining was done to analyze mitochondrial ROS production in hippocampus. Furthermore, mitochondrial autophagy was observed in hippocampus from amyloid precursor protein/pesenilin-1 AD modeled mice, and their cognitive function was assessed by Morris water maze. Mitochondrial damage, ROS production, and mitochondrial autophagy were observed in AD cell model induced by Aß1-42. In AD, signal transducer and activator of transcription 3 (STAT3) and transient receptor potential mucolipin-1 (TRPML1) expression was downregulated, although miR-204 expression was upregulated. TRPML1 overexpression, downregulation of miR-204, or STAT3 pathway activation reduced the Aß1-42-induced mitochondrial damage, along with ROS production and mitochondrial autophagy in vivo and in vitro. Silencing of miR-204 could upregulate TRPML1 expression, thus suppressing ROS production and mitochondrial autophagy in AD through STAT3 pathway.
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Circular RNA (circRNA/circ)ubiquitin associated protein 2 (UBAP2), a newly recognized circRNA, serves a functional role in several types of tumor, including ovarian cancer, colorectal cancer and osteosarcoma. However, the precise roles and molecular mechanism underlying circUBAP2 in osteosarcoma (OS) are not completely understood. In the present study, the expression levels of circUBAP2, microRNA (miR)2043p and (HMGA2) were evaluated via reverse transcriptionquantitative PCR in OS tissues and cells. OS cell proliferation, migration, invasion and apoptosis were assessed by performing Cell Counting Kit8, Transwell and flow cytometry assays, respectively. HMGA2 protein expression levels were determined via western blotting. Dualluciferase reporter assays were performed to verify the interaction between circUBAP2 and miR2043p, and between miR2043p and HMGA2. An RNA immunoprecipitation (RIP) assay was conducted to confirm the interaction between circUBAP2 and miR2043p. The results demonstrated that circUBAP2 expression was significantly upregulated in OS tissues and cell lines compared with paracancerous tissues and hFOB1.19 cells, respectively. In addition, high circUBAP2 expression levels in patients with OS were associated with a lower survival rate compared with lower expression levels in patients with OS. The functional assays revealed that circUBAP2 knockdown significantly inhibited OS cell proliferation, migration and invasion, but increased OS cell apoptosis compared with the small interfering RNAnegative control (siNC) group. The dualluciferase reporter and RIP assay results confirmed that circUBAP2 bound to miR2043p. Moreover, miR2043p expression was significantly downregulated in OS tissues compared with paracancerous tissues, and miR2043p expression was negatively correlated with circUBAP2 expression in OS tissues. Collectively, the results demonstrated that miR2043p was associated with circUBAP2 knockdownmediated inhibition of OS cell malignant behavior. Moreover, miR2043p was also identified as one of the direct targets of HMGA2. Collectively, the results indicated that compared with the siNC group, circUBAP2 knockdown significantly inhibited OS cell malignant behavior by binding to miR2043p, which subsequently regulated HMGA2 expression. Therefore, the present study demonstrated that circUBAP2 expression was upregulated in OS, and circUBAP2 regulated OS cell malignant behavior via the miR2043p/HMGA2 axis.
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Neoplasias Óseas/genética , Proteína HMGA2/genética , MicroARNs/metabolismo , Osteosarcoma/genética , ARN Circular/metabolismo , Adolescente , Adulto , Apoptosis/genética , Neoplasias Óseas/diagnóstico , Neoplasias Óseas/patología , Línea Celular Tumoral , Niño , Progresión de la Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Masculino , MicroARNs/genética , Estadificación de Neoplasias , Osteosarcoma/diagnóstico , Osteosarcoma/patología , ARN Circular/genética , Regulación hacia Arriba , Adulto JovenRESUMEN
BACKGROUND: Angiopoietin-like protein 2 (ANGPTL2) is linked to various tumors. MicroRNA-204 (miR-204) is associated with colorectal cancer (CRC). Bioinformatic analysis has demonstrated a targeting relationship between miR-204 and ANGPTL2. The present study aimed to investigate the role of miR-204 in the proliferation and apoptosis of colorectal tumor cells. METHODS: Colorectal tumor tissues were collected. Normal colon mucosa was used as a control. The relationship between miR-204 and ANGPTL2 expression and tumor stage and prognosis was analyzed. The dual-luciferase reporter assay confirmed targeted regulation between miR-204 and ANGPTL2. SW480 cells were allocated to the miR-NC group and the miR-204 mimic group, followed by apoptotic analysis using flow cytometry and cellular proliferation analysis using EdU staining. RESULTS: Compared with normal colonic mucosa, miR-204 expression was decreased in colorectal tumor tissues and ANGPTL2 expression was increased, which correlated with TNM staging. The prognosis of patients with low miR-204 expression and high ANGPTL2 expression was worse than for patients with high miR-204 expression and low ANGPTL2 expression. The dual-luciferase reporter assay confirmed a targeting regulation relationship between miR-204 and ANGPTL2. Transfection of miR-204 mimic significantly inhibited the expression of ANGPTL2 and cell proliferation in SW480 cells and promoted apoptosis. CONCLUSIONS: Downregulating miR-204 expression plays a vital role in upregulating ANGPTL2 expression and promoting the pathogenesis of CRC. MiR-204 is able to hinder the proliferation of colorectal tumor cells and encourage apoptosis by targeting the inhibition of ANGPTL2 expression.
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Accumulating studies highlight the role of long noncoding RNAs (lncRNAs)/microRNAs (miRNAs)/messenger RNAs (mRNAs) as important regulatory networks in various human cancers, including thyroid cancer (TC). This study aimed to investigate a novel regulatory network dependent on lncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in relation to TC development. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot were initially employed to detect the expression of MALAT1, insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2), and myelocytomatosis (MYC) in TC cells. Interactions among MALAT1, miR-204, and IGF2BP2 were then identified in vitro. The biological processes of proliferation, migration, invasion, and apoptosis were evaluated in vitro via gain- and loss-of-function experiments, followed by in vivo validation using xenograft mice. Our data indicated that MALAT1 and IGF2BP2 were highly expressed, while miR-204 was poorly expressed in TC. IGF2BP2 was verified as a target of miR-204. MALAT1 was found to upregulate IGF2BP2 and enhance MYC expression via m6A modification recognition by competitively binding to miR-204, conferring a stimulatory effect on proliferation, migration, and invasion of TC cells, which was accompanied by weakened tumor growth and cell apoptosis. Altogether, the central findings of our study suggest that MALAT1 contributes to TC progression through the upregulation of IGF2BP2 by binding to miR-204.
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The participation of long noncoding RNAs (lncRNAs) and microRNAs (miRs) in the progression of rheumatoid arthritis (RA) is a key area of investigation. The current study aimed to investigate the action of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in fibroblast-like synoviocyte (FLS) proliferation and synovitis in RA. A rat model of RA was established. LncRNA NEAT1 expression in the synovial tissues of patients with RA and FLSs from the RA rat model was determined using RT-qPCR. Next, dual luciferase reporter gene assay was applied to investigate the relationship between miR-129/204 and mitogen-activated protein kinase (MAPK)/extracellular regulated protein kinase (ERK). A putative binding relationship between miR-204 and lncRNA NEAT1 was evaluated by RIP assay, and miR-129 promoter methylation was determined using MSP. After the expression of lncRNA NEAT1, miR-129 or miR-204 was altered in FLSs, the extent of ERK1/2 phosphorylation was assessed. In addition, FLS synovitis and proliferation were determined by ELISA and EdU assay, respectively. In RA rats, lncRNA NEAT1 was silenced and miR-129/miR-204 was overexpressed to explore their roles in vivo. LncRNA NEAT1 was upregulated, while miR-129 and miR-204 were downregulated in RA synovial tissues and FLSs. MAPK1 was target gene of both miR-129 and miR-204. LncRNA NEAT1 bound to miR-204 and promoted miR-129 promoter methylation. Silencing lncRNA NEAT1 or overexpressing miR-129/miR-204 enhanced miR-129/miR-204 expression, but reduced the extent of ERK1/2 phosphorylation, proliferation of FLSs, and synovitis in RA. Collectively, silencing lncRNA NEAT1 promoted miR-129 and miR-204 to inhibit the MAPK/ERK signalling pathway, reducing FLS synovitis in RA.Abbreviations: ACR: American College of Rheumatology; ELISA: Enzyme-linked immunosorbent assay; ERK: extracellular signal-regulated kinase; FLS: fibroblast-like synoviocyte; GADPH: glyceraldehyde-3-phosphate dehydrogenase; HRP: horseradish peroxidase; IFA: Incomplete Freund's Adjuvant; lncRNAs: long noncoding RNAs; MSP: Methylation-specific PCR; NC: negative control; NEAT1: nuclear paraspeckle assembly transcript 1; OD: optical density; RA: rheumatoid arthritis; RIPA: Radio Immunoprecipitation Assay; RLU: relative light units; RT-qPCR: reverse transcription quantitative polymerase chain reaction; UTR: untranslated region.
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Artritis Reumatoide/patología , MicroARNs/genética , ARN Largo no Codificante/genética , Anciano , Animales , Artritis Reumatoide/genética , Artritis Reumatoide/metabolismo , Estudios de Casos y Controles , Células Cultivadas , Regulación hacia Abajo/genética , Quinasas MAP Reguladas por Señal Extracelular/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Regulación de la Expresión Génica , Humanos , Sistema de Señalización de MAP Quinasas/genética , Masculino , MicroARNs/metabolismo , Persona de Mediana Edad , Interferencia de ARN , Ratas , Ratas Wistar , Inducción de Remisión , Sinoviocitos/metabolismo , Sinoviocitos/patologíaRESUMEN
OBJECTIVE: Ankylosing spondylitis (AS) is a disease characterized by inflammation of the sacroiliac joint and the attachment point of the spine. This study aimed to investigate the effect of microRNA (miR)-204-targeted GSDMD on fibroblast-like synoviocytes (FLSs) in AS. METHODS: miR-204, GSDMD, pyrolysis-related genes (Caspase-1, Caspase-11 and NLRP3) in synovial tissues from AS patients were tested by RT-qPCR. Online website prediction and dual luciferase reporter gene assay were conducted to verify the binding relationship between miR-204 and GSDMD. FLSs were isolated from AS patients and transfected with miR-204- or GSDMD-related oligonucleotides, siRNA and plasmids to explore their roles in pyroptosis of FLSs. Intracellular [Ca2+] was detected by laser scanning confocal microscopy, reactive oxygen species (ROS) by DCFH-DA and pyrolysis by AO/EB staining and flow cytometry. RESULTS: Decreased miR-204 and elevated GSDMD were found in synovial tissue of patients with AS. miR-204 could directly target GSDMD and inhibit GSDMD protein expression. FLSs treated with miR-204 mimic inhibited the pyroptosis rate and Caspase-1/PI double-positive cells and reduced [Ca2+], ROS, NLRP3, Caspase-1 and Caspase-11 levels in FLSs. Up-regulating GSDMD blocked the effect of miR-204 overexpression on FLSs. CONCLUSION: Altogether, up-regulated miR-204 suppresses pyroptosis of FLSs in AS via suppressing GSDMD, which may help us to understand the mechanism of AS deeply.
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Péptidos y Proteínas de Señalización Intracelular/metabolismo , MicroARNs/metabolismo , Proteínas de Unión a Fosfato/metabolismo , Piroptosis , Espondilitis Anquilosante/metabolismo , Sinoviocitos/metabolismo , Adulto , Calcio/metabolismo , Estudios de Casos y Controles , Caspasa 1/genética , Caspasa 1/metabolismo , Caspasas/genética , Caspasas/metabolismo , Células Cultivadas , Femenino , Regulación de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , MicroARNs/genética , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Proteínas de Unión a Fosfato/genética , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Espondilitis Anquilosante/genética , Espondilitis Anquilosante/patología , Sinoviocitos/patología , Adulto JovenRESUMEN
Long noncoding (lnc)RNAs and microRNAs (miRNAs/miRs) have physiological and pathological functions in various diseases, including gastric cancer (GC). The current study explored the association between lncRNA small nucleolar RNA host gene 4 (SNHG4) and miR148a3p, and their functions in GC cells. SNHG4 expression and overall survival data were analyzed using bioinformatics, and the interaction of SNHG4 and miR148a3p was predicted using starBase and confirmed via a dualluciferase reporter assay. Cell viability, colony formation ability and apoptosis rate were detected using Cell Counting Kit8, colony formation and flow cytometry assays, respectively. Cell migration and invasion were determined via woundhealing and Transwell assays. mRNA and protein expression levels were determined via reverse transcriptionquantitative PCR and western blotting. The results demonstrated that in GC tissues and cell lines, SNHG4 was highly expressed, while miR2045p expression was decreased, and that the expression levels of SNHG4 and miR2045p were negatively correlated. The downregulated expression of SNHG4 decreased the effects of miR2045p inhibitor on promoting cell proliferation, migration, invasion and epithelialmesenchymal transition, but enhanced the inhibitory effect of miR2045p on GC cell apoptosis. The findings of the current study revealed the potential mechanism of the SNHG4miR2045p pathway in GC, which may be conducive to the development of novel drugs against GC growth.