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1.
EMBO J ; 42(1): e110937, 2023 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-36382717

RESUMO

Hutchinson-Gilford progeria syndrome (HGPS) is a lethal premature aging disorder without an effective therapeutic regimen. Because of their targetability and influence on gene expression, microRNAs (miRNAs) are attractive therapeutic tools to treat diseases. Here we identified that hsa-miR-59 (miR-59) was markedly upregulated in HGPS patient cells and in multiple tissues of an HGPS mouse model (LmnaG609G/G609G ), which disturbed the interaction between RNAPII and TFIIH, resulting in abnormal expression of cell cycle genes by targeting high-mobility group A family HMGA1 and HMGA2. Functional inhibition of miR-59 alleviated the cellular senescence phenotype of HGPS cells. Treatment with AAV9-mediated anti-miR-59 reduced fibrosis in the quadriceps muscle, heart, and aorta, suppressed epidermal thinning and dermal fat loss, and yielded a 25.5% increase in longevity of LmnaG609G/G609G mice. These results identify a new strategy for the treatment of HGPS and provide insight into the etiology of HGPS disease.


Assuntos
MicroRNAs , Progéria , Camundongos , Animais , Progéria/genética , Antagomirs/uso terapêutico , Senescência Celular/genética , MicroRNAs/genética , Fenótipo
2.
Blood ; 143(5): 429-443, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-37847858

RESUMO

ABSTRACT: Hematological malignancies such as Burkitt lymphoma (BL), Hodgkin lymphoma (HL), and diffuse large B-cell lymphoma (DLBCL) cause significant morbidity in humans. A substantial number of these lymphomas, particularly HL and DLBCLs have poorer prognosis because of their association with Epstein-Barr virus (EBV). Our earlier studies have shown that EBV-encoded nuclear antigen (EBNA2) upregulates programmed cell death ligand 1 in DLBCL and BLs by downregulating microRNA-34a. Here, we investigated whether EBNA2 affects the inducible costimulator (ICOS) ligand (ICOSL), a molecule required for efficient recognition of tumor cells by T cells through the engagement of ICOS on the latter. In virus-infected and EBNA2-transfected B-lymphoma cells, ICOSL expression was reduced. Our investigation of the molecular mechanisms revealed that this was due to an increase in microRNA-24 (miR-24) by EBNA2. By using ICOSL 3' untranslated region-luciferase reporter system, we validated that ICOSL is an authentic miR-24 target. Transfection of anti-miR-24 molecules in EBNA2-expressing lymphoma cells reconstituted ICOSL expression and increased tumor immunogenicity in mixed lymphocyte reactions. Because miR-24 is known to target c-MYC, an oncoprotein positively regulated by EBNA2, we analyzed its expression in anti-miR-24 transfected lymphoma cells. Indeed, the reduction of miR-24 in EBNA2-expressing DLBCL further elevated c-MYC and increased apoptosis. Consistent with the in vitro data, EBNA2-positive DLBCL biopsies expressed low ICOSL and high miR-24. We suggest that EBV evades host immune responses through EBNA2 by inducing miR-24 to reduce ICOSL expression, and for simultaneous rheostatic maintenance of proproliferative c-MYC levels. Overall, these data identify miR-24 as a potential therapeutically relevant target in EBV-associated lymphomas.


Assuntos
Infecções por Vírus Epstein-Barr , Doença de Hodgkin , Linfoma Difuso de Grandes Células B , MicroRNAs , Humanos , Antagomirs , Infecções por Vírus Epstein-Barr/complicações , Infecções por Vírus Epstein-Barr/genética , Infecções por Vírus Epstein-Barr/metabolismo , Antígenos Nucleares do Vírus Epstein-Barr/genética , Antígenos Nucleares do Vírus Epstein-Barr/metabolismo , Herpesvirus Humano 4/genética , Doença de Hodgkin/complicações , Ligantes , Linfoma Difuso de Grandes Células B/metabolismo , MicroRNAs/genética , Proteínas Virais/metabolismo
3.
Nucleic Acids Res ; 52(9): 5273-5284, 2024 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-38348876

RESUMO

RNA interference (RNAi) is an endogenous process that can be harnessed using chemically modified small interfering RNAs (siRNAs) to potently modulate gene expression in many tissues. The route of administration and chemical architecture are the primary drivers of oligonucleotide tissue distribution, including siRNAs. Independently of the nature and type, oligonucleotides are eliminated from the body through clearance tissues, where their unintended accumulation may result in undesired gene modulation. Divalent siRNAs (di-siRNAs) administered into the CSF induce robust gene silencing throughout the central nervous system (CNS). Upon clearance from the CSF, they are mainly filtered by the kidneys and liver, with the most functionally significant accumulation occurring in the liver. siRNA- and miRNA-induced silencing can be blocked through substrate inhibition using single-stranded, stabilized oligonucleotides called antagomirs or anti-siRNAs. Using APOE as a model target, we show that undesired di-siRNA-induced silencing in the liver can be mitigated through administration of liver targeting GalNAc-conjugated anti-siRNAs, without impacting CNS activity. Blocking unwanted hepatic APOE silencing achieves fully CNS-selective silencing, essential for potential clinical translation. While we focus on CNS/liver selectivity, coadministration of differentially targeting siRNA and anti-siRNAs can be adapted as a strategy to achieve tissue selectivity in different organ combinations.


Assuntos
Sistema Nervoso Central , Interferência de RNA , Animais , Humanos , Masculino , Camundongos , Acetilgalactosamina/química , Antagomirs/genética , Antagomirs/metabolismo , Apolipoproteínas E/genética , Sistema Nervoso Central/metabolismo , Inativação Gênica , Fígado/metabolismo , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo
4.
Proc Natl Acad Sci U S A ; 120(20): e2220334120, 2023 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-37155893

RESUMO

Esophageal squamous cell carcinoma (ESCC) is a deadly disease with few prevention or treatment options. ESCC development in humans and rodents is associated with Zn deficiency (ZD), inflammation, and overexpression of oncogenic microRNAs: miR-31 and miR-21. In a ZD-promoted ESCC rat model with upregulation of these miRs, systemic antimiR-31 suppresses the miR-31-EGLN3/STK40-NF-κB-controlled inflammatory pathway and ESCC. In this model, systemic delivery of Zn-regulated antimiR-31, followed by antimiR-21, restored expression of tumor-suppressor proteins targeted by these specific miRs: STK40/EGLN3 (miR-31), PDCD4 (miR-21), suppressing inflammation, promoting apoptosis, and inhibiting ESCC development. Moreover, ESCC-bearing Zn-deficient (ZD) rats receiving Zn medication showed a 47% decrease in ESCC incidence vs. Zn-untreated controls. Zn treatment eliminated ESCCs by affecting a spectrum of biological processes that included downregulation of expression of the two miRs and miR-31-controlled inflammatory pathway, stimulation of miR-21-PDCD4 axis apoptosis, and reversal of the ESCC metabolome: with decrease in putrescine, increase in glucose, accompanied by downregulation of metabolite enzymes ODC and HK2. Thus, Zn treatment or miR-31/21 silencing are effective therapeutic strategies for ESCC in this rodent model and should be examined in the human counterpart exhibiting the same biological processes.


Assuntos
Carcinoma de Células Escamosas , Neoplasias Esofágicas , Carcinoma de Células Escamosas do Esôfago , MicroRNAs , Humanos , Ratos , Animais , Neoplasias Esofágicas/tratamento farmacológico , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas do Esôfago/tratamento farmacológico , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/patologia , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Antagomirs , Zinco/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Inflamação/complicações , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Movimento Celular , Proteínas de Ligação a RNA/metabolismo
5.
Development ; 149(3)2022 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-35156681

RESUMO

Axolotls are an important model organism for multiple types of regeneration, including functional spinal cord regeneration. Remarkably, axolotls can repair their spinal cord after a small lesion injury and can also regenerate their entire tail following amputation. Several classical signaling pathways that are used during development are reactivated during regeneration, but how this is regulated remains a mystery. We have previously identified miR-200a as a key factor that promotes successful spinal cord regeneration. Here, using RNA-seq analysis, we discovered that the inhibition of miR-200a results in an upregulation of the classical mesodermal marker brachyury in spinal cord cells after injury. However, these cells still express the neural stem cell marker sox2. In vivo cell tracking allowed us to determine that these cells can give rise to cells of both the neural and mesoderm lineage. Additionally, we found that miR-200a can directly regulate brachyury via a seed sequence in the 3'UTR of the gene. Our data indicate that miR-200a represses mesodermal cell fate after a small lesion injury in the spinal cord when only glial cells and neurons need to be replaced.


Assuntos
MicroRNAs/metabolismo , Regeneração da Medula Espinal/genética , Medula Espinal/metabolismo , Regiões 3' não Traduzidas , Ambystoma mexicanum/metabolismo , Animais , Antagomirs/metabolismo , Diferenciação Celular , Proteínas Fetais/genética , Proteínas Fetais/metabolismo , Mesoderma/citologia , Mesoderma/metabolismo , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Neuroglia/citologia , Neuroglia/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Medula Espinal/citologia , Traumatismos da Medula Espinal/metabolismo , Traumatismos da Medula Espinal/patologia , Células-Tronco/citologia , Células-Tronco/metabolismo , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Cauda/fisiologia , Via de Sinalização Wnt , beta Catenina/antagonistas & inibidores , beta Catenina/química , beta Catenina/metabolismo
6.
J Cell Mol Med ; 28(3): e18074, 2024 02.
Artigo em Inglês | MEDLINE | ID: mdl-38186203

RESUMO

We previously found that miR-664a-5p is specifically expressed in urinary exosomes of idiopathic membranous nephropathy (IMN) patients. Homeodomain-interacting protein kinase 2 (HIPK2), a nuclear serine/threonine kinase, plays an important role in nephropathy. But the function of these factors and their connection in MN are unclear. To investigate the function and mechanism of miR-664a-5p in MN, the miR-664a-5p expression in HK-2 cells, exosomes, podocytes and renal tissues were studied, as well as cell growth and apoptosis of these cells, the binding of miR-664a-5p to HIPK2 mRNA, the levels of relative proteins and autophagy. The MN progression in MN mice model was also studied. Albumin increased the miR-664a-5p content and apoptosis of HK-2 cells, which was blocked by miR-664a-5p antagomir. miR-664a-5p bound to the 3' UTR of HIPK2 mRNA, resulting in the up-regulation of Calpain1, GSα shear and the inhibition of autophagy level. Autophagy inhibitor CQ blocked the protective effect of miR-664a-5p antagomir, HIPK2 overexpression, Calpain inhibitor SJA6017 on albumin-mediated injury. MiR-664a-5p from albumin-treated HK-2 cells could be horizontally transported to podocytes through exosomes. Exosomes from albumin-treated HK-2 cells promoted progression of MN mice, AAV-Anti-miR-664-5p (mouse homology miRNA) could improve them. Albumin increases the miR-664a-5p level and causes changes of HIPK2/Calpain1/GSα pathway, which leads to autophagy inhibition and apoptosis up-regulation of renal tubular epithelial cells. miR-664a-5p can horizontally enter podocytes through exosomes resulting in podocytes injury. Targeted inhibition of miR-664a-5p can reduce the apoptosis of renal tubule cells and podocytes, and may improve the MN progression.


Assuntos
Glomerulonefrite Membranosa , MicroRNAs , Animais , Humanos , Camundongos , Albuminas/metabolismo , Antagomirs , Apoptose , Autofagia , Proteínas de Transporte , Glomerulonefrite Membranosa/genética , MicroRNAs/genética , Proteínas Serina-Treonina Quinases/metabolismo , RNA Mensageiro
7.
Funct Integr Genomics ; 24(2): 32, 2024 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-38363406

RESUMO

Researchers have reported that miR-124-3p is highly expressed in patients with chronic endometritis. However, the underlying mechanism of miR-124-3p in the development of endometritis remains unclear. This study constructed an in vitro endometrial cell injury model by treating HEECs with 2 µg/mL LPS for 48 h. Then, 1 mg/kg LPS was injected into both sides of the mouse uterus to construct an in vivo endometrial injury model. The expression of miR-124-3p in human endometrial epithelial cells (HEECs) was assessed using RT‒qPCR. Exosomes were separated from bone marrow-derived mesenchymal stem cells (BMSCs) and cocultured with HEECs. A dual-luciferase reporter assay was performed to confirm the relationship between miR-124-3p and DUSP6. The results indicated that LPS inhibited HEEC viability in a time- and dose-dependent manner. The miR-124-3p inhibitor reversed the LPS-induced apoptosis and inhibition of HEEC viability. In addition, miR-124-3p could be transferred from BMSCs to HEECs by exosomes. Exosomes were derived from BMSCs treated with an NC inhibitor (BMSCs/NC Exo) or miR-124-3p inhibitor (BMSCs/anti-miR-124-3p Exo). In addition, BMSCs/anti-miR-124-3p Exo abolished the LPS-induced inhibition of HEEC viability and proliferation by inducing HEEC apoptosis. Moreover, BMSCs/anti-miR-124-3p Exo alleviated the LPS-induced inflammation of HEECs by upregulating DUSP6 and downregulating p-p65 and p-ERK. Furthermore, in an LPS-induced in vivo endometrial injury model, BMSCs/anti-miR-124-3p Exo increased the expression level of DUSP6 and decreased the expression levels of p-p65 and p-ERK. BMSCs/anti-miR-124-3p Exo protected against LPS-induced endometrial damage in vitro and in vivo by upregulating DUSP6 and downregulating p-p65 and p-ERK1/2. This study showed that BMSCs/anti-miR-124-3p Exo might be a potential alternative for the treatment of endometritis.


Assuntos
Endometrite , Exossomos , MicroRNAs , Feminino , Animais , Camundongos , Humanos , Antagomirs , Lipopolissacarídeos/toxicidade , Endometrite/induzido quimicamente , Endometrite/terapia , MicroRNAs/genética
8.
Development ; 148(5)2021 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-33472846

RESUMO

In mammals, sperm-borne regulators can be transferred to oocytes during fertilization and have different effects on the formation of pronuclei, the first cleavage of zygotes, the development of preimplantation embryos and even the metabolism of individuals after birth. The regulatory role of sperm microRNAs (miRNAs) in the development of bovine preimplantation embryos has not been reported in detail. By constructing and screening miRNA expression libraries, we found that miR-202 was highly enriched in bovine sperm. As a target gene of miR-202, co-injection of SEPT7 siRNA can partially reverse the accelerated first cleavage of bovine embryos caused by miR-202 inhibitor. In addition, both a miR-202 mimic and SEPT7 siRNA delayed the first cleavage of somatic cell nuclear transfer (SCNT) embryos, suggesting that miR-202-SEPT7 mediates the delay of first cleavage of bovine embryos. By further exploring the relationship between miR-202/SEPT7, HDAC6 and acetylated α-tubulin during embryonic development, we investigated how sperm-borne miR-202 regulates the first cleavage process of bovine embryos by SEPT7 and demonstrate the potential of sperm-borne miRNAs to improve the efficiency of SCNT.


Assuntos
Citoesqueleto/metabolismo , Embrião de Mamíferos/metabolismo , MicroRNAs/metabolismo , Septinas/metabolismo , Regiões 3' não Traduzidas , Acetilação , Animais , Antagomirs/metabolismo , Bovinos , Desenvolvimento Embrionário , Feminino , Fertilização in vitro , Desacetilase 6 de Histona/metabolismo , Masculino , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Gravidez , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Septinas/antagonistas & inibidores , Septinas/genética , Espermatozoides/metabolismo , Tubulina (Proteína)/metabolismo , Zigoto/metabolismo
9.
Brain Behav Immun ; 119: 919-944, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38718909

RESUMO

Neuroinflammation and accumulation of Amyloid Beta (Aß) accompanied by deterioration of special memory are hallmarks of Alzheimer's disease (AD). Effective preventative and treatment options for AD are still needed. Microglia in AD brains are characterized by elevated levels of microRNA-17 (miR-17), which is accompanied by defective autophagy, Aß accumulation, and increased inflammatory cytokine production. However, the effect of targeting miR-17 on AD pathology and memory loss is not clear. To specifically inhibit miR-17 in microglia, we generated mannose-coated lipid nanoparticles (MLNPs) enclosing miR-17 antagomir (Anti-17 MLNPs), which are targeted to mannose receptors readily expressed on microglia. We used a 5XFAD mouse model (AD) that recapitulates many AD-related phenotypes observed in humans. Our results show that Anti-17 MLNPs, delivered to 5XFAD mice by intra-cisterna magna injection, specifically deliver Anti-17 to microglia. Anti-17 MLNPs downregulated miR-17 expression in microglia but not in neurons, astrocytes, and oligodendrocytes. Anti-17 MLNPs attenuated inflammation, improved autophagy, and reduced Aß burdens in the brains. Additionally, Anti-17 MLNPs reduced the deterioration in spatial memory and decreased anxiety-like behavior in 5XFAD mice. Therefore, targeting miR-17 using MLNPs is a viable strategy to prevent several AD pathologies. This selective targeting strategy delivers specific agents to microglia without the adverse off-target effects on other cell types. Additionally, this approach can be used to deliver other molecules to microglia and other immune cells in other organs.


Assuntos
Doença de Alzheimer , Encéfalo , Modelos Animais de Doenças , Manose , Camundongos Transgênicos , MicroRNAs , Microglia , Nanopartículas , Animais , Doença de Alzheimer/metabolismo , Doença de Alzheimer/tratamento farmacológico , MicroRNAs/metabolismo , Nanopartículas/administração & dosagem , Camundongos , Microglia/metabolismo , Microglia/efeitos dos fármacos , Manose/farmacologia , Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Peptídeos beta-Amiloides/metabolismo , Lipídeos , Masculino , Antagomirs/farmacologia , Antagomirs/administração & dosagem
10.
Circ Res ; 131(1): 77-90, 2022 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-35534923

RESUMO

BACKGROUND: miRNA therapeutics have gained attention during the past decade. These oligonucleotide treatments can modulate the expression of miRNAs in vivo and could be used to correct the imbalance of gene expression found in human diseases such as obesity, metabolic syndrome, and atherosclerosis. The in vivo efficacy of current anti-miRNA technologies hindered by physiological and cellular barriers to delivery into targeted cells and the nature of miRNAs that allows one to target an entire pathway that may lead to deleterious off-target effects. For these reasons, novel targeted delivery systems to inhibit miRNAs in specific tissues will be important for developing effective therapeutic strategies for numerous diseases including atherosclerosis. METHODS: We used pH low-insertion peptide (pHLIP) constructs as vehicles to deliver microRNA-33-5p (miR-33) antisense oligonucleotides to atherosclerotic plaques. Immunohistochemistry and histology analysis was performed to assess the efficacy of miR-33 silencing in atherosclerotic lesions. We also assessed how miR-33 inhibition affects gene expression in monocytes/macrophages by single-cell RNA transcriptomics. RESULTS: The anti-miR-33 conjugated pHLIP constructs are preferentially delivered to atherosclerotic plaque macrophages. The inhibition of miR-33 using pHLIP-directed macrophage targeting improves atherosclerosis regression by increasing collagen content and decreased lipid accumulation within vascular lesions. Single-cell RNA sequencing analysis revealed higher expression of fibrotic genes (Col2a1, Col3a1, Col1a2, Fn1, etc) and tissue inhibitor of metalloproteinase 3 (Timp3) and downregulation of Mmp12 in macrophages from atherosclerotic lesions targeted by pHLIP-anti-miR-33. CONCLUSIONS: This study provides proof of principle for the application of pHLIP for treating advanced atherosclerosis via pharmacological inhibition of miR-33 in macrophages that avoid the deleterious effects in other metabolic tissues. This may open new therapeutic opportunities for atherosclerosis-associated cardiovascular diseases via selective delivery of other protective miRNAs.


Assuntos
Aterosclerose , MicroRNAs , Placa Aterosclerótica , Antagomirs/metabolismo , Antagomirs/uso terapêutico , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/terapia , Humanos , Macrófagos/metabolismo , MicroRNAs/metabolismo , Placa Aterosclerótica/patologia
11.
Circ Res ; 131(8): 687-700, 2022 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-36098045

RESUMO

BACKGROUND: Chronic heart failure (CHF) is associated with redox imbalance. Downregulation of Nrf2 (nuclear factor [erythroid-derived 2]-like 2) plays important roles in disrupting myocardial redox homeostasis and mediating sympathetic nerve activity in the setting of CHF. However, it is unclear if circulating extracellular vesicles (EVs) elicit sympathetic excitation in CHF by disrupting central redox homeostasis. We tested the hypothesis that cardiac-derived EVs circulate to the presympathetic rostral ventrolateral medulla and contribute to oxidative stress and sympathetic excitation via EV-enriched microRNA-mediated Nrf2 downregulation. METHODS: Data were collected on rats with CHF post-myocardial infarction (MI) and on human subjects with ischemic CHF. EVs were isolated from tissue and plasma, and we determined the miRNAs cargo that related to targeting Nrf2 translation. We tracked the distribution of cardiac-derived EVs using in vitro labeled circulating EVs and cardiac-specific membrane GFP+ transgenic mice. Finally, we tested the impact of exogenously loading of antagomirs to specific Nrf2-related miRNAs on CHF-EV-induced pathophysiological phenotypes in normal rats (eg, sympathetic and cardiac function). RESULTS: Nrf2 downregulation in CHF rats was associated with an upregulation of Nrf2-targeting miRNAs, which were abundant in cardiac-derived and circulating EVs from rats and humans. EVs isolated from the brain of CHF rats were also enriched with Nrf2-targeting miRNAs and cardiac-specific miRNAs. Cardiac-derived EVs were taken up by neurons in the rostral ventrolateral medulla. The administration of cardiac-derived and circulating EVs from CHF rats into the rostral ventrolateral medulla of normal rats evoked an increase in renal sympathetic nerve activity and plasma norepinephrine compared with Sham-operated rats, which were attenuated by exogenously preloading CHF-EVs with antagomirs to Nrf2-targeting miRNAs. CONCLUSIONS: Cardiac microRNA-enriched EVs from animals with CHF can mediate crosstalk between the heart and the brain in the regulation of sympathetic outflow by targeting the Nrf2/antioxidant signaling pathway. This new endocrine signaling pathway regulating sympathetic outflow in CHF may be exploited for novel therapeutics.


Assuntos
Vesículas Extracelulares , Insuficiência Cardíaca , MicroRNAs , Animais , Antagomirs/metabolismo , Antioxidantes/metabolismo , Vesículas Extracelulares/metabolismo , Insuficiência Cardíaca/metabolismo , Humanos , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Norepinefrina/metabolismo , Ratos , Sistema Nervoso Simpático
12.
Mol Ther ; 31(5): 1332-1345, 2023 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-37012704

RESUMO

Repeated use of opioids such as morphine causes changes in the shape and signal transduction pathways of various brain cells, including astrocytes and neurons, resulting in alterations in brain functioning and ultimately leading to opioid use disorder. We previously demonstrated that extracellular vesicle (EV)-induced primary ciliogenesis contributes to the development of morphine tolerance. Herein, we aimed to investigate the underlying mechanisms and potential EV-mediated therapeutic approach to inhibit morphine-mediated primary ciliogenesis. We demonstrated that miRNA cargo in morphine-stimulated-astrocyte-derived EVs (morphine-ADEVs) mediated morphine-induced primary ciliogenesis in astrocytes. CEP97 is a target of miR-106b and is a negative regulator of primary ciliogenesis. Intranasal delivery of ADEVs loaded with anti-miR-106b decreased the expression of miR-106b in astrocytes, inhibited primary ciliogenesis, and prevented the development of tolerance in morphine-administered mice. Furthermore, we confirmed primary ciliogenesis in the astrocytes of opioid abusers. miR-106b-5p in morphine-ADEVs induces primary ciliogenesis via targeting CEP97. Intranasal delivery of ADEVs loaded with anti-miR-106b ameliorates morphine-mediated primary ciliogenesis and prevents morphine tolerance. Our findings bring new insights into the mechanisms underlying primary cilium-mediated morphine tolerance and pave the way for developing ADEV-mediated small RNA delivery strategies for preventing substance use disorders.


Assuntos
Vesículas Extracelulares , MicroRNAs , Camundongos , Animais , Antagomirs/metabolismo , Morfina/farmacologia , Morfina/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Encéfalo/metabolismo , Vesículas Extracelulares/metabolismo
13.
Cell Mol Biol Lett ; 29(1): 55, 2024 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-38643118

RESUMO

BACKGROUND: Viral myocarditis (VMC) is a disease resulting from viral infection, which manifests as inflammation of myocardial cells. Until now, the treatment of VMC is still a great challenge for clinicians. Increasing studies indicate the participation of miR-29b-3p in various diseases. According to the transcriptome sequencing analysis, miR-29b-3p was markedly upregulated in the viral myocarditis model. The purpose of this study was to investigate the role of miR-29b-3p in the progression of VMC. METHODS: We used CVB3 to induce primary cardiomyocytes and mice to establish a model of viral myocarditis. The purity of primary cardiomyocytes was identified by immunofluorescence. The cardiac function of mice was detected by Vevo770 imaging system. The area of inflammatory infiltration in heart tissue was shown by hematoxylin and eosin (H&E) staining. The expression of miR-29b-3p and DNMT3A was detected by quantitative real time polymerase chain reaction (qRT-PCR). The expression of a series of pyroptosis-related proteins was detected by western blot. The role of miR-29b-3p/DNMT3A in CVB3-induced pyroptosis of cardiomyocytes was studied in this research. RESULTS: Our data showed that the expression of miR-29b-3p was upregulated in CVB3-induced cardiomyocytes and heart tissues in mice. To explore the function of miR-29b-3p in CVB3-induced VMC, we conducted in vivo experiments by knocking down the expression of miR-29b-3p using antagomir. We then assessed the effects on mice body weight, histopathology changes, myocardial function, and cell pyroptosis in heart tissues. Additionally, we performed gain/loss-of-function experiments in vitro to measure the levels of pyroptosis in primary cardiomyocytes. Through bioinformatic analysis, we identified DNA methyltransferases 3A (DNMT3A) as a potential target gene of miR-29b-3p. Furthermore, we found that the expression of DNMT3A can be modulated by miR-29b-3p during CVB3 infection. CONCLUSIONS: Our results demonstrate a correlation between the expression of DNMT3A and CVB3-induced pyroptosis in cardiomyocytes. These findings unveil a previously unidentified mechanism by which CVB3 induces cardiac injury through the regulation of miR-29b-3p/DNMT3A-mediated pyroptosis.


Assuntos
MicroRNAs , Miocardite , Animais , Camundongos , Antagomirs/metabolismo , Inflamação/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Miocardite/genética , Miocardite/metabolismo , Miócitos Cardíacos/metabolismo , Piroptose
14.
Nucleic Acids Res ; 50(2): 617-634, 2022 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-34967419

RESUMO

MicroRNAs (miRNAs) are short endogenously expressed RNAs that have the potential to regulate the expression of any RNA. This potential has led to the publication of several thousand papers each year connecting miRNAs to many different genes and human diseases. By contrast, relatively few papers appear that investigate the molecular mechanism used by miRNAs. There is a disconnect between rigorous understanding of mechanism and the extraordinary diversity of reported roles for miRNAs. Consequences of this disconnect include confusion about the assumptions underlying the basic science of human miRNAs and slow development of therapeutics that target miRNAs. Here, we present an overview of investigations into miRNAs and their impact on gene expression. Progress in our understanding of miRNAs would be aided by a greater focus on the mechanism of miRNAs and a higher burden of evidence on researchers who seek to link expression of a particular miRNA to a biological phenotype.


Assuntos
Inativação Gênica , MicroRNAs/genética , Interferência de RNA , Animais , Antagomirs/síntese química , Antagomirs/genética , Antagomirs/uso terapêutico , Pareamento de Bases , Sequência de Bases , Estudos Clínicos como Assunto , Desenvolvimento de Medicamentos , Avaliação Pré-Clínica de Medicamentos , Variação Genética , Humanos , MicroRNAs/síntese química , MicroRNAs/uso terapêutico , Relação Estrutura-Atividade , Resultado do Tratamento
15.
Acta Biochim Biophys Sin (Shanghai) ; 56(4): 607-620, 2024 04 25.
Artigo em Inglês | MEDLINE | ID: mdl-38414350

RESUMO

Myocardial damage is a critical complication and a significant contributor to mortality in sepsis. MicroRNAs (miRNAs) have emerged as key players in sepsis pathogenesis. In this study, we explore the effect and mechanisms of miR-29b-1-5p on sepsis-induced myocardial damage. Sepsis-associated Gene Expression Omnibus datasets (GSE72380 and GSE29914) are examined for differential miRNAs. The mouse sepsis-induced cardiac injury was established by Lipopolysaccharide (LPS) or cecal ligation and puncture (CLP). LPS-treated HL-1 mouse cardiomyocytes simulate myocardial injury in vitro. miR-29b-1-5p is co-upregulated in both datasets and in cardiac tissue from sepsis mouse and HL-1 cell models. miR-29b-1-5p expression downregulation was achieved by antagomir transduction and confirmed by real-time quantitative reverse transcription PCR. Survival analysis and echocardiography examination show that miR-29b-1-5p inhibition improves mice survival cardiac function in LPS- and CLP-induced sepsis mice. Hematoxylin and eosin and Masson's trichrome staining and Immunohistochemistry analysis of mouse myocardial α-smooth muscle actin show that miR-29b-1-5p inhibition reduces myocardial tissue injury and fibrosis. The inflammatory cytokines and cardiac troponin I (cTnI) levels in mouse serum and HL-1 cells are also decreased by miR-29b-1-5p inhibition, as revealed by enzyme-linked immunosorbent assay. The expressions of autophagy-lysosomal pathway-related and apoptosis-related proteins in the mouse cardiac tissues and HL-1 cells are evaluated by western blot analysis. The sepsis-induced activation of the autophagy-lysosomal pathway and apoptosis are also reversed by miR-29b-1-5p antagomir. MTT and flow cytometry measurement further confirm the protective role of miR-29b-1-5p antagomir in HL-1 cells by increasing cell viability and suppressing cell apoptosis. Metascape functionally enriches TargetScan-predicted miR-29b-1-5p target genes. TargetScan prediction and dual luciferase assay validate the targeting relationship between miR-29b-1-5p and telomeric repeat-binding factor 2 (TERF2). The expression and function of TERF2 in HL-1 cells and mice are also evaluated. MiR-29b-1-5p negatively regulates the target gene TERF2. TERF2 knockdown partly restores miR-29b-1-5p antagomir function in LPS-stimulated HL-1 cells. In summary, miR-29b-1-5p targetedly inhibits TERF2, thereby enhancing sepsis-induced myocardial injury.


Assuntos
MicroRNAs , Sepse , Camundongos , Animais , Lipopolissacarídeos/farmacologia , Antagomirs , MicroRNAs/genética , MicroRNAs/metabolismo , Regulação para Baixo , Sepse/complicações , Sepse/genética , Sepse/metabolismo
16.
PLoS Genet ; 17(12): e1009934, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34914716

RESUMO

MicroRNAs (miRNA) are short non-coding RNAs widely implicated in gene regulation. Most metazoan miRNAs utilize the RNase III enzymes Drosha and Dicer for biogenesis. One notable exception is the RNA polymerase II transcription start sites (TSS) miRNAs whose biogenesis does not require Drosha. The functional importance of the TSS-miRNA biogenesis is uncertain. To better understand the function of TSS-miRNAs, we applied a modified Crosslinking, Ligation, and Sequencing of Hybrids on Argonaute (AGO-qCLASH) to identify the targets for TSS-miRNAs in HCT116 colorectal cancer cells with or without DROSHA knockout. We observed that miR-320a hybrids dominate in TSS-miRNA hybrids identified by AGO-qCLASH. Targets for miR-320a are enriched for the eIF2 signaling pathway, a downstream component of the unfolded protein response. Consistently, in miR-320a mimic- and antagomir- transfected cells, differentially expressed gene products are associated with eIF2 signaling. Within the AGO-qCLASH data, we identified the endoplasmic reticulum (ER) chaperone calnexin as a direct miR-320a down-regulated target, thus connecting miR-320a to the unfolded protein response. During ER stress, but not amino acid deprivation, miR-320a up-regulates ATF4, a critical transcription factor for resolving ER stress. In summary, our study investigates the targetome of the TSS-miRNAs in colorectal cancer cells and establishes miR-320a as a regulator of unfolded protein response.


Assuntos
Fator 4 Ativador da Transcrição/genética , Neoplasias Colorretais/genética , MicroRNAs/genética , Ribonuclease III/genética , Antagomirs/genética , Proteínas Argonautas/genética , Calnexina/genética , Movimento Celular/genética , Proliferação de Células/genética , Neoplasias Colorretais/patologia , RNA Helicases DEAD-box/genética , Retículo Endoplasmático/genética , Estresse do Retículo Endoplasmático/genética , Fator de Iniciação 2 em Eucariotos/genética , Técnicas de Inativação de Genes , Células HCT116 , Humanos , Transdução de Sinais/genética , Sítio de Iniciação de Transcrição
17.
Proc Natl Acad Sci U S A ; 118(28)2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34260396

RESUMO

Pseudomonas aeruginosa is an opportunistic pathogen that forms antibiotic-resistant biofilms, which facilitate chronic infections in immunocompromised hosts. We have previously shown that P. aeruginosa secretes outer-membrane vesicles that deliver a small RNA to human airway epithelial cells (AECs), in which it suppresses the innate immune response. Here, we demonstrate that interdomain communication through small RNA-containing membrane vesicles is bidirectional and that microRNAs (miRNAs) in extracellular vesicles (EVs) secreted by human AECs regulate protein expression, antibiotic sensitivity, and biofilm formation by P. aeruginosa Specifically, human EVs deliver miRNA let-7b-5p to P. aeruginosa, which systematically decreases the abundance of proteins essential for biofilm formation, including PpkA and ClpV1-3, and increases the ability of beta-lactam antibiotics to reduce biofilm formation by targeting the beta-lactamase AmpC. Let-7b-5p is bioinformatically predicted to target not only PpkA, ClpV1, and AmpC in P. aeruginosa but also the corresponding orthologs in Burkholderia cenocepacia, another notorious opportunistic lung pathogen, suggesting that the ability of let-7b-5p to reduce biofilm formation and increase beta-lactam sensitivity is not limited to P. aeruginosa Here, we provide direct evidence for transfer of miRNAs in EVs secreted by eukaryotic cells to a prokaryote, resulting in subsequent phenotypic alterations in the prokaryote as a result of this interdomain communication. Since let-7-family miRNAs are in clinical trials to reduce inflammation and because chronic P. aeruginosa lung infections are associated with a hyperinflammatory state, treatment with let-7b-5p and a beta-lactam antibiotic in nanoparticles or EVs may benefit patients with antibiotic-resistant P. aeruginosa infections.


Assuntos
Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Vesículas Extracelulares/metabolismo , MicroRNAs/metabolismo , Pseudomonas aeruginosa/fisiologia , Antagomirs/farmacologia , Aztreonam/farmacologia , Biofilmes/efeitos dos fármacos , Vesículas Extracelulares/efeitos dos fármacos , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Humanos , MicroRNAs/genética , Plâncton/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/isolamento & purificação , beta-Lactamas/farmacologia
18.
Environ Toxicol ; 39(4): 2077-2085, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38100242

RESUMO

Colorectal cancer (CRC) exhibits highly metastatic potential even in the early stages of tumor progression. Gallic acid (GA), a common phenolic compound in plants, is known to possess potent antioxidant and anticancer activities, thereby inducing cell death or cell cycle arrest. However, whether GA reduces the invasiveness of CRC cells without inducing cell death remains unclear. Herein, we aimed to investigate the antimetastatic activity of low-dose GA on CRC cells and determine its underlying mechanism. Cell viability and tumorigenicity were analyzed by MTS, cell adhesion, and colony formation assay. Invasiveness was demonstrated using migration and invasion assays. Changes in protein phosphorylation and expression were assessed by Western blot. The involvement of microRNAs was validated by microarray analysis and anti-miR antagonist. Our findings showed that lower dose of GA (≤100 µM) did not affect cell viability but reduced the capabilities of colony formation, cell adhesion, and invasiveness in CRC cells. Cellularly, GA downregulated the cellular level of integrin αV/ß3, talin-1, and tensin and diminished the phosphorylated FAK, paxillin, Src, and AKT in DLD-1 cells. Microarray results revealed that GA increased miR-1247-3p expression, and pretreatment of anti-miR antagonist against miR-1247-3p restored the GA-reduced integrin αV/ß3 and the GA-inhibited paxillin activation in DLD-1 cells. Consistently, the in vivo xenograft model showed that GA administration inhibited tumor growth and liver metastasis derived from DLD-1 cells. Collectively, our findings indicated that GA inhibited the metastatic capabilities of CRC cells, which may result from the suppression of integrin/FAK axis mediated by miR1247-3p.


Assuntos
Neoplasias Colorretais , MicroRNAs , Humanos , Paxilina/genética , Paxilina/metabolismo , Integrinas/genética , Integrinas/metabolismo , Ácido Gálico/farmacologia , Antagomirs , Integrina alfaV/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias Colorretais/metabolismo , Movimento Celular , Regulação Neoplásica da Expressão Gênica
19.
Int J Mol Sci ; 25(14)2024 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-39062857

RESUMO

The delay in wound healing caused by chronic wounds or pathological scars is a pressing issue in clinical practice, imposing significant economic and psychological burdens on patients. In particular, with the aging of the population and the increasing incidence of diseases such as diabetes, impaired wound healing is one of the growing health problems. MicroRNA (miRNA) plays a crucial role in wound healing and regulates various biological processes. Our results show that miR-618 was significantly upregulated during the inflammatory phase of wound healing.Subsequently, miR-618 promotes the secretion of pro-inflammatory cytokines and regulates the proliferation and migration of keratinocytes. Mechanistically, miR-618 binds to the target gene-Atp11b and inhibits the PI3K-Akt signaling pathway, inhibiting the epithelial-mesenchymal transition (EMT) of keratinocytes. In addition, the PI3K-Akt signaling pathway induces the enrichment of nuclear miR-618, and miR-618 binds to the promoter of Lin7a to regulate gene transcription. Intradermal injection of miR-618 antagomir around full-thickness wounds in peridermal mice effectively accelerates wound closure compared to control. In conclusion, miR-618 antagomir can be a potential therapeutic agent for wound healing.


Assuntos
Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal , Queratinócitos , MicroRNAs , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Cicatrização , MicroRNAs/genética , MicroRNAs/metabolismo , Animais , Queratinócitos/metabolismo , Cicatrização/genética , Camundongos , Movimento Celular/genética , Fosfatidilinositol 3-Quinases/metabolismo , Transição Epitelial-Mesenquimal/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Humanos , Antagomirs/metabolismo , Antagomirs/farmacologia
20.
Int J Mol Sci ; 25(3)2024 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-38338785

RESUMO

Rheumatoid arthritis (RA) is an autoimmune disease that causes inflammation, pain, and ultimately, bone erosion of the joints. The causes of this disease are multifactorial, including genetic factors, such as the presence of the human leukocyte antigen (HLA)-DRB1*04 variant, alterations in the microbiota, or immune factors including increased cytotoxic T lymphocytes (CTLs), neutrophils, or elevated M1 macrophages which, taken together, produce high levels of pro-inflammatory cytokines. In this review, we focused on the function exerted by osteoclasts on osteoblasts and other osteoclasts by means of the release of exosomal microRNAs (miRNAs). Based on a thorough revision, we classified these molecules into three categories according to their function: osteoclast inhibitors (miR-23a, miR-29b, and miR-214), osteoblast inhibitors (miR-22-3p, miR-26a, miR-27a, miR-29a, miR-125b, and miR-146a), and osteoblast enhancers (miR-20a, miR-34a, miR-96, miR-106a, miR-142, miR-199a, miR-324, and miR-486b). Finally, we analyzed potential therapeutic targets of these exosomal miRNAs, such as the use of antagomiRs, blockmiRs, agomiRs and competitive endogenous RNAs (ceRNAs), which are already being tested in murine and ex vivo models of RA. These strategies might have an important role in reestablishing the regulation of osteoclast and osteoblast differentiation making progress in the development of personalized medicine.


Assuntos
Artrite Reumatoide , MicroRNAs , Humanos , Camundongos , Animais , Osteoclastos/patologia , MicroRNAs/genética , Artrite Reumatoide/genética , Artrite Reumatoide/patologia , Osteoblastos/patologia , Macrófagos/patologia , Antagomirs
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