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1.
Invest Ophthalmol Vis Sci ; 63(1): 27, 2022 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-35060996

RESUMO

Purpose: Erianin has been reported to inhibit tumor activity by suppressing the expression of integrins. It is hypothesized that erianin can inhibit retinal neovascularization in collagen by suppressing the expression of integrins. With an aim to test this hypothesis, the regulation of erianin on collagen-mediated retinal angiogenesis via the Ras homolog gene family member A (RhoA)/Rho-associated coiled-coil containing protein kinase 1 (ROCK1) signaling pathway induced by α2 and ß1 integrin-collagen interactions was investigated. Methods: The effects of erianin on human retinal vascular endothelial cells (HRVECs) were assessed in vitro using a hypoxia model in a three-dimensional cell culture induced by cobalt (II) chloride (CoCl2). A hypoxia-induced retinopathy model in adult zebrafish and zebrafish embryos was established to assess the antiangiogenic effect of erianin with and without vitreous collagen in vivo. The expression of α2 and ß1 integrin and RhoA/ROCK1 pathway in HRVECs and zebrafish retinas were analyzed. Results: In vitro, collagen improved the angiogenic potential of HRVECs, including migration, adhesion, and tube formation, in a three-dimensional cell culture model. Erianin suppressed the angiogenic processes of the CoCl2-induced hypoxia HRVEC model in a concentration-dependent manner. In vivo, erianin reduced retinal angiogenesis in the hypoxia-induced retinopathy model in adult and embryo zebrafish. Erianin inhibited the expression of α2 and ß1 integrin and RhoA/ROCK1 in a hypoxia-induced model in vitro in three-dimensional cell culture and in vivo in adult zebrafish. Conclusions: Collagen-mediated retinal angiogenesis may be regulated by erianin via the RhoA/ROCK1 signaling pathway induced by α2 and ß1 integrin-collagen interactions. These findings suggest that erianin has the therapeutic potential on intraocular collagen-mediated retinal angiogenesis.


Assuntos
Bibenzilas/farmacologia , Regulação da Expressão Gênica , Integrina alfa1/genética , Integrina beta1/genética , Proteínas Monoméricas de Ligação ao GTP/genética , Fenol/farmacologia , Neovascularização Retiniana/genética , Proteínas de Peixe-Zebra/genética , Quinases Associadas a rho/genética , Animais , Movimento Celular , Células Cultivadas , Colágeno/metabolismo , Modelos Animais de Doenças , Integrina alfa1/biossíntese , Integrina beta1/biossíntese , Proteínas Monoméricas de Ligação ao GTP/biossíntese , Neovascularização Retiniana/metabolismo , Neovascularização Retiniana/patologia , Transdução de Sinais , Peixe-Zebra , Proteínas de Peixe-Zebra/biossíntese , Quinases Associadas a rho/biossíntese
2.
Invest Ophthalmol Vis Sci ; 63(1): 13, 2022 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-35006271

RESUMO

Purpose: It was previously demonstrated that opticin (OPTC) inhibits the collagen-induced promotion of bioactivities of human retinal vascular endothelial cells (hRVECs). The present in vivo study aimed to further investigate the regulatory role of opticin in vitreous collagen-mediated retinal neovascularization and to elucidate its regulatory mechanisms with regard to integrin α2-I domain-GXXGER complex formation and RhoA/ROCK1 signal change. The regulatory role of Mg2+ on integrin α2-I domain-GXXGER complex formation in the above process was also investigated. Methods: The zebrafish model of hypoxia-induced retinopathy was established, and OPTC-overexpressing plasmids were intravitreally injected to assess the antiangiogenesis effect of opticin. The regulatory role of opticin in integrin α2-I domain-GXXGER complex formation in vivo was analyzed by mass spectrometry. The mRNA and protein expression of RhoA/ROCK1 were examined. The concentration of Mg2+ as an activator of the integrin α2-I domain-GXXGER complex was measured. Solid-phase binding assays were performed to investigate the interference of opticin in integrin α2 collagen binding and the regulatory role of Mg2+ in that process. Results: Opticin and OPTC-overexpressing plasmid injection reduced retinal neovascularization in the zebrafish model of hypoxia-induced retinopathy. Mass spectrometry revealed that opticin could inhibit integrin α2-I domain-GXXGER complex formation. The Mg2+ concentration was also decreased by opticin, which was another indication of the complex activation. Injection of OPTC-overexpressing plasmids inhibited mRNA and the protein expression of RhoA/ROCK1 in the zebrafish model of hypoxia-induced retinopathy. The solid-phase binding assay revealed that opticin could block integrin α2-collagen I binding in the presence of Mg2+. Conclusions: Opticin exerts its antiangiogenesis effect by interfering in the Mg2+-modulated integrin α2-I domain-collagen complex formation and suppressing the downstream RhoA/ ROCK1 signaling pathway.


Assuntos
Regulação da Expressão Gênica , Integrina alfa2/genética , Proteínas Monoméricas de Ligação ao GTP/genética , Proteoglicanas/genética , Neovascularização Retiniana/genética , Proteínas de Peixe-Zebra/genética , Quinases Associadas a rho/genética , Animais , Modelos Animais de Doenças , Integrina alfa2/biossíntese , Proteínas Monoméricas de Ligação ao GTP/biossíntese , Proteoglicanas/biossíntese , RNA/genética , Neovascularização Retiniana/metabolismo , Neovascularização Retiniana/patologia , Transdução de Sinais , Peixe-Zebra , Proteínas de Peixe-Zebra/biossíntese , Quinases Associadas a rho/biossíntese
3.
Respir Res ; 22(1): 248, 2021 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-34537072

RESUMO

BACKGROUND: Chemoresistance limits the therapeutic effect of cisplatin (DDP) on non-small cell lung cancer (NSCLC). Circular RNAs (circRNAs) function as important regulators in chemoresistance. This study aimed to explore the regulation of circRNA Phosphatidylinositol-4-Phosphate 5-Kinase Type 1 Alpha (circ_PIP5K1A) in DDP resistance. METHODS: The expression analysis of circ_PIP5K1A, micoRNA-493-5p (miR-493-5p) and Rho Associated Coiled-Coil Containing Protein Kinase 1 (ROCK1) was conducted through reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Cell sensitivity was determined using 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyl tetrazolium bromide (MTT) assay. Cell proliferation and cell viability were evaluated by colony formation assay and MTT assay, respectively. Cell cycle and apoptosis detection was performed via flow cytometry. Cell motility was examined by transwell migration or invasion assay. Dual-luciferase reporter assay was applied to confirm the target binding. ROCK1 protein level was assayed via Western blot. In vivo assay was carried out using xenograft model in mice. RESULTS: Circ_PIP5K1A level was abnormally increased in DDP-resistant NSCLC tissues and cells. Silencing circ_PIP5K1A reduced DDP resistance, proliferation, cell cycle progression and cell motility in DDP-resistant NSCLC cells. Circ_PIP5K1A directly interacted with miR-493-5p in NSCLC cells. The function of circ_PIP5K1A was dependent on the negative regulation of miR-493-5p. MiR-493-5p directly targeted ROCK1 and circ_PIP5K1A regulated the ROCK1 level via acting as a sponge of miR-493-5p. Overexpression of miR-493-5p inhibited chemoresistance and cancer progression by downregulating ROCK1 expression in DDP-resistant NSCLC cells. Circ_PIP5K1A regulated DDP sensitivity in vivo via the miR-493-5p/ROCK1 axis. CONCLUSION: These findings suggested that circ_PIP5K1A upregulated the ROCK1 expression to promote DDP resistance and cancer progression in NSCLC by sponging miR-493-5p.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Cisplatino/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Pulmonares/metabolismo , MicroRNAs/biossíntese , Fosfotransferases (Aceptor do Grupo Álcool)/biossíntese , Quinases Associadas a rho/biossíntese , Células A549 , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Cisplatino/farmacologia , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/fisiologia , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , RNA Circular/biossíntese , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
4.
Respir Res ; 22(1): 250, 2021 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-34548087

RESUMO

BACKGROUND: Mechanical ventilation can induce or aggravate lung injury, which is termed ventilator-induced lung injury (VILI). Piezo1 is a key element of the mechanotransduction process and can transduce mechanical signals into biological signals by mediating Ca2+ influx, which in turn regulates cytoskeletal remodeling and stress alterations. We hypothesized that it plays an important role in the occurrence of VILI, and investigated the underlying mechanisms. METHODS: High tidal volume mechanical ventilation and high magnitude cyclic stretch were performed on Sprague-Dawley rats, and A549 and human pulmonary microvascular endothelial cells, respectively, to establish VILI models. Immunohistochemical staining, flow cytometry, histological examination, enzyme-linked immunosorbent assay, western blotting, quantitative real-time polymerase chain reaction and survival curves were used to assess the effect of Piezo1 on induction of lung injury, as well as the signaling pathways involved. RESULTS: We observed that Piezo1 expression increased in the lungs after high tidal volume mechanical ventilation and in cyclic stretch-treated cells. Mechanistically, we observed the enhanced expression of RhoA/ROCK1 in both cyclic stretch and Yoda1-treated cells, while the deficiency or inhibition of Piezo1 dramatically antagonized RhoA/ROCK1 expression. Furthermore, blockade of RhoA/ROCK1 signaling using an inhibitor did not affect Piezo1 expression. GSMTx4 was used to inhibit Piezo1, which alleviated VILI-induced pathologic changes, water content and protein leakage in the lungs, and the induction of systemic inflammatory mediators, and improved the 7-day mortality rate in the model rats. CONCLUSIONS: These findings indicate that Piezo1 affects the development and progression of VILI through promotion of RhoA/ROCK1 signaling.


Assuntos
Mecanotransdução Celular/fisiologia , Proteínas de Membrana/biossíntese , Respiração Artificial/efeitos adversos , Lesão Pulmonar Induzida por Ventilação Mecânica/metabolismo , Proteínas rho de Ligação ao GTP/biossíntese , Quinases Associadas a rho/biossíntese , Células A549 , Animais , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Volume de Ventilação Pulmonar/fisiologia , Lesão Pulmonar Induzida por Ventilação Mecânica/patologia
5.
Mol Neurobiol ; 58(11): 6032-6048, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34436749

RESUMO

Long non-coding RNAs (lncRNAs) play a key role in a variety of disease processes. Plasmacytoma variant translocation 1 (PVT1), a lncRNA, is known to regulate cell functions and play a key role in the pathogenesis of many malignant tumors. The function and molecular mechanisms of lncRNA-PVT1 in cerebral ischemia remain unknown. Real-time PCR (qRT-PCR) was used to detect lncRNA-PVT1 and microRNA-30c-5p (miR-30c-5p) expression in the brain tissues of mice underwent middle cerebral artery occlusion/reperfusion (MCAO/R) and oxygen-glucose deprivation/reperfusion (OGD/R)-treated mouse primary brain neurons. Gain- or loss-of-function approaches were used to manipulate PVT1, miR-30c-5p, and Rho-associated protein kinase 2 (Rock2). The mechanism of PVT1 in ischemic stroke was evaluated both in vivo and in vitro via bioinformatics analysis, CCK-8, flow cytometry, TUNEL staining, luciferase activity assay, RNA FISH, and Western blot. PVT1 was upregulated in the brain tissues of mice treated with MCAO/R and primary cerebral cortex neurons of mice treated with OGD/R. Mechanistically, PVT1 knockdown resulted in a lower infarct volume and ameliorated neurobehavior in MCAO mice. Consistent with in vivo results, PVT1 upregulation significantly decreased the viability and induced apoptosis of neurons cultured in OGD/R. Moreover, we demonstrated that PVT1 acts as a competitive endogenous RNA (ceRNA) that competes with miR-30c-5p, thereby negatively regulating its endogenous target Rock2. Overexpression of miR-30c-5p significantly promoted cell proliferation and inhibited apoptosis. Meanwhile, PVT1 was confirmed to target miR-30c-5p, thus activating Rock2 expression, which finally led to the activation of MAPK signaling. We demonstrated that PVT1, as a ceRNA of miR-30c-5p, could target and regulate the level of Rock2, which aggravates cerebral I/R injury via activation of the MAPK pathway. These findings reveal a new function of PVT1, which helps to broadly understand cerebral ischemic stroke and provide a new treatment strategy for this disease.


Assuntos
Infarto da Artéria Cerebral Média/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , RNA Longo não Codificante/fisiologia , Traumatismo por Reperfusão/metabolismo , Quinases Associadas a rho/biossíntese , Animais , Apoptose , Divisão Celular , Hipóxia Celular , Células Cultivadas , Epigênese Genética , Mutação com Ganho de Função , Técnicas de Silenciamento de Genes , Glucose/farmacologia , Infarto da Artéria Cerebral Média/genética , Mutação com Perda de Função , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Oxigênio/farmacologia , Interferência de RNA , RNA Longo não Codificante/genética , RNA Interferente Pequeno/genética , Traumatismo por Reperfusão/genética , Frações Subcelulares/química , Regulação para Cima , Quinases Associadas a rho/genética
6.
Neuroreport ; 32(10): 888-893, 2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34050115

RESUMO

BACKGROUND: This study aimed to investigate the role of lncRNA RP11-390F4.3 in glioblastoma. METHODS: The expression levels of RP11-390F4.3, miR-148a and ROCK1 in glioblastoma and nontumor tissues were measured by performing quantitative PCR (qPCR) and data were compared using paired t test. Linear regression analysis was performed to analyze the correlations between RP11-390F4.3 and miR-148a/ROCK1 in glioblastoma tissues. The effects of overexpression of RP11-390F4.3, miR-148a and ROCK1 on U-373 MG cell invasion and migration were analyzed by Transwell assay. RESULTS: RP11-390F4.3 and ROCK1 were both upregulated in glioblastoma, while miR-148a was downregulated in glioblastoma. In glioblastoma, RP11-390F4.3 was positively correlated with ROCK1 but negatively correlated with miR-148a. In glioblastoma cells, overexpression of RP11-390F4.3 led to upregulated ROCK1 and downregulated miR-148a. Cell invasion and migration analysis showed that overexpression of RP11-390F4.3 and ROCK1 resulted in increased, and overexpression of miR-148a resulted in deceased invasion and migration rates of glioblastoma cells. CONCLUSION: Therefore, RP11-390F4.3 may upregulate ROCK1 by downregulating miR-148a to promote glioblastoma cell invasion and migration.


Assuntos
Neoplasias Encefálicas/metabolismo , Movimento Celular/fisiologia , Regulação Neoplásica da Expressão Gênica , Glioblastoma/metabolismo , RNA Longo não Codificante/biossíntese , Quinases Associadas a rho/biossíntese , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Glioblastoma/genética , Glioblastoma/patologia , Humanos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , RNA Longo não Codificante/genética , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/genética
7.
J Drug Target ; 29(5): 531-540, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33307856

RESUMO

OBJECTIVE: It is believed that microRNAs (miRNAs) participate in the pathogenesis of Alzheimer's disease (AD), but the specified function of miR-10b-5p in the disease has not been thoroughly understood. Thereafter, this research aimed to assess the function of miR-10b-5p in AD. METHODS: Rat AD models were established by injected with amyloid-ß1-42 (Aß1-42), which were mainly treated with lentivirus-miR-10b-5p inhibitor, or lentivirus-overexpressed homeobox D10 (HOXD10). MiR-10b-5p, HOXD10, RhoA, ROCK1 and ROCK2 expression in rat hippocampal tissues were determined. Afterwards, the behaviour of rats was tested, and neuronal apoptosis, pathological injury, and inflammatory factors and oxidative stress-related factors were all assessed. Finally, the target relation between miR-10b-5p and HOXD10 was detected. RESULTS: MiR-10b-5p was upregulated while HOXD10 was downregulated, and the Rho/ROCK signalling pathway was activated in hippocampal tissues of rats with AD. Inhibition of miR-10b-5p could attenuate the neuronal apoptosis, pathological injury, inflammation reaction, and oxidative stress by elevating HOXD10 and inhibiting the Rho/ROCK signalling pathway in AD rats. Moreover, HOXD10 was targeted by miR-10b-5p. CONCLUSION: Inhibited miR-10b-5p decelerated the development of AD by promoting HOXD10 and inactivating the Rho/ROCK signalling pathway, and our findings may contribute to the exploration of AD treatment.


Assuntos
Doença de Alzheimer/metabolismo , Proteínas de Homeodomínio/biossíntese , MicroRNAs/biossíntese , Fatores de Transcrição/biossíntese , Quinases Associadas a rho/biossíntese , Doença de Alzheimer/induzido quimicamente , Doença de Alzheimer/prevenção & controle , Peptídeos beta-Amiloides/toxicidade , Animais , MicroRNAs/antagonistas & inibidores , Fragmentos de Peptídeos/toxicidade , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Regulação para Cima/fisiologia , Quinases Associadas a rho/antagonistas & inibidores
8.
J Cell Mol Med ; 24(18): 10855-10865, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32776418

RESUMO

Curcumin treatment was reported to delay the progression of OA, but its underlying mechanism remains unclear. In this study, we aimed to investigate the molecular mechanism underlying the role of curcumin in OA treatment. Accordingly, by conducting MTT and flow cytometry assays, we found that the exosomes derived from curcumin-treated MSCs helped to maintain the viability while inhibiting the apoptosis of model OA cells. Additionally, quantitative real-time PCR and Western blot assays showed that the exosomes derived from curcumin-treated MSCs significantly restored the down-regulated miR-143 and miR-124 expression as well as up-regulated NF-kB and ROCK1 expression in OA cells. Mechanistically, curcumin treatment decreased the DNA methylation of miR-143 and miR-124 promoters. In addition, the 3' UTRs of NF-kB and ROCK1 were proven to contain the binding sites for miR-143 and miR-124, respectively. Therefore, the up-regulation of miR-143 and miR-124 in cellular and mouse OA models treated with exosomes remarkably restored the normal expression of NF-kB and ROCK1. Consequently, the progression of OA was attenuated by the exosomes. Our results clarified the molecular mechanism underlying the therapeutic role of MSC-derived exosomes in OA treatment.


Assuntos
Curcumina/farmacologia , Exossomos/fisiologia , Osteoartrite/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose , Células Cultivadas , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Curcumina/uso terapêutico , Metilação de DNA/efeitos dos fármacos , Exossomos/química , Vetores Genéticos , Humanos , Interleucina-1beta/farmacologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Camundongos , MicroRNAs/biossíntese , MicroRNAs/genética , NF-kappa B/biossíntese , NF-kappa B/genética , Osteoartrite/metabolismo , RNA/metabolismo , Receptor Toll-Like 9/biossíntese , Receptor Toll-Like 9/genética , Quinases Associadas a rho/biossíntese , Quinases Associadas a rho/genética
9.
J Cell Mol Med ; 24(18): 10693-10704, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32725958

RESUMO

Endometriosis is a benign gynaecological disease appearing with pelvic pain, rising dysmenorrhoea and infertility seriously impacting on 10% of reproductive-age females. This research attempts to demonstrate the function and molecular mechanism of RhoA/ROCK pathway on epithelial-mesenchymal transition (EMT) and proliferation in endometriosis. The expression of Rho family was abnormally changed in endometriotic lesions; in particular, RhoA and ROCK1/2 were significantly elevated. Overexpression of RhoA in human eutopic endometrial epithelial cells (eutopic EECs) enhanced the cell mobility, epithelial-mesenchymal transition (EMT) and proliferation, and RhoA knockdown exhibited the opposite function. Oestrogen up-regulated the RhoA activity and expression of RhoA and ROCK1/2. RhoA overexpression reinforced the effect of oestrogen on promoting EMT and proliferation, and RhoA knockdown impaired the effect of oestrogen. oestrogen receptor α (ERα) was involved with the regulation of oestrogen on EMT and proliferation and up-regulated RhoA activity and expression of RhoA and ROCK1/2. The function of ERα was modulated by the change in RhoA expression. Furthermore, phosphorylated ERK that was enhanced by oestrogen and ERα promoted the protein expression of RhoA/ROCK pathway. Endometriosis mouse model revealed that oestrogen enhanced the size and weight of endometriotic lesions. The expression of RhoA and phosphorylated ERK in mouse endometriotic lesions was significantly elevated by oestrogen. We conclude that abnormal activated RhoA/ROCK pathway in endometriosis is responsible for the function of oestrogen/ERα/ERK signalling, which promoted EMT and proliferation and resulted in the development of endometriosis.


Assuntos
Endometriose/patologia , Endométrio/patologia , Transição Epitelial-Mesenquimal/fisiologia , Estrogênios/fisiologia , Transdução de Sinais/fisiologia , Quinases Associadas a rho/fisiologia , Proteína rhoA de Ligação ao GTP/fisiologia , Adulto , Animais , Células Cultivadas , Modelos Animais de Doenças , Endometriose/cirurgia , Endométrio/efeitos dos fármacos , Endométrio/transplante , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/efeitos dos fármacos , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/fisiologia , Feminino , Regulação da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Cistos Ovarianos/etiologia , Cistos Ovarianos/cirurgia , Interferência de RNA , RNA Mensageiro/biossíntese , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Proteínas Recombinantes/efeitos dos fármacos , Proteínas Recombinantes/metabolismo , Transdução de Sinais/efeitos dos fármacos , Quinases Associadas a rho/biossíntese , Quinases Associadas a rho/genética , Proteína rhoA de Ligação ao GTP/antagonistas & inibidores , Proteína rhoA de Ligação ao GTP/biossíntese , Proteína rhoA de Ligação ao GTP/genética
10.
Inflammation ; 43(4): 1476-1487, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32240450

RESUMO

Sanggenon C (SC), a natural flavonoid extracted from Cortex Mori (Sang Bai Pi), is reported to possess anti-inflammatory and antioxidant properties in hypoxia. The present study aimed to investigate the therapeutic potential and the underlying mechanisms of SC in cerebral ischemia-reperfusion (I/R) injury. A rat model of reversible middle cerebral artery occlusion (MCAO) was used to induce cerebral I/R injury in vivo, and SC was administrated intragastrically. Brain injuries were evaluated using Bederson scores, brain water content, and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. The levels of inflammatory factors and oxidative stress were examined using corresponding kits. Cell apoptosis was evaluated by TUNEL. Moreover, the expressions of apoptosis-related and RhoA/ROCK signaling-related proteins were detected through western blotting. In vitro, RhoA was overexpressed in oxygen-glucose deprivation and reperfusion (OGD/R)-induced PC12 cells to confirm the contribution of RhoA-ROCK signaling inhibition by SC to the neuroprotective effects post OGD/R. Pretreatment with SC significantly ameliorated the neurologic impairment, brain edema, and cerebral infarction post MCAO-reperfusion, associated with reductions of inflammation, oxidative stress, and cell apoptosis in the brain. Furthermore, SC remarkably downregulated the expression of RhoA/ROCK signaling-related proteins post MCAO-reperfusion in rats, while overexpression of RhoA reversed the beneficial effects of SC on protecting against inflammation and oxidative stress in OGD/R-induced PC12 cells. Taken together, these findings demonstrated that SC exerts neuroprotective effects after cerebral I/R injury via inhibiting inflammation and oxidative stress through regulating RhoA-ROCK signaling, suggesting a therapeutic potential of SC in cerebral I/R injury.


Assuntos
Anti-Inflamatórios/uso terapêutico , Benzofuranos/uso terapêutico , Isquemia Encefálica/tratamento farmacológico , Cromonas/uso terapêutico , Estresse Oxidativo/efeitos dos fármacos , Traumatismo por Reperfusão/tratamento farmacológico , Proteínas rho de Ligação ao GTP/antagonistas & inibidores , Quinases Associadas a rho/antagonistas & inibidores , Animais , Anti-Inflamatórios/farmacologia , Benzofuranos/farmacologia , Isquemia Encefálica/metabolismo , Cromonas/farmacologia , Relação Dose-Resposta a Droga , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Masculino , Estresse Oxidativo/fisiologia , Células PC12 , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/metabolismo , Proteínas rho de Ligação ao GTP/biossíntese , Quinases Associadas a rho/biossíntese
11.
Pharmacol Rep ; 72(4): 903-911, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32144744

RESUMO

BACKGROUND: Chronic heart failure (CHF) is characterized by left ventricular dysfunction and altered autonomic control of cardiac function. This study aimed to investigate the effects of atorvastatin on left ventricular remodeling (LVR) and cardiac function in rats with isoproterenol-induced CHF and the possible mechanism. METHODS: An isoproterenol-induced CHF model was established in rata, which were subsequently treated with atorvastatin. Echocardiography, hemodynamic, and left ventricular mass indexes were assessed. The mRNA expression of RhoA, Rho kinase, and endothelial nitric oxide synthase (eNOS) was determined by RT-qPCR. The protein expression of myosin-binding subunit (MBS), MBS-P, eNOS, phosphorylated-eNOS, RhoA, and Rho kinase was measured by Western blot analysis. The relative activity of NADPH oxidase, ROS, and NO was assessed by ELISA. RESULTS: Isoproterenol-induced CHF rats treated with atorvastatin exhibited decreased left ventricular end-systolic dimension, left ventricular end-diastolic dimension, left ventricular end-diastolic pressure, left ventricular mass index, maximum fall rate of change in left ventricular pressure, heart rate (p < 0.001), expression of RhoA, Rho kinase, MBS and MBS-P (p < 0.01), and relative activity of NADPH oxidase, ROS and NO (p < 0.05) and increased left ventricular short axis fractional shortening, left ventricular end-systolic pressure, maximum rise rate of change in left ventricular pressure (p < 0.001) and expression of eNOS, and phosphorylated-eNOS ser1177 (all p < 0.05) compared with those of rats with isoproterenol-induced CHF. CONCLUSION: We demonstrated that atorvastatin inhibits LVR and improves cardiac function in rats with isoproterenol-induced CHF through inhibition of the RhoA/Rho kinase signaling pathway.


Assuntos
Atorvastatina/uso terapêutico , Insuficiência Cardíaca/tratamento farmacológico , Isoproterenol/toxicidade , Remodelação Ventricular/efeitos dos fármacos , Proteínas rho de Ligação ao GTP/antagonistas & inibidores , Quinases Associadas a rho/antagonistas & inibidores , Animais , Atorvastatina/farmacologia , Cardiotônicos/toxicidade , Doença Crônica , Ecocardiografia/métodos , Insuficiência Cardíaca/induzido quimicamente , Insuficiência Cardíaca/diagnóstico por imagem , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Masculino , Óxido Nítrico Sintase Tipo III/biossíntese , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Remodelação Ventricular/fisiologia , Proteínas rho de Ligação ao GTP/biossíntese , Quinases Associadas a rho/biossíntese
12.
J Neuroimmunol ; 341: 577172, 2020 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-32028123

RESUMO

Multiple sclerosis (MS) is a neurodegenerative disease of the central nervous system. The animal model of MS, experimental autoimmune encephalomyelitis (EAE), is commonly used for studies of human inflammatory demyelinating diseases and has been shown to be suitable for studying the effects of exercise on MS pathophysiology. The present study was conducted to determine the impact of forced swimming and voluntary running wheel exercises before and after the induction of EAE on expression of Nogo-A, NgR, and ROCK genes in the brain tissue. A total of 96 C57BL/6 mice were randomly divided into two groups, namely exercises before (EXb, n = 48) and after (EXa, n = 48) induction of EAE. Each group was divided into four subgroups: Forced Swimming + EAE (n = 12), Voluntary Running Wheel + EAE (n = 12), NoEX-EAE (n = 12), and Control group (n = 12). Animals performed either swimming exercise for 30 min per day or running wheel for one hour per day, five days per week for four weeks. Results of Luxal Fast Blue (LFB) staining demonstrated that the degree of demyelination was significantly less in the experimental exercised compared to NoEX-EAE groups (P < .05). Amazingly, both modes of exercise reduced the severity of MS symptoms in mice exposed to swimming and wheel running, evaluated as body weight, clinical scores, degree of demyelination, and gene expressions, regardless of whether the exercise was performed before or after EAE induction.


Assuntos
Encéfalo/metabolismo , Encefalomielite Autoimune Experimental/terapia , Terapia por Exercício , Esclerose Múltipla/terapia , Proteínas do Tecido Nervoso/biossíntese , Condicionamento Físico Animal , Animais , Peso Corporal , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/metabolismo , Feminino , Regulação da Expressão Gênica , Camundongos , Camundongos Endogâmicos C57BL , Bainha de Mielina/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas Nogo/biossíntese , Proteínas Nogo/genética , Receptor Nogo 1/biossíntese , Receptor Nogo 1/genética , Distribuição Aleatória , Reação em Cadeia da Polimerase em Tempo Real , Corrida , Natação , Volição , Quinases Associadas a rho/biossíntese , Quinases Associadas a rho/genética
13.
Mol Med Rep ; 21(3): 1304-1309, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31922232

RESUMO

Long noncoding (lnc)RNA HAND2­AS1 inhibits the development of several human malignancies. The role of HAND2­AS1 was investigated in hepatocellular carcinoma (HCC). It was found that levels of HAND2­AS1 in serum were significantly lower, while serum levels of Rho­associated protein kinase 2 (ROCK2) in HCC patients were significantly increased compared with hepatitis B (HB) patients and healthy controls. Decreased HAND2­AS1 levels distinguished HCC patients but not HB patients from healthy controls. A significant negative correlation between HAND2­AS1 and ROCK2 was found in HCC patients but not in HB patients or healthy controls. HAND2­AS1 overexpression inhibited, while ROCK2 overexpression promoted HCC cell migration, proliferation and invasion. HAND2­AS1 overexpression led to downregulated ROCK2 expression. ROCK2 overexpression did not significantly affect ROCK2 expression but attenuated the inhibitory effects of HAND2­AS1 overexpression. It was therefore concluded that HAND2­AS1 might mediate the downregulation of ROCK2 in HCC to inhibit cancer cell migration, proliferation and invasion.


Assuntos
Carcinoma Hepatocelular/metabolismo , Movimento Celular , Proliferação de Células , Regulação para Baixo , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/biossíntese , RNA Longo não Codificante/metabolismo , RNA Neoplásico/metabolismo , Quinases Associadas a rho/biossíntese , Adulto , Idoso , Carcinoma Hepatocelular/patologia , Feminino , Humanos , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica
14.
Pathol Oncol Res ; 26(2): 877-883, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30864107

RESUMO

Osteosarcoma (OS) is the most common primary malignant bone tumor with two peaks of incidence, in early adolescence and the elderly. Patients affected with this malignancy often present metastatic disease at diagnosis, and despite multimodality therapy, survival has not improved substantially over the past 3 decades. Recently, miR-138-5p, proposed as a crucial intracellular mediator of invasion, has been recognized to target the Rho-associated coiled-coil containing protein kinase 2 (ROCK2). Dysregulation of ROCK1 and ROCK2 was also described in OS, being associated to higher metastasis incidence and worse prognosis. Nonetheless, the specific roles of miR-138-5p in pediatric and young adult OS and its ability to modulate these kinases remain to be established. Thus, in the present study, the expression levels miR-138-5p were evaluated in a consecutive cohort of exclusively pediatric and young adult primary OS samples. In contrast to previous reports that included adult tissues, our results showed upregulation of miR-138-5p associated with reduced event-free survival and relapsed cases. In parallel, ROCK1 mRNA levels were significantly reduced in tumor samples and negatively correlated with miR-138-5p. Similar correlations were observed after studying the profiles of ROCK1 and ROCK2 by immunohistochemistry. Our data present miR-138-5p as a consistent prognostic factor in pediatric and young adult OS, reinforcing its participation in the post-transcriptional regulation of ROCK kinases.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Ósseas/patologia , MicroRNAs/genética , Osteossarcoma/patologia , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Prognóstico , Adulto Jovem , Quinases Associadas a rho/biossíntese
15.
Stem Cell Res ; 41: 101614, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31715427

RESUMO

Pluripotency of mouse embryonic stem cells is regulated by transcription factor regulatory networks as well as mechanical stimuli sensed by the cells. It has been unclear how the mechanical strain applied to the plasma membrane is transferred to the nucleus in mouse embryonic stem cells (mESCs). We here investigated the machinery of the mechanotransduction based on the finding that spontaneous differentiation of mESCs was inhibited with the downregulation of ROCK2 in cells attached to soft substrates. On examining the effects of actin bindings to both focal adhesions and cell junctions in cells on soft substrates, co-localization of actin filaments and α-catenin, which links actin to E-cadherin, decreased after differentiation induction. Also, disrupting actin-nucleus mechanical link through dominant negative assay of Nesprins helps to sustain the pluripotency genes; thus, revealing that mechanical strain relayed by actin-Nesprin connection is required for the initiation of the differentiation process.


Assuntos
Citoesqueleto de Actina/patologia , Diferenciação Celular , Núcleo Celular/metabolismo , Células-Tronco Embrionárias Murinas/metabolismo , Animais , Caderinas/metabolismo , Linhagem Celular , Regulação Enzimológica da Expressão Gênica , Camundongos , Células-Tronco Embrionárias Murinas/citologia , alfa Catenina/metabolismo , Quinases Associadas a rho/biossíntese
16.
J Alzheimers Dis ; 72(3): 815-822, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31658060

RESUMO

Upregulation of Rho-associated protein kinase 2 (ROCK2) hallmarks the progression of neurodegenerative diseases. However, the molecular mechanisms underlying the regulation of ROCK2 expression are not clear and thus addressed in the current study. We generated a subacute model of Parkinson's disease in mice with a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) method. The MPTP model was validated by impaired rotational behavior of the mice upon apomorphine exposure, astrocytic activation, and reduction in tyrosine-hydroxylase-positive neurons in the mouse striatum. Moreover, MPTP induced increases in ROCK2 protein but not in ROCK2 mRNA. Further analysis showed that MPTP inhibited the expression of microRNA-291 (miR-291), which suppressed ROCK2 mRNA via 3'-UTR-binding in neuronal cells to increase ROCK2 protein. Intracranial injection of miR-291 four days before MPTP alleviated the impaired rotational behavior of the mice upon apomorphine exposure, MPTP-induced astrocytic activation, and the reduction in tyrosine-hydroxylase-positive neurons in the mouse striatum. Together, these data suggest that miR-291 has a protective role in neurodegeneration, likely through regulation of ROCK2.


Assuntos
Epigênese Genética/fisiologia , MicroRNAs/biossíntese , Transtornos Parkinsonianos/metabolismo , Quinases Associadas a rho/biossíntese , Animais , Linhagem Celular , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismo , Transtornos Parkinsonianos/genética , Quinases Associadas a rho/genética
17.
Respir Res ; 20(1): 225, 2019 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-31638991

RESUMO

BACKGROUND: Significant evidence has shown that the miRNA pathway is an important component in the downstream signaling cascades of TGF-ß1 pathway. Our previous study has indicated that miR-335-5p expression was significantly down-regulated and acted as a vital player in the metastasis of non-small cell lung cancer (NSCLC), however the underlying mechanism remained unclear. METHODS: The differential expression level of miR-335-5p and ROCK1 were determined by qRT-PCR and IHC analysis in human tissue samples with or without lymph node metastasis. Transwell assay was conducted to determine cell ability of migration and invasion. SiRNA interference, microRNA transfection and western blot analysis were utilized to clarify the underlying regulatory mechanism. RESULTS: We showed that down-regulated expression of miR-335-5p and up-regulated expression of ROCK1 in NSCLC tissues were associated with lymph node metastasis. Over-expresion of miR-335-5p significantly inhibited TGF-ß1-mediated NSCLC migration and invasion. Furthermore, luciferase reporter assays proved that miR-335-5p can bind to 3'-UTR of ROCK1 directly. Moreover, we confirmed that siRNA-mediated silencing of ROCK1 significantly diminished TGF-ß1-mediated EMT and migratory and invasive capabilities of A549 and SPC-A1 cells. CONCLUSION: This is the first time to report that miR-335-5p regulates ROCK1 and impairs its functions, thereby playing a key role in TGF-ß1-induced EMT and cell migration and invasion in NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Neoplasias Pulmonares/metabolismo , MicroRNAs/biossíntese , Fator de Crescimento Transformador beta1/farmacologia , Quinases Associadas a rho/biossíntese , Células A549 , Carcinoma Pulmonar de Células não Pequenas/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Neoplasias Pulmonares/genética , MicroRNAs/genética , Quinases Associadas a rho/genética
18.
Mol Med Rep ; 20(4): 3440-3447, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31432120

RESUMO

Abnormal expression of microRNAs (miRNAs/miRs) has been previously reported in various types of human cancer, such as retinoblastoma (RB). Dysregulated miRNAs have been demonstrated to be important epigenetic regulators of numerous biological events associated with RB. Therefore, improved understanding of the precise roles of miRNAs in RB is required to develop novel therapeutic strategies for the treatment of patients with this disease. In the present study, reverse transcription­quantitative polymerase chain reaction (RT­qPCR) was performed to detect miR­330 expression in RB tissues and cell lines. The effects of miR­330 overexpression on the viability and invasion of RB cells were determined using MTT and Matrigel®­based invasion assays, respectively. The mechanisms underlying the activity of miR­330 in RB cells were investigated via bioinformatics analysis, luciferase reporter assays, and RT­qPCR and western blot analyses. It was revealed that the levels of miR­330 expression were significantly downregulated in RB tissues and cell lines compared with in control healthy tissues and cells, respectively. Overexpression of miR­330 in RB cells significantly reduced the viability and invasion of cells in vitro. Additionally, ρ­associated coiled­coil containing protein kinase 1 (ROCK1) was identified as a putative target of miR­330 using bioinformatics analysis. Subsequent experiments revealed that miR­330 interacted with the 3'­untranslated region of ROCK1 and downregulated its expression in RB cells. Furthermore, the expression levels of ROCK1 were increased in RB tissues compared with healthy controls and negatively correlated with miR­330 expression. Finally, upregulation of ROCK1 expression reversed the miR­330­induced inhibition of the viability and invasion of RB cells. Collectively, these results suggested that miR­330 exhibits tumor­suppressor activity in the development of RB by directly targeting ROCK1, indicating that restoration of miR­330 expression may be a promising therapeutic technique in the treatment of patients with RB.


Assuntos
Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , MicroRNAs/biossíntese , Proteínas de Neoplasias/biossíntese , RNA Neoplásico/biossíntese , Retinoblastoma/metabolismo , Quinases Associadas a rho/biossíntese , Adolescente , Adulto , Linhagem Celular Tumoral , Sobrevivência Celular , Feminino , Humanos , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteínas de Neoplasias/genética , RNA Neoplásico/genética , Retinoblastoma/genética , Retinoblastoma/patologia , Quinases Associadas a rho/genética
19.
Neurosci Lett ; 699: 177-183, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-30753912

RESUMO

Axon regeneration after cerebral ischemia in mammals is inadequate to restore function, illustrating the need to design better strategies for improving outcomes. Improvement of axon regeneration has been achieved through fastigial nucleus electrostimulation (FNS) in animal researches. However, the mechanisms underlying this neuroprotection remain poorly understood. Increasing the levels of the second messenger cyclic AMP (cAMP) enhances axon regeneration, making it an excellent candidate molecule that has therapeutic potential. In the present study, we examined the expression of cAMP signaling in ischemic brain tissues following focal cerebral ischemia. Adult rats were subjected to ischemia induced by middle cerebral artery occlusion (MCAO). A dipolar electrode was placed into the cerebellum to stimulate the cerebellar fastigial nucleus for 1 h after ischemia. Neurological deficits and the expressions of cAMP, PKA (protein kinase A) and ROCK (Rho-kinase) were determined. Axonal regeneration was measured by upregulation of growth-associated protein 43 (GAP43). The data indicated that FNS significantly enhanced axonal regeneration and motor function recovery after cerebral ischemia. FNS also significantly increased cAMP and PKA levels after ischemic brain injury. All the beneficial effects of FNS were blocked by Rp-cAMP, an antagonist of PKA. Our research suggested that the axonal regeneration conferred by FNS was likely achieved via the regulation of cAMP/PKA pathway.


Assuntos
Núcleos Cerebelares/fisiologia , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Terapia por Estimulação Elétrica , Infarto da Artéria Cerebral Média/terapia , Regeneração Nervosa , Transdução de Sinais , Animais , Núcleos Cerebelares/efeitos dos fármacos , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacologia , Proteína GAP-43/metabolismo , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/patologia , Masculino , Regeneração Nervosa/efeitos dos fármacos , Ratos , Recuperação de Função Fisiológica/efeitos dos fármacos , Tionucleotídeos/farmacologia , Regulação para Cima , Quinases Associadas a rho/biossíntese
20.
J Cardiothorac Surg ; 14(1): 22, 2019 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-30683137

RESUMO

BACKGROUND: Grafting vessel with LIMA to the left anterior descending coronary artery plays a most important role in the long-term prognosis of OPCABG surgery. The aim of this study was to compare the effects of isoflurane preconditioning on miRs and mRNAs levels in the left internal mammary arterie (LIMA) graft with propofol in patients undergoing off-pump coronary artery bypass surgery (OPCABG). METHODS: Patients were randomly assigned to receive either propofol (n = 15), or interrupted isoflurane (n = 15). In group P, propofol administration was continued at 3-5 mg/kg/h intravenous injection for the duration of surgery. Five minutes prior to incision, patients of the isoflurane group (group Iso) received 2 cycles of 1 MAC isoflurane. RESULTS: miR-221 were significantly lower in group Iso (P < 0 .05). E-selectin mRNA, RhoA mRNA and ROK mRNA were significantly lower at specimens of LIMA in group Iso compared with those in group P patients (P < 0 .05). The expression of NOS3 mRNA was significantly higher in group Iso patients (P < 0 .05). CONCLUSION: Our findings provide some insight that prior interrupted isoflurane administration could regulate miR-221, and downstream effectors (mRNAs) and resulted in actual attenuation of inflammation and spasm of LIMA in patients undergoing OPCABG surgery. TRIAL REGISTRATION: NCT No. ( ClinicalTrials.gov ): NCT02678650; Registration date: January 23, 2016.


Assuntos
Ponte de Artéria Coronária sem Circulação Extracorpórea/métodos , Doença da Artéria Coronariana/cirurgia , Vasos Coronários/metabolismo , Precondicionamento Isquêmico Miocárdico/métodos , Isoflurano/farmacologia , Óxido Nítrico/genética , Quinases Associadas a rho/genética , Idoso , Anestésicos Inalatórios/farmacologia , Doença da Artéria Coronariana/genética , Doença da Artéria Coronariana/metabolismo , Vasos Coronários/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Artéria Torácica Interna/metabolismo , Pessoa de Meia-Idade , Óxido Nítrico/biossíntese , Estudos Prospectivos , RNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Quinases Associadas a rho/biossíntese
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