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1.
Phytomedicine ; 129: 155597, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38643713

RESUMO

BACKGROUND: Sepsis-induced cardiac dysfunction (SICD) is a serious complication of sepsis that is associated with increased mortality. Ferroptosis has been reported in the SICD. TaoHe ChengQi decoction (THCQD), a classical traditional Chinese medicinal formula, has multiple beneficial pharmacological effects. The potential effects of THCQD on the SICD remain unknown. PURPOSE: To investigate the effect of THCQD on SICD and explore whether this effect is related to the regulation of myocardial ferroptosis through nuclear factor erythroid 2-related factor 2 (Nrf2) activation. METHODS: We induced sepsis in a mouse model using cecal ligation and puncture (CLP) and administered THCQD (2 and 4 g/kg) and dexamethasone (40 mg/kg). Mice mortality was recorded and survival curves were plotted. Echocardiography, hematoxylin and eosin staining, and analysis of serum myocardial injury markers and inflammatory factors were used to evaluate cardiac pathology. Myocardial ferroptosis was detected by quantifying specific biomarker content and protein levels. Through HPLC-Q-Exactive-MS analysis, we identified the components of the THCQD. Network pharmacology analysis and Cellular Thermal Shift Assay (CETSA) were utilized to predict the targets of THCQD for treating SICD. We detected the expression of Nrf2 using Western blotting or immunofluorescence. An RSL3-induced ferroptosis model was established using neonatal rat cardiomyocytes (NRCMs) to further explore the pharmacological mechanism of THCQD. In addition to measuring cell viability, we observed changes in NRCM mitochondria using electron microscopy and JC-1 staining. NRF2 inhibitor ML385 and Nrf2 knockout mice were used to validate whether THCQD exerted protective effects against SICD through Nrf2-mediated ferroptosis signaling. RESULTS: THCQD reduced mortality in septic mice, protected against CLP-induced myocardial injury, decreased systemic inflammatory response, and prevented myocardial ferroptosis. Network pharmacology analysis and CETSA experiments predicted that THCQD may protect against SICD by activating the Nrf2 signaling pathway. Western blotting and immunofluorescence showed that THCQD activated Nrf2 in cardiac tissue. THCQDs consistently mitigated RSL3-induced ferroptosis in NRCM, which is related to Nrf2. Furthermore, the pharmacological inhibition of Nrf2 and genetic Nrf2 knockout partially reversed the protective effects of THCQD on SICD and ferroptosis. CONCLUSION: The effect of THCQD on SICD was achieved by activating Nrf2 and its downstream pathways.


Assuntos
Modelos Animais de Doenças , Medicamentos de Ervas Chinesas , Ferroptose , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2 , Sepse , Animais , Sepse/complicações , Sepse/tratamento farmacológico , Fator 2 Relacionado a NF-E2/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Ferroptose/efeitos dos fármacos , Masculino , Camundongos , Ratos , Transdução de Sinais/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Miocárdio/metabolismo , Cardiopatias/tratamento farmacológico , Cardiopatias/etiologia , Farmacologia em Rede , Ratos Sprague-Dawley
2.
Phytomedicine ; 121: 155118, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37801895

RESUMO

BACKGROUND: With an increasing number of myocardial infarction (MI) patients, myocardial fibrosis is becoming a widespread health concern. It's becoming more and more urgent to conduct additional research and investigations into efficient treatments. Ethyl ferulate (EF) is a naturally occurring substance with cardioprotective properties. However, the extent of its impact and the underlying mechanism of its treatment for myocardial fibrosis after MI remain unknown. PURPOSE: The goal of this study was to look into how EF affected the signaling of the TGF-receptor 1 (TGFBR1) in myocardial fibrosis after MI. METHODS: Echocardiography, hematoxylin-eosin (HE) and Masson trichrome staining were employed to assess the impact of EF on heart structure and function in MI-affected mice in vivo. Cell proliferation assay (MTS), 5-Ethynyl-2'-deoxyuridine (EdU), and western blot techniques were employed to examine the influence of EF on native cardiac fibroblast (CFs) proliferation and collagen deposition. Molecular simulation and surface plasmon resonance imaging (SPRi) were utilized to explore TGFBR1 and EF interaction. Cardiac-specific Tgfbr1 knockout mice (Tgfbr1ΔMCK) were utilized to testify to the impact of EF. RESULTS: In vivo experiments revealed that EF alleviated myocardial fibrosis, improved cardiac dysfunction after MI and downregulated the TGFBR1 signaling in a dose-dependent manner. Moreover, in vitro experiments revealed that EF significantly inhibited CFs proliferation, collagen deposition and TGFBR1 signaling followed by TGF-ß1 stimulation. More specifically, molecular simulation, molecular dynamics, and SPRi collectively showed that EF directly targeted TGFBR1. Lastly, knocking down of Tgfbr1 partially reversed the inhibitory activity of EF on myocardial fibrosis in MI mice. CONCLUSION: EF attenuated myocardial fibrosis post-MI by directly suppressing TGFBR1 and its downstream signaling pathway.


Assuntos
Infarto do Miocárdio , Miocárdio , Humanos , Camundongos , Animais , Miocárdio/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I/uso terapêutico , Fibroblastos/metabolismo , Infarto do Miocárdio/tratamento farmacológico , Infarto do Miocárdio/metabolismo , Colágeno/metabolismo , Fibrose , Fator de Crescimento Transformador beta1/metabolismo
3.
J Opt Soc Am A Opt Image Sci Vis ; 40(1): 1-9, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-36607069

RESUMO

Aiming to solve the problem of low-light-level (LLL) images with dim overall brightness, uneven gray distribution, and low contrast, in this paper, we propose an effective LLL image enhancement method based on the guided filter and multi-scale fusion for contrast enhancement and detail preservation. First, a base image and detail image(s) are obtained by using the guided filter. After this procedure, the base image is processed by a maximum entropy-based Gamma correction to stretch the gray level distribution. Unlike the existing methods, we enhance the detail image(s) based on the guided filter kernel, which reflects the image area information. Finally, a new method is proposed to generate a sequence of artificial images to adjust the brightness of the output, which has a better performance in image detail preservation compared with other single-input algorithms. Experiments show that the proposed method can provide a more significant performance in enhancing contrast, preserving details, and maintaining the natural feeling of the image than the state of the art.

4.
Phytother Res ; 37(1): 35-49, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36059198

RESUMO

Myocardial infarction (MI) is the leading cause of death worldwide, and oxidative stress is part of the process that causes MI. Calycosin, a naturally occurring substance with cardioprotective properties, is one of the major active constituents in Radix Astragali. In this study, effect of Calycosin was investigated in vivo and in vitro to determine whether it could alleviate oxidative stress and oxidative stress-induced cardiac apoptosis in neonatal cardiomyocytes (NCMs) via activation of aldehyde dehydrogenase 2 (ALDH2). Calycosin protected against oxidative stress and oxidative stress-induced apoptosis in NCMs. Molecular docking revealed that the ALDH2-Calycosin complex had a binding energy of -9.885 kcal/mol. In addition, molecular docking simulations demonstrated that the ALDH2-Calycosin complex was stable. Using BLI assays, we confirmed that Calycosin could interact with ALDH2 (KD  = 1.9 × 10-4 M). Furthermore, an ALDH2 kinase activity test revealed that Calycosin increased ALDH2 activity, exhibiting an EC50 of 91.79 µM. Pre-incubation with ALDH2 inhibitor (CVT-10216 or disulfiram) reduced the cardio-protective properties Calycosin. In mice with MI, Calycosin therapy substantially reduced myocardial apoptosis, oxidative stress, and activated ALDH2. Collectively, our findings clearly suggest that Calycosin reduces oxidative stress and oxidative stress-induced apoptosis via the regulation of ALDH2 signaling, which supports potential therapeutic use in MI.


Assuntos
Infarto do Miocárdio , Miócitos Cardíacos , Camundongos , Animais , Aldeído-Desidrogenase Mitocondrial/metabolismo , Simulação de Acoplamento Molecular , Estresse Oxidativo , Apoptose , Aldeído Desidrogenase/metabolismo
5.
J Ethnopharmacol ; 302(Pt A): 115869, 2023 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-36309116

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Radix Glycyrrhizae (GL), a herbal medicine that is widely available, has shown advantages for a variety of inflammatory diseases. Toll like receptor 4 (TLR4) pathway has been shown to play a key role in the progression of inflammation. AIM OF THE STUDY: The purpose of this study was to investigate the involvement of TLR4 in the anti-inflammatory mechanism of GL extract and its active constituent on acute lung injury (ALI). MATERIALS AND METHODS: A model of inflammation produced by lipopolysaccharide (LPS) was established in C57BL/6 mice and macrophages derived from THP-1. To screen the active components of GL, molecular docking was used. Molecular dynamics and surface plasmon resonance imaging (SPRi) were used to study the interaction of a specific drug with the TLR4-MD2 complex. TLR4 was overexpressed by adenovirus to confirm TLR4 involvement in the anti-inflammatory activities of GL and the chosen chemical. RESULTS: We observed that GL extract significantly reduced both LPS-induced ALI and the production of pro-inflammatory factors including TNF-α, IL-6 and IL-1ß. Additionally, GL inhibited the binding of Alexa 488-labeled LPS (LPS-488) to the membrane of THP-1 derived macrophages. GL drastically reduce on the expression of TLR4 and the activation of mitogen-activated protein kinases (MAPKs) and nuclear factor-B (NF-κB). Furthermore, molecular docking revealed that Licochalcone A (LicoA) docked into the LPS binding site of TLR4-MD2 complex. MD2-LicoA binding conformation was found to be stable using molecular dynamic simulations. SPRi indicated that LicoA bound to TLR4-MD2 recombinant protein with a KD of 3.87 × 10-7 M. LicoA dose-dependently reduced LPS-488 binding to the cell membrane. LicoA was found to significantly inhibit LPS-induced lung damage and inflammation. Furthermore, LicoA inhibited TLR4 expression, MAPK and NF-κB activation in a dose-dependent manner. The inhibitory effects of GL and LicoA on LPS-induced inflammation and TLR4 signaling activation were partly eliminated by TLR4 overexpression. CONCLUSION: Our findings imply that GL and LicoA exert inhibitory effects on inflammation by targeting the TLR4 directly.


Assuntos
Lesão Pulmonar Aguda , Receptor 4 Toll-Like , Camundongos , Animais , Receptor 4 Toll-Like/metabolismo , Lipopolissacarídeos/toxicidade , NF-kappa B/metabolismo , Simulação de Acoplamento Molecular , Camundongos Endogâmicos C57BL , Antígeno 96 de Linfócito/metabolismo , Anti-Inflamatórios/efeitos adversos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Inflamação/induzido quimicamente
6.
Appl Opt ; 61(21): 6339-6348, 2022 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-36256249

RESUMO

Tone mapping operators (TMOs) aim to adjust high dynamic range (HDR) images to low dynamic range (LDR) ones so that they can be displayed on conventional devices with visual information retained. Nonetheless, existing TMOs can successfully tone-map only limited types of HDR images, and the parameters need to be manually adjusted to yield the best subjective-quality tone-mapped outputs. To cope with the aforementioned issues, an adaptive parameter-free and scene-adaptive TMO for dynamic range adjusting and detail enhancing is proposed to yield a high-resolution and high-subjective-quality tone-mapped output. This method is based on detail/base layer decomposition to decompose the input HDR image into coarse detail, fine detail, and base images. After that, we adopt different strategies to process each layer to adjust the overall brightness and contrast and to retain as much scene information. Finally, a new method, to the best of our knowledge, is proposed for visualization to generate a sequence of artificial images to adjust the brightness. Experiments with numerous HDR images and state-of-the-art TMOs are conducted; the results demonstrate that the proposed method consistently produces better quality tone-mapped images than the state-of-the-art methods.

7.
Food Funct ; 13(6): 3234-3246, 2022 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-35213678

RESUMO

Background: Abnormal proliferation of vascular smooth muscle cells (VSMCs) in the intimal region is a key event in the development of neointimal hyperplasia. 10-G, a bioactive compound found in ginger, exerted inhibitory effects on the proliferation of several cancer cells. However, the effect and mechanism of 10-G on neointimal hyperplasia are not clear. Purpose: To explore the suppressive effects of 10-G on the proliferation and migration of VSMCs, and investigate the underlying mechanisms. Methods: In vivo, a left common carotid artery ligation mouse model was used to observe the effects of neointimal formation through immunohistochemistry and hematoxylin-eosin staining. In vitro, the cell proliferation and migration of HASMCs and A7r5 cells were detected by MTS assay, EdU staining, wound healing assay, Transwell assay, and western blotting as well. Molecular docking, molecular dynamics simulations and surface plasmon resonance imaging were collectively used to evaluate the interaction of 10-G with AMP-activated protein kinase (AMPK). Compound C and si-AMPK were used to inhibit the expression of AMPK. Results: Treatment with 10-G significantly reduced neointimal hyperplasia in the left common carotid artery ligation mouse model. MST and EdU staining showed that 10-G inhibited the proliferation of VSMC cells A7r5 and HASMC. We also found that 10-G altered the expression of proliferation-related proteins, including CyclinD1, CyclinD2, CyclinD3, and CDK4. Molecular docking revealed that the binding energy between AMPK and 10-G is -7.4 kcal mol-1. Molecular simulations suggested that the binding between 10-G and AMPK is stable. Surface plasmon resonance imaging analysis also showed that 10-G has a strong binding affinity to AMPK (KD = 6.81 × 10-8 M). 10-G promoted AMPKα phosphorylation both in vivo and in vitro. Blocking AMPK by an siRNA or AMPK inhibitor pathway partly abolished the anti-proliferation effects of 10-G on VSMCs. Conclusion: These data showed that 10-G might inhibit neointimal hyperplasia and suppress VSMC proliferation by the activation of AMPK as a natural AMPK agonist.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Catecóis/farmacologia , Álcoois Graxos/farmacologia , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Neointima/patologia , Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Proteínas Quinases Ativadas por AMP/química , Animais , Catecóis/química , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ativação Enzimática , Álcoois Graxos/química , Humanos , Hiperplasia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Simulação de Acoplamento Molecular , Músculo Liso Vascular/efeitos dos fármacos , Fosforilação , Conformação Proteica , Ratos , Transdução de Sinais , Ressonância de Plasmônio de Superfície , Serina-Treonina Quinases TOR/metabolismo
8.
Phytomedicine ; 95: 153705, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34538671

RESUMO

BACKGROUND: Liver cancer is one of the leading causes of cancer-related death worldwide. Dihydrotanshinone I (DHI) was shown to inhibit the growth of several types of cancer. However, research related to hepatoma treatment using DHI is limited. PURPOSE: Here, we explored the inhibitory effect of DHI on the growth of hepatoma cells, and investigated the underlying molecular mechanisms. METHODS: The proliferation of Hep3B, SMCC-7721 and SK-Hep1 hepatoma cells was evaluated using the MTS and Edu staining assay. Hepatoma cell death was analyzed with a LIVE/DEAD Cell Imaging Kit. The relative expression and phosphorylation of proto-oncogene tyrosine-protein kinase Src (Src) and signal transducer and activator of transcription-3 (STAT3) proteins in hepatoma cells, as well as the expression of other protein components, were measured by western blotting. The structural interaction of DHI with Src proteins was evaluated by molecular docking, molecular dynamics simulation, surface plasmon resonance imaging and Src kinase inhibition assay. Src overexpression was achieved by infection with an adenovirus vector encoding human Src. Subsequently, the effects of DHI on tumor growth inhibition were further validated using mouse xenograft models of hepatoma. RESULTS: In vitro studies showed that treatment with DHI inhibited the proliferation and promoted cell death of Hep3B, SMCC-7721 and SK-Hep1 hepatoma cells. We further identified and verified Src as a direct target of DHI by using molecular stimulation, surface plasmon resonance image and Src kinase inhibition assay. Treatment with DHI reduced the in vitro phosphorylation levels of Src and STAT3, a transcription factor regulated by Src. In the xenograft mouse models, DHI dose-dependently suppressed tumor growth and Src and STAT3 phosphorylation. Moreover, Src overexpression partly abrogated the inhibitory effects of DHI on the proliferation and cell death in hepatoma cells. CONCLUSION: Our results suggest that DHI inhibits the growth of hepatoma cells by direct inhibition of Src.


Assuntos
Carcinoma Hepatocelular , Furanos/farmacologia , Fenantrenos , Quinonas/farmacologia , Quinases da Família src/antagonistas & inibidores , Animais , Carcinoma Hepatocelular/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células , Camundongos , Simulação de Acoplamento Molecular , Fenantrenos/farmacologia , Fosforilação , Fator de Transcrição STAT3/metabolismo , Quinases da Família src/metabolismo
9.
Medicine (Baltimore) ; 100(33): e26858, 2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34414936

RESUMO

BACKGROUND: The proportion of women is higher than men in depression. This is mainly due to women's physiological regulation is different from men, especially in puberty, menstruation, pregnancy, menopause, among others. Therefore, treating depressive women is still a health challenge. Besides, recent studies of exercise therapy have a more outstanding performance in treating depression, especially in contrast to drug therapy and psychotherapy. Its main advantages are convenience, quickness, no side effects, real-time, and long-term effectiveness. OBJECTIVE: The aim of this study was to systematically review the clinical efficacy of exercise on women with depressive symptoms. METHODS: Searching PubMed, The Cochrane Library, and Embase databases to collect randomized controlled trials about exercise in the treatment of depressive women. After literature screening, data extraction, and literature quality evaluation, the meta-analysis of acquirement data was performed with RevMan5.3 software. RESULTS: A total of 2294 patients were included in 25 different articles totally. Meta-analysis shows that compared with the control group, exercise could relieve female depression (standard mean difference [95% confidence interval, CI] = -0.64 [-0.89 to -0.39], Z = 4.99, P < .001). Subgroup analysis shows that different types of exercise have significant effects in improving depression symptoms. Exercise therapy has better effect on depressive patients induced by physiology or disease than ordinary depressive patients. CONCLUSION: Exercise can significantly improve depressive symptoms in women.


Assuntos
Depressão/terapia , Terapia por Exercício , Feminino , Humanos , Resultado do Tratamento
10.
Phytother Res ; 35(7): 3836-3847, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33792976

RESUMO

Melanoma is the most common type of skin cancer. Signal transducer and activator of transcription 3 (STAT3) signaling has been demonstrated to be a therapeutic target for melanoma. Dauricine (Dau), an alkaloid compound isolated from the root of Menispermum dauricum DC., has shown tumor-suppressing effects in multiple human cancers, but its potential in melanoma remains unexplored. In this study, we demonstrated that Dau significantly inhibited the viability and proliferation of A375 and A2058 melanoma cells. Death of melanoma cells was also markedly promoted by Dau. Moreover, Dau inhibited phosphorylation-mediated activation of STAT3 and Src in a dose-dependent manner. Notably, constitutive activation of Src partially abolished the antiproliferative and cytotoxic activities of Dau on melanoma cells. Molecular docking showed that Dau could dock on the kinase domain of Src with a binding energy of -10.42 kcal/mol. Molecular dynamics simulations showed that Src-Dau binding was stable. Surface plasmon resonance imaging analysis also showed that Dau has a strong binding affinity to Src. In addition, Dau suppressed the growth of melanoma cells and downregulated the activation of Src/STAT3 in a xenograft model in vivo. These data demonstrated that Dau inhibits proliferation and promotes cell death in melanoma cells by inhibiting the Src/STAT3 pathways.


Assuntos
Benzilisoquinolinas/farmacologia , Melanoma , Proteínas Proto-Oncogênicas pp60(c-src) , Fator de Transcrição STAT3 , Tetra-Hidroisoquinolinas/farmacologia , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Melanoma/tratamento farmacológico , Simulação de Acoplamento Molecular , Fosforilação , Proteínas Proto-Oncogênicas pp60(c-src)/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos
11.
Tumour Biol ; 2016 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-27743381

RESUMO

Rapidly accumulated evidence has shown that long non-coding RNA (lncRNAs) disregulation is involved in human tumorigenesis in many cancers, including prostate cancer (PCa). LncRNAs can regulate essential pathways that contribute to tumor initiation and progression with tissue specificity, which suggests that lncRNAs could be valuable biomarkers and therapeutic targets. Prostate cancer antigen 3 (PCA3), also known as differential display code 3 (DD3), is one such lncRNA that maps to chromosome 9q21-22. PCA3 expression is highly specific to PCa. In the present study, the level of PCA3 expression in prostate cancer cells was reduced by small interfering RNA (siRNA). Subsequently, the ability of LNCaP cell proliferation, invasion, and migration of PCa was compromised both in vivo and in vitro with the occurrence of cell autophagy. Recently, a novel regulatory mechanism has been proposed in which RNAs cross talk via competing with the shared microRNAs (miRNAs). In addition, lncRNAs can directly interact with RNA-binding proteins and then bind to the gene promoter region to further regulate gene expression. The proposed competitive endogenous RNAs mediate the bioavailability of miRNAs on their targets, thus imposing another level of post-transcriptional regulation. Here, we demonstrated that binding of Snail to the promoter region of PCA3 could activate the expression of PCA3. Down-regulation of PCA3 by silencing could increase the expression of the miRNA-1261, which then targeted at the PRKD3 gene (protein kinase D3) through competitive sponging. In summary, these results suggest that the transcription factor, Snail, activated the expression of lncRNA PCA3, which could inhibit the translation of PRKD3 protein via competitive miR-1261 sponging, and thus high expression of PRKD3 further promoted invasion and migration of prostate cancer.

12.
Biochem Biophys Res Commun ; 475(4): 329-34, 2016 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-27216460

RESUMO

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) and GATA Binding Protein 4 (GATA4) are important for the growth of cardiac fibroblasts (CFs). When deregulated, LOX-1 and GATA4 can cause cardiac remodeling. In the present study, we found novel evidence that GATA4 was required for the LOX-1 regulation of CF proliferation. The inhibition of LOX-1 by RNA interference LOX-1 lentivirus resulted in the loss of PI3K/Akt activation and GATA4 protein expression. The overexpression of LOX-1 by lentivirus rescued CF proliferation, PI3K/Akt activation, and GATA4 protein expression. Moreover, GATA4 overexpression enhanced CF proliferation with LOX-1 inhibition. We also found that the inhibition of PI3K/Akt activation by LY294002, a PI3K inhibitor, reduced cell proliferation and protein level of GATA4. In summary, GATA4 may play an important role in the LOX-1 and PI3K/Akt regulation of CF proliferation.


Assuntos
Proliferação de Células , Fibroblastos/citologia , Fator de Transcrição GATA4/metabolismo , Miocárdio/citologia , Receptores Depuradores Classe E/metabolismo , Animais , Células Cultivadas , Fibroblastos/metabolismo , Miocárdio/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais
13.
J Exp Clin Cancer Res ; 33: 72, 2014 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-25200485

RESUMO

Prostate cancer gene expression marker 1 (PCGEM1) is a long non-coding RNA (lncRNA) overexpressed in prostate cancer (PCa) cells that promotes PCa initiation and progression, and protects against chemotherapy-induced apoptosis. The microRNA miR-145 functions as a tumor suppressor in PCa. We speculate that reciprocal regulation of PCGEM1 and miR-145 promote proliferation of LNCaP prostate cancer cells. To test this hypothesis, the interaction between PCGEM1 and miR-145 was examined using a luciferase reporter assay. Expression levels were selectively altered in LNCaP cells and noncancerous RWPE-1 prostate cells by transfection of miR-145 or small interfering RNA sequences against (siRNA) PCGEM1. Relative expression levels were detected by RT-PCR, tumor cell growth and early apoptosis by the MTT assay and flow cytometry, respectively, and tumor cell migration and invasion properties by transwell assays. The effect of siRNA PCGEM1 and miR-145 transfection on prostate cancer growth in vivo was examined in the (nu/nu) mouse model. PCGEM1 and miR-145 exhibited reciprocal regulation; downregulation of PCGEM1 expression in LNCaP cells increased expression of miR-145, while overexpression of miR-145 decreased PCGEM1 expression. Transfection of the miR-145 expression vector and siRNA PCGEM1 inhibited tumor cell proliferation, migration, and invasion, and induced early apoptosis both in vitro. In contrast, there was no effect on RWPE-1 cells. We demonstrate a reciprocal negative control relationship between PCGEM1 and miR-145 that regulates both LNCaP cell proliferation and nu/nu PCa tumor growth. The results also identify PCGEM1 and associated regulators as possible targets for PCa therapy.


Assuntos
Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Neoplasias da Próstata/genética , RNA Longo não Codificante/genética , Regiões 3' não Traduzidas , Animais , Apoptose/genética , Sequência de Bases , Sítios de Ligação , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , Modelos Animais de Doenças , Progressão da Doença , Técnicas de Silenciamento de Genes , Xenoenxertos , Humanos , Masculino , Camundongos , MicroRNAs/química , Neoplasias da Próstata/patologia , RNA Longo não Codificante/química , RNA Mensageiro/química , RNA Mensageiro/genética , Transfecção , Carga Tumoral
14.
Yi Chuan ; 30(12): 1629-34, 2008 Dec.
Artigo em Chinês | MEDLINE | ID: mdl-19073581

RESUMO

To explore the feasibility and accuracy of MLPA-based array (Array-MLPA) in detecting sex chromosome abnormalities, MLPA probes were designed to target against three gene loci, TSPY (p11.2), PRY (q11), and RBMY (q11.2) in human Y chromosome. Array-MLPA approach was applied to test abnormalities of Y chromosome in 15 patient samples with known karyotypes. The data were compared with karyotyping and PCR analyses. The results showed that the copy number of each site detected by Array-MLPA was basically consistent with karyotyping analysis. Moreover, small deletions of chromosomes that were not found by routine karyotyping analysis were identified by the approach described, which fully agreed with PCR analysis, indicating that Array-MLPA was able to detect small abnormalities of chromosomes that cannot be found by karyotyping analysis. Compared to the routine karyotyping method, Array-MLPA has the advantages of high efficiency and reliability in chromosomal analysis, which has great potential in clinical application of diagnosis of chromosome abnormalities.


Assuntos
Aberrações Cromossômicas , Cromossomos Humanos Y/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Humanos , Cariotipagem , Reação em Cadeia da Polimerase
15.
Hum Mol Genet ; 16(21): 2616-25, 2007 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-17716993

RESUMO

The beta-thalassemia is associated with abnormality in beta-globin gene, leading to imbalanced synthesis of alpha-/beta-globin chains. Consequently, the excessive free alpha-globin chains precipitate to the erythrocyte membrane, resulting in hemolytic anemia. We have explored post-transcriptional strategies aiming at alpha-globin reduction and beta-globin enrichment on beta(654) (Hbb(th-4)/Hbb(+)) mouse, carrying a human splicing-deficient beta-globin allele (Hbb(th-4)). Lentiviral vectors of short hairpin RNA (shRNA) targeting alpha-globin and/or antisense RNA facilitating beta-globin correct splicing were microinjected into beta(654) single-cell embryos. Three transgenic strains were generated, as alpha(i)-Hbb(th-4)/Hbb(+)(shRNA), beta(a)-Hbb(th-4)/Hbb(+)(antisense) and alpha(i)beta(a)-Hbb(th-4)/Hbb(+)(both shRNA and antisense). Without notable abnormalities, all the founders and their offsprings showed sustained amelioration of hematologic parameters, ineffective erythropoiesis and extramedullary hematopoiesis. Augmented effects appeared in alpha(i)beta(a)-Hbb(th-4)/Hbb(+), which correlated with a better-balanced alpha-/beta-globin mRNA level. Among the transgenic mice integrated with shRNA and antisense RNA, one homozygous mouse (Hbb(th-4)/Hbb(th-4)) had been viable, and the 3-week survival rate for heterozygotes (Hbb(th-4)/Hbb(+)) was 97%, compared with 45.4% for untreated. Our data have demonstrated the feasibility of techniques for beta-thalassemia therapy by balancing the synthesis of alpha-/beta-globin chains.


Assuntos
Terapia Genética/métodos , Globinas/antagonistas & inibidores , RNA Antissenso/genética , RNA Interferente Pequeno/genética , Talassemia beta/terapia , Anemia/terapia , Animais , Medula Óssea/patologia , Eritropoese/genética , Expressão Gênica , Vetores Genéticos/genética , Globinas/genética , Hemoglobinas/genética , Humanos , Lentivirus/genética , Camundongos , Camundongos Transgênicos , Fenótipo , Splicing de RNA/genética
16.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 22(4): 431-4, 2005 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-16086284

RESUMO

OBJECTIVE: To evaluate the role of RNA interference (RNAi) in silencing the enhanced green fluorescent protein (eGFP) expression in 293T and Mel cells. METHODS: Nested-PCR was used to amplify H1 promoter from human 293T cells for driving RNAi synthesis. RNAi vectors (TR1) for silencing the eGFP expression was constructed. The eGFP vector and RNAi vector (TR1) were then co-transfected into the 293T and Mel cells, in which the silencing effect on eGFP expression was investigated by fluorescence microscopy, reverse transcription-PCR(RT-PCR), fluorescence-assited cell sorting(FACS) analysis and real-time RT-PCR. RESULTS: RNAi could effectively reduce more than 50 percent of eGFP expression in 293T cells as well as in Mel cells. CONCLUSION: The RNAi vector constructed in this way paper can effectively inhibit eGFP expression in cells.


Assuntos
Proteínas de Fluorescência Verde/genética , Interferência de RNA , Linhagem Celular , Citometria de Fluxo , Vetores Genéticos/genética , Humanos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
17.
Blood Cells Mol Dis ; 34(3): 220-5, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-15885605

RESUMO

We have studied the role of small interfering RNA (siRNA) in the suppression of the enhanced green fluorescent protein (eGFP) in vitro and in vivo. siRNA plasmids for suppressing the eGFP expression were constructed, in which H1 promoter amplified from human 293 T cells was used to drive the small interfering RNA (19-nt) synthesis. In vitro studies demonstrated that the constructed siRNA plasmids had an effective suppressive effect on eGFP expression in 293 T and Mel cells. When the siRNA plasmid was injected into erythroid-specific eGFP transgenic mice, the eGFP expression were significantly suppressed (over 20% reduction) in the recipient mice compared to the control mice and the suppressing effect lasted for 2 weeks after one single injection. Moreover, the suppressive effect could be re-generated or boosted with secondary injections. Our results demonstrate a tissue-specific gene suppression by siRNA treatment.


Assuntos
Regulação para Baixo/efeitos dos fármacos , Células Eritroides/metabolismo , Proteínas de Fluorescência Verde/genética , RNA Interferente Pequeno/farmacologia , Animais , Linhagem Celular , Regulação para Baixo/genética , Proteínas de Fluorescência Verde/antagonistas & inibidores , Humanos , Camundongos , Camundongos Transgênicos , Plasmídeos/administração & dosagem , Plasmídeos/genética , Regiões Promotoras Genéticas , RNA Interferente Pequeno/administração & dosagem , Fatores de Tempo
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