RESUMO
Neural-derived 17ß-estradiol (E2) plays an important role in the synaptic plasticity of the hippocampus and prefrontal cortex, but the mechanism is not well defined. This study was designed to explore the effect and mechanism of neural-derived E2 on synaptic plasticity of the hippocampus and prefrontal cortex. Primary cultured hippocampal and prefrontal cells in mice were randomly divided into the DMSO (D), aromatase (Rate-limiting enzymes for E2 synthesizes) inhibitor letrozole (L), and ERs antagonist (MPG) treated groups. After intervention for 48 h, the cell was collected, and then, the expressions of AMPA-receptor subunit GluR1 (GluR1), synaptophysin (SYN), p-21-Activated kinase (PAK) phosphorylation, Rho kinase (ROCK), p-Cofilin, F-actin, and G-actin proteins were detected. Letrozole or ER antagonists inhibited the expression of GluR1, F-actin/G-actin, p-PAK and p-Cofilin proteins in prefrontal cells significantly. And the expressions of GluR1 and F-actin/G-actin proteins were declined in hippocampal cells markedly after adding letrozole or ERs antagonists. In conclusion, neural-derived E2 and ERs regulated the synaptic plasticity, possibly due to promoting actin polymerization in prefrontal and hippocampal cells. The regional specificity in the effect of neural-derived E2 and ERs on the actin polymerization-related pathway may provide a theoretical basis for the functional differences between the hippocampus and prefrontal cortex.
Assuntos
Actinas/metabolismo , Estradiol/farmacologia , Hipocampo/efeitos dos fármacos , Plasticidade Neuronal/efeitos dos fármacos , Fatores de Despolimerização de Actina/antagonistas & inibidores , Fatores de Despolimerização de Actina/metabolismo , Actinas/antagonistas & inibidores , Animais , Células Cultivadas , Hipocampo/metabolismo , Letrozol/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Polimerização/efeitos dos fármacos , Proteínas Quinases/metabolismo , Receptores de AMPA/antagonistas & inibidores , Receptores de AMPA/metabolismoRESUMO
As one of the most commonly used nanoparticles, titanium dioxide nanoparticles (TiO2-NPs) are widely used as coating reagents in cosmetics, medicine and other industries. The increasing risk of exposure to TiO2-NPs raises concerns about their safety. In this study, we investigated the mechanism by which TiO2-NPs cross the blood-testis barrier (BTB). TM-4 cells were selected as an in vitro Sertoli cell model of BTB. Cell viability, cell morphological changes, apoptosis, oxidative damage, and the expression levels of actin regulatory and tight junction (TJ) proteins were assessed in TM-4 cells treated with 3-nm and 24-nm TiO2-NPs. Cells treated with 3-nm TiO2-NPs exhibited increased cytotoxicity and decreased Annexin II expression, whereas cells treated with 24-nm TiO2-NPs exhibited increased Arp 3 and c-Src expression. Both TiO2-NPs induced significant oxidative stress, decreased the expression of TJ proteins (occludin, ZO-1 and claudin 5), damaged the TJ structure, and exhibited enlarged gaps between TM-4 cells. Our results indicated that both TiO2-NPs crossed the BTB by disrupting actin-based adhesive junctions of TM-4 cells; however, apoptosis was not observed. Our results provide new insights into how TiO2-NPs cross the BTB.
Assuntos
Actinas/antagonistas & inibidores , Barreira Hematotesticular/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Nanopartículas Metálicas/efeitos adversos , Titânio/efeitos adversos , Actinas/metabolismo , Animais , Linhagem Celular , Relação Dose-Resposta a Droga , Masculino , Camundongos , Células de Sertoli/efeitos dos fármacos , Células de Sertoli/metabolismo , Proteínas de Junções Íntimas/metabolismoRESUMO
Aberrant angiogenesis lies at the heart of a wide range of ocular pathologies such as proliferative diabetic retinopathy, wet age-related macular degeneration and retinopathy of prematurity. This study explores the anti-angiogenic activity of a novel small molecule investigative compound capable of inhibiting profilin1-actin interaction recently identified by our group. We demonstrate that our compound is capable of inhibiting migration, proliferation and angiogenic activity of microvascular endothelial cells in vitro as well as choroidal neovascularization (CNV) ex vivo. In mouse model of laser-injury induced CNV, intravitreal administration of this compound diminishes sub-retinal neovascularization. Finally, our preliminary structure-activity relationship study (SAR) demonstrates that this small molecule compound is amenable to improvement in biological activity through structural modifications.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Neovascularização de Coroide/tratamento farmacológico , Neovascularização Retiniana/tratamento farmacológico , Actinas/antagonistas & inibidores , Animais , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neovascularização de Coroide/metabolismo , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Humanos , Injeções Intravítreas , Camundongos , Camundongos Endogâmicos C57BL , Profilinas/antagonistas & inibidores , Neovascularização Retiniana/metabolismo , Vasos Retinianos/citologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Degeneração Macular Exsudativa/tratamento farmacológico , Degeneração Macular Exsudativa/metabolismoRESUMO
Chlorophyll and heme are among the "pigments of life", tetrapyrrolic structures, without which life on Earth would not be possible. Their catabolites, the phyllobilins and the bilins, respectively, share not only structural features, but also a similar story: Long considered waste products of detoxification processes, important bioactivities for both classes have now been demonstrated. For phyllobilins, however, research on physiological roles is sparse. Here, we introduce actin, the major component of the cytoskeleton, as the first discovered target of phyllobilins and as a novel target of bilins. We demonstrate the inhibition of actin dynamics in vitro and effects on actin and related processes in cancer cells. A direct interaction with G-actin is shown by in silico studies and confirmed by affinity chromatography. Our findings open a new chapter in bioactivities of tetrapyrroles-especially phyllobilins-for which they form the basis for broad implications in plant science, ecology, and physiology.
Assuntos
Actinas/antagonistas & inibidores , Clorofila/química , Heme/química , Pigmentos Biológicos/farmacologia , Tetrapirróis/farmacologia , Actinas/metabolismo , Humanos , Pigmentos Biológicos/química , Tetrapirróis/químicaRESUMO
Cancer metastasis is a complex process involving highly motile tumor cells that breach tissue barriers, enter the bloodstream and lymphatic system, and disseminate throughout the body as circulating tumor cells. The primary cellular mechanism contributing to these critical events is the reorganization of the actin cytoskeleton. Mycalolide B (MycB) is an actin-targeting marine macrolide that can suppress proliferation, migration, and invasion of breast and ovarian cancer cells at low nanomolar doses. Through structure-activity relationship studies focused on the actin-binding tail region (C24-C35) of MycB, we identified a potent truncated derivative that inhibits polymerization of G-actin and severs F-actin by binding to actin's barbed end cleft. Biological analyses of this miniature MycB derivative demonstrate that it causes a rapid collapse of the actin cytoskeleton in ovarian cancer cells and impairs cancer cell motility and invasion of the extracellular matrix (ECM) by inhibiting invadopodia-mediated ECM degradation. These studies provide essential proof-of-principle for developing actin-targeting therapeutic agents to block cancer metastasis and establish a synthetically tractable barbed end-binding pharmacophore that can be further improved by adding targeting groups for precision drug design.
Assuntos
Actinas/antagonistas & inibidores , Antineoplásicos/farmacologia , Matriz Extracelular/efeitos dos fármacos , Toxinas Marinhas/farmacologia , Oxazóis/farmacologia , Actinas/metabolismo , Antineoplásicos/síntese química , Antineoplásicos/química , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Matriz Extracelular/metabolismo , Feminino , Humanos , Toxinas Marinhas/síntese química , Toxinas Marinhas/química , Modelos Moleculares , Estrutura Molecular , Oxazóis/síntese química , Oxazóis/química , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Repetitive uses of antifungals result in a worldwide crisis of drug resistance; therefore, natural fungicides with minimal side-effects are currently sought after. This study aimed to investigate antifungal property of 19, 20-epoxycytochalasin Q (ECQ), derived from medicinal mushroom Xylaria sp. BCC 1067 of tropical forests. In a model yeast Saccharomyces cerevisiae, ECQ is more toxic in the erg6∆ strain, which has previously been shown to allow higher uptake of many hydrophilic toxins. We selected one pathway to study the effects of ECQ at very high levels on transcription: the ergosterol biosynthesis pathway, which is unlikely to be the primary target of ECQ. Ergosterol serves many functions that cholesterol does in human cells. ECQ's transcriptional effects were correlated with altered sterol and triacylglycerol levels. In the ECQ-treated Δerg6 strain, which presumably takes up far more ECQ than the wild-type strain, there was cell rupture. Increased actin aggregation and lipid droplets assembly were also found in the erg6∆ mutant. Thereby, ECQ is suggested to sensitize yeast cells lacking ERG6 through actin-targeting and consequently but not primarily led to disruption of lipid homeostasis. Investigation of cytochalasins may provide valuable insight with potential biopharmaceutical applications in treatments of fungal infection, cancer or metabolic disorder.
Assuntos
Actinas/antagonistas & inibidores , Antifúngicos/farmacologia , Citocalasinas/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Saccharomyces cerevisiae/efeitos dos fármacos , Xylariales/metabolismo , Metiltransferases/genética , Saccharomyces cerevisiae/genéticaRESUMO
Osteoporosis is a metabolic bone-loss disease characterized by abnormally excessive osteoclast formation and bone resorption. Identification of natural medicines that can inhibit osteoclastogenesis, bone resorption, and receptor activator of nuclear factor-κB ligand (RANKL)-induced signaling is necessary for improved treatment of osteoporosis. In this study, hinokitiol, a tropolone-related compound extracted from the heart wood of several cupressaceous plants, was found to inhibit RANKL-induced osteoclast formation and bone resorption in vitro. Hinokitiol inhibited early activation of the ERK, p38, and JNK-MAPK pathways, thereby suppressing the activity and expression of downstream factors (c-Jun, c-Fos, and NFATC1). Consistent with the above in vitro findings, hinokitiol treatment protected against ovariectomy-induced bone loss in vivo. Collectively, our results imply that hinokitiol can potentially serve as an effective agent for treating osteoclast-induced osteoporosis.
Assuntos
Conservadores da Densidade Óssea/farmacologia , Reabsorção Óssea/prevenção & controle , Monoterpenos/farmacologia , Osteogênese/efeitos dos fármacos , Osteoporose/metabolismo , Osteoporose/prevenção & controle , Tropolona/análogos & derivados , Actinas/antagonistas & inibidores , Animais , Conservadores da Densidade Óssea/uso terapêutico , Reabsorção Óssea/diagnóstico por imagem , Reabsorção Óssea/etiologia , Linhagem Celular , Modelos Animais de Doenças , Feminino , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Monoterpenos/uso terapêutico , Fatores de Transcrição NFATC/antagonistas & inibidores , Osteoclastos/efeitos dos fármacos , Osteogênese/genética , Ovariectomia/efeitos adversos , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-fos/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-jun/antagonistas & inibidores , Ligante RANK/toxicidade , Fator de Transcrição AP-1/antagonistas & inibidores , Tropolona/farmacologia , Tropolona/uso terapêuticoRESUMO
This study aimed to assess the effects of acetylation levels on actomyosin disassociation and phosphorylation of lamb during incubation at 4 °C. Samples of whole proteins from lamb longissimus thoracis muscles were prepared and assigned into three treatments (high, middle and low acetylation groups). The results showed that deacetylation of myosin heavy chain and actin was inhibited by lysine deacetylase inhibitor trichostatin A and nicotinamide in this study. Phosphorylation levels of myosin heavy chain and actin were inhibited by their acetylation during incubation in vitro. Actomyosin disassociation degree in high acetylation group was significantly lower than that in middle and low acetylation groups (P < 0.05). The ATPase activity in high acetylation group was significantly higher than that in middle and low acetylation groups (P < 0.05). In conclusion, acetylation of myosin heavy chain and actin inhibited actomyosin dissociation by inhibiting their phosphorylation at 4 °C in vitro.
Assuntos
Actomiosina/metabolismo , Músculos/metabolismo , Acetilação/efeitos dos fármacos , Actinas/antagonistas & inibidores , Actinas/metabolismo , Actomiosina/antagonistas & inibidores , Animais , Sítios de Ligação , Temperatura Baixa , Ácidos Hidroxâmicos/farmacologia , Simulação de Dinâmica Molecular , Cadeias Pesadas de Miosina/antagonistas & inibidores , Cadeias Pesadas de Miosina/metabolismo , Niacinamida/farmacologia , Fosforilação , OvinosRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Curculigo orchioides Gaertn is used for the treatment of impotence, atrophic debility of bones (osteoporosis), limb limpness, and arthritis of the lumbar and knee joints in traditional Chinese medicine and Ayurvedic medical system. Curculigoside (Cur) from Curculigo orchioides Gaertn has been shown to have regulatory effects on bone metabolism via anti-oxidative activities in rats and osteoblasts. However, little is known about the molecular pharmacological activity of Cur in osteoclastic bone resorption. AIM: The aim of this work is to investigate the inhibitory effect of Cur against osteoclasts (OCs) under the oxidative stress status, and explore the possible underlying mechanism. MATERIALS AND METHODS: OCs were induced from RAW264.7 cells using RANKL and H2O2. The number of OCs was measured by tartrate-resistant acid phosphatase (TRAP) staining. F-Actin and nuclear translocation of P65 and Nrf2 were stained with immunofluorescence assay and observed under a laser confocal microscope. The biochemical parameters of OCs were detected with an ELISA kit. The expression of Nrf2 and NF-κB pathway-related proteins was analyzed by Western Blot. RESULTS: Cur inhibited the TRAP activity, release of degrading products from bone slices and the expression of NFATc1, c-Fos, Cathepsin K (Ctsk) and matrix metallopeptidase 9 (MMP9) of OCs induced with RANKL and H2O2. In addition, Cur suppressed the ROS level and NADPH oxidase 1(NOX1) and NADPH oxidase 4 (NOX4) activities of OCS. More importantly, Cur enhanced the expression and nucleus translocation of Nrf2 and activities of its regulatory cytoprotective enzymes, and reduced the NF-κB expression and phosphorylation and nucleus translocation of p65 in OCs. Furthermore, the Nrf2 inhibitor ML385 and NF-κB inhibitor Bay11-7082 counteracted the effect of Cur in OCs. CONCLUSION: Cur mitigated oxidative stress and osteoclastogenesis by activating Nrf2 and inhibiting the NF-κB pathway, suggesting that Cur may prove to be a promising candidate for the treatment of osteoporosis. Our findings may also help partially explain the rationale behind the traditional use of Curculigo orchioides Gaertn.
Assuntos
Benzoatos/farmacologia , Glucosídeos/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Osteogênese/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Acetilcisteína/farmacologia , Actinas/antagonistas & inibidores , Actinas/metabolismo , Animais , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/metabolismo , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Peróxido de Hidrogênio/farmacologia , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Camundongos , NADPH Oxidase 1/metabolismo , NADPH Oxidase 2/metabolismo , NADPH Oxidase 4/metabolismo , NF-kappa B/antagonistas & inibidores , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Ligante RANK/farmacologia , Células RAW 264.7 , Espécies Reativas de Oxigênio/metabolismo , Fosfatase Ácida Resistente a Tartarato/metabolismoRESUMO
RATIONALE: Higenamine (HG), is one of the main active components in many widely used Chinese herbs, and a common ingredient of health products in Europe and North America. Several groups, including our own, have previously shown the beneficial effects of HG against cardiomyocyte death during acute ischemic damage. However, the effect of HG on chronic cardiac remodeling, such as cardiac fibrosis, remains unknown. OBJECTIVE: Herein, we aim to investigate the role of HG in cardiac fibrosis in vivo as well as its cellular and molecular mechanisms. METHODS AND RESULTS: Chronic pressure overload with transverse aortic constriction (TAC) significantly increased cardiac hypertrophy, fibrosis, and cardiac dysfunction in mice, which were significantly attenuated by HG. Consistently, cardiac fibrosis induced by the chronic infusion of isoproterenol (ISO), was also significantly reduced by HG. Interestingly, our results showed that HG had no effect on adult mouse CM hypertrophy in vitro. However, HG suppressed the activation of cardiac fibroblasts (CFs) in vitro. Furthermore, TGF-ß1-induced expression of ACTA2, a marker of fibroblast activation, was significantly suppressed by HG. Concomitantly, HG inhibited TGF-ß1-induced phosphorylation of Smad2/3 in CFs. HG also reduced the expression of extracellular matrix molecules such as collagen I and collagen III. To our surprise, the inhibitory effect of HG on CFs activation was independent of the activation of the beta2 adrenergic receptor (ß2-AR) that is known to mediate the effect of HG on antagonizing CMs apoptosis. CONCLUSION: Our findings suggest that HG ameliorates pathological cardiac fibrosis and dysfunction at least partially by suppressing TGF-ß1/Smad signaling and CFs activation.
Assuntos
Alcaloides/farmacologia , Fibrinolíticos/farmacologia , Fibroblastos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/antagonistas & inibidores , Tetra-Hidroisoquinolinas/farmacologia , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Actinas/antagonistas & inibidores , Agonistas Adrenérgicos beta , Animais , Aorta/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Cardiomegalia/induzido quimicamente , Cardiomegalia/prevenção & controle , Fibrose/prevenção & controle , Cardiopatias/induzido quimicamente , Cardiopatias/prevenção & controle , Hipertensão/induzido quimicamente , Hipertensão/tratamento farmacológico , Isoproterenol , Camundongos , Ratos , Ratos Sprague-DawleyRESUMO
ACTG1 is a member of the actin family but is not a muscle actin gene. The ACTG1 mutation leads to hearing loss in humans, and the knockdown of ACTG1 suppresses the proliferation and migration of tumor cells; however, its role in intervertebral disc degeneration (IDD) is yet unclear. Bioinformatics methods revealed that ACTG1 might be a hub gene in IDD. Furthermore, the expression ACTG1 in severely degenerated nucleus pulposus (NP) tissues (Pfirrmann grade IV and V) was low as compared to that in mildly degenerated samples (Pfirrmann grade II and III). Moreover, the ACTG1 level was negatively correlated with human disc degeneration grades. The low expression of ACTG1 is also found in degenerated NP tissues in the rat. To further explore the function of ACTG1 in IDD, the gene expression was depleted in human NP cells via siRNA transfection. The ablation of ACTG1 increased MMP3 expression but decreased the level of collagen II. Excessive apoptosis was observed in ACTG1 knockdown groups, indicating that the absence of ACTG1 exacerbated IDD. GO function and pathway enrichment analysis for differentially expressed genes (DEGs) of two microarray datasets (GSE56081 and GSE42611) indicated that inflammatory response plays a crucial role in IDD. Interestingly, in the protein-protein interaction (PPI) network, ACTG1 is connected to the proteins of inflammation-related pathways. Furthermore, ACTG1 knockdown upregulated P-P65 level but suppressed P-Akt expression. These data collectively demonstrated that ACTG1 regulated the development of IDD through the NF-κB-p65 and Akt pathways, and ACTG1 may be a novel marker and therapeutic target of IDD in the future.
Assuntos
Actinas/genética , Actinas/metabolismo , Degeneração do Disco Intervertebral/genética , Degeneração do Disco Intervertebral/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição RelA/metabolismo , Actinas/antagonistas & inibidores , Adulto , Idoso , Animais , Células Cultivadas , Modelos Animais de Doenças , Feminino , Técnicas de Silenciamento de Genes , Humanos , Degeneração do Disco Intervertebral/patologia , Masculino , Pessoa de Meia-Idade , Núcleo Pulposo/metabolismo , Núcleo Pulposo/patologia , Mapas de Interação de Proteínas , RNA Interferente Pequeno/genética , Ratos , Ratos Sprague-Dawley , Transdução de SinaisRESUMO
OBJECTIVE: In recent years, the roles of Neuregulin-1 (NRG-1) in optic nerve injury and retinal cells have been investigated. However, the molecular mechanism by which NRG-1 affects optic nerve injury remains elusive and merits deeper exploration. Hence, this study examined the specific function of NRG-1 in the RhoA/cofilin/F-actin axis in optic nerve injury. METHODS: Retinal cells were isolated and identified for subsequent experimental uses. Reverse transcription quantitative polymerase chain reaction and Western blot assays were performed to measure NRG-1 expression in retinal cells which were cultured under elevated pressure. TUNEL staining was used to detect the cell apoptosis rate, and Western blot assay was performed to detect the expression of related genes. The axon growth was examined by immunofluorescence. The effects of NRG-1 on RhoA activity, cofilin phosphorylation, and F-actin were detected by Western blot assay. In other studies we established a rat model of acute optic nerve injury, and tested for beneficial effects of NRG-1 in vivo. RESULTS: High expression of NRG-1 was evident in the retinal tissues of rats with optic nerve injury. Overexpressing NRG-1 successfully inhibited RhoA activity and the phosphorylation of cofilin and promoted F-actin expression. In cell experiments, overexpressed NRG-1 suppressed the apoptosis of retinal cells and promoted axon growth through the RhoA/cofilin/F-actin axis. In animal experiments, overexpressed NRG-1 relieved retinal injury. CONCLUSION: Our results strongly suggest that overexpressed NRG-1 is highly effective in the protection of normal optic nerve function by suppressing RhoA activity and the phosphorylation of cofilin and rescuing F-actin function.
Assuntos
Fatores de Despolimerização de Actina/metabolismo , Actinas/metabolismo , Neuregulina-1/biossíntese , Traumatismos do Nervo Óptico/metabolismo , Traumatismos do Nervo Óptico/prevenção & controle , Proteínas rho de Ligação ao GTP/metabolismo , Fatores de Despolimerização de Actina/antagonistas & inibidores , Actinas/antagonistas & inibidores , Animais , Células Cultivadas , Masculino , Fármacos Neuroprotetores/metabolismo , Ratos , Ratos Sprague-Dawley , Regulação para Cima/fisiologia , Proteínas rho de Ligação ao GTP/antagonistas & inibidoresRESUMO
The interaction of actin and myosin is essential for cell migration. We have identified kaempferol and pentahalogenated pseudilins as efficient inhibitors of migration of MDA-MB-231 breast adenocarcinoma cells. The compounds were studied with respect to possible effects on myosin-2-ATPase activity. The pentahalogenated pseudilins inhibited the enzyme activity in vitro. Flavonoids showed no effect on enzyme activity. The polymerization dynamics of actin was measured to test whether the integrity of F-actin is essential for the migration of MDA-MB-231 cells. Quercetin and kaempferol depolymerized F-actin with similar efficiencies as found for the pentahalogenated pseudilins, whereas epigallocatechin showed the weakest effect. As the inhibitory effect on cell migration may be caused by a toxic effect, we have performed a cytotoxicity test and, furthermore, investigated the influence of the test compounds on cardiac function in eleutheroembryos of medaka (Oryzias latipes). Compared with the pentahalogenated pseudilins, the cytotoxic and cardiotoxic effects of flavonoids on medaka embryos were found to be moderate.
Assuntos
Actinas/antagonistas & inibidores , Quempferóis/farmacologia , Miosinas/antagonistas & inibidores , Quercetina/farmacologia , Actinas/metabolismo , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Quempferóis/química , Estrutura Molecular , Miosinas/metabolismo , Quercetina/química , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Human adipose-derived stem cells (hASCs) are ideal seed cells for tissue engineering due to their multidirectional differentiation potential. Microfilaments, microtubules, and intermediate filaments are responsible for supporting the intracellular space. Vimentin, a type III intermediate filament protein that is specifically expressed in cells of mesenchymal origin, can function as a scaffold and endow cells with tension and shear stress resistance. Actin stress fibers (ASF) act as an important physical device in stress signal transduction, providing stiffness for cells, and promoting osteogenesis. Through direct physical contact, cross-linkers, and spatial interactions, vimentin and actin networks exist as intersecting entities. Spatial interactions occur in the overlapping area of cytoskeleton subsystems, which could affect cell morphology, cell mechanics, and cell fate. However, how does the spatial organization between the cytoskeletal subsystems changed during osteogenesis, especially between vimentin and ASF, is still not understood, and its mechanism effect on cell fate remains unclear. In our study, WB experiment was used to detect the expression changes in Vimentin, ASF, and other proteins. Cells were reconstructed by three-dimensional scanning with fluorescence microscope, and the spatial thickness of vimentin and ASF cytoskeletons and the thickness of the overlapping area between them were calculated, respectively, so as to observe the spatial reorganization of vimentin and ASF in cells. Cytochalasin D (an inhibitor of actin polymerization) and vimentin upregulated/downregulated cells were used to verify the change in the spatial organization between vimentin and ASF and its influence on osteogenesis. Then, heat shock protein 27 (HSP27) was downregulated to illuminate the regulatory mechanisms of spatial organization between vimentin and ASF during osteogenesis. The amounts and the spatial positions of vimentin and actin stress fiber exhibited opposite trends during osteogenesis. Through controlling the anchor sites on the nucleus, intermediate filaments vimentin can reduce the spatial proportion of actin stress fibers, which can be regulated by HSP27. In addition, depolymerization of actin stress fibers lead to lower osteogenic differentiation ability, resulting in osteogenesis and lipogenesis existed simultaneously, that can be resisted by vimentin. Our data indicate that the spatial reorganization of vimentin and actin stress fibers is a key factor in the regulation of the differentiation state of hASCs. And their spatial overlapping area is detrimental to hASCs osteogenesis, providing a new perspective for further exploring the mechanism underlying hASCs osteogenesis.
Assuntos
Actinas/metabolismo , Tecido Adiposo/citologia , Diferenciação Celular/genética , Osteogênese/genética , Transdução de Sinais/genética , Células-Tronco/metabolismo , Fibras de Estresse/metabolismo , Vimentina/metabolismo , Actinas/antagonistas & inibidores , Diferenciação Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Células Cultivadas , Citocalasina D/farmacologia , Citoplasma/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Filamentos Intermediários/metabolismo , Microscopia de Fluorescência , Microtúbulos/metabolismo , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Osteogênese/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transfecção , Vimentina/genéticaRESUMO
Clear-cell renal cell carcinoma (ccRCC), the most common subtype of renal cancer, has a poor clinical outcome. A hallmark of ccRCC is genetic loss-of-function of VHL (von Hippel-Lindau) that leads to a highly vascularized tumor microenvironment. Although many ccRCC patients initially respond to antiangiogenic therapies, virtually all develop progressive, drug-refractory disease. Given the role of dysregulated expressions of cytoskeletal and cytoskeleton-regulatory proteins in tumor progression, we performed analyses of The Cancer Genome Atlas (TCGA) transcriptome data for different classes of actin-binding proteins to demonstrate that increased mRNA expression of profilin1 (Pfn1), Arp3, cofilin1, Ena/VASP, and CapZ, is an indicator of poor prognosis in ccRCC. Focusing further on Pfn1, we performed immunohistochemistry-based classification of Pfn1 staining in tissue microarrays, which indicated Pfn1 positivity in both tumor and stromal cells; however, the vast majority of ccRCC tumors tend to be Pfn1-positive selectively in stromal cells only. This finding is further supported by evidence for dramatic transcriptional up-regulation of Pfn1 in tumor-associated vascular endothelial cells in the clinical specimens of ccRCC. In vitro studies support the importance of Pfn1 in proliferation and migration of RCC cells and in soluble Pfn1's involvement in vascular endothelial cell tumor cell cross-talk. Furthermore, proof-of-concept studies demonstrate that treatment with a novel computationally designed Pfn1-actin interaction inhibitor identified herein reduces proliferation and migration of RCC cells in vitro and RCC tumor growth in vivo Based on these findings, we propose a potentiating role for Pfn1 in promoting tumor cell aggressiveness in the setting of ccRCC.
Assuntos
Carcinoma de Células Renais/patologia , Neoplasias Renais/patologia , Profilinas/metabolismo , Actinas/antagonistas & inibidores , Actinas/metabolismo , Animais , Proteína de Capeamento de Actina CapZ/genética , Proteína de Capeamento de Actina CapZ/metabolismo , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Cofilina 1/genética , Cofilina 1/metabolismo , Bases de Dados Genéticas , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Humanos , Neoplasias Renais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Profilinas/antagonistas & inibidores , Profilinas/genética , Prognóstico , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Microambiente Tumoral , Regulação para CimaRESUMO
Actin's interactions with myosin and other actin-binding proteins are essential for cellular viability in numerous cell types, including muscle. In a previous high-throughput time-resolved FRET (TR-FRET) screen, we identified a class of compounds that bind to actin and affect actomyosin structure and function. For clinical utility, it is highly desirable to identify compounds that affect skeletal and cardiac muscle differently. Because actin is more highly conserved than myosin and most other muscle proteins, most such efforts have not targeted actin. Nevertheless, in the current study, we tested the specificity of the previously discovered actin-binding compounds for effects on skeletal and cardiac α-actins as well as on skeletal and cardiac myofibrils. We found that a majority of these compounds affected the transition of monomeric G-actin to filamentous F-actin, and that several of these effects were different for skeletal and cardiac actin isoforms. We also found that several of these compounds affected ATPase activity differently in skeletal and cardiac myofibrils. We conclude that these structural and biochemical assays can be used to identify actin-binding compounds that differentially affect skeletal and cardiac muscles. The results of this study set the stage for screening of large chemical libraries for discovery of novel compounds that act therapeutically and specifically on cardiac or skeletal muscle.
Assuntos
Actinas , Músculo Esquelético/metabolismo , Miocárdio/metabolismo , Miofibrilas/metabolismo , Miosinas , Bibliotecas de Moléculas Pequenas , Actinas/antagonistas & inibidores , Actinas/química , Actinas/metabolismo , Animais , Bovinos , Avaliação Pré-Clínica de Medicamentos , Transferência Ressonante de Energia de Fluorescência , Miosinas/química , Miosinas/metabolismo , CoelhosRESUMO
The uterus plays an important and unique role during pregnancy and is a dynamic organ subjected to mechanical stimuli. It has been reported that infertility occurs when the peristalsis is prevented, although its mechanisms remain unknown. In this study, we found that mechanical strain mimicking the peristaltic motion of the uterine smooth muscle layer enabled the endometrial stromal cells to acquire contractility. In order to mimic the peristalsis induced by uterine smooth muscle cells, cyclic tensile stretch was applied to human endometrial stromal cells. The results showed that the strained cells exerted greater contractility in three-dimensional collagen gels in the presence of oxytocin, due to up-regulated alpha-smooth muscle actin expression via the cAMP signaling pathway. These in vitro findings underscore the plasticity of the endometrial stromal cell phenotype and suggest the possibility of acquired contractility by these cells in vivo and its potential contribution to uterine contractile activity. This phenomenon may be a typical example of how a tissue passively acquires new contractile functions under mechanical stimulation from a neighboring tissue, enabling it to support the adjacent tissue's functions.
Assuntos
Endométrio/citologia , Miócitos de Músculo Liso/citologia , Células Estromais/fisiologia , Resistência à Tração , Actinas/antagonistas & inibidores , Actinas/metabolismo , Adulto , Células Cultivadas , Colágeno Tipo I/metabolismo , AMP Cíclico/metabolismo , Feminino , Regulação da Expressão Gênica , Humanos , Ionomicina/farmacologia , Isoquinolinas/farmacologia , Pessoa de Meia-Idade , Músculo Liso , Ocitocina/farmacologia , Peristaltismo , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Sulfonamidas/farmacologia , Regulação para CimaRESUMO
We report benzeneselenol-modified gold nanoclusters (Se_Au NCs) that have significant anticancer activity by inducing autophagy and disassembly of F-actin in cancer cells. Upon treatment of different tumor models on mice, it has been found that Se_Au NCs exhibit excellent antitumor efficacy, high stability, and extremely low toxicity, which are the requirements for an ideal antineoplastic drug. Our study is a successful attempt in exploring novel anticancer agents with high biosafety.
Assuntos
Antineoplásicos/farmacologia , Derivados de Benzeno/farmacologia , Doxorrubicina/farmacologia , Ouro/farmacologia , Nanopartículas Metálicas/química , Compostos Organosselênicos/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Actinas/antagonistas & inibidores , Actinas/metabolismo , Animais , Antineoplásicos/química , Autofagia/efeitos dos fármacos , Derivados de Benzeno/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Doxorrubicina/química , Portadores de Fármacos/química , Portadores de Fármacos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Ouro/química , Humanos , Camundongos , Neoplasias Experimentais/diagnóstico por imagem , Neoplasias Experimentais/tratamento farmacológico , Imagem Óptica , Compostos Organosselênicos/química , Neoplasias Pancreáticas/diagnóstico por imagem , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Memory transience is essential to gain cognitive flexibility. Recently, hippocampal neurogenesis is emerging as one of the mechanisms involved in the balance between persistence and forgetting. Social recognition memory (SRM) has its duration prolonged by neurogenesis. However, it is still to be determined whether boosting neurogenesis in distinct phases of SRM may favor forgetting over persistence. In the present study, we used enriched environment (EE) and memantine (MEM) to increase neurogenesis. SRM was ubiquitously prolonged by both, while EE after the memory acquisition did not favor forgetting. Interestingly, the proportion of newborn neurons with mature morphology in the dorsal hippocampus was higher in animals where persistence prevailed. Finally, one of the main factors for dendritic growth is the formation of cytoskeleton. We found that Latrunculin A, an inhibitor of actin polymerization, blunted the promnesic effect of EE. Altogether, our results indicate that the mechanisms triggered by EE to improve SRM are not limited to increasing the number of newborn neurons.
Assuntos
Memória/fisiologia , Neurogênese/fisiologia , Neurônios/fisiologia , Reconhecimento Psicológico/fisiologia , Comportamento Social , Actinas/antagonistas & inibidores , Amnésia/induzido quimicamente , Amnésia/psicologia , Animais , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Dendritos/efeitos dos fármacos , Proteína Duplacortina , Meio Ambiente , Antagonistas de Aminoácidos Excitatórios/farmacologia , Hipocampo/citologia , Hipocampo/efeitos dos fármacos , Imuno-Histoquímica , Memantina/farmacologia , Consolidação da Memória/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Tiazolidinas/farmacologiaRESUMO
BACKGROUND: The protein plasminogen activator inhibitor-1 (PAI-1), an inhibitor specific for urokinase plasminogen activator (uPA) and tissue plasminogen activator (tPA), has been shown to have a key role in cancer metastases. Currently, it is unknown as to whether the exocellular inhibition of PAI-1 can inhibit the migration of cancer cells. METHODS: By fusing the mutated serine protease domain (SPD) of uPA and human serum albumin (HSA), PAItrap3, a protein that traps PAI-1, was synthesized and experiments were conducted to determine if exocellular PAItrap3 attenuates PAI-1-induced cancer cell migration in vitro. RESULTS: PAItrap3 (0.8 µM) significantly inhibited the motility of MCF-7, MDA-MB-231, HeLa and 4T1 cancer cells, by 90%, 50%, 30% and 20%, respectively, without significantly altering their proliferation. The PAI-1-induced rearrangement of F-actin was significantly inhibited by PAItrap3, which produced a decrease in the number of cell protrusions by at least 20%. CONCLUSIONS: In vitro, PAItrap3 inhibited PAI-1-induced cancer cell migration, mainly through inhibiting the rearrangement of F-actin. Overall, these results, provided they can be extrapolated to humans, suggest that the PAItrap3 protein could be used as an exocellular inhibitor to attenuate cancer metastases.