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1.
Phys Rev Lett ; 127(10): 108101, 2021 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-34533352

RESUMO

We investigate the rheological properties of interpenetrating networks reconstituted from the main cytoskeletal components: filamentous actin, microtubules, and vimentin intermediate filaments. The elastic modulus is determined largely by actin, with little contribution from either microtubules or vimentin. However, vimentin dramatically impacts the relaxation, with even small amounts significantly increasing the relaxation time of the interpenetrating network. This highly unusual decoupling between dissipation and elasticity may reflect weak attractive interactions between vimentin and actin networks.


Assuntos
Filamentos Intermediários/química , Modelos Químicos , Vimentina/química , Actinas/química , Actinas/metabolismo , Citoesqueleto/química , Citoesqueleto/metabolismo , Células Eucarióticas , Filamentos Intermediários/metabolismo , Microtúbulos/química , Microtúbulos/metabolismo , Reologia/métodos , Vimentina/metabolismo
2.
Nat Struct Mol Biol ; 28(8): 681-693, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34373646

RESUMO

The synaptonemal complex (SC) is a supramolecular protein assembly that mediates synapsis between homologous chromosomes during meiosis. SC elongation along the chromosome length (up to 24 µm) depends on its midline α-fibrous component SYCE2-TEX12. Here, we report X-ray crystal structures of human SYCE2-TEX12 as an individual building block and on assembly within a fibrous lattice. We combine these structures with mutagenesis, biophysics and electron microscopy to reveal the hierarchical mechanism of SYCE2-TEX12 fiber assembly. SYCE2-TEX12's building blocks are 2:2 coiled coils that dimerize into 4:4 hetero-oligomers and interact end-to-end and laterally to form 10-nm fibers that intertwine within 40-nm bundled micrometer-long fibers that define the SC's midline structure. This assembly mechanism bears striking resemblance with intermediate filament proteins vimentin, lamin and keratin. Thus, SYCE2-TEX12 exhibits behavior typical of cytoskeletal proteins to provide an α-fibrous SC backbone that structurally underpins synaptic elongation along meiotic chromosomes.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Pareamento Cromossômico/fisiologia , Complexo Sinaptonêmico/metabolismo , Cristalografia por Raios X , Proteínas do Citoesqueleto/metabolismo , Humanos , Queratinas/metabolismo , Laminas/metabolismo , Meiose/fisiologia , Estrutura Quaternária de Proteína , Vimentina/metabolismo
3.
Biomolecules ; 11(8)2021 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-34439816

RESUMO

Cataracts are a leading cause of blindness worldwide. Surgical removal of cataracts is a safe and effective procedure to restore vision. However, a large number of patients later develop vision loss due to regrowth of lens cells and subsequent degradation of the visual axis leading to visual disability. This postsurgical complication, known as posterior capsular opacification (PCO), occurs in up to 30% of cataract patients and has no clinically proven pharmacological means of prevention. Despite the availability of many compounds capable of preventing early steps in PCO development, there is currently no effective means to deliver such therapies into the eye for a suitable duration. To model a solution to this unmet medical need, we fabricated acrylic substrates as intraocular lens (IOL) mimics scaled to place into the capsular bag of the mouse lens following a mock-cataract surgery. Substrates were coated with a hydrophilic crosslinked acrylate nanogel designed to elute Sorbinil, an aldose reductase inhibitor previously shown to suppress PCO. Insertion of the Sorbinil-eluting device into the lens capsule at the time of cataract surgery resulted in substantial prevention of cellular changes associated with PCO development. This model demonstrates that a cataract inhibitor can be delivered into the postsurgical lens capsule at therapeutic levels.


Assuntos
Opacificação da Cápsula/prevenção & controle , Extração de Catarata/efeitos adversos , Portadores de Fármacos , Inibidores Enzimáticos/farmacologia , Imidazolidinas/farmacologia , Lentes Intraoculares , Actinas/genética , Actinas/metabolismo , Animais , Caderinas/genética , Caderinas/metabolismo , Opacificação da Cápsula/etiologia , Opacificação da Cápsula/genética , Opacificação da Cápsula/patologia , Catarata/genética , Catarata/metabolismo , Catarata/patologia , Extração de Catarata/métodos , Modelos Animais de Doenças , Fibronectinas/genética , Fibronectinas/metabolismo , Regulação da Expressão Gênica , Humanos , Cristalino/metabolismo , Cristalino/patologia , Cristalino/cirurgia , Camundongos , Nanogéis/administração & dosagem , Nanogéis/química , Transdução de Sinais , Vimentina/genética , Vimentina/metabolismo
4.
Exp Cell Res ; 406(1): 112759, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34332984

RESUMO

The Serum Amyloid A (SAA) family of proteins is associated with various pathological conditions, including cancer. However, their role in cancer is incompletely understood. Here, we investigated the role of SAA1 in cell cycle regulation, apoptosis, survival signaling, metabolism, and metastasis in models of triple-negative breast cancer (TNBC), using RNAi. Our data show that in untransformed epithelial cells (MCF12A), the knockdown of SAA1 induces the expression of cell cycle regulators (MCM2, p53), the activation of DNA repair (PARP synthesis), and survival signaling (NFκB). In contrast, knockdown of SAA1 in the TNBC cell line (MDA-MB-231) induced the expression p16 and shifted cells in the cell cycle from the S to G2/M phase, without the activation of DNA repair. Moreover, in SAA1-deficient MDA-MB-231 and HCC70 cells, metabolism (NADH oxidation) continually increased while cell migration (% wound closure and the rate of wound closure) decreased. However, silencing of SAA1 altered epithelial and mesenchymal markers in MCF12A (E-cadherin, Laminin 1ß, Vimentin) and MDA-MB-231 (α-Smooth muscle actin) cells, associated with the metastatic program of epithelial-mesenchymal transition. Nonetheless, our data provide evidence that SAA1 could potentially serve as a therapeutic target in TNBC.


Assuntos
Apoptose/genética , Ciclo Celular/genética , Movimento Celular/genética , Células Epiteliais/metabolismo , Proteína Amiloide A Sérica/genética , Actinas/genética , Actinas/metabolismo , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Células Epiteliais/patologia , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , Humanos , Laminina/genética , Laminina/metabolismo , Componente 2 do Complexo de Manutenção de Minicromossomo/genética , Componente 2 do Complexo de Manutenção de Minicromossomo/metabolismo , Modelos Biológicos , NF-kappa B/genética , NF-kappa B/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína Amiloide A Sérica/antagonistas & inibidores , Proteína Amiloide A Sérica/metabolismo , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Vimentina/genética , Vimentina/metabolismo
5.
Nan Fang Yi Ke Da Xue Xue Bao ; 41(7): 995-1001, 2021 Jul 20.
Artigo em Chinês | MEDLINE | ID: mdl-34308848

RESUMO

OBJECTIVE: To explore the role of human short stature homeobox 2 (SHOX2) in regulating the migration, invasion and stemness of human bladder cancer cells. METHODS: We analyzed SHOX2 gene expression in bladder cancer and adjacent tissues based on TCGA database. Univariate survival analysis of SHOX2 gene expression in TCGA-BLCA data was performed using GEPIA. The probable function of SHOX2 was predicted using GSEA. Human bladder cancer T24 cell models of SHOX2 knockdown or overexpression were assessed for changes in migration and invasion abilities using wound healing assay and Transwell assay, and their cancer stem cell-like characteristics were evaluated using tumorsphere formation assay and colony formation assay. Western blotting was used to detect the expressions of epithelial mesenchymal transition (EMT) markers Ecadherin and vimentin and the TGF-ß signaling network component TßR-I in the cells. RESULTS: SHOX2 expression was significantly higher in bladder cancer tissues than in the adjacent tissues (P < 0.05), especially in paired tissue specimens (P < 0.01), and was negatively correlated with the overall survival of the patients (P < 0.05). SHOX2 gene expression was correlated positively with EMT-related (P < 0.05) and stemness-related gene signatures (P < 0.01). In T24 cells, SHOX2 knockdown significantly suppressed cell migration and invasion, which was significantly enhanced by SHOX2 overexpression (P < 0.01). The cancer stem cell-like characteristics of T24 cells was repressed by SHOX2 knockdown but significantly enhanced by SHOX2 overexpression (P < 0.01). SHOX2 knockdown induced morphological changes of the cells into epithelioid cells, whereas SHOX2 overexpression induced a mesenchymal morphology of the cells. SHOX2 knockdown increased E-cadherin expression and decreased vimentin and TßR-I expression, while SHOX2 overexpression increased the expressions of vimentin and TßR-I in the cells. CONCLUSION: SHOX2 promotes the migration, invasion and stemness of human bladder cancer cells possibly by regulating EMT via the TGF-ß signaling pathway.


Assuntos
Neoplasias da Bexiga Urinária , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/genética , Humanos , Neoplasias da Bexiga Urinária/genética , Vimentina/genética , Vimentina/metabolismo
6.
Int J Mol Sci ; 22(13)2021 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-34201927

RESUMO

Polylactide (PLA), widely used in bioengineering and medicine, gained popularity due to its biocompatibility and biodegradability. Natural origin and eco-friendly background encourage the search of novel materials with such features, such as polyhydroxyoctanoate (P(3HO)), a polyester of bacterial origin. Physicochemical features of both P(3HO) and PLA have an impact on cellular response 32, i.e., adhesion, migration, and cell morphology, based on the signaling and changes in the architecture of the three cytoskeletal networks: microfilaments (F-actin), microtubules, and intermediate filaments (IF). To investigate the role of IF in the cellular response to the substrate, we focused on vimentin intermediate filaments (VIFs), present in mouse embryonic fibroblast cells (MEF). VIFs maintain cell integrity and protect it from external mechanical stress, and also take part in the transmission of signals from the exterior of the cell to its inner organelles, which is under constant investigation. Physiochemical properties of a substrate have an impact on cells' morphology, and thus on cytoskeleton network signaling and assembly. In this work, we show how PLA and P(3HO) crystallinity and hydrophilicity influence VIFs, and we identify that two different types of vimentin cytoskeleton architecture: network "classic" and "nutshell-like" are expressed by MEFs in different numbers of cells depending on substrate features.


Assuntos
Materiais Biocompatíveis , Citoesqueleto/metabolismo , Poliésteres , Vimentina/metabolismo , Animais , Materiais Biocompatíveis/química , Linhagem Celular , Células Cultivadas , Citoesqueleto/química , Fibroblastos/metabolismo , Imunofluorescência , Expressão Gênica , Camundongos , Vimentina/genética
7.
Chem Biol Interact ; 346: 109570, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34217686

RESUMO

Verapamil is reported to prevent scar formation. However, whether verapamil is involved in the ureteral stricture scar and the underlying mechanism need further investigation. Fibroblasts were isolated from ureteral scar tissues. TGF-ß1 stimulation was used to induce fibrosis of fibroblasts. Inhibition of CaMK II was achieved by shRNA transfection. CCK-8 was performed to evaluate cell viability. qRT-PCR was applied to determine the level of mRNA while western blotting was used to determine the level of proteins. Immunofluorescence was used to detect the level of vimentin, collagen I and collagen III. Primary fibroblasts was successfully isolated from ureteral scar tissues. TGF-ß1 stimulation was capable to induce collagen production and fibrosis in primary fibroblasts while inhibition of CaMK II attenuate collagen production. Overexpression of wild type CaMK II lead to further increase of collagen production upon TGF-ß1 stimulation while the mutated CaMK II did not exert this promotion. Treatment of verapamil inhibits TGF-ß1 induced collagen production via inhibiting CaMK II. In present study, we revealed a vital role of Verapamil and CaMK II in the formation of ureteral scar. Verapamil inhibited TGF-ß1 induced collagen fiber formation by regulating CaMK II. Our finding might provide new insight into mechanism of prevention and treatment of ureteral scar.


Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Verapamil/farmacologia , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/antagonistas & inibidores , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Células Cultivadas , Cicatriz/tratamento farmacológico , Cicatriz/metabolismo , Cicatriz/patologia , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Mutagênese , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Regulação para Cima/efeitos dos fármacos , Verapamil/uso terapêutico , Vimentina/metabolismo
8.
Int J Mol Sci ; 22(14)2021 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-34299089

RESUMO

The cytoskeletal protein vimentin is secreted under various physiological conditions. Extracellular vimentin exists primarily in two forms: attached to the outer cell surface and secreted into the extracellular space. While surface vimentin is involved in processes such as viral infections and cancer progression, secreted vimentin modulates inflammation through reduction of neutrophil infiltration, promotes bacterial elimination in activated macrophages, and supports axonal growth in astrocytes through activation of the IGF-1 receptor. This receptor is overexpressed in cancer cells, and its activation pathway has significant roles in general cellular functions. In this study, we investigated the functional role of extracellular vimentin in non-tumorigenic (MCF-10a) and cancer (MCF-7) cells through the evaluation of its effects on cell migration, proliferation, adhesion, and monolayer permeability. Upon treatment with extracellular recombinant vimentin, MCF-7 cells showed increased migration, proliferation, and adhesion, compared to MCF-10a cells. Further, MCF-7 monolayers showed reduced permeability, compared to MCF-10a monolayers. It has been shown that the receptor binding domain of SARS-CoV-2 spike protein can alter blood-brain barrier integrity. Surface vimentin also acts as a co-receptor between the SARS-CoV-2 spike protein and the cell-surface angiotensin-converting enzyme 2 receptor. Therefore, we also investigated the permeability of MCF-10a and MCF-7 monolayers upon treatment with extracellular recombinant vimentin, and its modulation of the SARS-CoV-2 receptor binding domain. These findings show that binding of extracellular recombinant vimentin to the cell surface enhances the permeability of both MCF-10a and MCF-7 monolayers. However, with SARS-CoV-2 receptor binding domain addition, this effect is lost with MCF-7 monolayers, as the extracellular vimentin binds directly to the viral domain. This defines an influence of extracellular vimentin in SARS-CoV-2 infections.


Assuntos
Neoplasias da Mama/patologia , Mama/patologia , Permeabilidade da Membrana Celular , Matriz Extracelular/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Vimentina/metabolismo , Mama/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Células Cultivadas , Feminino , Humanos , Domínios Proteicos , Glicoproteína da Espícula de Coronavírus/genética , Vimentina/genética
9.
Nutrients ; 13(5)2021 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-34069461

RESUMO

Investigating dietary polyphenolic compounds as antitumor agents are rising due to the growing evidence of the close association between immunity and cancer. Cancer cells elude immune surveillance for enhancing their progression and metastasis utilizing various mechanisms. These mechanisms include the upregulation of programmed death-ligand 1 (PD-L1) expression and Epithelial-to-Mesenchymal Transition (EMT) cell phenotype activation. In addition to its role in stimulating normal embryonic development, EMT has been identified as a critical driver in various aspects of cancer pathology, including carcinogenesis, metastasis, and drug resistance. Furthermore, EMT conversion to another phenotype, Mesenchymal-to-Epithelial Transition (MET), is crucial in developing cancer metastasis. A central mechanism in the upregulation of PD-L1 expression in various cancer types is EMT signaling activation. In breast cancer (BC) cells, the upregulated level of PD-L1 has become a critical target in cancer therapy. Various signal transduction pathways are involved in EMT-mediated PD-L1 checkpoint overexpression. Three main groups are considered potential targets in EMT development; the effectors (E-cadherin and Vimentin), the regulators (Zeb, Twist, and Snail), and the inducers that include members of the transforming growth factor-beta (TGF-ß). Meanwhile, the correlation between consuming flavonoid-rich food and the lower risk of cancers has been demonstrated. In BC, polyphenols were found to downregulate PD-L1 expression. This review highlights the effects of polyphenols on the EMT process by inhibiting mesenchymal proteins and upregulating the epithelial phenotype. This multifunctional mechanism could hold promises in the prevention and treating breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Polifenóis/farmacologia , Receptor de Morte Celular Programada 1/metabolismo , Antígenos CD , Antígeno B7-H1/metabolismo , Neoplasias da Mama/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Receptor de Morte Celular Programada 1/genética , Transdução de Sinais , Fator de Crescimento Transformador beta , Vimentina/metabolismo
10.
Cell Biochem Funct ; 39(6): 813-820, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34128234

RESUMO

Prostate cancer (PCa) is the second most prevalent cancer in men worldwide. Most cases of death from PCa are due to metastasis. Early stages of metastasis are mediated by epithelial-mesenchymal transition (EMT) process through which cancer cells acquire motility and invasive characteristics. Thus, more potent and novel therapeutic strategies must be designed based on the inhibition of EMT or metastasis. Herein, we employ a co-culture system to evaluate the anti-EMT effects of human amniotic mesenchymal stromal cells (hAMSCs) on LNCaP PCa cells. The RNA of treated (sample) and untreated cancer cells (control) and whole-cell lysates of related cells were prepared and analysed through quantitative real-time polymerase chain reaction (qRT-PCR) and western blot, respectively. Based on the results, the expression of vimentin, Snail and Zeb1 in LNCaP cells decreased and the expression of E-cadherin increased after treatment with hAMSCs. Furthermore, induction of the cellular apoptosis in LNCaP cells was detected. The anti-cancer activity of conditioned medium from hAMSCs was shown using hanging drop technique (a 3D cell culture model). Our findings support the idea that stem cells can be considered as a novel therapeutic approach to inhibit prostate cancer cells. SIGNIFICANCE OF THE STUDY: The anti-tumour activity of hAMSCs on LNCaP prostate cancer cells using 2D and 3D cell culture models via induction of apoptosis, suppression of EMT process and down-regulation of EGFR was shown. The results of the present study support this idea that hAMSCs may be a potent therapeutic tool to suppress tumour growth in LNCaP prostate cancer cells.


Assuntos
Apoptose/efeitos dos fármacos , Meios de Cultivo Condicionados/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Fatores de Transcrição da Família Snail/antagonistas & inibidores , Vimentina/antagonistas & inibidores , Homeobox 1 de Ligação a E-box em Dedo de Zinco/antagonistas & inibidores , Técnicas de Cocultura , Meios de Cultivo Condicionados/química , Regulação para Baixo/efeitos dos fármacos , Humanos , Células-Tronco Mesenquimais/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Células Tumorais Cultivadas , Vimentina/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo
11.
Redox Biol ; 44: 102014, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34062408

RESUMO

The cytoskeleton is a supramolecular structure consisting of interacting protein networks that support cell dynamics in essential processes such as migration and division, as well as in responses to stress. Fast cytoskeletal remodeling is achieved with the participation of regulatory proteins and posttranslational modifications (PTMs). Redox-related PTMs are emerging as critical players in cytoskeletal regulation. Here we used a cellular model of mild nitroxidative stress in which a peroxynitrite donor induced transient changes in the organization of three key cytoskeletal proteins, i.e., vimentin, actin and tubulin. Nitroxidative stress-induced reconfiguration of intermediate filaments, microtubules and actin structures were further correlated with their PTM profiles and dynamics of the PTM landscape. Using high-resolution mass spectrometry, 62 different PTMs were identified and relatively quantified in vimentin, actin and tubulin, including 12 enzymatic, 13 oxidative and 2 nitric oxide-derived modifications as well as 35 modifications by carbonylated lipid peroxidation products, thus evidencing the occurrence of a chain reaction with formation of numerous reactive species and activation of multiple signaling pathways. Our results unveil the presence of certain modifications under basal conditions and their modulation in response to stress in a target-, residue- and reactive species-dependent manner. Thus, some modifications accumulated during the experiment whereas others varied transiently. Moreover, we identified protein PTM "hot spots", such as the single cysteine residue of vimentin, which was detected in seven modified forms, thus, supporting its role in PTM crosstalk and redox sensing. Finally, identification of novel PTMs in these proteins paves the way for unveiling new cytoskeleton regulatory mechanisms.


Assuntos
Proteínas do Citoesqueleto , Processamento de Proteína Pós-Traducional , Proteínas do Citoesqueleto/metabolismo , Citoesqueleto/metabolismo , Filamentos Intermediários/metabolismo , Vimentina/metabolismo
12.
Nat Commun ; 12(1): 3799, 2021 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-34145230

RESUMO

The cytoskeleton determines cell mechanics and lies at the heart of important cellular functions. Growing evidence suggests that the manifold tasks of the cytoskeleton rely on the interactions between its filamentous components-actin filaments, intermediate filaments, and microtubules. However, the nature of these interactions and their impact on cytoskeletal dynamics are largely unknown. Here, we show in a reconstituted in vitro system that vimentin intermediate filaments stabilize microtubules against depolymerization and support microtubule rescue. To understand these stabilizing effects, we directly measure the interaction forces between individual microtubules and vimentin filaments. Combined with numerical simulations, our observations provide detailed insight into the physical nature of the interactions and how they affect microtubule dynamics. Thus, we describe an additional, direct mechanism by which cells establish the fundamental cross talk of cytoskeletal components alongside linker proteins. Moreover, we suggest a strategy to estimate the binding energy of tubulin dimers within the microtubule lattice.


Assuntos
Citoesqueleto de Actina/metabolismo , Filamentos Intermediários/metabolismo , Microtúbulos/metabolismo , Vimentina/metabolismo , Animais , Fenômenos Biofísicos/fisiologia , Citoesqueleto/metabolismo , Eletricidade Estática
13.
Int J Mol Sci ; 22(10)2021 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-34065633

RESUMO

The circadian clock coordinates biological and physiological functions to day/night cycles. The perturbation of the circadian clock increases cancer risk and affects cancer progression. Here, we studied how BMAL1 knockdown (BMAL1-KD) by shRNA affects the epithelial-mesenchymal transition (EMT), a critical early event in the invasion and metastasis of colorectal carcinoma (CRC). In corresponding to a gene set enrichment analysis, which showed a significant enrichment of EMT and invasive signatures in BMAL1_high CRC patients as compared to BMAL1_low CRC patients, our results revealed that BMAL1 is implicated in keeping the epithelial-mesenchymal equilibrium of CRC cells and influences their capacity of adhesion, migration, invasion, and chemoresistance. Firstly, BMAL1-KD increased the expression of epithelial markers (E-cadherin, CK-20, and EpCAM) but decreased the expression of Twist and mesenchymal markers (N-cadherin and vimentin) in CRC cell lines. Finally, the molecular alterations after BMAL1-KD promoted mesenchymal-to-epithelial transition-like changes mostly appeared in two primary CRC cell lines (i.e., HCT116 and SW480) compared to the metastatic cell line SW620. As a consequence, migration/invasion and drug resistance capacities decreased in HCT116 and SW480 BMAL1-KD cells. Together, BMAL1-KD alerts the delicate equilibrium between epithelial and mesenchymal properties of CRC cell lines, which revealed the crucial role of BMAL1 in EMT-related CRC metastasis and chemoresistance.


Assuntos
Fatores de Transcrição ARNTL/genética , Fatores de Transcrição ARNTL/metabolismo , Neoplasias do Colo/genética , Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/genética , Antígenos CD/metabolismo , Caderinas/metabolismo , Moléculas de Adesão Celular/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Movimento Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Bases de Dados Genéticas , Molécula de Adesão da Célula Epitelial/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Técnicas de Silenciamento de Genes , Humanos , Queratina-20/metabolismo , Invasividade Neoplásica/genética , Oxaliplatina/farmacologia , Transporte Proteico , Vimentina/metabolismo , beta Catenina/metabolismo
14.
Int J Mol Sci ; 22(10)2021 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-34070214

RESUMO

During tubo-ovarian high-grade serous carcinoma (HGSC) progression, tumoral cells undergo phenotypic changes in their epithelial marker profiles, which are essential for dissemination processes. Here, we set out to determine whether standard epithelial markers can predict HGSC patient prognosis. Levels of E-CADH, KRT7, KRT18, KRT19 were quantified in 18 HGSC cell lines by Western blot and in a Discovery cohort tissue microarray (TMA) (n = 101 patients) using immunofluorescence. E-CADH and KRT7 levels were subsequently analyzed in the TMA of the Canadian Ovarian Experimental Unified Resource cohort (COEUR, n = 1158 patients) and in public datasets. Epithelial marker expression was highly variable in HGSC cell lines and tissues. In the Discovery cohort, high levels of KRT7 and KRT19 were associated with an unfavorable prognosis, whereas high E-CADH expression indicated a better outcome. Expression of KRT7 and E-CADH gave a robust combination to predict overall survival (OS, p = 0.004) and progression free survival (PFS, p = 5.5 × 10-4) by Kaplan-Meier analysis. In the COEUR cohort, the E-CADH-KRT7 signature was a strong independent prognostic biomarker (OS, HR = 1.6, p = 2.9 × 10-4; PFS, HR = 1.3, p = 0.008) and predicted a poor patient response to chemotherapy (p = 1.3 × 10-4). Our results identify a combination of two epithelial markers as highly significant indicators of HGSC patient prognosis and treatment response.


Assuntos
Antígenos CD/metabolismo , Biomarcadores Tumorais/metabolismo , Caderinas/metabolismo , Cistadenocarcinoma Seroso/metabolismo , Neoplasias das Tubas Uterinas/metabolismo , Queratina-7/metabolismo , Neoplasias Ovarianas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Estudos de Coortes , Cistadenocarcinoma Seroso/mortalidade , Neoplasias das Tubas Uterinas/mortalidade , Feminino , Humanos , Estimativa de Kaplan-Meier , Queratina-19/metabolismo , Neoplasias Pulmonares/metabolismo , Pessoa de Meia-Idade , Neoplasias Ovarianas/mortalidade , Prognóstico , Intervalo Livre de Progressão , Neoplasias Gástricas/metabolismo , Vimentina/metabolismo
15.
BMC Cancer ; 21(1): 492, 2021 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-33941102

RESUMO

BACKGROUND: Mycosis fungoides (MF) is a primary cutaneous T-cell lymphoma (CTCL) that transforms from mature, skin-homing T cells and progresses during the early stages in the skin. The role of the skin microenvironment in MF development is unclear, but recent findings in a variety of cancers have highlighted the role of stromal fibroblasts in promoting or inhibiting tumorigenesis. Stromal fibroblasts are an important part of the cutaneous tumor microenvironment (TME) in MF. Here we describe studies into the interaction of TME-fibroblasts and malignant T cells to gain insight into their role in CTCL. METHODS: Skin from normal (n = 3) and MF patients (n = 3) were analyzed for FAPα by immunohistochemistry. MyLa is a CTCL cell line that retains expression of biomarkers TWIST1 and TOX that are frequently detected in CTCL patients. MyLa cells were cultured in the presence or absence of normal or MF skin derived fibroblasts for 5 days, trypsinized to detached MyL a cells, and gene expression analyzed by RT-PCR for MF biomarkers (TWIST1 and TOX), Th1 markers (IFNG, TBX21), Th2 markers (GATA3, IL16), and proliferation marker (MKI67). Purified fibroblasts were assayed for VIM and ACTA2 gene expression. Cellular senescence assay was performed to assess senescence. RESULTS: MF skin fibroblast showed increased expression of FAP-α with increasing stage compared to normal. Normal fibroblasts co-cultured with MyLa cells suppressed expression of TWIST1 (p < 0.0006), and TOX (p < 0.03), GATA3 (p < 0.02) and IL16 (p < 0.03), and increased expression of IFNG (p < 0.03) and TBX21 (p < 0.03) in MyLa cells. In contrast, MyLa cells cultured with MF fibroblasts retained high expression of TWIST1, TOX and GATA3. MF fibroblasts co-culture with MyLa cells increased expression of IL16 (p < 0.01) and IL4 (p < 0.02), and suppressed IFNG and TBX21 in MyLa cells. Furthermore, expression of MKI67 in MyLa cells was suppressed by normal fibroblasts compared to MF fibroblasts. CONCLUSION: Skin fibroblasts represent important components of the TME in MF. In co-culture model, normal and MF fibroblasts have differential influence on T-cell phenotype in modulating expression of Th1 cytokine and CTCL biomarker genes to reveal distinct roles with implications in MF progression.


Assuntos
Fibroblastos Associados a Câncer/metabolismo , Proteínas de Grupo de Alta Mobilidade/metabolismo , Linfoma Cutâneo de Células T/metabolismo , Proteínas Nucleares/metabolismo , Neoplasias Cutâneas/metabolismo , Microambiente Tumoral , Proteína 1 Relacionada a Twist/metabolismo , Actinas/genética , Actinas/metabolismo , Idoso , Fibroblastos Associados a Câncer/patologia , Linhagem Celular Tumoral , Senescência Celular , Técnicas de Cocultura , Endopeptidases/genética , Endopeptidases/metabolismo , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Fator de Transcrição GATA3/genética , Fator de Transcrição GATA3/metabolismo , Expressão Gênica , Proteínas de Grupo de Alta Mobilidade/genética , Humanos , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-16/genética , Interleucina-16/metabolismo , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Linfoma Cutâneo de Células T/genética , Linfoma Cutâneo de Células T/patologia , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Micose Fungoide/genética , Micose Fungoide/metabolismo , Micose Fungoide/patologia , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Pele/citologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Linfócitos T/metabolismo , Proteína 1 Relacionada a Twist/genética , Vimentina/genética , Vimentina/metabolismo
16.
Exp Cell Res ; 405(1): 112666, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34052237

RESUMO

Vimentin protein is one of the main cytoskeleton and plays an important role in cell motility and metastasis. Nowadays, vimentin is widely studied as an epithelial-mesenchymal transition (EMT) marker of cancer cells while its involvement in cancer proliferation is poorly understood. In this study, we investigated the participation of vimentin in regulating cancer proliferation by silencing VIM gene in four cancer cell lines. Our results demonstrated that vimentin loss significantly induced cancer cell proliferation both in vitro and in vivo, which has not been reported so far. Mechanistically, knockdown of vimentin expression activated AKT phosphorylation and its downstream ß-catenin signaling. Nuclear translocation and transcriptional activity of ß-catenin was enhanced after silencing vimentin expression. Furthermore, vimentin loss could prevent Rictor from autophagy-dependent degradation via reducing AMPK-mediated autophagy signaling. AICAR, an AMPK activator, down-regulated Rictor and p-AKT levels while vimentin knockdown could rescue the effects. In vivo, it was also found that Ki67 expression and p-AKT/ß-catenin signaling pathway were obviously up-regulated in the tumor tissues in which vimentin was silenced compared to control groups. Taken together, these data showed the novel function of vimentin in regulating cancer proliferation via Rictor/AKT/ß-catenin signaling pathway, which suggested that it need more careful consideration before inhibiting metastatic cancers through targeting vimentin.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteína Companheira de mTOR Insensível à Rapamicina/metabolismo , Vimentina/deficiência , beta Catenina/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Movimento Celular , Proliferação de Células , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Transição Epitelial-Mesenquimal , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Fosforilação , Proteínas Proto-Oncogênicas c-akt/genética , Proteína Companheira de mTOR Insensível à Rapamicina/genética , Células Tumorais Cultivadas , Vimentina/genética , Vimentina/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/genética
17.
Exp Eye Res ; 210: 108643, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34058231

RESUMO

Choroidal melanoma is a devastating disease that causes visual loss and a high mortality rate due to metastasis. Luteolin, a potential anticancer compound, is widely found in natural plants. The aim of this study was to evaluate the antiproliferative, antiadhesive, antimigratory and anti-invasive effects of luteolin on choroidal melanoma cells in vitro and to explore its potential mechanism. Cell counting kit-8 (CCK-8) assays, 5-ethynyl-2'-deoxyuridine (EdU) assays, Cell adhesion, migration, and invasion assays were performed to examine the inhibitory effects of luteolin on cell cell viability, proliferation, adhesion, migration and invasion capacities, respectively. Considering the correlation between Matrix metalloenzymes and tumor metastasis, Enzyme-linked immunosorbent assays (ELISAs) were used to assess matrix metalloproteases MMP-2 and MMP-9 secretion. Western blotting was performed to detect p-PI3K P85, Akt, and p-Akt protein expression. The cytoskeletal proteins vimentin were observed with cell immunofluorescence staining. Luteolin can inhibit OCM-1 cell proliferation, migration, invasion and adhesion and C918 cell proliferation, migration, and invasion through the PI3K/Akt signaling pathway. Therefore, Luteolin may have potential as a therapeutic medication for Choroidal melanoma.


Assuntos
Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias da Coroide/tratamento farmacológico , Luteolina/uso terapêutico , Melanoma/tratamento farmacológico , Western Blotting , Sobrevivência Celular/efeitos dos fármacos , Neoplasias da Coroide/enzimologia , Neoplasias da Coroide/patologia , Ensaio de Imunoadsorção Enzimática , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Melanoma/enzimologia , Melanoma/patologia , Microscopia de Fluorescência , Invasividade Neoplásica/prevenção & controle , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Células Tumorais Cultivadas , Vimentina/metabolismo
18.
Anticancer Res ; 41(5): 2211-2215, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33952447

RESUMO

Elastofibroma dorsi is an uncommon benign fibroblastic pseudotumor that typically occurs in the subscapular region of middle-aged or older individuals. The pathogenesis is still unclear and a matter of debate. Magnetic resonance imaging can be used as a first-line investigation of the lesion and reveals a lenticular soft-tissue mass with a signal intensity similar to that of skeletal muscle interlaced with strands of fat. Biopsy is not necessary if all pathognomonic criteria are present. A conservative "wait and see" attitude is reasonable and immediate surgery is no more the standard treatment of elastofibroma dorsi. This review provides an updated overview of the diagnosis, management and pathogenesis of elastofibroma dorsi. We also discuss recent advances in our understanding of genomic alterations in elastofibroma dorsi.


Assuntos
Fibroma/diagnóstico , Fibroma/terapia , Neoplasias de Tecidos Moles/diagnóstico , Neoplasias de Tecidos Moles/terapia , Antígenos CD34/metabolismo , Aberrações Cromossômicas , Variações do Número de Cópias de DNA , Fibroma/genética , Humanos , Imuno-Histoquímica , Imageamento por Ressonância Magnética , Muramidase/metabolismo , Neoplasias de Tecidos Moles/metabolismo , Vimentina/metabolismo
19.
Adv Exp Med Biol ; 1269: 179-183, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33966214

RESUMO

Epithelial-mesenchymal transition (EMT), which is involved in metastasis formation, requires reprogramming of gene expression mediated by key EMT transcription factors. However, signals from the cellular microenvironment, including hypoxia, can also modulate the process of EMT. Hypoxia is often associated with a reduction in the extracellular pH of the tumor microenvironment (acidosis). Whether acidosis alone has an impact on the expression of the EMT markers E-cadherin, N-cadherin, and vimentin was studied in NCI-H358 lung cancer cells. Reducing extracellular pH decreased E-cadherin mRNA, while vimentin and N-cadherin mRNA were doubled. However, at the protein level, E-cadherin and N-cadherin were both reduced, and only vimentin was upregulated. E-cadherin and N-cadherin expression at the cell surface, which is the relevant parameter for cell-cell and cell-matrix interaction, decreased too. The reduction of cell surface proteins was due to diminished protein expression and not changes in cellular localization, since localization of EMT markers in general was not affected by acidosis. Acidosis also affected NCI-H358 cells functionally. Adhesion was decreased when the cells were primed in an acidic medium before measuring cell adherence, which is in line with the reduced expression of cadherins at the cell surface. Additionally, migration was decreased after acidic priming. A possible mechanism for the regulation of EMT markers involves the action of microRNA-203a (miR-203a). In NCI-H358 lung cancer cells, miR-203a expression was repressed by acidosis. Since a decrease in the level of miR-203a has been shown to induce EMT, it might be involved in the modulation of EMT marker expression, adhesion, and migration by the acidic tumor microenvironment in NCI-H358 lung cancer cells.


Assuntos
Transição Epitelial-Mesenquimal , Neoplasias Pulmonares , Biomarcadores , Linhagem Celular Tumoral , Movimento Celular , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/genética , Microambiente Tumoral/genética , Vimentina/genética , Vimentina/metabolismo
20.
Toxins (Basel) ; 13(3)2021 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-33808971

RESUMO

Mammalian animal toxicity of ochratoxin A (OTA) has focused largely in the past half-century on pigs because of initial recognition of it as a principal cause of intermittent growth suppression and renal disease caused by mouldy feed. Subsequent classical toxicology has used laboratory rodents because renal pathology in pigs raised questions concerning possible involvement in the human idiopathic bilateral renal atrophy of Balkan endemic nephropathy for which OTA was a focus of attention for human nephropathy through 1980s and into 2000s. Emphasis on human nephropathy has more recently concerned the plant metabolite aristolochic acid. Recognition that agricultural management can often minimise food and feed-stuff spoilage by OTA-producing Aspergilli and Penicillia has moderated some of the risks for animals. Legislation for human food safety combined with sophisticated analysis generally provides safety in the developed world. Chronic experimental exposure of male rats, in the absence of clinical dis-ease, specifically causes renal cancer. The possibility of this as a unique model for the human has generated considerable experimental evidence which may be more directly relevant for carcinogenesis in the complex kidney than that obtained from biochemical toxicities in vitro. Nevertheless, there does not appear to be any case of human renal or urinary tract cancer for which there is verified etiological proof for causation by OTA, contrary to much claim in the literature. To contribute to such debate, histopathology review of OTA/rat renal cancers, augmented where appropriate by immune profiles, has been completed for all remaining tumours in our research archive. Overall consistency of positivity for vimentin, is matched with occasional positives either for CD10 or the cytokeratin MNF 116. The current situation is discussed. Suggestion that OTA could cause human testicular cancer has also been challenged as unsupported by any experimental findings in rats, where the Leydig cell tumour immune profile does not match that of human germ cell neoplasms.


Assuntos
Testes de Carcinogenicidade , Neoplasias Renais/induzido quimicamente , Ocratoxinas/toxicidade , Neoplasias Testiculares/induzido quimicamente , Animais , Exposição Dietética , Humanos , Queratinas/metabolismo , Neoplasias Renais/metabolismo , Neoplasias Renais/patologia , Masculino , Neprilisina/metabolismo , Ratos Endogâmicos F344 , Medição de Risco , Fatores de Risco , Neoplasias Testiculares/metabolismo , Neoplasias Testiculares/patologia , Fatores de Tempo , Vimentina/metabolismo
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