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1.
Bioengineered ; 10(1): 282-291, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31311401

RESUMO

Transforming growth factor (TGF)-ß1 plays a crucial role in the epithelial-to-mesenchymal transition (EMT) in many cancer types and in thyroid cancers. Epigallocatechin-3-gallate (EGCG), the most important ingredient in the green tea, has been reported to possess antioxidant and anticancer activities. However, the cellular and molecular mechanisms explaining its action have not been completely understood. In this study, we found that EGCG significantly suppresses EMT, invasion and migration in anaplastic thyroid carcinoma (ATC) 8505C cells in vitro by regulating the TGF-ß/Smad signaling pathways. EGCG significantly inhibited TGF-ß1-induced expression of EMT markers (E-cadherin reduction and vimentin induction) in 8505C cells in vitro. Treatment with EGCG completely blocked the phosphorylation of Smad2/3, translocation of Smad4. Taken together, these results suggest that EGCG suppresses EMT and invasion and migration by blocking TGFß/Smad signaling pathways.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Catequina/análogos & derivados , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Transdução de Sinais/efeitos dos fármacos , Células Epiteliais da Tireoide/efeitos dos fármacos , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/genética , Caderinas/metabolismo , Catequina/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Humanos , Fosforilação/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína Smad3/genética , Proteína Smad3/metabolismo , Proteína Smad4/genética , Proteína Smad4/metabolismo , Células Epiteliais da Tireoide/metabolismo , Células Epiteliais da Tireoide/patologia , Fator de Crescimento Transformador beta1/farmacologia , Vimentina/agonistas , Vimentina/genética , Vimentina/metabolismo
2.
Life Sci ; 230: 197-207, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31150688

RESUMO

AIMS: Increased amounts of protein, in particular albumin within renal tubular cells (TBCs), induce the expression of inflammatory and fibrogenic mediators, which are adverse prognostic factors in tubulointerstitial fibrosis and diabetic nephropathy (DN). We sought to assess the participation of the thiol-linked tertiary structure of albumin in the mechanism of protein toxicity in a model of TBCs. MATERIALS AND METHODS: Cultured human renal proximal tubular cells, HK-2, were exposed to isolated albumin from patients with and without DN (Stages 0, 1 and 4). The magnitude of change of the albumin tertiary structure, cell viability (LDH leakage), apoptosis (Annexin V), transdifferentiation and reticulum endoplasmic stress (Western blot and flow cytometry) and lysosomal enzyme activity were assessed. KEY FINDINGS: We found that albumin from Stage 4 patients presented >50% higher thiol-dependent changes of tertiary structure compared to Stages 0 and 1. Cells incubated with Stage 4 albumin displayed 5 times less viability, accompanied by an increased number of apoptotic cells; evidence of profibrogenic markers E-cadherin and vimentin and higher expression of epithelial-to-mesenchymal transition markers α-SMA and E-cadherin and of endoplasmic reticulum stress protein GRP78 were likewise observed. Moreover, we found that cathepsin B activity in isolated lysosomes showed a significant inhibitory effect on albumin from patients in advanced stages of DN and on albumin that was intentionally modified. SIGNIFICANCE: Overall, this study showed that thiol-dependent changes in albumin's tertiary structure interfere with the lysosomal proteolysis of renal TBCs, inducing molecular changes associated with interstitial fibrosis and DN progression.


Assuntos
Nefropatias Diabéticas/metabolismo , Lisossomos/fisiologia , Albumina Sérica Humana/fisiologia , Adulto , Idoso , Albuminas/metabolismo , Apoptose/efeitos dos fármacos , Caderinas/metabolismo , Linhagem Celular , Sobrevivência Celular , Transdiferenciação Celular , Nefropatias Diabéticas/fisiopatologia , Estresse do Retículo Endoplasmático , Células Epiteliais/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Fibrose , Humanos , Túbulos Renais/patologia , Masculino , Pessoa de Meia-Idade , Cultura Primária de Células , Estrutura Terciária de Proteína/fisiologia , Albumina Sérica Humana/metabolismo , Transdução de Sinais/efeitos dos fármacos , Vimentina/metabolismo
3.
Gene ; 710: 186-192, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31175924

RESUMO

EMT (Epithelial-Mesenchymal Transition) is one of the factors in the pathogenesis of adenomyosis. FMNL2 induced invasion of cancer cell through promoting EMT, but it is unclear the role of FMNL2 in the adenomyosis. By IHC staining, we found the expression level of FMNL2 was significantly higher in the ectopic endometrial stromal cells from women with adenomyosis when compared with normal endometrial stromal cells. Knockdown of FMNL2 inhibited the invasion and migration of ectopic endometrial stromal cells and promoted the protein levels of E-cadherin and Vimentin. Meanwhile, inhibition of FMNL2 could induce the cell membrane localization of E-cadherin. Our findings reveal that the aberrant activation of FMNL2 promotes the pathogenesis of adenomyosis through inducing the EMT process. On the contrary, inhibition of FMNL2 promotes the transition of ectopic endometrial stromal cells to epithelial cells in adenomyosis through a MET-like process.


Assuntos
Adenomiose/metabolismo , Células Epiteliais/metabolismo , Proteínas/metabolismo , Células Estromais/metabolismo , Regulação para Cima , Adenomiose/genética , Adulto , Antígenos CD/metabolismo , Caderinas/metabolismo , Movimento Celular , Proliferação de Células , Células Cultivadas , Células Epiteliais/citologia , Transição Epitelial-Mesenquimal , Feminino , Técnicas de Silenciamento de Genes , Humanos , Pessoa de Meia-Idade , Proteínas/genética , Transdução de Sinais , Células Estromais/citologia , Vimentina/metabolismo
4.
Cell Physiol Biochem ; 53(1): 87-100, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31204440

RESUMO

BACKGROUND/AIMS: Different components of the tumor microenvironment can be either tumor-promoting or tumor-suppressive agents depending on factors which are not fully understood. Fibulins are components of the extracellular matrix from different tissues and constitute a clear example of this dual function. In fact, fibulins may either support tumor growth or abolish progression of malignant cells depending on the crosstalk between tumor cells and their surrounding stroma through mechanisms that remain to be elucidated. Among all fibulins, fibulin-5 contains a particular structural hallmark which consists in the presence of a RGD motif within its architecture. Previous reports have highlighted the importance of the interaction of this motif with integrins, and not only in normal functions but also in a tumor context. METHODS: Site-Directed Mutagenesis technique was employed to introduce the change RGD to RGE (RGD-to-RGE) within Fbln5 cDNA sequence. Cell proliferation was measured using the MTT assay or by counting Ki-67 positive cell nuclei. Cell adhesion was analysed using culture plates coated with different extracellular matrix components. Cell invasion was evaluated using 24-well Matrigel-coated invasion chambers, and mammosphere formation was monitored using ultralow attachment culture plates. BALB/c mice were employed to induce subcutaneous tumors. RESULTS: The RGD-to-RGE change alters the capacity of breast cancer cells to adhere to different extracellular matrix proteins as well as to αvß3 and α5ß1 integrins, and promotes protumor effects using different cell-based assays. Moreover, 4T1 cells, a mouse breast cancer cell line model, shows an increased capacity to generate tumors when exogenously expresses fibulin-5 with a RGD-to-RGE change, and such capacity is similar to that shown for 4T1 cells with an interfered Fbln5 gene. CONCLUSION: These data highlight the importance of the RGD motif of fibulin-5 to induce antitumor effects and provide new insights into the involvement of fibulins in tumor processes.


Assuntos
Adesão Celular/efeitos dos fármacos , Proteínas da Matriz Extracelular/farmacologia , Oligopeptídeos/metabolismo , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/efeitos dos fármacos , Proteínas da Matriz Extracelular/metabolismo , Proteínas da Matriz Extracelular/uso terapêutico , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mutagênese Sítio-Dirigida , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Oligopeptídeos/genética , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Transplante Homólogo , Vimentina/metabolismo
5.
Chem Biol Interact ; 307: 158-166, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31059706

RESUMO

Metastatic osteosarcoma usually has an unsatisfactory response to the current standard chemotherapy and causes poor prognosis. Currently, epithelial-mesenchymal transition (EMT) is reported as a critical event in osteosarcoma metastasis. Glaucocalyxin A, a bioactive ent-kauranoid diterpenoid, exerts anti-cancer effect on osteosarcoma by inducing apoptosis in previous study. However, the effect of Glaucocalyxin A on EMT and metastasis of osteosarcoma is unclear. In this study, we investigated the potential mechanisms of Glaucocalyxin A on EMT and metastasis of osteosarcoma. We found that Glaucocalyxin A inhibited migration and invasion of MG-63 and 143B cells. Moreover, Glaucocalyxin A increased the protein and mRNA levels of E-cadherin and decreased the protein and transcription expression of N-cadherin, Vimentin. Glaucocalyxin A also inhibited the protein and mRNA levels of EMT-associated transcription factor including Snail and Slug. Furthermore, Glaucocalyxin A inhibited transforming growth factor-ß1 (TGF-ß1)-induced migration, invasion and EMT of low-metastatic osteosarcoma U2OS cells. Glaucocalyxin A inhibited TGF-ß-induced phosphorylation of Smad 2/3 in osteosarcoma U2OS cells. Finally, we established transplanted metastatic models of highly metastatic osteosarcoma 143B cells. Glaucocalyxin A inhibited lung metastasis in vivo. Interestingly, Glaucocalyxin A increased the protein expression of E-cadherin and reduced the protein expression of N-cadherin and Vimentin. Glaucocalyxin A inhibited the protein expression of Snail and Slug in vivo. In summary, this study demonstrated that Glaucocalyxin A inhibited EMT and TGF-ß1-induced EMT by inhibiting TGF-ß1/Smad2/3 signaling pathway in osteosarcoma. Therefore, Glaucocalyxin A might be a promising candidate against the metastasis of human osteosarcoma.


Assuntos
Diterpenos de Caurano/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/farmacologia , Animais , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Diterpenos de Caurano/química , Diterpenos de Caurano/uso terapêutico , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Nus , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Fosforilação/efeitos dos fármacos , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Vimentina/metabolismo
6.
Nanomedicine (Lond) ; 14(10): 1247-1265, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31084385

RESUMO

Aim: Plant virus-like particles (VLPs) have emerged as a novel platform for delivery of drugs/antibodies. The aim of the present investigation is to establish the entry mechanism of flexuous rod-shaped virus particles into mammalian cells. Methods: Far-Western blot analysis, pull-down and ELISA were used to characterize vimentin and Hsp60 interaction with VLPs. The mode/kinetics of internalization of VLPs was deciphered using pharmacological inhibitors/endosomal markers. Results & discussion: The flexuous rod-shaped VLPs of Pepper vein banding virus (PVBV) enter HeLa and HepG2 cells via cell-surface proteins: vimentin and Hsp60, respectively. VLPs internalize via different modes of endocytosis in HeLa, HepG2 cells and are biodegradable. Vimentin and Hsp60 could be potential epithelial ligands that facilitate targeting of nanoparticles to tumor cells.


Assuntos
Endocitose , Células Epiteliais/metabolismo , Nanopartículas/metabolismo , Potyvirus/fisiologia , Animais , Transporte Biológico , Chaperonina 60/metabolismo , Citoplasma/metabolismo , Células HeLa , Células Hep G2 , Humanos , Cinética , Vimentina/metabolismo , Vírion/fisiologia , Internalização do Vírus
7.
APMIS ; 127(7): 491-502, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30942913

RESUMO

Secretory carcinoma (SC) of salivary glands is a newly described low-grade malignancy characterized by the presence of ETV6 rearrangement. Only a few cases and very small series with cytomorphology were reported so far. Six cases of fine-needle aspirations (FNAs) from afterward histologically, immunohistochemically and genetically confirmed SCs were retrieved from the archives of the authors. Ancillary immunocytochemistry (ICC) and translocation detection were performed on cell blocks (CBs). All aspirates were sufficiently cellular and cells were arranged in more or less cohesive groups with only mild nuclear polymorphism. The cytoplasm was eosinophilic, granulated and vacuolated, especially in CBs. Secretory material within the microcystic spaces was periodic acid-Schiff (PAS) positive. Triple positivity of immunomarkers S-100 protein, mammaglobin and vimentin was present. The proliferation index was low. Ancillary techniques suggested the possibility of SC in a few cytology cases; nevertheless, the final diagnosis was based on histomorphology, immunohistochemistry and genetics. The SC of salivary glands is detectable pre-operatively using ICC and genetics. The presence of the diagnostic ETV6 rearrangement increases the accuracy of FNA to the maximum. According to the Milan system, cases genetically not confirmed should be categorized as Suspicious for Malignancy or Salivary Gland Neoplasm of Uncertain Malignant Potential (SUMP), both requiring surgery.


Assuntos
Carcinoma/diagnóstico , Carcinoma/patologia , Neoplasias das Glândulas Salivares/diagnóstico , Neoplasias das Glândulas Salivares/patologia , Glândulas Salivares/patologia , Adulto , Idoso , Biomarcadores Tumorais/metabolismo , Biópsia por Agulha Fina/métodos , Carcinoma/metabolismo , Citodiagnóstico/métodos , Feminino , Humanos , Imuno-Histoquímica/métodos , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-ets/metabolismo , Proteínas Repressoras/metabolismo , Proteínas S100/metabolismo , Neoplasias das Glândulas Salivares/metabolismo , Glândulas Salivares/metabolismo , Vimentina/metabolismo , Adulto Jovem
8.
Bull Exp Biol Med ; 166(6): 811-815, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31020581

RESUMO

We assessed changes of olfactory bulbs in rata with 6-hydroxydopamine destruction of the substantia nigra. The expression of marker proteins of immature and differentiated neurons and glia (vimentin, PSA-NCAM, tyrosine hydroxylase, and S100) was analyzed by immunohistochemical and morphometric methods. The number of periglomerular dopamine neurons and astroglia in the olfactory bulbs increased on the side of toxin injection and expression of PSA-NCAM and vimentin increased in the rostral migratory stream. Destruction of the substantia nigra shifted differentiation of neuronal progenitors towards the dopaminergic phenotype and increased their survival in the olfactory bulbs, which can be explained by increased expression of PSA-NCAM.


Assuntos
Neuroglia/patologia , Neurônios/patologia , Bulbo Olfatório/patologia , Doença de Parkinson Secundária/patologia , Substância Negra/patologia , Adaptação Fisiológica , Animais , Biomarcadores/metabolismo , Modelos Animais de Doenças , Expressão Gênica , Humanos , Imuno-Histoquímica , Injeções Intraventriculares , Masculino , Atividade Motora/fisiologia , Molécula L1 de Adesão de Célula Nervosa/genética , Molécula L1 de Adesão de Célula Nervosa/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Bulbo Olfatório/metabolismo , Oxidopamina/administração & dosagem , Doença de Parkinson Secundária/induzido quimicamente , Doença de Parkinson Secundária/genética , Doença de Parkinson Secundária/metabolismo , Ratos , Ratos Wistar , Proteínas S100/genética , Proteínas S100/metabolismo , Ácidos Siálicos/genética , Ácidos Siálicos/metabolismo , Técnicas Estereotáxicas , Substância Negra/metabolismo , Tirosina 3-Mono-Oxigenase/genética , Tirosina 3-Mono-Oxigenase/metabolismo , Vimentina/genética , Vimentina/metabolismo
9.
Bull Exp Biol Med ; 166(6): 774-778, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31028580

RESUMO

We performed immunohistochemical analysis of the expression of TGFß, galectin-1, vimentin, and thrombospondin in the mucosa in gastric cancer of diffuse and intestinal type associated with systemic undifferentiated connective tissue dysplasia. In diffuse gastric cancer, both with and without association with connective tissue dysplasia, a higher level of expression of TGFß, galectin-1, vimentin, and thrombospondin in the tumor was detected in comparison with the perifocal and tumor zones in intestinal gastric cancer, which may reflect the pathogenetic peculiarities of the two histotypes of gastric cancer. Intestinal type of gastric cancer associated with connective tissue dysplasia is characterized by a high level of expression of galectin-1 and vimentin in the perifocal zone and TGFß in the tumor zone. The pattern of expression of the studied markers can reflect both the pathogenetic peculiarities of the two histotypes of gastric cancer and peculiar expression of some growth factors, cytoskeleton proteins, and matrix-cell proteins associated with undifferentiated connective tissue dysplasia which may contribute to epithelial-mesenchymal transition.


Assuntos
Biomarcadores Tumorais/genética , Galectina 1/genética , Neoplasias Gástricas/genética , Trombospondinas/genética , Fator de Crescimento Transformador beta/genética , Doenças do Tecido Conjuntivo Indiferenciado/genética , Vimentina/genética , Adulto , Idoso , Biomarcadores Tumorais/metabolismo , Transição Epitelial-Mesenquimal , Feminino , Galectina 1/metabolismo , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Neoplasias Gástricas/complicações , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/patologia , Trombospondinas/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Doenças do Tecido Conjuntivo Indiferenciado/complicações , Doenças do Tecido Conjuntivo Indiferenciado/diagnóstico , Doenças do Tecido Conjuntivo Indiferenciado/patologia , Vimentina/metabolismo
10.
Mol Med Rep ; 19(5): 4101-4108, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30942437

RESUMO

Although forkhead box protein M1 (FOXM1) is markedly upregulated in human premalignant and oral squamous cell carcinoma (OSCC) tissues and cultured cells, the association of FOXM1 expression with OSCC prognosis is not well understood. The present study investigated the possible association of FOXM1 expression in patients with OSCC with their clinicopathological characteristics and clinical outcomes. The expression of FOXM1 protein in OSCC tissues from 119 patients was evaluated by immunohistochemistry, and the results demonstrated that FOXM1 overexpression in patients with OSCC was associated with tumour recurrence and poor prognosis. To study the in vitro effects of FOXM1, its expression was decreased by small interfering RNA (siRNA) in OSCC cell lines, and FOXM1 knockdown decreased the proliferative, migratory and invasive capacities of cells. FOXM1 inhibition by siRNA gave rise to reduced expression of vimentin and increased expression of E­cadherin. The present study reported FOXM1 as a novel predictor of tumour recurrence in patients with OSCC and its potential involvement in epithelial­mesenchymal transition in OSCC cells.


Assuntos
Carcinoma de Células Escamosas/patologia , Transição Epitelial-Mesenquimal/genética , Proteína Forkhead Box M1/metabolismo , Neoplasias Bucais/patologia , Caderinas/genética , Caderinas/metabolismo , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/mortalidade , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Proteína Forkhead Box M1/antagonistas & inibidores , Proteína Forkhead Box M1/genética , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/metabolismo , Neoplasias Bucais/mortalidade , Recidiva Local de Neoplasia , Estadiamento de Neoplasias , Prognóstico , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Vimentina/genética , Vimentina/metabolismo
11.
Zhonghua Zhong Liu Za Zhi ; 41(3): 193-199, 2019 Mar 23.
Artigo em Chinês | MEDLINE | ID: mdl-30917455

RESUMO

Objective: To investigate the role of microRNA-96-5p in the proliferation and invasion of gastric cancer cells and its molecular mechanism. Methods: From June 2015 to January 2017, 53 resected specimens were collected. The transcriptional levels of microRNA-96-5p and forkhead box Q1 (FoxQ1) in gastric cancer tissues and the matched para-cancerous tissues were quantified by quantitative real-time PCR (qRT-PCR). The expression of FoxQ1 protein was also detected by immunohistochemistry (IHC). The relationship between microRNA-96-5p expression and the clinicopathological features of gastric cancer and its correlation with FoxQ1 expression were analyzed. The expressions of miRNA-96-5p in gastric cancer tissue and adjacent normal tissue were detected by qRT-PCR. miRNA-96-5p mimics was transfected to BGC-823 gastric cancer cells. The effects of miRNA-96-5p on cell proliferation and invasion were detected by cell counting kit-8 (CCK-8) assay and Transwell assay, respectively. The protein expressions of FoxQ1, E-cadherin and vimentin were determined by western blot. The relationship between FoxQ1 and miRNA-96-5p expressed in BGC-823 cells was detected by dual-luciferase reporter assay. Results: The median expression of miRNA-96-5p in gastric cancer tissue was 1.05, significantly lower than 3.23 of para-cancerous tissues (P<0.05). The positive rate of FoxQ1 expression in gastric cancer tissue was 71.7%, significantly higher than 28.3% of para-cancerous tissues (P<0.05). The expression of FoxQ1 was negatively corelated with the level of miRNA-96-5p (r=-0.613, P=0.006). The expression of miRNA-96-5p in gastric cancer cell BGC-823 was significantly decreased compared with normal gastric epithelial cell (0.96±0.08 vs 2.84±0.15, P<0.05). The results of CCK-8 assay and Transwell assay showed that overexpression of miRNA-96-5p significantly reduced the proliferation and invasion abilities of gastric cancer cells (P<0.05). Overexpression of miRNA-96-5p decreased the protein level of FoxQ1. Moreover, it upregulated the expression of E-cadherin and downregulated the expression of vimentin. The result of dual-luciferase-3'-UTR reporter assay confirmed that miRNA-96-5p binds to the 3'UTR of FoxQ1. Conclusion: miRNA-96-5p may suppress the proliferation, migration and epithelial-mesenchymal transition (EMT) of gastric cancer cell by down-regulation of FoxQ1.


Assuntos
Movimento Celular , Proliferação de Células , Fatores de Transcrição Forkhead/metabolismo , MicroRNAs/metabolismo , Invasividade Neoplásica , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Caderinas/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , Humanos , Lesões Pré-Cancerosas/metabolismo , Lesões Pré-Cancerosas/patologia , Regulação para Cima , Vimentina/metabolismo
12.
Cell Physiol Biochem ; 52(3): 595-605, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30897324

RESUMO

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is the fifth most common malignancy and most frequently develops in patients with cirrhosis. Surveillance strategies are recommended in high-risk groups because early detection of small lesions improves the likelihood of curative treatment. This study investigated the prospective clinical significance of serum levels of anti-Ku86 and plasma levels of lamin B1and vimentin as early markers of HCC. METHODS: We recruited 74 patients at Assiut University Hospital-37 with HCC and 37 with chronic liver disease (liver cirrhosis patients)-and 36 age- and sex-matched healthy controls. Lamin B1 and vimentin mRNA expression levels were evaluated by reverse transcription-PCR and serum levels of anti-Ku86 were measured by enzyme-linked immunosorbent assay. RESULTS: Compared with liver disease patients and controls, HCC patients showed higher levels of lamin B1 mRNA (sensitivity, 96%; specificity, 65%), vimentin mRNA (sensitivity, 94%; specificity, 92%), and anti-Ku86 (sensitivity, 94%; specificity, 80%). LaminB1 levels were significantly higher in patients with a tumor size < 2 cm than in patients with tumors 2-5 cm and >5cm in size. Lamin B1 had significant positive correlations with alpha-fetoprotein (AFP) (P=0.034) and anti-Ku86 (P=0.002). Receiver operating characteristic curves for differentiating HCCfrom liver cirrhosis revealed a higher area under the curve(AUC).for vimentin than for AFP, lamin B1, and anti-Ku86 for the diagnosis of HCC (P<0.001). CONCLUSION: Circulating levels of anti-Ku86, lamin B1,and vimentin might be potential surrogate markers of HCC, either alone or in combination with AFP. However, independent and discriminative serological biomarkers with higher sensitivity and specificity are still needed for the early detection of HCC.


Assuntos
Anticorpos/sangue , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/diagnóstico , Hepatite C/diagnóstico , Lamina Tipo B/metabolismo , Neoplasias Hepáticas/diagnóstico , Vimentina/metabolismo , Adulto , Área Sob a Curva , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/complicações , Carcinoma Hepatocelular/patologia , Estudos de Casos e Controles , Feminino , Hepatite C/complicações , Humanos , Autoantígeno Ku/imunologia , Lamina Tipo B/genética , Neoplasias Hepáticas/complicações , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Curva ROC , Vimentina/genética , alfa-Fetoproteínas/análise
13.
Theriogenology ; 130: 120-124, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30884332

RESUMO

Microminipigs are one of the smallest miniature pigs characterized as sexually precocious; the males achieve sexual maturity at around 3-4.5 months of age. However, the physiology of this sexual precocity is still unclear. To understand sexual precocity in male microminipigs, we analyzed their testes at five developmental stages: neonatal (<7 days), 30-day-old, 45-day-old, 80-day-old, and adult (>24 months) stages. We used 4 pigs in each of the stages. To analyze testicular development histologically, the seminiferous tubule diameter (SD) was measured, and the presence or absence of the seminiferous lumen was confirmed. Changes in the expression of pluripotency markers, DBA, UCHL1, ZBTB16, and vimentin, were evaluated immunohistologically. For the analyses, cells positive for DBA, UCHL1, and ZBTB16 per 150 round seminiferous tubules in cross sections from each testis were counted to evaluate the total number of positive cells. The number of positive cells per 100 Sertoli cells (DBA+/Sertoli, UCHL1+/Sertoli, and ZBTB16+/Sertoli) was calculated to compare the five developmental stages. Histologically, SDs became larger with piglet growth, and precocity was confirmed; seminiferous lumens were observed from the 30-day-old stage. Immunohistologically, the number of DBA+/Sertoli, which indicates the number of gonocytes, decreased rapidly to an undetectable level by the 45-day-old stage. In the same period, the number of UCHL1+/Sertoli, which indicates total SSCs, increased significantly, suggesting that the proliferation of SSCs was accelerated before 30 days of age. Consequently, our study clarified that differentiation of SSCs in microminipigs started during the fetal period, the differentiation of gonocytes and proliferation of SSCs was then accelerated before 30 days of age, and the early phase of spermatogenesis was finally completed at around 45 days after birth. Consequently, sexual precocity in male microminipigs was characterized by a shorter duration of the early phase of spermatogenesis.


Assuntos
Células-Tronco Germinativas Adultas/metabolismo , Maturidade Sexual/fisiologia , Suínos/fisiologia , Animais , Biomarcadores , Diferenciação Celular , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Espermatogênese/fisiologia , Porco Miniatura , Ubiquitina Tiolesterase/genética , Ubiquitina Tiolesterase/metabolismo , Vimentina/genética , Vimentina/metabolismo
14.
BMC Cancer ; 19(1): 248, 2019 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-30894168

RESUMO

BACKGROUND: Garlic has been used for centuries for its flavour and health promoting properties that include protection against cancer. The vinyl disulfide-sulfoxide ajoene is one of the phytochemicals found in crushed cloves, hypothesised to act by S-thiolating reactive cysteines in target proteins. METHODS: Using our fluorescently labelled ajoene analogue called dansyl-ajoene, ajoene's protein targets in MDA-MB-231 breast cancer cells were tagged and separated by 2D electrophoresis. A predominant band was identified by MALDI-TOF MS/MS to be vimentin. Target validation experiments were performed using pure recombinant vimentin protein. Computational modelling of vimentin bound to ajoene was performed using Schrödinger and pKa calculations by Epik software. Cytotoxicity of ajoene in MDA-MB-231 and HeLa cells was measured by the MTT assay. The vimentin filament network was visualised in ajoene-treated and non-treated cells by immunofluorescence and vimentin protein expression was determined by immunoblot. The invasion and migration activity was measured by wound healing and transwell assays using wildtype cells and cells in which the vimentin protein had been transiently knocked down by siRNA or overexpressed. RESULTS: The dominant protein tagged by dansyl-ajoene was identified to be the 57 kDa protein vimentin. The vimentin target was validated to reveal that ajoene and dansyl-ajoene covalently bind to recombinant vimentin via a disulfide linkage at Cys-328. Computational modelling showed Cys-328 to be exposed at the termini of the vimentin tetramer. Treatment of MDA-MB-231 or HeLa cells with a non-cytotoxic concentration of ajoene caused the vimentin filament network to condense; and to increase vimentin protein expression. Ajoene inhibited the invasion and migration of both cancer cell lines which was found to be dependent on the presence of vimentin. Vimentin overexpression caused cells to become more migratory, an effect that was completely rescued by ajoene. CONCLUSIONS: The garlic-derived phytochemical ajoene targets and covalently modifies vimentin in cancer cells by S-thiolating Cys-328. This interaction results in the disruption of the vimentin filament network and contributes to the anti-metastatic activity of ajoene in cancer cells.


Assuntos
Movimento Celular/efeitos dos fármacos , Dissulfetos/farmacologia , Alho/química , Neoplasias/tratamento farmacológico , Vimentina/metabolismo , Linhagem Celular Tumoral , Simulação por Computador , Dissulfetos/metabolismo , Dissulfetos/uso terapêutico , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Invasividade Neoplásica/prevenção & controle , Neoplasias/patologia , Ligação Proteica , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Vimentina/isolamento & purificação
15.
Reprod Domest Anim ; 54(6): 835-845, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30907027

RESUMO

Equine endometrosis, a frequent cause of subfertility, is characterized by periglandular fibrosis, and no treatment exists. Endometrial biopsies not only contain diseased glands, but also contain healthy glands and stroma. Myoepithelial (ME) and myofibroblastic (MF) markers are calponin, smooth muscle actin (SMA), desmin and glial fibrillary acidic protein (GFAP). Epithelial vimentin expression indicates epithelial to mesenchymal transition (EMT). The aim of this immunohistochemical study was to investigate whether biopsies with endometrosis express MF and ME markers and vimentin. Compared to healthy areas, significantly higher percentages of endometrotic glands were lined by calponin- and vimentin-positive epithelial cells, whereas periglandular fibrosis contained significantly higher percentages of stromal cells positive for vimentin, desmin and SMA and significantly less calponin-positive stromal cells. The rare GFAP expression was restricted to endometrotic glands. Of these, the most frequent features of endometrotic glands were higher percentages of SMA- and vimentin-positive stromal cells and the prominent epithelial calponin staining that occurred in 100%, 93% and 95% of examined biopsies. Results indicate plasticity of equine endometrial epithelial and stromal cells. Particularly, endometrotic glands show evidence for ME differentiation and EMT. The different expression of MF markers between stromal cells from healthy and endometrotic areas suggests functional differences. The characteristic changes in the expression of SMA, vimentin and calponin between endometrotic glands and healthy areas can be helpful to confirm early stages of endometrosis. The characterization of cellular differentiation may help to decipher the pathogenesis of endometrosis and could lead to therapeutic strategies.


Assuntos
Endometriose/veterinária , Transição Epitelial-Mesenquimal , Doenças dos Cavalos/patologia , Actinas/metabolismo , Animais , Biomarcadores , Biópsia/veterinária , Proteínas de Ligação ao Cálcio/metabolismo , Desmina/metabolismo , Endometriose/metabolismo , Endometriose/patologia , Feminino , Doenças dos Cavalos/metabolismo , Cavalos , Imuno-Histoquímica/veterinária , Proteínas dos Microfilamentos/metabolismo , Células Estromais , Vimentina/metabolismo
16.
Int J Mol Med ; 43(5): 2075-2085, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30864678

RESUMO

von Willebrand factor C and EGF domain­containing protein (URG11), a cell growth regulator, is involved in the progression of a variety of types of cancer, including prostate cancer (Pca). However, the functions of the URG11 gene in Pca cells require in­depth investigation. The mRNA and protein levels of URG11 were measured by reverse transcription quantitative polymerase chain reaction (RT­qPCR) and western blot analysis. Cell Counting kit­8 (CCK­8), wound­healing and Transwell assays were used to detect cell viability, migration and invasion, respectively. Apoptosis and cell cycle analyses were performed using flow cytometry. The mRNA and protein expression levels of epithelial (E)­cadherin, vimentin, α­smooth muscle actin (α­SMA), cyclin D1 and MYC proto­oncogene protein (c­Myc) were analyzed by RT­qPCR and western blot analysis. In the present study, the mRNA and protein levels of URG11 were markedly upregulated in Pca cell lines compared with those in the normal prostate epithelial cell line. With functional experiments, the cell viability, migration and invasion of Pca cells were markedly promoted by URG11 overexpression. The cell cycle was effectively induced by URG11 and apoptosis was inhibited by the overexpression of URG11. Concomitantly, the epithelial marker E­cadherin was downregulated, and the mesenchymal markers vimentin and α­SMA were upregulated following URG11 overexpression. By contrast, genetic knockout of URG11 elicited the opposite effects. The present study also identified that the downstream effector genes of the Wnt/ß­catenin signal pathway, cyclin D1 and c­Myc, were increased following the overexpression of endogenous URG11, which are known to regulate cell proliferation. In addition, the Wnt/ß­catenin inhibitor FH535 ameliorated the promotive effects of URG11 on LNCaP cells viability, migration and invasion, and the Wnt/ß­catenin agonist LiCl reversed the inhibitory effects of siURG11 in LNCaP cells on cell viability, migration and invasion. The present study demonstrated that URG11 served an oncogenic role in the development of Pca cells and provided evidence that URG11 has potential as a novel therapeutic target in Pca.


Assuntos
Apoptose , Transativadores/metabolismo , Actinas/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/genética , Caderinas/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Ciclina D1/metabolismo , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Cloreto de Lítio/farmacologia , Masculino , Invasividade Neoplásica , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Interferente Pequeno/metabolismo , Sulfonamidas/farmacologia , Transativadores/genética , Vimentina/metabolismo
17.
Invest Ophthalmol Vis Sci ; 60(4): 1010-1020, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30884531

RESUMO

Purpose: The purpose of this study was to evaluate the effect of removal of Descemet's basement membrane and endothelium compared with removal of the endothelium alone on posterior corneal fibrosis. Methods: Twelve New Zealand White rabbits were included in the study. Six eyes had removal of the Descemet's membrane-endothelial complex over the central 8 mm of the cornea. Six eyes had endothelial removal with an olive-tipped cannula over the central 8 mm of the cornea. All corneas developed stromal edema. Corneas in both groups were cryofixed in optimum cutting temperature (OCT) formula at 1 month after surgery. Immunohistochemistry (IHC) was performed for α-smooth muscle actin (SMA), keratocan, CD45, nidogen-1, vimentin, and Ki-67, and a TUNEL assay was performed to detect apoptosis. Results: Six of six corneas that had Descemet's membrane-endothelial removal developed posterior stromal fibrosis populated with SMA+ myofibroblasts, whereas zero of six corneas that had endothelial removal alone developed fibrosis or SMA+ myofibroblasts (P < 0.01). Myofibroblasts in the fibrotic zone of corneas that had Descemet's membrane-endothelial removal were undergoing both mitosis and apoptosis at 1 month after surgery. A zone between keratocan+ keratocytes and SMA+ myofibroblasts contained keratocan-SMA-vimentin+ cells that were likely CD45- corneal fibroblasts and CD45+ fibrocytes. Conclusions: Descemet's basement membrane has an important role in modulating posterior corneal fibrosis after injury that is analogous to the role of the epithelial basement membrane in modulating anterior corneal fibrosis after injury. Fibrotic areas had myofibroblasts undergoing mitosis and apoptosis, indicating that fibrosis is in dynamic flux.


Assuntos
Substância Própria/patologia , Lâmina Limitante Posterior/fisiologia , Actinas/metabolismo , Animais , Apoptose/fisiologia , Edema da Córnea/etiologia , Substância Própria/metabolismo , Lâmina Limitante Posterior/cirurgia , Feminino , Fibrose/metabolismo , Fibrose/patologia , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Antígeno Ki-67/metabolismo , Antígenos Comuns de Leucócito/metabolismo , Glicoproteínas de Membrana/metabolismo , Mitose/fisiologia , Miofibroblastos/citologia , Proteoglicanas/metabolismo , Coelhos , Vimentina/metabolismo
18.
Cancer Biother Radiopharm ; 34(4): 245-251, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30888215

RESUMO

Background: The plant homeodomain finger 6 (PHF6) was originally identified as single gene mutated in Börjeson-Forssman-Lehmann syndrome, which was reported to be a tumor suppressor in T-cell acute lymphoblastic leukemia. However, the biological function of PHF6 in hepatocellular carcinoma (HCC) has been poorly characterized. Materials and Methods: In this study, we first determined the mRNA levels of PHF6 in HCC tissues and adjacent normal tissues using quantitative real-time PCR. Then the expression of PHF6 was knocked down in HCC cell lines (HepG2, SMMC-7721, and Bel-7402) by siRNA transfection. A series of functional experiments, including EdU proliferation assay, colony formation assay, and Transwell assay, were performed in HCC cells. Western blot analysis was used to detect the expression of PHF6, E-cadherin, and Vimentin. Results: We found that PHF6 was significantly elevated in HCC tissues and positively correlated with TNM stage, differentiation, and lymph node metastasis. Silencing PHF6 significantly inhibited cell proliferation, colony formation, and migration in HCC cells. Furthermore, silencing PHF6 obviously increased E-cadherin and decreased Vimentin expression. Conclusions: These findings suggest that PHF6 plays a positive role in the growth of HCC cells, and targeting PHF6 could serve as a promising therapeutic strategy for human HCC.


Assuntos
Carcinoma Hepatocelular/patologia , Proteínas de Transporte/metabolismo , Neoplasias Hepáticas/patologia , Metástase Linfática/patologia , Antígenos CD/metabolismo , Caderinas/metabolismo , Proteínas de Transporte/genética , Movimento Celular , Proliferação de Células , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Masculino , Estadiamento de Neoplasias , RNA Interferente Pequeno/metabolismo , Regulação para Cima , Vimentina/metabolismo
19.
Pathol Res Pract ; 215(5): 1020-1032, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30833029

RESUMO

BACKGROUND: Although previous researchers have analyzed the expression level of vimentin in nasopharyngeal carcinoma (NPC), the sample size of each study was too small, and there was no further in-depth study utilizing microarray and RNA-sequencing data. More importantly, the role and molecular mechanism of vimentin in NPC have not yet been addressed comprehensively. Accordingly, the aim of the present research was to conduct a full exploration of the clinical significance of vimentin in NPC in a large sample size. MATERIALS AND METHODS: Immunohistochemistry was used to test the expression of vimentin in clinical samples. Data from relevant microarray and RNA-sequencing datasets were screened and extracted to explore the clinical role of vimentin in NPC. Subsequently, vimentin-related signaling pathways were investigated via in-silico approaches. RESULTS: The clinical immunohistochemistry detection showed the positive expression ratio of vimentin was 24.6% (14/57) of the NPC specimens, whereas vimentin expression was negative in nasopharyngitis (NPG) tissues (0/20, P = 0.016). The mRNA and protein levels of vimentin were both remarkably up-regulated in NPC based on 196 and 1566 cases, respectively. The protein level of vimentin was also a risky factor for the prognosis prediction of NPC with the hazard ratios (HR) being 3.831. Gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) analyses, the localization of vimentin was in both the cytoplasm and the cytoskeleton, and vimentin was involved in the regulation of molecular function, the execution phase of apoptosis, and the regulation of cellular component organization. CONCLUSION: The high expression of vimentin plays a pivotal role in the development and poor progression of NPC, which indicates that vimentin may be an effective predictive indicator for NPC.


Assuntos
Carcinoma Nasofaríngeo/patologia , Neoplasias Nasofaríngeas/patologia , Vimentina/metabolismo , Adulto , Idoso , Biomarcadores Tumorais/análise , Biologia Computacional , Conjuntos de Dados como Assunto , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade
20.
Clin Exp Rheumatol ; 37(3): 358-366, 2019 May-Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30767876

RESUMO

OBJECTIVES: Non-radiographic (nr-axSpA) and radiographic (AS) forms of axial spondyloarthritis (axSpA) share clinical features, but have different radiographic patterns. Radiographic progression is not associated with the current disease activity biomarkers. We investigated a matrix metalloproteinase mediated metabolite of C-reactive protein (CRPM) and two biomarkers of citrullinated vimentin (VICM and anti-MCV) as novel biomarkers of disease activity. METHODS: AxSpA patients (n=121 nr-axSpA and n=72 AS) were characterised by activity (AS disease activity score with CRP [ASDAS-CRP], Bath AS disease activity index [BASDAI] and functional index [BASFI]), radiographic scores and quality of life questionnaires. CRPM, VICM and anti-MCV levels were analysed by ELISA in serum. Asymptomatic controls (n=100) were used as reference. Multiple regression investigated association with disease activity and diagnostic potential. RESULTS: CRPM and VICM levels were increased in AS compared to nr-axSpA (11.9nM vs. 10.2nM, p<0.001 and 4.92nM vs. 3.77nM, p=0.0025). Anti-MCV was similar in both axSpA subgroups, but lowered in controls. In nr-axSpA, CRPM correlated with CRP (ρ=0.33, p<0.001) and VICM (ρ=0.29, p=0.001); in AS, VICM correlated with CRP (ρ=0.34, p=0.0032) and ESR (ρ=0.38, p<0.001). ASDAS-CRP correlated with CRPM and anti-MCV, but when adjusting for CRP the correlation only remained with CRPM. CRPM and VICM separated the subgroups with odds ratios of 1.19 and 1.10 adjusted for age, gender, BMI, and disease duration. VICM lost significance when adjusting for CRP. CONCLUSIONS: CRPM was associated with disease activity in axSpA, and CRPM and VICM separated the axSpA groups. This study indicates that serological biomarkers may be novel biomarkers in axSpA.


Assuntos
Proteína C-Reativa/metabolismo , Espondilartrite/metabolismo , Vimentina/metabolismo , Humanos , Qualidade de Vida , Espondilartrite/patologia , Espondilite Anquilosante/metabolismo , Espondilite Anquilosante/patologia
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