Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 2.933
Filtrar
1.
Anticancer Res ; 41(9): 4365-4375, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34475056

RESUMO

BACKGROUND/AIM: Ouabain has been shown to induce human cancer cell death via apoptosis. Still, its anti-metastatic effect on cell migration and invasion of human gastric cancer cells has not been addressed. MATERIALS AND METHODS: Cell proliferation and viability were measured by the MTT assay and flow cytometry, respectively. Cell motitlity was analysed by wound healing assay. Cell migration and invasion were analysed by the transwell system. Protein expression was assayed by western blotting. RESULTS: Ouabain decreased AGS cell proliferation, cell viability, and motility. In addition, ouabain inhibited AGS cell migration and invasion. Furthermore, ouabain decreased matrix metalloproteinase-2 (MMP-2) activity at 48 h. Ouabain reduced the levels of proteins associated with PI3K/AKT and p38/MAPK pathways. In addition, ouabain decreased the expressions of N-cadherin, tissue inhibitor of metalloproteinases-1 (TIMP-1), urokinase-type plasminogen activator (c-uPA), and MMP-2 at 48 h. CONCLUSION: Ouabain suppresses cell metastasis through multiple signaling pathways in AGS cells.


Assuntos
Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Metaloproteinase 2 da Matriz/metabolismo , Ouabaína/farmacologia , Neoplasias Gástricas/metabolismo , Antígenos CD/metabolismo , Caderinas/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Invasividade Neoplásica , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/tratamento farmacológico , Inibidor Tecidual de Metaloproteinase-1/metabolismo
2.
Int J Mol Sci ; 22(17)2021 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-34502227

RESUMO

Tissue Inhibitor of Metalloproteases 1, also known as TIMP-1, is named for its well-established function of inhibiting the proteolytic activity of matrix metalloproteases. Given this function, many studies were carried out to verify if TIMP-1 was able to interrupt processes such as tumor cell invasion and metastasis. In contrast, many studies have shown that TIMP-1 expression is increased in several types of tumors, and this increase was correlated with a poor prognosis and lower survival in cancer patients. Later, it was shown that TIMP-1 is also able to modulate cell behavior through the induction of signaling pathways involved in cell growth, proliferation, and survival. The mechanisms involved in the regulation of the pleiotropic functions of TIMP-1 are still poorly understood. Thus, this review aimed to present literature data that show its ability to form a membrane complex with CD63 and ß1-integrin, and point to N-glycosylation as a potential regulatory mechanism of the functions exerted by TIMP-1. This article reviewed the characteristics and functions performed individually by TIMP1, CD63, and ß1-integrin, the roles of the TIMP-1/CD63/ß1-integrin complex, both in a physiological context and in cancer, and the regulatory mechanisms involved in its assembly.


Assuntos
Integrina beta1/metabolismo , Neoplasias/patologia , Domínios e Motivos de Interação entre Proteínas , Tetraspanina 30/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Humanos , Neoplasias/metabolismo , Transdução de Sinais
3.
Int J Mol Sci ; 22(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34445312

RESUMO

"Neuroplasticity" is often evoked to explain adaptation and compensation after acute lesions of the Central Nervous System (CNS). In this study, we investigated the modification of 80 genes involved in synaptic plasticity at different times (24 h, 8 and 45 days) from the traumatic spinal cord injury (SCI), adopting a bioinformatic analysis. mRNA expression levels were analyzed in the motor cortex, basal ganglia, cerebellum and in the spinal segments rostral and caudal to the lesion. The main results are: (i) a different gene expression regulation is observed in the Spinal Cord (SC) segments rostral and caudal to the lesion; (ii) long lasting changes in the SC includes the extracellular matrix (ECM) enzymes Timp1, transcription regulators (Egr, Nr4a1), second messenger associated proteins (Gna1, Ywhaq); (iii) long-lasting changes in the Motor Cortex includes transcription regulators (Cebpd), neurotransmitters/neuromodulators and receptors (Cnr1, Gria1, Nos1), growth factors and related receptors (Igf1, Ntf3, Ntrk2), second messenger associated proteins (Mapk1); long lasting changes in Basal Ganglia and Cerebellum include ECM protein (Reln), growth factors (Ngf, Bdnf), transcription regulators (Egr, Cebpd), neurotransmitter receptors (Grin2c). These data suggest the molecular mapping as a useful tool to investigate the brain and SC reorganization after SCI.


Assuntos
Encéfalo/metabolismo , Plasticidade Neuronal/genética , Traumatismos da Medula Espinal/metabolismo , Medula Espinal/metabolismo , Transcriptoma , Animais , Feminino , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/metabolismo , Neurotransmissores/genética , Neurotransmissores/metabolismo , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/genética , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
4.
Am J Trop Med Hyg ; 105(3): 638-642, 2021 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-34280134

RESUMO

Chagas disease (CD) mainly conveys stroke risk through structural cardiac disease. However, stroke and cognitive impairment are seen in CD independently of cardiac disease severity. Chronic inflammation may be an explanation for this association, because inflammation plays an important role in the pathogenesis of acute ischemic stroke and dementia. In the present study, we selected five candidate biomarkers for Chagas disease: interleukin-6, membrane metalloproteinase-9, tissue inhibitor of metalloproteinase-1 (TIMP1), orosomucoid, and neprilysin. We sought to determine if mean levels of proinflammatory biomarkers are higher in patients with heart failure (HF) associated with Chagas disease when compared with other etiologies of HF. Patients were consecutively enrolled from subspecialty HF outpatient clinics at two university-based hospitals. Serum biomarker levels from blood samples were analyzed by ELISA. Severity of HF on echocardiography was worse in non-CD when compared with CD patients. No significant difference was observed in the levels of candidate biomarkers between the CD and non-CD groups. We found a significantly 2.2 ng/mL higher level of TIMP1 in CD when compared with non-CD patients with HF after adjustment for age and gender (95% confidence interval = 0.1 to 4.5, P = 0.037). In patients with heart failure, serum TIMP1 is increased in Chagas patients despite a lower myocardial disease severity on echocardiography when compared with non-Chagas patients. TIMP1 is probably one of multiple mediators of inflammatory injury.


Assuntos
Cardiomiopatia Chagásica/metabolismo , Insuficiência Cardíaca/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Adulto , Idoso , Cardiomiopatia Chagásica/diagnóstico por imagem , Feminino , Insuficiência Cardíaca/diagnóstico por imagem , Humanos , Inflamação/metabolismo , Interleucina-6/metabolismo , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Pessoa de Meia-Idade , Neprilisina/metabolismo , Orosomucoide/metabolismo
5.
Sci Rep ; 11(1): 13376, 2021 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-34183752

RESUMO

MMP-9 plays a number of important physiological functions but is also responsible for many pathological processes, including cancer invasion, metastasis, and angiogenesis. It is, therefore, crucial to understand its enzymatic activity, including activation and inhibition mechanisms. This enzyme may also be partially involved in the "cytokine storm" that is characteristic of COVID-19 disease (SARS-CoV-2), as well as in the molecular mechanisms responsible for lung fibrosis. Due to the variety of processing pathways involving MMP-9 in biological systems and its uniqueness due to the O-glycosylated domain (OGD) and fibronectin-like (FBN) domain, specific interactions with its natural TIMP-1 inhibitor should be carefully studied, because they differ significantly from other homologous systems. In particular, earlier experimental studies have indicated that the newly characterised circular form of a proMMP-9 homotrimer exhibits stronger binding properties to TIMP-1 compared to its monomeric form. However, molecular structures of the complexes and the binding mechanisms remain unknown. The purpose of this study is to fill in the gaps in knowledge. Molecular modelling methods are applied to build the inhibitory and non-inhibitory MMP-9-TIMP-1 complexes, which allows for a detailed description of these structures and should allow for a better understanding of the regulatory processes in which MMP-9 is involved.


Assuntos
Metaloproteinase 9 da Matriz/metabolismo , Simulação de Dinâmica Molecular , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Precursores Enzimáticos/química , Precursores Enzimáticos/metabolismo , Humanos , Metaloproteinase 9 da Matriz/química , Ligação Proteica , Domínios Proteicos , Multimerização Proteica , Eletricidade Estática , Inibidor Tecidual de Metaloproteinase-1/antagonistas & inibidores
6.
Exp Parasitol ; 225: 108112, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33964315

RESUMO

Matrix metalloproteinases (MMPs), are implicated in the pathogenesis of multiple sclerosis (MS) and in its animal model, experimental autoimmune encephalomyelitis (EAE). Our aim was to investigate whether amelioration of EAE in Dark Agouti (DA) rats, induced by Trichinella spiralis muscle larvae excretory-secretory products (ES L1), could be related to the level and activity of gelatinases, MMP-9 and MMP-2. Serum levels of MMP-9, MMP-2, NGAL/MMP-9, TIMP-1, and cytokines, evaluated by gel-zymography or ELISA, as well as gelatinases and TIMP-1 expression in the spinal cord (SC), were determined in: i) EAE induced, ii) ES L1-treated EAE induced animals. Milder clinical signs in ES L1-treated EAE induced DA rats were accompanied with lower serum levels of MMP-9 and NGAL/MMP-9 complex. However, the correlation between the severity of EAE and the level of serum MMP-9 was found only in the peak of the disease, with MMP-9/TIMP-1 ratio higher in EAE animals without ES L1 treatment. Lower expression of MMP-9 in SC of ES L1-treated, EAE induced rats, correlated with the reduced number of SC infiltrating cells. In SC infiltrates, in the effector and the recovery phase, production of anti-inflammatory cytokines IL-4 and IL-10 was higher in animals treated with ES L1 prior to EAE induction, compared to untreated EAE animals. Reduced expression of MMP-9 in SC tissue, which correlated with the reduced number of infiltrating cells, might be ascribed to regulatory mechanisms, among which is IL-10.


Assuntos
Antígenos de Helmintos/uso terapêutico , Encefalomielite Autoimune Experimental/metabolismo , Proteínas de Helminto/uso terapêutico , Metaloproteinase 9 da Matriz/metabolismo , Trichinella spiralis/metabolismo , Animais , Citocinas/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Encefalomielite Autoimune Experimental/prevenção & controle , Inflamação , Interleucina-10/metabolismo , Ratos , Índice de Gravidade de Doença , Medula Espinal/imunologia , Medula Espinal/metabolismo , Medula Espinal/patologia , Inibidor Tecidual de Metaloproteinase-1/metabolismo
7.
Biomed Pharmacother ; 140: 111700, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34044279

RESUMO

BACKGROUND: Frozen shoulders (FS) is a major clinical concern, where chronic synovial inflammation, abnormal angiogenesis, and fibrosis represent the critical pathologies in the glenohumeral capsule. However, no pharmacotherapy has been introduced to treat this pathology. Tetrandrine (TET) has been proposed as a treatment for many diseases due to its strong anti-inflammatory, anti-angiogenic, and anti-fibrotic effects. PURPOSE: To study the anti-inflammatory, anti-angiogenic, and anti-fibrotic effects of TET on FS, and identify whether TET can prevent the development of FS in rats. STUDY DESIGN: A controlled laboratory study. METHODS: Forty-eight male Sprague-Dawley (SD) rats were randomly divided into control, TET, and FS groups. The TET group was intraperitoneally injected with TET every 2 days. TET and saline treatment were started on the day of FS surgery. After 8 weeks, the animals were sacrificed, and samples were collected for X-ray examination, glenohumeral range of motion (ROM) evaluation, histology and immunohistochemistry analysis, transmission electron microscopy (TEM) observation, and profibrogenic factors as well as proinflammatory cytokines measurements. RESULTS: No significant difference in shoulder ROM was observed between the TET and control groups, but a significant difference was noted between these groups and the FS group (P < 0.01). Immunohistochemical staining showed no abnormal angiogenesis or fibrosis in the TET group or the control group. However, significant angiogenesis, collagen remodeling, and fibrosis were observed in the FS group, and the expression and proportion of type I and type III collagen in the FS group were significantly higher than those in the TET group or the control group (P < 0.01). TEM observation showed that TET protected the ultrastructure of collagen fibrous reticular arrangement of the articular capsule and prevented the formation of scar-like fibrotic structures, which are unique to FS. The significantly increased expression of Smad7 and the suppressed expression of Smad 2 in the TET group compared with that of the FS group indicated that TET also significantly inhibited the TGF-ß1 intracellular signal pathway. The expression of profibrogenic factors and proinflammatory cytokines in the TET group and the control group was significantly lower than that in the TET group (P < 0.01). CONCLUSION: The results demonstrated that TET protected the normal reticular structure of the capsule during the freezing period and prevented the development of FS by inhibiting inflammation, angiogenesis, and fibrosis in a rat FS model. CLINICAL RELEVANCE: TET may be a safe and effective clinical medication for preventing and treating FS.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Benzilisoquinolinas/uso terapêutico , Bursite/tratamento farmacológico , Inibidores da Angiogênese/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Benzilisoquinolinas/farmacologia , Bursite/metabolismo , Bursite/patologia , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Fibrose , Cápsula Articular/efeitos dos fármacos , Cápsula Articular/metabolismo , Cápsula Articular/patologia , Cápsula Articular/ultraestrutura , Masculino , Metaloproteinase 3 da Matriz/metabolismo , Ratos Sprague-Dawley , Inibidor Tecidual de Metaloproteinase-1/metabolismo
8.
Res Vet Sci ; 137: 30-39, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33932820

RESUMO

The aim of this study was to characterize the protein expression of matrix metalloproteinase-2 (MMP-2) and -- 9 and their inhibitors (TIMP-1 and -2) in mammary tissue of dairy cows with naturally occurring chronic S. aureus intramammary infections (IMI) during active involution. Moreover, the gelatinolytic activity of MMP-2 and -9 in mammary secretions was evaluated. Cows in late lactation that were either uninfected or with chronic naturally acquired S. aureus IMI were included in this study. Protein expression of MMP-2 and -9 in mammary tissues was significantly higher in S. aureus-infected than uninfected quarters at day 14 and 21 of involution. Protein expression of TIMP-1 and -2 was significantly higher in S. aureus-infected than uninfected quarters at day 7, 14 and 21 of involution. The MMP-2/TIMP-1, MMP-2/TIMP-2, MMP-9/TIMP-1 and MMP-9/TIMP-2 ratios were significantly higher in S. aureus-infected compared with uninfected quarters at day 14 of involution. The MMP-2 activity was significantly higher in mammary secretions from S. aureus-infected compared with uninfected quarters at day 1, 2, 7 and 14 of involution. The MMP-9 activity was significantly higher in mammary secretions from infected quarters compared with uninfected quarters at day 7, 14 and 21 of involution. The increased expression of MMP-2 and -9 in mammary tissue as well as the high levels of activity observed in mammary secretion from infected quarters compared with uninfected quarters during active involution, strongly suggests that these gelatinases could contribute to degradation of mammary tissue components during chronic S. aureus IMI. The MMPs/TIMPs imbalance could lead to greater proteolysis and potentially more damage to mammary tissue in S. aureus-infected quarters.


Assuntos
Mastite Bovina/enzimologia , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Staphylococcus aureus , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Animais , Bovinos , Feminino , Regulação Enzimológica da Expressão Gênica , Lactação , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/microbiologia , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Infecções Estafilocócicas/veterinária , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-2/genética
9.
Int J Mol Sci ; 22(7)2021 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-33808256

RESUMO

Histiocytic sarcomas refer to highly aggressive tumors with a poor prognosis that respond poorly to conventional treatment approaches. Oncolytic viruses, which have gained significant traction as a cancer therapy in recent decades, represent a promising option for treating histiocytic sarcomas through their replication and/or by modulating the tumor microenvironment. The live attenuated canine distemper virus (CDV) vaccine strain Onderstepoort represents an attractive candidate for oncolytic viral therapy. In the present study, oncolytic virotherapy with CDV was used to investigate the impact of this virus infection on tumor cell growth through direct oncolytic effects or by virus-mediated modulation of the tumor microenvironment with special emphasis on angiogenesis, expression of selected MMPs and TIMP-1 and tumor-associated macrophages in a murine xenograft model of canine histiocytic sarcoma. Treatment of mice with xenotransplanted canine histiocytic sarcomas using CDV induced overt retardation in tumor progression accompanied by necrosis of neoplastic cells, increased numbers of intratumoral macrophages, reduced angiogenesis and modulation of the expression of MMPs and TIMP-1. The present data suggest that CDV inhibits tumor growth in a multifactorial way, including direct cell lysis and reduction of angiogenesis and modulation of MMPs and their inhibitor TIMP-1, providing further support for the concept of its role in oncolytic therapies.


Assuntos
Sarcoma Histiocítico/metabolismo , Neoplasias/metabolismo , Terapia Viral Oncolítica/métodos , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Cinomose/metabolismo , Cinomose/virologia , Vírus da Cinomose Canina/patogenicidade , Doenças do Cão/imunologia , Cães , Feminino , Xenoenxertos , Sarcoma Histiocítico/veterinária , Sarcoma Histiocítico/virologia , Metaloendopeptidases/metabolismo , Camundongos , Camundongos SCID , Necrose/metabolismo , Neoplasias/virologia , Neovascularização Patológica/metabolismo , Vírus Oncolíticos , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Microambiente Tumoral/fisiologia , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Molecules ; 26(6)2021 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-33799348

RESUMO

Nasopharyngeal carcinoma (NPC) is a common malignant head and neck tumor. Drug resistance and distant metastasis are the predominant cause of treatment failure in NPC patients. Hispidulin is a flavonoid extracted from the bioassay-guided separation of the EtOH extract of Salvia plebeia with strong anti-proliferative activity in nasopharyngeal carcinoma cells (CNE-2Z). In this study, the effects of hispidulin on proliferation, invasion, migration, and apoptosis were investigated in CNE-2Z cells. The [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (MTT) assay and the colony formation assay revealed that hispidulin could inhibit CNE-2Z cell proliferation. Hispidulin (25, 50, 100 µM) also induced apoptosis in a dose-dependent manner in CNE-2Z cells. The expression of Akt was reduced, and the expression of the ratio of Bax/Bcl-2 was increased. In addition, scratch wound and transwell assays proved that hispidulin (6.25, 12.5, 25 µM) could inhibited the migration and invasion in CNE-2Z cells. The expressions of HIF-1α, MMP-9, and MMP-2 were decreased, while the MMPs inhibitor TIMP1 was enhanced by hispidulin. Moreover, hispidulin exhibited potent suppression tumor growth and low toxicity in CNE-2Z cancer-bearing mice at a dosage of 20 mg/kg/day. Thus, hispidulin appears to be a potentially effective agent for NPC treatment.


Assuntos
Apoptose/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Flavonas/farmacologia , Flavonoides/farmacologia , Carcinoma Nasofaríngeo/tratamento farmacológico , Salvia/química , Animais , Linhagem Celular Tumoral , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Carcinoma Nasofaríngeo/metabolismo , Neoplasias Nasofaríngeas/tratamento farmacológico , Neoplasias Nasofaríngeas/metabolismo , Invasividade Neoplásica/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo
11.
J Cell Mol Med ; 25(10): 4572-4582, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33733597

RESUMO

In this study, we aimed to explore the molecular mechanisms underlying the development of osteoporosis in post-menopausal females. Real-time PCR was conducted to measure the expression of potential lncRNAs involved in the osteoporosis of post-menopausal females. In addition, Western blot and IHC assays were used to study the possible correlation among HOTAIR, miR-138 and TIMP1, while a computational analysis was carried out to predict the 'seed sequence' responsible for the binding between miR-138 and HOTAIR/TIMP1. Furthermore, luciferase reporter assays were conducted to validate the negative regulatory relationship between miR-138 and TIMP1/HOTAIR. To evaluate the effect of oestrogen on the function of HOATIR and its downstream effectors, luciferase activity was measured in cells cotransfected with different vectors or treated with different doses of oestrogen. The results of the luciferase assay were further validated by real-time PCR, Western blot, MTT assay and flow cytometry. Among the candidate lncRNAs, HOTAIR was the only lncRNA down-regulated in post-menopausal females. HOTAIR bound to miR-138 and negatively regulated its expression. Meanwhile, miR-138 could also bind to TIMP1 mRNA and reduce its expression. Furthermore, a dose-dependent up-regulation of HOTAIR was observed in cells treated with oestrogen, and the elevated HOTAIR increased the level of TIMP1 by targeting miR-138. In addition, oestrogen promoted cell viability and suppressed cell apoptosis, and effects of oestrogen were blocked by the silencing of HOTAIR. Therefore, it can be concluded that oestrogen deficiency could induce the apoptosis of osteoblasts and lead to osteoporosis in post-menopausal females via modulation of the HOTAIR/miR-138/TIMP1 signalling axis.


Assuntos
Biomarcadores/metabolismo , Estrogênios/deficiência , MicroRNAs/genética , Osteoblastos/patologia , Pós-Menopausa , RNA Longo não Codificante/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Adulto , Apoptose , Proliferação de Células , Células Cultivadas , Regulação da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Pré-Menopausa , Inibidor Tecidual de Metaloproteinase-1/genética
12.
J Ethnopharmacol ; 272: 113938, 2021 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-33610708

RESUMO

ETHNOBOTANICAL RELEVANCE: Tectona grandis L.f (or syn: Jatus grandis (L.f.) Kuntze Revis), from family Lamiaceae, also known as Teak, is widely recognized in ayurvedic system of medicine and confer curative potential against inflammation, liver disorders, biliousness, diabetes, bronchitis, leprosy and dysentery. Its leaves are rich source of edible food colorant and reported nontoxic for liver and various organs. AIM OF STUDY: Hepatic injury progression to liver cirrhosis and cancer is a serious health issue across the world. Currently, anti-fibrotic therapeutic options are limited and expensive with no FDA approved direct anti-hepato-fibrotic drug validated in clinic. Thus, the aim of this study was to understand ameliorative effect of Tectona grandis L.f, leaves in early liver fibrosis. METHOD AND RESULTS: C57BL/6 mice suffering from CCl4 induced liver injury, were orally administered at three different doses (50, 100 & 200 mg/kg) of Tectona grandis L.f, leaf extract, thrice a week, up to 4 and 8 weeks. Anti-fibrotic effect was evaluated through animal body/liver weight measurements, serological tests (AST, ALT, GSH, MDA and LDH assays), tissue hydroxyproline content, and histochemical analysis (H&E, Masson trichrome, Sirius red and αSMA localization). Moreover, transcriptional and post-transcriptional expression of fibrosis associated biomarkers and TGF-ß/Smad cascade were analyzed. It was observed that 100 mg/kg dose optimally downregulated TGF-ß1/Smad2 with upregulation of Smad7 and regulated αSMA, Col 1, PDGF, TIMP1 and MMP3 expression, post 8 weeks of treatment. In addition, MMP3/TIMP1 ratio was upregulated to 0.7, 2.5 and 1.7 fold at 50 mg/kg, 100 mg/kg & 200 mg/kg treatments respectively, in comparison to untreated liver fibrosis models. The extract contains gallic acid, caffeic acid, sinapinic acid and myricetin when analyzed through high performance liquid chromatography. CONCLUSION: Tectona grandis L.f, leaves have potential to ameliorate liver fibrosis induced by CCl4 in mice via modulation of TGF-ß1/Smad pathway and upregulated MMP3/TIMP1 ratio.


Assuntos
Lamiaceae/química , Cirrose Hepática/prevenção & controle , Metaloproteinase 3 da Matriz/metabolismo , Substâncias Protetoras/farmacologia , Substâncias Protetoras/envenenamento , Transdução de Sinais/efeitos dos fármacos , Proteína Smad2/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Tetracloreto de Carbono/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Colágeno/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Células Hep G2 , Células Estreladas do Fígado/efeitos dos fármacos , Humanos , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Metaloproteinase 3 da Matriz/genética , Camundongos Endogâmicos C57BL , Extratos Vegetais/administração & dosagem , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Folhas de Planta/química , Substâncias Protetoras/química , Proteína Smad2/genética , Inibidor Tecidual de Metaloproteinase-1/genética , Transaminases/sangue , Fator de Crescimento Transformador beta/genética , Células Vero
13.
J Tradit Chin Med ; 41(1): 98-106, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33522202

RESUMO

OBJECTIVE: To explore the effects of Taohong Siwu decoction (, THSWD) on the extracellular matrix of endometrium in rats following drug-induced abortion. METHODS: Thirty-six pregnant female rats were administered mifepristone and misoprostol to induce abortion, and amounts of uterine bleeding were recorded. Pathological damage and collagen accumulation were detected by hematoxylin-eosin staining and Masson's trichrome staining in uterus, respectively. Myeloperoxidase was evaluated by immunohistochemistry.The expression levels of fibronectin, laminin, matrix metalloproteinase 9 (MMP-9), and tissue inhibitor of metalloproteinase 1 (TIMP-1) were quantified using western blotting. RESULTS: THSWD could promote endometrial protection in rats following drug-induced abortion. The contents of cellulose and myeloperoxidase were significantly decreased in uterine tissue of THSWD-treated groups. Moreover, THSWD significantly decreased the expression levels of fibronectin, laminin, and TIMP-1. THSWD also significantly increased MMP-9 expression and the MMP-9/TIMP1 ratio. CONCLUSION: THSWD plays a critical role in endometrial protection by reducing extracellular matrix deposition and uterine fibrosis. These effects may have been achieved by increasing MMP-9, reducing TIMP-1, and/or altering the ratio of MMP-9/TIMP-1.


Assuntos
Aborto Espontâneo/tratamento farmacológico , Medicamentos de Ervas Chinesas/administração & dosagem , Endométrio/efeitos dos fármacos , Matriz Extracelular/efeitos dos fármacos , Aborto Induzido , Aborto Espontâneo/metabolismo , Animais , Endométrio/metabolismo , Matriz Extracelular/metabolismo , Feminino , Humanos , Masculino , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Gravidez , Ratos , Ratos Sprague-Dawley , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Útero/efeitos dos fármacos , Útero/metabolismo
14.
Biomed Pharmacother ; 137: 111337, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33582453

RESUMO

BACKGROUND: Senna alata L. Roxb or candle bush is a traditional medicinal plant with a wide range of biological activities including anti-inflammatory, antimicrobial and antifungal. Leaf extract of S. alata showed the anti-tumor activity in various cancer cell lines. In this study, we focused on the inhibitory mechanism of S. alata extract (SAE) on cancer metastasis including cell migration, cell invasion and signaling pathways in chondrosarcoma SW1353 cells. PURPOSE: This study aimed to evaluate the anti-metastatic mechanisms of Senna alata extract on chondrosarcoma SW1353 cells. METHODS: Screening for phytochemicals in biologically active fraction of SAE was analysed by 1H NMR spectroscopy. Cell viability and cytoxicity were determined by using MTT assay. Cell migration was observed by scratch wound healing and transwell migration assay. Cell invasion and cell adhesion assay were examined by Matrigel coated transwell chambers or plates. The expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), MAPKs and PI3K/Akt signaling pathways and NF-κB were detected by Western blot analysis. RESULTS: The SAE treatment at the sub-cytoxic and non-cytotoxic concentrations significantly inhibited cell migration, cell invasion and cell adhesion of SW1353 cells in a dose-dependent manner. The results from Western blot analysis showed decreased MMP-2 and MMP-9 expression, while increased TIMP-1 and TIMP-2 expression in SAE treated cells. Moreover, SAE suppressed phosphorylation of ERK1/2, p38 and Akt but decreased NF-κB transcription factor expression in SW1353 cells. CONCLUSION: These results revealed that SAE could reduce MMP-2 and MMP-9 expression by downregulation of NF-κB which is downstream of MAPKs and PI3K/Akt signaling pathway in SW1353 cells resulting in reduced cancer cell migration and invasion. Therefore, SAE may have the potential use as an alternative treatment of chondrosarcoma metastasis.


Assuntos
Condrossarcoma/tratamento farmacológico , Metástase Neoplásica/tratamento farmacológico , Extrato de Senna/farmacologia , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Condrossarcoma/metabolismo , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Proteína Oncogênica v-akt/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Extrato de Senna/química , Transdução de Sinais/efeitos dos fármacos , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Inibidor Tecidual de Metaloproteinase-2/efeitos dos fármacos , Inibidor Tecidual de Metaloproteinase-2/metabolismo
15.
Biomolecules ; 11(2)2021 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-33573220

RESUMO

Metalloproteinases (MPs) are proteolytic enzymes involved in extracellular matrix deposition, regulation of cellular signals of inflammation, proliferation, and apoptosis. Metalloproteinases are classified into three families: Matrix-MPs (MMPs), A-Disintegrin-and-Metalloprotease (ADAMs), and the A-Disintegrin-and-Metalloproteinase-with-Thrombospondin-1-like-Domains (ADAMTS). Previous studies showed that MPs are involved in the development of aortic aneurysms (AA) and, concomitantly, in the onset of chronic kidney disease (CKD). CKD has been, per se, associated with an increased risk for AA. The aim of this review is to examine the pathways that may associate MPs with CKD and AA. Several MMPs, such as MMP-2, -8, -9, and TIMP-1 have been shown to damage the AA wall and to have a toxic effect on renal tubular cells, leading to fibrosis. Similarly, ADAM10 and 17 have been shown to degrade collagen in the AA wall and to worsen kidney function via pro-inflammatory stimuli, the impairment of the Renin-Angiotensin-Aldosterone System, and the degradation of structural proteins. Moreover, MMP-2 and -9 inhibitors reduced aneurysm growth and albuminuria in experimental and human studies. It would be important, in the future, to expand research on MPs from both a prognostic, namely, to refine risk stratification in CKD patients, and a predictive perspective, likely to improve prognosis in response to targeted treatments.


Assuntos
Aneurisma Aórtico/fisiopatologia , Taxa de Filtração Glomerular , Metaloproteases/metabolismo , Insuficiência Renal Crônica/fisiopatologia , Aneurisma/fisiopatologia , Animais , Aneurisma Aórtico/complicações , Aneurisma Aórtico/enzimologia , Apoptose , Proliferação de Células , Progressão da Doença , Transição Epitelial-Mesenquimal , Matriz Extracelular/metabolismo , Fibrose , Humanos , Inflamação , Falência Renal Crônica/fisiopatologia , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 8 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Prognóstico , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/enzimologia , Sistema Renina-Angiotensina , Risco , Inibidor Tecidual de Metaloproteinase-1/metabolismo
16.
Zhongguo Fei Ai Za Zhi ; 24(2): 69-77, 2021 Feb 20.
Artigo em Chinês | MEDLINE | ID: mdl-33478195

RESUMO

BACKGROUND: Cullin1 is a representative member of the Cullin family, and it plays an important role in the ubiquitination of cell cycle, transcription and signal transduction related proteins. Cullin1 is closely related to the occurrence and development of a variety of malignant tumors. The aim of this study is to investigate the effects of Cullin1 on biological function of lung adenocarcinoma A549 and H1395 Cells. METHODS: The expression of Cullin1 mRNA was detected by quantitative Real-time polymerase chain reaction in lung adenocarcinoma cells (A549, H358, H1395, H1650) and human normal lung epithelial cells BEAS-2B, siRNA technology was used to interfere with lung adenocarcinoma cells with relatively high expression of Cullin1 mRNA; cell proliferation, cell cycle distribution, early cell apoptosis, invasion and migration ability were detected by methyl thiazolyl tetrazolium assay (MTT), flow cytometry and Transwell experiment; Western blot was used to detect the expression levels of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), tissue inhibitor of metalloproteinase-1 (TIMP-1), Cyclin D1, Cyclin E2, p21 and p27. RESULTS: Compared with the BEAS-2B cell, Cullin1 mRNA was highly expressed in lung adenocarcinoma cells, especially in lung adenocarcinoma A549 and H1395 cells (P<0.05). The proliferation ability of lung adenocarcinoma cells was inhibited after interference with Cullin1, and the number of cells in G1 phase increased, the number of cells in S phase decreased, and the early apoptosis rate of lung adenocarcinoma cells is significantly increased (P<0.05); The invasion and migration ability of lung adenocarcinoma cells decreased (P<0.05). After interference with Cullin1, the protein expression of MMP-9, MMP-2, CyclinD1 and CyclinE2 decreased (P<0.05), while the expression of TIMP-1, p21 and p27 protein increased (P<0.05). CONCLUSIONS: Interference with Cullin1 inhibits the proliferation, invasion and migration of lung adenocarcinoma A549 and H1395 cells, Cullin1 plays a role in promoting cancer in lung adenocarcinoma.


Assuntos
Proteínas Culina/metabolismo , Neoplasias Pulmonares/metabolismo , Células A549 , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/metabolismo , Adenocarcinoma de Pulmão/fisiopatologia , Apoptose , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Proteínas Culina/genética , Ciclina D1/genética , Ciclina D1/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/fisiopatologia , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Invasividade Neoplásica , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo
17.
Int J Mol Sci ; 22(3)2021 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-33498591

RESUMO

The periodontal ligament (PDL) responds to applied orthodontic forces by extracellular matrix (ECM) remodeling, in which human periodontal ligament-derived mesenchymal stromal cells (hPDL-MSCs) are largely involved by producing matrix metalloproteinases (MMPs) and their local inhibitors (TIMPs). Apart from orthodontic forces, the synthesis of MMPs and TIMPs is influenced by the aseptic inflammation occurring during orthodontic treatment. Interleukin (IL)-1ß is one of the most abundant inflammatory mediators in this process and crucially affects the expression of MMPs and TIMPs in the presence of cyclic low-magnitude orthodontic tensile forces. In this study we aimed to investigate, for the first time, how IL-1ß induced expression of MMPs, TIMPs and how IL-1ß in hPDL-MSCs was changed after applying in vitro low-magnitude orthodontic tensile strains in a static application mode. Hence, primary hPDL-MSCs were stimulated with IL-1ß in combination with static tensile strains (STS) with 6% elongation. After 6- and 24 h, MMP-1, MMP-2, TIMP-1 and IL-1ß expression levels were measured. STS alone had no influence on the basal expression of investigated target genes, whereas IL-1ß caused increased expression of these genes. In combination, they increased the gene and protein expression of MMP-1 and the gene expression of MMP-2 after 24 h. After 6 h, STS reduced IL-1ß-induced MMP-1 synthesis and MMP-2 gene expression. IL-1ß-induced TIMP-1 gene expression was decreased by STS after 6- and 24-h. At both time points, the IL-1ß-induced gene expression of IL-1ß was increased. Additionally, this study showed that fetal bovine serum (FBS) caused an overall suppression of IL-1ß-induced expression of MMP-1, MMP-2 and TIMP-1. Further, it caused lower or opposite effects of STS on IL-1ß-induced expression. These observations suggest that low-magnitude orthodontic tensile strains may favor a more inflammatory and destructive response of hPDL-MSCs when using a static application form and that this response is highly influenced by the presence of FBS in vitro.


Assuntos
Interleucina-1beta/farmacologia , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 2 da Matriz/genética , Células-Tronco Mesenquimais/metabolismo , Ligamento Periodontal/citologia , Estresse Mecânico , Células Cultivadas , Humanos , Metaloproteinase 1 da Matriz/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo
18.
Biochem Biophys Res Commun ; 534: 843-848, 2021 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-33183761

RESUMO

Multiple myeloma (MM) is an intractable hematological malignancy characterized by abnormal plasma cells in the bone marrow (BM) and increased osteolytic lesions. Within the BM niche, mesenchymal stem cells (MSCs) have been proposed to contribute to functionally important MM-MSC interactions. However, despite various studies on MM pathology, the impact of MM on MSCs during the early stages of malignancy has not been adequately addressed. We previously identified tissue inhibitor of matrix metalloproteinase 1 (TIMP-1) as a cytokine that is modulated in vivo within the MM BM niche, and highlighted its potential relevance in MM. Given the role of TIMP-1 in preventing migration of breast cancer cells, this study aimed to investigate the relationship between MSC-secreted TIMP-1 and MM progression. Here, we examined the effect of MSC-derived TIMP-1 on MM cell migration, and found that TIMP-1 secreted by human MSCs play a role in preventing migration of MM cells by reducing the levels of MM cell-derived MMP-9. We also investigated how MM cells regulate expression of TIMP-1 in MSCs. Using a knockdown approach in MSCs, we implicated TGF-B activated kinase 1 binding protein 1 (TAB1) as an upstream effector of TIMP-1 that was downregulated in the presence of MM cells, which resulted in reduced TIMP-1 secretion. Overall, our findings uncover how MSCs in the MM BM niche are modulated to promote MM progression, and unravel a previously unreported role of the TAB1-TIMP-1 axis in the context of the MM BM niche.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Células-Tronco Mesenquimais/patologia , Mieloma Múltiplo/patologia , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Movimento Celular , Técnicas de Cocultura , Regulação Neoplásica da Expressão Gênica , Humanos , Metaloproteinase 9 da Matriz/metabolismo , Células-Tronco Mesenquimais/metabolismo , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo , Inibidor Tecidual de Metaloproteinase-1/genética
19.
Environ Toxicol ; 36(2): 168-176, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32918399

RESUMO

Non-alcoholic fatty liver disease (NAFLD) affects around 25% of the worldwide population. Non-alcoholic steatohepatitis (NASH), the more progressive variant of NAFLD, is characterized by steatosis, cellular ballooning, lobular inflammation, and may culminate on hepatic stellate cell (HSC) activation, thus increasing the risk for fibrosis, cirrhosis, and HCC development. Conversely, the antifibrotic effects of sorafenib, an FDA-approved drug for HCC treatment, have been demonstrated in 2D cell cultures and animal models, but its mechanisms in a NAFLD-related microenvironment in vitro requires further investigation. Thus, a human 3D co-culture model of fatty hepatocytes and HSC was established by culturing hepatoma C3A cells, pre-treated with 1.32 mM oleic acid, with HSC LX-2 cells. The fatty C3A/LX-2 spheroids showed morphological and molecular hallmarks of altered lipid metabolism and steatosis-induced fibrogenesis, similarly to the human disease. Sorafenib (15 µM) for 72 hours reduced fatty spheroid viability, and upregulated the expression of lipid oxidation- and hydrolysis-related genes, CPT1 and LIPC, respectively. Sorafenib also inhibited steatosis-induced fibrogenesis by downregulating COL1A1, TGFB1, PDGF, and TIMP1 and by decreasing protein levels of IL-6, TGF-ß1, and TNF-α in fatty spheroids. The demonstration of the antifibrotic properties of sorafenib on steatosis-induced fibrogenesis in a 3D in vitro model of NAFLD supports its clinical use as a therapeutic agent for the treatment of NAFLD/NASH patients.


Assuntos
Células Estreladas do Fígado/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Cirrose Hepática/prevenção & controle , Fígado/efeitos dos fármacos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Sorafenibe/farmacologia , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Fibrose , Células Estreladas do Fígado/patologia , Hepatócitos/patologia , Humanos , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
20.
J Ethnopharmacol ; 267: 113506, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33148433

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Alstonia scholaris (L.) R. Br. (Apocynaceae) is a Dai folk medicine for the treatment of lung diseases in China. AIM OF THE STUDY: The present study investigated the anti-pulmonary fibrosis effects of total alkaloids (TA) and the potential active ingredients and its possible mechanism. MATERIALS AND METHODS: After intratracheal instillation of bleomycin (BLM, 5 mg/kg), mice were divided into ten groups, and orally treated with the corresponding samples once daily for 28 days. The effect of indole alkaloids was determined through analysis of cytokines, as well as histopathological examinations and gene expressions. RESULTS: Severe lung fibrosis was observed in the BLM-treated mice on day 28. However, the administration of TA significantly ameliorated the pathological changes in the lungs, decreased the content of Krebs von den Lungen-6, lactate dehydrogenase, transforming growth factor-ß (TGF-ß), hydroxyproline, type I collagen, and malonaldehyde, and enhanced the activity of superoxide dismutase in the serum and lung tissues. In addition, the enhanced TGF-ß and matrix metalloproteinase-1 (MMP-1) expressions in BLM-induced mice were obviously weakened by indole alkaloids, as well as the ratio of matrix metalloproteinase-1 to tissue inhibitor of metalloproteinase-1 was decreased. Moreover, picrinine and scholaricine yielded markedly better values in the aforementioned indices than those in other samples, indicating that they may be the active ingredients of alkaloids. CONCLUSIONS: TA exerted protective effects against BLM-induced pulmonary fibrosis by reducing collagen deposition through TGF-ß/MMP-1 pathway.


Assuntos
Alstonia , Alcaloides Indólicos/farmacologia , Pulmão/efeitos dos fármacos , Extratos Vegetais/farmacologia , Fibrose Pulmonar/prevenção & controle , Alstonia/química , Animais , Bleomicina , Colágeno/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica , Alcaloides Indólicos/isolamento & purificação , Mediadores da Inflamação/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Masculino , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , Camundongos Endogâmicos ICR , Extratos Vegetais/isolamento & purificação , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Transdução de Sinais , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...