RESUMEN
The role extracellular matrix (ECM) in multiple events of morphogenesis has been well described, little is known about its specific role in early eye development. One of the first morphogenic events in lens development is placodal thickening, which converts the presumptive lens ectoderm from cuboidal to pseudostratified epithelium. This process occurs in the anterior pre-placodal ectoderm when the optic vesicle approaches the cephalic ectoderm and is regulated by transcription factor Pax6 and secreted BMP4. Since cells and ECM have a dynamic relationship of interdependence and modulation, we hypothesized that the ECM evolves with cell shape changes during lens placode formation. This study investigates changes in optic ECM including both protein distribution deposition, extracellular gelatinase activity and gene expression patterns during early optic development using chicken and mouse models. In particular, the expression of Timp2, a metalloprotease inhibitor, corresponds with a decrease in gelatinase activity within the optic ECM. Furthermore, we demonstrate that optic ECM remodeling depends on BMP signaling in the placode. Together, our findings suggest that the lens placode plays an active role in remodeling the optic ECM during early eye development.
Asunto(s)
Matriz Extracelular , Regulación del Desarrollo de la Expresión Génica , Cristalino , Factor de Transcripción PAX6 , Animales , Matriz Extracelular/metabolismo , Ratones , Cristalino/metabolismo , Cristalino/crecimiento & desarrollo , Cristalino/citología , Factor de Transcripción PAX6/metabolismo , Factor de Transcripción PAX6/genética , Proteínas del Ojo/metabolismo , Proteínas del Ojo/genética , Proteína Morfogenética Ósea 4/metabolismo , Proteína Morfogenética Ósea 4/genética , Embrión de Pollo , Proteínas de Homeodominio/metabolismo , Proteínas de Homeodominio/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/genética , Factores de Transcripción Paired Box/metabolismo , Factores de Transcripción Paired Box/genética , Proteínas Represoras/metabolismo , Proteínas Represoras/genética , Transducción de Señal , Pollos/genética , Ojo/metabolismo , Ojo/crecimiento & desarrollo , Ojo/embriologíaRESUMEN
Matrix metalloproteinases (MMPs) play a crucial role in the degenerative course of rheumatic disorders. They are responsible for cartilage and other joint-associated tissues breakdown. Amid arthritis treatments, photobiostimulation (PBM), a non-thermal and non-invasive low-power laser application, appears to be an outstanding therapy alternative once it has succeeded in MMPs modulation. Thus, this study aimed to evaluate the PBM effects of low infrared laser (830 nm), testing two different energy densities (3 and 30 Jcm-2) in MMP-2, MMP-9, MMP-13, and MMP-14 as well as the inhibitor TIMP-2 expressions using zymosan-induced arthritis model. C57BL/6 mice were distributed into four groups (n = 8): zymosan-induced arthritis without treatment; zymosan-induced arthritis and dexamethasone-treated; zymosan-induced arthritis and PBM at energy density of 3 Jcm-2 treated; and zymosan-induced arthritis and PBM at energy density of 30 Jcm-2 treated. MMPs and TIMP-2 mRNA relative levels by qRT-PCR and proteins expression by immunohistochemical and Western blotting techniques were performed after PBM treatment in the inflamed joint. Our results demonstrated PBM could modulate both mRNA relative levels and proteins expression of the MMP-2, -9, -13, -14, and TIMP-2 in joint tissues, decreasing MMP-9 protein expression and increasing TIMP-2 protein expression. PBM promotes a better arthritis prognostic, modulating metalloproteinase and its inhibitor, especially MMP-9 and TIMP-2 protein expression that is important inflammatory markers. These findings may also corroborate that PBM may regulate MMPs expression using different pathways.
Asunto(s)
Artritis , Terapia por Luz de Baja Intensidad , Animales , Ratones , Artritis/inducido químicamente , Artritis/genética , Artritis/radioterapia , Metaloproteinasas de la Matriz/genética , Metaloproteinasas de la Matriz/metabolismo , Ratones Endogámicos C57BL , ARN Mensajero/genética , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , ZimosanRESUMEN
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.
Asunto(s)
Mastitis Bovina/enzimología , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Staphylococcus aureus , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Animales , Bovinos , Femenino , Regulación Enzimológica de la Expresión Génica , Lactancia , Glándulas Mamarias Animales/metabolismo , Glándulas Mamarias Animales/microbiología , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/genética , Infecciones Estafilocócicas/veterinaria , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genéticaRESUMEN
To assist in evaluating and quantifying tissue changes, fractal dimension (FD) is a useful method for assessing the organization in an image from fractals that describes the amount of space and the self-similarity of the structure, once FD detects subtle morphological changes and performs functional quantitative measures. Here, we hypothesized that fractal analysis may be different in functional and regressing bovine corpus luteum (CL) and may be correlated with differential expression of genes involved in extracellular matrix remodeling. CL presents two developmental stages, the functional and regressing CL, according to progesterone levels and morphology. First, we found a lower FD in functional CL using HE staining and picrosirius red approach. Additionally, we found a great amount of total collagen in regressing CL. Regarding gene expression, we showed an up regulation of COL1A1, COL1A2, MMP2, and MMP14 and a down regulation of TIMP1 and TIMP2 in regressing CL compared to the functional one. Thus, we concluded that differential FD observed during luteal regression is an effective method to evaluate the tissue changes observed during luteal development in cattle and is related to differential quantity of genes involved in extracellular matrix remodeling.
Asunto(s)
Colágeno Tipo I/genética , Cuerpo Lúteo/metabolismo , Matriz Extracelular/metabolismo , Regulación del Desarrollo de la Expresión Génica , Luteólisis/metabolismo , Animales , Compuestos Azo , Bovinos , Colágeno Tipo I/metabolismo , Cuerpo Lúteo/crecimiento & desarrollo , Cuerpo Lúteo/ultraestructura , Eosina Amarillenta-(YS) , Matriz Extracelular/ultraestructura , Femenino , Fractales , Hematoxilina , Histocitoquímica/métodos , Luteólisis/genética , Metaloproteinasa 14 de la Matriz/genética , Metaloproteinasa 14 de la Matriz/metabolismo , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismoRESUMEN
OBJECTIVE: Endometriosis is characterized by the growth of endometrial tissue outside the uterine cavity. The prevalence of endometriosis among women experiencing pain, infertility, or both is as high as 35% to 50%. The most common symptoms of endometriosis are dysmenorrhea, dyspareunia, chronic pelvic pain, and infertility. Evidence has suggested that endometriosis symptoms result from a local inflammatory peritoneal reaction caused by ectopic endometrial implants that undergo cyclic bleeding. On the other hand, regular physical exercise seems to have protective effects against diseases that involve inflammatory processes such as type 2 diabetes and colon and breast cancer. On this basis, it is possible that the practice of physical exercise may have beneficial effects on endometriosis. Therefore, the objective of this study was to evaluate the possible anti-inflammatory effect of physical exercise on endometriosis experimentally induced in rats. STUDY DESIGN: Seventy female Wistar rats were divided into 7groups of 10 animals each. Animals performed light exercise (swimming once a week), moderate exercise (swimming 3 times a week), and intense exercise (swimming 5 times a week) before or after endometriosis induction. RESULTS: At the end of the experimental protocol, a reduction in the size of endometriotic lesions was observed after physical exercise regardless of its frequency, with a greater reduction in the groups practicing moderate and intense activity; an increase in FAS levels and a decrease in matrix metalloproteinases 9 and proliferating cell nuclear antigen (PCNA)levels was also observed. The immunohistochemistry results did not lead to conclusive results. As expected, oxidative stress was reduced in all groups. These results show that the practice of physical exercise could be beneficial, at least in part, for the treatment of endometriosis.
Asunto(s)
Endometriosis/terapia , Condicionamiento Físico Animal , Animales , Modelos Animales de Enfermedad , Endometriosis/metabolismo , Endometriosis/patología , Endometrio/química , Femenino , Inflamación/prevención & control , Metaloproteinasa 9 de la Matriz/análisis , Metaloproteinasa 9 de la Matriz/genética , Estrés Oxidativo/fisiología , Antígeno Nuclear de Célula en Proliferación/análisis , Antígeno Nuclear de Célula en Proliferación/genética , ARN/análisis , Ratas , Ratas Wistar , Natación , Inhibidor Tisular de Metaloproteinasa-2/análisis , Inhibidor Tisular de Metaloproteinasa-2/genética , Receptor fas/análisis , Receptor fas/genéticaRESUMEN
Dysregulated expression of tissue inhibitors of matrix metalloproteinases (TIMPs) is associated with systolic dysfunction and worsening heart failure (HF). However, no study has assessed the relationship between TIMP polymorphisms and chronic HF. In this study, 300 HF outpatients with reduced left ventricular ejection fraction and 304 healthy blood donors were genotyped for the 372 T > C polymorphism (Phe124Phe; rs4898) in the TIMP-1 gene and the -418 G > C polymorphism (rs8179090) in the TIMP-2 gene to investigate whether these polymorphisms are associated with HF susceptibility and prognosis. The genotype and allele frequencies of the 372 T > C polymorphism in HF patients were not significantly different from those observed among healthy subjects, and the C allele of the -418 G > C polymorphism was very rare in our population (frequency < 1%). After a median follow-up duration of 5.5 years, 121 patients (40.3%) died (67 of them from HF). Survival analysis did not show statistically significant differences in all-cause death and HF-related death between patients with and without the T allele (P > 0.05 for all comparisons). Thus, our findings do not support the hypothesis that the 372 T > C (Phe124Phe) polymorphism in the TIMP-1 gene and the -418 G > C polymorphism in the TIMP-2 gene are associated with HF susceptibility and prognosis in Southern Brazilians.
Asunto(s)
Insuficiencia Cardíaca/genética , Polimorfismo de Nucleótido Simple , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Adulto , Femenino , Humanos , MasculinoRESUMEN
Liver fibrosis occurs in the presence of continuous insults, including toxic or biological agents. Novel treatments must focus on ceasing the progression of cellular damage, promoting the regeneration of the parenchyma and inhibition of the fibrotic process. The present study analyzed the effect of bone morphogenetic protein (BMP)7 gene therapy with or without cotreatment with doxazosin in a model of liver cirrhosis in hamsters. The serum alanine aminotransferase, aspartate aminotransferase and albumin levels were analyzed spectrophotometrically. Tissue hepatic samples were analyzed by hematoxylin and eosin for parenchymal structure and Sirius red for collagen fiber content. BMP7 and αsmooth muscle actin (SMA)positive cells were detected by immunohistochemistry. BMP7 and collagen type I content in hepatic tissue were analyzed by western blotting, and tissue inhibitor of metalloproteinases (TIMP)2 and matrix metalloproteinase (MMP)13 expression levels were detected by reverse transcriptionquantitative polymerase chain reaction. The present study detected a significant reduction of collagen type I deposits in the group treated with adenoviraltransduction with BMP7 and doxazosin. In animals with BMP7 and doxazosin therapy, αSMApositive cells were 31.7 and 29% significantly decreased compared with animals with placebo, respectively. AdenoviralBMP7 transduction and/or doxazosin treatments actively induced decrement in type I collagen deposition via increased MMP13 and reduced TIMP2 expression. In conclusion, the adenovirusBMP7 gene therapy and the doxazosin therapy are potential candidates for the diminution of fibrosis in the liver, although combination of both therapies does not improve the individual antifibrotic effect once cirrhosis is established.
Asunto(s)
Adenoviridae/metabolismo , Proteína Morfogenética Ósea 7/metabolismo , Doxazosina/uso terapéutico , Cirrosis Hepática/tratamiento farmacológico , Antagonistas Adrenérgicos alfa/farmacología , Animales , Colágeno Tipo I/metabolismo , Cricetinae , Modelos Animales de Enfermedad , Doxazosina/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Células Estrelladas Hepáticas/efectos de los fármacos , Células Estrelladas Hepáticas/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Hígado/fisiopatología , Cirrosis Hepática/patología , Cirrosis Hepática/fisiopatología , Masculino , Metaloproteinasa 13 de la Matriz/genética , Metaloproteinasa 13 de la Matriz/metabolismo , Receptores Adrenérgicos alfa/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Transducción GenéticaRESUMEN
Abstract Anterior open bite (AOB) has a multifactorial etiology caused by the interaction of sucking habits and genetic factors. The aim of this study was to evaluate the association between AOB and polymorphisms in genes that encode Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs). Four hundred and seventy-two children that presented at least one sucking habit were evaluated. Children were examined clinically for the presence of AOB. Genomic DNA was extracted from saliva. Genotyping of the selected polymorphisms in MMP2, MMP3, MMP9, TIMP1 and TIMP2 was carried out by real-time PCR using the TaqMan method. Allele and genotype frequencies were compared between the groups with and without AOB using the PLINK® software in a free and in a recessive model using a chi-square test. Logistic regression analysis was implemented (p≤0.05). Two hundred nineteen children had AOB while 253 did not. The polymorphism rs17576 in MMP9 was significantly associated with AOB (p=0.009). In a recessive model GG genotype was a protective factor for AOB (p=0.014; OR 4.6, 95%CI 1.3-16.2). In the logistic regression analysis, none of the genes was associated with AOB. In conclusion, the polymorphism rs17576 (glutamine for arginine substitution) in MMP9 was a protective factor for AOB.
Resumo A mordida aberta anterior apresenta uma etiologia multifatorial causada pela interação entre hábitos de sucção e fatores genéticos. O objetivo deste estudo foi avaliar a associação entre mordida aberta anterior e polimorfismo nos genes que codificam as metaloproteinases da matriz (MMPs) e seus inibidores teciduais (TIMPs). Foram avaliadas 472 crianças que apresentvam pelo menos um hábito de sucção. As crianças foram clinicamente examinadas para avaliar a presença de mordida aberta anterior. DNA genômico foi extraído da saliva. A genotipagem dos polimorfismos selecionados em MMP2, MMP3, MMP9, TIMP1 e TIMP2 foi realizada por PCR em tempo real, usando o método de TaqMan. As frequências alélicas e genotípicas foram comparadas entre os grupos com e sem mordida aberta anterior usando o software PLINK®. Duzentas e dezenove crianças apresentavam mordida aberta anterior enquanto 253 não a apresentavam. O polimorfismo rs17576 em MMP9 estava significativamente associado com mordida aberta anterior (p=0,009). No modelo recessivo (GG versus AG+AA) o genótipo GG foi um fator protetor para mordida aberta anterior (p=0,014; OR 4,6; 95%CI 1,3- 16,2). Concluindo, o polimorfismo rs17576 (substituição de glutamina por arginina) em MMP9 está associado com mordida aberta anterior. Os resultados obtidos suportam a hipótese de que fatores genéticos estão envolvidos com a etiologia da mordida aberta anterior.
Asunto(s)
Humanos , Masculino , Femenino , Preescolar , Niño , Metaloproteinasa 9 de la Matriz/genética , Polimorfismo de Nucleótido Simple , Mordida Abierta/etiología , Metaloproteinasa 3 de la Matriz/genética , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Metaloproteinasa 2 de la Matriz/genética , Mordida Abierta/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Succión del Dedo , Frecuencia de los Genes , Genotipo , Modelos GenéticosRESUMEN
OBJECTIVE: To evaluate the association between polymorphisms in DLX1, DLX2, MMP13, TIMP1 and TIMP2 genes with dental fluorosis (DF) phenotype. DESIGN: Four hundred and eighty one subjects (108 with DF and 373 DF free) from 6 to 18 years of age were recruited. This population lived in Rio de Janeiro, a city with fluoridation of public water supplies. DF was assessed using the Deans index modified. Only erupted permanent teeth were assessed. Genetic polymorphisms in DLX1, DLX2, MMP13, TIMP1 and TIMP2 were analyzed by real-time PCR from genomic DNA. Association between DF, genotype, and allele distribution were evaluated using chi-square and logistic regression analyses with an alpha level of 5%. RESULTS: DF was more prevalent in Afro-descendants than in Caucasians (p=0.08; OR=1.83; CI 95%=1.18-2.82). Logistic regression analysis adjusted by the ethnicity demonstrated a statistical difference for TIMP1 genotype (p=0.033; OR=2.93, 95%CI, 1.09-7.90). When only the severer cases of DF were analyzed, polymorphisms in DLX1 and DLX2 were associated with DF (p<0.05). CONCLUSION: Our results provided evidence that polymorphisms in TIMP1, DLX1 and DLX2 genes may be associated with DF phenotypes.
Asunto(s)
Amelogénesis/genética , Fluorosis Dental/genética , Adolescente , Alelos , Brasil , Niño , Estudios Transversales , Femenino , Genotipo , Proteínas de Homeodominio/genética , Humanos , Masculino , Metaloproteinasa 1 de la Matriz/genética , Fenotipo , Polimorfismo Genético , Reacción en Cadena en Tiempo Real de la Polimerasa , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Factores de Transcripción/genéticaRESUMEN
Anterior open bite (AOB) has a multifactorial etiology caused by the interaction of sucking habits and genetic factors. The aim of this study was to evaluate the association between AOB and polymorphisms in genes that encode Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs). Four hundred and seventy-two children that presented at least one sucking habit were evaluated. Children were examined clinically for the presence of AOB. Genomic DNA was extracted from saliva. Genotyping of the selected polymorphisms in MMP2, MMP3, MMP9, TIMP1 and TIMP2 was carried out by real-time PCR using the TaqMan method. Allele and genotype frequencies were compared between the groups with and without AOB using the PLINK® software in a free and in a recessive model using a chi-square test. Logistic regression analysis was implemented (p≤0.05). Two hundred nineteen children had AOB while 253 did not. The polymorphism rs17576 in MMP9 was significantly associated with AOB (p=0.009). In a recessive model GG genotype was a protective factor for AOB (p=0.014; OR 4.6, 95%CI 1.3-16.2). In the logistic regression analysis, none of the genes was associated with AOB. In conclusion, the polymorphism rs17576 (glutamine for arginine substitution) in MMP9 was a protective factor for AOB.
Asunto(s)
Metaloproteinasa 9 de la Matriz/genética , Mordida Abierta/etiología , Polimorfismo de Nucleótido Simple , Niño , Preescolar , Femenino , Succión del Dedo , Frecuencia de los Genes , Genotipo , Humanos , Masculino , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 3 de la Matriz/genética , Modelos Genéticos , Mordida Abierta/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genéticaRESUMEN
The BRAF(V600E) mutation confers constitutive kinase activity and accounts for >90% of BRAF mutations in melanoma. This genetic alteration is a current therapeutic target; however, the antitumorigenic effects of the BRAF(V600E) inhibitor vemurafenib are short-lived and the majority of patients present tumor relapse in a short period after treatment. Characterization of vemurafenib resistance has been essential to the efficacy of next generation therapeutic strategies. Herein, we found that acute BRAF inhibition induced a decrease in active MMP-2, MT1-MMP and MMP-9, but did not modulate the metalloproteinase inhibitors TIMP-2 or RECK in naïve melanoma cells. In vemurafenib-resistant melanoma cells, we observed a lower growth rate and an increase in EGFR phosphorylation followed by the recovery of active MMP-2 expression, a mediator of cancer metastasis. Furthermore, we found a different profile of MMP inhibitor expression, characterized by TIMP-2 downregulation and RECK upregulation. In a 3D spheroid model, the invasion index of vemurafenib-resistant melanoma cells was more evident than in its non-resistant counterpart. We confirmed this pattern in a matrigel invasion assay and demonstrated that use of a matrix metalloproteinase inhibitor reduced the invasion of vemurafenib resistant melanoma cells but not drug naïve cells. Moreover, we did not observe a delimited group of cells invading the dermis in vemurafenib-resistant melanoma cells present in a reconstructed skin model. The same MMP-2 and RECK upregulation profile was found in this 3D skin model containing vemurafenib-resistant melanoma cells. Acute vemurafenib treatment induces the disorganization of collagen fibers and consequently, extracellular matrix remodeling, with this pattern observed even after the acquisition of resistance. Altogether, our data suggest that resistance to vemurafenib induces significant changes in the tumor microenvironment mainly by MMP-2 upregulation, with a corresponding increase in cell invasiveness.
Asunto(s)
Antineoplásicos/farmacología , Indoles/farmacología , Metaloproteinasa 2 de la Matriz/metabolismo , Melanoma/patología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Sulfonamidas/farmacología , Línea Celular Tumoral , Resistencia a Antineoplásicos/fisiología , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/metabolismo , Humanos , Interleucina-8/metabolismo , Metaloproteinasa 14 de la Matriz/genética , Metaloproteinasa 14 de la Matriz/metabolismo , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/metabolismo , Melanoma/genética , Melanoma/metabolismo , Invasividad Neoplásica , Proteínas Proto-Oncogénicas B-raf/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Microambiente Tumoral/efectos de los fármacos , Regulación hacia Arriba , VemurafenibRESUMEN
The aim of this study was to determine the effect of artesunate on extracellular matrix (ECM) accumulation and the expression of collagen-IV, matrix metalloproteinase (MMP), and tissue inhibitor of matrix metalloproteinase (TIMP) to understand the pharmacological role of artesunate in pulmonary fibrosis. Eighty Sprague-Dawley rats were randomly assigned to four groups that were administered saline alone, bleomycin (BLM) alone, BLM + artesunate, or artesunate alone for 28 days. Lung tissues from 10 rats in each group were used to obtain lung fibroblast (LF) primary cells, and the rest were used to analyze protein expression. The mRNA expression of collagen-IV, MMP-2, MMP-9, TIMP-1, and TIMP-2 in lung fibroblasts was detected by real-time quantitative reverse transcriptase polymerase chain reaction. The protein levels of collagen-IV, MMP-2, MMP-9, TIMP-1, and TIMP-2 protein in lung tissues were analyzed by western blotting. Artesunate treatment alleviated alveolitis and pulmonary fibrosis induced by bleomycin in rats, as indicated by a decreased lung coefficient and improvement of lung tissue morphology. Artesunate treatment also led to decreased collagen-IV protein levels, which might be a result of its downregulated expression and increased MMP-2 and MMP-9 protein and mRNA levels. Increased TIMP-1 and TIMP- 2 protein and mRNA levels were detected after artesunate treatment in lung tissues and primary lung fibroblast cells and may contribute to enhanced activity of MMP-2 and -9. These findings suggested that artesunate attenuates alveolitis and pulmonary fibrosis by regulating expression of collagen-IV, TIMP-1 and 2, as well as MMP-2 and -9, to reduce ECM accumulation.
Asunto(s)
Colágeno Tipo IV/biosíntesis , Metaloproteinasa 2 de la Matriz/biosíntesis , Metaloproteinasa 9 de la Matriz/biosíntesis , Fibrosis Pulmonar/tratamiento farmacológico , Inhibidor Tisular de Metaloproteinasa-1/biosíntesis , Inhibidor Tisular de Metaloproteinasa-2/biosíntesis , Animales , Artemisininas/administración & dosificación , Artesunato , Colágeno Tipo IV/genética , Modelos Animales de Enfermedad , Matriz Extracelular/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Pulmón/efectos de los fármacos , Pulmón/patología , Metaloproteinasa 2 de la Matriz/genética , Fibrosis Pulmonar/genética , Fibrosis Pulmonar/patología , ARN Mensajero/biosíntesis , Ratas , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genéticaRESUMEN
The aim of the current study was to examine matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-2 (TIMP-2) expression in patients with cervical disc herniation (CDH). A total of 127 specimens from CDH patients undergoing posterior spinal surgery were obtained for the case group, which was divided into three subgroups: lateral protrusion (N = 102), median protrusion (N = 18), and paramedian protrusion (N = 7). Another 55 specimens from subjects who had cervical spine trauma and underwent spinal canal decompression were obtained for the control group. Routine hematoxylin and eosin staining was performed for pathological diagnosis. Immunohistochemical (IHC) analysis was used to determine MMP-2 and TIMP-2 expression. Under light microscopy, MMP-2-positive cells presented brown-yellow or dark brown staining in the cell membrane or cytoplasm. MMP-2 expression in the case group was significantly higher than that in controls (P < 0.05). Furthermore, MMP-2 expression in the lateral and median protrusion groups was significantly higher compared to that in the paramedian protrusion group (both P < 0.05), while there was no apparent difference in MMP-2 expression between the lateral and median protrusion groups (P > 0.05). IHC results showed that TIMP-2 expression in cases was significantly lower than that in controls (P < 0.05). Spearman correlation analysis indicated that MMP- 2 was negatively correlated with TIMP-2 expression (r = -0.418, P < 0.001). In conclusion, MMP-2 expression increased, whereas TIMP- 2 expression decreased in CDH patients, suggesting that MMP-2 and TIMP-2 expression may contribute to CDH development.
Asunto(s)
Desplazamiento del Disco Intervertebral/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Adulto , Estudios de Casos y Controles , Vértebras Cervicales/metabolismo , Vértebras Cervicales/patología , Femenino , Humanos , Desplazamiento del Disco Intervertebral/genética , Desplazamiento del Disco Intervertebral/patología , Masculino , Metaloproteinasa 2 de la Matriz/genética , Persona de Mediana Edad , Inhibidor Tisular de Metaloproteinasa-2/genéticaRESUMEN
BACKGROUND: Matrix metalloproteinases and their inhibitors might be involved in enamel formation. AIM: This study aimed to evaluate the association between polymorphisms in MMP2, MMP3, MMP9, MMP20, TIMP1, and TIMP2 with white spot lesions (WSL) and early childhood caries (ECC). DESIGN: A cross-sectional study was performed on 786 children aged from 2 to 6 years in Brazil. After clinical evaluation, they were classified into groups with disease (the presence of WSL and/or ECC) and without disease (the absence of WSL or ECC). Genotyping of the selected polymorphisms was carried out with TaqMan real-time PCR, using genomic DNA extracted from buccal cells. Allele and genotype frequencies were compared between groups. Chi-square test, odds ratio (OR), and logistic regression were used (P ≤ 0.05). RESULTS: The dmft score was 1.3 (SD: 2.4), and 41.34% of the children have at least one caries lesion. In MMP9, the GG genotype was more frequent in the group without disease (P = 0.006). In a recessive model, WSL was associated with the marker rs1711437 in MMP20 (P = 0.019; OR = 1.20, 95% CI 1.02-1.42). The marker rs1784418 in MMP20 showed an association between the allele G distribution for the WSL group (P = 0.020; OR = 0.73, 95% CI 0.55-0.96). CONCLUSION: MMP9 and MMP20 are involved in WSL and ECC development.
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Caries Dental/genética , Metaloproteinasas de la Matriz/genética , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Alelos , Niño , Preescolar , Estudios Transversales , ADN/genética , Caries Dental/enzimología , Femenino , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Isoenzimas , Masculino , Mucosa Bucal , Polimorfismo Genético , Reacción en Cadena en Tiempo Real de la Polimerasa , Factores de RiesgoRESUMEN
Liver fibrosis occurring as an outcome of non-alcoholic steatohepatitis (NASH) can precede the development of cirrhosis. We investigated the effects of sorafenib in preventing liver fibrosis in a rodent model of NASH. Adult Sprague-Dawley rats were fed a choline-deficient high-fat diet and exposed to diethylnitrosamine for 6 weeks. The NASH group (n=10) received vehicle and the sorafenib group (n=10) received 2.5 mg·kg(-1)·day(-1) by gavage. A control group (n=4) received only standard diet and vehicle. Following treatment, animals were sacrificed and liver tissue was collected for histologic examination, mRNA isolation, and analysis of mitochondrial function. Genes related to fibrosis (MMP9, TIMP1, TIMP2), oxidative stress (HSP60, HSP90, GST), and mitochondrial biogenesis (PGC1α) were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Liver mitochondrial oxidation activity was measured by a polarographic method, and cytokines by enzyme-linked immunosorbent assay (ELISA). Sorafenib treatment restored mitochondrial function and reduced collagen deposition by nearly 63% compared to the NASH group. Sorafenib upregulated PGC1α and MMP9 and reduced TIMP1 and TIMP2 mRNA and IL-6 and IL-10 protein expression. There were no differences in HSP60, HSP90 and GST expression. Sorafenib modulated PGC1α expression, improved mitochondrial respiration and prevented collagen deposition. It may, therefore, be useful in the treatment of liver fibrosis in NASH.
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Cirrosis Hepática/tratamiento farmacológico , Mitocondrias Hepáticas/efectos de los fármacos , Niacinamida/análogos & derivados , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Compuestos de Fenilurea/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Animales , Chaperonina 60/análisis , Chaperonina 60/genética , Dieta Alta en Grasa/métodos , Dietilnitrosamina , Modelos Animales de Enfermedad , Colágenos Fibrilares/efectos de los fármacos , Glutatión Transferasa/análisis , Glutatión Transferasa/genética , Proteínas HSP90 de Choque Térmico/análisis , Proteínas HSP90 de Choque Térmico/genética , Interleucina-10/análisis , Interleucina-10/genética , Interleucina-6/análisis , Interleucina-6/genética , Cirrosis Hepática/etiología , Cirrosis Hepática/patología , Metaloproteinasa 9 de la Matriz/análisis , Metaloproteinasa 9 de la Matriz/genética , Mitocondrias Hepáticas/metabolismo , Niacinamida/uso terapéutico , Enfermedad del Hígado Graso no Alcohólico/inducido químicamente , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Polarografía , ARN Mensajero/aislamiento & purificación , Ratas Sprague-Dawley , Sorafenib , Inhibidor Tisular de Metaloproteinasa-1/análisis , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/análisis , Inhibidor Tisular de Metaloproteinasa-2/genética , Factores de Transcripción/análisis , Factores de Transcripción/genéticaAsunto(s)
Hipertensión/genética , Infarto del Miocardio/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Anciano , Estudios de Casos y Controles , Femenino , Frecuencia de los Genes , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Masculino , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple , Regiones Promotoras GenéticasRESUMEN
Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) are involved in several reproductive events like oocyte-spermatozoa interaction and semen liquefaction. In order to study their role in the llama oviductal reproductive process, MMP activity in oviductal fluid (OF) was assayed. Considering that llama genome sequences are partially known, a strategy to procure cDNA sequences of MMP-2, MMP-9, TIMP-1 and TIMP-2 was designed. Afterwards, their expression patterns in the different llama oviductal segments were assayed. Gelatine zymograms detected 62 and 94 kDa protease activities that matched MMP-2 and pro-MMP-9, respectively. Expression pattern analysis showed that MMP and TIMP mRNAs were present in ampulla, isthmus, utero-tubal junction (UTJ) and papilla. Altogether, these findings support the argument that MMPs/TIMPs are produced in the oviduct and secreted into the oviductal lumen. Our results encourage further studies to elucidate the role of these proteins in reproductive oviductal events.
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Camélidos del Nuevo Mundo , Trompas Uterinas/enzimología , Metaloproteinasa 2 de la Matriz/análisis , Metaloproteinasa 9 de la Matriz/análisis , Inhibidor Tisular de Metaloproteinasa-1/análisis , Inhibidor Tisular de Metaloproteinasa-2/análisis , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Trompas Uterinas/química , Femenino , Expresión Génica , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/genética , Datos de Secuencia Molecular , ARN Mensajero/análisis , Análisis de Secuencia de ADN , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genéticaRESUMEN
The invasive phenotype of many tumors is associated with an imbalance between the matrix metalloproteinases (MMPs) and their inhibitors, tissue inhibitors of metalloproteinases (TIMPs), and the membrane-anchored reversion-inducing cysteine-rich protein with Kazal motifs (RECK). RECK inhibits MMP-2, MMP-9, and MT1-MMP, and has been linked to patient survival and better prognosis in several types of tumors. However, despite the wide implication of these MMPs in melanoma establishment and progression, the role of RECK in this type of tumor is still unknown. Here, we analyzed the expression of RECK, TIMP1, TIMP2, TIMP3, MT1MMP, MMP2, and MMP9 in two publicly available melanoma microarray datasets and in a panel of human melanoma cell lines. We found that RECK is downregulated in malignant melanoma, accompanied by upregulation of MT1MMP and TIMP2. In both datasets, we observed that the group of samples displaying higher RECK levels show lower median expression levels of MT1MMP and TIMP2 and higher levels of TIMP3. When tested in a sample-wise manner, these correlations were statistically significant. Inverse correlations between RECK, MT1MMP, and TIMP2 were verified in a panel of human melanoma cell lines and in a further reduced model that includes a pair of matched primary tumor-derived and metastasis-derived cell lines. Taken together, our data indicate a consistent correlation between RECK, MT1MMP, and TIMP2 across different models of clinical samples and cell lines and suggest evidence of the potential use of this subset of genes as a gene signature for diagnosing melanoma.
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Proteínas Ligadas a GPI/biosíntesis , Metaloproteinasa 14 de la Matriz/biosíntesis , Melanoma/genética , Melanoma/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/biosíntesis , Línea Celular Tumoral , Regulación hacia Abajo , Proteínas Ligadas a GPI/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Metaloproteinasa 14 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/biosíntesis , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/biosíntesis , Metaloproteinasa 9 de la Matriz/genética , Melanoma/enzimología , Fenotipo , Pronóstico , ARN Mensajero/genética , ARN Mensajero/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/biosíntesis , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/genética , Inhibidor Tisular de Metaloproteinasa-3/biosíntesis , Inhibidor Tisular de Metaloproteinasa-3/genéticaRESUMEN
DNA methylation is a fundamental epigenetic mechanism in regulating the expression of genes controlling crucial cell functions in cancer development. Gene silencing via CpG island methylation/demethylation in the promoter region is one of the mechanisms by which different genes are inactivated/activated in human cancers. Tissue inhibitor of metalloproteinase-2 (TIMP-2) is known to antagonize matrix metalloproteinase (MMP) activity and to suppress tumor growth, angiogenesis, invasion, and metastasis. TIMP-2 expression has been found to be both upregulated and downregulated in various cancers. The inconsistent TIMP-2 expression and unclear epigenetic regulation lead us to investigate its role in colorectal cancer in the presence of a methylating agent. Highly invasive human colorectal cells SW-620 were treated with the methyl donor S-adenosylmethionine (SAM) and its effect was evaluated by cell proliferation, cell cycle, invasion and migration assay. The ability of SAM to down regulate a panel of activated prometastatic, angiogenesis and growth- and cell cycle-regulatory genes was evaluated using end-point and real-time PCR. Treatment of SW-620 with SAM diminished cell proliferation and altered cell cycle kinetic G2/M phase cell cycle arrest. An in vitro matrigel invasion assay of SAM-treated cells showed a significant reduction in the invasive potential compared to untreated SW-620 cells. Treatment of SW-620 cells with SAM resulted in activation of TIMP-2 and inhibition of the expression of genes such as MMP (MMP-2, MT1-MMP), urokinase plasminogen activator, and vascular endothelial growth factors. The level of expression of tumor suppressor and apoptotic genes was not significantly higher compared to the untreated control. No changes in the levels of expression of genes (growth and cell cycle regulator), such as TGF-ß, Smad2, Smad4, and p21 were observed. Our data support the hypothesis that TIMP-2, along with other prometastatic genes, is hypomethylated and expressed differently in colorectal cancer. Further in-depth analysis is warranted to confirm the promoter region CpG methylation pattern of the TIMP-2 gene.
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Neoplasias Colorrectales/genética , S-Adenosilmetionina/farmacología , Inhibidor Tisular de Metaloproteinasa-2/genética , Apoptosis/efectos de los fármacos , Apoptosis/genética , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Neoplasias Colorrectales/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Inhibidor Tisular de Metaloproteinasa-2/metabolismoRESUMEN
BACKGROUND: Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases with an important role in physiological and pathological remodeling. Their activity is regulated by tissue inhibitors of matrix metalloproteinases (TIMPs). Excess MMPs and myeloperoxidase (MPO) activity have been associated with loss of tooth supporting tissues in periodontal disease (PD). We investigate the changes in salivary MMP-8, MMP-9, TIMP-1, TIMP-2, and MPO concentrations during PD treatment and compare results with plasma levels. METHODS: MMP-8, MMP-9, TIMP-1 and TIMP-2 were analyzed by ELISA. Gelatinolytic activity of MMP-9 forms was determined by zymography, and the MPO activity was determined by colorimetric assay. RESULTS: Subjects were divided into 2 groups: PD and control, which were further divided into 2 subgroups each, namely PD before (PB) and after 3 months (PA) of non-surgical periodontal therapy, and healthy volunteers at baseline (CB) and 3months after baseline (CA). Subgroup PA presented lower gelatinolytic activity and MMP-8 and TIMP-2 concentrations in the saliva compared with PB (p<0.05). The MPO activity was higher in PB compared with CB (p<0.05). There were significant correlations between the gelatinolytic activity of the saliva and MMP-8 and MMP-9 plasma levels. There was a significant correlation between plasma and saliva TIMP-2 levels. CONCLUSION: These results suggest attenuation of some inflammatory markers in the saliva and plasma after PD treatment. Moreover, correlations between salivary and plasma levels exist for some of these markers.