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BACKGROUND: Integrating aerobic exercise (AE) into rehabilitation programs for post-stroke individuals could enhance motor recovery and cardiovascular health by increasing brain-derived neurotrophic factor (BDNF) and the myokine irisin. Chronic stroke survivors typically exhibit elevated matrix metalloproteinase-9 (MMP-9) activity, which is negatively correlated with steps and time in medium cadence, although the impact of AE on this biomarker remains unclear. OBJECTIVE: To evaluate the effect of high-intensity AE training prior to modified constraint-induced movement therapy (mCIMT) on BDNF and irisin concentration, and on MMP-2 and MMP-9 activity in chronic post-stroke individuals and to associate these results with functional improvements. METHODS: Nine participants received AE combined with mCIMT for two weeks, while the control group (n = 7) received mCIMT alone. Manual dexterity and functional capacity were assessed before and after the intervention. Serum samples were analyzed for BDNF, irisin, MMP-2 and MMP-9. RESULTS: There were no significant main effects of assessment, group or interaction on molecular biomarkers. However, the AE group had a significant increase in MMP-9 activity post-intervention (p = .033; d = 0.67). For the Box and Block Test, there were significant main effects of assessment (F [1, 14] = 33.27, p = .000, ηp2 = 0.70) and group (F [1, 14] = 5.43, p = .035, ηp2 = .28). No correlations were found between biomarkers and clinical assessments. CONCLUSION: AE prior to mCIMT did not influence circulating BDNF and irisin levels but did induce an acute rise in MMP-9 activity, suggesting potential effects on cardiovascular remodeling in this population.
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BACKGROUND: Ameloblastoma is a locally destructive benign odontogenic tumor. While the neoplastic cells of conventional ameloblastoma can infiltrate the connective tissue and bone, in unicystic ameloblastoma the epithelium is encapsulated. The mechanisms driving ameloblastoma's bone resorption remains unclear. METHODS: RNA sequencing (RNA-seq) was performed in a discovery cohort of conventional ameloblastoma, and pathway enrichment analysis was carried out. mRNA levels of MMP13, a gene associated with bone resorption, were assessed using RT-qPCR in a larger cohort of conventional ameloblastoma and in unicystic ameloblastoma. Zymogram gels and the immunoexpression profile of collagenase 3 (encoded by MMP13 gene) were evaluated as well. RESULTS: Enriched pathways related to bone mineralization and upregulation of MMP13 were observed in ameloblastomas. Collagenolytic activity of collagenase 3 was detected in the tumor lysates. Collagenase 3 immunopositivity was observed in ameloblastomatous epithelium infiltrating the fibrous capsule of unicystic ameloblastoma. At the tumor-bone interface, collagenase 3 expression was detected in stromal cells, osteoblasts, and osteocytes. CONCLUSION: The results indicate a potential involvement of MMP13 in ameloblastoma-related bone resorption and progression.
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Ameloblastoma , Resorción Ósea , Metaloproteinasa 13 de la Matriz , Ameloblastoma/patología , Humanos , Metaloproteinasa 13 de la Matriz/metabolismo , Resorción Ósea/patología , Neoplasias Maxilomandibulares/patología , Neoplasias Maxilomandibulares/metabolismo , Masculino , Femenino , Adulto , Persona de Mediana Edad , ARN MensajeroRESUMEN
A Disintegrin and Metalloproteinase 10 (ADAM10) is a crucial transmembrane protein involved in diverse cellular processes, including cell adhesion, migration, and proteolysis. ADAM10's ability to cleave over 100 substrates underscores its significance in physiological and pathological contexts, particularly in Alzheimer's disease (AD). This review comprehensively examines ADAM10's multifaceted roles, highlighting its critical function in the non-amyloidogenic processing of the amyloid precursor protein (APP), which mitigates amyloid beta (Aß) production, a critical factor in AD development. We summarize the regulation of ADAM10 at multiple levels: transcriptional, translational, and post-translational, revealing the complexity and responsiveness of its expression to various cellular signals. A standardized nomenclature for ADAM10 isoforms is proposed to improve clarity and consistency in research, facilitating better comparison and replication of findings across studies. We address the challenges in detecting ADAM10 isoforms using antibodies, advocating for standardized detection protocols to resolve discrepancies in results from different biological matrices. By highlighting these issues, this review underscores the potential of ADAM10 as a biomarker for early diagnosis and a therapeutic target in AD. By consolidating current knowledge on ADAM10's regulation and function, we aim to provide insights that will guide future research and therapeutic strategies in the AD context.
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Proteína ADAM10 , Enfermedad de Alzheimer , Isoformas de Proteínas , Humanos , Enfermedad de Alzheimer/metabolismo , Proteína ADAM10/metabolismo , Animales , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Anticuerpos , Proteínas de la Membrana/metabolismo , Relevancia ClínicaRESUMEN
This systematic review evaluated the efficacy of doxycycline in MMP inhibition, its antibacterial action, and other properties relevant to dental materials testing. The study protocol was registered at the Open Science Framework ( https://doi.org/10.17605/OSF.IO/ZVK2T ). Reporting was based on PRISMA statement. The search was carried out in the databases: PubMed, Scopus, Web of Science, Embase, Lilacs, and Google Scholar. Articles were restricted to Portuguese, English, and Spanish, with no date limit. In vitro studies were selected based on the following outcomes: DOX antibacterial and anti-metalloproteinase activity and its influence in physico-chemical properties. Two researchers independently selected the articles and collected the data. Of 1507 documents, 82 were fully evaluated and 21 were included. Different forms of doxycycline incorporation were found, both as free form and incorporated into carrier agents. The drug was tested as primers, incorporated in adhesive or glass ionomer cement. No studies were found that evaluated its incorporation in resin composite or resin cement. The results confirmed the therapeutic properties of the medication, with more significant results when incorporated in an adhesive. However, although promising, the use of this substance requires standardization in application methods and adopted concentrations, allowing for more direct comparisons between studies. Furthermore, long-term studies are interesting to conduct, ensuring biocompatibility and complete understanding of long-term effects on dental materials.
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Central obesity is characterized by visceral adipose tissue (VAT) expansion, considered one of the main risk factors for metabolic complications. In recent years, new drugs have been studied for obesity treatment. Liraglutide (LGT), a GLP-1 agonist, decreases body weight, however, several mechanisms of action on VAT are still unknown. Aim: to study the effect of LGT on factors associated with VAT remodeling and mitochondrial dynamics in mice fed a high-fat diet (HFD). Methods: C57BL/6 mice were divided into Control (C) and HFD. After 15 weeks of feeding, each group was subdivided according to LGT administration for 5 weeks: C, C + LGT, HFD, and HFD + LGT. In epididymal AT (EAT) we evaluated histological and mitochondrial characteristics, vascularity, gelatinase activity (MMPs), and galectin-3 expression. Results: HFD presented larger adipocytes (p < 0.05), and lower vascular density and MMP-9 activity (p < 0.01) than C, while a major number of smaller adipocytes (p < 0.05) and an increase in vascularity (p < 0.001) and MMP-9 activity (p < 0.01) was observed in HFD + LGT. Collagen content was higher (p < 0.05) in EAT from HFD and decreased in HFD + LGT. In C, C + LGT, and HFD + LGT, mitochondria were predominantly tubular-shaped while in HFD mitochondria were mostly spherical (p < 0.001). Conclusion: LGT positively influences VAT behavior by modulating gelatinase activity, enhancing vascularization, and improving adipocyte histological characteristics. Additionally, LGT improves mitochondrial dynamics, a process that would favor VAT functionality.
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AIMS: Our aim was to evaluate whether the hydrogen sulfide (H2S) donor, 4-carboxyphenyl-isothiocyanate (4-CPI), exerts cardioprotective effect in the two kidney- one clip (2K-1C) rats through oxidative stress and MMP-2 activity attenuation and compare it with the classical H2S donor, Sodium Hydrosulfide (NaHS). MATERIALS AND METHODS: Renovascular hypertension (two kidneys-one clip; 2K-1C) was surgically induced in male Wistar rats. After two weeks, normotensive (2K) and hypertensive rats were intraperitoneally treated with vehicle (0.6 % dimethyl sulfoxide), NaHS (0.24 mg/Kg/day) or with 4-CPI (0.24 mg/Kg/day), for more 4 weeks. Systolic blood pressure (SBP) was evaluated weekly by tail-cuff plethysmography. Heart function was assessed by using the Millar catheter. Cardiac hypertrophy and fibrosis were evaluated by hematoxylin and eosin, and Picrosirius Red staining, respectively. The H2S was analyzed using WSP-1 fluorimetry and the cardiac oxidative stress was measured by lucigenin chemiluminescence and Amplex Red. MMP-2 activity was measured by in-gel gelatin or in situ zymography assays. Nox1, gp91phox, MMP-2 and the phospho-p65 subunit (Serine 279) nuclear factor kappa B (NF-κB) levels were evaluated by Western blotting. KEY FINDINGS: 4-CPI reduced blood pressure in hypertensive rats, decreased cardiac remodeling and promoted cardioprotection through the enhancement of cardiac H2S levels. An attenuation of oxidative stress, with inactivation of the p65-NF-κB/MMP-2 axis was similarly observed after NaHS or 4-CPI treatment in 2K-1C hypertension. SIGNIFICANCE: H2S is a mediator that promotes cardioprotective effects and decreases blood pressure, and 4-CPI seems to be a good candidate to reverse the maladaptive remodeling and cardiac dysfunction in renovascular hypertension.
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Presión Sanguínea , Sulfuro de Hidrógeno , Metaloproteinasa 2 de la Matriz , FN-kappa B , Estrés Oxidativo , Animales , Masculino , Ratas , Presión Sanguínea/efectos de los fármacos , Cardiotónicos/farmacología , Sulfuro de Hidrógeno/farmacología , Sulfuro de Hidrógeno/metabolismo , Hipertensión/tratamiento farmacológico , Hipertensión/metabolismo , Hipertensión Renovascular/tratamiento farmacológico , Hipertensión Renovascular/metabolismo , Hipertensión Renovascular/fisiopatología , Isotiocianatos/farmacología , Metaloproteinasa 2 de la Matriz/metabolismo , FN-kappa B/metabolismo , Estrés Oxidativo/efectos de los fármacos , Ratas Wistar , Sulfuros/farmacologíaRESUMEN
The hypothesis that physiological changes in women can affect periodontal tissues is the subject of this study, and inflammatory markers such as matrix metalloproteinase-8 can measure susceptibility to inflammation. The study aimed to analyze MMP-8 levels in periodontal sites of postpartum women and women without a history of pregnancy, comparing health parameters and periodontal disease. This is a case-control study with 40 participants, 20 cases (women in the postpartum period) and 20 controls (women without any pregnancy), who underwent clinical periodontal examination and the collection of crevicular gingival fluid. The ELISA test was used to detect MMP-8 levels. Postpartum women had worse periodontal parameters, such as bleeding index on probing, number of sites with CAL ≥ 3, and fewer teeth present. In the group of women without a history of pregnancy, a significantly lower MMP-8 level was observed in healthy sites and a higher one was observed in periodontal pockets (p < 0.01). In contrast, in postpartum women, MMP-8 levels were elevated in both healthy sites and periodontal pockets (p > 0.01). The MMP-8 levels in gingival fluid appear to be related to periodontal clinical parameters and may be a possible marker of enzymatic changes involved in periodontal tissue destruction in postpartum women.
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Líquido del Surco Gingival , Metaloproteinasa 8 de la Matriz , Periodo Posparto , Humanos , Femenino , Metaloproteinasa 8 de la Matriz/metabolismo , Metaloproteinasa 8 de la Matriz/análisis , Adulto , Estudios de Casos y Controles , Líquido del Surco Gingival/enzimología , Embarazo , Enfermedades Periodontales/enzimología , Biomarcadores/metabolismo , Adulto JovenRESUMEN
Surgeries that require general anesthesia occur in 1.5-2% of gestations. Isoflurane is frequently used because of its lower possibility of affecting fetal growth. Therefore, we examined the isoflurane anesthesia-induced effects on maternal hemodynamic and vascular changes. We hypothesized that isoflurane would enhance endothelium-dependent vasodilation as a consequence of increased nitric oxide and decreased metalloproteinases (MMPs). Female rats (n=28) were randomized into 4 groups (7 rats/group): conscious (non-anesthetized) non-pregnant group, non-pregnant anesthetized group, conscious pregnant group, and pregnant anesthetized group. Anesthesia was performed on the 20th pregnancy day, and hemodynamic parameters were monitored. Nitric oxide metabolites, gelatinolytic activity of MMP-2 and MMP-9, and the vascular function were assessed. Isoflurane caused no significant hemodynamic changes in pregnant compared with non-pregnant anesthetized group. Impaired acetylcholine-induced relaxations were observed only in conscious non-pregnant group (by approximately 62%) versus 81% for other groups. Phenylephrine-induced contractions were greater in endothelium-removed aorta segments of both pregnant groups (with or without isoflurane) compared with non-pregnant groups. Higher nitric oxide metabolites were observed in anesthetized pregnant in comparison with the other groups. Reductions in the 75 kDa activity and concomitant increases in 64 kDa MMP-2 isoforms were observed in aortas of pregnant anesthetized (or not) groups compared with conscious non-pregnant group. Isoflurane anesthesia shows stable effects on hemodynamic parameters and normal MMP-2 activation in pregnancy. Furthermore, there were increases in nitric oxide bioavailability, suggesting that isoflurane provides protective actions to the endothelium in pregnancy.
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Isoflurano , Metaloproteinasa 2 de la Matriz , Óxido Nítrico , Vasodilatación , Animales , Femenino , Embarazo , Ratas , Anestésicos por Inhalación/farmacología , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/metabolismo , Hemodinámica/efectos de los fármacos , Isoflurano/farmacología , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Óxido Nítrico/metabolismo , Vasodilatación/efectos de los fármacos , Ratas WistarRESUMEN
Mammary tumors are the most frequent type of neoplasms in intact female dogs. New therapies that target neoplastic cells without affecting normal cells are highly sought. The Bacillus anthracis toxin has been reengineered to target tumor cells that express urokinase plasminogen activators and metalloproteinases. In previous studies carried out in our laboratory, the reengineered anthrax toxin had inhibitory effects on canine oral mucosal melanoma and canine osteosarcoma cells. In this study, five canine neoplastic epithelial cell lines (four adenocarcinomas and one adenoma) and one non-neoplastic canine mammary epithelial cell line were treated with different concentrations of reengineered anthrax toxin components. Cell viability was quantified using an MTT assay and half-maximal inhibitory concentration (IC50) values. Cell lines were considered sensitive when the IC50 was lower than 5000 ng/ml. One canine mammary adenocarcinoma cell line and one mammary adenoma cell line showed significantly decreased viability after treatment, whereas the non-neoplastic cell line was resistant. We conclude that the reengineered anthrax toxin may be considered a targeted therapy for canine mammary neoplasms while preserving normal canine mammary epithelial cells.
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Antígenos Bacterianos , Toxinas Bacterianas , Enfermedades de los Perros , Neoplasias Mamarias Animales , Animales , Perros , Neoplasias Mamarias Animales/tratamiento farmacológico , Toxinas Bacterianas/farmacología , Femenino , Antígenos Bacterianos/farmacología , Línea Celular Tumoral , Enfermedades de los Perros/tratamiento farmacológico , Células Epiteliales/efectos de los fármacos , Adenocarcinoma/veterinaria , Adenocarcinoma/tratamiento farmacológico , Adenocarcinoma/patología , Supervivencia Celular/efectos de los fármacos , Adenoma/veterinaria , Adenoma/tratamiento farmacológico , Adenoma/patologíaRESUMEN
Snake venom metalloproteases (SVMPs) are hydrolytic enzymes dependent on metal binding, primarily zinc (Zn2+), at their catalytic site. They are classified into three classes (P-I to P-III). BjussuMP-II, a P-I SVMP isolated from Bothrops jararacussu snake venom, has a molecular mass of 24 kDa. It exhibits inhibitory activity on platelet aggregation and hydrolyzes fibrinogen. TNF-α upregulates the expression of adhesion molecules on endothelial cell surfaces, promoting leukocyte adhesion and migration during inflammation. Literature indicates that SVMPs may cleave the TNF-α precursor, possibly due to significant homology between metalloproteases from mammalian extracellular matrix and SVMPs. This study aimed to investigate BjussuMP-II's effects on human umbilical vein endothelial cells (HUVEC), focusing on viability, detachment, adhesion, release, and cleavage of TNF-α, IL-1ß, IL-6, IL-8, and IL-10. HUVEC were incubated with BjussuMP-II (1.5-50 µg/mL) for 3-24 h. Viability was determined using LDH release, MTT metabolization, and 7AAD for membrane integrity. Adhesion and detachment were assessed by incubating cells with BjussuMP-II and staining with Giemsa. Cytokines were quantified in HUVEC supernatants using EIA. TNF-α cleavage was evaluated using supernatants from PMA-stimulated cells or recombinant TNF-α. Results demonstrated BjussuMP-II's proteolytic activity on casein. It was not toxic to HUVEC at any concentration or duration studied but interfered with adhesion and promoted detachment. PMA induced TNF-α release by HUVEC, but this effect was not observed with BjussuMP-II, which cleaved TNF-α. Additionally, BjussuMP-II cleaved IL-1ß, IL-6, and IL-10. These findings suggest that the zinc metalloprotease BjussuMP-II could be a valuable biotechnological tool for treating inflammatory disorders involving cytokine deregulation.
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Adhesión Celular , Citocinas , Células Endoteliales de la Vena Umbilical Humana , Metaloproteasas , Humanos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Citocinas/metabolismo , Metaloproteasas/metabolismo , Adhesión Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Bothrops/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Venenos de Crotálidos/metabolismo , Venenos de Crotálidos/toxicidad , Proteolisis/efectos de los fármacosRESUMEN
PURPOSE: Resistance training (RT) induces muscle growth at varying rates across RT phases, and evidence suggests that the muscle-molecular responses to training bouts become refined or attenuated in the trained state. This study examined how proteolysis-related biomarkers and extracellular matrix (ECM) remodeling factors respond to a bout of RT in the untrained (UT) and trained (T) state. METHODS: Participants (19 women and 19 men) underwent 10 weeks of RT. Biopsies of vastus lateralis were collected before and after (24 h) the first (UT) and last (T) sessions. Vastus lateralis cross-sectional area (CSA) was assessed before and after the experimental period. RESULTS: There were increases in muscle and type II fiber CSAs. In both the UT and T states, calpain activity was upregulated and calpain-1/-2 protein expression was downregulated from Pre to 24 h. Calpain-2 was higher in the T state. Proteasome activity and 20S proteasome protein expression were upregulated from Pre to 24 h in both the UT and T. However, proteasome activity levels were lower in the T state. The expression of poly-ubiquitinated proteins was unchanged. MMP activity was downregulated, and MMP-9 protein expression was elevated from Pre to 24 h in UT and T. Although MMP-14 protein expression was acutely unchanged, this marker was lower in T state. TIMP-1 protein levels were reduced Pre to 24 h in UT and T, while TIMP-2 protein levels were unchanged. CONCLUSION: Our results are the first to show that RT does not attenuate the acute-induced response of proteolysis and ECM remodeling-related biomarkers.
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Biomarcadores , Matriz Extracelular , Proteolisis , Entrenamiento de Fuerza , Humanos , Masculino , Femenino , Entrenamiento de Fuerza/métodos , Matriz Extracelular/metabolismo , Biomarcadores/metabolismo , Adulto , Calpaína/metabolismo , Músculo Esquelético/metabolismo , Adulto Joven , Complejo de la Endopetidasa Proteasomal/metabolismoRESUMEN
Angiogenesis is a process that is controlled by a delicate combination of proangiogenic and antiangiogenic molecules and can be disrupted in various illnesses, including cancer. Non-cancerous diseases can also have an abnormal or insufficient vascular growth, inflammation and hypoxia, which exacerbate angiogenesis. These conditions include atherosclerosis, psoriasis, endometriosis, asthma, obesity and AIDS. Based on that, the present work assessed the in vitro and ex vivo antiangiogenic properties stemming from BthMP, a P-I metalloproteinase from Bothrops moojeni snake venom, via the VEGF pathway. BthMP at a concentration of 5 and 40 µg/mL showed no toxicity to endothelial cells (HUVEC) in the MTT assay and was not able to induce necrosis and colony proliferation. Interestingly, BthMP inhibited adhesion, migration and invasion of HUVECs in Matrigel and arrested in vitro angiogenesis by reducing the average number of nodules in toxin-treated cells by 9.6 and 17.32 at 5 and 40 µg/mL, respectively, and the number of tubules by 15.9 at 5 µg/mL and 21.6 at 40 µg/mL in a VEGF-dependent way, an essential proangiogenic property. Furthermore, BthMP inhibited the occurrence of the angiogenic process in an ex vivo aortic ring test by decreasing new vessel formation by 52% at 5 µg/mL and by 66% at 40 µg/mL and by increasing the expression of an antiangiogenic gene, SFLT-1, and decreasing the expression of the proangiogenic genes VEGFA and ANGPT-1. Finally, this toxin reduces the production of nitric oxide, a marker that promotes angiogenesis and VEGF modulation, and decreases the protein expression of VEGFA in the supernatant of the HUVEC culture by about 30 %. These results suggest that BthMP has a promising antiangiogenic property and proves to be a biotechnological mechanism for understanding the antiangiogenic responses induced by snake venom metalloproteinases, which could be applied to a variety of diseases that exhibit an imbalance of angiogenesis mechanisms.
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Bothrops , Células Endoteliales , Serpientes Venenosas , Animales , Femenino , Humanos , Células Endoteliales/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Bothrops/metabolismo , Metaloproteasas/metabolismo , Venenos de Serpiente , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Inhibidores de la Angiogénesis/farmacologíaRESUMEN
We evaluated the effect of the total macerate (TM) and seed oil (SO) of mature Carica candamarcensis fruits, on the release of Matrix metalloproteinase 9 (MMP9) and the phosphorylation of MAPK in neutrophils. The antioxidant capacity of these extracts was evaluated by ABTS assay. Neutrophils stimulated with different dilutions of TM or SO were analyzed for cytotoxicity, MMP9 release, and MAPK phosphorylation, using trypan blue exclusion assays, zymography, and immunoblotting, respectively. Both extracts show antioxidant activity, being higher in TM; none presented cytotoxic effect. The 5% and 2.5% dilutions of TM significantly reduced MMP9 release, and all decreased MAPK phosphorylation. SO significantly increased the release o f MMP9 and MAPK phosphorylation, the effect being greater when they were prestimulated with lipopolysaccharide.TM may have anti - inflammatory potential, while SO could have a priming effect that needs to be confirmed
Evaluamos el efecto del macerado total (MT) y aceite de semillas (AV) de frutos maduros de Carica candamarcensis , en la liberación de Matriz metaloproteinasa 9 (MMP9) y la fosfor ilación de MAPK en neutrófilos. La capacidad antioxidante de estos extractos se evaluó por ensayo ABTS. En neutrófilos estimulados con diferentes diluciones de MT o AV se analizó la citotoxicidad, liberación de MMP9 y fosforilación de MAPK, mediante ensayo s de exclusión con azul de tripano, zimografía e inmunotransferencia, respectivamente. Ambos extractos muestran actividad antioxidante, siendo mayor en MT; ninguno presentó efecto citotóxico. Las diluciones 5% y 2,5% de MT redujeron significativamente la l iberación de MMP9, y todas disminuyeron la fosforilación de MAPK. El AV incrementó significativamente la liberación de MMP9 y la fosforilación de MAPK, el efecto fue mayor cuando se preestimularon con lipopolisacárido. El MT puede tener potencial antiinfla matorio, mientras que el AV podría tener un efecto "priming" que necesita ser corroborado.
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Humanos , Extractos Vegetales/farmacología , Carica/enzimología , Neutrófilos/efectos de los fármacos , Fosforilación , Proteína Quinasa 1 Activada por Mitógenos , Metaloproteinasa 9 de la Matriz/metabolismo , Látex/análisisRESUMEN
In this study, a methodology is proposed, combining ligand- and structure-based virtual screening tools, for the identification of phosphorus-containing compounds as inhibitors of zinc metalloproteases. First, we use Dragon molecular descriptors to develop a Linear Discriminant Analysis classification model, which is widely validated according to the OECD principles. This model is simple, robust, stable and has good discriminating power. Furthermore, it has a defined applicability domain and it is used for virtual screening of the DrugBank database. Second, docking experiments are carried out on the identified compounds that showed good binding energies to the enzyme thermolysin. Considering the potential toxicity of phosphorus-containing compounds, their toxicological profile is evaluated according to Protox II. Of the five molecules evaluated, two show carcinogenic and mutagenic potential at small LD50, not recommended as drugs, while three of them are classified as non-toxic, and could constitute a starting point for the development of new vasoactive metalloprotease inhibitor drugs. According to molecular dynamics simulation, two of them show stable interactions with the active site maintaining coordination with the metal. A high agreement is evident between QSAR, docking and molecular dynamics results, demonstrating the potentialities of the combination of these tools.
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Simulación de Dinámica Molecular , Relación Estructura-Actividad Cuantitativa , Simulación del Acoplamiento Molecular , Ligandos , Metaloproteasas , FósforoRESUMEN
OBJECTIVE: The present study was designed to investigate the role of macrophage migration inhibitory factor (MIF) in the exacerbation of pregestational periodontal disease (PGPD). BACKGROUND: Periodontitis (PT) is a severe stage of periodontal disease characterized by inflammation of the supporting tissues of the teeth, which usually worsens during pregnancy. MIF is a proinflammatory cytokine that is significantly elevated in periodontitis, both at the beginning and at the end of pregnancy. Although periodontitis usually presents with greater severity during pregnancy, the participation of MIF in the evolution of periodontitis has not been established. METHODS: To analyze the relevance of MIF in the exacerbation of PGPD, we employed a model of PGPD in WT and Mif-/- mice, both with a BALB/c genetic background. PT was induced with nylon suture ligatures placed supramarginally around the second upper right molar. For PGPD, PT was induced 2 weeks before mating. We evaluated histological changes and performed histometric analysis of the clinical attachment loss, relative expression of MMP-2 and MMP-13 by immunofluorescence, and relative expression of the cytokines mif, tnf-α, ifn-γ, and il-17 by quantitative real-time polymerase chain reaction (qRT-PCR). RESULTS: Our data revealed that periodontal tissue from PGPD WT mice produced a twofold increase in MIF compared with PT WT mice. Moreover, the evolution of periodontitis in Mif-/- mice was less severe than in PGDP WT mice. Periodontal tissue from Mif-/- mice with PGPD produced 80% less TNF-α and no IFN-γ, as well as 50% lower expression of matrix metalloproteinase (MMP)-2 and 25% less MMP-13 compared to WT PGDP mice. CONCLUSIONS: Our study suggests that MIF plays an important role in the exacerbation of periodontitis during pregnancy and that MIF is partially responsible for the inflammation associated with the severity of periodontitis during pregnancy.
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Factores Inhibidores de la Migración de Macrófagos , Periodontitis , Animales , Femenino , Ratones , Embarazo , Inflamación/metabolismo , Factores Inhibidores de la Migración de Macrófagos/metabolismo , Metaloproteinasa 13 de la Matriz , Periodontitis/metabolismo , Factor de Necrosis Tumoral alfaRESUMEN
Abstract Objectives This study sought to determine effects of Thai propolis extract mixed in mineral trioxide aggregate (MTA) on matrix metalloproteinase-2 (MMP-2) expression and its activity in inflamed human dental pulp cells (HDPCs). Materials and Methods Interleukin-1β-primed HDPCs were treated with either the eluate of MTA mixed with distilled water, of MTA mixed with 0.75 mg/ml of the propolis extract, or of Dycal®, 0.75 mg/ml of the propolis extract, or 0.2% (v/v) of chlorhexidine for 24 or 72 h. The viability of HDPCs was determined by the PrestoBlue® cytotoxic assay. HDPCs' lysates were analyzed for MMP-2 mRNA expression by RT-qPCR, while their supernatants were measured for MMP-2 activity by gelatin zymography. Results At 24 and 72 h, a non-toxic dose of the propolis extract at 0.75 mg/ml by itself or mixed in MTA tended to reduce MMP-2 expression upregulated by MTA, while it further decreased the MMP-2 activity as compared to that of MTA mixed with distilled water. The MMP-2 activity of interleukin-1β-primed HDPCs treated with the eluate of the propolis extract mixed in MTA was significantly lower than that of interleukin-1β-primed HDPCs at 24 h (p=0.012). As a control, treatment with chlorhexidine significantly inhibited MMP-2 expression induced by MTA and MMP-2 activity enhanced by interleukin-1β (p<0.05). Treatment with Dycal® caused a significant increase in HDPC's death, resulting in a significant decrease in MMP-2 expression and activity (p<0.05). Conclusions MTA mixed with Thai propolis extract can reduce MMP-2 mRNA expression and activity when compared to MTA mixed with distilled water in inflamed HDPCs.
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Angiogenesis is a process that is controlled by a delicate combination of proangiogenic and antiangiogenic molecules and can be disrupted in various illnesses, including cancer. Non-cancerous diseases can also have an abnormal or insufficient vascular growth, inflammation and hypoxia, which exacerbate angiogenesis. These conditions include atherosclerosis, psoriasis, endometriosis, asthma, obesity and AIDS. Based on that, the present work assessed the in vitro and ex vivo antiangiogenic properties stemming from BthMP, a P–I metalloproteinase from Bothrops moojeni snake venom, via the VEGF pathway. BthMP at a concentration of 5 and 40 μg/mL showed no toxicity to endothelial cells (HUVEC) in the MTT assay and was not able to induce necrosis and colony proliferation. Interestingly, BthMP inhibited adhesion, migration and invasion of HUVECs in Matrigel and arrested in vitro angiogenesis by reducing the average number of nodules in toxin-treated cells by 9.6 and 17.32 at 5 and 40 μg/mL, respectively, and the number of tubules by 15.9 at 5 μg/mL and 21.6 at 40 μg/mL in a VEGF-dependent way, an essential proangiogenic property. Furthermore, BthMP inhibited the occurrence of the angiogenic process in an ex vivo aortic ring test by decreasing new vessel formation by 52% at 5 μg/mL and by 66% at 40 μg/mL and by increasing the expression of an antiangiogenic gene, SFLT-1, and decreasing the expression of the proangiogenic genes VEGFA and ANGPT-1. Finally, this toxin reduces the production of nitric oxide, a marker that promotes angiogenesis and VEGF modulation, and decreases the protein expression of VEGFA in the supernatant of the HUVEC culture by about 30 %. These results suggest that BthMP has a promising antiangiogenic property and proves to be a biotechnological mechanism for understanding the antiangiogenic responses induced by snake venom metalloproteinases, which could be applied to a variety of diseases that exhibit an imbalance of angiogenesis mechanisms.
RESUMEN
Snakebite envenomation is classified as a Neglected Tropical Disease. Bothrops jararaca venom induces kidney injury and coagulopathy. HF3, a hemorrhagic metalloproteinase of B. jararaca venom, participates in the envenomation pathogenesis. We evaluated the effects of HF3 in mouse kidney and blood plasma after injection in the thigh muscle, mimicking a snakebite. Transcriptomic analysis showed differential expression of 31 and 137 genes related to kidney pathology after 2 h and 6 h, respectively. However, only subtle changes were observed in kidney proteome, with differential abundance of 15 proteins after 6 h, including kidney injury markers. N-terminomic analysis of kidney proteins showed 420 proteinase-generated peptides compatible with meprin specificity, indicating activation of host proteinases. Plasma analysis revealed differential abundance of 90 and 219 proteins, respectively, after 2 h and 6 h, including coagulation-cascade and complement-system components, and creatine-kinase, whereas a semi-specific search of N-terminal peptides indicated activation of endogenous proteinases. HF3 promoted host reactions, altering the gene expression and the proteolytic profile of kidney tissue, and inducing plasma proteome imbalance driven by changes in abundance and proteolysis. The overall response of the mouse underscores the systemic action of a hemorrhagic toxin that transcends local tissue damage and is related to known venom-induced systemic effects.
Asunto(s)
Bothrops , Venenos de Crotálidos , Ratones , Animales , Proteoma , Multiómica , Metaloproteasas/metabolismo , Venenos de Serpiente/toxicidad , Péptidos , Plasma/metabolismo , Riñón/metabolismo , Bothrops/metabolismo , Venenos de Crotálidos/toxicidad , Venenos de Crotálidos/metabolismoRESUMEN
INTRODUCTION: Increased matrix metalloproteinase (MMP)-2 activity contributes to increase vascular smooth muscle cell (VSMC) proliferation in the aorta in early hypertension by cleaving many proteins of the extracellular matrix. Cleaved products from type I collagen may activate focal adhesion kinases (FAK) that trigger migration and proliferation signals in VSMC. We therefore hypothesized that increased activity of MMP-2 proteolyzes type I collagen in aortas of hypertensive rats, and thereby, induces FAK activation, thus leading to increased VSMC proliferation and hypertrophic remodeling in early hypertension. METHODS: Male Sprague-Dawley rats were submitted to renovascular hypertension by the two kidney-one clip (2K1C) model and treated with doxycycline (30 mg/kg/day) by gavage from the third to seventh-day post-surgery. Controls were submitted to sham surgery. Systolic blood pressure (SBP) was measured daily by tail-cuff plethysmography and the aortas were processed for zymography and Western blot for MMP-2, pFAK/FAK, integrins and type I collagen. Mass spectrometry, morphological analysis and Ki67 immunofluorescence were also done to identify collagen changes and VSMC proliferation. A7r5 cells were stimulated with collagen and treated with the MMP inhibitors (doxycycline or ARP-100), and with the FAK inhibitor PND1186 for 24 h. Cells were lysed and evaluated by Western blot for pFAK/FAK. RESULTS: 2K1C rats developed elevated SBP in the first week as well as increased expression and activity of MMP-2 in the aorta (p < 0.05 vs. Sham). Treatment with doxycycline reduced both MMP activity and type I collagen proteolysis in aortas of 2K1C rats (p < 0.05). Increased pFAK/FAK and increased VSMC proliferation (p < 0.05 vs. Sham groups) were also seen in the aortas of 2K1C and doxycycline decreased both parameters (p < 0.05). Higher proliferation of VSMC contributed to hypertrophic remodeling as seen by increased media/lumen ratio and cross sectional area (p < 0.05 vs Sham groups). In cell culture, MMP-2 cleaves collagen, an effect reversed by MMP inhibitors (p < 0.05). Increased levels of pFAK/FAK were observed when collagen was added in the culture medium (p < 0.05 vs control) and MMP and FAK inhibitors reduced this effect. CONCLUSIONS: Increase in MMP-2 activity proteolyzes type I collagen in the aortas of 2K1C rats and contributes to activate FAK and induces VSMC proliferation during the initial phase of hypertension.