RESUMEN
Understanding the mechanisms of modulators' action on enzymes is crucial for optimizing and designing pharmaceutical substances. The acute inflammatory response, in particular, is regulated mainly by a disintegrin and metalloproteinase (ADAM) 17. ADAM17 processes several disease mediators such as TNFα and APP, releasing their soluble ectodomains (shedding). A malfunction of this process leads to a disturbed inflammatory response. Chemical protease inhibitors such as TAPI-1 were used in the past to inhibit ADAM17 proteolytic activity. However, due to ADAM17's broad expression and activity profile, the development of active-site-directed ADAM17 inhibitor was discontinued. New 'exosite' (secondary substrate binding site) inhibitors with substrate selectivity raised the hope of a substrate-selective modulation as a promising approach for inflammatory disease therapy. This work aimed to develop a high-throughput screen for potential ADAM17 modulators as therapeutic drugs. By combining experimental and in silico methods (structural modeling and docking), we modeled the kinetics of ADAM17 inhibitor. The results explain ADAM17 inhibition mechanisms and give a methodology for studying selective inhibition towards the design of pharmaceutical substances with higher selectivity.
Asunto(s)
Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/efectos de los fármacos , Proteína ADAM17/metabolismo , Proteínas ADAM/metabolismo , Sitios de Unión/efectos de los fármacos , Dominio Catalítico/efectos de los fármacos , Simulación por Computador , Evaluación Preclínica de Medicamentos/métodos , Células HEK293 , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Cinética , Inhibidores de Proteasas/farmacología , Especificidad por Sustrato/efectos de los fármacosRESUMEN
Novel therapeutic strategies are needed to attenuate increased systemic and gut inflammation that contribute to morbidity and mortality in chronic HIV infection despite potent antiretroviral therapy (ART). The goal of this study is to use preclinical models of chronic treated HIV to determine whether the antioxidant and anti-inflammatory apoA-I mimetic peptides 6F and 4F attenuate systemic and gut inflammation in chronic HIV. We used two humanized murine models of HIV infection and gut explants from 10 uninfected and 10 HIV infected persons on potent ART, to determine the in vivo and ex vivo impact of apoA-I mimetics on systemic and intestinal inflammation in HIV. When compared to HIV infected humanized mice treated with ART alone, mice on oral apoA-I mimetic peptide 6F with ART had consistently reduced plasma and gut tissue cytokines (TNF-α, IL-6) and chemokines (CX3CL1) that are products of ADAM17 sheddase activity. Oral 6F attenuated gut protein levels of ADAM17 that were increased in HIV-1 infected mice on potent ART compared to uninfected mice. Adding oxidized lipoproteins and endotoxin (LPS) ex vivo to gut explants from HIV infected persons increased levels of ADAM17 in myeloid and intestinal cells, which increased TNF-α and CX3CL1. Both 4F and 6F attenuated these changes. Our preclinical data suggest that apoA-I mimetic peptides provide a novel therapeutic strategy that can target increased protein levels of ADAM17 and its sheddase activity that contribute to intestinal and systemic inflammation in treated HIV. The large repertoire of inflammatory mediators involved in ADAM17 sheddase activity places it as a pivotal orchestrator of several inflammatory pathways associated with morbidity in chronic treated HIV that make it an attractive therapeutic target.
Asunto(s)
Apolipoproteína A-I , Infecciones por VIH/patología , Inflamación/patología , Intestinos/efectos de los fármacos , Péptidos/farmacología , Proteína ADAM17/efectos de los fármacos , Animales , Fármacos Anti-VIH/farmacología , Humanos , RatonesRESUMEN
Astragaloside III (AS-III) is a triterpenoid saponin contained in Astragali Radix and has potent anti-inflammatory effects on vascular endothelial cells; however, underlying mechanisms are unclear. In this study, we provided the first piece of evidence that AS-III induced phosphorylation of TNF-α converting enzyme (TACE) at Thr735 and enhanced its sheddase activity. As a result, AS-III reduced surface TNFR1 level and increased content of sTNFR1 in the culture media, leading to the inhibition of NF-κB signaling pathway and attenuation of downstream cytokine gene expression. Furthermore, AS-III induced TACE-dependent epidermal growth factor receptor (EGFR) transactivation and activation of downstream ERK1/2 and AKT pathways. Finally, AS-III induced activation of p38. Both TACE activation and EGFR transactivation induced by AS-III were significantly inhibited by p38 inhibitor SB203580. Taken together, we concluded that AS-III activates TACE-dependent anti-inflammatory and growth factor signaling in vascular endothelial cells in a p38-dependent fashion, which may contribute to its cardiovascular protective effect.
Asunto(s)
Proteína ADAM17/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Saponinas/uso terapéutico , Animales , Humanos , Ratones , Saponinas/farmacología , Transducción de Señal/efectos de los fármacosRESUMEN
BACKGROUND: In myocardial infarction (MI), inflammatory cytokine including tumor necrosis factor-α (TNF-α) plays pivotal role throughout worsening processes and recovery, whereas its cleavage is controlled by a disintegrin and metalloproteinases-17 (ADAM-17). Syzygium polyanthum (Wight) is widely used plant in Indonesia and Southeast Asian countries to treat various illnesses. OBJECTIVE: This study aimed to analyze the effect of S. polyanthum extract towards TNF-α and ADAM17 expressions in MI rat model. METHODS: Myocardial infarction were surgically induced in wistar rats by ligating left anterior descending coronary artery in both treatment and control group. Around 3.6 mg of S. polyanthum extract (SPE) was administered via nasogastric tube to treatment groups, while control group received only saline. Serum TNF-α level and expression of TNF-α and ADAM17 in blanching heart muscle was examined in both groups on day 1, day 4, day 7, and day 14 after treatment with SPE. RESULTS: Reduction of serum TNF-α levels was markedly seen from day 4 in treatment group and was lower than in control group since day 4. Expression of ADAM17 was reduced and expression of TNF-α was only seen in myocardial membrane in group with SPE treatment. CONCLUSION: Syzygium polyanthum exerts its anti-inflammatory effect by decreasing ADAM17 expression subsequently lowering TNF-α regulation in MI rat model.
Asunto(s)
Proteína ADAM17/efectos de los fármacos , Proteína ADAM17/metabolismo , Antiinflamatorios/uso terapéutico , Infarto del Miocardio/fisiopatología , Extractos Vegetales/uso terapéutico , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Humanos , Indonesia , Masculino , Modelos Animales , Fitoterapia , Plantas Medicinales/química , Ratas , Ratas Wistar , Syzygium/químicaRESUMEN
Vulnerable plaques in advanced atherosclerosis have defective efferocytosis. The role of ANG II in the progression of atherosclerosis is not fully understood. Herein, we investigated the effects and the underlying mechanisms of ANG II on macrophage efferocytosis in advanced atherosclerosis. ANG II decreased the surface expression of Mer tyrosine kinase (MerTK) in macrophages through a disintegrin and metalloproteinase17 (ADAM17)-mediated shedding of the soluble form of MerTK (sMer) in the medium, which led to efferocytosis suppression. ANG II-activated ADAM17 required reactive oxygen species (ROS) and p38 MAPK phosphorylation. Selective angiotensin II type 1 receptor (AT1R) blocker losartan suppressed ROS production, and ROS scavenger N-acetyl-l-cysteine (NAC) prevented p38 MAPK phosphorylation. In addition, mutant MERTKΔ483-488 was resistant to ANG II-induced MerTK shedding and efferocytosis suppression. The advanced atherosclerosis model that is characterized by larger necrotic cores, and less collagen content was established by feeding apolipoprotein E knockout (ApoE-/-) mice with a high-fat diet for 16 wk. NAC and losartan oral administration prevented atherosclerotic lesion progression. Meanwhile, the inefficient efferocytosis represented by decreased macrophage-associated apoptotic cells and decreased MerTK+CD68+double-positive macrophages in advanced atherosclerosis were prevented by losartan and NAC. Additionally, the serum levels of sMer were increased and positively correlated with the upregulated levels of ANG II in acute coronary syndrome (ACS) patients. In conclusion, ANG II promotes MerTK shedding via AT1R/ROS/p38 MAPK/ADAM17 pathway in macrophages, which led to defective efferocytosis and atherosclerosis progression. Defining the molecular mechanisms of defective efferocytosis may provide a promising prognosis and therapy for ACS patients.
Asunto(s)
Proteína ADAM17/efectos de los fármacos , Angiotensina II/farmacología , Aterosclerosis/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Tirosina Quinasa c-Mer/efectos de los fármacos , Animales , Aterosclerosis/tratamiento farmacológico , Humanos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones Transgénicos , Fagocitosis/efectos de los fármacos , Proteínas Tirosina Quinasas/efectos de los fármacos , Proteínas Tirosina Quinasas/metabolismo , Proteínas Tirosina Quinasas Receptoras/efectos de los fármacos , Proteínas Tirosina Quinasas Receptoras/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Chlorpyrifos (CPF) is an organophosphorus pesticide widely used in agricultural applications and household environments. 6-Gingerol-rich fraction from Zingiber officinale (Ginger, 6-GRF) has been reported to possess potent anti-oxidative, anti-inflammatory and anti-apoptotic properties. Here, we investigated the protective properties of 6-GRF on CPF-induced oxidative damage and inflammation in the brain, ovary and uterus of rats. Five groups of rats containing 14 rats/group received corn oil (control), CPF (5 mg/kg), 6-GRF (100 mg/kg), CPF (5 mg/kg) + 6-GRF (50 mg/kg) and CPF (5 mg/kg) + 6-GRF (100 mg/kg) through gavage once per day for 35 days respectively. The results showed that 6-GRF protected against CPF-induced increases in oxidative stress ((hydrogen peroxide (H2O2) and malondialdehyde (MDA)), inflammatory (myeloperoxidase (MPO), nitric oxide (NO) and tumour necrosis factor-α (TNF- α)), and apoptotic (caspase-3) markers. Also, 6-GRF improved the activities of antioxidant enzymes catalase, superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione S-transferase (GST) as well as glutathione (GSH) level in the brain, ovary and uterus of rats exposed to CPF (p < 0.05). Overall, the protective effects of 6-GRF on CPF-induced toxicity in the brain and reproductive organs of rats may be due to its potent antioxidative, anti-inflammatory and antiapoptotic properties.
Asunto(s)
Encéfalo/efectos de los fármacos , Catecoles/uso terapéutico , Alcoholes Grasos/uso terapéutico , Ovario/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Extractos Vegetales/farmacología , Útero/efectos de los fármacos , Zingiber officinale/química , Proteína ADAM17/efectos de los fármacos , Animales , Peso Corporal/efectos de los fármacos , Encéfalo/patología , Cloropirifos/toxicidad , Femenino , Inflamación/tratamiento farmacológico , Ovario/patología , Extractos Vegetales/uso terapéutico , Ratas , Ratas Wistar , Útero/patologíaRESUMEN
OBJECTIVE: Intraluminal pancreatic trypsin and other digestive enzymes injure the gut barrier following trauma-hemorrhagic shock (T/HS). Intestinal proteases (sheddases) exert important effects on normal gut function but may cause barrier disruption due to exaggerated production following T/HS. We hypothesized that the protective mechanism of TXA on the gut barrier following T/HS includes inhibition of these "downstream" proteases. This was studied in vitro. METHODS: Trypsin, matrix metalloproteinase (MMP-9) and ADAM-17 activity were measured in intestinal epithelial cells (IEC) exposed to HR + trypsin. TXA was added to IEC subsets. Pulmonary microvascular endothelial cells (HMVEC) were exposed to IEC supernatants and syndecan release and ICAM-1 expression determined. RESULTS: Trypsin activity and the activity of the "downstream" sheddases ADAM-17, MMP was increased in IEC lysates following exposure to HR + trypsin. Syndecan and ICAM-1 were increased in HMVEC exposed to IEC supernatants. TXA administration 'early' abrogated these effects. CONCLUSIONS: TXA administration early after shock protects the gut barrier by inhibiting trypsin uptake and activity and the subsequent downstream protease cascade. To be effective, TXA should be administered early in all "at risk" patients.
Asunto(s)
Antifibrinolíticos/farmacología , Ácido Tranexámico/farmacología , Tripsina/metabolismo , Proteína ADAM17/efectos de los fármacos , Proteína ADAM17/metabolismo , Línea Celular , Células Cultivadas , Células Epiteliales/metabolismo , Humanos , Molécula 1 de Adhesión Intercelular/efectos de los fármacos , Molécula 1 de Adhesión Intercelular/metabolismo , Intestinos/citología , Pulmón/citología , Metaloproteinasa 9 de la Matriz/efectos de los fármacos , Metaloproteinasa 9 de la Matriz/metabolismo , Choque Hemorrágico/complicaciones , Sindecanos/metabolismo , Tripsina/efectos de los fármacosRESUMEN
Angiotensin II (AngII) has been strongly implicated in hypertension and its complications. Evidence suggests the mechanisms by which AngII elevates blood pressure and enhances cardiovascular remodeling and damage may be distinct. However, the signal transduction cascade by which AngII specifically initiates cardiovascular remodeling, such as hypertrophy and fibrosis, remains insufficiently understood. In vascular smooth muscle cells, a metalloproteinase ADAM17 mediates epidermal growth factor receptor transactivation, which may be responsible for cardiovascular remodeling but not hypertension induced by AngII. Thus, the objective of this study was to test the hypothesis that activation of vascular ADAM17 is indispensable for vascular remodeling but not for hypertension induced by AngII. Vascular ADAM17-deficient mice and control mice were infused with AngII for 2 weeks. Control mice infused with AngII showed cardiac hypertrophy, vascular medial hypertrophy, and perivascular fibrosis. These phenotypes were prevented in vascular ADAM17-deficient mice independent of blood pressure alteration. AngII infusion enhanced ADAM17 expression, epidermal growth factor receptor activation, and endoplasmic reticulum stress in the vasculature, which were diminished in ADAM17-deficient mice. Treatment with a human cross-reactive ADAM17 inhibitory antibody also prevented cardiovascular remodeling and endoplasmic reticulum stress but not hypertension in C57Bl/6 mice infused with AngII. In vitro data further supported these findings. In conclusion, vascular ADAM17 mediates AngII-induced cardiovascular remodeling via epidermal growth factor receptor activation independent of blood pressure regulation. ADAM17 seems to be a unique therapeutic target for the prevention of hypertensive complications.
Asunto(s)
Proteína ADAM17/efectos de los fármacos , Proteína ADAM17/metabolismo , Angiotensina II/farmacología , Cardiomegalia/metabolismo , Receptores ErbB/metabolismo , Hipertensión/complicaciones , Animales , Cardiomegalia/prevención & control , Células Cultivadas , Modelos Animales de Enfermedad , Fibrosis/metabolismo , Fibrosis/patología , Humanos , Hipertensión/inducido químicamente , Hipertensión/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , Terapia Molecular Dirigida , Miocitos Cardíacos/metabolismo , Distribución Aleatoria , Sistema Renina-Angiotensina/fisiología , Sensibilidad y Especificidad , Transducción de Señal/efectos de los fármacos , Remodelación Vascular/efectos de los fármacos , Remodelación Vascular/fisiología , Remodelación Ventricular/efectos de los fármacos , Remodelación Ventricular/fisiologíaRESUMEN
Betel quid (BQ) chewing is an etiologic factor of oral submucous fibrosis (OSF) and oral cancer. There are 600 million BQ chewers worldwide. The mechanisms for the toxic and inflammatory responses of BQ are unclear. In this study, both areca nut (AN) extract (ANE) and arecoline stimulated epidermal growth factor (EGF) and interleukin-1α (IL-1α) production of gingival keratinocytes (GKs), whereas only ANE can stimulate a disintegrin and metalloproteinase 17 (ADAM17), prostaglandin E2 (PGE2) and 8-isoprostane production. ANE-induced EGF production was inhibited by catalase. Addition of anti-EGF neutralizing antibody attenuated ANE-induced cyclooxygenase-2 (COX-2), mature ADAM9 expression and PGE2 and 8-isoprostane production. ANE-induced IL-1α production was inhibited by catalase, anti-EGF antibody, PD153035 (EGF receptor antagonist) and U0126 (MEK inhibitor) but not by α-naphthoflavone (cytochrome p450-1A1 inhibitor). ANE-induced ADAM17 production was inhibited by pp2 (Src inhibitor), U0126, α-naphthoflavone and aspirin. AG490 (JAK inhibitor) prevented ANE-stimulated ADAM17, IL-1α, PGE2 production, COX-2 expression, ADAM9 maturation, and the ANE-induced decline in keratin 5 and 14, but showed little effect on cdc2 expression and EGF production. Moreover, ANE-induced 8-isoprostane production by GKs was inhibited by catalase, anti-EGF antibody, AG490, pp2, U0126, α-naphthoflavone, Zinc protoporphyrin (ZnPP) and aspirin. These results indicate that AN components may involve in BQ-induced oral cancer by induction of reactive oxygen species, EGF/EGFR, IL-1α, ADAMs, JAK, Src, MEK/ERK, CYP1A1, and COX signaling pathways, and the aberration of cell cycle and differentiation. Various blockers against ROS, EGF, IL-1α, ADAM, JAK, Src, MEK, CYP1A1, and COX can be used for prevention or treatment of BQ chewing-related diseases.