Nuestros investigadores

Josune Orbe Lopategui

Publicaciones científicas más recientes (desde 2010)

Autores: Rodríguez, José Antonio; Fernández-Seara, M. A.; et al.
Revista: SCIENTIFIC REPORTS
ISSN 2045-2322  2020 
Matrix metalloproteinases have been implicated in diabetic microvascular complications. However, little is known about the pathophysiological links between MMP-10 and the renin-angiotensin system (RAS) in diabetic kidney disease (DKD). We tested the hypothesis that MMP-10 may be up-regulated in early stage DKD, and could be down-regulated by angiotensin II receptor blockade (telmisartan). Serum MMP-10 and TIMP-1 levels were measured in 268 type 2 diabetic subjects and 111 controls. Furthermore, histological and molecular analyses were performed to evaluate the renal expression of Mmp10 and Timp1 in a murine model of early type 2 DKD (db/db) after telmisartan treatment. MMP-10 (473¿±¿274¿pg/ml vs. 332¿±¿151; p¿=¿0.02) and TIMP-1 (573¿±¿296¿ng/ml vs. 375¿±¿317; p¿<¿0.001) levels were significantly increased in diabetic patients as compared to controls. An early increase in MMP-10 and TIMP-1 was observed and a further progressive elevation was found as DKD progressed to end-stage renal disease. Diabetic mice had 4-fold greater glomerular Mmp10 expression and significant albuminuria compared to wild-type, which was prevented by telmisartan. MMP-10 and TIMP-1 are increased from the early stages of type 2 diabetes. Prevention of MMP-10 upregulation observed in diabetic mice could be another protective mechanism of RAS blockade in DKD.
Autores: Roncal, Carmen; Martinez-Aguilar, E.; Orbe, J; et al.
Revista: SCIENTIFIC REPORTS
ISSN 2045-2322  Vol. 9  2019  págs. 15580
Peripheral artery disease (PAD) is a major cause of acute and chronic illness, with extremely poor prognosis that remains underdiagnosed and undertreated. Trimethylamine-N-Oxide (TMAO), a gut derived metabolite, has been associated with atherosclerotic burden. We determined plasma levels of TMAO by mass spectrometry and evaluated their association with PAD severity and prognosis. 262 symptomatic PAD patients (mean age 70 years, 87% men) categorized in intermittent claudication (IC, n = 147) and critical limb ischemia (CLI, n = 115) were followed-up for a mean average of 4 years (min 1-max 102 months). TMAO levels were increased in CLI compared to IC (P < 0.001). Receiver operating characteristic (ROC) curves for severity (CLI) rendered a cutoff of 2.26 mu mol/L for TMAO (62% sensitivity, 76% specificity). Patients with TMAO > 2.26 mu mol/L exhibited higher risk of cardiovascular death (sub-hazard ratios >= 2, P < 0.05) that remained significant after adjustment for confounding factors. TMAO levels were associated to disease severity and CV-mortality in our cohort, suggesting an improvement of PAD prognosis with the measurement of TMAO. Overall, our results indicate that the intestinal bacterial function, together with the activity of key hepatic enzymes for TMA oxidation (FMO3) and renal function, should be considered when designing therapeutic strategies to control gut-derived metabolites in vascular patients.
Autores: Roncal, Carmen; Salicio, Agustina; et al.
Revista: TRANSLATIONAL STROKE RESEARCH
ISSN 1868-4483  Vol. 10  Nº 4  2019  págs. 389 - 401
Diabetes is an important risk factor for ischemic stroke (IS). Tissue-type plasminogen activator (tPA) has been associated with less successful revascularization and poor functional outcome in diabetes. We assessed whether a new thrombolytic strategy based on MMP10 was more effective than tPA in a murine IS model of streptozotocin (STZ)-induced diabetes. Wild-type mice were administered a single dose of streptozotocin (STZ) (180mg/kg) to develop STZ-induced diabetes mellitus. Two weeks later, IS was induced by thrombin injection into the middle cerebral artery and the effect of recombinant MMP10 (6.5 mu g/kg), tPA (10mg/kg) or tPA/MMP10 on brain damage and functional outcome were analysed. Motor activity was assessed using the open field test. Additionally, we studied plasminogen activator inhibitor-1 (PAI-1) and thrombin-antithrombin complex levels (TAT) by ELISA and oxidative stress and blood-brain barrier (BBB) integrity by immunohistochemistry and western blot. MMP10 treatment was more effective at reducing infarct size and neurodegeneration than tPA 24h and 3days after IS in diabetic mice. Locomotor activity was impaired by hyperglycemia and ischemic injury, but not by the thrombolytic treatments. Additionally, TAT, oxidative stress and BBB permeability were reduced by MMP10 treatment, whereas brain bleeding or PAI-1 expression did not differ between treatments. Thrombolytic treatment with MMP10 was more effective than tPA at reducing stroke and neurodegeneration in a diabetic murine model of IS, without increasing haemorrhage. Thus, we propose MMP10 as a potential candidate for the clinical treatment of IS in diabetic patients.
Autores: Lorente, L., (Autor de correspondencia); Martin, M. M.; Ramos, L.; et al.
Revista: JOURNAL OF CRITICAL CARE
ISSN 0883-9441  Vol. 51  2019  págs. 117 - 121
Purpose: Previously, higher circulating levels of matrix metalloproteinase (MMP)-9 and tissue inhibitor matrix metalloproteinases (TIMP)-1 were reported in the first hours after TBI in blood samples from patients with poor prognosis. Thus, the objectives of this study were to determine whether MMP-9 and TIMP-1 levels during the first week of a severe TBI could be used as biomarker predictive of mortality. Methods: We included patients with severe TBI (defined as Glasgow Coma Scale lower than 9), and with Injury Severity Score in non-cranial aspects lower than 9. We determined serum concentrations of MMP-9 and TIMP-1 at days 1, 4 and 8 of TBI. Results: TIMP-1 concentrations at days 1 (p < .001), 4 (p = .001), and 8 (p = .01) of TBI were higher in nonsurviving (n = 34) than in surviving (n = 90) patients. ROC curve analyses showed an area under curve of TIMP-1 concentrations at days 1, 4, and 8 of TBI to predict 30-day mortality of 78% (p < .001), 76% (p <.001) and 71% (p= .02) respectively. Conclusions: The most relevant new findings of our study were that TIMP-1 levels during the first week of a severe TBI were higher in non-surviving than in surviving patients and that could be used as biomarker predictive of mortality. (C) 2019 Elsevier Inc. All rights reserved.
Autores: Lorente, L., (Autor de correspondencia); Martin, M. M. ; Ramos, L.; et al.
Revista: BMC NEUROLOGY
ISSN 1471-2377  Vol. 19  2019  págs. 167
Background: Higher circulating levels of tissue inhibitor of matrix metalloproteinases (TIMP)-1 early after ischemic stroke have been associated with lower survival. The objectives of this study were to determine serum TIMP-1 levels during the first week of a severe cerebral infarction in surviving and non-surviving patients, and whether those levels during the first week could be used as a mortality biomarker for these patients. Methods: We included patients with severe malignant middle cerebral artery infarction (MMCAI) defined as computer tomography showing ischaemic changes in more than 50% of the middle cerebral artery territory and Glasgow Coma Scale (GCS) <= 8. We measured serum levels of matrix metalloproteinases (MMP)-9 and TIMP-1. End-point study was 30-day mortality. Results: We found higher TIMP-1 concentrations at days 1 (p < 0.001), 4 (p = 0.001), and 8 (p = 0.03) of MMCAI in non-urviving (n = 34) than in surviving (n = 34) patients. We found lower serum MMP-9 concentrations at day 1 (p = 0.03) of MMCAI and no significant differences at days 4 and 8. ROC curve analysis of TIMP-1 concentrations performed at days 1, 4, and 8 of MMCAI showed an area under curve to predict 30-day mortality of 81% (p < 0.001), 80% (p < 0.001) and 72% (p = 0.07) respectively. Conclusions: The new findings of our study were that non-surviving MMCAI patients showed higher serum TIMP-1 levels during the first week of MMCAI that surviving patients, and those levels during the first week of MMCAI could be used as mortality biomarkers.
Autores: Marcos-Jubilar, María; Orbe, J; Roncal, Carmen; et al.
Revista: CLINICA E INVESTIGACION EN ARTERIOSCLEROSIS
ISSN 0214-9168  Vol. 31  Nº 4  2019  págs. 152 - 159
Introduction: Monocytes play an important role in atherosclerotic progression having both pro and anti-inflammatory effects depending on different circulating monocyte subpopulations. The objective of this study is to characterize these subpopulations and their association with cardiovascular risk factors. Methods: Transversal study including 102 selected patients, mean age: 65 years-old (range 41-86), 69% males. A set of specific antibodies against classical monocytes (Mont, CD14+CD16- CD300e+HLADR+), intermediate (Mon2, CD14+CD16+CD300e+HLADR+) and nonclassical (Mon3, CD14 CD16+CD300e+HLADR+) was assayed. Three groups of patients were included: 17 asymptomatic with more than one cardiovascular risk factor (group 1), 56 subjects asymptomatic but with vascular pathology assessed by ultrasound or microalbuminuria (group 2) and 19 patients with a previous atherothrombotic event (group 3). The cardiovascular risk was determined by Framingham and REGICOR scores. Results: An association between study groups and the percentage of Mon1 and Mon2 was observed (ANOVA, p <.05), being independent of age and sex for Mon2. Likewise Mont and Mon2 subpopulations were associated with cardiovascular adverse events (beta=0.86, p=.02 beta-0.1 p=.002, respectively), independently of age and sex in the case of Mon2. Moreover the percentage of Mon3 was associated with the presence of several cardiovascular risk factors ((3 = 0.21, p =.04) in the univariate analysis. In addition, there was a correlation between the levels of Mon1 and Mon2 and leukocytes (r =0.7, p <.001 and r =0.26, p =.01, respectively). Conclusions: The analysis of monocyte subpopulations may be clinically useful to stratify the inflammatory profile related to the different cardiovascular risk groups.
Autores: Marcos-Jubilar, María; Pastrana, Juan Carlos; Orbe, J; et al.
Revista: HAEMATOLOGICA
ISSN 0390-6078  Vol. 104  2019  págs. 144 - 144
Autores: Orbe, J; Aguilar, E. M.; et al.
Revista: ATHEROSCLEROSIS
ISSN 0021-9150  Vol. 287  2019  págs. E141 - E141
Autores: David, L.; Maillo, A.; et al.
Revista: ATHEROSCLEROSIS
ISSN 0021-9150  Vol. 287  2019  págs. E65 - E65
Autores: Enguita, J. M.; Saldise, M. B.; et al.
Revista: HAEMATOLOGICA
ISSN 0390-6078  Vol. 104  2019  págs. 135 - 136
Autores: Hernandez, M.; Fernandez-Robredo, P; Recalde, Sergio; et al.
Revista: INVESTIGATIVE OPHTHALMOLOGY AND VISUAL SCIENCE
ISSN 0146-0404  Vol. 60  Nº 9  2019 
Autores: Rodríguez, José Antonio; Orbe, J; Saenz-Pipaon, G.; et al.
Revista: THROMBOSIS RESEARCH
ISSN 0049-3848  Vol. 170  2018  págs. 1 - 9
Introduction: Wound healing after myocardial infarction (MI) is mediated by different cell types, secreted proteins, components of the extracellular matrix (ECM) and, as increasing evidences suggest, extracellular vesicles (EVs). We aim to determine the dynamics of release and origin of EVs after MI, as well as their biological activity on endothelial cells (ECs). Methods: MI was induced in WT mice and blood and tissues collected at baseline, 3, 15 and 30 days post-ligation for cardiac function (echocardiography) and histological evaluation. Circulating EVs subpopulations were measured by flow cytometry in mouse, and in a small cohort of patients with ST-segment elevation MI (STEMI, n= 6). In vitro, EVs were isolated from a cardiomyocyte cell line (HL1) and their function assayed on ECs. Results: Leukocyte and endothelial EVs increased concomitant to inflammatory and angiogenic processes triggered by ischemia. More strikingly, cardiomyocyte EVs (connexin43+) were detected in STEMI patients and in murine MI, where a significant increase in their levels was reported at day 15 post-ischemia (p < 0.05 vs baseline). In vitro, HL1EVs induced ECs migration (p= 0.05) and proliferation (p < 0.05), but impaired tube formation. These apparent contradictory results could be partially explained by the upregulation of MMP3, and the apoptosis and senescence genes, p53 and p16, induced by HL1EVs on ECs (p < 0.05). Conclusions: MI induces the release of different EVs subpopulations, including those of cardiac origin, in a preclinical model of MI and STEMI patients. In vitro, cardiomyocyte derived EVs are able to modulate endothelial function, suggesting their active role in heart repair after ischemia.
Autores: de Miguel, I.; Orbe, J; Sanchez-Arias, J. A. ; et al.
Revista: ACS MEDICINAL CHEMISTRY LETTERS
ISSN 1948-5875  Vol. 9  Nº 5  2018  págs. 428 - 433
In an effort to find novel chemical series as antifibrinolytic agents, we explore alpha-phenylsulfonyl-alpha-spiropiperidines bearing different zinc-binding groups (ZBGs) to target those metalloproteinases involved in the fibrinolytic process: MMP3 and MMP10. Surprisingly, all these new chemical series were inactive against these metalloproteinases; however, several new molecules retained the antifibrinolytic activity in a phenotypic functional assay using thromboelastometry and human whole blood. Further optimization led to compound 38 as a potent antifibrinolytic agent in vivo, three times more efficacious than the current standard-of-care (tranexamic acid, TXA) at 300 times lower dose. Finally, in order to decipher the underlying mode-of action leading to this phenotypic response, an affinity-based probe 39 was successfully designed to identify the target involved in this response: a potentially unknown mechanism-of-action in the fibrinolytic process.
Autores: Roncal, Carmen; Orbe, J; et al.
Revista: ATHEROSCLEROSIS
ISSN 0021-9150  Vol. 278  2018  págs. 124 - 134
Background and aims: Matrix metalloproteinases (MMPs) have been implicated in atherosclerosis and vascular calcification. Among them, we reported that MMP10 is present in human atheroma, associated with atherosclerosis. However, it remains unclear whether MMP10 is involved in atherogenesis and vascular calcification. Methods: MMP10 was measured in serum from patients with subclinical atherosclerosis and analyzed in carotid endarterectomies by immunostaining. ApoE-deficient mice (Apoe(-/-)) were crossed to MMP10-deficient (Mmp10(-/-)) mice and followed up to 20 months. Plaque area and composition were assessed by histology and immunohistochemistry. Inflammatory markers were measured in atherosclerotic plaques by RT-qPCR, and leukocyte subpopulations were analyzed by flow cytometry. In vitro calcification assays were performed in aortic vascular smooth muscle cells (VSMC). Results: MMP10 serum levels were associated with coronary calcification in subjects with subclinical atherosclerosis. Immunostaining revealed MMP10 expression in human atheromas, spatially associated with calcification areas, and complicated plaques released higher amounts of MMP10 than non-diseased segments. Interestingly, vascular MMP10 expression was confined to the atherosclerotic lesion in Apoe(-/-) mice, and Apoe(-/-) Mmp10(-/-) showed a substantial reduction in atherosclerotic lesion size, macrophage content and plaque calcification. Reduced local and systemic inflammatory markers could be demonstrated in Apoe(-/-) Mmp10(-/-) by gene expression and flow cytometry analysis. Calcium phosphate deposition and vascular calcification markers were downregulated in VSMC from Apoe(-/-) Mmp10(-/-) mice. Conclusions: Delayed plaque progression and altered cellular composition in the absence of MMP10 suggests that MMP10 plays a role in atherosclerosis, favoring inflammation, development and complication of the plaque.
Autores: Reyes, R.; Rodríguez, José Antonio; Orbe, J; et al.
Revista: DRUG DELIVERY
ISSN 1071-7544  Vol. 25  Nº 1  2018  págs. 750 - 756
The effect of dual delivery of bone morphogenetic protein-2 (BMP-2) and matrix metalloproteinase 10 (MMP10) on bone regeneration was investigated in a murine model of calvarial critical-size defect, hypothesizing that it would result in an enhanced bone formation. Critical-size calvarial defects (4 mm diameter) were created in mice and PLGA microspheres preloaded with either BMP-2, MMP10 or a microsphere combination of both were transplanted into defect sites at different doses. Empty microspheres were used as the negative control. Encapsulation efficiency was assessed and in vivo release kinetics of BMP-2 and MMP10 were examined over 14 days. Histological analyses were used to analyze bone formation after four and eightweeks. Combination with MMP10 (30 ng) significantly enhanced BMP-2 (600ng)-mediated osteogenesis, as confirmed by the increase in percentage of bone fill (p < .05) at four weeks. Moreover, it also increased mineral apposition rate (p < .05), measured by double labeling with tetracycline and calceine. MMP10 accelerates bone repair by enhancing BMP-2-promoted bone healing and improving the mineralization rate. In conclusion combination of MMP10 and BMP-2 may become a promising strategy for repair and regeneration of bone defects.
Autores: Salicio, M.; Zandio, B.; et al.
Revista: HAEMATOLOGICA
ISSN 0390-6078  Vol. 103  Nº Supl. 2  2018  págs. 2 - 2
Autores: Roncal, Carmen; Martinez-Aguilar, E. ; Orbe, J; et al.
Revista: EUROPEAN JOURNAL OF CLINICAL INVESTIGATION
ISSN 0014-2972  Vol. 48  Nº Supl. 1  2018  págs. 121 - 122
Autores: Martinez-Aguilar, E.; Orbe, J; Fernández-Montero, Alejandro; et al.
Revista: JOURNAL OF VASCULAR SURGERY
ISSN 0741-5214  Vol. 66  Nº 5  2017  págs. 1527 - 1533
Objective The prognosis of patients with peripheral arterial disease (PAD) is characterized by an exceptionally high risk for myocardial infarction, ischemic stroke, and death; however, studies in search of new prognostic biomarkers in PAD are scarce. Even though low levels of high-density lipoprotein cholesterol (HDL-C) have been associated with higher risk of cardiovascular (CV) complications and death in different atherosclerotic diseases, recent epidemiologic studies have challenged its prognostic utility. The aim of this study was to test the predictive value of HDL-C as a risk factor for ischemic events or death in symptomatic PAD patients. Methods Clinical and demographic parameters of 254 symptomatic PAD patients were recorded. Amputation, ischemic coronary disease, cerebrovascular disease, and all-cause mortality were recorded during a mean follow-up of 2.7 years. Results Multivariate analyses showed that disease severity (critical limb ischemia) was significantly reduced in patients with normal HDL-C levels compared with the group with low HDL-C levels (multivariate analysis odds ratio, 0.09; 95% confidence interval [CI], 0.03-0.24). A decreased risk for mortality (hazard ratio, 0.46; 95% CI, 0.21-0.99) and major adverse CV events (hazard ratio, 0.38; 95% CI, 0.16-0.86) was also found in patients with normal vs reduced levels of HDL-C in both Cox proportional hazards models and Kaplan-Meier estimates, after adjustment for confounding factors. Conclusions Reduced HDL-C levels were significantly associated with higher risk for development of CV complications as well as with mortality in PAD patients. These findings highlight the usefulness of this simple test for early identification of PAD patients at high risk for development of major CV events.
Autores: Demuth, H. U. ; Dijkhuizen, R. M. ; Farr, T. D. ; et al.
Revista: RESTORATIVE NEUROLOGY AND NEUROSCIENCE
ISSN 0922-6028  Vol. 35  Nº 1  2017  págs. 87 - 103
The already established and widely used intravenous application of recombinant tissue plasminogen activator as a re-opening strategy for acute vessel occlusion in ischemic stroke was recently added by mechanical thrombectomy, representing a fundamental progress in evidence-based medicine to improve the patient's outcome. This has been paralleled by a swift increase in our understanding of pathomechanisms underlying many neurovascular diseases and most prevalent forms of dementia. Taken together, these current advances offer the potential to overcome almost two decades of marginally successful translational research on stroke and dementia, thereby spurring the entire field of translational neuroscience. Moreover, they may also pave the way for the renaissance of classical neuroprotective paradigms. This review reports and summarizes some of the most interesting and promising recent achievements in neurovascular and dementia research. It highlights sessions from the 9th International Symposium on Neuroprotection and Neurorepair that have been discussed from April 19th to 22nd in Leipzig, Germany. To acknowledge the emerging culture of interdisciplinary collaboration and research, special emphasis is given on translational stories ranging from fundamental research on neurodeand -regeneration to late stage translational or early stage clinical investigations.
Autores: Roncal, Carmen; Salicio, Agustina; et al.
Revista: CARDIOVASCULAR RESEARCH
ISSN 0008-6363  Vol. 113  Nº 10  2017  págs. 1219 - 1229
Aims Early reperfusion with tissue-type plasminogen activator (tPA) is an effective therapeutic strategy to treat acute ischemic stroke, but only 1/3 of tPA-treated patients recover and are free from disability. tPA has also shown neurotoxicity in experimental models of cerebral ischemia. Considering that MMP-10 improves stroke injury, we have examined the therapeutic and protective effect of MMP10 and tPA/MMP10 as clot-dissolving and neuroprotective agent in an experimental model of ischemic stroke and studied in vitro the molecular pathways involved in MMP10-mediated effects. Methods and results Cerebral ischemia was induced by the local injection of thrombin into the middle cerebral artery followed by reperfusion with MMP10 (6.5 mu g/kg) and tPA (10 mg/kg) alone or in combination with MMP10. Cell cultures were also performed to determine the effect of MMP10 and tPA/MMP10 on brain endothelial cells and neurons. tPA/MMP10 significantly reduced the infarct size in the ischemic stroke model compared with tPA alone (P < 0.05). In vitro, MMP10 reduced the tPA-promoted endothelial ionic permeability, preserved the expression of claudin-5 and decreased ERK1/2 activation. Moreover, combination of tPA/MMP10 prevented tPA-mediated neuronal excitotoxicity and calcium influx. These effects were reversed by blocking MMP10 activity with a monoclonal antibody. Conclusion These results show that MMP10, either alone or in combination with tPA, might represent a new strategy for thrombolysis in ischemic stroke, providing higher protection against cerebrovascular damage.
Autores: Rodríguez, José Antonio; Sobrino, T.; López-Arias, E.; et al.
Revista: AMERICAN HEART ASSOCIATION. JOURNAL. CARDIOVASCULAR AND CEREBROVASCULAR DISEASE
ISSN 2047-9980  Vol. 6  Nº 6  2017  págs. e006042
BACKGROUND: Intracerebral hemorrhage (ICH) is an acute neurological disorder with high mortality and no effective treatment. In addition to the initial bleeding event, rebleeding and hematoma expansion are associated with poor outcome in these patients. We studied the effectiveness of the new antifibrinolytic agent CM352, a short-half-life matrix metalloproteinase inhibitor, for achieving early hemostasis and improving functional recovery in a rat model of collagenase-induced ICH. METHODS AND RESULTS: ICH was induced by striatal injection of collagenase, and 1 hour later, rats received an intravenous injection of saline (n=6) or CM352 (1 mg/kg, n=6). Hematoma (basal and after 3 and 24 hours) and lesion (14 days) volumes were quantified on T2-weighted (T2) magnetic resonance images. Neurological and functional recovery was evaluated by using Bederson score and a cylinder test (basal, 24 hours, and 14 days). Early treatment (1 hour) with CM352 was efficient reducing hematoma expansion at 3 hours (P<0.01) and, more markedly, at 24 hours (P<0.01). Decreased bleeding after antifibrinolytic treatment was accompanied by reduced interleukin-6 levels at 3 hours (P<0.05) and smaller lesion volume at 14 days (P<0.01). CM352 drastically reduced sensorimotor impairment (cylinder test) after ICH in rats at 24 hours (P<0.01) and 14 days (P<0.01). Similarly, it also attenuated neurological deficit (Bederson scale) at 24 hours (P<0.01) and 14 days (P<0.01). Interestingly, late (3 hours) CM352 administration also resulted in reduced lesion size and better functional outcome. CONCLUSIONS: CM352, a new antifibrinolytic agent and matrix metalloproteinase inhibitor, effectively prevented hematoma growth and reduced lesion size in ICH in association with improved functional and neurological recovery.
Autores: Orset, C.; Haelewyn, B.; Allan, S. M.; et al.
Revista: STROKE
ISSN 0039-2499  Vol. 47  Nº 5  2016  págs. 1312 - 1318
Background and Purpose The debate over the fact that experimental drugs proposed for the treatment of stroke fail in the translation to the clinical situation has attracted considerable attention in the literature. In this context, we present a retrospective pooled analysis of a large data set from preclinical studies, to examine the effects of early versus late administration of intravenous recombinant tissue-type plasminogen activator. Methods We collected data from 26 individual studies from 9 international centers (13 researchers; 716 animals) that compared recombinant tissue-type plasminogen activator with controls, in a unique mouse model of thromboembolic stroke induced by an in situ injection of thrombin into the middle cerebral artery. Studies were classified into early (<3 hours) versus late (3 hours) drug administration. Final infarct volumes, assessed by histology or magnetic resonance imaging, were compared in each study, and the absolute differences were pooled in a random-effect meta-analysis. The influence of time of administration was tested. Results When compared with saline controls, early recombinant tissue-type plasminogen activator administration was associated with a significant benefit (absolute difference, -6.63 mm(3); 95% confidence interval, -9.08 to -4.17; I-2=76%), whereas late recombinant tissue-type plasminogen activator treatment showed a deleterious effect (+5.06 mm(3); 95% confidence interval, +2.78 to +7.34; I-2=42%; P-int<0.00001). Results remained unchanged after subgroup analyses. Conclusions Our results provide the basis needed for the design of future preclinical studies on recanalization therapies using this model of thromboembolic stroke in mice. The power analysis reveals that a multicenter trial would require 123 animals per group instead of 40 for a single-center trial.
Autores: Sobrino, T.; Rodríguez, José Antonio; Vieites-Prado, A.; et al.
Revista: JOURNAL OF CEREBRAL BLOOD FLOW AND METABOLISM
ISSN 0271-678X  Vol. 36  Nº Supl. 1  2016  págs. 58 - 59
Autores: Rivera-Torres, J; Guzman-Martinez, G; Villa-Bellosta, R; et al.
Revista: JOURNAL OF HYPERTENSION
ISSN 0263-6352  Vol. 33  Nº 4  2015  págs. 843 - 850
OBJECTIVE: The Notch pathway has been linked to pulmonary hypertension, but its role in systemic hypertension and, in particular in left ventricular hypertrophy (LVH), remains poorly understood. The main objective of this work was to analyse the effect of inhibiting the Notch pathway on the establishment and maintenance of angiotensin II (Ang-II)-induced arterial hypertension and LVH in adult mice with inducible genetic deletion of ¿-secretase, and to test preclinically the therapeutic efficacy of ¿-secretase inhibitors (GSIs). BASIC METHODS: We analysed Ang-II responses in primary cultures of vascular smooth muscle cells obtained from a novel mouse model with inducible genetic deletion of the ¿-secretase complex, and the effects of GSI treatment on a mouse cardiac cell line. We also investigated Ang-II-induced hypertension and LVH in our novel mouse strain lacking the ¿-secretase complex and in GSI-treated wild-type mice. Moreover, we analysed vascular tissue from hypertensive patients with and without LVH. MAIN RESULTS: Vascular smooth muscle cells activate the Notch pathway in response to Ang-II both 'in vitro' and 'in vivo'. Genetic deletion of ¿-secretase in adult mice prevented Ang-II-induced hypertension and LVH without causing major adverse effects. Treatment with GSI reduced Ang-II-induced hypertrophy of a cardiac cell line 'in vitro' and LVH in wild-type mice challenged with Ang-II. We also report elevated expression of the Notch target HES5 in vascular tissue from hypertensive patients with LVH compared with those without LVH. CONCLUSION: The Notch pathway is activated in the vasculature of mice with hypertension and LVH, and its inhibition via inducible genetic ¿-secretase deletion protects against both conditions. Preliminary observations in hypertensive patients with LVH support the translational potential of these findings. Moreover, GSI treatment protects wild-type mice from Ang-II-induced LVH without affecting blood pressure. Our results unveil the potential use of GSIs in the treatment of hypertensive patients with LVH.
Autores: García, Oihane; Latasa, María Ujué; Carotti, S.; et al.
Revista: HEPATOLOGY
ISSN 0270-9139  Vol. 62  Nº 1  2015  págs. 166 - 178
Matrix metalloproteinases (MMPs) participate in tissue repair after acute injury, but also participate in cancer by promoting a protumorigenic microenvironment. Previously, we reported on a key role for MMP10 in mouse liver regeneration. Herein, we investigated MMP10 expression and function in human hepatocellular carcinoma (HCC) and diethylnitrosamine (DEN)-induced mouse hepatocarcinogenesis. MMP10 was induced in human and murine HCC tissues and cells. MMP10-deficient mice showed less HCC incidence, smaller histological lesions, reduced tumor vascularization, and less lung metastases. Importantly, expression of the protumorigenic, C-X-C chemokine receptor-4 (CXCR4), was reduced in DEN-induced MMP10-deficient mice livers. Human HCC cells stably expressing MMP10 had increased CXCR4 expression and migratory capacity. Pharmacological inhibition of CXCR4 significantly reduced MMP10-stimulated HCC cell migration. Furthermore, MMP10 expression in HCC cells was induced by hypoxia and the CXCR4 ligand, stromal-derived factor-1 (SDF1), through the extracellular signal-regulated kinase 1/2 pathway, involving an activator protein 1 site in MMP10 gene promoter. CONCLUSION: MMP10 contributes to HCC development, participating in tumor angiogenesis, growth, and dissemination. We identified a new reciprocal crosstalk between MMP10 and the CXCR4/SDF1 axis contributing to HCC progression and metastasis. To our knowledge, this is the first report addressing the role of a MMP in hepatocarcinogenesis in the corresponding genetic mouse model.
Autores: Orbe, J; Martínez-Aguilar, E.; Rodríguez, José Antonio; et al.
Revista: FASEB JOURNAL
ISSN 0892-6638  Vol. 29  Nº 3  2015  págs. 960 - 972
We studied the role of matrix metalloproteinase-10 (MMP-10) during skeletal muscle repair after ischemia using a model of femoral artery excision in wild-type (WT) and MMP-10 deficient (Mmp10(-/-)) mice. Functional changes were analyzed by small animal positron emission tomography and tissue morphology by immunohistochemistry. Gene expression and protein analysis were used to study the molecular mechanisms governed by MMP-10 in hypoxia. Early after ischemia, MMP-10 deficiency resulted in delayed tissue reperfusion (10%, P < 0.01) and in increased necrosis (2-fold, P < 0.01), neutrophil (4-fold, P < 0.01), and macrophage (1.5-fold, P < 0.01) infiltration. These differences at early time points resulted in delayed myotube regeneration in Mmp10(-/-) soleus at later stages (regenerating myofibers: 30 ± 9% WT vs. 68 ± 10% Mmp10(-/-), P < 0.01). The injection of MMP-10 into Mmp10(-/-) mice rescued the observed phenotype. A molecular analysis revealed higher levels of Cxcl1 mRNA (10-fold, P < 0.05) and protein (30%) in the ischemic Mmp10(-/-) muscle resulting from a lack of transcriptional inhibition by MMP-10. This was further confirmed using siRNA against MMP-10 in vivo. Our results demonstrate an important role of MMP-10 for proper muscle repair after ischemia, and suggest that chemokine regulation such as Cxcl1 by MMP-10 is involved in muscle regeneration
Autores: Lorente, L.; Martin, M. M.; Ramos, L.; et al.
Revista: BMC NEUROLOGY
ISSN 1471-2377  Vol. 15  2015  págs. 111
BACKGROUND: In the last years, circulating matrix metalloproteinases (MMP)-9 levels have been associated with functional outcome in ischemic stroke patients. However the prognostic value of circulating levels of tissue inhibitor of matrix metalloproteinases (TIMP)-1 and MMP-10 in functional outcome of ischemic stroke patients has been scarcely studied. In addition, to our knowledge, serum MMP-9, MMP-10 and TIMP-1 levels in patients with malignant middle cerebral artery infarction (MMCAI) for mortality prediction have not been studied, and these were the objectives of this study. METHODS: This was a multicenter, observational and prospective study carried out in six Spanish Intensive Care Units. We included patients with severe MMCAI defined as Glasgow Coma Scale (GCS) lower than 9. We measured circulating levels of MMP-9, MMP-10, TIMP-1, in 50 patients with severe MMCAI at diagnosis and in 50 healthy subjects. Endpoint was 30-day mortality. RESULTS: Patients with severe MMCAI showed higher serum levels of MMP-9 (p = 0.001), MMP-10 (p < 0.001), and TIMP-1 (p = 0.02) than healthy subjects. Non-surviving MMCAI patients (n = 26) compared to survivor ones (n = 24) showed higher circulating levels of TIMP-1 (p < 0.001), MMP-10 (p = 0.02) and PAI-1(p = 0.02), and lower MMP-9 levels (p = 0.04). Multiple binomial logistic regression analysis showed that serum TIMP-1 levels > 239 ng/mL are associated with 30-day mortality (OR = 5.82; 95% CI = 1.37-24.73; P = 0.02) controlling for GCS and age. The area under the curve for TIMP-1 as predictor of 30-day mortality was 0.81 (95% CI = 0.67-0.91; P < 0.001). We found an association between circulating levels of TIMP-1 and MMP-10 (rho = 0.45; P = 0.001), plasminogen activator inhibitor (PAI)-1 (rho = 0.53; P < 0.001), and tumor necrosis factor (TNF)-alpha (rho = 0.70; P < 0.001). CONCLUSIONS: The most relevant and new findings of our study, were that serum TIMP-1 levels in MMCAI patients were associated with mortality, and could be used as a prognostic biomarker of mortality in MMCAI patients.
Autores: García, Oihane; Latasa, María Ujué; Carotti, S.; et al.
Revista: HEPATOLOGY
ISSN 0270-9139  Vol. 62  Nº 1  2015  págs. 166 - 178
Matrix metalloproteinases (MMPs) participate in tissue repair after acute injury, but also participate in cancer by promoting a protumorigenic microenvironment. Previously, we reported on a key role for MMP10 in mouse liver regeneration. Herein, we investigated MMP10 expression and function in human hepatocellular carcinoma (HCC) and diethylnitrosamine (DEN)-induced mouse hepatocarcinogenesis. MMP10 was induced in human and murine HCC tissues and cells. MMP10-deficient mice showed less HCC incidence, smaller histological lesions, reduced tumor vascularization, and less lung metastases. Importantly, expression of the protumorigenic, C-X-C chemokine receptor-4 (CXCR4), was reduced in DEN-induced MMP10-deficient mice livers. Human HCC cells stably expressing MMP10 had increased CXCR4 expression and migratory capacity. Pharmacological inhibition of CXCR4 significantly reduced MMP10-stimulated HCC cell migration. Furthermore, MMP10 expression in HCC cells was induced by hypoxia and the CXCR4 ligand, stromal-derived factor-1 (SDF1), through the extracellular signal-regulated kinase 1/2 pathway, involving an activator protein 1 site in MMP10 gene promoter. Conclusion: MMP10 contributes to HCC development, participating in tumor angiogenesis, growth, and dissemination. We identified a new reciprocal crosstalk between MMP10 and the CXCR4/SDF1 axis contributing to HCC progression and metastasis. To our knowledge, this is the first report addressing the role of a MMP in hepatocarcinogenesis in the corresponding genetic mouse model. (Hepatology 2015;62:166-178)
Autores: De Gonzalo-Calvo, D.; Llorente-Cortés, V.; Orbe, J; et al.
Revista: INTERNATIONAL JOURNAL OF CARDIOLOGY
ISSN 0167-5273  Vol. 178  2015  págs. 102 - 104
Autores: Martínez-Aguilar, E.; Gómez-Rodríguez, V.; Orbe, J; et al.
Revista: JOURNAL OF VASCULAR SURGERY
ISSN 0741-5214  Vol. 61  Nº 2  2015  págs. 428 - 435
Objective: Peripheral arterial disease ( PAD) is associated with poor prognosis in terms of cardiovascular (CV) morbidity and mortality. Matrix metalloproteinases (MMPs) contribute to vascular remodeling by degrading extracellular matrix components and play a role in atherosclerosis as demonstrated for MMP-10 (stromelysin-2). This study analyzed MMP-10 levels in PAD patients according to disease severity and CV risk factors and evaluated the prognostic value of MMP-10 for CV events and mortality in lower limb arterial disease after a follow-up period of 2 years. Methods: MMP-10 was measured by enzyme-linked immunosorbent assay in 187 PAD patients and 200 sex-matched controls. Results: PAD patients presented with increased levels of MMP-10 (702 +/- 326 pg/mL control vs 946 +/- 473 pg/mL PAD; P < .001) and decreased levels of tissue inhibitor of matrix metalloproteinase 1 (312 +/- 117 ng/mL control vs 235 +/- 110 ng/mL PAD; P <.001) compared with controls. Among PAD patients, those with critical limb ischemia (n = 88) showed higher levels of MMP-10 (1086 6 478 pg/mL vs 822 6 436 pg/mL; P < .001) compared with those with intermittent claudication (n = 99), whereas the MMP-10/tissue inhibitor of matrix metalloproteinase 1 ratio remained similar. The univariate analysis showed an association between MMP-10, age (P = .015), hypertension (P = .021), and ankle-brachial index (P = .006) in PAD patients that remained significantly associated with PAD severity after adjustment for other CV risk factors. Patients with the highest MMP-10 tertile had an increased incidence of all-cause mortality and CV mortality (P < .03). Conclusions: Our results suggest that MMP-10 is associated with severity and poor outcome in PAD.
Autores: Orbe, J; Alexandru, N.; Roncal, Carmen; et al.
Revista: THROMBOSIS RESEARCH
ISSN 0049-3848  Vol. 136  Nº 2  2015  págs. 445 - 450
BACKGROUND: Thrombin-activatable fibrinolysis inhibitor (TAFI) plays an important role in coagulation and fibrinolysis. Whereas TAFI deficiency may lead to a haemorrhagic tendency, data from TAFI knockout mice (TAFI-/-) are controversial and no differences have been reported in these animals after ischemic stroke. There are also no data regarding the role of circulating microparticles (MPs) in TAFI-/-. OBJECTIVES: to examine the effect of tPA on the rate of intracranial haemorrhage (ICH) and on MPs generated in a model of ischemic stroke in TAFI-/- mice. METHODS: Thrombin was injected into the middle cerebral artery (MCA) to analyse the effect of tPA (10mg/Kg) on the infarct size and haemorrhage in the absence of TAFI. Immunofluorescence for Fluoro-Jade C was performed on frozen brain slides to analyse neuronal degeneration after ischemia. MPs were isolated from mouse blood and their concentrations calculated by flow cytometry. RESULTS: Compared with saline, tPA significantly increased the infarct size in TAFI-/- mice (p<0.05). Although plasma fibrinolytic activity (fibrin plate assay) was higher in these animals, no macroscopic or microscopic ICH was detected. A positive signal for apoptosis and degenerating neurons was observed in the infarct area, being significantly higher in tPA treated TAFI-/- mice (p<0.05). Interestingly, higher numbers of MPs were found in TAFI-/- plasma as compared to wild type, after stroke (p<0.05). CONCLUSIONS: TAFI deficiency results in increased brain damage in a model of thrombolysis after ischemic stroke, which was not associated with bleeding but with neuronal degeneration and MP production.
Autores: Orbe, J; Sánchez-Arias, J. A.; Rodríguez, José Antonio; et al.
Revista: JOURNAL OF MEDICINAL CHEMISTRY
ISSN 0022-2623  Vol. 58  Nº 5  2015  págs. 2465 - 2488
Growing evidence suggests that matrix metalloproteinases (MMP) are involved in thrombus dissolution; then, considering that new therapeutic strategies are required for controlling hemorrhage, we hypothesized that MMP inhibition may reduce bleeding by delaying fibrinolysis. Thus, we designed and synthesized a novel series of MMP inhibitors to identify potential candidates for acute treatment of bleeding. Structure-based and knowledge-based strategies were utilized to design this novel chemical series, ¿-spiropiperidine hydroxamates, of potent and soluble (>75 ¿g/mL) pan-MMP inhibitors. The initial hit, 12, was progressed to an optimal lead 19d. Racemic 19d showed a remarkable in vitro phenotypic response and outstanding in vivo efficacy; in fact, the mouse bleeding time at 1 mg/kg was 0.85 min compared to 29.28 min using saline. In addition, 19d displayed an optimal ADME and safety profile (e.g., no thrombus formation). Its corresponding enantiomers were separated, leading to the preclinical candidate 5 (described in Drug Annotations series, J. Med. Chem. 2015, ).
Autores: Orbe, J; Rodríguez, José Antonio; Sánchez-Arias, J. A.; et al.
Revista: JOURNAL OF MEDICINAL CHEMISTRY
ISSN 0022-2623  Vol. 58  Nº 7  2015  págs. 2941 - 2957
Discovery of potent and safe therapeutics that improve upon currently available antifibrinolytics, e.g., tranexamic acid (TXA, 1) and aprotinin, has been challenging. Matrix metalloproteinases (MMPs) participate in thrombus dissolution. Then we designed a novel series of optimized MMP inhibitors that went through phenotypic screening consisting of thromboelastometry and mouse tail bleeding. Our optimized lead compound, CM-352 (2), inhibited fibrinolysis in human whole blood functional assays and was more effective than the current standard of care, 1, in the tail-bleeding model using a 30¿000 times lower dose. Moreover, 2 reduced blood loss during liver hepatectomy, while 1 and aprotinin had no effect. Molecule 2 displayed optimal pharmacokinetic and safety profiles with no evidence of thrombosis or coagulation impairment. This novel mechanism of action, targeting MMP, defines a new class of antihemorrhagic agents without interfering with normal hemostatic function. Furthermore, 2 represents a preclinical candidate for the acute treatment of bleeding.
Autores: Martínez-Aguilar, E.; Orbe, J; Rodríguez, José Antonio; et al.
Revista: JOURNAL OF VASCULAR SURGERY
ISSN 0741-5214  Vol. 61  Nº 2  2015  págs. 428 - 435
Objective Peripheral arterial disease (PAD) is associated with poor prognosis in terms of cardiovascular (CV) morbidity and mortality. Matrix metalloproteinases (MMPs) contribute to vascular remodeling by degrading extracellular matrix components and play a role in atherosclerosis as demonstrated for MMP-10 (stromelysin-2). This study analyzed MMP-10 levels in PAD patients according to disease severity and CV risk factors and evaluated the prognostic value of MMP-10 for CV events and mortality in lower limb arterial disease after a follow-up period of 2 years. Methods MMP-10 was measured by enzyme-linked immunosorbent assay in 187 PAD patients and 200 sex-matched controls. Results PAD patients presented with increased levels of MMP-10 (702 ± 326 pg/mL control vs 946 ± 473 pg/mL PAD; P <.001) and decreased levels of tissue inhibitor of matrix metalloproteinase 1 (312 ± 117 ng/mL control vs 235 ± 110 ng/mL PAD; P <.001) compared with controls. Among PAD patients, those with critical limb ischemia (n = 88) showed higher levels of MMP-10 (1086 ± 478 pg/mL vs 822 ± 436 pg/mL; P <.001) compared with those with intermittent claudication (n = 99), whereas the MMP-10/tissue inhibitor of matrix metalloproteinase 1 ratio remained similar. The univariate analysis showed an association between MMP-10, age (P =.015), hypertension (P =.021), and ankle-brachial index (P =.006) in PAD patients that remained significantly associated with PAD severity after adjustment for other CV risk factors. Patients with the highest MMP-10 tertile had an increased incidence of all-cause mortality and CV mortality (P <.03). Conclusions Our results suggest that MMP-10 is associated with severity and poor outcome in PAD.
Autores: Gleeson, E. M. ; Dichiara, M. G.; Salicio, Agustina; et al.
Revista: BLOOD
ISSN 0006-4971  Vol. 126  Nº 7  2015  págs. 915 - 919
Activated protein C (APC) is an anticoagulant protease that initiates cell signaling via protease-activated receptor 1 (PAR1) to regulate vascular integrity and inflammatory response. In this study, a recombinant APC variant (APC(N329Q)) mimicking the naturally occurring APC-beta plasma glycoform was found to exhibit superior PAR1 proteolysis at a cleavage site that selectively mediates cytoprotective signaling. APC(N329Q) also enhanced integrin alpha(M)beta(2)-dependent PAR1 proteolysis to exert significantly improved antiinflammatory activity on macrophages compared with wild-type APC. Recent therapeutic applications of recombinant APC in ischemic stroke models have used APC variants with limited anticoagulant activity to negate potential bleeding side effects. Using a mouse model of ischemic stroke and late t-PA intervention, the neuroprotective activity of a murine APC variant with limited anticoagulant activity (mAPC(PS)) was compared with an identical APC variant except for the absence of glycosylation at the APC-beta sequon (mAPC(PS/N329Q)). Remarkably, mAPC(PS/N329Q) limited cerebral ischemic injury and reduced brain lesion volume significantly more effectively than mAPC(PS). Collectively, this study reveals the importance of APC glycosylation in controlling the efficacy of PAR1 proteolysis by APC and demonstrates the potential of novel APC variants with superior cytoprotective signaling function as enhanced therapeutic agents for the treatment of ischemic stroke.
Autores: Orbe, J; Rodríguez, José Antonio; Sanchez, J. A.; et al.
Revista: JOURNAL OF THROMBOSIS AND HAEMOSTASIS
ISSN 1538-7933  Vol. 13  Nº Supl. 2  2015  págs. 81 - 82
Autores: García, Oihane; Latasa, María Ujué; Carotti, S.; et al.
Revista: JOURNAL OF HEPATOLOGY
ISSN 0168-8278  Vol. 62  Nº Supl. 2  2015  págs. S242
Autores: Bobadilla, Miriam; Sáinz, Neira; Abizanda, Gloria María; et al.
Revista: STEM CELLS AND DEVELOPMENT
ISSN 1547-3287  Vol. 23  Nº 12  2014  págs. 1417-1427
The CXCR4/SDF1 axis participates in various cellular processes, including cell migration, which is essential for skeletal muscle repair. Although increasing evidence has confirmed the role of CXCR4/SDF1 in embryonic muscle development, the function of this pathway during adult myogenesis remains to be fully elucidated. In addition, a role for CXCR4 signaling in muscle maintenance and repair has only recently emerged. Here, we have demonstrated that CXCR4 and stromal cell-derived factor-1 (SDF1) are up-regulated in injured muscle, suggesting their involvement in the repair process. In addition, we found that notexin-damaged muscles showed delayed muscle regeneration on treatment with CXCR4 agonist (AMD3100). Accordingly, small-interfering RNA-mediated silencing of SDF1 or CXCR4 in injured muscles impaired muscle regeneration, whereas the addition of SDF1 ligand accelerated repair. Furthermore, we identified that CXCR4/SDF1-regulated muscle repair was dependent on matrix metalloproteinase-10 (MMP-10) activity. Thus, our findings support a model in which MMP-10 activity modulates CXCR4/SDF1 signaling, which is essential for efficient skeletal muscle regeneration.
Autores: Hernández, C.; Roncal, Carmen; Orbe, J; et al.
Revista: THROMBOSIS AND HAEMOSTASIS
ISSN 0340-6245  Vol. 111  Nº 1  2014  págs. 182
Autores: Bobadilla, Miriam; Sáinz, Neira; Rodríguez, José Antonio; et al.
Revista: STEM CELL
ISSN 1945-4570  Vol. 32  Nº 2  2014  págs. 447 - 461
Matrix metalloproteinases (MMPs), a family of endopeptidases that are involved in the degradation of extracellular matrix components, have been implicated in skeletal muscle regeneration. Among the MMPs, MMP-2 and MMP-9 are upregulated in Duchenne muscular dystrophy (DMD), a fatal X-linked muscle disorder. However, inhibition or overexpression of specific MMPs in a mouse model of DMD (mdx) has yielded mixed results regarding disease progression, depending on the MMP studied. Here, we have examined the role of MMP-10 in muscle regeneration during injury and muscular dystrophy. We found that skeletal muscle increases MMP-10 protein expression in response to damage (notexin) or disease (mdx mice), suggesting its role in muscle regeneration. In addition, we found that MMP-10-deficient muscles displayed impaired recruitment of endothelial cells, reduced levels of extracellular matrix proteins, diminished collagen deposition, and decreased fiber size, which collectively contributed to delayed muscle regeneration after injury. Also, MMP-10 knockout in mdx mice led to a deteriorated dystrophic phenotype. Moreover, MMP-10 mRNA silencing in injured muscles (wild-type and mdx) reduced muscle regeneration, while addition of recombinant human MMP-10 accelerated muscle repair, suggesting that MMP-10 is required for efficient muscle regeneration. Furthermore, our data suggest that MMP-10-mediated muscle repair is associated with VEGF/Akt signaling. Thus, our findings indicate that MMP-10 is
Autores: Llorente-Cortés, V.; De Gonzalo-Calvo, D.; Orbe, J; et al.
Revista: EUROPEAN JOURNAL OF CLINICAL INVESTIGATION
ISSN 0014-2972  Vol. 44  Nº 6  2014  págs. 539 - 548
BACKGROUND: Peripheral arterial disease is a relevant public health problem associated with increased risk of morbimortality. Most of the patients with this condition are asymptomatic. Therefore, the development of accessible biochemical markers seems to be necessary to anticipate diagnosis. Our hypothesis is that asymptomatic subjects with objectively confirmed femoral artery atherosclerosis could be distinguished from control subjects by gene expression analysis in peripheral blood mononuclear cells (PBMC). MATERIALS AND METHODS: A total of 37 asymptomatic males over 50 years old were recruited at the University Clinic of Navarra (Spain). Nineteen participants were free from atherosclerotic vascular disease and 18 participants presented subclinical femoral artery atherosclerosis defined by means of Doppler ultrasound. PBMC were isolated from blood and the RNA extracted. A panel of atherosclerotic-related genes were evaluated by Taqman low-density array. RESULTS: In univariate logistic regression models, we found a direct relationship between IL4, ITGAM and TLR2 expression levels in PBMC and femoral atherosclerosis, even when the models were adjusted for age and hypertension prevalence. Multivariate logistic regression models showed that elevated IL4 expression levels were intimately associated with subclinical femoral atherosclerosis after adjusting for the same potential confounders. CONCLUSIONS: Current data suggest that gene expression in PBMC, in particular IL4 expression, could be a useful tool in the diagnosis of femoral artery atherosclerosis in asymptomatic patients. Furthermore, in patients with no differences in cardiovascular risk factors except for hypertension, the results point to the immune and inflammatory deregulation as a feature of subclinical peripheral atherosclerosis.
Autores: Lorente, L.; Martín, M. M.; Solé-Violán, J.; et al.
Revista: PLOS ONE
ISSN 1932-6203  Vol. 9  Nº 4  2014  págs. e94318
Objective: Higher circulating levels of tissue inhibitor of matrix metalloproteinases (TIMP)-1 at the time of severe sepsis diagnosis have been reported in nonsurviving than in surviving patients. However, the following questions remain unanswered: 1) Does TIMP-1/MMP-9 ratio differ throughout the first week of intensive care between surviving and nonsurviving patients? 2) Is there an association between TIMP-1/MMP-9 ratio and sepsis severity and mortality during such period? 3) Could TIMP-1/MMP-9 ratio during the first week be used as an early biomarker of sepsis outcome? 4) Is there an association between TIMP-1/MMP-9 ratio and coagulation state and circulating cytokine levels during the first week of intensive care in these patients? The present study sought to answer these questions. Methods: Multicenter, observational and prospective study carried out in six Spanish Intensive Care Units (ICUs) of 295 patients with severe sepsis. Were measured circulating levels of TIMP-1, MMP-9, tumour necrosis factor (TNF)-alpha, interleukin (IL)-10 and plasminogen activator inhibitor (PAI)-1 at day 1, 4 and 8. End-point was 30-day mortality. Results: We found higher TIMP-1/MMP-9 ratio during the first week in non-surviving (n = 98) than in surviving patients (n = 197) (p<0.01). Logistic regression analyses showed that TIMP-1/MMP-9 ratio at days 1, 4 and 8 was associated with mortality. Receiver operating characteristic (ROC) curves showed that TIMP-1/MMP-9 ratio at days 1, 4 and 8 could predict mortality. There was an association between TIMP-1/MMP-9 ratio and TNF-alpha, IL-10, PAI-1 and lactic acid levels, SOFA score and platelet count at days 1, 4 and 8. Conclusions: The novel findings of our study were that non-surviving septic patients showed persistently higher TIMP-1/MMP-9 ratio than survivors ones during the first week, which was associated with severity, coagulation state, circulating cytokine levels and mortality; thus representing a new biomarker of sepsis outcome.
Autores: De Gonzalo-Calvo, D.; Llorente-Cortés, V.; Orbe, J; et al.
Revista: INTERNATIONAL JOURNAL OF CARDIOLOGY
ISSN 0167-5273  Vol. 173  Nº 2  2014  págs. 337 - 338
Autores: Orbe, J; Rodríguez, José Antonio; Sanchez, J. A.; et al.
Revista: CARDIOVASCULAR RESEARCH
ISSN 0008-6363  Vol. 103  Nº Supl. 1  2014  págs. P230
Autores: García, Oihane; Carotti, S.; Latasa, María Ujué; et al.
Revista: JOURNAL OF HEPATOLOGY
ISSN 0168-8278  Vol. 60  Nº 1  2014  págs. S87 - S87
Autores: Orbe, J; Rodríguez, José Antonio; Sanchez, J. A.; et al.
Revista: ABSTRACTS OF PAPERS OF THE AMERICAN CHEMICAL SOCIETY
ISSN 0065-7727  Vol. 248  2014 
Autores: Rodríguez, José Antonio, (Autor de correspondencia); Sobrino T.; Orbe, J; et al.
Revista: JOURNAL OF THROMBOSIS AND HAEMOSTASIS
ISSN 1538-7933  Vol. 11  Nº 8  2013  págs. 1464 - 1473
Background: Matrix metalloproteinases (MMPs) mediate tissue injury during stroke but also neurovascular remodeling and we have shown that MMP-10 is involved in atherothrombosis. Objective: The purpose of this study was to examine the relationship between proMMP-10 and clinical outcome, assessing inflammatory and proteolytic markers, in patients with acute ischemic stroke. Methods: We prospectively studied 76 patients with ischemic stroke treated with tPA within the first 3 h from symptom onset, compared with 202 non-tPA-treated ischemic stroke patients and 83 asymptomatic subjects. Stroke severity was assessed with the National Institutes of Health Stroke Scale (NIHSS). Hemorrhagic transformation (HT) and severe brain edema were diagnosed by cranial CT. Good functional outcome was defined as a modified Rankin scale score ¿ 2 at 90 days. Serum levels of MMP-9, proMMP-10, TIMP-1, tumor necrosis factor-¿ (TNF¿), interleukin-6 and cellular fibronectin were measured at admission. The effect of TNF¿ on endothelial proMMP-10 was assessed in vitro. Results: Serum proMMP-10 concentration in ischemic stroke patients, non-treated or treated with t-PA, which was higher than age-matched healthy subjects (P < 0.0001), was independently associated with higher infarct volume, severe brain edema, neurological deterioration and poor functional outcome at 3 months (all P < 0.05), but not with HT. proMMP-10 levels were also independently and positively associated with circulating levels of TNF¿ (P < 0.0001), which induced its endothelial expression in vitro, both mRNA and protein. MMP-9, however, was only associated with HT and severe edema (all P < 0.05). Conclusions: Increased serum proMMP-10 after acute ischemic stroke, associated with TNF¿, is a new marker of brain damage and poor outcome.
Autores: García, Oihane; Carotti, S.; Latasa, María Ujué; et al.
Revista: LIVER INTERNATIONAL
ISSN 1478-3223  Vol. 34  Nº 7  2013  págs. e257 - e270
Background & Aims Upon tissue injury, the liver mounts a potent reparative and regenerative response. A role for proteases, including serine and matrix metalloproteinases ( MMPs), in this process is increasingly recognized. We have evaluated the expression and function of MMP10 (stromelysin-2) in liver wound healing and regeneration. Methods The hepatic expression of MMP10 was examined in two murine models: liver regeneration after two-thirds partial hepatectomy (PH) and bile duct ligation (BDL). MMP10 was detected in liver tissues by qPCR, western blotting and immunohistochemistry. The effect of growth factors and toll-like receptor 4 (TLR4) agonists on MMP10 expression was studied in cultured parenchymal and biliary epithelial cells and macrophages respectively. The role of MMP10 was evaluated by comparing the response of Mmp10+/+ and Mmp10¿/¿ mice to PH and BDL. The intrahepatic turnover of the extracellular matrix proteins fibrin (ogen) and fibronectin was examined. Results MMP10 mRNA was readily induced after PH and BDL. MMP10 protein was detected in hepatocytes, cholangiocytes and macrophages. In cultured liver epithelial cells, MMP10 expression was additively induced by transforming growth factor-ß and epidermal growth factor receptor ligands. TLR4 ligands also stimulated MMP10 expression in macrophages. Lack of MMP10 resulted in increased liver injury upon PH and BDL. Resolution of necrotic areas was impaired, and Mmp10¿/¿ mice showed increased fibrogenesis and defective turnover of fibrin (ogen) and fibronectin. Conclusions MMP10 expression is induced during mouse liver injury and participates in the hepatic wound healing response. The profibrinolytic activity of MMP10 may be essential in this novel hepatoprotective role.
Autores: Hernández, C. ; Orbe, J; Roncal, Carmen; et al.
Revista: THROMBOSIS AND HAEMOSTASIS
ISSN 0340-6245  Vol. 110  Nº 3  2013  págs. 598 - 608
Abstract A prothrombotic state is one of the hallmarks of malignancy and a major contributor to morbidity and mortality in cancer patients.Tissue factor (TF) is often overexpressed in malignancy and is a prime candidate in predicting the hypercoagulable state. Moreover, increased number of TF-exposing microparticles (MPs) in cancer patients may contribute to venous thromboembolism (VTE). We have conducted a prospective cohort study to determine whether elevated TF antigen, TF activity and TF associated to MPs (MPs-TF) are predictive of VTE and mortality in cancer patients. The studied population consisted of 252 cancer patients and 36 healthy controls. TF antigen and activity and MPs-TF were determined by ELISA and chromogenic assays. During a median follow-up of 10 months, 40 thrombotic events were recorded in 34 patients (13.5%), and 73 patients (28.9%) died. TF antigen and activity were significantly higher in patients than in controls (p<0.01) mainly in patients with advanced stages, whereas no differences were observed for TF activity of isolated MPs. We did not find a statistically significant association of TF variables with the risk of VTE. Multivariate analysis adjusting for age, sex, type of cancer and other confounding variables showed that TF activity (p<0.01) and MPs-TF activity (p<0.05) were independently associated with mortality. In conclusion, while TF variables were not associated with future VTE in cancer patients, we found a strong association of TF and MPs-TF activity with mortality, thus suggesting they might be good prognostic markers in cancer patients.
Autores: Lorente, L.; Martin, M.; Plasencia, F.; et al.
Revista: CRITICAL CARE
ISSN 1466-609X  Vol. 17  Nº 3  2013  págs. R94
INTRODUCTION: Previous studies have found higher circulating levels of tissue inhibitor of matrix metalloproteinase (TIMP)-1 in nonsurviving septic patients than in surviving septic patients, and an association between the 372 T/C genetic polymorphism of TIMP-1 and the risk of developing certain diseases. However, the relationship between genetic polymorphisms of TIMP-1, circulating TIMP-1 levels and survival in patients with severe sepsis has not been examined, and this was the objective of the study. METHODS: This multicentre, prospective, observational study was carried out in six Spanish ICUs. We determined the 372 T/C genetic polymorphism of TIMP-1 (rs4898), serum levels of TIMP-1, matrix metalloproteinase (MMP)-9, MMP-10, TNF¿, IL-10 and plasma plasminogen activator inhibitor-1 (PAI-1). Survival at 30 days from ICU admission was the endpoint assessed. The association between continuous variables was carried out using Spearman's rank correlation coefficient or Spearman's rho coefficient. Multivariate logistic regression analysis was applied to determine the association between the 372 T/C genetic polymorphism and survival 30 days from ICU admission. RESULTS: Of 275 patients with severe sepsis, 80 had genotype CC, 55 had genotype CT and 140 had genotype TT of the 372 T/C genetic polymorphism of TIMP-1. Patients with the T allele showed higher serum levels of TIMP-1 than patients without the T allele (P=0.004). Multiple logistic regression analysis showed that the T allele was associated with higher mortality at 30 days (odds ratio=2.08; 95% confidence interval=1.06 to 4.09; P=0.03). Survival analysis showed that patients with the T allele presented lower 30-day survival than patients without the T allele (¿2=5.77; P=0.016). We found an association between TIMP-1 levels and levels of MMP-9 (¿=-0.19; P=0.002), MMP-10 (¿=0.55; P<0.001), TNF¿ (¿=0.56; P<0.001), IL-10 (¿=0.48; P<0.001) and PAI-1 (¿=0.49; P<0.001). CONCLUSION: The novel findings of our study are that septic patients with the T allele in the 372 T/C genetic polymorphism of TIMP-1 showed higher serum TIMP-1 levels and lower survival rate. The determination of the 372 T/C genetic polymorphism of TIMP-1 thus has prognostic implications and could help in the selection of patients who may benefit from modulation of the MMP/TIMP balance.
Autores: García, Oihane; Latasa, María Ujué; et al.
Revista: JOURNAL OF HEPATOLOGY
ISSN 0168-8278  Vol. 58  2013  págs. S126 - S126
Autores: Roncal, Carmen; calvayrac, o; et al.
Revista: Arteriosclerosis, thrombosis and vascular biology (Print)
ISSN 1079-5642  Vol. 32  Nº 6  2012  págs. 1477 - 1487
Autores: Mazo, Manuel María; Gavira, Juan José; Abizanda, Gloria María; et al.
Revista: CELL TRANSPLANTATION
ISSN 0963-6897  Vol. 21  Nº 5  2012  págs. 1023 - 1037
Fresh adipose-derived cells have been shown to be effective in the treatment of acute myocardial infarction (MI), but their role in the chronic setting is unknown. We sought to determine the long-term effect of the adipose derived-stromal vascular fraction (SVF) cell transplantation in a rat model of chronic MI. MI was induced in 82 rats by permanent coronary artery ligation and 5 weeks later rats were allocated to receive an intramyocardial injection of 10(7) GFP-expressing fresh SVF cells or culture media as control. Heart function and tissue metabolism were determined by echocardiography and F-18-FDG-microPET, respectively, and histological studies were performed for up to 3 months after transplantation. SVF induced a statistically significant long-lasting (3 months) improvement in cardiac function and tissue metabolism that was associated with increased revascularization and positive heart remodeling, with a significantly smaller infarct size, thicker infarct wall, lower scar fibrosis, and lower cardiac hypertrophy. Importantly, injected cells engrafted and were detected in the treated hearts for at least 3 months, directly contributing to the vasculature and myofibroblasts and at negligible levels to cardiomyocytes. Furthermore, SVF release of angiogenic (VEGF and HGF) and proinflammatory (MCP-1) cytokines, as well as TIMP1 and TIMP4, was demonstrated in vitro and in vivo, strongly suggesting that they have a trophic effect. These results show the potential of SVF to contribute to the regeneration of ischemic tissue and to provide a long-term functional benefit in a rat model of chronic MI, by both direct and indirect mechanisms.
Autores: Orbe, J; Barrenetxe, J; Rodríguez, José Antonio; et al.
Revista: Circulation
ISSN 0009-7322  Vol. 124  Nº 25  2011  págs. 2909 - 2919
BACKGROUND: The fibrinolytic and matrix metalloproteinase (MMP) systems cooperate in thrombus dissolution and extracellular matrix proteolysis. The plasminogen/plasmin system activates MMPs, and some MMPs have been involved in the dissolution of fibrin by targeting fibrin(ogen) directly or by collaborating with plasmin. MMP-10 has been implicated in inflammatory/thrombotic processes and vascular integrity, but whether MMP-10 could have a profibrinolytic effect and represent a promising thrombolytic agent is unknown. METHODS AND RESULTS: The effect of MMP-10 on fibrinolysis was studied in vitro and in vivo, in MMP-10-null mice (Mmp10(-/-)), with the use of 2 different murine models of arterial thrombosis: laser-induced carotid injury and ischemic stroke. In vitro, we showed that MMP-10 was capable of enhancing tissue plasminogen activator-induced fibrinolysis via a thrombin-activatable fibrinolysis inhibitor inactivation-mediated mechanism. In vivo, delayed fibrinolysis observed after photochemical carotid injury in Mmp10(-/-) mice was reversed by active recombinant human MMP-10. In a thrombin-induced stroke model, the reperfusion and the infarct size in sham or tissue plasminogen activator-treated animals were severely impaired in Mmp10(-/-) mice. In this model, administration of active MMP-10 to wild-type animals significantly reduced blood reperfusion time and infarct size to the same extent as tissue plasminogen activator and was associated with shorter bleeding time and no intracranial hemorrhage. This effect was not observed in thrombin-activatable fibrinolysis inhibitor-deficient mice, suggesting thrombin-activatable fibrinolysis inhibitor inactivation as one of the mechanisms involved in the MMP-10 profibrinolytic effect. CONCLUSIONS: A novel profibrinolytic role for MMP-10 in experimental ischemic stroke is described, opening new pathways for innovative fibrinolytic strategies in arterial thrombosis.
Autores: Lorente, L; Martin, M; Varo, N; et al.
Revista: Critical care medicine
ISSN 0090-3493  Vol. 15  Nº 2  2011  págs. 1 - 8
Autores: Calvayrac, O; Rodriguez-Calvo, R; Alonso, J; et al.
Revista: ARTERIOSCLEROSIS THROMBOSIS AND VASCULAR BIOLOGY
ISSN 1079-5642  Vol. 31  Nº 11  2011  págs. 2733 - 2741
Autores: Coll, B.; Rodríguez, José Antonio; Craver, L.; et al.
Revista: KIDNEY INTERNATIONAL
ISSN 0085-2538  Vol. 78  Nº 12  2010  págs. 1275 - 1280
Cardiovascular disease is the leading cause of mortality in chronic kidney disease (CKD). As matrix metalloproteinases have a major role in atherosclerosis, we hypothesized that alterations in metalloproteinases-8, -10 and their tissue inhibitor-1 can be associated with the severity of atherosclerosis in patients with kidney disease. This was evaluated in a cross-sectional, observational study of 111 patients with stages I-V kidney disease, 217 patients on dialysis and 50 healthy controls. The severity of atherosclerosis was estimated with the atherosclerosis score (AS), combining the results of ankle-brachial index and carotid ultrasound. Serum levels of the two metalloproteinases and tissue inhibitor-1 were measured by enzyme-linked immunosorbent assay and were significantly increased in patients with kidney disease compared with the healthy controls, and higher in patients on dialysis than in earlier stages of CKD. The severity of the AS was also more prevalent in the dialysis group, in which serum levels of both metalloproteinases and tissue inhibitor-1 were significantly higher. After multivariate analysis, metalloproteinase-10, dialysis, C-reactive protein, age, and male gender were associated with increased risk of atherosclerosis. Thus, patients with CKD exhibit elevated levels of circulating metalloproteinase-10, and this was independently associated with the severity of atherosclerosis and may represent a new biomarker of atherosclerotic diseases.
Autores: Páramo, José Antonio; Orbe, J; Rodríguez, José Antonio;
Revista: HAEMATOLOGICA
ISSN 1138-0381  Vol. 95  Nº Extra 1  2010  págs. 445 - 449