A novel truncated form of apolipoprotein A-I transported by dense LDL is increased in diabetic patients

Diabetic (DM) patients have exacerbated atherosclerosis and high CVD burden. Changes in lipid metabolism, lipoprotein structure, and dysfunctional HDL are characteristics of diabetes. Our aim was to investigate whether serum ApoA-I, the main protein in HDL, was biochemically modified in DM patients. By using proteomic technologies, we have identified a 26 kDa ApoA-I form in serum. MS analysis revealed this 26 kDa form as a novel truncated variant lacking amino acids 1-38, ApoA-IΔ(1-38). DM patients show a 2-fold increase in ApoA-IΔ(1-38) over nondiabetic individuals. ApoA-IΔ(1-38) is found in LDL, but not in VLDL or HDL, with an increase in LDL3 and LDL4 subfractions. To identify candidate mechanisms of ApoA-I truncation, we investigated potentially involved enzymes by in silico data mining, and tested the most probable molecule in an established animal model of diabetes. We have found increased hepatic cathepsin D activity as one of the potential proteases involved in ApoA-I truncation. Cathepsin D-cleaved ApoA-I exhibited increased LDL binding affinity and decreased antioxidant activity against LDL oxidation. In conclusion, we show for the first time: a) presence of a novel truncated ApoA-I form, ApoA-IΔ(1-38), in human serum; b) ApoA-IΔ(1-38) is transported by LDL; c) ApoA-IΔ(1-38) is increased in dense LDL fractions of DM patients; and d) cathepsin D-ApoA-I truncation may lead to ApoA-IΔ(1-38) binding to LDLs, increasing their susceptibility to oxidation and contributing to the high cardiovascular risk of DM patients. This research was originally published in Lipid Research. A novel truncated form of apolipoprotein A-I transported by dense LDL is increased in diabetic patients. Journal of Lipid Research. 2015. 56: 1762-1773 © the American Society for Biochemistry and Molecular Biology.Peer Reviewe

Identificación de nuevos biomarcadores en los síndromes coronarios agudos

[spa] La aterosclerosis, engrosamiento de la pared arterial, puede progresar gradualmente hasta complicarse con la aparición de un trombo ocasionando una obstrucción brusca de una arteria con obliteración del flujo en el órgano irrigado por dicha arteria. Según su localización, esta oclusión puede dar lugar a un accidente cerebrovascular (CVA), una obstrucción arterial periférica o a un síndrome coronario agudo (ACS). Dentro de los ACS podemos encontrar la angina inestable, el infarto agudo de miocardio con y sin elevación de ST y la muerte súbita. Estas manifestaciones agudas de la enfermedad cardíaca comparten un mismo fenómeno patofisiológico: la aterotrombosis coronaria y resultan de una isquemia aguda del miocardio. Aunque se han producido grandes avances en el tratamiento de esta enfermedad, la medicina actual no es capaz de predecir el riesgo de sufrir patología cardiovascular. Pese a la gran influencia de los factores de riesgo clásicos tales como la dislipemia, la diabetes, la hipertensión y la obesidad, una proporción importante de los eventos ocurren en individuos que no presentan ninguno de estos factores. De ello deriva la actual necesidad de identificar nuevos biomarcadores que mejoren la predicción global de riesgo de enfermedad cardiovascular. En los últimos años la proteómica se ha convertido en una estrategia básica para el estudio del perfil proteico y de asociaciones proteicas complejas en una muestra biológica. Mediante el uso de técnicas proteómicas se pueden determinar modificaciones en la estructura de una proteína, así como sus niveles de expresión y la presencia de modificaciones post-traduccionales, que pueden estar asociadas a una patología determinada y tener valor diagnóstico, pronóstico y terapéutico. Se han propuesto muchas moléculas como marcadores de la aterotrombosis, pero los resultados no son consistentes y la mayoría de ellos no ha llegado a utilizarse en la práctica clínica. Por eso cada vez hay una mayor necesidad de integrar las técnicas proteómicas en el descubrimiento de nuevos biomarcadores lo que permitirá conocer nuevos mediadores y vías patofisiológicas sin una asociación previa a las patologías cardiovasculares. En este trabajo se han aplicado técnicas de proteómica diferencial para el estudio de los cambios que se dan a nivel sérico en la fase aguda tras un infarto agudo de miocardio (AMI) de nueva presentación. Mediante la comparación del perfil proteómico de pacientes AMI e individuos sanos se han detectado importantes cambios en el patrón de distribución de dos grupos de proteínas: relacionadas con el metabolismo de las lipoproteínas de alta densidad (HDL) y relacionadas con el metabolismo del retinol, vía no asociada hasta el momento con el AMI. Dentro de las proteínas del metabolismo de las HDL encontramos cambios en la fase aguda post-AMI en la apolipoproteína J y la transtiretina (TTR). Ésta última también muestra cambios en situaciones de alto riesgo cardiovascular como la hipercolesterolemia familiar. A su vez, también se han detectado cambios en la proteína mayoritaria de las HDL, la apolipoproteína A-I, pero en este caso en una fase más tardía post-AMI y en pacientes diabéticos. Las proteínas relacionadas con el metabolismo del retinol que muestran un perfil diferencial son la TTR y la proteína plasmática de unión a retinol (RBP4). En una segunda parte de este trabajo se han estudiado los cambios proteómicos que se dan a nivel de tejido cardíaco en un modelo experimental porcino de AMI y se ha encontrado por primera vez una asociación de la vía de señalización del receptor del hidrocarburo de arilo con el daño producido por la reperfusión posterior a un AMI.. Además se ha visto que la aplicación de un procedimiento cardioprotector como es el post-condicionamiento isquémico atenua la activación de dicha vía inducida por la reperfusión.[eng] Cardiovascular diseases (CVD) are the leading cause of morbidity and mortality. Atherosclerosis, the underlying mechanism of cardiovascular diseases, progresses gradually until the blood flow is compromissed leading to the precipitaton of an acute ischemic event such as acute coronary syndrome (ACS) or ictus. Identifying subjects at risk of developing an acute ischemic event remains one of the great challenges of cardiovascular medicine. Classical approaches, such as the presence of cardiovascular risk factors, are unable to accurately predict cardiovascular events. During the last years several studies have been focussed on the search for new plasma biomarkers of acute ischemic events. Although many potential molecules have been described, the results have not been consistent enough and most of them are not used in clinical practice. Until now, the only group of accepted biomarkers for the diagnosis of acute myocardial ischemic events are troponins. Nevertheless, these structural proteins are released to the circulation as a consequence of an irreversible injury of the myocardium. Therefore there is still a need for the identification of new biomarkers that will allow the early detection of an ischemic event before the irreversible necrosis of the myocardium occurs. Proteomic technologies allow the identification of molecules related to new pathways that together with traditional markers could act as a multibiomarker for diagnosis, prognosis and treatment, and therefore become a key tool for the development of new approaches in the prevention of cardiovascular diseases. In this study by applying proteomic technologies such as bi-dimensional electrophoresis we have compared the serum proteomic profile of patients with an acute new-onset myocardial infarction (AMI) to that of healthy individuals and found important changes in two main group of proteins: HDL-related proteins (apolipoproteins J and A-I, and TTR) and retinol metabolism associated proteins (TTR and RBP4). In a second part of the work by the study of proteomic changes that occur in the myocardial tissue after an AMI in a swine experimental model we have found, for the first time, an association of the aryl hidrocarbon receptor signalling pathway with the reperfusion injury after an AMI. Moreover, we have found an attenuation of the same pathway in response to a cardioprotective approach such as ischemic post-conditioning

Methods and kits for the diagnosis and risk stratification of patients with ischemia

The invention relates to methods for the diagnosis of ischemia or ischemic tissue damage, methods for predicting the progression of ischemia in a patient having suffered an ischemic event, for determining the prognosis of a patient having suffered an ischemic event and for determining the risk that a patient suffering from stable coronary disease suffers a recurrent ischemic event based on the detection of the levels of glycosylated Apo J. The invention relates as well to a method for the determination of glycosylated Apo J in a sample.Institut Catalá de Ciències Cardiovasculars, Consejo Superior de Investigaciones Científicas (España)A1 Solicitud de patente con informe sobre el estado de la técnic

Uso de las isoformas de Apo J como biomarcadores de lesión tisular

La invención se refiere al uso de las formas glicosiladas de la Apolipoproteína J como marcadores de daño tisular, más concretamente producido de infarto agudo de miocardio, así como a un método de diagnóstico de dicho daño y a un kit que comprende los elementos necesarios para llevar a cabo dicho diagnóstico. En la presente invención se demuestra que las diversas formas glicosiladas varían su expresión si existe daño tisular lo que permite utilizarlas como marcadores en caso de existir este daño.Peer reviewedConsejo Superior de Investigaciones Científicas (España), Institut Catalá de Ciencies CardiovascularsA1 Solicitud de patente con informe sobre el estado de la técnic

Uso de las iso formas de apo j como biomarcadores de lesión tisular

[EN] The present invention relates to use of the glycosylated fonns of apolipoprotein J as markers of tissue damage and of the acute inflammation said damage entails, more particularly the product of acute myocardial infarction, together with a method of diagnosis of said damage and/or inflammation and a kit comprising the elements necessary to effect said diagnosis. In the present invention it is demonstrated that the diverse glycosylated fonns vary their expression should there exist tissue damage and/or acute inflammation, pennitting the utilisation thereof as markers in the case ofthe existence of damage and/or inflammation.[ES] La invención se refiere al uso de las fonnas glicosiladas de la Apolipoproteina J como marcadores de daño tisular así como de la inflamación aguda que conlleva dicho daño, más concretamente producido de infarto agudo de miocardio, así como a un método de diagnóstico de dicho daño y/o inflamación y a un kit que comprende los elementos necesarios para llevar a cabo dicho diagnóstico. En la presente invención se demuestra que las diversas fonnas glicosiladas varían su expresión si existe daño tisular y/o inflamación aguda lo que pennite utilizarlas como marcadores en caso de existir este daño y/o inflamación.Peer reviewedConsejo Superior de Investigaciones Científicas (España), Institut De Ciencies CardiovascularsA1 Solicitud de patente con informe sobre el estado de la técnic

Methods and kits for the diagnosis and risk stratification of patients with ischemia

The invention relates to methods for the diagnosis of ischemia or ischemic tissue damage, methods for predicting the progression of ischemia in a patient having suffered an ischemic event, for determining the prognosis of a patient having suffered an ischemic event and for determining the risk that a patient suffering from stable coronary disease suffers a recurrent ischemic event based on the detection of the levels of glycosylated Apo J. The invention relates as well to a method for the determination of glycosylated Apo J in a sample.Institut Catalá de Ciències Cardiovasculars, Consejo Superior de Investigaciones Científicas (España)A1 Solicitud de patente con informe sobre el estado de la técnic

Methods and kits for the diagnosis and risk stratification of patients with ischemia

The invention relates to methods for the diagnosis of ischemia or ischemic tissue damage, methods for predicting the progression of ischemia in a patient having suffered an ischemic event, for determining the prognosis of a patient having suffered an ischemic event and for determining the risk that a patient suffering from stable coronary disease suffers a recurrent ischemic event based on the detection of the levels of glycosylated Apo J. The invention relates as well to a method for the determination of glycosylated Apo J in a sample.Peer reviewedInstitut Catalá de Ciències Cardiovasculars, Consejo Superior de Investigaciones Científicas (España)E Solicitud de patente europe

Antibodies specific for glycosylated apoj and uses thereof

The invention relates to new antibodies against specific glycosylation sites within the ApoJ protein as well as their application thereof in the diagnosis and prognosis of ischemia and the determination of the risk of a recurrent ischemic event.Peer reviewedGlycardial Diagnostics S.L., Fundació Institut de Recerca de L'Hospital de la Santa Creu i Santa Pau, Consejo Superior de Investigaciones Científicas (España)A1 Solicitud de patente con informe sobre el estado de la técnic

Detrimental effect of hypercholesterolemia on high-density lipoprotein particle remodeling in pigs

[Background] Beneficial effects of high-density lipoproteins (HDL) seem altered in patients with symptomatic cardiovascular disease. We recently demonstrated in a swine model of ischemia-reperfusion (IR) that hypercholesterolemia abolishes HDL-related cardioprotection. [Objectives] This study sought to investigate, using the same animal model, whether the reported impairment of HDL cardioprotective function was associated with alterations in HDL remodeling and functionality. [Methods] Pigs were fed a normocholesterolemic (NC) or hypercholesterolemic (HL) diet for 10 days, reaching non-HDL cholesterol concentrations of 38.2 ± 3.5 mg/dl and 218.6 ± 27.6 mg/dl, respectively (p < 0.0001). HDLs were isolated, and lipidomics and differential proteomics tests were performed to determine HDL molecular changes. HDL functionality and particle size were determined. [Results] Using principal component analysis, we identified 255 molecular lipid species differentially clustered in NC-HDL and HL-HDL. Ninety lipid metabolites were differentially expressed, and 50 showed at least 1.5-fold variation (false discovery rate adjustment q value <0.05). HL-HDLs presented a core enriched in cholesteryl esters and a surface depleted of phosphatidylcholine species containing polyunsaturated and long-chain fatty acids, indicating the presence of mature HDL particles with low surface fluidity. Hypercholesterolemia induced an important change in HDL-transported proteins (576 spots in HL-HDL vs. 621 spots in NC-HDL). HL-HDLs showed a reduced content of lipocalin retinol binding protein 4 and apolipoprotein M and in the retinoic acid-transporter cellular retinoic acid binding protein 1 (p < 0.05 vs. NC-HDL). No changes were observed in apolipoprotein A-I content and profile. Functionally, HL-HDL showed lower antioxidant activity (−35%) and a reduced capacity to efflux cholesterol (−60%) compared to NC-HDL (p < 0.05). Hypercholesterolemia induced larger HDL particles. [Conclusions] We demonstrate that hypercholesterolemia induces HDL lipidomic changes, losing phosphatidylcholine-lipid species and gaining cholesteryl esters, and proteomic changes, with losses in cardioprotective proteins. These remodeling changes shifted HDL particles toward a dysfunctional state.The authors thank the Fundación Investigación Cardiovascular and the Fundación Jesús Serra for their continuous support and CERCA Programme/Generalitat de Catalunya.Peer reviewe