RESUMEN
An association between high serum calcium/phosphate and cardiovascular events or death is well-established. However, a mechanistic explanation of this correlation is lacking. Here, we examined the role of calciprotein particles (CPPs), nanoscale bodies forming in the human blood upon its supersaturation with calcium and phosphate, in cardiovascular disease. The serum of patients with coronary artery disease or cerebrovascular disease displayed an increased propensity to form CPPs in combination with elevated ionised calcium as well as reduced albumin levels, altogether indicative of reduced Ca2+-binding capacity. Intravenous administration of CPPs to normolipidemic and normotensive Wistar rats provoked intimal hyperplasia and adventitial/perivascular inflammation in both balloon-injured and intact aortas in the absence of other cardiovascular risk factors. Upon the addition to primary human arterial endothelial cells, CPPs induced lysosome-dependent cell death, promoted the release of pro-inflammatory cytokines, stimulated leukocyte adhesion, and triggered endothelial-to-mesenchymal transition. We concluded that CPPs, which are formed in the blood as a result of altered mineral homeostasis, cause endothelial dysfunction and vascular inflammation, thereby contributing to the development of cardiovascular disease.
Asunto(s)
Angina de Pecho/fisiopatología , Isquemia Encefálica/fisiopatología , Cloruro de Calcio/sangre , Enfermedad de la Arteria Coronaria/fisiopatología , Células Endoteliales/patología , Infarto del Miocardio/fisiopatología , Fosfatos/sangre , Angina de Pecho/sangre , Angina de Pecho/genética , Animales , Aorta/metabolismo , Aorta/patología , Isquemia Encefálica/sangre , Isquemia Encefálica/genética , Cloruro de Calcio/química , Estudios de Casos y Controles , Muerte Celular , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/genética , Células Endoteliales/metabolismo , Transición Epitelial-Mesenquimal , Floculación , Regulación de la Expresión Génica , Humanos , Inflamación , Molécula 1 de Adhesión Intercelular/genética , Molécula 1 de Adhesión Intercelular/metabolismo , Leucocitos/metabolismo , Leucocitos/patología , Lisosomas/metabolismo , Lisosomas/patología , Masculino , Infarto del Miocardio/sangre , Infarto del Miocardio/genética , Fosfatos/química , Cultivo Primario de Células , Ratas , Ratas Wistar , Factores de Transcripción de la Familia Snail/genética , Factores de Transcripción de la Familia Snail/metabolismo , Túnica Íntima/metabolismo , Túnica Íntima/patología , Molécula 1 de Adhesión Celular Vascular/genética , Molécula 1 de Adhesión Celular Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Currently, an ultrastructural analysis of cardiovascular tissues is significantly complicated. Routine histopathological examinations and immunohistochemical staining suffer from a relatively low resolution of light microscopy, whereas the fluorescence imaging of plaques and bioprosthetic heart valves yields considerable background noise from the convoluted extracellular matrix that often results in a low signal-to-noise ratio. Besides, the sectioning of calcified or stent-expanded blood vessels or mineralised heart valves leads to a critical loss of their integrity, demanding other methods to be developed. Here, we designed a conceptually novel approach that combines conventional formalin fixation, sequential incubation in heavy metal solutions (osmium tetroxide, uranyl acetate or lanthanides, and lead citrate), and the embedding of the whole specimen into epoxy resin to retain its integrity while accessing the region of interest by grinding and polishing. Upon carbon sputtering, the sample is visualised by means of backscattered scanning electron microscopy. The technique fully preserves calcified and stent-expanded tissues, permits a detailed analysis of vascular and valvular composition and architecture, enables discrimination between multiple cell types (including endothelial cells, vascular smooth muscle cells, fibroblasts, adipocytes, mast cells, foam cells, foreign-body giant cells, canonical macrophages, neutrophils, and lymphocytes) and microvascular identities (arterioles, venules, and capillaries), and gives a technical possibility for quantitating the number, area, and density of the blood vessels. Hence, we suggest that our approach is capable of providing a pathophysiological insight into cardiovascular disease development. The protocol does not require specific expertise and can be employed in virtually any laboratory that has a scanning electron microscope.
RESUMEN
Calcium phosphate bions (CPB) are biomimetic mineralo-organic nanoparticles which represent a physiological mechanism regulating the function, transport and disposal of calcium and phosphorus in the human body. We hypothesised that CPB may be pathogenic entities and even a cause of cardiovascular calcification. Here we revealed that CPB isolated from calcified atherosclerotic plaques and artificially synthesised CPB are morphologically and chemically indistinguishable entities. Their formation is accelerated along with the increase in calcium salts-phosphates/serum concentration ratio. Experiments in vitro and in vivo showed that pathogenic effects of CPB are defined by apoptosis-mediated endothelial toxicity but not by direct tissue calcification or functional changes in anti-calcification proteins. Since the factors underlying the formation of CPB and their pathogenic mechanism closely resemble those responsible for atherosclerosis development, further research in this direction may help us to uncover triggers of this disease.