A Semi-Automated and Reproducible Biological-Based Method to Quantify Calcium Deposition In Vitro

Journal of Visualized Experiments

Abstract

Vascular calcification involves a series of degenerative pathologies, including inflammation, changes to cellular phenotype, cell death, and the absence of calcification inhibitors, that concomitantly lead to a loss of vessel elasticity and function. Vascular calcification is an important contributor to morbidity and mortality in many pathologies, including chronic kidney disease, diabetes mellitus, and atherosclerosis. Current research models to study vascular calcification are limited and are only viable at the late stages of calcification development in vivo. In vitro tools for studying vascular calcification use end-point measurements, increasing the demands on biological material and risking the introduction of variability to research studies. We demonstrate the application of a novel fluorescently labeled probe that binds to in vitro calcification development on human vascular smooth muscle cells and determines the real-time development of in vitro calcification. In this protocol, we describe the application of our newly developed calcification assay, a novel tool in disease modeling that has potential translational applications. We envisage this assay to be relevant in a broader spectrum of mineral deposition research, including applications in bone, cartilage, or dental research.

Authors 7

1. Armand M. G. Jaminon

   Maastricht University

   Affiliation as printed

   Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University

2. Nikolas Rapp

   Maastricht University

   Affiliation as printed

   Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University

3. Asim Cengiz Akbulut

   Maastricht University

   Affiliation as printed

   Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University

4. Robert Dzhanaev Aachen BioInterface Group

   RWTH Aachen University

   Affiliation as printed

   Helmholtz Institute for Biomedical Engineering, Biointerface Group, RWTH Aachen University

5. Chris P.M. Reutelingsperger

   Maastricht University

   Affiliation as printed

   Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University

6. Willi Jahnen‐Dechent Aachen BioInterface Group

   RWTH Aachen University

   Affiliation as printed

   Helmholtz Institute for Biomedical Engineering, Biointerface Group, RWTH Aachen University; willi.jahnen@rwth-aachen.de

   Helmholtz Institute for Biomedical Engineering, Biointerface Group, RWTH Aachen University

7. Leon J. Schurgers Aachen Institute of Experimental Medicine and Systems Biology

   Maastricht University · RWTH Aachen University

   Affiliation as printed

   Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University; Institute of Experimental Medicine and Systems Biology, RWTH Aachen University; l.schurgers@maastrichtuniversity.nl

   Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University

   Institute of Experimental Medicine and Systems Biology, RWTH Aachen University