MODELING OF ENDOTHELIAL DYSFUNCTION AND SEARCH FOR ITS CIRCULATING BIOMARKERS
Authors: Shishkova Daria K. , Frolov Alexey V. , Markova Victoria E. , Markova Yulia O. , Kanonykina Anastasia Yu. , Lazebnaya Anastasia I. , Matveeva Vera G. , Torgunakova Evgenia A. , Kutikhin Anton G. 
Published: February 2025

Highlights

Modeling proinflammatory endothelial dysfunction in vitro is achieved by inducing mitochondrial-oxidative stress (alkylating agent mitomycin C), lysosomal-calcium stress (calciprotein particles), cytokine stress (lipopolysaccharide addition), or metabolic stress (palmitic acid addition). In vivo, this can be modeled by comparing aged and young laboratory rodents (specifically hyperlipidemic mice).

To model vasospastic endothelial dysfunction in vitro, it is advisable to use inhibitors of endothelial NO synthase. For in vivo modeling, rats with stress-induced hereditary arterial hypertension (SIHAH) should be used, comparing them with normotensive Wistar rats in experiments.

For modeling prothrombotic endothelial dysfunction, it is appropriate to use the S1 subunit of the Spike protein of the SARS-CoV-2 virus or its receptor-binding domain (RBD), as well as transgenic K18-hACE2 mice expressing the human ACE2 receptor.

 

Abstract

Despite its high clinical relevance to a range of acute (COVID-19, sepsis, multiple organ failure) and chronic (arterial hypertension, frailty syndrome, deep vein thrombosis) conditions and its direct impact on the development of adverse outcomes, the concept of endothelial dysfunction remains rather vague. Considering the various triggers, development mechanisms, molecular markers, and pathological consequences, it is reasonable to classify endothelial dysfunction as a typical pathological process into three types, determined by the leading pathogenetic factor: proinflammatory, vasospastic, and prothrombotic. Except for the hemostatically active high-molecular-weight multimers of von Willebrand factor accompanying the development of prothrombotic endothelial dysfunction in COVID-19, reliable and clinically applicable circulating markers of endothelial dysfunction remain unknown, significantly complicating the study of therapeutic correction methods. This review discusses approaches to modeling these types of endothelial dysfunction in cell cultures and animal models, as well as clinical scenarios for verifying potential markers identified in experiments. For modeling proinflammatory endothelial dysfunction in vitro, it is optimal to use mitomycin C, calciprotein particles, lipopolysaccharide, and palmitic acid; for vasospastic dysfunction – endothelial NO synthase inhibitors; for prothrombotic dysfunction – the S1 subunit of the SARS-CoV-2 Spike protein or its receptor-binding domain. For modeling proinflammatory endothelial dysfunction in vivo, mitomycin C is proposed; for vasospastic dysfunction – rats with hereditary baseline and stress-induced arterial hypertension; for prothrombotic dysfunction – intravenous administration of the S1 subunit or its receptor-binding domain to transgenic mice expressing the human ACE2 receptor (K18-hACE2). The systematic search for markers of endothelial dysfunction is based on step-by-step mass spectrometric profiling and dot-blot profiling (or multiplex immunofluorescent assay based on magnetic microspheres) of the cellular secretome (in serum-free culture medium) and blood serum (after fractionation and removal of high-molecular-weight proteins and supramolecular complexes), followed by verification of the selected markers using enzyme-linked immunosorbent assay.