Quantification of sterol-specific response in human macrophages using automated imaged-based analysis

Abstract

BackgroundThe transformation of normal macrophage cells into lipid-laden foam cells is an important step in the progression of atherosclerosis. One major contributor to foam cell formation in vivo is the intracellular accumulation of cholesterol.MethodsHere, we report the effects of various combinations of low-density lipoprotein, sterols, lipids and other factors on human macrophages, using an automated image analysis program to quantitatively compare single cell properties, such as cell size and lipid content, in different conditions.ResultsWe observed that the addition of cholesterol caused an increase in average cell lipid content across a range of conditions. All of the sterol-lipid mixtures examined were capable of inducing increases in average cell lipid content, with variations in the distribution of the response, in cytotoxicity and in how the sterol-lipid combination interacted with other activating factors. For example, cholesterol and lipopolysaccharide acted synergistically to increase cell lipid content while also increasing cell survival compared with the addition of lipopolysaccharide alone. Additionally, ergosterol and cholesteryl hemisuccinate caused similar increases in lipid content but also exhibited considerably greater cytotoxicity than cholesterol.ConclusionsThe use of automated image analysis enables us to assess not only changes in average cell size and content, but also to rapidly and automatically compare population distributions based on simple fluorescence images. Our observations add to increasing understanding of the complex and multifactorial nature of foam-cell formation and provide a novel approach to assessing the heterogeneity of macrophage response to a variety of factors.Electronic supplementary materialThe online version of this article (10.1186/s12944-017-0629-9) contains supplementary material, which is available to authorized users.

A Genome-Wide Screen of Deletion Mutants in the Filamentous Saccharomyces cerevisiae Background Identifies Ergosterol as a Direct Trigger of Macrophage Pyroptosis

“…Thus, if our model is correct, the effects of Upc2 mutants on pyroptosis are readily explained by alterations in the levels of overall cellular ergosterol. Second, a head-to-head comparison of the toxicity of sterols toward macrophages indicates that ergosterol is more toxic to macrophages than cholesterol ( 46 ). Although the mechanism of this toxicity was not characterized further, it is interesting to consider the fact that ergosterol does not activate liver X receptor (LXR), a key regulator of sterol efflux in liver and macrophage cells ( 47 ).…”

Biomimetic 3D Models for Investigating the Role of Monocytes and Macrophages in Atherosclerosis

“…Furthermore, these platforms must be thoroughly characterized, and a profound understanding of individual contributions of the cells involved in atherosclerosis development must be reached before venturing to co-culture models. Most atherosclerosis studies are carried on 2D tissue culture plates [16]. While meaningful contributions have been made towards 3D modeling atherosclerosis in vitro, most studies have focused on co-culture of endothelial cells (ECs), smooth muscle cells (SMCs), and monocytes, in either physiological [18] or synthetic [20] scaffolds, while detailed individual contributions of macrophages still remain unclear.…”

“…Animal models have provided abundant information for atherosclerosis research, however, to understand atherosclerosis healing mechanistically, relevant in vitro models are a must. Models of atherosclerosis have progressed from simple 2D culture to 3D multi-cellular cultures [ 16 , 17 ], the latter a better representation considering the dimensionality of the disease and anatomical features involved. To mimic in vivo-like conditions, in vitro models have encapsulated the relevant cells of interest within naturally-derived and polymer-based hydrogels.…”

In Vitro Immune Organs-on-Chip for Drug Development: A Review

“…These technological processes have enabled the development of fine structures that closely resemble those in vivo [19,20]. Furthermore, powerful computational techniques that allow the quantification of cellular behavior even at the single cell level are now being available and can be combined with microfluidic technology to provide a comprehensive description of cellular behavior under various stimuli [21,22,23].…”