Paper-based 3D cell culture platform for cholesterol efflux capacity assay

Elevated high-density lipoprotein (HDL) levels do not correlate with reduced cardiovascular disease risk, suggesting that measuring HDL levels is an inadequate biomarker. Consequently, cholesterol efflux capacity (CEC), also referred to as HDL function, has emerged as a more relevant indicator of atherosclerosis risk. Most conventional assays utilize two-dimensional (2D) cell culture, which does not accurately represent the in vivo microenvironment. In this study, a paper membrane scaffold was developed to form foam cells by culturing THP-1 cells on Whatman No. 1 paper. Differentiation of monocytes into macrophages, which serve as cholesterol donors, was achieved by treating the cells with 100 nM phorbol 12-myristate 13-acetate (PMA). The macrophages were treated with 50 mM BODIPY-cholesterol to induce foam cell formation. Cell adhesion between the cells and the fibrous paper was examined using environmental scanning electron microscopy (ESEM). CD marker expression in monocytes and macrophages was analyzed using fluorescence imaging. CEC was calculated as the percentage of BODIPY-cholesterol released relative to total cholesterol, based on fluorescence intensity (Ex 482 nm, Em 522 nm). The CEC values from 38 volunteers did not differ significantly from those obtained with the commercial cell-based method ( p < 0.0001). This paper-based THP-1-derived macrophage platform enables foam cell formation and CEC measurement.