Time-resolved metabolite and lipid profiling depicts macrophage continuum with apoptotic and ferroptotic heterogeneity along foam cell formation

Atherosclerosis is a chronic inflammatory disease driven by the maladaptive lipid metabolism and inflammatory response of macrophages. In the progression of atherosclerosis, macrophages work as a lipid scavenger and immune regulator by dynamic lipid and metabolic reprogramming in response to the microenvironment. Nevertheless, we are only starting to characterize the function of the heterogeneity in metabolite and lipid compositions. Herein, we developed a four-dimensional micro-manipulation platform for single-cell sample preparation, and observed the macrophage heterogeneity along foam cell formation via a single-cell time-resolved metabolite and lipid profiling. By integrating bulk metabolomic and lipidomic data with single-cell metabolomic and lipidomic data, we were surprised to uncover that macrophages have different fate outcomes during the late foam cell formation process. Some foam cells were more prone to apoptosis, while others are more susceptible to ferroptosis, suggesting heterogeneity in cell fate determination could have important implications for understanding atherosclerosis progression. Single-cell transcriptome sequencing was further performed on late-stage foam cells derived from both THP-1 and peripheral blood mononuclear cells, jointly confirming the divergent cell fates toward apoptosis or ferroptosis. Lastly, we validated the heterogeneity of late-stage foam cell differentiation through a series of in vitro experiments, including caspase activity assay and lipid peroxidation assay. In summary, single-cell multi-omics depict the molecular choreography that dictates the cell death in late atherosclerosis.

Keywords: Single-cell analysis; Metabolomics; Lipidomics; Foam cell; Ferroptosis.