Co-culture of primary human endothelial and vascular smooth muscle cells sensitized endothelial cells to obesity-associated metabolic and humoral stress, doubling the number of regulated genes.
Co-culture of endothelial and vascular smooth muscle cells sensitizes endothelial cells to obesity-related stressors, demonstrating that cellular crosstalk is crucial for stress-induced phenotypical changes.
Abstract Background Obesity is associated with type 2 diabetes (DMT2), subsequently leading to vascular dysfunction and finally renal end-organ damage. Previous in vivo studies in animal models suggest that vascular smooth muscle cells (VSMC) and endothelial cells (EC) interact during obesity/DMT2-induced changes in vascular phenotype. Purpose We investigated the possible importance of EC-VSMC-crosstalk for obesity/DMT2-associated metabolic and humoral stress in primary human EC and VSMC. Our focus was on the early-phase impact of obesity-associated stressors. Methods The role of EC-VSMC-crosstalk during exposure to obesity/DMT2-associated stress factors was investigated in primary human EC and VSMC, comparing cells in monoculture with cells kept in bilayer co-culture. To do so, cells were exposed for 48h to a combination of metabolic and humoral stressors (high glucose concentration, addition of free fatty acids, angiotensin II and norepinephrine), which reflects conditions of obese patients, or kept under control conditions. Various parameters (e.g. viability, proliferation, secretion of inflammation markers) as well as transcriptome changes were then assessed. For co-cultivated VSMC and EC, stressors-induced changes in their proteome and secretome were also measured. Omics results were integrated to identify putative mediators of the VSMC-EC crosstalk. Results Already under control conditions, vascular cells kept in co-culture differed from cells in monoculture regarding extracellular milieu composition and mitochondrial function, as well as gene expression, with hints for culture condition-dependent regulation of their cell cycle and extracellular matrix production. Additionally, obesity-associated stressors induced changes in gene expression of different magnitudes in cells kept in mono- or co-culture. The number of genes regulated in EC was thus twice as high in co-cultured cells compared to those in monoculture, suggesting that interaction with VSMCs sensitizes them to the applied stress factors. Subsequent functional analysis suggested that co-culture potentiated the effect of the stressors, including on EC proliferation, what was confirmed experimentally. Furthermore, the stressor-induced changes in the proteome and secretome of co-cultured cells were linked to the predicted regulators of changes in gene expression. This omics data integration predicted GDF15 (growth differentiation factor 15), HMGB1 (High-mobility group box 1) and PLAU (plasminogen activator) as putative mediators of the VSMC-EC crosstalk under obesity-related stress. Conclusions VSMC and EC kept in bilayer co-culture inherently differ from vascular cells in monoculture, and display different responsiveness to external stressors, as shown here for obesity-related stressors. These results highlight how cellular crosstalk may be required for stress-induced phenotypical changes, which could thus not be predicted from studies focused on single-cell types only.
Dubourg et al. (2026) studied Obesity/DMT2-associated vascular dysfunction. Exposure to metabolic and humoral stressors (high glucose, free fatty acids, angiotensin II, norepinephrine) in co-culture vs. Monoculture and/or control conditions was evaluated on Transcriptome changes, viability, proliferation, secretion of inflammation markers, proteome, and secretome. Co-culture of primary human endothelial and vascular smooth muscle cells sensitized endothelial cells to obesity-associated metabolic and humoral stress, doubling the number of regulated genes.