Rational: Peripheral arterial disease is a common disorder among diabetic and chronic kidney disease patients. As a characteristic sign of peripheral artery disease, vascular calcification (VC) has been associated with a poor prognosis. We hypothesize that hyperglycemia is driving vascular calcification through an altered metabolomics profil.

Methods and Results: Human coronary artery smooth muscle cells (SMCs) were cultured with 0 mM, 5.5 mM, 25 mM glucose under calcifying conditions (3 mM calcium, 2 mM phosphate, CaP). Glucose promoted extracellular matrix calcification in SMCs in a dose- and time-dependent manner, visualized by live-fluorescence imaging of the fetuin-A-probe and alizarin red staining. Furthermore, the absence of glucose abolished SMC calcification. Glucose did not affect CaP complex formation assessed by T50 assay nor cellular viability. However, 0 mM glucose reduced SMC proliferation in control (-2.0 fold, p = 0.029) and calcified conditions (-1.7 fold, p = 0.042) compared to 25 mM glucose. To explore the underlying mechanisms of glucose-dependent VC we applied untargeted metabolomics and transcriptomics. Multi-omics data integration revealed key players from the hypotaurine/taurine metabolic pathway as the center hub of the reconstructed network. Several metabolites from the hypotaurine/taurine metabolic pathway showed altered abundance. Glucose promoted hypotaurine secretion dose-dependently, while its intracellular abundance was not altered. Blocking hypotaurine production by propargylglycine increased extracellular matrix mineralization (+1.3 fold, p = 0.023), while hypotaurine treatment prevented it (12.5 fold, p < 0.001). Both hypotaurine and taurine decreased extracellular matrix mineralization dose-dependently. Furthermore, the omics data suggested an energy remodeling in calcified SMCs under hypoglycemia. Transcriptomics overrepresentation analysis revealed significant dysregulation of fatty acids and cholesterol metabolism. Beta-oxidation molecules were among the most dysregulated metabolites by glucose. Moreover, real-time extracellular efflux analysis showed decreased mitochondrial respiration, measured by ATP production (-67 %, p < 0.001), maximal respiration (-66 %, p < 0.001) and non-mitochondrial respiration (-49 %, p < 0.001) and decreased glycolysis (-54%, p = 0.039) in calcified SMCs. Finally, to identify novel molecules targeting glucose-dependent VC, we performed a drug repurposing analysis with the Cmap database based on signature matching. We identified a PKC inhibitor among the top predicted drugs to reverse calcification by 25 mM glucose.

Conclusion: Our multi-omics analysis revealed a functional role of the hypotaurine/taurine metabolic pathway in glucose-mediated VC. Moreover, our data suggest an energy remodeling in calcified SMCs treated with different glucose concentrations, suggesting potential novel therapeutic targets that warrant further investigation in hyperglycemia-dependent VC.