Vascular calcification (VC) is a multifactorial pathological deposition of calcium in the vasculature and is associated with severe cardiovascular outcomes, particularly in patients with chronic kidney disease (CKD). Various vitamin K analogs have been found to influence the development of VC. We utilized a high-phosphate-induced VC model in mouse vascular smooth muscle cells (VSMCs) and developed an in vivo VC model using ApoE^−/− mice subjected to 5/6 nephrectomy and fed an oral high-phosphorus diet to evaluate the effect of the vitamin K3 analog phthiocol. Transdermal glomerular filtration rate measurement, pulse wave velocity for aortic stiffness assessment, blood biochemical analysis, and pathological examinations were conducted. Phthiocol suppressed reactive oxygen species production and reduced subsequent cell death and calcification in a dose-dependent manner. It inhibited osteogenic trans-differentiation by restoring the PI3K/Akt pathway, activating Nrf2/HO-1 antioxidation signaling, and downregulating IL-1β and TNF-α. The high-phosphate diet in ApoE^−/− CKD mice significantly induced dyslipidemia, renal impairment, hyperphosphatemia, aortic stiffness, and calcium deposition in aortic tissue compared to the control group. Phthiocol treatment markedly improved dyslipidemia, hyperphosphatemia, and aortic stiffness. The vitamin K3 analog phthiocol ameliorates phosphate-induced osteogenic trans-differentiation of VSMCs and subsequent VC by restoring the PI3K/Akt pathway and enhancing Nrf2/HO-1 antioxidant activity.

Vascular calcification (VC) is a major complication of chronic kidney disease (CKD) that is strongly associated with cardiovascular mortality. While indoxyl sulfate (IS), a protein-bound uremic toxin, has been implicated in the progression of VC, the underlying molecular mechanisms remain unclear. We investigated the procalcific effects of IS using a two-step nephrectomy-induced CKD mouse model and cultured vascular smooth muscle cells. In vivo, progressive renal impairment was associated with elevated circulating IS levels and enhanced VC. In vitro, IS dose- and time-dependently induced calcium deposition, increased reactive oxygen species (ROS) production, and upregulated osteogenic markers, including runt-related transcription factor 2 (RUNX2), bone morphogenetic protein 2 (BMP2), and osteocalcin (OCN), whereas N-acetyl-L-cysteine (NAC) partially attenuated IS-induced ROS accumulation and cell injury. Mechanistically, IS exposure activated extracellular signal-regulated kinase (ERK) and Wnt/β-catenin signaling while suppressing nuclear factor erythroid 2-related factor 2 (Nrf2)-related antioxidant responses, as reflected by reductions in the phosphorylated Nrf2 (pNrf2) to total Nrf2 and heme oxygenase-1 (HO-1) expression. Pharmacological inhibition of ERK and Wnt/β-catenin signaling attenuated IS-induced osteogenic responses. These findings indicate that IS promotes VC in association with increased oxidative stress, activation of ERK and Wnt/β-catenin signaling, and impaired Nrf2-related antioxidant defense. These integrated findings provide mechanistic insight into IS-associated VC and highlight oxidative stress-related signaling networks as potential therapeutic targets in CKD.