Astaxanthin Counteracts Vascular Calcification In Vitro Through an Early Up-Regulation of SOD2 Based on a Transcriptomic Approach.

Chia-Ter Chao,Huei-Wen Chen,You-Tien Tsai,Tzu-Hang Yuan,Jenq-Wen Huang,Hsiang-Yuan Yeh,Min-Tser Liao

doi:10.3390/ijms21228530

Abstract

Vascular calcification (VC) is a critical contributor to the rising cardiovascular risk among at-risk populations such as those with diabetes or renal failure. The pathogenesis of VC involves an uprising of oxidative stress, for which antioxidants can be theoretically effective. However, astaxanthin, a potent antioxidant, has not been tested before for the purpose of managing VC. To answer this question, we tested the efficacy of astaxanthin against VC using the high phosphate (HP)-induced vascular smooth muscle cell (VSMC) calcification model. RNAs from treated groups underwent Affymetrix microarray screening, with intra-group consistency and inter-group differential expressions identified. Candidate hub genes were selected, followed by validation in experimental models and functional characterization. We showed that HP induced progressive calcification among treated VSMCs, while astaxanthin dose-responsively and time-dependently ameliorated calcification severities. Transcriptomic profiling revealed that 3491 genes exhibited significant early changes during VC progression, among which 26 potential hub genes were selected based on closeness ranking and biologic plausibility. SOD2 was validated in the VSMC model, shown to drive the deactivation of cellular senescence and enhance antioxidative defenses. Astaxanthin did not alter intracellular reactive oxygen species (ROS) levels without HP, but significantly lowered ROS production in HP-treated VSMCs. SOD2 knockdown prominently abolished the anti-calcification effect of astaxanthin on HP-treated VSMCs, lending support to our findings. In conclusion, we demonstrated for the first time that astaxanthin could be a potential candidate treatment for VC, through inducing the up-regulation of SOD2 early during calcification progression and potentially suppressing vascular senescence.

Highlights

Vascular calcification (VC) is an important complication that occurs frequently in patients of advanced age, with diabetes mellitus (DM), or chronic kidney disease (CKD) [1]
We showed that higher concentrations of astaxanthin dose-dependently attenuated the severity of calcification over time, with less Alizarin red positive nodules in cells exposed to higher doses (Figure 1A,B)
We discovered that 0.1, 1, and 5 μM of astaxanthin abolished 33%, 70%, and 67%, respectively, of calcification severity compared with the high phosphate (HP) only group after 7 days of treatment (Figure 1C)

Summary

Introduction

Vascular calcification (VC) is an important complication that occurs frequently in patients of advanced age, with diabetes mellitus (DM), or chronic kidney disease (CKD) [1]. The pathogenesis of VC has evolved from passive calcium deposition to the involvement of active osteoid secretion by trans-differentiated vascular smooth muscle cells (VSMCs) Triggering of such phenotypic changes is multi-faceted, including inflammatory stimuli, excessive calcium and/or phosphate exposure, prolonged exposure to pro-calcific and pro-fibrotic mediators, and so on, assisted simultaneously by the decline in anti-calcific defending molecules [7]. Sources of reactive oxygen species (ROS) ranging from peroxides, advanced oxidized protein products (AOPPs), to advanced glycation endproducts (AGEs) have all been shown to precipitate bio-mineralization among different types of cells subject to adverse stimuli [9]. From this perspective, anti-oxidants are purported as promising therapeutic options for managing VC of diverse origins [10]

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