Identification of cell-biologic mechanisms of coronary artery spasm and its ex vivo diagnosis using peripheral blood-derived iPSCs.

ABSTRACT

BACKGROUND: Although vasospastic angina (VSA) is known to be caused by coronary artery spasm, no study has fully elucidated the exact underlying mechanism. Moreover, in order to confirm VSA, patients should undergo invasive coronary angiography with spasm provocation test. Herein, we investigated the pathophysiology of VSA using peripheral blood-derived induced pluripotent stem cells (iPSCs) and developed an ex vivo diagnostic method for VSA. METHODS AND RESULTS: With 10 mL of peripheral blood from patients with VSA, we generated iPSCs and differentiated these iPSCs into target cells. As compared with vascular smooth muscle cells (VSMCs) differentiated from iPSCs of normal subjects with negative provocation test, VSA patient-specific iPSCs-derived VSMCs showed very strong contraction in response to stimulants. Moreover, VSA patient-specific VSMCs exhibited a significant increase in stimulation-induced intracellular calcium efflux (Changes in the relative fluorescence unit [ΔF/F]; Control group vs. VSA group, 2.89 ± 0.34 vs. 10.32 ± 0.51, p < 0.01), and exclusively induced a secondary or tertiary peak of calcium efflux, suggesting that those findings could be diagnostic cut-off values for VSA. The observed hyperreactivity of VSA patient-specific VSMCs were caused by the upregulation of sarco/endoplasmic reticulum Ca2+-ATPase 2a (SERCA2a) due to its enhanced small ubiquitin-related modifier (SUMO)ylation. This increased activity of SERCA2a was reversed by treatment with ginkgolic acid, an inhibitor of SUMOylated E1 molecules (pi/µg protein; VSA group vs. VSA + ginkgolic acid, 52.36 ± 0.71 vs. 31.93 ± 1.13, p < 0.01). CONCLUSIONS: Our findings showed that abnormal calcium handling in sarco/endoplasmic reticulum could be induced by the enhanced SERCA2a activity in patients with VSA, leading to spasm. Such novel mechanisms of coronary artery spasm could be useful for drug development and diagnosis of VSA.