After treatment, HUVECs or HAECs in 100-mm dishes were washed twice with PBS, and continued culture in serum-free EC medium for 24 h. The 24-h EC-conditioned medium was collected individually. To inactivate secreting miRNA complex, the collected EC-conditioned medium was treated with proteinase K, RNase or both, followed by determination of miR-126-3p levels and used to determine the effect on HASMC calcification.

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