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. 2021 Nov 29;53(11):1781–1791. doi: 10.1038/s12276-021-00708-6

Fig. 7. In vivo adenoviral overexpression of either Msx1 or Msx2 potentiates vitamin D3-induced vascular calcification in an MDM2-dependent manner in vivo.

Fig. 7

a Timeline for the induction of vascular calcification and adenoviral gene delivery. b Alizarin red S staining of aortas. Vascular calcification induced by injection of vitamin D3 (3 × 105 IU kg−1) was further enhanced by overexpression of either Msx1 or Msx2 in aortas from Mdm2fl/fl mice. However, this enhancement was not observed in SM22α-cre;Mdm2fl/fl mice. c Calcium content in the aorta (n = 3–11). d Immunohistochemical analysis with fluorescent antibodies showing the expression of Runx2 in Mdm2fl/fl (upper panels) and SM22α-cre;Mdm2fl/fl mice (lower panel). Scale bar = 100 μm. e Western blot analysis showing the protein expression of Mdm2, Hdac1, and Runx2. f Schematic diagram. Calcification stress induces the expression of the key transcription factors MSX1 and MSX2, which then redundantly activate the transcription of MDM2. MDM2 regulates HDAC1 expression and thereby the RUNX2 protein level in an E3 ligase activity-dependent manner, which results in vascular calcification.