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. 2022 Jul 26;146(21):1591–1609. doi: 10.1161/CIRCULATIONAHA.121.057623

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© 2022 The Authors.

Circulation is published on behalf of the American Heart Association, Inc., by Wolters Kluwer Health, Inc. This is an open access article under the terms of the Creative Commons Attribution Non-Commercial-NoDerivs License, which permits use, distribution, and reproduction in any medium, provided that the original work is properly cited, the use is noncommercial, and no modifications or adaptations are made.

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Figure 6. — NR1D1 deficiency results in early changes in mitochondrial function on day 3 after AngII infusion before a change in AAAs. Abdominal aortas from ApoE^−/−/Nr1d1^flox/flox and ApoE^−/−/Nr1d1^ΔSMC mice were harvested on the morning of day 3 after AngII infusion before a change in AAAs. A, Representative images of the macroscopic features and HE staining of suprarenal abdominal aortas in ApoE^−/−/Nr1d1^flox/flox and ApoE^−/−/Nr1d1^ΔSMC mice on day 3 after AngII infusion. B, Quantification of the maximal diameter of suprarenal abdominal aortas in indicated groups (n=10 per group). Data were analyzed by Student t test. C, Representative images of MMP activity and in situ DHE staining in indicated groups. D, Left, MMP-2 protein expression measured by Western blot and MMP activity measured by zymography in indicated groups. Right, Quantification of MMP-2 protein expression and MMP activity in indicated groups (n=6 per group). Data were analyzed by Student t test. **P<0.01; ***P<0.001. E, IL-6, TNF-α, and MCP-1 expression in abdominal aortas assessed by ELISAs in indicated groups (n=10 per group). Data were analyzed by Student t test. **P<0.01; ***P<0.001. F, IL-6, TNF-α, and MCP-1 expression measured by immunohistochemistry in indicated groups. G and H, Aco2 mRNA expression and ACO2 activity measured by RT-qPCR and ELISAs in abdominal aortas from ApoE^−/−/Nr1d1^flox/flox and ApoE^−/−/Nr1d1^ΔSMC mice on day 3 after AngII infusion (n=6 per group). Data were analyzed by Student t test. ***P<0.001. I, Left, ACO2 protein expression measured by Western blot in abdominal aortas from ApoE^−/−/Nr1d1^flox/flox and ApoE^−/−/Nr1d1^ΔSMC mice on day 3 after AngII infusion. Right, Quantification of ACO2 protein expression measured by Western blot in indicated groups (n=6 per group). Data were analyzed by Student t test. **P<0.01. J, Representative images of ACO2 expression by immunofluorescence staining in indicated groups. K, Left, Representative images of mitochondrial ∆Ψm by JC-1 staining in indicated groups. Right, Quantification of JC-1 staining (n=6 per group). Data were analyzed by Student t test. ***P<0.001. L, Left, Protein expression of mitochondrial complexes in the abdominal aortas from AngII-infused ApoE^−/−/Nr1d1^flox/flox and ApoE^−/−/Nr1d1^ΔSMC mice on day 3 after AngII infusion. Right, Quantification of protein expression of mitochondrial complexes in indicated groups (n=6 per group). Data were analyzed by Student t test. *P<0.05; **P<0.01; ***P<0.001. Data are expressed as mean ± SEM. ACO2 indicates aconitase-2; AngII, angiotensin II; DHE, dihydroethidium; IL-6, interleukin-6; MCP-1, monocyte chemoattractant protein-1; MMP, matrix metalloproteinase; NR1D1, nuclear receptor subfamily 1 group D member 1; NS, nonsignificant; RT-qPCR, real-time quantitative polymerase chain reaction; SMC, smooth muscle cell; and TNF-α, tumor necrosis factor-α.