Fig 2: Vcan haploinsufficiency protects MFS mice from aortic dilation.(A) Representative in vivo ultrasound images of the AsAo and AbAo from 12-week-old WT, Vcanhfd/+, MFS, and MFS Vcanhdf/+ mice. Yellow dashed lines delineate lumen boundaries and green dashed lines denote lumen diameters. Scale bar, 1 mm. (B–D) Maximal AsAo and AbAo diameters in Vcan haploinsufficient mice aged (B) 12, (C) 24, and (D) 36 weeks. Data information: (B–D) data are shown as box-and-whisker plots. The box itself spans from the first quartile (25%) to the third quartile (75%), representing the interquartile range where the central 50% of data values fall. Inside the box, a line denotes the median value. The whiskers of the boxplot extend from the ends of the box to the minimum and maximum values. Each data point denotes an individual mouse (12-week-old mice: WT, n = 13; Vcanhdf/+, n = 14; MFS, n = 15; MFS;Vcanhdf/+, n = 18. 24-week-old mice: WT, n = 20; Vcanhdf/+, n = 11; MFS, n = 18; MFS;Vcanhdf/+, n = 21. 36-week-old mice: WT, n = 23; Vcanhdf/+, n = 15; MFS, n = 25; MFS;Vcanhdf/+, n = 21). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 versus WT mice; #P < 0.05, ##P < 0.01, ####P < 0.0001 versus MFS mice (two-way ANOVA with Tukey’s post hoc test or the Kruskal–Wallis test with Dunn’s multiple comparison test, as appropriate). Source data are available online for this figure.

Fig 3: VCAN triggers aortic dilation via Akt activation and Nos2 induction in MFS mice.(A) Representative immunoblot analysis (left panel) and quantification of the immunoblot signal (right panel) of p-Akt-S473, total Akt, and tubulin in protein extracts from VSMCs grown in serum-starved conditions overnight and subsequently seeded for 1 h on plates precoated with 20 µg/ml VCAN V3 or treated with PBS as a control (n = 3 independent experiments). (B) Experimental design: 12-week-old WT and MFS mice were treated on 4 consecutive days with 20 mg/kg/day AZD8055. Longitudinal ultrasound (Eco) was performed at the indicated time points, and mice were euthanized after 4 days of treatment. (C) Maximal AsAo and AbAo diameters of WT Veh (n = 6), WT AZD8055 (n = 5), MFS Veh (n = 6), and MFS AZD8055 (n = 6) mice at the indicated time points. (D) Representative images of Nos2 immunofluorescence (red), elastin autofluorescence (green), and Hoechst-stained nuclei (blue) in mouse aortic sections. Scale bar, 50 μm. The same images are also presented in the bottom line of Fig. EV4. (E) Quantification of Nos2 immunofluorescence in AsAo from WT Veh (n = 6), WT AZD8055 (n = 6), MFS Veh (n = 6), and MFS AZD8055 (n = 5) mice. (F) Representative images of pAKT-S473 immunofluorescence (red), elastin autofluorescence (green), and Hoechst-stained nuclei (blue) in the medial layer of human aortic tissue from control donors and MFS patients. Scale bar, 50 µm. The same images are also presented in the first line of Fig. EV5. (G) Quantification of pAKT-S473 immunofluorescence in aortas from control donors (n = 7) and MFS patients (n = 8). Data information: (A) data are shown relative to non-coated plates (control) as mean ± s.e.m. Each data point denotes an independent experiment. *P < 0.05 (Student t test). (C)Data are shown as mean ± s.e.m. *P < 0.05, ***P < 0.001, ****P < 0.0001 versus MFS Veh mice (repeated-measurements two-way ANOVA with Tukey’s post hoc test). (E) Data are shown relative to Veh-treated WT mice as mean ± s.e.m. Each data point denotes an individual mouse. *p < 0.05 versus Veh-treated WT mice; #P < 0.05 versus Veh-treated MFS mice (Kruskal–Wallis test with Dunn’s multiple comparison test). (G) Data are shown relative to control donors as mean ± s.e.m. Each data point denotes an individual. **P < 0.01 (unpaired t test with Welch’s correction). Source data are available online for this figure.

Fig 4: Accumulation of Vcan in aortas of MFS mice at 12 weeks but not at 4 weeks of age.(A) Representative images of Vcan immunofluorescence (red), elastin autofluorescence (green), and Hoechst-stained nuclei (blue) in aortic sections from 4- or 12-week-old WT and MFS mice. Scale bar, 50 μm. (B) Quantification of Vcan immunofluorescence in mouse AsAo (4-week-old: WT, n = 4; MFS, n = 3. 12-week-old: WT, n = 4; MFS, n = 3). Data information: (B) data are shown relative to 12-week-old WT mice as mean ± s.e.m. Each data point denotes an individual mouse. *P < 0.05 (two-way ANOVA with Tukey’s post hoc test). Source data are available online for this figure.

Fig 5: Vcan silencing reduces Nos2 expression in aortas from MFS mice.(A) Representative images of Nos2 immunofluorescence (red), Hoechst-stained nuclei (blue), and elastin autofluorescence (green) in aortic sections from 16-week-old MFS and WT mice. Individual channels are shown followed by a composite image. Scale bar, 50 μm. The 4 merged images are identical to those shown in Fig. 7D. (B, C) Representative images of (B) Sma or (C) Cd31 (pale gray), Nos2 immunofluorescence (red), elastin autofluorescence (green), and Hoechst-stained nuclei (blue) in aortic sections from 12-week-old WT and MFS mice. Scale bar, 50 μm. Source data are available online for this figure.