Spontaneous Right Ventricular Pseudoaneurysms and Increased Arrhythmogenicity in a Mouse Model of Marfan Syndrome

Figure A1. Macroscopic analysis of the cardiac dimensions of the dissected hearts of WT and MFS mice at the age of 1, 2, and 3 months (n = 12/group). No significant difference in cardiac dimensions (length and width) was observed between WT and MFS mice (independent samples test; p ≥ 0.132) (a): dot plot visualizes the individual lengths of the dissected hearts and mean value per age group and genotype. (b): dot plot visualizes individual width of the dissected hearts and mean value per age group and genotype. WT and MFS measurements are indicated in grey and black, respectively. Error bars represent standard error. WT, wild-type; MFS, Marfan syndrome.

Figure A2. Representative traces of arrhythmic events. (a): murine ECG trace in sinus rhythm. (a’): two-fold magnification of the murine ECG trace of a single heart beat in sinus rhythm marked with the different components: the P-wave (P), the QRS-complex (QRS), the J-wave (J), and the T-wave (T). (b): murine ECG trace with extrasystole, single. An extrasystole is described as an extra heart beat either originating from supraventricular, ventricular or near junctional cardiac tissue. This is represented as an extra QRS-complex with an abnormal morphology, mostly followed by a deviant compensatory longer RR-interval (indicated with brackets). (c): murine ECG trace with extrasystole, doublet. A doublet is represented as two consecutive extrasystoles. (d): murine ECG trace with three consecutive extrasystoles. (e): murine ECG trace with sinus arrest. A sinus arrest is represented by a long RR-interval within the ECG trace in the absence of any ECG activity. (f): murine ECG trace with blocked P-wave. A blocked P-wave is represented by the presence of a P-wave (indicated with arrow) without subsequent QRS-complex. The resulting long RR-interval is not a multiple of the normal mean RR-interval.…

Figure 1. Macroscopic characterization of the heart of WT and MFS mice at 1, 2, and 3 months of age. Dot plot visualizes individual heart weight/tibia length ratios and mean value per age and genotype (n = 12 per age and genotype). WT and MFS measurements are indicated in grey and black, respectively. Error bars represent standard error. Associated p-values are annotated at the top of the graph (independent samples test on the mean values of the HW/TL). At the bottom of the figure, representative macroscopic pictures of WT and MFS mice hearts collected at 1, 2, and 3 months of age are depicted. Scale bar: 2 mm. HW/TL, heart weight to tibia length ratio (g/mm); WT, wild-type; MFS, Marfan syndrome.

Figure 2. Macroscopic analysis of the heart of MFS mice. (a,b): macroscopic view of the heart of a MFS mouse presenting a tear at the level of the right ventricular wall. The tear is indicated with a white arrow in panel (b). (c,d): macroscopic view of the heart of a MFS mouse presenting vesicles at the level of the right ventricular wall. Vesicles are indicated with a white arrowhead in panel (d). (e): number of WT and MFS mice presenting cardiac lesions per age group. Scale bar of panels (a,c) = 1 mm, scale bar of panels (b,d) = 250 µm.

Figure 3. Microscopic analysis of tissue sections at the level of a myocardial tear and vesicle in the right ventricle free wall of the heart of MFS mice. (a,c): standard HE staining of representative myocardial tissue sections presenting a myocardial tear (indicated with *) and a vesicle (indicated with •). The black arrow in panel (c) indicates the narrow duct in between cardiac muscle lamellae connecting the lumen of the vesicle with the lumen of the right ventricle. Black boxes indicate localization of panels (b,d,e,g) within the tissue sections. (b,d,e,g): detailed images of HE staining of the myocardial lesions and the adjacent tissue. (f,h): picrosirius red staining (collagen deposition) of myocardial tissue sections at the level of a tear and vesicle, respectively. (i–l): double fluorescent staining of vimentin and collagen (stained red and green, respectively) in myocardial tissue sections presenting a myocardial tear and vesicle. Nuclei are stained with DAPI (blue). (m–p): fluorescent staining of α-SMA (red) and collagen (green), with nuclei stained blue. Arrowheads indicate α-SMA staining of vascular smooth muscle cells in the blood vessel wall. (q,s): immunofluorescence…

Figure 4. Decreased compaction of the ventricular myocardial tissue in MFS mice correlates with a decreased presence of fibrillin-1 staining. (a,b): overview picture of fibrillin-1 (red) staining of myocardial tissue sections of a 3-month-old WT and MFS mouse, respectively. (c,d): representative overview pictures of HE-stained myocardial tissue sections of a 3-month-old WT and MFS mouse, respectively. (e): graphical presentation of the myocardial compaction level through the measurement of the average non-compaction index of the myocardial tissue sections (n/age group = 3). Within each age group, values are normalized to the WT values. WT and MFS measurements are indicated in grey and black, respectively. Error bars represents standard error. * p = 0.032 (independent samples test). RV, right ventricle; LV, left ventricle; WT, wild-type; MFS, Marfan syndrome. Scale bar = 500 µm.

Figure 5. Canonical and non-canonical TGFβ-dependent signaling in the myocardial tissue of 3-month-old male WT and MFS mice (n = 4 and 5, respectively). (a): no significant difference could be observed in Smad2 activation (pSmad2/Total Smad2) between the MFS mice and their WT littermates (Mann–Whitney U test; p = 0.1111). (b): no significant difference could be observed in Smad3 activation (pSmad3/Total Smad3) between the MFS mice and their WT littermates (Mann–Whitney U test; p = 0.9048). (c): no difference in ERK activation (pERK/ERK ratio) could be observed between the MFS mice and their WT littermates (Mann–Whitney U test; p = 0.4127). Dot plots visualize normalized individual ratios and mean value per genotype. WT and MFS measurements are indicated in grey and black, respectively. Error bars represent standard error. WT, wild-type; MFS, Marfan syndrome.