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. Author manuscript; available in PMC: 2023 Dec 6.
Published in final edited form as: Sci Signal. 2022 Dec 6;15(763):eabn2743. doi: 10.1126/scisignal.abn2743

Figure 1. TSC2 deficiency results in increased PAVSMC proliferation, vascular remodeling and PH.

Figure 1.

a, b. (a) Immunohistochemical analysis of human lung tissue sections to detect TSC2 (magenta), SMA (green) and DAPI (blue). Bar, 100 μm. Images are representative from 3 subjects/group, 6 PAs/subject. (b) Optical density (OD) measurement of TSC2 fluorescent signal in SMA-positive regions of human small PAs. Data are means±SE from 3 subjects/group, 6 PAs/subject, 12 areas/PA. PAH was normalized to control, which was set at 1. P value for PAH compared to control was determined by Mann Whitney U test. Images from different parts of the same gel are separated by dotted line.

c, d. Immunoblot analysis of small PAs. Data are means±SE from n=3 subjects/group. P value for PAH compared to control was determined by Mann Whitney U test.

e, f. Immunoblot analysis of human non-diseased (control) (white bars) and PAH (grey bars) PAVSMCs. Data are means±SE. PAH was normalized to control which was set to 1. N=5 subjects/group. P value for PAH compared to control was determined by Mann Whitney U test.

g. Proliferation of human control and PAH PAVSMCs as measured by Ki67 staining. Data represent % of Ki67-positive cells per total number of cells (as detected by DAPI) and are means±SE from n=6 subjects/group. P value for PAH compared to control was determined by Mann Whitney U test.

h. Equal quantity of cells was plated on each well of 6-well plate (day 0), cell counts were performed at days 3 and 6. Data are fold change to day 0. Data are means±SE from n=5 subjects/group. P values for PAH compared to control were determined by Mann Whitney U test.

i-k. Immunoblot (i), proliferation as measured by BrdU incorporation (j) or cell counting (k) performed on control human PAVSMCs transfected with siRNA directed against TSC2 (siTSC2) or control siRNA GLO (siCont) for 48 hours. Data are means±SE from n=3 subjects/group. P values for siTSC2 compared to siCont were determined by Mann Whitney U test.

l-o. Morphological and hemodynamic analysis of nine week old male SM22-Tsc2+/− and wild-type (WT) mice. Images (l) are representative of 6 mice/group, 12 PAs/mouse. Scale bar, 30 μm. Graphs should PA medial thickness (PA MT) (m), systolic RV pressure (sRVP) (n), and Fulton index (right ventricle (RV)/(left ventricle (LV) + septum) (o). Data are means±SE for n=6 WT mice and n=7 SM22-Tsc2+/− mice. P values for SM22-Tsc2+/− compared to WT by Mann Whitney U test.