An antiproliferative BMP-2/PPARγ/apoE axis in human and murine SMCs and its role in pulmonary hypertension
J. Clin. Invest. Georg Hansmann, et al. 118:1846 doi:10.1172/JCI32503 [Go to this article.]

Figure 1
Antiproliferative effects of BMP-2 (A, C, D, and F), the PPARγ agonist rosiglitazone (Rosi; B), and apoE (E) on PDGF-BB–induced proliferation of human (A, B, C, and E) and murine (D and F) PASMCs. PASMCs were seeded at 2.5 × 10⁴ cells per well of a 24-well plate in 500 μl of growth medium and allowed to adhere overnight. The cells were washed with PBS prior to the addition of starvation media (0.1% FBS) and incubated for 24 hours (murine PASMCs) or 48 hours (HPASMCs) and then stimulated with PDGF-BB (20 ng/ml) for 72 hours. BMP-2 (10 ng/ml), rosiglitazone (1 μM), and recombinant human apoE (1–10 μM) were added to quiescent cells 30 minutes prior to PDGF-BB stimulation. The PPARγ antagonist GW9662 (GW; 1 μM) was added 24 hours prior to the addition of BMP-2. Cells were finally washed twice with PBS, trypsinized, and counted in a hemacytometer (4 counts per well). Cell numbers in controls at time points 0 (CON) and 72 hours were not significantly different. A: shLacZi, HPASMCs transfected with short hairpin LacZi pLentivirus 6 (control); shBMP-RIIi, HPASMCs transfected with short hairpin pLentivirus 6 BMP-RIIi (i.e., BMP-RII–deficient PASMCs). D: Littermates, littermate control PASMCs; SMC PPARγ^–/–, PASMCs isolated from SM22α Cre PPARγ^flox/flox mice. F: C57BL/6, control murine PASMCs; apoE^–/–, PASMCs isolated from apoE-deficient mice. Bars represent mean ± SEM (n = 3 in A, D, and F; n = 4 in B and C; n = 6 in E; n = 12 in controls of A). *P < 0.05; **P < 0.01; ***P < 0.001 as indicated; ANOVA with Bonferroni’s multiple comparison test.