Abstract P063: BMP-2 Upregulates PTEN Expression and Induces Apoptosis of Pulmonary Artery Smooth Muscle Cells Under Hypoxia

Abstract

Background: Primary pulmonary hypertension is mainly caused by increased proliferation and decreased apoptosis in pulmonary artery smooth muscle cells (PASMCs). Aim: To investigate the role of BMP-2 in regulation of PTEN and apoptosis of PASMCs under hypoxia. Methods: Normal human PASMCs were cultured in basal medium (BM) or growth medium (GM) and treated with BMP-2 from 5–80 ng/ml under hypoxia (5% CO 2 + 94% N 2 +1% O 2 ) for 72 hours. Gene expression of PTEN, caspase−3, −8, and −9, AKT-1 and -2 were determined by quantitative RT-PCR (QRT-PCR). Protein expression levels of PTEN, AKT and phosph-AKT (pAKT) were determined. Apoptosis of PASMCs were determined by measuring activities of caspases-3, -8 and -9. siRNA-smad-4 was used to determine whether Smad signaling pathway was involved in the regulation of PTEN expression by BMP-2. bpV(HOpic) (PTEN inhibitor) and GW9662 (PPAR-r antagonist) were also used. Results: Proliferation of PASMCs showed dose dependence of BMP-2, the lowest proliferative rate was achieved at 40–60ng/ml under hypoxia (BM=77.2±4%, GM=80±2.8%). Increased gene expression levels of PTEN, caspases−3, −8 and −9 were found, while AKT-1, and -2 did not change. Consistently, the PTEN protein expression also showed dose dependence of BMP-2. Though AKT was unchanged in all treated samples, reduced pAKT was found in BMP-2 treated PASMCs. Increased activities of caspase-3, -8 and -9 of PASMCs were found after cultured with BMP-2 in both mediums. PTEN expression was unchanged when Smad-4 expression was inhibited. However pre-treat PASMCs with bpV(HOpic) and GW9662 (PPAR-r inhibitor) inhibited PTEN protein expression and recovered PASMCs proliferation rate. Conclusion: BMP-2 can increase PTEN expression under hypoxia in a dose dependent pattern. BMP-2 can increase apoptosis of PASMCs under hypoxia. The increased PTEN expression may be mediated through PPAR-r signalling pathway, instead of BMP/Smad signalling pathway.