P3490Phenotypic specification of endothelial cells in chronic thromboembolic pulmonary hypertension

Abstract

Results: We found that severe OSA could damage cardiac function and aggravate collagen deposition via EndMT. These features were improved by PHD3 overexpression. In vitro, PHD3 overexpression in cultured HUVECs ameliorated OSA-induced EndMT through inactivating HIF-1α and Smad2/3. Conclusion: We conclude that PHD3 overexpression might be beneficial in the prevention of OSA-induced cardiac fibrosis by inhibiting EndMT through inactivating HIF-1α and Smad2/3. Introduction: Endothelial erosion of plaques is the underlying mechanism that causes approximately 30% of heart attacks. It is mostly observed overlying stenotic plaques (expected to experience elevated shear stress) of smokers (known to experience endothelial dysfunction). Smokers also have elevated circulating cytokine levels, e.g.TNFα, resulting from inflammation in the lungs. Purpose: To develop a model to study human coronary artery endothelial cells (HCAEC) at elevated shear stress and investigate the effects of cigarette smoke extract (CSE) and TNFα. Methods: Confluent monolayers of HCAEC were under oscillatory (0±5 dynes/cm 2 , 1Hz -OSS), normal laminar (15 dynes/cm 2 , -LSS), or elevated laminar shear stress (75 dynes/cm 2 , -ESS), using a parallel plate flow apparatus. Cell were cultured for 24 hours to allow cells to adapt to their shear environment before three treatments of vehicle (control), TNFα (5ng/ml), CSE (10%), or both (T+C) were administered into the flow apparatus 16 hours apart. Cells were analysed 16 hours after the last treatment. Results: LSS and ESS induced alignment of HCAEC with the direction of flow, which was unaffected with treatment of TNFα or CSE. Combine exposure of T+C resulted in ∼30% cell loss under elevated flow conditions, with no cell loss observed at OSS. Treatment with apoptosis inhibitor (Z-VAD-FMK) or matrix metalloproteinase inhibitor (GM6001) did not prevent cell loss. A robust activation of Nrf2-regulated genes e.g. HMOX1, was observed by CSE, which was amplified at LSS, and ESS by TNFα. Inclusion of NRF2 activators sulforaphane (2.5μM) or isoliquiritigenin (10μM) triggered ∼80% cell loss at elevated shear stress with T+C, implying that hyperactivation of the Nrf2 system, may promote, rather than protect from cell detachment. In addition, adenoviral overexpression of wild type or dominant negative Nrf2, inhibited HCAEC proliferation. Conclusions:We have identified that the combination of CSE and inflammatory cytokine TNFα trigger cell detachment in HCAEC adapted to elevated shear stress. Cell detachment appeared independent of apoptosis, or MMP activity. Treatment of CSE increased the expression of several Nrf2-regulated genes, an effect amplified by TNFα at elevated shear stress. Further activation of Nrf2 resulted in almost complete cell detachment, suggesting it may play a role in HCAEC cell loss. Modulation of Nrf2 activity outside its normal range reduced cell proliferation, which may limit the ability of the endothelium to repair damage. Taken together, sustained hyperactivation of Nrf2 may promote endothe...

Cell landscape atlas for patients with chronic thromboembolic pulmonary hypertension after pulmonary endarterectomy constructed using single-cell RNA sequencing

“…Furthermore, specific marker genes of fibroblast/smooth muscle cells were identified, such as PDGFRB, COL1A1, and COL3A1, which were significantly enriched for multiple functions associated with smooth muscle cell migration. It has been reported that CTEPH endothelial cells can exhibit pro-fibrotic and pro-inflammatory phenotypes, as demonstrated by the expression of genes (COL1A1 and COL3A1) involved in extracellular matrix production and fibril organization [20]. In addition, PDGF is a potent mitogen for cells of mesenchymal origin, such as fibroblasts and smooth muscle cells [21].…”