Imbalanced prostanoid release mediates cigarette smoke-induced human pulmonary artery cell proliferation

Abstract

Background Pulmonary hypertension is a common and serious complication of chronic obstructive pulmonary disease (COPD). Studies suggest that cigarette smoke can initiate pulmonary vascular remodelling by stimulating cell proliferation; however, the underlying cause, particularly the role of vasoactive prostanoids, is unclear. We hypothesize that cigarette smoke extract (CSE) can induce imbalanced vasoactive prostanoid release by differentially modulating the expression of respective synthase genes in human pulmonary artery smooth muscle cells (PASMCs) and endothelial cells (PAECs), thereby contributing to cell proliferation. Methods Aqueous CSE was prepared from 3R4F research-grade cigarettes. Human PASMCs and PAECs were treated with or without CSE. Quantitative real-time RT-PCR and Western blotting were used to analyse the mRNA and protein expression of vasoactive prostanoid syhthases. Prostanoid concentration in the medium was measured using ELISA kits. Cell proliferation was assessed using the cell proliferation reagent WST-1. Results We demonstrated that CSE induced the expression of cyclooxygenase-2 (COX-2), the rate-limiting enzyme in prostanoid synthesis, in both cell types. In PASMCs, CSE reduced the downstream prostaglandin (PG) I synthase (PGIS) mRNA and protein expression and PGI2 production, whereas in PAECs, CSE downregulated PGIS mRNA expression, but PGIS protein was undetectable and CSE had no effect on PGI2 production. CSE increased thromboxane (TX) A synthase (TXAS) mRNA expression and TXA2 production, despite undetectable TXAS protein in both cell types. CSE also reduced microsomal PGE synthase-1 (mPGES-1) protein expression and PGE2 production in PASMCs, but increased PGE2 production despite unchanged mPGES-1 protein expression in PAECs. Furthermore, CSE stimulated proliferation of both cell types, which was significantly inhibited by the selective COX-2 inhibitor celecoxib, the PGI2 analogue beraprost and the TXA2 receptor antagonist daltroban. Conclusions These findings provide the first evidence that cigarette smoke can induce imbalanced prostanoid mediator release characterized by the reduced PGI2/TXA2 ratio and contribute to pulmonary vascular remodelling and suggest that TXA2 may represent a novel therapeutic target for pulmonary hypertension in COPD.

Increased Thromboxane A2 Levels in Pulmonary Artery Smooth Muscle Cells Isolated from Patients with Chronic Obstructive Pulmonary Disease

“…Human pulmonary artery smooth muscle cells from three patients with COPD and from three healthy donors were used in the current study. Cells from healthy donors were obtained as previously described [ 5 ] from Thermo Fisher Scientific, Waltham, MA, USA, Cell Applications, San Diego, CA, USA, and Lonza Group, Basel, Switzerland, with no history of respiratory disease or evidence of pulmonary vascular abnormalities. Human pulmonary artery smooth muscle cells from three smokers with a confirmed diagnosis of COPD ( Table 1 ) were supplied by Prof Linhua Pang and Dr Rachel L. Clifford (University of Nottingham, Nottingham, UK).…”

“…Human pulmonary artery smooth muscle cells from three smokers with a confirmed diagnosis of COPD ( Table 1 ) were supplied by Prof Linhua Pang and Dr Rachel L. Clifford (University of Nottingham, Nottingham, UK). Both healthy and diseased cells were received at passage 3, sub-cultured in Dulbecco’s Modified Eagles Medium (DMEM) with 20% foetal bovine serum (FBS) as previously described [ 5 ] and used at passage 6. Pulmonary artery smooth muscle cells were cultured in a six-well plate with DMEM with 20% FBS for 48 h until the cells reached 100% confluence.…”

“…Cigarette smoke, the most prominent risk factor for COPD, has been shown to induce pulmonary vascular remodelling, which leads to increased pulmonary vascular resistance, pulmonary arterial pressure and, ultimately, pulmonary hypertension [ 2 , 3 , 4 ]. Findings reported by our group and others revealed that cigarette smoke extract could induce the proliferation of pulmonary artery cells, including pulmonary artery smooth muscle cells [ 5 , 6 , 7 ]. Although the underlying mechanism is not completely understood, our previous findings strongly suggest that imbalanced prostanoids play a key role in the cigarette smoke-induced proliferation of pulmonary artery cells [ 5 ].…”

“…Findings reported by our group and others revealed that cigarette smoke extract could induce the proliferation of pulmonary artery cells, including pulmonary artery smooth muscle cells [ 5 , 6 , 7 ]. Although the underlying mechanism is not completely understood, our previous findings strongly suggest that imbalanced prostanoids play a key role in the cigarette smoke-induced proliferation of pulmonary artery cells [ 5 ].…”

“…It has been demonstrated that cigarette smoke, the strongest risk factor for COPD, can induce imbalanced prostanoids, characterised mainly by increased thromboxane A 2 production in healthy human pulmonary artery smooth muscle cells and pulmonary artery endothelial cells [ 5 ]. More importantly, it has also been reported that blocking the effect of increased thromboxane A 2 using daltroban, a thromboxane A 2 receptor antagonist, can inhibit the cigarette smoke-induced proliferation of pulmonary artery smooth muscle cells and pulmonary artery endothelial cells [ 5 ]. However, it is largely unknown whether thromboxane A 2 is greater in pulmonary artery cells from smokers with COPD.…”