DataSheet1_Inhibition of the Proliferation of Human Lung Fibroblasts by Prostacyclin Receptor Agonists is Linked to a Sustained cAMP Signal in the Nucleus.PDF

Idiopathic pulmonary fibrosis is a chronic and progressive fibrotic lung disease, and current treatments are limited by their side effects. Proliferation of human lung fibroblasts in the pulmonary interstitial tissue is a hallmark of this disease and is driven by prolonged ERK signalling in the nucleus in response to growth factors such as platelet-derived growth factor (PDGF). Agents that increase cAMP have been suggested as alternative therapies, as this second messenger can inhibit the ERK cascade. We previously examined a panel of eight Gαs-cAMP-coupled G protein-coupled receptors (GPCRs) endogenously expressed in human lung fibroblasts. Although the cAMP response was important for the anti-fibrotic effects of GPCR agonists, the magnitude of the acute cAMP response was not predictive of anti-fibrotic efficacy. Here we examined the reason for this apparent disconnect by stimulating the Gαs-coupled prostacyclin receptor and measuring downstream signalling at a sub-cellular level. MRE-269 and treprostinil caused sustained cAMP signalling in the nucleus and complete inhibition of PDGF-induced nuclear ERK and fibroblast proliferation. In contrast, iloprost caused a transient increase in nuclear cAMP, there was no effect of iloprost on PDGF-induced ERK in the nucleus, and this agonist was much less effective at reversing PDGF-induced proliferation. This suggests that sustained elevation of cAMP in the nucleus is necessary for efficient inhibition of PDGF-induced nuclear ERK and fibroblast proliferation. This is an important first step towards understanding of the signalling events that drive GPCR inhibition of fibrosis.

特发性肺纤维化是一种慢性且渐进性的肺纤维化疾病，其治疗手段受限于其副作用。人类肺成纤维细胞在肺间质组织的增殖是该疾病的特征之一，且由对血小板衍生生长因子（PDGF）等生长因子反应的长期ERK信号传导在细胞核内驱动。有研究表明，提高cAMP水平的药物可作为替代疗法，因为这种第二信使能够抑制ERK级联反应。我们先前曾检查了在人类肺成纤维细胞中内源表达的一种Gαs-cAMP偶联G蛋白偶联受体（GPCR）的八种受体面板。尽管cAMP反应对于GPCR激动剂的抗纤维化作用至关重要，但急性cAMP反应的幅度并不能预测抗纤维化疗效。在本研究中，我们通过刺激Gαs偶联的前列环素受体并测量亚细胞水平的下游信号传导，来探究这种看似脱节的根本原因。MRE-269和treprostinil在细胞核内引起了持续的cAMP信号传导，并完全抑制了PDGF诱导的细胞核ERK和成纤维细胞的增殖。相比之下，iloprost仅导致细胞核cAMP的短暂增加，对PDGF诱导的细胞核ERK没有抑制作用，且这种激动剂在逆转PDGF诱导的增殖方面的效果显著较差。这表明，在细胞核内持续提高cAMP水平对于有效抑制PDGF诱导的细胞核ERK和成纤维细胞增殖是必要的。这是理解驱动GPCR抑制纤维化的信号事件的重要第一步。