Prostacyclin Analogs Inhibit Fibroblast Contraction of Collagen Gels through the cAMP-PKA Pathway

Abstract

Prostacyclin is an arachidonic acid metabolite that modulates vascular tone within the lung. The current study evaluated the hypothesis that prostacyclin can also modulate tissue remodeling by affecting fibroblast-mediated contraction of extracellular matrix. To accomplish this, fibroblasts were cultured in three-dimensional native type I collagen gels in the presence of prostacyclin analogs: carbaprostacyclin, iloprost, and beraprost. All three analogs significantly inhibited contraction of the three-dimensional collagen gels mediated by three different fibroblasts. All three analogs significantly inhibited fibronectin release and reduced fibroblast fibronectin mRNA expression. Addition of exogenous fibronectin restored the contractile activity to fibroblasts incubated in the presence of all three analogs. Iloprost and beraprost significantly activated cAMP-dependent protein kinase-A (PKA), and an action through this pathway was confirmed by blockade of the inhibitory effect on contraction and fibronectin release with the PKA inhibitor KT-5720. In contrast, carbaprostacyclin, which is not as selective for the prostacyclin (IP) receptor, did not activate PKA, and its effects on contraction and fibronectin release were not fully blocked by KT-5720. Finally, the cAMP analogs N 6 -Benzoyl-(6-Bnz-) cAMP and dibutyryl-cAMP inhibited contraction, and this contrasted with the activity of an Epac selective agonist 8-pCPT-2-O-Me-cAMP, which had no effect. Taken together, these results indicate that prostacyclin, acting through the IP receptor and by activating PKA, can lead to inhibition of fibronectin release and can subsequently inhibit fibroblastmediated collagen gel contraction. The ability of prostacyclin to modulate fibroblast function suggests that prostacyclin can contribute to tissue remodeling.

Mechanisms of bradykinin-induced contraction in human fetal lung fibroblasts

“…This function may be increased by a variety of mediators, including platelet-derived growth factor and transforming growth factor-b, and downregulated by badrenergic agonists and prostaglandin E 2 [24,25]. In agreement with a previous observation, we found that a significant increase in BK-induced fibroblast-mediated collagen-gel contraction is detectable at BK concentrations similar to those found in human sputum [26] and after a relatively short period of time (60 min) [15].…”

“…The BK-induced expression of a-SMA and its polymerisation stress fibres were also completely abolished by EGTA, while MLC phosphorylation and collagen-gel contraction were partly inhibited by EGTA. These results are not unexpected findings considering the complexity of the different intracellular pathways involved in fibroblast activation [15,20,25].…”

Prostacyclin analogs stimulate VEGF production from human lung fibroblasts in culture

“…The ability of prostacyclin to modulate fibroblasts through a cAMP-mediated pathway suggests that responses in addition to VEGF production may also be regulated. In this regard, a number of agents that increase cAMP have been demonstrated to inhibit several profibrotic responses of fibroblasts, including chemotaxis (21-23), proliferation (3,30,38), production of ECM (30,38), and contraction of 3-D collagen gels (17,23,28). Prostacyclin analogs, acting through a PKA-dependent pathway, have recently been demonstrated to inhibit fibroblastmediated contraction of 3-D collagen gels (17).…”

“…Prostacyclin analogs have been shown to inhibit the contraction of 3-D collagen gels in a concentration-dependent manner (17). However, the response of fibroblasts to exogenous mediators in 3-D culture can differ from the response in monolayer culture.…”

International Union of Basic and Clinical Pharmacology. LXXXIII: Classification of Prostanoid Receptors, Updating 15 Years of Progress

“…The IP receptor has long been known to be both G s -and G q -coupled, resulting in increased cAMP formation, PI turnover, and Ca 2ϩ signaling (Coleman et al, 1994;Namba et al, 1994;Narumiya et al, 1999). IP receptor activation of the cAMP-PKA pathway seems to be exclusively responsible for some prostacyclin-mediated events (Nasrallah et al, 2001;Ritchie et al, 2004;Kamio et al, 2007;Muja et al, 2007). Solution structure studies indicate that the first and third intracellular loops of the IP receptor are in contact with C-terminal residues of G␣ s to initiate cAMP signaling (Zhang et al, 2006a;Zhang et al, 2006b).…”