Dopamine Synthesis and D1 Receptor Agonism are Essential for Resolution of Experimental Lung Fibrosis

Abstract

Rationale Idiopathic Pulmonary fibrosis (IPF) is characterized by progressive and ultimately fatal end-stage lung scarring associated with uncontrolled deposition, stiffening and remodeling of fibrous connective tissue proteins. We have previously shown that agonism of the dopamine receptor D1 attenuates experimental lung fibrosis in mice, and that the lungs of IPF patients display reduced transcript and protein levels of DOPA decarboxylase (DDC), the enzyme that catalyzes the biosynthesis of dopamine. Here we investigate the role of endogenous dopamine production and signaling in the resolution of experimental lung fibrosis. Methods Using a self-resolving model of experimental lung fibrosis, we administered bleomycin (1.1U/kg) to 10-week old mice intratracheally at day 0, and collected lung tissue from sham and bleo groups at day 21 to establish the level of baseline fibrosis. We then administered the DDC inhibitor carbidopa (20mg/kg, IP), the D1 receptor antagonist SCH 39166 (0.5mg/kg, SC), or vehicle control daily from day 21 to 42. Lung tissue was collected for hydroxyproline and histological analysis. In order to investigate dopamine synthesis and signaling ex vivo, we generated precision-cut lung slices (PCLS) from healthy mice and stimulated them +/- TGFβ for 72 hours and collected total RNA from the lung slices and conditioned media to measure dopamine production by ELISA. Results As previously observed, lung tissue collected at day 21 post injury displayed enhanced markers of fibrosis measured by hydroxyproline (478% increase over sham) and H&E stained histology. By day 42 the vehicle control mice resolved this fibrosis. In contrast, the mice treated with carbidopa or SCH 39166 maintained the fibrosis observed at day 21 (514% and 592% increase compared to sham, respectively). Consistent with our prior observation in IPF patient lung, PCLS cultured ex vivo with TGFβ expressed significantly less Ddc (75% reduction compared to control) and total dopamine abundance in the conditioned media dropped from ~120ng/mL to ~32ng/mL. Conclusions Dopamine synthesis through the activity of DDC, and agonism of the dopamine receptor D1 are essential to pulmonary fibrosis resolution after injury. PCLS represents a useful model to further investigate the molecular mechanisms leading to reduced expression of DDC, and test pharmacological approaches to stimulate dopamine signaling as a potential therapy for IPF.