Dysregulated zinc and sphingosine-1-phosphate signalling in pulmonary hypertension: potential effects by targeting of bone morphogenetic protein receptor type 2 in pulmonary microvessels

Abstract

Recently identified molecular targets in pulmonary artery hypertension (PAH) include sphingosine-1-phosphate (S1P) and zinc transporter ZIP12 signalling. This study sought to determine linkages between these pathways, and with BMPR2 signalling. Lung tissues from a rat model of monocrotaline-induced PAH and therapeutic treatment with bone marrow-derived endothelial-like progenitor cells transduced to over-express BMPR2 were studied. Multifluorescence quantitative confocal microscopy (MQCM) was applied for analysis of protein expression and localisation of markers of vascular remodelling (αSMA and BMPR2), parameters of zinc homeostasis (zinc transporter SLC39A/ZIP family members 1, 10, 12 and 14; and metallothionein MT3) and S1P extracellular signalling (SPHK1, SPNS2, S1P receptor isoforms 1, 2, 3, 5) in 20-200µm pulmonary microvessels. ZIP12 expression in whole lung tissue lysates was assessed by Western blot. Spearman nonparametric correlations between MQCM readouts and haemodynamic parameters, Fulton index (FI) and right ventricular systolic pressure (RVSP) were measured. In line with PAH status, pulmonary microvessels in monocrotaline-treated animals demonstrated significant (p<0.05, n=6 per group) upregulation of αSMA (two fold) and downregulation of BMPR2 (20%). Upregulated ZIP12 (92%), MT3 (57.7%), S1PR2 (54.8%) and S1PR3 (30.3%) were also observed. Significant positive and negative correlations were demonstrated between parameters of zinc homeostasis (ZIP12, MT3), S1P signalling (S1PRs, SPNS2), and vascular remodelling (αSMA, FI, RVSP). MQCM and Western blot analysis showed that monocrotaline-induced ZIP12 upregulation could be partially negated by BMPR2-targeted therapy. Our results indicate that altered zinc transport/storage and S1P signalling in the monocrotaline-induced PAH rat model are linked to each other, and could be alleviated by BMPR2-targeted therapy. This article is protected by copyright. All rights reserved.