Abstract

Background Pulmonary arterial hypertension (PAH) is a deadly disease without effective non-invasive diagnostic and prognostic testing. Inflammatory activation is a part of PAH pathogenesis. However, questions exist regarding whether vasodilator therapy reverses such reprogramming. Single-cell profiling of inflammatory cells in blood could clarify these mechanisms of PAH and be leveraged to advance clinical testing.

Methods We evaluated a UPMC cohort consisting of idiopathic PAH and scleroderma associated PAH (iPAH and sscPAH) patients and non-PAH controls. We performed single cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) from non-diseased control individuals (N=3) as well as from individuals with PAH (iPAH and sscPAH), naïve to treatment (N=4), 3 months after phosphodiesterase 5 inhibitor (PDE5i) treatment (N=7), and 3 months after treatment with PDE5i plus an endothelin receptor antagonist, macitentan (N=6). We compared the transcriptional profiles of PBMCs from iPAH and sscPAH patients in each treatment group in 5 different PBMC cell types to observe their responses to PDE5i and macitentan. To bolster our single cell analyses of differential gene expression, we utilized network analysis to illuminate the altered connectivity of biological networks in this complex disease and its responses to treatment.

Results We defined differential gene expression and perturbed network connectivity in immune cell populations of PAH patients following either PDE5i or combination of PDE5i and macitentan. We identified differences in iPAH and sscPAH patients’ baseline characteristics and response to PDE5i and macitentan. Importantly, we identified significant inflammatory reversal of key transcripts and pathways in the combined PAH patient cohort after vasodilator therapy in every PBMC type assessed. Namely, the ‘Glucagon Signaling in Metabolic Regulation’ pathway in monocytes was reversed after vasodilator therapy via two independent data analysis modalities.

Conclusion Via a systems biology scRNA-seq approach, we define inflammatory reprogramming in blood of PAH patients and the anti-inflammatory activity of vasodilator drugs in this deadly disease. Such findings establish diagnostic and prognostic blood-based tools of single cell resolution for tracking inflammatory progression of PAH and response to therapy.

Principal Investigators

Stephen Y. Chan, Vascular Medicine Institute at the University of Pittsburgh School of Medicine

Seungchan Kim, Electrical and Computer Engineering Dept., Roy G. Perry College of Engineering, Praie View A&M University

Satoshi Okawa, Vascular Medicine Institute at the University of Pittsburgh School of Medicine