Conference: 2018 PHA International PH Conference & Scientific Sessions
Release Date: 07.28.2018
Presentation Type: Abstracts
File Download: Conference 2018_1003
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Abstract presented at the 2018 International PH Conference and Scientific Sessions in Orlando, Fla., June 28-July 1, 2018.
Bardoxolone methyl (BARD) is an Nrf2 activator currently under investigation in the LARIAT and CATALYST clinical trials for the treatment of connective tissue disease associated pulmonary arterial hypertension (CTD-PAH). Vasoconstriction is an important feature of PAH and several drugs with vasodilatory activity have been approved for PAH treatment. However, alterations in other molecular and cellular processes –including inflammation, mitochondrial metabolism, oxidative stress, abnormal proliferation, and fibrosis–also play key roles in PAH and underlie tissue damage and remodeling. Nrf2 activation is known to promote the resolution of inflammation, improve mitochondrial function, and reduce oxidative stress. In this study, we asked whether the Nrf2 activator BARD improves the molecular and cellular parameters associated with PAH.
THP-1 monocytes and RAW 264.7 macrophages were stimulated with IFNγ ± LPS to activate inflammatory signaling pathways. Mitochondrial function was assessed in RAW 264.7 macrophages and C2C12 skeletal muscle cells using Seahorse XF Technology. Reactive oxygen species were measured in H9c2 cardiomyocytes and C2C12 skeletal muscle cells using fluorescent molecular probes. Pulmonary arterial smooth muscle cells (PASMCs) were cultured under normoxic (21% O2) or hypoxic (2% O2) conditions and hypoxia-induced cell proliferation was assessed by BrdU incorporation. Pulmonary artery endothelial cells (PAECs) were treated with TGFβ, IL-1β, and TNFα to induce endothelial to mesenchymal transition (EndMT) and fibrosis. Expression of Nrf2 target genes, pro-inflammatory cytokines, and markers of EndMT was measured by reverse transcription and quantitative PCR. Protein levels of pro-inflammatory cytokines were measured by ELISA or HTRF.
Treatment with BARD increased the expression of Nrf2 target genes in monocytes, macrophages, PAECs, PASMCs, cardiomyocytes, and skeletal muscle cells. At the same concentrations that increased Nrf2 activity, BARD suppressed the levels of pro-inflammatory cytokines in several cell types. BARD restored compromised mitochondrial function in activated macrophages and skeletal muscle cells. In addition, BARD reduced reactive oxygen species in skeletal muscle cells and cardiomyocytes. Treatment with BARD prevented hypoxia-induced proliferation in PASMCs without affecting their viability. Finally, BARD inhibited markers of EndMT and fibrosis in PAECs treated with TGFβ, IL-1β, and TNFα.
Taken together, these results demonstrate that BARD may have the potential to improve multiple dysfunctional molecular and cellular parameters that are associated with pulmonary arterial hypertension.