Essential role of protein kinase R in the pathogenesis of pulmonary veno-occlusive disease
Amit Prabhakar, Rahul Kumar, Meetu Wadhwa, Abhilash Barpanda, Joseph Lyons, Asavari Gowda, Simren Gupta, Ananyaa Arvind, Prajakta Ghatpande, Arun P. Wiita, Brian B. Graham, Giorgio Lagna, Akiko Hata
Amit Prabhakar, Rahul Kumar, Meetu Wadhwa, Abhilash Barpanda, Joseph Lyons, Asavari Gowda, Simren Gupta, Ananyaa Arvind, Prajakta Ghatpande, Arun P. Wiita, Brian B. Graham, Giorgio Lagna, Akiko Hata
View: Text | PDF
Research Article Cell biology Vascular biology

Essential role of protein kinase R in the pathogenesis of pulmonary veno-occlusive disease

Abstract

Pulmonary veno-occlusive disease (PVOD) is a rare and severe subtype of pulmonary arterial hypertension, characterized by progressive remodeling of small pulmonary arteries and veins with no therapies. Using a mitomycin C–induced (MMC-induced) rat model, we previously demonstrated that protein kinase R–mediated (PKR-mediated) integrated stress response (ISR) drives endothelial dysfunction and vascular remodeling. To determine whether PKR is the primary mediator of ISR and the pathogenesis, we treated control (Ctrl) and PKR-knockout (KO) mice with the same dose of MMC. Consistent with rat data, Ctrl mice displayed ISR activation, vascular remodeling, and pulmonary hypertension after MMC treatment, while KO mice showed none of these phenotypes. Proteomic analysis revealed that MMC-mediated ISR activation attenuated protein synthesis in Ctrl but not in KO mice. These findings underscore the critical role of PKR-dependent ISR activation and subsequent perturbation of proteostasis as central mechanisms driving PVOD pathogenesis and identify PKR as a promising therapeutic target.

Authors

Amit Prabhakar, Rahul Kumar, Meetu Wadhwa, Abhilash Barpanda, Joseph Lyons, Asavari Gowda, Simren Gupta, Ananyaa Arvind, Prajakta Ghatpande, Arun P. Wiita, Brian B. Graham, Giorgio Lagna, Akiko Hata

×

Figure 3

MMC treatment does not impair pulmonary vascular endothelium in PKR-deficient mice following MMC treatment.

Options: View larger image (or click on image) Download as PowerPoint
MMC treatment does not impair pulmonary vascular endothelium in PKR-defi...
(A) PVECs were isolated from the lungs of Ctrl and KO mice administered with vehicle or MMC and subjected to IF staining with anti–VE-Cad antibody (green) and DAPI (blue) for nuclei. Scale bars: 10 μm. (B) PVECs isolated from the mice administered vehicle or MMC were subjected to IF staining for p-PKR (green, left), ATF4 (red, right), and with DAPI (blue) for nuclei. Scale bars: 10 μm. (C) The plasma isolated from Veh- or MMC-treated Ctrl and KO mice on day 5 were subjected to IP with an anti-Rad51 antibody or nonspecific IgG (control), followed by immunoblot analysis with an anti–VE-Cad (for VRC) and anti-Rad51 antibody to detect the interaction between these proteins. Immunoblot with an anti-transferrin antibody (TF) is shown as loading control. n = 2 plasma samples from Veh-treated Ctrl or KO mice and 3 plasma samples from MMC-treated Ctrl or KO mice (left). The relative amounts of the indicated proteins normalized to TF are demonstrated as mean ± SEM (right). n = 4 plasma samples from Veh-treated Ctrl or KO mice and 6 plasma samples from MMC-treated Ctrl or KO mice (left). Statistical analysis was performed using 2-way ANOVA with Tukey’s multiple-comparison test.