Frataxin deficiency promotes endothelial senescence in pulmonary hypertension
Miranda K. Culley, … , Thomas Bertero, Stephen Y. Chan
Miranda K. Culley, … , Thomas Bertero, Stephen Y. Chan
Published April 27, 2021
Citation Information: J Clin Invest. 2021;131(11):e136459. https://doi.org/10.1172/JCI136459.
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Research Article Pulmonology Vascular biology

Frataxin deficiency promotes endothelial senescence in pulmonary hypertension

Abstract

The dynamic regulation of endothelial pathophenotypes in pulmonary hypertension (PH) remains undefined. Cellular senescence is linked to PH with intracardiac shunts; however, its regulation across PH subtypes is unknown. Since endothelial deficiency of iron-sulfur (Fe-S) clusters is pathogenic in PH, we hypothesized that a Fe-S biogenesis protein, frataxin (FXN), controls endothelial senescence. An endothelial subpopulation in rodent and patient lungs across PH subtypes exhibited reduced FXN and elevated senescence. In vitro, hypoxic and inflammatory FXN deficiency abrogated activity of endothelial Fe-S–containing polymerases, promoting replication stress, DNA damage response, and senescence. This was also observed in stem cell–derived endothelial cells from Friedreich’s ataxia (FRDA), a genetic disease of FXN deficiency, ataxia, and cardiomyopathy, often with PH. In vivo, FXN deficiency–dependent senescence drove vessel inflammation, remodeling, and PH, whereas pharmacologic removal of senescent cells in Fxn-deficient rodents ameliorated PH. These data offer a model of endothelial biology in PH, where FXN deficiency generates a senescent endothelial subpopulation, promoting vascular inflammatory and proliferative signals in other cells to drive disease. These findings also establish an endothelial etiology for PH in FRDA and left heart disease and support therapeutic development of senolytic drugs, reversing effects of Fe-S deficiency across PH subtypes.

Authors

Miranda K. Culley, Jingsi Zhao, Yi Yin Tai, Ying Tang, Dror Perk, Vinny Negi, Qiujun Yu, Chen-Shan C. Woodcock, Adam Handen, Gil Speyer, Seungchan Kim, Yen-Chun Lai, Taijyu Satoh, Annie M.M. Watson, Yassmin Al Aaraj, John Sembrat, Mauricio Rojas, Dmitry Goncharov, Elena A. Goncharova, Omar F. Khan, Daniel G. Anderson, James E. Dahlman, Aditi U. Gurkar, Robert Lafyatis, Ahmed U. Fayyaz, Margaret M. Redfield, Mark T. Gladwin, Marlene Rabinovitch, Mingxia Gu, Thomas Bertero, Stephen Y. Chan

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Figure 5

Acute FXN knockdown promotes replication stress and S-phase arrest.

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Acute FXN knockdown promotes replication stress and S-phase arrest. (A–G...
(A–G) All experiments were performed 48 hours after transfection in PAECs with or without FXN inhibition by siRNA. (A) Colorimetric BrdU incorporation (n = 6/group). (B) Manual PAEC count (n = 3/group). (C) Flow cytometric analysis of FXN-deficient or control PAECs pulsed with BrdU and the DNA marker 7-AAD (n = 6/group). (D) Immunoblot and quantification of the replication stress marker, phosphorylated RPA32 (p-RPA32) (n = 3/group). (E) Representative confocal imaging and quantification of replication rate (kb/ min) in FXN-deficient or control PAECs pulsed with CldU (20 minutes, 50 μM; green) followed by IdU (20 minutes, 250 μM; red) with hydroxyurea (2 mM; HU) (n = 175 versus n = 204 forks). Scale bar: 10 μm. (F) Quantified immunoblot of DNA damage response markers (p-ATR, CHK1, Ub-γH2AX) (n = 3/group). (G) Immunofluorescence staining and confocal microscopy of nuclear 53BP1 foci (red) within DAPI-stained nuclei (blue) (n = 37 versus n = 34). Scale bar: 10 μm. Two-tailed Student’s t test with error bars that reflect mean ± SD.