Gene therapy targeting survivin selectively induces pulmonary vascular apoptosis and reverses pulmonary arterial hypertension
M. Sean McMurtry, … , Lakshmi Puttagunta, Evangelos D. Michelakis
M. Sean McMurtry, … , Lakshmi Puttagunta, Evangelos D. Michelakis
Published June 1, 2005
Citation Information: J Clin Invest. 2005;115(6):1479-1491. https://doi.org/10.1172/JCI23203.
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Gene therapy targeting survivin selectively induces pulmonary vascular apoptosis and reverses pulmonary arterial hypertension

Abstract

Pulmonary arterial hypertension (PAH) is characterized by genetic and acquired abnormalities that suppress apoptosis and enhance cell proliferation in the vascular wall, including downregulation of the bone morphogenetic protein axis and voltage-gated K+ (Kv) channels. Survivin is an “inhibitor of apoptosis” protein, previously thought to be expressed primarily in cancer cells. We found that survivin was expressed in the pulmonary arteries (PAs) of 6 patients with PAH and rats with monocrotaline-induced PAH, but not in the PAs of 3 patients and rats without PAH. Gene therapy with inhalation of an adenovirus carrying a phosphorylation-deficient survivin mutant with dominant-negative properties reversed established monocrotaline-induced PAH and prolonged survival by 25%. The survivin mutant lowered pulmonary vascular resistance, RV hypertrophy, and PA medial hypertrophy. Both in vitro and in vivo, inhibition of survivin induced PA smooth muscle cell apoptosis, decreased proliferation, depolarized mitochondria, caused efflux of cytochrome c in the cytoplasm and translocation of apoptosis-inducing factor into the nucleus, and increased Kv channel current; the opposite effects were observed with gene transfer of WT survivin, both in vivo and in vitro. Inhibition of the inappropriate expression of survivin that accompanies human and experimental PAH is a novel therapeutic strategy that acts by inducing vascular mitochondria-dependent apoptosis.

Authors

M. Sean McMurtry, Stephen L. Archer, Dario C. Altieri, Sebastien Bonnet, Alois Haromy, Gwyneth Harry, Sandra Bonnet, Lakshmi Puttagunta, Evangelos D. Michelakis

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Figure 4

Ad-GFP-S-M infection induces PASMC mitochondria-dependent apoptosis.

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Ad-GFP-S-M infection induces PASMC mitochondria-dependent apoptosis. (A)...
(A) PASMCs effectively infected with Ad-GFP-S (green) show slight hyperpolarization of mitochondrial membrane potential (increased TMRM fluorescence) compared with noninfected cells. In contrast, mitochondria of Ad-GFP-S-M–infected cells (but not neighboring noninfected cells) are less red, indicating depolarized mitochondria. Mean data are shown on the right (arbitrary fluorescence units [FU], means from 15 plates per group; *P < 0.01). Immunoblots show that, in contrast to expression of WT survivin, expression of survivin mutant in PASMCs grown in 10% FBS induces activation of caspase-9 and caspase-3. (B) PASMCs infected with Ad-GFP-S demonstrate sequestered cytochrome c within mitochondria, as shown by the punctate pattern of staining. In contrast, mitochondria of Ad-GFP-S-M–infected cells show cytochrome c–positive staining diffusely throughout the cell, indicating leakage of cytochrome c from mitochondria into the cytoplasm. Magnification: left and middle panels, ×75; right panels, ×125. (C) In contrast to infection with Ad-GFP-S, infection with Ad-GFP-S-M induces translocation of the mitochondria-based apoptosis-inducing factor (AIF) in the nucleus, where it initiates caspase-independent apoptosis. Magnification, ×100.