Genetic ablation of Nrf2 enhances susceptibility to cigarette smoke–induced emphysema in mice
Tirumalai Rangasamy, … , Rubin M. Tuder, Shyam Biswal
Tirumalai Rangasamy, … , Rubin M. Tuder, Shyam Biswal
Published November 1, 2004
Citation Information: J Clin Invest. 2004;114(9):1248-1259. https://doi.org/10.1172/JCI21146.
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Genetic ablation of Nrf2 enhances susceptibility to cigarette smoke–induced emphysema in mice

Abstract

Although inflammation and protease/antiprotease imbalance have been postulated to be critical in cigarette smoke–induced (CS-induced) emphysema, oxidative stress has been suspected to play an important role in chronic obstructive pulmonary diseases. Susceptibility of the lung to oxidative injury, such as that originating from inhalation of CS, depends largely on its upregulation of antioxidant systems. Nuclear factor, erythroid-derived 2, like 2 (Nrf2) is a redox-sensitive basic leucine zipper protein transcription factor that is involved in the regulation of many detoxification and antioxidant genes. Disruption of the Nrf2 gene in mice led to earlier-onset and more extensive CS-induced emphysema than was found in wild-type littermates. Emphysema in Nrf2-deficient mice exposed to CS for 6 months was associated with more pronounced bronchoalveolar inflammation; with enhanced alveolar expression of 8-oxo-7,8-dihydro-2′-deoxyguanosine, a marker of oxidative stress; and with an increased number of apoptotic alveolar septal cells — predominantly endothelial and type II epithelial cells — as compared with wild-type mice. Microarray analysis identified the expression of nearly 50 Nrf2-dependent antioxidant and cytoprotective genes in the lung that may work in concert to counteract CS-induced oxidative stress and inflammation. The responsiveness of the Nrf2 pathway may act as a major determinant of susceptibility to tobacco smoke–induced emphysema by upregulating antioxidant defenses and decreasing lung inflammation and alveolar cell apoptosis.

Authors

Tirumalai Rangasamy, Chung Y. Cho, Rajesh K. Thimmulappa, Lijie Zhen, Sorachai S. Srisuma, Thomas W. Kensler, Masayuki Yamamoto, Irina Petrache, Rubin M. Tuder, Shyam Biswal

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Figure 2

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Cigarette smoke exposure causes lung cell apoptosis as assessed by TUNEL...
Cigarette smoke exposure causes lung cell apoptosis as assessed by TUNEL in Nrf2–/– lungs. (A) Lung sections (n = 5 per group) of room air– or CS-exposed (6 months) Nrf2+/+ or Nrf2–/– mice were subjected to TUNEL (left column) and DAPI stain (middle column). Merged images are shown in the right column. CS-exposed Nrf2–/– mice show abundant TUNEL-positive cells (arrows) in the alveolar septa. Magnification, ×20. (B) Quantification of TUNEL-positive cells (per 1,000 DAPI-stained cells). The number of TUNEL-positive cells was significantly higher in the CS-exposed Nrf2–/– mice as compared with their wild-type counterparts (*P – 0.05). Values represent mean ± SEM. (C) Identification of apoptotic (TUNEL-positive) type II epithelial cells (left column), endothelial cells (middle column), and alveolar macrophages (right column) in the lungs of CS-exposed (6 months) Nrf2+/+ and Nrf2–/– mice. Type II epithelial cells, endothelial cells, and alveolar macrophages were detected with anti-SpC, anti-CD34, and anti–Mac-3 antibodies, respectively, as outlined in Methods. Nuclei were detected with DAPI (blue). Shown are the merged images, with colocalization (yellow arrows) of cell-specific markers (cytoplasmic red signal) and apoptosis (nuclear green + blue DAPI signal, resulting in a lavender-like signal); non-apoptotic (TUNEL-negative) cells with positive cell specific marker (red signal) are highlighted with a red arrow. TUNEL-positive apoptotic cells lacking a cell-specific marker are highlighted by white arrowheads. The majority of TUNEL-positive cells consisted of endothelial and type II epithelial cells, whereas most alveolar macrophages were TUNEL negative. Scale bars: 5 μm.