Cigarette smoke-induced autophagy is regulated by SIRT1–PARP-1-dependent mechanism: Implication in pathogenesis of COPD
Introduction
Cigarette smoke (CS)1 contains numerous oxidants/free radicals as well as chemical compounds that induce oxidative stress and are involved in the pathogenesis of lung and heart diseases. CS accelerates cell death and senescence both through direct mechanisms mediated by oxidants/free radicals as well as via generation of oxidants from inflammatory cells in chronic inflammatory diseases including chronic obstructive pulmonary disease (COPD) and cardiovascular co-morbidities [1], [2]. Although there is emerging evidence that CS-mediated cell death and senescence increase the susceptibility to diseases, the exact mechanism by which cigarette smoking accelerates cell death and senescence remains unclear.
Autophagy is a fundamental cellular process that eliminates long-lived proteins and damaged organelles through a lysosomal degradation pathway, and has been suggested to have an essential function in maintaining cellular homeostasis [3]. The autophagic process is initiated by sequestering redundant cytoplasmic contents within double-membrane structures termed autophagosomes. The autophagosome fuses with a lysosome, and its contents are degraded and recycled [4]. Although autophagy occurs at basal levels in all cells to maintain cellular homeostasis, recent reports show that autophagy is also induced in response to environmental stresses, such as pathogen infections, starvation and oxidative stress [4], [5], [6], [7]. Although, autophagy plays a protective role in overcoming the exogenous stress, prolonged and excessive autophagy can lead to cell death [8], [9]. Failure to regulate autophagy has been implicated in pathogenesis of cancer, cardiovascular failure, immune disease, skeletal muscle atrophy and neurodegenerative disorders [10], [11], [12], [13], [14], [15], [16], [17]. Recent studies have shown that increased autophagy occurs in lungs of patients with COPD and in lung cells of mouse exposed to CS [18], [19]. However, the underlying mechanism for CS-induced autophagy was not studied.
Sirtuin 1 (SIRT1), the mammalian ortholog of yeast silent information regulator 2 (Sir2), is an NAD+-dependent deacetylase which is shown to be an anti-inflammatory and anti-aging protein [20]. SIRT1 is involved in diverse physiological functions, including gene silencing, stress resistance, apoptosis, inflammation, senescence and aging [21], [22], [23], [24]. These physiological functions of SIRT1 are mediated by deacetylation of histones and several important transcription factors such as forkhead box O3 (FoxO3), p53 and nuclear factor-κB (NF-κB) [20], [22], [23], [25], [26], [27]. SIRT1 activity is also regulated by NAD+ depletion induced by oxidative stress or activation of the NAD+-dependent enzyme poly(ADP-ribose)-polymerase-1 (PARP-1) [28], [29]. It has recently been shown that SIRT1 regulates autophagy under calorie restriction/starvation [30]. Moreover, we have recently shown that SIRT1 levels/activity is decreased in response to CS exposure in vitro in macrophages and epithelial cells as well as in lungs of smokers and patients with COPD [20], [24], [28]. However, the role of SIRT1 and PARP-1 on CS-mediated autophagy is not known. Therefore, we hypothesized that SIRT1 plays an important role in regulating CS-mediated autophagy in lung cells. We studied the effect of CS on induction of autophagy in different lung cell types and macrophages in vitro and in mouse lung in vivo, and determined the role of SIRT1–PARP-1 axis in regulation of autophagy.
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Reagents
Penicillin–Streptomycin, l-glutamine and RPMI-1640 were obtained from Gibco BRL (Grand Island, NY). Fetal bovine serum (FBS) was obtained from HyClone Laboratories (Logan, UT). Dulbecco’s modified Eagle’s medium-Ham’s F12 50:50 mixture (DMEM-F12) was purchased from Mediatech (Manassas, VA). Amphotericin B was purchased from Lonza (Walkersville, MD). Resveratrol was purchased from Biomol (Plymouth Meeting, PA). Sirtinol was procured from Sigma (St. Louis, MO). 3-Aminobenzamide (3-AB) was
Cigarette smoke extract (CSE) induces autophagy in different lung cell types and macrophages
We investigated whether CSE could affect the induction of autophagy in different lung cell types (epithelial cells and fibroblasts), and in macrophages. Treatment of human bronchial epithelial cells (H292) with CSE caused a dose- and time-dependent increase in the conversion of LC3-I to LC3-II, a hallmark of autophagic activity (Fig. 1A) [37]. At the concentration of 1% CSE, approximately 5-fold increase in the amount of LC3-II/LC3-I was found as compared to controls. CSE (1%) time-dependently
Discussion
Recent studies have reported that down-regulation of histone deacetylase (HDAC) activity can induce autophagy. HDAC inhibitors, such as sodium butyrate and suberoylanilide hydroxamic acid (SAHA) can induce autophagy [40], [41]. In addition, Chen et al. demonstrated that decreased HDAC activity in response to CS triggers autophagy [19]. Despite increasing reports of the association between decreased HDAC activity and induction of autophagy, little is known about the relationship between
Acknowledgments
This work was supported by NIH1R01HL085613, 1R01HL097751, 1R01HL092842, and NIEHSES01247.
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