PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-25 (37)
 

Clipboard (0)
None

Select a Filter Below

Year of Publication
1.  Carbon Monoxide in Exhaled Breath Testing and Therapeutics 
Journal of breath research  2013;7(1):017111.
Carbon monoxide (CO), a low molecular weight gas, is a ubiquitous environmental product of organic combustion, which is also produced endogenously in the body, as the byproduct of heme metabolism. CO binds to hemoglobin, resulting in decreased oxygen delivery to bodily tissues at toxicological concentrations. At physiological concentrations, CO may have endogenous roles as a potential signaling mediator in vascular function and cellular homeostasis. Exhaled CO (eCO), similar to exhaled nitric oxide (eNO), has been evaluated as a candidate breath biomarker of pathophysiological states, including smoking status, and inflammatory diseases of the lung and other organs. eCO values have been evaluated as potential indicators of inflammation in asthma, stable COPD and exacerbations, cystic fibrosis, lung cancer, or during surgery or critical care. The utility of eCO as a marker of inflammation, and potential diagnostic value remains incompletely characterized. Among other candidate “medicinal gases” with therapeutic potential, (e.g., NO and H2S), CO has been shown to act as an effective anti-inflammatory agent in preclinical animal models of inflammatory disease, acute lung injury, sepsis, ischemia/reperfusion injury and organ graft rejection. Current and future clinical trials will evaluate the clinical applicability of this gas as a biomarker and/or therapeutic in human disease.
doi:10.1088/1752-7155/7/1/017111
PMCID: PMC3651886  PMID: 23446063
Carbon Monoxide; Exhaled Breath; Heme Oxygenase-1
2.  The Impact of Autophagy on Cell Death Modalities 
Autophagy represents a homeostatic cellular mechanism for the turnover of organelles and proteins, through a lysosome-dependent degradation pathway. During starvation, autophagy facilitates cell survival through the recycling of metabolic precursors. Additionally, autophagy can modulate other vital processes such as programmed cell death (e.g., apoptosis), inflammation, and adaptive immune mechanisms and thereby influence disease pathogenesis. Selective pathways can target distinct cargoes (e.g., mitochondria and proteins) for autophagic degradation. At present, the causal relationship between autophagy and various forms of regulated or nonregulated cell death remains unclear. Autophagy can occur in association with necrosis-like cell death triggered by caspase inhibition. Autophagy and apoptosis have been shown to be coincident or antagonistic, depending on experimental context, and share cross-talk between signal transduction elements. Autophagy may modulate the outcome of other regulated forms of cell death such as necroptosis. Recent advances suggest that autophagy can dampen inflammatory responses, including inflammasome-dependent caspase-1 activation and maturation of proinflammatory cytokines. Autophagy may also act as regulator of caspase-1 dependent cell death (pyroptosis). Strategies aimed at modulating autophagy may lead to therapeutic interventions for diseases in which apoptosis or other forms of regulated cell death may play a cardinal role.
doi:10.1155/2014/502676
PMCID: PMC3932252  PMID: 24639873
3.  Autophagic proteins 
Autophagy  2012;8(3):426-428.
Oxygen (O2), while essential for aerobic life, can also cause metabolic toxicity through the excess generation of reactive oxygen species (ROS). Pathological changes in ROS production can originate through the partial reduction of O2 during mitochondrial electron transport, as well as from enzymatic sources. This phenomenon, termed the oxygen paradox, has been implicated in aging and disease, and is especially evident in critical care medicine. Whereas high O2 concentrations are utilized as a life-sustaining therapeutic for respiratory insufficiency, they in turn can cause acute lung injury. Alveolar epithelial cells represent a primary target of hyperoxia-induced lung injury. Recent studies have indicated that epithelial cells exposed to high O2 concentrations die by apoptosis, or necrosis, and can also exhibit mixed-phenotypes of cell death (aponecrosis). Autophagy, a cellular homeostatic process responsible for the lysosomal turnover of organelles and proteins, has been implicated as a general response to oxidative stress in cells and tissues. This evolutionarily conserved process is finely regulated by a complex interplay of protein factors. During autophagy, senescent organelles and cellular proteins are sequestered in autophagic vacuoles (autophagosomes) and subsequently targeted to the lysosome, where they are degraded by lysosomal hydrolases, and the breakdown products released for reutilization in anabolic pathways. Autophagy has been implicated as a cell survival mechanism during nutrient-deficiency states, and more generally, as a determinant of cell fate. However, the mechanisms by which autophagy and/or autophagic proteins potentially interact with and/or regulate cell death pathways during high oxygen stress, remain only partially understood.
doi:10.4161/auto.19258
PMCID: PMC3337844  PMID: 22302001
acute lung injury; Apoptosis; autophagy; caveolin-1; Fas; hyperoxia; LC3B
4.  Carbon monoxide: present and future indications for a medical gas 
Gaseous molecules continue to hold new promise in molecular medicine as experimental and clinical therapeutics. The low molecular weight gas carbon monoxide (CO), and similar gaseous molecules (e.g., H2S, nitric oxide) have been implicated as potential inhalation therapies in inflammatory diseases. At high concentration, CO represents a toxic inhalation hazard, and is a common component of air pollution. CO is also produced endogenously as a product of heme degradation catalyzed by heme oxygenase enzymes. CO binds avidly to hemoglobin, causing hypoxemia and decreased oxygen delivery to tissues at high concentrations. At physiological concentrations, CO may have endogenous roles as a signal transduction molecule in the regulation of neural and vascular function and cellular homeostasis. CO has been demonstrated to act as an effective anti-inflammatory agent in preclinical animal models of inflammation, acute lung injury, sepsis, ischemia/reperfusion injury, and organ transplantation. Additional experimental indications for this gas include pulmonary fibrosis, pulmonary hypertension, metabolic diseases, and preeclampsia. The development of chemical CO releasing compounds constitutes a novel pharmaceutical approach to CO delivery with demonstrated effectiveness in sepsis models. Current and pending clinical evaluation will determine the usefulness of this gas as a therapeutic in human disease.
doi:10.3904/kjim.2013.28.2.123
PMCID: PMC3604600  PMID: 23525151
Acute lung injury; Carbon monoxide; Heme oxygenase (decyclizing); Reperfusion injury; Sepsis
5.  Carbon Monoxide Protects against Hepatic Ischemia/Reperfusion Injury via ROS-Dependent Akt Signaling and Inhibition of Glycogen Synthase Kinase 3β 
Carbon monoxide (CO) may exert important roles in physiological and pathophysiological states through the regulation of cellular signaling pathways. CO can protect organ tissues from ischemia/reperfusion (I/R) injury by modulating intracellular redox status and by inhibiting inflammatory, apoptotic, and proliferative responses. However, the cellular mechanisms underlying the protective effects of CO in organ I/R injury remain incompletely understood. In this study, a murine model of hepatic warm I/R injury was employed to assess the role of glycogen synthase kinase-3 (GSK3) and phosphatidylinositol 3-kinase (PI3K)-dependent signaling pathways in the protective effects of CO against inflammation and injury. Inhibition of GSK3 through the PI3K/Akt pathway played a crucial role in CO-mediated protection. CO treatment increased the phosphorylation of Akt and GSK3-beta (GSK3β) in the liver after I/R injury. Furthermore, administration of LY294002, an inhibitor of PI3K, compromised the protective effect of CO and decreased the level of phospho-GSK3β after I/R injury. These results suggest that CO protects against liver damage by maintaining GSK3β phosphorylation, which may be mediated by the PI3K/Akt signaling pathway. Our study provides additional support for the therapeutic potential of CO in organ injury and identifies GSK3β as a therapeutic target for CO in the amelioration of hepatic injury.
doi:10.1155/2013/306421
PMCID: PMC3880761  PMID: 24454979
6.  Autophagy: An Integral Component of the Mammalian Stress Response 
Mammalian cells and tissues respond to chemical and physical stress by inducing adaptive or protective mechanisms that prolong survival. Among these, the major stress inducible proteins (heat shock proteins, glucose regulated proteins, heme oxygenase-1) provide cellular protection through protein chaperone and/or anti-oxidative and anti-inflammatory functions. In recent years it has become clear that autophagy, a genetically-programmed and evolutionarily-conserved cellular process represents another adaptive response to cellular stress. During autophagy cytosolic material, including organelles, proteins, and foreign pathogens, are sequestered into membrane-bound vesicles termed autophagosomes, and then delivered to the lysosome for degradation. Through recycling of cellular biochemicals, autophagy provides a mechanism for adaptation to starvation. Recent research has uncovered selective autophagic pathways that target distinct cargoes to autophagosomes, including mechanisms for the clearance of aggregated protein, and for the removal of dysfunctional mitochondria (mitophagy). Autophagy can be induced by multiple forms of chemical and physical stress, including endoplasmic reticulum stress and oxidative stress, and plays an integral role in the mammalian stress response. Understanding of the interaction and co-regulation of autophagy with other stress-inducible systems will be useful in the design and implementation of therapeutics targeting this pathway.
PMCID: PMC3865984  PMID: 24358454
apoptosis; autophagy; endoplasmic reticulum; mitochondria; oxidative stress; stress proteins
7.  Heme Oxygenase-1 as a Novel Metabolic Player 
doi:10.1155/2013/814058
PMCID: PMC3871917  PMID: 24381718
8.  Bile Pigments in Pulmonary and Vascular Disease 
The bile pigments, biliverdin, and bilirubin, are endogenously derived substances generated during enzymatic heme degradation. These compounds have been shown to act as chemical antioxidants in vitro. Bilirubin formed in tissues circulates in the serum, prior to undergoing hepatic conjugation and biliary excretion. The excess production of bilirubin has been associated with neurotoxicity, in particular to the newborn. Nevertheless, clinical evidence suggests that mild states of hyperbilirubinemia may be beneficial in protecting against cardiovascular disease in adults. Pharmacological application of either bilirubin and/or its biological precursor biliverdin, can provide therapeutic benefit in several animal models of cardiovascular and pulmonary disease. Furthermore, biliverdin and bilirubin can confer protection against ischemia/reperfusion injury and graft rejection secondary to organ transplantation in animal models. Several possible mechanisms for these effects have been proposed, including direct antioxidant and scavenging effects, and modulation of signaling pathways regulating inflammation, apoptosis, cell proliferation, and immune responses. The practicality and therapeutic-effectiveness of bile pigment application to humans remains unclear.
doi:10.3389/fphar.2012.00039
PMCID: PMC3296960  PMID: 22408625
antioxidant; biliverdin; bilirubin; cardiovascular disease; pulmonary disease
9.  Therapeutic Potential of Heme Oxygenase-1/Carbon Monoxide in Lung Disease 
Heme oxygenase (HO), a catabolic enzyme, provides the rate-limiting step in the oxidative breakdown of heme, to generate carbon monoxide (CO), iron, and biliverdin-IXα. Induction of the inducible form, HO-1, in tissues is generally regarded as a protective mechanism. Over the last decade, considerable progress has been made in defining the therapeutic potential of HO-1 in a number of preclinical models of lung tissue injury and disease. Likewise, tissue-protective effects of CO, when applied at low concentration, have been observed in many of these models. Recent studies have expanded this concept to include chemical CO-releasing molecules (CORMs). Collectively, salutary effects of the HO-1/CO system have been demonstrated in lung inflammation/acute lung injury, lung and vascular transplantation, sepsis, and pulmonary hypertension models. The beneficial effects of HO-1/CO are conveyed in part through the inhibition or modulation of inflammatory, apoptotic, and proliferative processes. Recent advances, however, suggest that the regulation of autophagy and the preservation of mitochondrial homeostasis may serve as additional candidate mechanisms. Further preclinical and clinical trials are needed to ascertain the therapeutic potential of HO-1/CO in human clinical disease.
doi:10.1155/2012/859235
PMCID: PMC3296197  PMID: 22518295
10.  Autophagy in Inflammatory Diseases 
Autophagy provides a mechanism for the turnover of cellular organelles and proteins through a lysosome-dependent degradation pathway. During starvation, autophagy exerts a homeostatic function that promotes cell survival by recycling metabolic precursors. Additionally, autophagy can interact with other vital processes such as programmed cell death, inflammation, and adaptive immune mechanisms, and thereby potentially influence disease pathogenesis. Macrophages deficient in autophagic proteins display enhanced caspase-1-dependent proinflammatory cytokine production and the activation of the inflammasome. Autophagy provides a functional role in infectious diseases and sepsis by promoting intracellular bacterial clearance. Mutations in autophagy-related genes, leading to loss of autophagic function, have been implicated in the pathogenesis of Crohn's disease. Furthermore, autophagy-dependent mechanisms have been proposed in the pathogenesis of several pulmonary diseases that involve inflammation, including cystic fibrosis and pulmonary hypertension. Strategies aimed at modulating autophagy may lead to therapeutic interventions for diseases associated with inflammation.
doi:10.1155/2011/732798
PMCID: PMC3235460  PMID: 22190939
11.  Quercetin Induces Mitochondrial Biogenesis through Activation of HO-1 in HepG2 Cells 
The regeneration of mitochondria by regulated biogenesis plays an important homeostatic role in cells and tissues and furthermore may provide an adaptive mechanism in certain diseases such as sepsis. The heme oxygenase (HO-1)/carbon monoxide (CO) system is an inducible cytoprotective mechanism in mammalian cells. Natural antioxidants can provide therapeutic benefit, in part, by inducing the HO-1/CO system. This study focused on the mechanism by which the natural antioxidant quercetin can induce mitochondrial biogenesis in HepG2 cells. We found that quercetin treatment induced expression of mitochondrial biogenesis activators (PGC-1α, NRF-1, TFAM), mitochondrial DNA (mtDNA), and proteins (COX IV) in HepG2 cells. The HO inhibitor SnPP and the CO scavenger hemoglobin reversed the effects of quercetin on mitochondrial biogenesis in HepG2 cells. The stimulatory effects of quercetin on mitochondrial biogenesis could be recapitulated in vivo in liver tissue and antagonized by SnPP. Finally, quercetin conferred an anti-inflammatory effect in the liver of mice treated with LPS and prevented impairment of mitochondrial biogenesis by LPS in vivo. These salutary effects of quercetin in vivo were also antagonized by SnPP. Thus, our results suggest that quercetin enhances mitochondrial biogenesis mainly via the HO-1/CO system in vitro and in vivo. The beneficial effects of quercetin may provide a therapeutic basis in inflammatory diseases and sepsis.
doi:10.1155/2013/154279
PMCID: PMC3833383  PMID: 24288584
12.  Deacetylation of p53 induces autophagy by suppressing Bmf expression 
The Journal of Cell Biology  2013;201(3):427-437.
IFN-γ induces the interaction of HDAC1 and p53, leading to p53 deacetylation, which facilitates autophagy via Bmf suppression.
Interferon γ (IFN-γ)–induced cell death is mediated by the BH3-only domain protein, Bik, in a p53-independent manner. However, the effect of IFN-γ on p53 and how this affects autophagy have not been reported. The present study demonstrates that IFN-γ down-regulated expression of the BH3 domain-only protein, Bmf, in human and mouse airway epithelial cells in a p53-dependent manner. p53 also suppressed Bmf expression in response to other cell death–stimulating agents, including ultraviolet radiation and histone deacetylase inhibitors. IFN-γ did not affect Bmf messenger RNA half-life but increased nuclear p53 levels and the interaction of p53 with the Bmf promoter. IFN-γ–induced interaction of HDAC1 and p53 resulted in the deacetylation of p53 and suppression of Bmf expression independent of p53’s proline-rich domain. Suppression of Bmf facilitated IFN-γ–induced autophagy by reducing the interaction of Beclin-1 and Bcl-2. Furthermore, autophagy was prominent in cultured bmf−/− but not in bmf+/+ cells. Collectively, these observations show that deacetylation of p53 suppresses Bmf expression and facilitates autophagy.
doi:10.1083/jcb.201205064
PMCID: PMC3639396  PMID: 23629966
13.  Beclin 1 deficiency is associated with increased hypoxia-induced angiogenesis 
Autophagy  2011;7(8):829-839.
Beclin 1, a tumor suppressor protein, acts as an initiator of autophagy in mammals. Heterozygous disruption of Beclin 1 accelerates tumor growth, but the underlying mechanisms remain unclear. We examined the role of Beclin 1 in tumor proliferation and angiogenesis, using a primary mouse melanoma tumor model. Beclin 1 (Becn1+/−) hemizygous mice displayed an aggressive tumor growth phenotype with increased angiogenesis under hypoxia, associated with enhanced levels of circulating erythropoietin but not vascular endothelial growth factor, relative to wild-type mice. Using in vivo and ex vivo assays, we demonstrated increased angiogenic activity in Becn1+/− mice relative to wild-type mice. Endothelial cells from Becn1+/− mice displayed increased proliferation, migration and tube formation in response to hypoxia relative to wild-type cells. Moreover, Becn1+/− cells subjected to hypoxia displayed increased hypoxia-inducible factor-2α (HIF-2α) expression relative to HIF-1α. Genetic interference of HIF-2α but not HIF-1α, dramatically reduced hypoxia-inducible proliferation, migration and tube formation in Becn1+/− endothelial cells. We demonstrated that mice deficient in the autophagic protein Beclin 1 display a pro-angiogenic phenotype associated with the upregulation of HIF-2α and increased erythropoietin production. These results suggest a relationship between Beclin 1 and the regulation of angiogenesis, with implications in tumor growth and development.
doi:10.4161/auto.7.8.15598
PMCID: PMC3149693  PMID: 21685724
angiogenesis; autophagy; beclin 1; hypoxia-inducible factor
14.  Heme Oxygenase-1, a Critical Arbitrator of Cell Death Pathways in Lung Injury and Disease 
Increases in cell death by programmed (ie., apoptosis, autophagy) or non-programmed mechanisms (ie., necrosis) occur during tissue injury, and may contribute to the etiology of several pulmonary or vascular disease states. The low molecular weight stress protein heme oxygenase-1 (HO-1) confers cytoprotection against cell death in various models of lung and vascular injury by inhibiting apoptosis, inflammation, and cell proliferation. HO-1 serves a vital metabolic function as the rate-limiting step in the heme degradation pathway and in the maintenance of iron homeostasis. The transcriptional induction of HO-1 occurs in response to multiple forms of chemical and physical cellular stress. The cytoprotective functions of HO-1 may be attributed to heme turnover, as well as to beneficial properties of its enzymatic reaction products: biliverdin-IXα, iron, and carbon monoxide (CO). Recent studies have demonstrated that HO-1 or CO inhibits stress-induced extrinsic and intrinsic apoptotic pathways in vitro. A variety of signaling molecules have been implicated in the cytoprotection conferred by HO-1/CO, including autophagic proteins, p38 mitogen activated protein kinase, signal transducer and activator of transcription proteins, nuclear factor-κB, phosphatydylinositol-3-kinase/Akt, and others. Enhanced HO-1 expression or the pharmacological application of HO end-products affords protection in preclinical models of tissue injury, including experimental and transplant-associated ischemia/reperfusion injury, promising potential future therapeutic applications.
doi:10.1016/j.freeradbiomed.2009.04.007
PMCID: PMC3078523  PMID: 19362144
15.  Autophagy in the Lung 
Autophagy is a cellular process for the disposal of damaged organelles or denatured proteins through a lysosomal degradation pathway. By reducing endogenous macromolecules to their basic components (i.e., amino acids, lipids), autophagy serves a homeostatic function by ensuring cell survival during starvation. Increased autophagy can be found in dying cells, although the relationships between autophagy and programmed cell death remain unclear. To date, few studies have examined the regulation and functional significance of autophagy in human lung disease. The lung, a complex organ that functions primarily in gas exchange, consists of diverse cell types (i.e., endothelial, epithelial, mesenchymal, inflammatory). In lung cells, autophagy may represent a general inducible adaptive response to injury resulting from exposure to stress agents, including hypoxia, oxidants, inflammation, ischemia–reperfusion, endoplasmic reticulum stress, pharmaceuticals, or inhaled xenobiotics (i.e., air pollution, cigarette smoke). In recent studies, we have observed increased autophagy in mouse lungs subjected to chronic cigarette smoke exposure, and in pulmonary epithelial cells exposed to cigarette smoke extract. Knockdown of autophagic proteins inhibited apoptosis in response to cigarette smoke exposure in vitro, suggesting that increased autophagy was associated with epithelial cell death. We have also observed increased morphological and biochemical markers of autophagy in human lung specimens from patients with chronic obstructive pulmonary disease (COPD). We hypothesize that increased autophagy contributes to COPD pathogenesis by promoting epithelial cell death. Further research will examine whether autophagy plays a homeostatic or maladaptive role in COPD and other human lung diseases.
doi:10.1513/pats.200909-101JS
PMCID: PMC3137145  PMID: 20160144
autophagy; apoptosis; pulmonary disease
16.  Autophagy in Vascular Disease 
Autophagy, or “self eating,” refers to a regulated cellular process for the lysosomal-dependent turnover of organelles and proteins. During starvation or nutrient deficiency, autophagy promotes survival through the replenishment of metabolic precursors derived from the degradation of endogenous cellular components. Autophagy represents a general homeostatic and inducible adaptive response to environmental stress, including endoplasmic reticulum stress, hypoxia, oxidative stress, and exposure to pharmaceuticals and xenobiotics. Whereas elevated autophagy can be observed in dying cells, the functional relationships between autophagy and programmed cell death pathways remain incompletely understood. Preclinical studies have identified autophagy as a process that can be activated during vascular disorders, including ischemia–reperfusion injury of the heart and other organs, cardiomyopathy, myocardial injury, and atherosclerosis. The functional significance of autophagy in human cardiovascular disease pathogenesis remains incompletely understood, and potentially involves both adaptive and maladaptive outcomes, depending on model system. Although relatively few studies have been performed in the lung, our recent studies also implicate a role for autophagy in chronic lung disease. Manipulation of the signaling pathways that regulate autophagy could potentially provide a novel therapeutic strategy in the prevention or treatment of human disease.
doi:10.1513/pats.200909-100JS
PMCID: PMC3137148  PMID: 20160147
autophagy; apoptosis; vascular disease
17.  Carbon Monoxide and Bilirubin 
Heme oxygenase (HO)-1, an inducible, low–molecular-weight stress protein, confers cellular and tissue protection in multiple models of injury and disease, including oxidative or inflammatory lung injury, ischemia/reperfusion (I/R) injuries, and vascular injury/disease. The tissue protection provided by HO-1 potentially relates to the endogenous production of the end products of its enzymatic activity: namely, biliverdin (BV)/bilirubin (BR), carbon monoxide (CO), and iron. Of these, CO and BV/BR show promise as possible therapeutic agents when applied exogenously in models of lung or vascular injury. CO activates intracellular signaling pathways that involve soluble guanylate cyclase and/or p38 mitogen-activated protein kinase. Although toxic at elevated concentrations, low concentrations of CO can confer antiinflammatory, antiapoptotic, antiproliferative, and vasodilatory effects. BV and BR are natural antioxidants that can provide protection against oxidative stress in cell culture and in plasma. Application of BV or BR protects against I/R injury in several organ models. Recent evidence has also demonstrated antiinflammatory and antiproliferative properties of these pigments. To date, evidence has accumulated for salutary effects of CO, BV, and/or BR in lung/vascular injury models, as well as in models of transplant-associated I/R injury. Thus, the exogenous application of HO end products may provide an alternative to pharmacologic or gene therapy approaches to harness the therapeutic potential of HO-1.
doi:10.1165/rcmb.2006-0333TR
PMCID: PMC2176112  PMID: 16980550
bilirubin; carbon monoxide; heme oxygenase-1; inflammation; ischemia/reperfusion
18.  Hyperoxia-Induced LC3B Interacts with the Fas Apoptotic Pathway in Epithelial Cell Death 
Epithelial cell death plays a critical role in hyperoxia-induced lung injury. We investigated the involvement of the autophagic marker microtubule-associated protein-1 light chain-3B (LC3B) in epithelial cell apoptosis after hyperoxia. Prolonged hyperoxia (>95% O2), which causes characteristic lung injury in mice, activated morphological and biochemical markers of autophagy. Hyperoxia induced the time-dependent expression and conversion of LC3B-I to LC3B-II in mouse lung in vivo and in cultured epithelial cells (Beas-2B, human bronchial epithelial cells) in vitro. Hyperoxia increased autophagosome formation in Beas-2B cells, as evidenced by electron microscopy and increased GFP-LC3 puncta. The augmented LC3B level after hyperoxia was transcriptionally regulated and dependent in part on the c-Jun N-terminal kinase pathway. We hypothesized that LC3B plays a regulatory role in hyperoxia-induced epithelial apoptosis. LC3B siRNA promoted hyperoxia-induced cell death in epithelial cells, whereas overexpression of LC3B conferred cytoprotection after hyperoxia. The autophagic protein LC3B cross-regulated the Fas apoptotic pathway by physically interacting with the components of death-inducing signaling complex. This interaction was mediated by caveolin-1 tyrosine 14, which is a known target of phosphorylation induced by hyperoxia. Taken together, hyperoxia-induced LC3B activation regulates the Fas apoptotic pathway and thus confers cytoprotection in lung epithelial cells. The interaction of LC3B and Fas pathways requires cav-1.
doi:10.1165/rcmb.2009-0415OC
PMCID: PMC3359946  PMID: 22095627
apoptosis; autophagy; hyperoxia; lung injury; caveolin-1
19.  Carbon Monoxide Activates Autophagy via Mitochondrial Reactive Oxygen Species Formation 
Autophagy, an autodigestive process that degrades cellular organelles and protein, plays an important role in maintaining cellular homeostasis during environmental stress. Carbon monoxide (CO), a toxic gas and candidate therapeutic molecule, confers cytoprotection in animal models of acute lung injury. The mechanisms underlying CO-dependent lung cell protection and the role of autophagy in this process remain unclear. Here, we demonstrate that CO exposure time-dependently increased the expression and activation of the autophagic protein, microtubule-associated protein–1 light chain-3B (LC3B) in mouse lung, and in cultured human alveolar (A549) or human bronchial epithelial cells. Furthermore, CO increased autophagosome formation in epithelial cells by electron microscopy and green fluorescent protein (GFP)-LC3 puncta assays. Recent studies indicate that reactive oxygen species (ROS) play an important role in the activation of autophagy. CO up-regulated mitochondria-dependent generation of ROS in epithelial cells, as assayed by MitoSOX fluorescence. Furthermore, CO-dependent induction of LC3B expression was inhibited by N-acetyl-L-cysteine and the mitochondria-targeting antioxidant, Mito-TEMPO. These data suggest that CO promotes the autophagic process through mitochondrial ROS generation. We investigated the relationships between autophagic proteins and CO-dependent cytoprotection using a model of hyperoxic stress. CO protected against hyperoxia-induced cell death, and inhibited hyperoxia-associated ROS production. The ability of CO to protect against hyperoxia-induced cell death and caspase-3 activation was compromised in epithelial cells infected with LC3B-small interfering (si)RNA, indicating a role for autophagic proteins. These studies uncover a new mechanism for the protective action of CO, in support of potential therapeutic application of this gas.
doi:10.1165/rcmb.2010-0352OC
PMCID: PMC3208612  PMID: 21441382
apoptosis; autophagy; carbon monoxide; epithelial cells; hyperoxia
20.  Autophagic Protein LC3B Confers Resistance against Hypoxia-induced Pulmonary Hypertension 
Rationale: Pulmonary hypertension (PH) is a progressive disease with unclear etiology. The significance of autophagy in PH remains unknown.
Objectives: To determine the mechanisms by which autophagic proteins regulate tissue responses during PH.
Methods: Lungs from patients with PH, lungs from mice exposed to chronic hypoxia, and human pulmonary vascular cells were examined for autophagy using electron microscopy and Western analysis. Mice deficient in microtubule-associated protein-1 light chain-3B (LC3B−/−), or early growth response-1 (Egr-1−/−), were evaluated for vascular morphology and hemodynamics.
Measurements and Main Results: Human PH lungs displayed elevated lipid-conjugated LC3B, and autophagosomes relative to normal lungs. These autophagic markers increased in hypoxic mice, and in human pulmonary vascular cells exposed to hypoxia. Egr-1, which regulates LC3B expression, was elevated in PH, and increased by hypoxia in vivo and in vitro. LC3B−/− or Egr-1−/−, but not Beclin 1+/−, mice displayed exaggerated PH during hypoxia. In vitro, LC3B knockdown increased reactive oxygen species production, hypoxia-inducible factor-1α stabilization, and hypoxic cell proliferation. LC3B and Egr-1 localized to caveolae, associated with caveolin-1, and trafficked to the cytosol during hypoxia.
Conclusions: The results demonstrate elevated LC3B in the lungs of humans with PH, and of mice with hypoxic PH. The increased susceptibility of LC3B−/− and Egr-1−/− mice to hypoxia-induced PH and increased hypoxic proliferation of LC3B knockdown cells suggest adaptive functions of these proteins during hypoxic vascular remodeling. The results suggest that autophagic protein LC3B exerts a protective function during the pathogenesis of PH, through the regulation of hypoxic cell proliferation.
doi:10.1164/rccm.201005-0746OC
PMCID: PMC3081281  PMID: 20889906
autophagy; hypoxia; hypertension, pulmonary
21.  Autophagy proteins regulate innate immune response by inhibiting NALP3 inflammasome-mediated mitochondrial DNA release 
Nature immunology  2010;12(3):222-230.
Autophagy, a cellular process for organelle and protein turnover, regulates innate immune responses. We demonstrate that depletion of autophagic proteins microtubule associated protein-1 light chain 3B (LC3B) and Beclin 1 enhances caspase-1 activation and secretion of interleukin-1β and interleukin-18. Autophagic protein depletion promoted accumulation of dysfunctional mitochondria and cytosolic translocation of mitochondrial DNA (mtDNA) in response to lipopolysaccharide (LPS) and ATP in macrophages. Release of mtDNA into the cytosol depended on the NALP3 inflammasome and mitochondrial ROS. Cytosolic mtDNA contributed to IL-1β and IL-18 secretion in response to LPS and ATP. LC3B-deficient mice produced more caspase-1-dependent cytokines in two sepsis models and were susceptible to LPS-induced mortality. Our study suggests that autophagic proteins regulate NALP3-dependent inflammation by preserving mitochondrial integrity.
doi:10.1038/ni.1980
PMCID: PMC3079381  PMID: 21151103
22.  Autophagy in cigarette smoke-induced chronic obstructive pulmonary disease 
The molecular and cellular mechanisms underlying the pathogenesis of chronic obstructive pulmonary disease (COPD) remain incompletely understood. We have investigated the potential role of macro-autophagy, a cellular homeostatic mechanism, in COPD and cigarette smoke-induced lung-cell injury. Autophagy is a dynamic process for the turnover of organelles and proteins, which regenerates metabolic precursors through the lysosomal-dependent catabolism of cellular macromolecules. It is typically associated with survival pathways, especially in nutrient deficiency states. The role of autophagy in human diseases is less clear, and has been associated with both protective and detrimental consequences, depending on the disease model. While autophagy is considered cytoprotective, this process is often found in association with cell death, and the relationships between autophagy and cell death remain ambiguous. We have found elevated autophagy in COPD lung specimens, as well as in response to cigarette smoke exposure in vitro and in vivo. In our studies, the activation of autophagic proteins was associated with epithelial cell apoptosis in response to cigarette smoke, with pathogenic implications in COPD. Further studies are needed to determine the functional significance of autophagy in COPD and other diseases of the lung.
doi:10.1586/ers.10.61
PMCID: PMC3081520  PMID: 20923337
apoptosis; autophagy; chronic obstructive pulmonary disease; cigarette smoke; emphysema
23.  Carbon monoxide inhibits Fas activating antibody-induced apoptosis in endothelial cells 
Background
The extrinsic apoptotic pathway initiates when a death ligand, such as the Fas ligand, interacts with its cell surface receptor (ie., Fas/CD95), forming a death-inducing signaling complex (DISC). The Fas-dependent apoptotic pathway has been implicated in several models of lung or vascular injury. Carbon monoxide, an enzymatic product of heme oxygenase-1, exerts antiapoptotic effects at low concentration in vitro and in vivo.
Methods
Using mouse lung endothelial cells (MLEC), we examined the antiapoptotic potential of carbon monoxide against apoptosis induced by the Fas/CD95-activating antibody (Jo2). Carbon monoxide was applied to cell cultures in vitro. The expression and/or activation of apoptosis-related proteins and signaling intermediates were determined using Western Immunoblot and co-immunoprecipitation assays. Cell death was monitored by lactate dehydrogenase (LDH) release assays. Statistical significance was determined by student T-test and a value of P < 0.05 was considered significant.
Results
Treatment of MLEC with Fas-activating antibody (Jo2) induced cell death associated with the formation of the DISC, and activation of caspases (-8, -9, and -3), as well as the pro-apoptotic Bcl-2 family protein Bax. Exposure of MLEC to carbon monoxide inhibited Jo2-induced cell death, which correlated with the inhibition of DISC formation, cleavage of caspases-8, -9, and -3, and Bax activation. Carbon monoxide inhibited the phosphorylation of the Fas-associated death domain-containing protein, as well as its association with the DISC. Furthermore, carbon monoxide induced the expression of the antiapoptotic protein FLIP and increased its association with the DISC.
CO-dependent cytoprotection against Fas mediated apoptosis in MLEC depended in part on activation of ERK1/2-dependent signaling.
Conclusions
Carbon monoxide has been proposed as a potential therapy for lung and other diseases based in part on its antiapoptotic effects in endothelial cells. In vitro, carbon monoxide may inhibit both Fas/caspase-8 and Bax-dependent apoptotic signaling pathways induced by Fas-activating antibody in endothelial cells. Strategies to block Fas-dependent apoptotic pathways may be useful in development of therapies for lung or vascular disorders.
doi:10.1186/2045-9912-1-8
PMCID: PMC3231877  PMID: 22146483
24.  Carbon monoxide prevents ventilator induced lung injury via caveolin-1 
Critical care medicine  2009;37(5):1708-1715.
Objectives
Carbon monoxide (CO) can confer anti-inflammatory protection in rodent models of ventilator-induced lung injury (VILI). Caveolin-1 exerts a critical role in cellular responses to mechanical stress, and has been shown to mediate cytoprotective effects of CO in vitro. We sought to determine the role of caveolin-1 in lung susceptibility to VILI in mice. Furthermore, we assessed the role of caveolin-1 in the tissue protective effects of CO in the VILI model.
Design
Prospective experimental study
Setting
University laboratory
Subjects
Wild type (wt) and caveolin-1 deficient (cav-−/−) mice
Interventions
Mice were subjected to tracheostomy and arterial cannulation. Wt and cav-1−/− mice were ventilated with a tidal volume of 12 ml/kg body weight and a frequency of 80/min for 5 min as control, or for 8h with air in the absence or presence of CO (250 parts per million). Bronchoalveolar lavage (BAL) and histology were used to determine lung injury. Lung sections or homogenates were analyzed for caveolin-1 expression by immunohistochemical staining or Western Blotting, respectively.
Measurements and Main Results
Ventilation led to an increase in BAL protein concentration, cell count, neutrophil recruitment, and edema formation that was prevented in the presence of CO. While ventilation alone slightly induced caveolin-1 expression in epithelial cells, the application of CO during the ventilation significantly increased the expression of caveolin-1. In comparison to wt mice, mechanical ventilation of cav-1−/− mice led to a significantly higher degree of lung injury as compared to wt mice. In contrast to its effectiveness in wt mice, CO-administration failed to reduce lung injury markers in cav-1−/− mice.
Conclusions
Caveolin-1 null mice are more susceptible to VILI. Carbon monoxide executes lung protective effects during mechanical ventilation that are dependent in part, on caveolin-1 expression.
doi:10.1097/CCM.0b013e31819efa31
PMCID: PMC3086639  PMID: 19325477
ventilator induced lung injury; mechanical ventilation; carbon monoxide; caveolin-1; mechanotransduction; acute lung injury
25.  Heme Oxygenase-1/Carbon Monoxide 
Heme oxygenase-1 (HO-1), a ubiquitous inducible stress-response protein, serves a major metabolic function in heme turnover. HO activity cleaves heme to form biliverdin-IXα, carbon monoxide (CO), and iron. Genetic experiments have revealed a central role for HO-1 in tissue homeostasis, protection against oxidative stress, and in the pathogenesis of disease. Four decades of research have witnessed not only progress in elucidating the molecular mechanisms underlying the regulation and function of this illustrious enzyme, but also have opened remarkable translational applications for HO-1 and its reaction products. CO, once regarded as a metabolic waste, can act as an endogenous mediator of cellular signaling and vascular function. Exogenous application of CO by inhalation or pharmacologic delivery can confer cytoprotection in preclinical models of lung/vascular injury and disease, based on anti-apoptotic, anti-inflammatory, and anti-proliferative properties. The bile pigments, biliverdin and bilirubin, end products of heme degradation, have also shown potential as therapeutics in vascular disease based on anti-inflammatory and anti-proliferative activities. Further translational and clinical trials research will unveil whether the HO-1 system or any of its reaction products can be successfully applied as molecular medicine in human disease.
doi:10.1165/rcmb.2009-0170TR
PMCID: PMC2742746  PMID: 19617398
carbon monoxide; bilirubin; heme oxygenase-1; lung injury

Results 1-25 (37)