Tejaswini Kulkarni

Heme oxygenase-1-mediated autophagy protects against pulmonary endothelial cell death and development of emphysema in cadmium-treated mice

Pulmonary exposure to cadmium, a major component of cigarette smoke, has a dramatic impact on lung function and the development of emphysema. Cigarette smoke exposure induces heme oxygenase-1 (HO-1), a cytoprotective enzyme. In this study, we employed a truncated mouse model of emphysema by intratracheal instillation of cadmium (CdCl2) solution (0.025% per 1 mg/kg body wt) in HO-1(+/+), HO-1(-/-), and overexpressing humanized HO-1 bacterial artificial chromosome (hHO-1BAC) mice. We evaluated the role of HO-1 in cadmium-induced emphysema in mice by analyzing histopathology, micro-computed tomography scans, and lung function tests. CdCl2-exposed HO-1(-/-) mice exhibited more severe emphysema compared with HO-1(+/+) or hHO-1BAC mice. Loss of pulmonary endothelial cells (PECs) from the alveolar capillary membrane is recognized to be a target in emphysema. PECs from HO-1(+/+), HO-1(-/-), and hHO-1BAC were employed to define the underlying molecular mechanism for the protection from emphysema by HO-1. Electron microscopy, expression of autophagic markers (microtubule-associated protein 1B-light chain 3 II, autophagy protein 5, and Beclin1) and apoptotic marker (cleaved caspase 3) suggested induction of autophagy and apoptosis in PECs after CdCl2 treatment. CdCl2-treated HO-1(-/-) PECs exhibited downregulation of autophagic markers and significantly increased cleaved caspase 3 expression and activity (∼4-fold higher). Moreover, hHO-1BAC PECs demonstrated upregulated autophagy and absence of cleaved caspase 3 expression or activity. Pretreatment of HO-1(+/+) PECs with rapamycin induced autophagy and resulted in reduced cell death upon cadmium treatment. Induction of autophagy following CdCl2 treatment was found to be protective from apoptotic cell death. HO-1 induced protective autophagy in PECs and mitigated cadmium-induced emphysema.

Matrix Remodeling in Pulmonary Fibrosis and Emphysema

Pulmonary fibrosis and emphysema are chronic lung diseases characterized by a progressive decline in lung function, resulting in significant morbidity and mortality. A hallmark of these diseases is recurrent or persistent alveolar epithelial injury, typically caused by common environmental exposures such as cigarette smoke. We propose that critical determinants of the outcome of the injury-repair processes that result in fibrosis versus emphysema are mesenchymal cell fate and associated extracellular matrix dynamics. In this review, we explore the concept that regulation of mesenchymal cells under the influence of soluble factors, in particular transforming growth factor-β1, and the extracellular matrix determine the divergent tissue remodeling responses seen in pulmonary fibrosis and emphysema.

Characteristics of patients with pulmonary venoocclusive disease awaiting transplantation.

RATIONALE Pulmonary venoocclusive disease (PVOD) is an uncommon cause of pulmonary arterial hypertension (PAH). However, unlike PAH, treatment options for PVOD are usually quite limited. The impact of the lung allocation score on access to transplantation for patients with PVOD and the clinical course of these patients have not been well-described. OBJECTIVES To examine the association between the diagnosis of PVOD and lung transplantation for patients on the transplant waiting list. METHODS Patients with a diagnosis of PVOD and PAH registered on the United Network for Organ Sharing wait list for transplantation from May 4, 2005 to May 3, 2013 were included. Lung transplantation was the primary outcome measure. Multivariable analyses were performed to determine the odds of dying or receiving a lung transplant after listing. Survival was compared using Kaplan-Meier and competing risks methods. RESULTS Of 12,251 patients listed for lung transplantation, 49 with PVOD and 647 with PAH were identified. There were no significant differences in the lung allocation score between patients with PVOD and PAH at listing, transplant, or wait list removal for death/too sick for transplant. By 6 months, 22.6% of patients with PVOD had been removed from the wait list due to death, compared with 11.0% of patients with PAH (Chi-square P = 0.03). Patients with PVOD who died or were considered too sick for transplant were removed from the waiting list sooner after listing (22 vs. 105 d, P = 0.08). There was no difference in the proportion of patients with PVOD and PAH transplanted (50.0 vs. 47.6%, P = 0.60). CONCLUSIONS In the lung allocation score era, patients with PVOD may be at higher risk for death while on the transplant waiting list. After wait list registration, close monitoring for disease progression is advised.

Heme Oxygenase-1 Protects Corexit 9500A-Induced Respiratory Epithelial Injury across Species

The effects of Corexit 9500A (CE) on respiratory epithelial surfaces of terrestrial mammals and marine animals are largely unknown. This study investigated the role of CE-induced heme oxygenase-1 (HO-1), a cytoprotective enzyme with anti-apoptotic and antioxidant activity, in human bronchial airway epithelium and the gills of exposed aquatic animals. We evaluated CE-mediated alterations in human airway epithelial cells, mice lungs and gills from zebrafish and blue crabs. Our results demonstrated that CE induced an increase in gill epithelial edema and human epithelial monolayer permeability, suggesting an acute injury caused by CE exposure. CE induced the expression of HO-1 as well as C-reactive protein (CRP) and NADPH oxidase 4 (NOX4), which are associated with ROS production. Importantly, CE induced caspase-3 activation and subsequent apoptosis of epithelial cells. The expression of the intercellular junctional proteins, such as tight junction proteins occludin, zonula occludens (ZO-1), ZO-2 and adherens junctional proteins E-cadherin and Focal Adhesion Kinase (FAK), were remarkably inhibited by CE, suggesting that these proteins are involved in CE-induced increased permeability and subsequent apoptosis. The cytoskeletal protein F-actin was also disrupted by CE. Treatment with carbon monoxide releasing molecule-2 (CORM-2) significantly inhibited CE-induced ROS production, while the addition of HO-1 inhibitor, significantly increased CE-induced ROS production and apoptosis, suggesting a protective role of HO-1 or its reaction product, CO, in CE-induced apoptosis. Using HO-1 knockout mice, we further demonstrated that HO-1 protected against CE-induced inflammation and cellular apoptosis and corrected CE-mediated inhibition of E-cadherin and FAK. These observations suggest that CE activates CRP and NOX4-mediated ROS production, alters permeability by inhibition of junctional proteins, and leads to caspase-3 dependent apoptosis of epithelial cells, while HO-1 and its reaction products protect against oxidative stress and apoptosis.

Alveolar epithelial disintegrity in pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease characterized by progressive decline in lung function, resulting in significant morbidity and mortality. Current concepts of the pathogenesis of IPF primarily center on dysregulated epithelial cell repair and altered epithelial-mesenchymal communication and extracellular matrix deposition following chronic exposure to cigarette smoke or environmental toxins. In recent years, increasing attention has been directed toward the role of the intercellular junctional complex in determining the specific properties of epithelia in pulmonary diseases. Additionally, recent genomewide association studies suggest that specific genetic variants predictive of epithelial cell dysfunction may confer susceptibility to the development of sporadic idiopathic pulmonary fibrosis. A number of genetic disorders linked to pulmonary fibrosis and familial interstitial pneumonias are associated with loss of epithelial integrity. However, the potential links between extrapulmonary clinical syndromes associated with defects in epithelial cells and the development of pulmonary fibrosis are not well understood. Here, we report a case of hereditary mucoepithelial dysplasia that presented with pulmonary fibrosis and emphysema on high-resolution computed tomography. This case illustrates a more generalizable concept of epithelial disintegrity in the development of fibrotic lung diseases, which is explored in greater detail in this review article.

Autoimmunity to Vimentin Is Associated with Outcomes of Patients with Idiopathic Pulmonary Fibrosis

Autoimmunity has been implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF); however, the repertoire of autoantigens involved in this disease and the clinical relevance of these autoimmune responses are still being explored. Our initial discovery assays demonstrated that circulating and intrapulmonary vimentin levels are increased in IPF patients. Subsequent studies showed native vimentin induced HLA-DR–dependent in vitro proliferation of CD4 T cells from IPF patients and enhanced the production of IL-4, IL-17, and TGF-β1 by these lymphocytes in contrast to normal control specimens. Vimentin supplementation of IPF PBMC cultures also resulted in HLA-DR–dependent production of IgG with anti-vimentin specificities. Circulating anti-vimentin IgG autoantibody levels were much greater in IPF subjects from the University of Alabama at Birmingham (n = 102) and the University of Pittsburgh (U. Pitt., n = 70) than in normal controls. Anti-vimentin autoantibody levels in IPF patients were HLA biased and inversely correlated with physiological measurements of lung function (i.e., forced expiratory volumes and diffusing capacities). Despite considerable intergroup differences in transplant-free survival between these two independent IPF cohorts, serious adverse outcomes were most frequent among the patients within each population that had the highest anti-vimentin autoantibody levels (University of Alabama at Birmingham: hazard ratio 2.5, 95% confidence interval 1.2–5.3, p = 0.012; University of Pittsburgh: hazard ratio 2.7, 95% confidence interval 1.3–5.5, p = 0.006). These data show that anti-vimentin autoreactivity is prevalent in IPF patients and is strongly associated with disease manifestations. These findings have implications with regard to the pathogenesis of this enigmatic disease and raise the possibility that therapies specifically directed at these autoimmune processes could have therapeutic efficacy.

Flexible Bronchoscopy Is Safe and Effective in Adult Subjects Supported With Extracorporeal Membrane Oxygenation

BACKGROUND: Previous studies have demonstrated the safety of flexible bronchoscopy (FB) in mechanically ventilated subjects. However, the safety of FB in adult subjects receiving extracorporeal membrane oxygenation (ECMO) has not been described previously. METHODS: A retrospective review was conducted of all adult subjects who underwent FB while receiving ECMO support at the University of Alabama at Birmingham Hospital from January 1, 2013, to December 31, 2014. Physiologic variables, pre- and post-FB ECMO, and ventilator settings were recorded. RESULTS: 79 adult subjects underwent FB receiving ECMO with a total of 223 bronchoscopies. The most common indications for bronchoscopy included diagnostic evaluation of infection in subjects with pneumonia (29%) and clearance of excessive secretions (22%). In 70% of subjects, moderate or greater amounts of secretions were noted. FB yielded positive culture data in 37 subjects (47%), which resulted in a change to the antibiotic regimen in 14 subjects (38%) with positive culture data. No significant differences in mean PaO2/FIO2, mean ECMO flow, mean sweep gas, ventilator settings, or hemodynamic parameters (heart rate, oxygen saturation, and mean blood pressure) were noted before and after FB. Complications were mild and transient: blood-tinged secretions after FB in 21% cases, which resolved spontaneously, intraprocedural hypoxemia in 2.2% of cases, and dysrhythmia in <1% of cases. There were no episodes of ECMO cannula dislodgement or inadvertent extubation. CONCLUSIONS: FB can be used safely in adult subjects supported with ECMO and is not associated with significant hemodynamics changes, bleeding, or mechanical complications during ECMO support.

Attenuated heme oxygenase-1 responses predispose the elderly to pulmonary nontuberculous mycobacterial infections

Pulmonary infections with nontuberculous mycobacteria (P-NTM), such as by Mycobacterium avium complex (M. avium), are increasingly found in the elderly, but the underlying mechanisms are unclear. Recent studies suggest that adaptive immunity is necessary, but not sufficient, for host defense against mycobacteria. Heme oxygenase-1 (HO-1) has been recognized as a critical modulator of granuloma formation and programmed cell death in mycobacterial infections. Old mice (18-21 mo) infected with M. avium had attenuated HO-1 response with diffuse inflammation, high burden of mycobacteria, poor granuloma formation, and decreased survival (45%), while young mice (4-6 mo) showed tight, well-defined granuloma, increased HO-1 expression, and increased survival (95%). To further test the role of HO-1 in increased susceptibility to P-NTM infections in the elderly, we used old and young HO-1+/+ and HO-1-/- mice. The transcriptional modulation of the JAK/STAT signaling pathway in HO-1-/- mice due to M. avium infection demonstrated similarities to infected wild-type old mice with upregulation of SOCS3 and inhibition of Bcl2. Higher expression of SOCS3 with downregulation of Bcl2 resulted in higher macrophage death via cellular necrosis. Finally, peripheral blood monocytes (PBMCs) from elderly patients with P-NTM also demonstrated attenuated HO-1 responses after M. avium stimulation and increased cell death due to cellular necrosis (9.69% ± 2.02) compared with apoptosis (4.75% ± 0.98). The augmented risk for P-NTM in the elderly is due, in part, to attenuated HO-1 responses, subsequent upregulation of SOCS3, and inhibition of Bcl2, leading to programmed cell death of macrophages, and sustained infection.