The Role Of Peroxiredoxin 4 And Er Redox Stress In Idiopathic Pulmonary Fibrosis

Idiopathic Pulmonary Fibrosis (IPF) is a disease that is characterized by excessive scar formation in the lung. IPF is a disease associated with aging and is believed to be the manifestation of repeated micro-injuries and lack of adequate repair over time. Notably increases in endoplasmic reticulum (ER) stress of epithelial cells and excessive epithelial cell death have been identified as processes that drive the progression of fibrosis. Every year 34,000 people in the United States are diagnosed with Idiopathic Pulmonary Fibrosis (IPF). The average survival time for these patients is only 3-5 years after diagnosis. Two available drugs stabilize patients for some time but not influence overall survival, pointing to a strong need for development of new treatment modalities. ER stress has come to the forefront of IPF research. ER stress occurs in familial and sporadic IPF, and pharmacological inducers of ER stress augment bleomycin-induced fibrosis in mice. Recent studies have shown that triggering of the ER stress response or epithelial cell death are sufficient to induce pulmonary fibrosis in mouse models. The ER is an oxidizing environment that allows disulfide bridge formation in proteins that are localized to the cell surface or secreted, in order to provide structural stability. Therefore, the ER has a unique complement of oxidoreductases required to maintain redox homeostasis. Peroxiredoxin 4 (PRDX4) is amongst the highest expressed ER oxidoreductases in the lung. Little information is known about the role of PRDX4 in the pathogenesis of pulmonary fibrosis or in epithelial cell responses to pro-fibrotic insults. In this thesis, I demonstrated that with age there is a significant decrease in the mRNA expression of a large subset of oxidoreductase genes in the lung, one of which is PRDX4. Interestingly there was also a significant increase in genes associated with the extracellular matrix and the expression of the profibrotic growth factor, transforming growth factor beta 1 (TGFB1), which has been shown to play an important role in IPF. I identified an increase in the oligomerization of PRDX4 in IPF patient samples, which points to an alteration in ER redox homeostasis in the lungs of IPF patients compared to normal controls. The stabilization of high molecular weight (HMW) species of PRDX4 in response to an increased oxidative burden was confirmed and the mechanism by which PRDX4 oligomerizes was unraveled via mutagenesis studies. Treatment of mouse lung epithelial cells with TGFB1 caused the formation of HMW species of PRDX4, induced the unfolded protein response (UPR), and led to apoptosis. I also demonstrated that TGFB1 induced PRDX4 HMW species formation via the increased expression of ER oxidoreductin 1 alpha (ERO1A). Furthermore, the overexpression of PRDX4 alleviated TGFB1 induced ER stress, and the inhibition of ERO1A decreased lung epithelial cell death in response to TGFB1. Overall, these findings point to the importance of the ER oxidative environment in modulating epithelial cell fate in settings of pulmonary fibrosis

Glutathione S-transferases and their implications in the lung diseases asthma and chronic obstructive pulmonary disease: Early life susceptibility?

Our lungs are exposed daily to airborne pollutants, particulate matter, pathogens as well as lung allergens and irritants. Exposure to these substances can lead to inflammatory responses and may induce endogenous oxidant production, which can cause chronic inflammation, tissue damage and remodeling. Notably, the development of asthma and Chronic Obstructive Pulmonary Disease (COPD) is linked to the aforementioned irritants. Some inhaled foreign chemical compounds are rapidly absorbed and processed by phase I and II enzyme systems critical in the detoxification of xenobiotics including the glutathione-conjugating enzymes Glutathione S-transferases (GSTs). GSTs, and in particular genetic variants of GSTs that alter their activities, have been found to be implicated in the susceptibility to and progression of these lung diseases. Beyond their roles in phase II metabolism, evidence suggests that GSTs are also important mediators of normal lung growth. Therefore, the contribution of GSTs to the development of lung diseases in adults may already start in utero, and continues through infancy, childhood, and adult life. GSTs are also known to scavenge oxidants and affect signaling pathways by protein-protein interaction. Moreover, GSTs regulate reversible oxidative post-translational modifications of proteins, known as protein S-glutathionylation. Therefore, GSTs display an array of functions that impact the pathogenesis of asthma and COPD. In this review we will provide an overview of the specific functions of each class of mammalian cytosolic GSTs. This is followed by a comprehensive analysis of their expression profiles in the lung in healthy subjects, as well as alterations that have been described in (epithelial cells of) asthmatics and COPD patients. Particular emphasis is placed on the emerging evidence of the regulatory properties of GSTs beyond detoxification and their contribution to (un)healthy lungs throughout life. By providing a more thorough understanding, tailored therapeutic strategies can be designed to affect specific functions of particular GSTs

ER Oxidative Stress Promotes Glutathione-Dependent Oxidation of Collagen-1A1 and Promotes Lung Fibroblast Activation

Changes in the oxidative (redox) environment accompany idiopathic pulmonary fibrosis (IPF). S-glutathionylation of reactive protein cysteines is a post-translational event that transduces oxidant signals into biological responses. We recently demonstrated that increases in S-glutathionylation promote pulmonary fibrosis, which was mitigated by the deglutathionylating enzyme glutaredoxin (GLRX). However, the protein targets of S-glutathionylation that promote fibrogenesis remain unknown. In the present study we addressed whether the extracellular matrix is a target for S-glutathionylation. We discovered increases in collagen 1A1 S-glutathionylation (COL1A1-SSG) in lung tissues from IPF subjects compared to control subjects in association with increases in ER oxidoreductin 1 (ERO1A) and enhanced oxidation of ER-localized peroxiredoxin 4 (PRDX4) reflecting an increased oxidative environment of the endoplasmic reticulum (ER). Human lung fibroblasts exposed to transforming growth factor beta 1 (TGFB1) show increased secretion of COL1A1-SSG. Pharmacologic inhibition of ERO1A diminished oxidation of PRDX4, attenuated COL1A1-SSG and total COL1A1 levels and dampened fibroblast activation. Absence of Glrx enhanced COL1A1-SSG and overall COL1A1 secretion and promoted activation of mechanosensing pathways. Remarkably, COL1A1-SSG resulted in marked resistance to collagenase degradation. Compared to COL1, lung fibroblasts plated on COL1-SSG proliferated more rapidly, and increased expression of genes encoding extracellular matrix crosslinking enzymes and genes linked to mechanosensing pathways. Overall, these findings suggest that glutathione-dependent oxidation of COL1A1 occurs in settings of IPF in association with enhanced ER oxidative stress and may promote fibrotic remodeling due to increased resistance to collagenase-mediated degradation and fibroblast activation

COVID-19 vaccination elicits an evolving, cross-reactive antibody response to epitopes conserved with endemic coronavirus spike proteins.

The COVID-19 pandemic has triggered the first widespread vaccination campaign against a coronavirus. Many vaccinated subjects are previously naive to SARS-CoV-2; however, almost all have previously encountered other coronaviruses (CoVs), and the role of this immunity in shaping the vaccine response remains uncharacterized. Here, we use longitudinal samples and highly multiplexed serology to identify mRNA-1273 vaccine-induced antibody responses against a range of CoV Spike epitopes, in both phylogenetically conserved and non-conserved regions. Whereas reactivity to SARS-CoV-2 epitopes shows a delayed but progressive increase following vaccination, we observe distinct kinetics for the endemic CoV homologs at conserved sites in Spike S2: these become detectable sooner and decay at later time points. Using homolog-specific antibody depletion and alanine-substitution experiments, we show that these distinct trajectories reflect an evolving cross-reactive response that can distinguish rare, polymorphic residues within these epitopes. Our results reveal mechanisms for the formation of antibodies with broad reactivity against CoVs