title: Molecular Basis of Apoptotic-Resistance in Idiopathic Pulmonary Fibrosis Fibroblasts
Creators: name: (:Unkn) Unknown
Publisher: name: George Mason University
Year: 2019

description: Idiopathic Pulmonary Fibrosis (IPF) is a chronic and fatal form of interstitial lung disease. IPF is characterized by excessive remodeling of the lung parenchyma, which impairs gaseous exchange and proper oxygenation as a result of scar tissue in the interstitium. The fibrotic process is distinct from normal physiological wound healing due to the abundance of active fibroblasts that persist in densely fibrotic regions called the fibrotic foci and evade apoptosis through mechanisms not yet well-understood. In typical cell aging, the cell is primed with death signals to promote cell-turnover. However, fibroblasts at the foci displaying hallmarks of senescence (telomere attrition, p16+, p21+) are producing more collagen and migrating more despite the loss of proteostasis and organelle perturbation. This suggests activated fibroblasts likely require tonic expression of prosurvival proteins involved in cellular stress management in order to ensure survival. We postulate that the dysregulation of organelles, such as the mitochondria and endoplasmic reticulum in aging cells, is a source of apoptotic-resistance observed in senescent IPF fibroblasts. Here, we characterize the mitochondrial dynamics of IPF fibroblasts (fission, fusion, spatial redistribution, and network morphology) in the presence of Tunicamycin-induced ER stress. We found that mitochondria in fibroblast populations of the IPF lung favored survival by maintaining mitochondrial membrane potential and retaining cytochrome c localization in the mitochondria. Mitochondrial genome expression was also correlated with patient prognosis, as measured by %FEV, FVC, and 6-minute walk test. The difference in mitochondrial response to ER stress can also be partially attributed to a maladaptive unfolded protein response (UPR) pathway in IPF fibroblasts. The ER stress sensor IREα, once activated, acts as an endoribonuclease and splices XBP1 into a functional transcription factor capable of regulating proapoptotic gene expression. We report a reduction in spliced XBP (sXBP1) —suggesting altered IRE-α regulation between IPF fibroblasts and normal fibroblasts. We demonstrate that Bax-Inhibitor-1 (BI-1), a protein overexpressed in IPF lungs, decreases ER stress sensitivity by negatively regulating IREα; knockdown of BI-1 resulted in the rescue of a normal fibroblast phenotype demonstrated by abrogating the antiapoptotic phenotype, reducing cell migratory capacity, and attenuating activated morphology. Upon exploring other chaperone proteins, FKBP10 was identified as a potential therapeutic target due to its role in collagen synthesis. We confirmed FKBP10-LH2 protein:protein interaction by proximity assay and interfering with that binding using Tacrolimus and its non-immunosuppressant analogue resulted in decreased collagen secretion and deposition. Finally, to selectively deliver drugs, mitotropic liposomes were prepared by thin-film hydration, characterized, and tested in vitro. Trafficking was directed using the novel theory that, conjugation of a TPP cation to the surface of a lipid-based nanocarrier would enhance drug efficacy by delivering directly to IPF fibroblasts with hyperpolarized mitochondria. Collectively our data characterizes multiple senescence-associated apoptosis resistance mechanisms in IPF fibroblasts and demonstrates the utility of targeted, small molecule intervention of the observed pathways.