Targeting redox to resorb ECM

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PROJECT SUMMARY Idiopathic Pulmonary Fibrosis (IPF) is a rapidly progressing and incurable disease. In healthy tissue, fibroblasts balance depositing and resorbing extracellular matrix, yet in fibrosis they produce excessive amounts of extracellular matrix while ECM degrading enzymes are downregulated. Many of these effects may be linked to a shift in balance to a more oxidative state due to dysregulated metabolism, including inhibition of Cathepsin K, a collagenolytic enzyme essential to normal lung function and development. Metabolic shifts are known to increase reactive oxygen species, which themselves alter the overall tissue oxidation-reduction (redox) state. We believe that to resolve fibrosis, therapeutics should restore the redox balance and repair defective metabolism. In this project, I will focus on identifying mechanisms regulating the redox state of the fibroblasts and transitions in this state in vivo during fibrosis and resolution. I will longitudinally monitor Col1a1-GFP+ fibroblasts through a thoracic window and image the autofluorescence of NADH and FAD, two indicators of the optical redox ratio. Changes to the redox ratio can be indicative of increased ROS. Therefore, by using a thoracic window to image the cellular redox ratio I can identify changes to the cellular stress in real-time, allowing me to directly compare changes during fibrosis progression to resolution phases. To further explore the relationship of fibroblast redox state to fibrosis resolution, I will explore these changes in mouse models of accelerated and non-resolving fibrosis. I will then mechanistically test the role of PGC1a as a regulator of the redox state in fibroblasts and will investigate the role the redox ratio plays in regulating Cathepsin K activity as a key link to fibrosis resolution. I hypothesize that endogenous or exogenous mechanisms that shift the redox state to be more reductive are essential to restore the fibroblast metabolic state to promote fibrosis resolution. I propose to test this hypothesis in two specific aims combining intravital imaging, in vitro cell culture, and precision cut lung slices in combination with selective activation or inhibition of proposed mechanistic pathway components. Together these combined studies will test the redox ratio as a required regulator of fibrosis resolution.