Project Summary/Abstract Survivors of acute respiratory failure (ARF) often face a daunting return to normal life as they have significant impairment of physical function. Over half of these patients are never able to return to work and often sustain significant skeletal muscle loss. Reliable algorithms to predict long-term physical function are currently unavailable. We previously described how metabolomics are predictive of acute outcomes in sepsis and ARF. Many of these metabolomic changes are related to dysfunction of β-oxidation, the citric acid cycle (TAC) and nicotinamide adenine dinucleotide (NADH) metabolism. Mitochondrial dysfunction is now considered to be a major driver of sepsis outcomes and may play a role in sepsis-induced muscle dysfunction. Furthermore, mitochondrial DNA (mtDNA) damage-associated molecular patterns (DAMPs) predicts patient outcomes in critical illness and trauma. We have developed a novel and innovative RNA target-bait capture deep sequencing and bioinformatics protocol for accurate quantitation and differentiation of mtDNA DAMPs and nuclear mitochondrial pseudogenes We hypothesize that mtDNA damage-associated molecular patterns (DAMPs) or metabolomic changes are reflective of mitochondrial-related bioenergetic status, and will also predict physical function in six-month survivors of ARF. In support of this hypothesis, we found in a small pilot study that mitochondrial related metabolites measured at patient discharge correlated with poor physical function in six- month survivors of ARF. In this study, we propose to expand upon the metabolomic and mtDNA results by leveraging patient data and plasma samples that were collected within the single center, randomized clinical trial at Wake Forest Baptist Medical Center (TARGET; ClinicalTrials.gov Identifier: NCT00976833). This study was designed to test if physical therapy initiated in the ICU reduced ICU length-of-stay and improved longterm quality- of-life in survivors. In Aim 1, we will establish that increased concentrations of mtDNA DAMPs in cell-free plasma predict physical function in survivors of ARF. Aim 2 will investigate the link between the concentration of mitochondrial-related bioenergetics metabolites and the abundance of mtDNA DAMPs in cell-free plasma. The results from this study will provide new pathophysiologic insight into the role of mtDNA DAMPs and mitochondrial-associated metabolites in ARF-induced poor physical function. New pharamcological therapies have the potential to improve physical function through improved mitochondrial function, biogenesis, and reduction of mtDNA DAMPs, while NAD-derivatives or androgenic steroid repletion could improve physical strength and function.