The cell type specific sequences of transcriptional programs during lung regeneration have remained elusive. Using time-series single cell RNA-seq of the bleomycin lung injury model, we resolved transcriptional dynamics for 28 cell types. Trajectory modeling together with lineage tracing revealed that airway and alveolar stem cells converge on a unique Krt8 + transitional stem cell state during alveolar regeneration. These cells have squamous morphology, feature p53 and NFkB activation and display transcriptional features of cellular senescence. The Krt8+ state appears in several independent models of lung injury and persists in human lung fibrosis, creating a distinct cell-cell communication network with mesenchyme and macrophages during repair. We generated a model of gene regulatory programs leading to Krt8+ transitional cells and their terminal differentiation to alveolar type-1 cells. We propose that in lung fibrosis, perturbed molecular checkpoints on the way to terminal differentiation can cause aberrant persistence of regenerative intermediate stem cell states.

In the heart, autophagy is required for normal cardiac function and also has been implicated in cardiovascular disease.

Transcriptional regulatory mechanisms of cardiac oxidative stress resistance are not well defined. FoxO transcription factors are critical mediators of oxidative stress resistance in multiple cell types, but cardioprotective functions have not been reported previously. FoxO function in oxidative stress resistance was investigated in cultured cardiomyocytes and in mice with cardiomyocyte-specific combined deficiency of FoxO1 and FoxO3 subjected to myocardial infarction (MI) or acute ischemia/reperfusion (I/R) injury. Induction of oxidative stress in cardiomyocytes promotes FoxO1 and FoxO3 nuclear localization and target gene activation. Infection of cardiomyocytes with a dominant-negative FoxO1(⌬256) adenovirus results in a significant increase in reactive oxygen species and cell death, whereas increased FoxO1 or FoxO3 expression reduces reactive oxygen species and cell death. Mice generated with combined conditional deletion of FoxO1 and FoxO3 specifically in cardiomyocytes were subjected to I/R or MI. Loss of FoxO1 and FoxO3 in cardiomyocytes results in a significant increase in infarct area with decreased expression of the antiapoptotic molecules, PTEN-induced kinase1 (PINK1) and CBP/P300-interacting transactivator (CITED2). Expressions of the antioxidants catalase and manganese superoxide dismutase-2 (SOD2) and the autophagy-related proteins LC3II and Gabarapl1 also are decreased following I/R compared with controls. Mice with cardiomyocyte-specific FoxO deficiency subjected to MI have reduced cardiac function, increased scar formation, induction of stress-responsive signaling, and increased apoptotic cell death relative to controls. These data support a critical role for FoxOs in promoting cardiomyocyte survival during conditions of oxidative stress through induction of antioxidants and cell survival pathways.Ischemic heart disease or myocardial ischemia/reperfusion (I/R) 2 injury is a leading cause of mortality worldwide (1).Cardiac oxidative injury can lead to cardiomyocyte cell death followed by fibrosis, hypertrophy, ventricular chamber dilation, decreased cardiac function, and ultimately heart failure.Oxidative stress is associated with increased formation of reactive oxygen species (ROS) that contribute to the pathophysiology of I/R injury (1). Increased ROS leads to DNA, protein and lipid modifications, as well as activates stress-signaling pathways leading to heart failure in I/R injury (2). Protective factors in ROS-mediated cardiac injury include AMP-activated protein kinase (AMPK) and sirtuins (Sirts) (3, 4). Understanding the intracellular processes that protect the heart from oxidative damage following I/R injury is important for the development of therapies to prevent the progression of heart disease. FoxO transcription factors (FoxO1, FoxO3, FoxO4, and FoxO6) belong to the forkhead family of transcriptional regulators, of which FoxO1 and FoxO3 are expressed in developing and adult cardiomyocytes (5, 6). Mice lacking FoxO1 are embryonic lethal by E10.5 due to impaired vasculogenesis, and mice l...