Single Cell Transcriptomics of Fibrotic Lungs Unveils Aging-associated Alterations in Endothelial and Epithelial Cell Regeneration

Abstract

Lung regeneration deteriorates with aging leading to increased susceptibility to pathologic conditions, including fibrosis. Here, we investigated bleomycin-induced lung injury responses in young and aged mice at single-cell resolution to gain insights into the cellular and molecular contributions of aging to fibrosis. Analysis of 52,542 cells in young (8 weeks) and aged (72 weeks) mice identified 15 cellular clusters, many of which exhibited distinct injury responses that associated with age. We identified Pdgfra+ alveolar fibroblasts as a major source of collagen expression following bleomycin challenge, with those from aged lungs exhibiting a more persistent activation compared to young ones. We also observed age-associated transcriptional abnormalities affecting lung progenitor cells, including ATII pneumocytes and general capillary (gCap) endothelial cells (ECs). Transcriptional analysis combined with lineage tracing identified a sub-population of gCap ECs marked by the expression of Tropomyosin Receptor Kinase B (TrkB) that appeared in bleomycin-injured lungs and accumulated with aging. This newly emerged TrkB+ EC population expressed common gCap EC markers but also exhibited a distinct gene expression signature associated with aberrant YAP/TAZ signaling, mitochondrial dysfunction, and hypoxia. Finally, we defined ACKR1+ venous ECs that exclusively emerged in injured lungs of aged animals and were closely associated with areas of collagen deposition and inflammation. Immunostaining and FACS analysis of human IPF lungs demonstrated that ACKR1+ venous ECs were dominant cells within the fibrotic regions and accumulated in areas of myofibroblast aggregation. Together, these data provide high-resolution insights into the impact of aging on lung cell adaptability to injury responses.

A high-resolution view of the heterogeneous aging endothelium

“…This study, in which most patients were of older age, also observed a loss of general capillary and aerocyte ECs in both age-related diseases compared to healthy control tissue, as well as an increase of venous EC populations. Similar observations were made in drug-induced IPF in mice, where the influence of age on disease outcome was observable by reduced aerocyte abundance, an increase of venous ECs, and remodeling of the vasculature in 18-month old mice, stressing the impact of ECs in this condition [ 77 ]. In fibrotic conditions (COVID-19, IPF), an enrichment of genes involved in cellular stress, as well as signatures suggestive of hampered immunoregulation and loss of vessel wall integrity were furthermore found [ 7 ].…”

“…From a human point-of-view, while extensive atlasing efforts [ 9 , 75 , 76 ] allowed for an all-encompassing overview of endothelial populations in healthy human lung tissue, a thorough interrogation of how aging affects these is still lacking. However, multiple studies analyzed age-related pulmonary diseases, such as IPF (idiopathic pulmonary fibrosis), COPD (chronic obstructive pulmonary disorder) and COVID-19 [ 7 , 66 , 77 , 78 ], and helped reveal EC phenotypes and gene expression signatures that may be at play in healthy aging as well. For instance, in human lung tissue from IPF, smokers, and COPD patients, a specialized systemic/peribronchial EC population was found, defined by the marker gene COL15A1 , which was enriched in IPF lungs around fibrotic areas [ 66 ].…”

Stricturing Crohn’s disease single-cell RNA sequencing reveals fibroblast heterogeneity and intercellular interactions

“…20 ADAMDEC1 is critical in the fibroblastic response to tissue remodeling and healing during colitis 21 and PDGFRA+ fibroblasts display a proliferative phenotype and are a major source of collagen. 22,23 Based on the prominent role of fibroblasts in CDs signaling, the differences largely restricted to the LP1/2 fraction, the over-representation of distinct fibroblast types, and the strong contribution to transcriptional changes in CDs, we sought a unifying molecule and identified CDH11 as the only cell surface receptor shared by the CXCL14+ and MMP/WNT5A+ fibroblasts. While also expressed in macrophages, CDH11 is primarily responsible for mediating homophilic stromal cell-cell interactions, [24][25][26][27] is upregulated in fibrotic disorders of the lung, 24 liver, 25 skin, 26 intestine, 27 and its inhibition attenuates fibrosis in multiple animal models.…”

Multi-omics analysis of human tendon adhesion reveals ACKR1 migrates macrophages involve in endogenous regeneration

“…Endothelial cells were detected in multiple organs fibrosis and adhesion tissues, although their function is unknown. For example, RGCC was linked to renal fibrosis, while ACKR1 + ECs promoted pulmonary fibrosis 61,62 . Besides, the similar profibrotic phenomenon of ACKR1 + ECs was also found in hepatic fibrosis 6 .…”