mTORC1 activation in lung mesenchyme drives sex- and age-dependent pulmonary structure and function decline

Abstract

Lymphangioleiomyomatosis (LAM) is a rare fatal cystic lung disease due to bi-allelic inactivating mutations in tuberous sclerosis complex (TSC1/TSC2) genes coding for suppressors of the mechanistic target of rapamycin complex 1 (mTORC1). The origin of LAM cells is still unknown. Here, we profile a LAM lung compared to an age- and sex-matched healthy control lung as a hypothesis-generating approach to identify cell subtypes that are specific to LAM. Our single-cell RNA sequencing (scRNA-seq) analysis reveals novel mesenchymal and transitional alveolar epithelial states unique to LAM lung. This analysis identifies a mesenchymal cell hub coordinating the LAM disease phenotype. Mesenchymal-restricted deletion of Tsc2 in the mouse lung produces a mTORC1-driven pulmonary phenotype, with a progressive disruption of alveolar structure, a decline in pulmonary function, increase of rapamycin-sensitive expression of WNT ligands, and profound female-specific changes in mesenchymal and epithelial lung cell gene expression. Genetic inactivation of WNT signaling reverses age-dependent changes of mTORC1-driven lung phenotype, but WNT activation alone in lung mesenchyme is not sufficient for the development of mouse LAM-like phenotype. The alterations in gene expression are driven by distinctive crosstalk between mesenchymal and epithelial subsets of cells observed in mesenchymal Tsc2-deficient lungs. This study identifies sex- and age-specific gene changes in the mTORC1-activated lung mesenchyme and establishes the importance of the WNT signaling pathway in the mTORC1-driven lung phenotype.

CrossTORC and WNTegration in Disease: Focus on Lymphangioleiomyomatosis

“…However, this was unable to be investigated in vivo since LAM lung cells have not been able to be grown in mice [ 7 , 8 ]. Wnt pathway genes were also observed in LAM-specific mesenchymal lung cells by two important single-cell RNA sequencing (scRNAseq) studies in LAM [ 44 , 45 ].…”

“…Our research showed upregulation of the Wnt pathway in LAM-lung-specific mesenchymal cells [ 45 ]. To follow this up in vivo, we deleted the Tsc2 gene, specifically in mouse lung mesenchymal progenitor cells, resulting in progressive age-dependent enlargement of alveolar spaces, as well as pulmonary lung function decline [ 45 ].…”

“…Our research showed upregulation of the Wnt pathway in LAM-lung-specific mesenchymal cells [ 45 ]. To follow this up in vivo, we deleted the Tsc2 gene, specifically in mouse lung mesenchymal progenitor cells, resulting in progressive age-dependent enlargement of alveolar spaces, as well as pulmonary lung function decline [ 45 ]. Selected genes from the Wnt pathway, including Wnt3a, Wnt4, Wnt5a , and Wnt10b , encoding for Wnt ligands, Fzd1, Fzd2 , and Fzd8 , encoding for FZD receptors, were elevated in the Tsc2 -knockout lung mesenchyme, while Wnt3a and Wnt5a gene expression appeared to be downregulated by in vivo rapamycin treatment.…”

“…Rapamycin, however, did not reverse but slowed down the alveolar space enlargement phenotype. We highlighted elevated crosstalk between Tsc2 -deficient mesenchymal-derived cells and surrounding epithelial cells via canonical Wnt ligand/receptor signaling [ 45 ]. Genetic inactivation of Wnt signaling by deletion of the β-catenin-encoding gene ( cttnb1 ), in addition to mTORC1 activation in the mouse lung mesenchyme, reversed age-dependent alveolar enlargement.…”

“…Genetic inactivation of Wnt signaling by deletion of the β-catenin-encoding gene ( cttnb1 ), in addition to mTORC1 activation in the mouse lung mesenchyme, reversed age-dependent alveolar enlargement. However, Wnt pathway activation by genetically stabilized β-catenin in the lung mesenchyme was not sufficient for the development of the observed phenotype in this model [ 45 ]. Together, these data suggested that activation of both mTORC1 and canonical Wnt/β-catenin pathways act synergistically in triggering the development of the LAM-like phenotype.…”