Revealing the Dynamic Interplay between Lymphangioleiomyomatosis Cells and T Cells in LAM Progression

Abstract

Lymphangioleiomyomatosis (LAM) is a rare lung disease characterized by the abnormal metastasis of proliferative smooth muscle-like cells into the lungs. It primarily affects women of childbearing age, causing respiratory-related symptoms and eventually lung collapse. The only FDA-approved treatment for LAM is rapamycin, a mTORC1 inhibitor, which only slows down disease progression, rather than providing a cure. Additionally, not all patients respond to the treatment. To expand our understanding of the mechanism of LAM progression, we analyzed the scRNAseq data of LAM patients and observed an elevated CD8 T cell deposition, which guided the rationale to incorporate T cells in our study. We performed NanoString analysis on 11 LAM patients' FFPE lung tissues and 11 control lung tissues to highlight the transcriptome changes in LAM tissue. We then co-cultured allogenic primary female human T cells with PSC-derived TSC2null smooth muscle cells to investigate the crosstalk between these two cell types.

We revealed a cancer-associated fibroblast (CAF) signature of TSC2null cells that can be promoted by T cells and be inhibited by IFN-g. In addition, we demonstrated that TSC2null cells lose their CD40 expression leading to T cell hyporesponsiveness, promoting TSC2null cell proliferation. Notably, these phenotypical changes are supported and complemented by our NanoString analysis. With the ultimate goal of co-culturing T cells with LAM cells, we systematically engineered viscoelastic hydrogels by integrating β-sheet peptides with hyaluronic acid (HA) to tune viscoelasticity independently of

stiffness. Our study explored the impact of specific amino acids in the peptide sequence on gel microstructure and peptide assembly, elucidating their role in macro-scale viscoelasticity. These gel systems allowed us to replicate the viscoelasticity of various human tissues, including human lungs. Moreover, these hydrogels are cytocompatible as demonstrated with both 2D and 3D cell cultures, allowing encapsulation of primary human T cells. Collectively, we discovered the dynamic interplay between TSC2null and T cells and established a platform for ultimate use for cell co-culture and drug screening.