Single-Cell Omics Approaches in Disease Mechanism Discovery

Abstract

Single-cell omics technologies--encompassing single-cell RNA sequencing (scRNA-seq), single-cell ATAC-seq
(scATAC-seq), single-cell proteomics, spatial transcriptomics, and multi-modal CITE-seq--have fundamentally
transformed the resolution at which cellular heterogeneity, developmental trajectories, and disease mechanisms can be
characterised, enabling the dissection of complex tissues into their constituent cell types and states at unprecedented
granularity. This study applies an integrated single-cell multi-omics framework--combining scRNA-seq (10x Genomics
Chromium, 3' v3.1), scATAC-seq, and CITE-seq with 200 surface protein markers--to 847,293 single cells from tissue
biopsies of three disease conditions: non-alcoholic steatohepatitis (NASH, liver; n=42 patients), idiopathic pulmonary
fibrosis (IPF, lung; n=38 patients), and treatment-naive colorectal cancer (CRC, colon; n=45 patients), compared against
matched healthy controls (n=41). Unbiased clustering identified 94 distinct cell clusters including 23 novel cell states not
previously described in existing single-cell atlases for these tissues. Disease-specific regulatory networks reconstructed
from scATAC-seq chromatin accessibility data identified 1,847 disease-associated transcription factor binding sites, of
which 312 co-localised with GWAS risk loci, providing mechanistic links between genetic predisposition and gene
regulatory changes at single-cell resolution. Trajectory analysis revealed a previously undescribed hepatic stellate cell
activation intermediate state in NASH that precedes full myofibroblast differentiation and represents a potential
therapeutic intervention window. Ligand-receptor interactome analysis identified 47 novel cross-cell-type signalling axes
disrupted in disease, including a macrophage-to-fibroblast TGFB1-TGFBR2 axis in both IPF and NASH that may
represent a shared therapeutic target across fibrotic diseases.