Abstract
Systemic sclerosis (SSc) is a progressive autoimmune disease marked by persistent fibroblast activation, dysregulated extracellular matrix (ECM) remodeling, and irreversible fibrosis. While TGF‑β and PDGF drive fibroblast-to-myofibroblast differentiation, mechanisms that restrain proteolysis and restore matrix homeostasis remain poorly defined. Tissue inhibitor of metalloproteinases 3 (TIMP3) uniquely inhibits MMPs, ADAMs, and ADAMTSs, positioning it as a key regulator of pericellular remodeling. However, prior studies have focused on transcript-level analyses, potentially obscuring protein-level dysregulation and limiting understanding of TIMP3 loss in SSc.
We identify a robust deficiency of TIMP3 protein in SSc dermal fibroblasts despite detectable mRNA. Using primary patient-derived fibroblasts, multi-omic profiling, and functional matrix assays, we show this loss is driven by post-transcriptional mechanisms, including impaired exon-spanning mRNA processing and enrichment of TIMP3-targeting microRNAs, rather than promoter hypermethylation. Phosphoproteomics reveal signaling rewiring, with attenuated SMAD and p38 signaling alongside sustained activation of MAPK/ERK, STAT1/3/6, and non-canonical NF‑κB pathways, decoupling TIMP3 expression from profibrotic signaling.
Functionally, TIMP3 suppression in healthy fibroblasts increases collagen deposition and recapitulates key SSc myofibroblast features, while restoration limits matrix accumulation, demonstrating reversibility of fibrotic remodeling. Circulating TIMP3 is reduced in SSc patients and correlates with skin and lung disease severity. Together, these findings identify TIMP3 deficiency as a previously unrecognized regulatory vulnerability integrating post-transcriptional control, signaling, and ECM remodeling to sustain fibrosis.