A comprehensive analysis of human infrapatellar fat pad tissue using single-nucleus RNA sequencing and spatial transcriptomics revealed new insights into its cellular composition and potential role in knee osteoarthritis. The study, published in Annals of the Rheumatic Diseases, identified distinct cell populations, transcriptomic profiles, and metabolic alterations in IFP tissue that varied based on osteoarthritis status, sex, and obesity.
Researchers analyzed infrapatellar fat pad (IFP) samples from 21 individuals (15 with knee osteoarthritis (KOA), 6 healthy controls) using single-nucleus RNA sequencing (snRNA-seq), spatial transcriptomics, and bioinformatics approaches. They sequenced a total of 82,924 nuclei, with 73,808 nuclei analyzed after filtering. Eight cell types were identified based on canonical markers: fibroblasts (44.35% of nuclei analyzed), macrophages (19.44%), adipocytes (16.41%), endothelial cells (12.12%), dendritic cells, smooth muscle cells, lymphocytes, and mast cells.
Five distinct fibroblast subclusters were characterized, each with unique transcriptomic profiles. The proportions of these subclusters ranged from 30.32% (subcluster 0) to 9.63% (subcluster 4). Pseudotime trajectory analysis suggested that fibroblasts expressing universal markers (DPP4+PI16+CD34+) may act as precursors for other fibroblast subclusters and adipocytes.
Comparing KOA to healthy control IFPs, 38 differentially expressed genes were identified in fibroblasts. The proportion of fibroblast subcluster 3 was significantly higher in healthy control IFPs compared to KOA samples. Pathway analysis of these genes revealed enrichment for cell adhesion and signaling pathways.
Sex-based differences in KOA IFP fibroblasts included 105 differentially expressed genes between female and male samples. Notably, the transcription factor CREB5 and lubricin-encoding gene PRG4 were upregulated in female KOA IFPs, suggesting potentially enhanced joint lubrication mechanisms.
When comparing obese (BMI 30-40 kg/m2) to normal BMI (18.5-24.9 kg/m2) KOA IFPs, 21 differentially expressed genes were identified in fibroblasts. CREB5 and PRG4 were downregulated in obese samples, indicating possible reduced joint lubrication in obese individuals with KOA.
Metabolomic analysis of fibroblast culture supernatants revealed 5 metabolites at significantly different levels between obese and normal BMI KOA IFPs. These included upregulated triglycerides and linoleic acid, and downregulated homoarginine in obese samples. Treatment with TGF-β or TNF-α further altered metabolite profiles, with 9 and 10 differentially detected metabolites, respectively.
The study took a multi-omic approach to provide a comprehensive map of cellular and transcriptomic diversity in human IFP tissue, highlighting potential mechanisms by which IFP fibroblasts may contribute to KOA pathogenesis. The study's findings suggested IFP fibroblasts play a significant role in IFP biology and OA pathology through transcriptional, metabolic, and cell differentiation mechanisms influenced by KOA status, sex, and obesity.
The authors noted some limitations, including the relatively small sample size and narrow age range of KOA participants. They suggested future studies focus on understanding the impact of age on IFP transcriptomics and investigating other major cell types within the tissue.
The authors declared having no competing interests.