Dermal fibroblasts cultured from aged human skin display a pro‐inflammatory phenotype

Abstract Dermal fibroblasts synthesize, organize, and reside within the dermal extracellular matrix (ECM), which provides mechanical support to the skin and a microenvironment critical for dermal cell function. During aging, the ECM becomes fragmented, a change associated with clinical manifestations such as thin, fragile skin and altered fibroblast function, including reduced expression of ECM structural genes and increased expression of pro‐inflammatory mediators. We investigated whether functional alterations of fibroblasts in aged skin reflect adaptation to the aged dermal ECM microenvironment or intrinsic aging responses. Primary dermal fibroblast cultures were established from young (20–30 years; 6 females, 6 males) and aged (>80 years; 6 females, 6 males) sun‐protected skin. No differences in early growth kinetics were observed between groups; however, fibroblasts from aged individuals grew more slowly after approximately 25 population doublings and had a 20% lower final population doubling level. Prior to passage, after brief primary culture (6–8 days), the average telomere length of fibroblasts from aged donors was reduced by approximately 1000 base pairs (∼1 kb). Global gene expression revealed 477 differentially expressed genes (DEGs; padj ≤0.05, absolute log2FC ≥ 0.58), comprising 300 upregulated and 177 downregulated transcripts. Upregulated DEGs and associated protein–protein interaction networks were enriched in cytokine‐ and chemokine‐mediated signaling pathways ( p < 10 −5 ). Downregulated DEGs were enriched in lipid biosynthesis and fatty acid metabolism. Notably, fibroblasts from young and aged skin expressed similar levels of genes encoding ECM structural proteins, including collagens (42 genes), proteoglycans (29 genes), and glycoproteins (157 genes). These findings indicate that fibroblasts from aged skin retain a pro‐inflammatory transcriptional phenotype in primary culture, resembling their phenotype in vivo. In contrast, primary fibroblasts from aged skin express youthful levels of ECM structural genes in primary culture, suggesting that reduced ECM gene expression in vivo reflects dynamic adaptation to the aged dermal microenvironment rather than permanent reduction of synthetic capacity.