Proteomic and phenotypic profiling of replicative-senescent human foreskin fibroblasts under brief heat shock.

As the largest human organ, the skin experiences lifelong exposure to intrinsic/extrinsic factors that over time diminish its functional capacity and structural integrity. Skin aging involves cellular dysfunction and the loss or fragmentation of extracellular matrix (ECM) fibers, clinically presenting as wrinkles, slackening, and pigmentary abnormalities. The heat shock response (HSR) is a gene regulatory program that controls the expression of molecular chaperones associated with aging, cancer, and neurodegenerative disorders. By maintaining cellular homeostasis and facilitating DNA repair, HSR exerts protective effects against skin aging, as utilized in aesthetic technologies such as radiofrequency and focused ultrasound. This study aimed to investigate the mechanism, optimal conditions, and potential risks of short-term heat shock (HS) on the senescence process of human foreskin fibroblasts (HFF-1), providing experimental evidence to support the application of thermal stress in delaying skin aging. A replicative senescence model of HFF-1 cells was first established. Subsequently, cells were subjected to HS at 41 °C, 45 °C, and 49 °C, with a control group maintained at 37 °C. Assessments, including cell proliferation and viability assays, apoptosis analysis, reactive oxygen species (ROS) levels, Western blot, and heat shock proteins (HSPs) mRNA expression, demonstrated that 49 °C HS induced irreversible cellular damage. In contrast, 30 min HS at 41 °C and 45 °C attenuated senescence-associated phenotypes to varying extents under our experimental conditions.