Regenerative Potential of Fibroblast Secretome: Keratinocyte Growth Factor (KGF) and Total Protein Effects on Senescent Cell Proliferation via Mek 1/2 Activation and Enhanced Collagen Type 1 Utilization

Aging leads to reduced physical, mental, and social performance, lowering productivity. The fibroblast secretome, rich in bioactive proteins, growth factors, and enzymes, is essential for cell repair and regeneration. This study evaluated its regenerative potential by examining MEK1/2 pathway activation and type I collagen utilization to enhance senescent fibroblast proliferation.

Young (passage 3) and senescent (passage 24) fibroblasts were cultured in DMEM. Senescent cells were treated with 10% or 20% fibroblast secretome. Culture media were collected on days 1, 3, 5, and 7 for analysis of total protein, KGF, cell proliferation, MEK phosphorylation, and collagen concentration.

The 20% secretome contained 5830,67 ± 181,62 µg/mL total protein and 1712,67 ± 7,19 pg/mL KGF. Senescent fibroblasts treated with 20% secretome showed a 44% proliferation increase on day 5 (21135,67 ± 1392,89 cells/well) versus controls (14637 ± 2250,57 cells/well, p<0,05). MEK1/2 phosphorylation rose from 0,01 ± 0,00 pg/mL at 45 minutes to 0,18 ± 0,03 pg/mL at 24 hours (p<0,05), indicating sustained activation.

Secretome treatment enhanced MEK1/2 signaling and proliferation, with a 12% reduction in extracellular type I collagen on day 5 (2,67 ± 0,03 ng/mL vs. 3,02 ± 0,03 ng/mL, p<0,05), reflecting increased collagen utilization and matrix remodeling.

The 20% fibroblast secretome enriched with KGF and bioactive proteins promotes senescent fibroblast proliferation via MEK1/2 activation and optimized collagen dynamics, supporting its potential in regenerative medicine for aging and tissue repair.