A naturally occurring urinary collagen type I alpha 1-derived peptide inhibits collagen type I-induced endothelial cell migration at physiological concentrations
Collagen type I (COL(I)) is a key component of the extracellular matrix (ECM) and is involved in cell signalling and migration through cell receptors. Collagen degradation produces bioactive peptides (matrikines), which influence cellular processes. In this study, we investigated the biological effects of nine most abundant, naturally occurring urinary COL(I)-derived peptides on human endothelial cells at physiological concentrations, using cell migration assays, mass spectrometry-based proteomics, flow cytometry, and AlphaFold3. While none of the peptides significantly altered endothelial migration by themselves at physiological concentrations, full-length COL(I) increased cell migration, which was inhibited by Peptide 1 (229NGDDGEAGKPGRPGERGPpGp249). This peptide uniquely contains the DGEA and GRPGER motifs, interacting with integrin 2{beta}1. Flow cytometry confirmed the presence of integrin 2{beta}1 on human endothelial cells, and AlphaFold3 modelling predicted an interaction between Peptide 1 and integrin 2. Mass spectrometry-based proteomics investigating signalling pathways revealed that COL(I) triggered phosphorylation events linked to integrin 2{beta}1 activation and cell migration, which were absent in COL(I) plus peptide 1-treated cells. These findings identify Peptide 1 as a biologically active COL(I)-derived peptide at a physiological concentration capable of modulating collagen-induced cell migration, and provide a foundation for further investigation into its mechanisms of action and role in urine excretion.