Investigation of Collagen Network Microstructure and Vascular Sprouting On-Chip Using Non-enzymatic Glycation

Non-enzymatic glycation of collagen, a post-translational modificationprocess, creates extra crosslinks between collagen molecules. The accumulation of advanced glycation end-product crosslinks is one of the main phenomena observed in connective tissue of subjects with diabetes and aging, and many studies showed the alteration of microstructure of collagen network by these glycation-mediated crosslinks. However, the effects of the glycation-induced collagen network changes on adjacent cells, particularly in growth and tissue formation, remain largely unexplored. Here, we present a microfluidic model to investigate vascular sprouting in glycated collagen networks under well-controlled conditions. Using scanning electron microscopy, fluorescence labeling microscopy (NHS-ester), and label-free microscopy (second harmonic generation microscopy), we characterized the microstructure of collagen networks across different ribose concentrations. Three-dimensional vascular sprouting analysis revealed that the higher ribose concentrations resulted in longer and thicker vascular sprouts. Finally, we investigated the alteration of gene expressions in endothelial cells by glycated collagen hydrogels with VEGF gradients. This study enhances our understanding of how glycation-induced changes in collagen affect vascular growth, providing insights into tissue formation in pathological conditions.