An Integrin-Specific Collagen-Mimetic Peptide Approach for Optimizing Hep3B Liver Cell Adhesion, Proliferation, and Cellular Functions

Abstract

This study focused on mimicking collagen structurally and biologically using various peptide sequences toward realizing an artificial collagen-like biomaterial. Collagen-mimetic peptides (CMPs) incorporating integrin-specific glycine-phenylalanine-hydroxyproline-glycine-glutamate-arginine (GFOGER) sequence from residues 502 to 507 of collagen alpha(1)(I) were used as a bioadhesive matrix and grafted onto poly(3-hydroxybutyrate-co3-hydroxyvalerate) microspheres to optimize cell adhesion, proliferation, and functions. Cell recognition of these biomolecules appeared to be conformation dependent, with the CMP1 of higher triple helix stability being preferred. Absence of the GFOGER hexapeptide in the CMP1' and CMP2' caused an adverse effect on the level of cell adhesion (<10%). The GFOGER-containing triple-helical CMPs effectively inhibited cell adhesion to collagen in a competition assay. The cell-adhesion activity of the CMP1 was approximately 50% of that of collagen. The cell spreading on the CMP1 was comparable with that observed on collagen. The presence of the CMP1 promoted cell attachment and spreading on the microspheres and extensive cell proliferation and bridging. Slower cell proliferation was observed on the blank microspheres. Live-dead assay showed that most cells are viable after 10-day culture. The presence of CMP1 on the microspheres maintained the albumin secretion and P-450 activity levels of the liver cells for up to 14 days. Our results established the potential of CMP1 to create a collagen-like microenvironment for optimizing cellular responses for liver tissue engineering.

Biodegradable poly(ethylene glycol)–peptide hydrogels with well-defined structure and properties for cell delivery

“…The existence of micro-fibrils may be caused by the interaction between the peptide molecules. A similar phenomenon was reported by Tong et al on collagen mimetic peptides, which formed 3-dimensional nano/micro-fibril network upon being grafted onto polymer microspheres [35]. The formation of fibrils by peptides was reported to be folded in a b-sheet conformation [36].…”

Integrin α3β1 Binding to Fibronectin Is Dependent on the Ninth Type III Repeat

“…Further evidence suggesting that alternate conformational states induce integrin "switching" comes from studies in which conformational stability was conferred to the ninth or tenth type III repeats, resulting in modulation of integrin affinity. In these studies, the ninth type III repeat was stabilized via a Leu-Pro mutation at amino acid 1408 (22,23), or the tenth type III repeat was stabilized through facilitation of greater hydrogen bonding within the repeat (24). In both cases, stabilization of the relative positions of the two repeats resulted in increased affinity for integrin ␣5␤1 over integrin ␣v␤3.…”

Emerging Concepts in Engineering Extracellular Matrix Variants for Directing Cell Phenotype

“…The effect of force-mediated unfolding of FN on integrin engagement and activity levels may be further explained by studies demonstrating that the degrees of conformational stability of the ninth (or tenth) FNIII repeat can modulate integrin accessibility of the RGD motif. A stabilization of the ninth FNIII repeat via a Leu–Pro point mutation at amino acid 1408 [34], or stabilization of the hydrogen bonding within the tenth type-III repeat [35], increases affinity for α 5 β 1 over α V β 3 .…”

“…It has been shown that integrin recognition of this sequence is entirely dependent on the conformation of the sequence; it must be presented in a triple-helical conformation similar to that of native collagen [35,55]. Tong et al produced a series of collagen-mimetic peptides containing the GFOGER sequence and showed that while the GFOGER sequence is critical for cell binding, the triple-helical context was critical for integrin recognition; cells were found to bind more tightly to peptides with higher triple-helix stability.…”