Collagen binding and mimetic peptide-functionalized self-assembled peptide hydrogel enhance chondrogenic differentiation of human mesenchymal stem cells.

The avascular structure and low cell migration to the damaged area due to the low number of cells do not allow spontaneous repair of the articular cartilage tissue. Therefore, functional scaffolds obtained from biomaterials are used for the regeneration of cartilage tissue. Here, we functionalized one of the self-assembling peptide (SAP) scaffolds KLD (KLDLKLDLKLDL) with short bioactive motifs, which are the α1 chain of type II collagen binding peptide WYRGRL (C1) and the triple helical collagen mimetic peptide GFOGER (C2) by direct coupling. Our goal was to develop injectable functional SAP hydrogels with proper mechanical characteristics that would improve chondrogenesis. Scanning electron microscopy (SEM) was used to observe the integration of peptide scaffold structure at the molecular level. To assure the stability of SAPs, the rheological characteristics and degradation profile of SAP hydrogels were assessed. The biochemical study of the DNA, glycosaminoglycan (GAG), and collagen content revealed that the developed bioactive SAP hydrogels greatly increased hMSCs proliferation compared with KLD scaffolds. Moreover, the addition of bioactive peptides to KLD dramatically increased the expression levels of important chondrogenic markers such as aggrecan, SOX-9, and collagen Type II as evaluated by real-time polymerase chain reaction (PCR). We showed that hMSC proliferation and chondrogenic differentiation were encouraged by the developed SAP scaffolds. Although the chondrogenic potentials of WYRGRL and GFOGER were previously investigated, no study compares the effect of the two peptides integrated into 3-D SAP hydrogels in chondrogenic differentiation. Our findings imply that these specifically created bioactive peptide scaffolds might help enhance cartilage tissue regeneration.

Role of functionalized self-assembled peptide hydrogels in in vitro vasculogenesis.

Fabrication of vascularized tissue constructs plays an integral role in creating clinically relevant tissues. Scaffold materials should be sufficiently vascularized to mimic functional and complex native tissues. Herein, we report the development of bioactive and biomimetic self-assembled peptide (SAP) hydrogels that allow the rapid formation of a vascular structure in vitro. The KLDLKLDLKLDL (KLD peptide) SAP was functionalized with laminin derived peptides IKVAV (V1) and YIGSR (V2) through direct coupling to mimic the natural extracellular matrix (ECM) and human umbilical endothelial cells (HUVECs) and mesenchymal stem cells (MSCs) cultured in 0.5% and 1% SAP hydrogels organized into vascularized structures. Atomic force microscopy (AFM) and scanning electron microscopy (SEM) images proved the molecular integration of the nanofibrous structure in SAP hydrogels. The stability of SAP hydrogels was confirmed by rheological and degradation measurements. Bioactive peptide scaffolds enhanced significantly HUVEC/hMSC proliferation depicted by MTT analysis compared to KLD. Furthermore, the real time quantitative polymerase chain reaction (rt-PCR) was performed to analyse vascular gene expressions such as platelet/endothelial cell adhesion molecule-1 (PECAM-1), von Willebrand factor (vWF), and vascular endothelial cadherin (VE-cadherin). The results indicated that the KLD-V2 hydrogel significantly induced vasculogenesis in hMSC/HUVEC co-culture compared to KLD-V1, Biogelx and KLD because YIGSR in KLD-V2 promoted cell population and ECM secretion by the interaction with cells and increased vasculogenesis. Overall, the designed SAP hydrogel represents an effective scaffold for vascularization of tissue constructs with useful tissue engineering applications.

Enhanced osteogenesis of human mesenchymal stem cells by self-assembled peptide hydrogel functionalized with glutamic acid templated peptides.

Self-assembling peptide (SAP) hydrogel has been shown to be an excellent biological material for three-dimensional cell culture and stimulatie cell migration and differentiation into the scaffold, as well as for repairing bone tissue defects. Herein, we designed one of the SAP scaffolds KLD (KLDLKLDLKLDL) through direct coupling to short bioactive motif O1 (EEGGC) and O2 (EEEEE) of which bioactivity on osteogenic differentiation was previously demonstrated and self-assembled in different concentrations (0.5%, 1%, and 2%). Our aim was to enhance osteogenesis and biomineralization of injectable SAP hydrogels with controlled mechanical properties so that the peptide hydrogel also becomes capable of being injected to bone defects. The molecular integration of the nanofibrous peptide scaffolds was observed using atomic force microscopy (AFM) and scanning electron microscopy (SEM). The rheological properties and degradation profile of SAP hydrogels were evaluated to ensure stability of SAPs. Compared with pure KLD scaffold, we found that these designed bioactive peptide scaffolds significantly promoted hMSCs proliferation depicted by biochemical analysis of alkaline phosphatase (ALP) activity, total calcium deposition. Moreover, key osteogenic markers of ALP activity, collagen type I (COL-1), osteopontin (OP), and osteocalcin (OCN) expression levels determined by real-time polymerase chain reaction (PCR) and immunofluorescence analysis were also significantly increased with the addition of glutamic acid residues to KLD. We demonstrated that the designed SAP scaffolds promoted the proliferation and osteogenic differentiation of hMSCs. Our results suggest that these designed bioactive peptide scaffolds may be useful for promoting bone tissue regeneration.