Effect of surface modification of nanofibres with glutamic acid peptide on calcium phosphate nucleation and osteogenic differentiation of marrow stromal cells

Abstract

Biomineralization is mediated by extracellular matrix (ECM) proteins with amino acid sequences rich in glutamic acid. The objective of this study was to investigate the effect of calcium phosphate deposition on aligned nanofibres surface-modified with a glutamic acid peptide on osteogenic differentiation of rat marrow stromal cells. Blend of EEGGC peptide (GLU) conjugated low molecular weight polylactide (PLA) and high molecular weight poly(lactide-co-glycolide) (PLGA) was electrospun to form aligned nanofibres (GLU-NF). The GLU-NF microsheets were incubated in a modified simulated body fluid for nucleation of calcium phosphate crystals on the fibre surface. To achieve a high calcium phosphate to fibre ratio, a layer-by-layer approach was used to improve diffusion of calcium and phosphate ions inside the microsheets. Based on dissipative particle dynamics simulation of PLGA/PLA-GLU fibres, > 80% of GLU peptide was localized to the fibre surface. Calcium phosphate to fibre ratios as high as 200%, between those of cancellous (160%) and cortical (310%) bone, was obtained with the layer-by-layer approach. The extent of osteogenic differentiation and mineralization of marrow stromal cells seeded on GLU-NF microsheets was directly related to the amount of calcium phosphate deposition on the fibres prior to cell seeding. Expression of osteogenic markers osteopontin, alkaline phosphatase (ALP), osteocalcin and type 1 collagen increased gradually with calcium phosphate deposition on GLU-NF microsheets. Results demonstrate that surface modification of aligned synthetic nanofibres with EEGGC peptide dramatically affects nucleation and growth of calcium phosphate crystals on the fibres leading to increased osteogenic differentiation of marrow stromal cells and mineralization.

A developmentally inspired combined mechanical and biochemical signaling approach on zonal lineage commitment of mesenchymal stem cells in articular cartilage regeneration

“…Stained samples were imaged with a Nikon Eclipse Ti-E inverted fluorescent microscope as described. 45 …”

Devitalized Stem Cell Microsheets for Sustainable Release of Osteogenic and Vasculogenic Growth Factors and Regulation of Anti‐Inflammatory Immune Response

“…Synthesis of Mineralized Nanofiber Microsheets : Low molecular weight poly( d,l ‐lactide) (LMPLA) was synthesized by ring‐opening polymerization of LA monomer as previously described . LMPLA was reacted with AC to form acrylate‐terminated LMPLA as previously described .…”

“…LMPLA was reacted with AC to form acrylate‐terminated LMPLA as previously described . Number average molecular weight () and polydispersity index of Ac‐LMPLA, determined from the chemical shifts in 1 H‐NMR spectrum as described previously, were 5.3 and 1.2 kDa, respectively. Amino acid sequence Glu‐Glu‐Gly‐Gly‐Cys (GLU peptide) was synthesized manually on Rink Amide NovaGel resin as previously described .…”

“…Number average molecular weight () and polydispersity index of Ac‐LMPLA, determined from the chemical shifts in 1 H‐NMR spectrum as described previously, were 5.3 and 1.2 kDa, respectively. Amino acid sequence Glu‐Glu‐Gly‐Gly‐Cys (GLU peptide) was synthesized manually on Rink Amide NovaGel resin as previously described . GLU peptide was conjugated to Ac‐LMPLA by Michael addition reaction to produce GLU‐LMPLA conjugate as previously described .…”

Multiscale assembly for tissue engineering and regenerative medicine

“…Biomaterial synthesis focuses on capturing cell-specific ECM features by incorporating cell-specific signaling molecules, matrix stiffness, and porosity. The biological properties of synthetic hydrogels can be tuned by incorporating peptides that enhance cell adhesion [e.g., arginine–glycine–aspartic acid (RGD) and glutamic acid peptide] [111]. Microenvironments can be engineered with photopatterning, where ECM proteins or soluble factors can be spatially organized within 3D PEG-based hydrogels, thereby controlling the distribution of encapsulated mesenchymal stem cells [112].…”