Promoting differentiation of cultured myoblasts using biomimetic surfaces that present alpha-laminin-2 peptides

Abstract

Traditionally, muscle cell lines are cultured on glass coverslips and differentiated to investigate myoblast fusion and differentiation. Efficient differentiation of myoblasts produces a dense network of myotubes with the correct organisation for contraction. Here we have tested the ability of artificially generated, precisely controlled peptide surfaces to enhance the efficiency of myoblast differentiation. We focused on specific short peptides from α-laminin-2 (IKVSV, VQLRNGFPYFSY and GLLFYMARINHA) as well as residues 15–155 from FGF1. We tested if these peptides in isolation, and/or in combination promoted muscle differentiation in culture, by promoting fusion and/or by improving sarcomere organisation. The majority of these peptides promoted fusion and differentiation in two different mouse myogenic cell lines and in primary human myoblasts. The additive effects of all four peptides gave the best results for both mouse cell lines tested, while primary human cell cultures differentiated equally well on most peptide surfaces tested. These data show that a mixture of short biomimetic peptides can reliably promote differentiation in mouse and human myoblasts.

A1603P and K1617del, Mutations in β-Cardiac Myosin Heavy Chain that Cause Laing Early-Onset Distal Myopathy, Affect Secondary Structure and Filament Formation In Vitro and In Vivo

“…C2C12 myoblasts were seeded at the same cell density (1 × 10 5 cells/ml) on coverslips coated with laminin-1 (Sigma). After 24-h infection with adenovirus, the growth medium was exchanged for differentiation medium and the cells were incubated at 37 °C for 5 days to allow for differentiation into skeletal muscle myotubes, before being fixed as described [34] .…”

Uniaxially crumpled graphene as a platform for guided myotube formation

“…In addition to gene expression, the effect of extracellular matrix components on differentiation and subsequent production by the cells can be studied in the future. Previous researchers reported that the laminin protein significantly promotes cell adhesion and differentiation of C2C12 cells 26,50,51 . Furthermore, we expect that these electrically active, nanotopographic crumpled graphene substrates can electrically stimulate cells.…”

Application of biomimetic surfaces and 3D culture technology to study the role of extracellular matrix interactions in neurite outgrowth and inhibition

“…Previously, we have demonstrated that peptide motifs can be presented to cells, by use of an engineered variant of outer membrane protein A (OmpA) as a scaffold protein for anchoring the motifs in a structurally intact, correctly orientated and highly functional manner as selfassembled monolayers (SAMs) [51,52]. This "ORLASurf®" technology has been used to generate biomimetic surfaces that can be used in cell culture applications, and have previously been shown to promote neural differentiation [53,54] and cellular adhesion [55], depending upon the ECM-derived motif presented to cells.…”