Initial Cell Adhesion onto a Phospholipid Polymer Brush Surface Modified with a Terminal Cell Adhesion Peptide

Abstract

Dynamic changes in the properties of adsorbed protein layers at material surfaces make it difficult to analyze a cell adhesion behavior. Adhesion is affected by the ligand molecules in the adsorbed protein layers on the material's surface. This study aimed to quantitatively analyze the initial cell adhesion onto a polymeric surface modified with immobilized cell adhesion molecules with a well-defined structure. Peptides containing an arginine-glycine-aspartic acid (RGD) sequence were introduced at almost all the termini of the grafted poly(2-methacryloyloxyethyl phosphorylcholine) [poly(MPC)] chains using a click reaction at a highly protein-resistant poly(MPC) brush layer. Thus, the surface could bind to the cell membrane proteins only through the immobilized RGD. Furthermore, the degree of polymerization of the grafted poly(MPC) chains could control the hydrated poly(MPC) brush layer softness, as determined by measuring the dissipation energy loss using a quartz crystal microbalance. At the initial stage of cell adhesion, the density of cells adhering to the RGD-immobilized poly(MPC) brush layers did not depend on the poly(MPC) brush layer softness. However, spreading of the adherent cells was inhibited on the RGD-immobilized poly(MPC) brush layers with a higher softness. Hence, the results suggested that the layer softness did not affect the binding number between the RGD and cell membrane protein during initial cell adhesion; however, the intracellular signaling triggered by the RGD-receptor interaction was inhibited. The poly(MPC) brush surface carrying immobilized cell adhesion molecules has the potential to analyze precisely the effect of the properties of cell adhesion molecules on initial cell adhesion.

Interface of Phospholipid Polymer Grafting Layers to Analyze Functions of Immobilized Oligopeptides Involved in Cell Adhesion

“…The procedure to immobilize the functional peptides at the terminal of the surface PMPC chains is briefly described as follows. , A methanol solution of NaN 3 was added to the polymerization solution to adjust the final concentration to 50 mmol/L, and then stirred at 40 °C overnight. After the substrates were removed from the solution and rinsed with methanol (PMPC-N 3 graft substrate), the substrates were immersed in a mixture of dimethyl sulfoxide (DMSO) and water (80/20 by volume) containing 0.30 mmol/L of each oligopeptide, 1.0 mmol/L Cu­(II)­SO 4 5H 2 O, and 10 mmol/L l -ascorbic acid sodium salt.…”

“…Surface-initiated atom transfer polymerization (SI-ATRP) was carried out using a glass substrate coated with gold. 59 Gold thin films were formed on glass substrates using the high-frequency magnetron sputtering system (VTR-150M/SRF (SCOTT-C3), ULVAC, Inc., Kanagawa, Japan). First, chromium was sputtered at a thickness of 10 nm on the glass substrate (1.2 × 2.4 × 0.2 cm 3 ), and then gold was sputtered at a thickness of 200 nm.…”

“…So far, it has been reported that polymers with a phosphorylcholine group, a sulfobetaine group, and a carboxybetaine group suppress protein adsorption. − Polymers composed of the 2-methacryloyloxyethyl phosphorylcholine (MPC) unit, a common polymer with a phosphorylcholine group, can be shaped into a desired structure relatively easily due to the polymerization mode of MPC being radical polymerization. , In particular, it has been shown that the application of a living radical polymerization method can accurately define the molecular weight of the MPC polymer and the chemical structure of the terminal functional group. , When this polymerization method is used for the initiation reaction from the surface of the substrate, a homogeneous polymer graft layer can be constructed on the surface. − The cell response is governed by the terminal part of the polymer layer, and specific biomolecules are immobilized to control cell functions and responses. Thus, the terminal functional groups of the polymer chains grafted to the surface can also be converted by a chemical reaction so that the biomolecules can be bound. ,− Using this method, a substrate was prepared to observe the state of a biomolecule that specifically acts on cells in detail. The effects of density, movability, and orientation of the cell adhesive peptide on cell adhesion in the early stage were examined.…”

Interfacial nanoarchitectonics for responsive cellular biosystems

“…Nanoarchitectonics essence can be introduced to intercell spaces within cell assemblies to produce artificial cell-cell communication for the cell-to-cell functional relations. In addition, material developments for nanoarchitected platforms for cell adhesion such as self-assembled phosphate-polyamine networks [ 155 ] and biocompatible polymer brushes [ [156] , [157] , [158] ] have been continuously researched. These efforts would bring us much closer to the final goals of nanoarchitectonics, the creation of living creature–like functional systems.…”

E7‐Modified Substrates to Promote Adhesion and Maintain Stemness of Mesenchymal Stem Cells

“…In this regard, massive research reported that high affinity to cells could be achieved by endowing a substrate with adhesive molecules such as fibronectin [ 22 ] and Arg‐Gly‐Asp (RGD). [ 23,24 ] Unfortunately, such biomolecules lack cell specificity due to the widespread receptors on multiple cell types, and even run the risk of inducing differentiation of MSCs. [ 25,26 ] To overcome these limitations, a peptide sequence EPLQLKM (E7) [ 27 ] with high and specific affinity to the rat bone marrow‐derived mesenchymal stem cells (rBMSCs) has been discovered and used.…”