Artificial cell membrane binding thrombin constructs drive in situ fibrin hydrogel formation

Abstract

Cell membrane re-engineering is emerging as a powerful tool for the development of next generation cell therapies, as it allows the user to augment therapeutic cells to provide additional functionalities, such as homing, adhesion or hypoxia resistance. To date, however, there are few examples where the plasma membrane is re-engineered to display active enzymes that promote extracellular matrix protein assembly. Here, we report on a self-contained matrix-forming system where the membrane of human mesenchymal stem cells is modified to display a novel thrombin construct, giving rise to spontaneous fibrin hydrogel nucleation and growth at near human plasma concentrations of fibrinogen. The cell membrane modification process is realised through the synthesis of a membrane-binding supercationic thrombin-polymer surfactant complex. Significantly, the resulting robust cellular fibrin hydrogel constructs can be differentiated down osteogenic and adipogenic lineages, giving rise to self-supporting monoliths that exhibit Young’s moduli that reflect their respective extracellular matrix compositions.

Proteolytic Rafts for Improving Intraparenchymal Migration of Minimally Invasively Administered Hydrogel-Embedded Stem Cells

“…At present, there are several techniques that use immobilization of relatively expensive enzymes in solid phase of carriers to operate in areas of biotransformation, food processing, medical diagnostics, and pharmacy [15]. In the field of cellular therapies, Deller et al modified cellular membrane of MSCs by thrombin immobilization to improve cell adhesion, homing, and resilience to hypoxia [17]. Functionalization of biomaterials opens a new possibility to improve cell transplantation.…”

Tailoring Gelation Mechanisms for Advanced Hydrogel Applications

“…[243] A similar approach was used to form DNA-polyelectrolyte hydrogels on the surface of various mammalian cell lines, including smooth muscle cells, hematopoietic cells, and human mesenchymal stem cells. Cholesterol was used to anchor a single DNA strand into cell membranes, where it was able to hybridize free DNA hairpins to create an alginate-DNA polymer network hydrogel that could be further [241] Reproduced under the terms of the Creative Commons CC-BY-NC license. [241] Copyright 2019, The Authors, published by Springer Nature Limited.…”

“…Cholesterol was used to anchor a single DNA strand into cell membranes, where it was able to hybridize free DNA hairpins to create an alginate-DNA polymer network hydrogel that could be further [241] Reproduced under the terms of the Creative Commons CC-BY-NC license. [241] Copyright 2019, The Authors, published by Springer Nature Limited. B) Schematic of the strategy taken by Shi et al to form an alginate-DNA polymer network hydrogel that could be further crosslinked with polylysine around the surface of a cell.…”

“…The encapsulated cells were shown to proliferate for several weeks and could undergo both adipogenic and osteogenic differentiation. [ 241 ] In another work, Sakai et al used HRP conjugated to a biocompatible anchor molecule, which could be inserted into the membrane of various cell lines, such as mouse embryonic fibroblasts. In the presence of millimolar concentrations of hydrogen peroxide, the HRP enzyme was able to catalyze the oxidative coupling of phenols to form polyphenolic networks.…”

Fibrin Matrices as (Injectable) Biomaterials: Formation, Clinical Use, and Molecular Engineering

“…Finally, it is worth mentioning a recent approach by the group of Perriman, who instead of dispersing cells within a gelling fibrinogen solution, have nucleated fibrin around them: using a complex between a heavily cationized thrombin and a PEGylated surfactant that accumulates on cell surfaces, fibrin was produced directly on the surface of mesenchymal stem cells. With this method, 3D constructs can be produced where cells are individually encapsulated, and then driven toward various differentiation paths through an appropriate choice of the culture media …”