Hydrogel Modulus Affects Proliferation Rate and Pluripotency of Human Mesenchymal Stem Cells Grown in Three-Dimensional Culture

Abstract

Hydrogels have been used extensively with various cell types in three-dimensional (3D) culture, including with human mesenchymal stem cells (hMSCs). Here, we report on the use of poly­(ethylene glycol) (PEG)-conjugated fibrinogen hydrogels to grow bone marrow-derived hMSCs in 3D culture. The initial modulus of the PEG-fibrinogen (PF) hydrogels was varied to study the influence of the stiffness on the proliferation response of the cells growing within. Shear rheology was used to quantify the changes to the initial material properties; the shear storage modulus of the PF was controlled by changing the concentration of synthetic PEG cross-linker, while keeping the fibrinogen concentration constant. Cell culture was performed during a 14-day experiment to quantify the cell proliferation response in the different modulus materials tested. The hMSCs were recovered from the hydrogels by mild enzymatic dissolution, and characterized for proliferation and cell number using cytometry. The results indicate a modulus-dependent response from the cells, and the ability to preferentially define initial hydrogel modulus that favors higher proliferation and multipotency of the hMSCs. Bioactive supplements added to the hydrogels, including exogenous fibronectin (Fn) and von willebrand factor (vWf) were used to further stimulate the proliferation response of the hMSCs in the hydrogel cultures, without altering their multipotency. These insights underscore the importance of mechanical properties in regulating cell proliferation in a 3D culture milieu. The versatility of the hydrogel culture environment and the ability to control mechanical properties for cell-fate determination further highlight the significance of a modular approach when developing materials that ultimately optimize stem cell cultures.

The cell in the ink: Improving biofabrication by printing stem cells for skeletal regenerative medicine

“…Crucially, even before printing, the hydrogel moduli affect the cell viability when encapsulated. For example, low moduli hydrogels (<1 kPa) can often provide a better environment for cells to attach, expand and proliferate [ 72 , 73 ]. Physically stronger gels can be loaded with cells with the help of mechanical methods (mixing, centrifugation or vortexing) ensuring high printing fidelity, nevertheless cell encapsulation still remains a crucial and unresolved issue.…”

Role of Nanoparticle–Polymer Interactions on the Development of Double-Network Hydrogel Nanocomposites with High Mechanical Strength

“…Researchers have also taken advantage of the ability of hydrogels to swell or shrink in response to external stimuli (e.g., pH, temperature) to develop biosensors for the detection of biomolecules [4,5]. Additionally, their highly porous and hydrated polymer structure mimics the extracellular cellular matrix and renders them highly suitable for in vitro cell culture, and several studies have demonstrated the successful use of hydrogels to encapsulate mammalian cells in a 3D physiological-like environment and develop in vitro models of cell proliferation, migration, and differentiation [6][7][8][9][10]. However, hydrogels have poor mechanical strength, which limits their broad applicability for tissue engineering [11].…”

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

“…Due to the benign character of photopolymerization, living cells can be easily encapsulated and imaged in these materials, that can also be fashioned in the form of microparticles or spheroid/microspheres . These PEGylated fibrinogen gels have been employed in a variety of in vitro models, for example, including pluripotency maintenance (human mesenchymal stem cells) or neural growth neural outgrowth, and have been applied in vivo for BMP‐2 stimulated bone repair, for the generation of skeletal muscles from encapsulated mesangioblasts, for the regeneration of sciatic nerve, or for myocardial regeneration (no cell encapsulated) after infarction …”