Cationic Hydrogels Modulate Neural Stem and Progenitor Cell Proliferation and Differentiation Behavior in Dependence of Cationic Moiety Concentration in 2D Cell Culture.

The central nervous system (CNS) has a limited regenerative capacity because a hostile environment prevents tissue regeneration after damage or injury. Neural stem/progenitor cells (NSPCs) are considered a potential resource for CNS repair, which raises the issue of adequate cultivation and expansion procedures. Cationic charge supports the survival and adhesion of NSPCs. Typically, tissue culture plates with cationic coatings, such as poly-l-ornithine (PLO), have been used to culture these cell types (NSPCs). Yet presently, little is known about the impact of cationic charge concentration on the viability, proliferation, and differentiation capacity of NSPCs. Therefore, we have recently developed well-defined, fully synthetic hydrogel systems G1 (gel 1) to G6 (gel 6) that allow for the precise control of the concentration of the cationic trimethylaminoethyl acrylate (TMAEA) molecule associated with the polymer in a range from 0.06 to 0.91 µmol/mg. When murine NSPCs were cultured on these gels under differentiation conditions, we observed a strong correlation of cationic charge concentration with NSPC survival. In particular, neurons were preferentially formed on gels with a higher cationic charge concentration, whereas astrocytes and oligodendrocytes favored weakly charged or even neutral gel surfaces. To test the properties of the gels under proliferative conditions, the NSPCs were cultivated in the presence of fibroblast growth factor 2 (FGF2). The cytokine significantly increased the number of NSPCs but delayed the differentiation toward neurons and glia cells. Thus, the hydrogels are compatible with the survival, expansion, and differentiation of NSPCs and may be useful to create supportive environments in transplantation approaches.

Concentration Dependent Effect of Quaternary Amines on the Adhesion of U251-MG Cells.

Cationic gels have seen increasing interest in recent years for 2D cell cultivation since they may represent an alternative to the well-known RGD-peptide motif functionalized gels. However, few hydrogel systems with adjustable cationic strength have been fabricated and investigated so far. In this work, eight gels with defined concentrations of cationic groups, two of which also contained the RGD peptide, were prepared from three well-defined, soluble precursor copolymers with thiol-functionalities and PEGDA3500 as a crosslinker via thiol-ene chemistry. Live/dead stainings of U-251-MG cells on the hydrogels with different concentrations of the cationic motif were made after 3 days and 7 days of cultivation. The results show a high dependence of the number of adhesive cells and their morphology, cluster versus spread cells, on the concentration of cationic groups in the gel. This effect was more pronounced when the gels were not further dialyzed before usage. In addition, a synergistic effect of the two motifs, cationic group and RGD peptide, could be demonstrated, which together induce stronger cell adhesion than either motif alone.

Synthesis and Characterization of Cationic Hydrogels from Thiolated Copolymers for Independent Manipulation of Mechanical and Chemical Properties of Cell Substrates.

Cells sense both mechanical and chemical properties in their environment and respond to these inputs with altered phenotypes. Precise and selective experimental manipulations of these environmental cues require biocompatible synthetic materials, for which multiple properties can be fine-tuned independently from each other. For example, cells typically show critical thresholds for cell adhesion as a function of substrate parameters such as stiffness and the degree of functionalization. However, the choice of tailor-made, defined materials to produce such cell adhesion substrates is still very limited. Here, a platform of synthetic hydrogels based on well-defined thiolated copolymers is presented. Therefore, four disulfide crosslinked hydrogels of different composition by free radical polymerization are prepared. After cleavage with dithiothreitol, four soluble copolymers P1-P4 with 0-96% cationic monomer content are obtained. P1 and P4 are then combined with PEGDA3500 as a crosslinker, to fabricate 12 hydrogels with variable elasticity, ranging from 8.1 to 26.3 kPa and cationic group concentrations of up to 350 µmol cm-3 . Systematic analysis using COS7 cells shows that all of these hydrogels are nontoxic. However, successful cell adhesion requires both a minimal elasticity and a minimal cationic group concentration.

Hydrogels Derivatized With Cationic Moieties or Functional Peptides as Efficient Supports for Neural Stem Cells.

The increasing incidence of neurodegenerative diseases such as Alzheimer's or Parkinson's disease represents a significant burden for patients and national health systems. The conditions are primarily caused by the death of neurons and other neural cell types. One important aim of current stem cell research is to find a way to replace the lost cells. In this perspective, neural stem cells (NSCs) have been considered as a promising tool in the field of regenerative medicine. The behavior of NSCs is modulated by environmental influences, for example hormones, growth factors, cytokines, and extracellular matrix molecules or biomechanics. These factors can be studied by using well-defined hydrogels, which are polymeric networks of synthetic or natural origin with the ability to swell in water. These gels can be modified with a variety of molecules and optimized with regard to their mechanical properties to mimic the natural extracellular environment. In particular modifications applying distinct units such as functional domains and peptides can modulate the development of NSCs with regard to proliferation, differentiation and migration. One well-known peptide sequence that affects the behavior of NSCs is the integrin recognition sequence RGD that has originally been derived from fibronectin. In the present review we provide an overview concerning the applications of modified hydrogels with an emphasis on synthetic hydrogels based on poly(acrylamides), as modified with either cationic moieties or the peptide sequence RGD. This knowledge might be used in tissue engineering and regenerative medicine for the therapy of spinal cord injuries, neurodegenerative diseases and traumata.