In Vitro Simulated Neuronal Environmental Conditions Qualify Umbilical Cord Derived Highly Potent Stem Cells for Neuronal Differentiation.

The healing of neuronal injuries is still an unachieved goal. Medicine-based therapies can only extend the survival of patients, but not finally lead to a healing process. Currently, a variety of stem cell-based tissue engineering developments are the subject of many research projects to bridge this gap. As yet, neuronal differentiation of induced pluripotent stem cells (iPS), embryonic cell lines, or neuronal stem cells could be accomplished and produce functional neuronally differentiated cells. However, clinical application of cells from these sources is hampered by ethical considerations. To overcome these hurdles numerous studies investigated the potential of adult mesenchymal stem cells (MSCs) as a potential stem cell source. Adult MSCs have been approved as cellular therapeutical products due to their regenerative potential and immunomodulatory properties. Only a few of these studies could demonstrate the capacity to differentiate MSCs into active firing neuron like cells. With this study we investigated the potential of Wharton's Jelly (WJ) derived stem cells and focused on the intrinsic pluripotent stem cell pool and their potential to differentiate into active neurons. With a comprehensive neuronal differentiation protocol comprised of mechanical and biochemical inductive cues, we investigated the capacity of spontaneously forming stem cell spheroids (SCS) from cultured WJ stromal cells in regard to their neuronal differentiation potential and compared them to undifferentiated spheroids or adherent MSCs. Spontaneously formed SCSs show pluripotent and neuroectodermal lineage markers, meeting the pre-condition for neuronal differentiation and contain a higher amount of cells which can be differentiated into cells whose functional phenotypes in calcium and voltage responsive electrical activity are similar to neurons. In conclusion we show that up-concentration of stem cells from WJ with pluripotent characteristics is a tool to generate neuronal cell replacement.

Improvements of Microcontact Printing for Micropatterned Cell Growth by Contrast Enhancement.

Patterned neuronal cell cultures are important tools for investigating neuronal signal integration, network function, and cell-substrate interactions. Because of the variable nature of neuronal cells, the widely used coating method of microcontact printing is in constant need of improvements and adaptations depending on the pattern, cell type, and coating solutions available for a certain experimental system. In this work, we report on three approaches to modify microcontact printing on borosilicate glass surfaces, which we evaluate with contact angle measurements and by determining the quality of patterned neuronal growth. Although background toxification with manganese salt does not result in the desired pattern enhancement, a simple heat treatment of the glass substrates leads to improved background hydrophobicity and therefore neuronal patterning. Thirdly, we extended a microcontact printing process based on covalently linking the glass surface and the coating molecule via an epoxysilane. This extension is an additional hydrophobization step with dodecylamine. We demonstrate that shelf life of the silanized glass is at least 22 weeks, leading to consistently reliable neuronal patterning by microcontact printing. Thus, we compared three practical additions to microcontact printing, two of which can easily be implemented into a workflow for the investigation of patterned neuronal networks.