Mimicking the neural stem cell niche: An engineer’s view of cell: material interactions

Neural stem cells have attracted attention in recent years to treat neurodegeneration. There are two neurogenic regions in the brain where neural stem cells reside, one of which is called the subventricular zone (SVZ). The SVZ niche is a complicated microenvironment providing cues to regulate self-renewal and differentiation while maintaining the neural stem cell’s pool. Many scientists have spent years understanding the cellular and structural characteristics of the SVZ niche, both in homeostasis and pathological conditions. On the other hand, engineers focus primarily on designing platforms using the knowledge they acquire to understand the effect of individual factors on neural stem cell fate decisions. This review provides a general overview of what we know about the components of the SVZ niche, including the residing cells, extracellular matrix (ECM), growth factors, their interactions, and SVZ niche changes during aging and neurodegenerative diseases. Furthermore, an overview will be given on the biomaterials used to mimic neurogenic niche microenvironments and the design considerations applied to add bioactivity while meeting the structural requirements. Finally, it will discuss the potential gaps in mimicking the microenvironment

Investigating Alternative Measures of Functional Recovery in Rat Sciatic Nerve Injury

There is a pressing need for advancements in peripheral nerve repair techniques and functional recovery evaluation methods. The rat sciatic nerve injury model is a well examined model for peripheral nerve repair. One measure of functional recovery after nerve damage, the sciatic functional index (SFI), fails in the presence of self-mutilation, toe contracture, and other abnormalities in gait. In this IACUC approved study, the sciatic nerve was severed in four experimental groups (n=5). The nerves were repaired with Arginylglycylaspartic acid-poly(ε-caprolactone) (RGD-PCL) peptide functionalized nanofibers, non-functionalized PCL control nanofibers, an isograft, and a negative control empty conduit. Video walking track analysis allowed for a retrospective analysis with three other evaluation techniques: imbalance coupling (IC), stance factor (SF), and toe out angle (TOA). While these techniques are independent of self-mutilation and toe contracture, walking speed remained as a confounding variable. One way repeated measures ANOVA tests showed no significant difference between treatments or subjects in SFI, SF, or TOA. For SFI, 6 and 12 week trials both saw significant increases over time (p=0.00 for both). A significant difference was found between treatments in IC (p=0.03). Imbalance coupling showed promising Pearson correlation with the current industry standard, SFI (p=0.03). In a regression model, SFI over time had an R-squared value of 94.5%. IC, SF, and TOA had low R-squared values. Future investigation with updated protocol is necessary to confirm the degree of correlation and to evaluate the potential for a new industry standard for evaluating nerve repair

Developing AFM Techniques for Testing PEG Hydrogels

Many instruments are used to find elastic properties of biological samples using methods such as tensile and bending tests, but using the atomic force microscope (AFM) is considered a non-destructive method because it can provide repeated local stiffness information without damaging the sample. It additionally allows the sample to be tested in an aqueous environment, which is optimal for soft materials such as hydrogels. The nanoindentation is performed via cantilever, measuring the deflection of the cantilever during the contact of the sample using a laser. Compared to hard samples, testing soft materials can present more challenges when working with the AFM, creating the need for a refined technique.[1] This study will explore ways to improve the accuracy and feasibility of testing hydrogels, which are significant in biomaterials research as they offer the ability to be altered mechanically and chemically to fit the needs of cells.[2] The technique for testing the hydrogels will be refined through a process moving from dry to wet samples, attempting to repeatedly and successfully obtain topography and elastic properties through high resolution topography scans and force curves. References: Radmacher, M., Tillamnn, R.W., Fritz, M., and Gaub, H.E. (1992), From molecules to cells: imaging soft samples with the atomic force microscope. Science. 257: 1900-1905 Flake, M. M., Nguyen, P. K., Scott, R. A., Vandiver, L. R., Willits, R. K., and Elbert, D. L. Poly(ethylene glycol) microparticles produced by precipitation polymerization in aqueous solution. Biomacromolecules 12 (844-850). 2011

Inherent Interfacial Mechanical Gradients in 3D Hydrogels Influence Tumor Cell Behaviors

Cells sense and respond to the rigidity of their microenvironment by altering their morphology and migration behavior. To examine this response, hydrogels with a range of moduli or mechanical gradients have been developed. Here, we show that edge effects inherent in hydrogels supported on rigid substrates also influence cell behavior. A Matrigel hydrogel was supported on a rigid glass substrate, an interface which computational techniques revealed to yield relative stiffening close to the rigid substrate support. To explore the influence of these gradients in 3D, hydrogels of varying Matrigel content were synthesized and the morphology, spreading, actin organization, and migration of glioblastoma multiforme (GBM) tumor cells were examined at the lowest (<50 µm) and highest (>500 µm) gel positions. GBMs adopted bipolar morphologies, displayed actin stress fiber formation, and evidenced fast, mesenchymal migration close to the substrate, whereas away from the interface, they adopted more rounded or ellipsoid morphologies, displayed poor actin architecture, and evidenced slow migration with some amoeboid characteristics. Mechanical gradients produced via edge effects could be observed with other hydrogels and substrates and permit observation of responses to multiple mechanical environments in a single hydrogel. Thus, hydrogel-support edge effects could be used to explore mechanosensitivity in a single 3D hydrogel system and should be considered in 3D hydrogel cell culture systems

Polyethlyene Glycol Microgels to Deliver Bioactive Nerve Growth Factor

Delivery of bioactive molecules is a critical step in fabricating materials for regenerative medicine, yet, this step is particularly challenging in hydrated scaffolds such as hydrogels. Although bulk photocrosslinked poly(ethylene glycol) (PEG) hydrogels have been used for a variety of tissue engineering applications, their capability as drug delivery scaffolds has been limited due to undesirable release profiles and reduction in bioactivity of molecules. To solve these problems, this article presents the fabrication of degradable PEG microgels, which are micron-sized spherical hydrogels, to deliver bioactive nerve growth factor (NGF). NGF release and activity was measured after encapsulation in microgels formed from either 3 kDa or 6 kDa PEG to determine the role of hydrogel mesh size on release. Microgels formed from 6 kDa PEG were statistically larger and had a higher swelling ratio than 3 kDa PEG. The 6 kDa PEG microgels provided a Fickian release with a reduced burst effect and 3 kDa microgels provided anomalous release over ≥20 days. Regardless of molecular weight of PEG, NGF bioactivity was not significantly reduced compared to unprocessed NGF. These results demonstrate that microgels provide easy mechanisms to control the release while retaining the activity of growth factors. As this microgel-based delivery system can be injected at the site of nerve injury to promote nerve repair, the potential to deliver active growth factors in a controlled manner may reduce healing time for neural tissue engineering applications

Effects of a mechanical response-contingent surrogate on the development of behaviors in nursery-reared rhesus macaques (Macaca mulatta).

Nursery-reared infants have several behavioral and physiologic differences from their mother-reared counterparts. We investigated whether a response-contingent surrogate mitigated some of those differences by decreasing fearfulness and partner-clinging and increasing environmental exploration in nursery-reared infants continuously paired with a peer. Six nursery-reared infant rhesus macaques (in pairs) were given a mechanical responsive surrogate (RS), and 6 (in pairs) were given an identical but nonresponsive surrogate (NRS). The 2 treatment groups were compared and then combined into a single group of all 12 of surrogate-exposed animals (CS) that was compared with a nonsurrogate control group (NS) of 10 nursery-reared infants. Results showed significant differences between CS and NS infants but no significant differences between the RS and NRS infants. As compared with NS infants, CS infants showed less partner-clinging, less affiliation directed toward only partner, and more foraging and tactile-oral exploration of the environment. These advantageous effects support additional research to develop improved surrogate and the implementation of surrogate programs for nursery-reared infants

Dorsal root ganglia neurite outgrowth measured as a function of changes in microelectrode array resistance.

Current research in prosthetic device design aims to mimic natural movements using a feedback system that connects to the patient's own nerves to control the device. The first step in using neurons to control motion is to make and maintain contact between neurons and the feedback sensors. Therefore, the goal of this project was to determine if changes in electrode resistance could be detected when a neuron extended a neurite to contact a sensor. Dorsal root ganglia (DRG) were harvested from chick embryos and cultured on a collagen-coated carbon nanotube microelectrode array for two days. The DRG were seeded along one side of the array so the processes extended across the array, contacting about half of the electrodes. Electrode resistance was measured both prior to culture and after the two day culture period. Phase contrast images of the microelectrode array were taken after two days to visually determine which electrodes were in contact with one or more DRG neurite or tissue. Electrodes in contact with DRG neurites had an average change in resistance of 0.15 MΩ compared with the electrodes without DRG neurites. Using this method, we determined that resistance values can be used as a criterion for identifying electrodes in contact with a DRG neurite. These data are the foundation for future development of an autonomous feedback resistance measurement system to continuously monitor DRG neurite outgrowth at specific spatial locations