Use of weak DC electric fields to rapidly align mammalian cells.

Objective.The ability to modulate cell morphology has clinical relevance in regenerative biology. For example, cells of the skeletal muscle, peripheral nerve and vasculature have specific oriented architectures that emerge from unique structure-function relationships. Methods that can induce similar cell morphologiesin vitrocan be of use in the development of biomimetic constructs for the repair or replacement of damaged tissues. In this work, we demonstrate that direct current (DC) electric fields (EFs) can be used as a tool to globally align cell populationsin vitro. Approach.Using a 2D culture chamber system, we were able to quickly (within hours) align Schwann cells at different culture densities with an application of steady EFs at 200-500 mV mm-1.Main results.Cellular alignment was perpendicular to the field vector and varied proportionately as a function of field magnitude. In addition, the degree of cellular alignment was also dependent on cellular density. Even well-established Schwann cell monolayers were responsive to the applied DC fields with cells retracting parallel oriented processes (with respect to the imposed field) and re-extending them along the perpendicular axis. When the DC field was removed, monolayers retained the aligned morphology for many days afterwards, likely due to contact inhibition. We further show the method is applicable to other field-responsive cells, such as 3T3 fibroblasts.Significance.The patterned cells provided nanoscale haptotactic cues and can be subsequently used as a basal layer for co-culturing or manipulated for other applications. DC fields represent a rapid, simple, and efficient technique compared to other cell patterning methods such as substrate manipulation.

DC Electric Fields Induce Perpendicular Alignment and Enhanced Migration in Schwann Cell Cultures.

Schwann cells (SCs) are PNS glia that play numerous support functions including myelination of axons. After PNS injury, SCs facilitate regeneration by phagocytosing cellular debris and providing physical and biochemical cues to guide axon growth. This reparative phenotype suggests SCs could be critical cellular targets for enhancing nerve regeneration. One method for altering cell morphology and motility is the application of direct current (DC) electric fields (EFs). Endogenous EFs have physiologic relevance during embryogenesis and serve as guidance and polarization cues. While much literature exists on EFs and CNS and PNS neurons, the effects of EFs on SCs have not been extensively studied. In this work, cell alignment, migration, and morphology of rat SCs were measured in response to several EF stimulation regimes including constant DC, 50% duty cycle DC and oscillating DC. SCs were found to re-orient perpendicular to field lines and respond to DC EFs as low as 75 mV/mm. EF exposure promoted directed migration, with travel towards the cathode at a mean rate of 7.5 µm/h. The data highlight the utility of EFs in modulating SC morphology, alignment and migration. Results may have implications for using EFs to attract and realign SCs at the site of PNS trauma.

Effect of Molecular Weight and Functionality on Acrylated Poly(caprolactone) for Stereolithography and Biomedical Applications.

Degradable polymers are integral components in many biomedical polymer applications. The ability of these materials to decompose in situ has become a critical component for tissue engineering, allowing scaffolds to guide cell and tissue growth while facilitating gradual regeneration of native tissue. The objective of this work is to understand the role of prepolymer molecular weight and functionality of photocurable poly(caprolactone) (PCL) in determining reaction kinetics, mechanical properties, polymer degradation, biocompatibility, and suitability for stereolithography. PCL, a degradable polymer used in a number of biomedical applications, was functionalized with acrylate groups to enable photopolymerization and three-dimensional printing via stereolithography. PCL prepolymers with different molecular weights and functionalities were studied to understand the role of molecular structure in reaction kinetics, mechanical properties, and degradation rates. The mechanical properties of photocured PCL were dependent on cross-link density and directly related to the molecular weight and functionality of the prepolymers. High-molecular weight, low-functionality PCLDA prepolymers exhibited a lower modulus and a higher strain at break, while low-molecular weight, high-functionality PCLTA prepolymers exhibited a lower strain at break and a higher modulus. Additionally, degradation profiles of cross-linked PCL followed a similar trend, with low cross-link density leading to degradation times up to 2.5 times shorter than those of more highly cross-linked polymers. Furthermore, photopolymerized PCL showed biocompatibility both in vitro and in vivo, causing no observed detrimental effects on seeded murine-induced pluripotent stem cells or when implanted into pig retinas. Finally, the ability to create three-dimensional PCL structures is shown by fabrication of simple structures using digital light projection stereolithography. Low-molecular weight, high-functionality PCLTA prepolymers printed objects with feature sizes near the hardware resolution limit of 50 µm. This work lays the foundation for future work in fabricating microscale PCL structures for a wide range of tissue regeneration applications.