Our Parkinson's Place: 3-D neural structure guided with biocompatible nanofiber scaffolds and hydrogels

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Sunday, April 5, 2015

3-D neural structure guided with biocompatible nanofiber scaffolds and hydrogels

Neurons cultivated with the help of ordinary skin cells create a 3-D network on a chip. Credit: (c) Edinson Lucumi Moreno, LCSB

Last updated: Sunday 5 April 2015

Damage to neural tissue is typically permanent and causes lasting disability in patients, but a new approach has recently been discovered that holds incredible potential to reconstruct neural tissue at high resolution in three dimensions. Research recently published in the Journal of Neural Engineering demonstrated a method for embedding scaffolding of patterned nanofibers within three-dimensional (3D) hydrogel structures, and it was shown that neurite outgrowth from neurons in the hydrogel followed the nanofiber scaffolding by tracking directly along the nanofibers, particularly when the nanofibers were coated with a type of cell adhesion molecule called laminin. It was also shown that the coated nanofibers significantly enhanced the length of growing neurites, and that the type of hydrogel could significantly affect the extent to which the neurites tracked the nano fibers.

"Neural stem cells hold incredible potential for restoring damaged cells in the nervous system, and 3D reconstruction of neural tissue is essential for replicating the complex anatomical structure and function of the brain and spinal cord," said Dr. McMurtrey, author of the study and director of the research institute that led this work. "So it was thought that the combination of induced neuronal cells with micropatterned biomaterials might enable unique advantages in 3D cultures, and this research showed that not only can neuronal cells be cultured in 3D conformations, but the direction and pattern of neurite outgrowth can be guided and controlled using relatively simple combinations of structural cues and biochemical signaling factors."

The next step will be replicating more complex structures using a patient's own induced stem cells to reconstruct damaged or diseased sites in the nervous system. These 3D reconstructions can then be used to implant into the damaged areas of neural tissue to help reconstruct specific neuroanatomical structures and integrate with the proper neural circuitry in order to restore function. Successful restoration of function would require training of the new neural circuitry over time, but by selecting the proper neurons and forming them into native architecture, implanted neural stem cells would have a much higher chance of providing successful outcomes. The scaffolding and hydrogel materials are biocompatible and biodegradable, and the hydrogels can also help to maintain the microstructure of implanted cells and prevent them from washing away in the cerebrospinal fluid that surrounds the brain and spinal cord.

McMurtrey also noted that by making these site-specific reconstructions of neural tissue, not only can neural architecture be rebuilt, but researchers can also make models for studying disease mechanisms and developmental processes just by using skin cells that are induced into pluripotent stem cells and into neurons from patients with a variety of diseases and conditions. "The 3D constructs enable a realistic replication of the innate cellular environment and also enable study of diseased human neurons without needing to biopsy neurons from affected patients and without needing to make animal models that can fail to replicate the full array of features seen in humans," said McMurtrey.

The ability to engineer neural tissue from stem cells and biomaterials holds great potential for regenerative medicine. The combination of stem cells, functionalized hydrogel architecture, and patterned and functionalized nanofiber scaffolding enables the formation of unique 3D tissue constructs, and these engineered constructs offer important applications in brain and spinal cord tissue that has been damaged by trauma, stroke, or degeneration. In particular, this work may one day help in the restoration of functional neuroanatomical pathways and structures at sites of spinal cord injury, traumatic brain injury, tumor resection, stroke, or neurodegenerative diseases of Parkinson's, Huntington's, Alzheimer's, or amyotrophic lateral sclerosis.

http://www.medicalnewstoday.com/releases/291845.php?tw

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The progressive loss of neurons in the brain of Parkinson's patients is slow yet inexorable. So far, there are no drugs that can halt this insidious process. Researchers at the Luxembourg Centre for Systems Biomedicine (LCSB) of the University of Luxembourg have now managed to grow the types of neurons affected starting from neuronal stem cells in a three-dimensional cell culture system. The scientists working with Dr. Ronan Fleming of the LCSB research group Systems Biochemistry are confident this system could greatly facilitate the continuing search for therapeutic agents in future as it models the natural conditions in the brain more realistically than other systems available so far. It is also significantly cheaper to employ in the laboratory.

The results were recently published in the journal Lab on a Chip.

Parkinson's disease is characterised in particular by the death of dopamine-producing neurons in the Substantia nigra of the midbrain. It is already possible to grow these dopaminergic neurons in cell cultures. "But most such cell cultures are two-dimensional, with the cells growing along the base of a petri dish, for example," group leader Fleming explains. "Instead, we have the neurons grow in a gel that yields a far better model of their natural, three-dimensional environment."

As the starting point for cultivating the target neurons, the scientists use ordinary skin cells. They convert these through conventional methods into induced pluripotent stem cells, or iPSCs for short. For the development of this technology Japanese scientist Shinya Yamanaka was awarded the Nobel Prize for Physiology or Medicine in 2012 together with John Gurdon. "By adding suitable growth factors, the iPSCs can then be converted in a second step into neural stem cells," says Prof. Jens Schwamborn, head of the LCSB research group Developmental & Cellular Biology, which is responsible for the differentiation of the cells. "These are the starting cells we use in the microfluidic culture."

The researchers first mix the cells with a liquid, which they then fill into little test vessels called bioreactors. "You can imagine such a bioreactor as a tunnel separated down the middle by a flat barrier," LCSB researcher Edinson Lucumi Moreno, first author of the study, explains. "One side of the tunnel we load the liquid with the cells, where it hardens into a gel under controlled temperatures. The other side we load with a medium to which we can add nutrients and substances for further differentiation of the neuronal stem cells as required."

After only a few hours, the researchers already observe changes in the neuronal stem cells: The cells begin to form little protrusions, which develop over the following days into axons and dendrites - the long extensions typical of neurons. After 30 days, 91 percent of the cells are neurons, about 20 percent of which are the desired dopaminergic neurons. This has been confirmed in morphological and immunological tests.

One of the major advantages of this 3D cell culture system is that it can already be automated in its present form: The bioreactors are placed on commercially available plates that can be processed and read out by laboratory robots. "In drug development, dozens of chemical substances can therefore be tested for possible therapeutic effects in a single step," says Ronan Fleming. "Because we use far smaller amounts of substances than in conventional cell culture systems, the costs drop to about one tenth the usual."

A further advantage is that the bioreactors can be loaded with cells originating from the skin cells of individual Parkinson's patients. "This is an important step towards personalised drug development," Fleming asserts. As a next step, Fleming's team and their international collaborators want to study cells from patients and to test potential active pharmaceutical ingredients. Promising substances will then be tested in mice.