A new model to study fibrosis treatments without using animals

To find treatments for connective tissue disorders like fibrosis, scientists need models that can replicate the structure and function of human tissue when healthy or unhealthy, and respond to drugs as human tissue would. sick. But most models are animal-based and have significant limitations.

A new lab test model developed by researchers at Brown University uses human cells and replicates not only the structure of human tissue, but also its mechanics.

The researchers describe the model in a Advanced sciences study published on Tuesday, February 1.

“This model gives researchers a new tool to not only explore the mechanisms underlying fibrosis and inherited extracellular matrix diseases, but also to test potential treatments for them,” said lead author Jeff Morgan, professor of pathology and laboratory medicine at Brown University, and of engineering.

This development is crucial, Morgan added, because there is no cure for fibrosis and extracellular matrix disorders like Ehlers-Danlos syndrome and Marfan syndrome need new treatments.

Frame the problem

Key to the functionality of the new model is that it does not include an external artificial ‘scaffolding’ for the cells; it uses a novel approach in which cells are harnessed to make their own natural extracellular matrix.

Most tissue engineering approaches rely on the use of protein or polymer scaffolds, explained study co-author Ben Wilks, who earned a Ph.D. in biomedical engineering at Brown and is now a research fellow at Harvard Medical School and Massachusetts General Hospital. Conventional methods involve culturing cells on plastic, while newer approaches embed cells in a collagen hydrogel to mimic the extracellular matrix. This new approach goes much further: it allows cells to synthesize and assemble their own human extracellular matrix.

Over the past few decades, there has been a shift in the scientific understanding of the tissue extracellular matrix. Not only does the matrix provide structural support, but it also communicates with cells through the transmission of mechanical and biochemical signals. This dynamic, bidirectional communication between matrix and cells plays a crucial role in maintaining cellular homeostasis and tissue function, Wilks said.

“We are interested in how changes in nutrients, growth factors, or drug treatments affect cell synthesis and extracellular matrix remodeling and the resulting mechanical properties of tissue constructs,” Wilks said. “Therefore, a scaffold-free approach is much better suited to investigating the questions we ask.”

Researchers in Morgan’s lab at Brown have been studying scaffold-free tissue engineering for more than 15 years. The lab is focused on developing tools that allow scientists to leverage the intrinsic properties of cells to assemble 3D tissues and synthesize their own extracellular matrix, Morgan explained. The lab has developed technology that allows researchers to control the 3D shape of artificial tissue constructs, forming spheres, rings or more complex geometries, by exploiting a phenomenon they call cellular self-assembly.

However, cellular self-assembly seemed to work differently with fibroblasts, a highly contractile cell found throughout the body that plays an important role in wound healing, extracellular matrix synthesis and breakdown, and tissue homeostasis.

The tissue stiffness characteristic of progressive fibrosis, for example, is due to the abnormal behavior of fibroblasts that accumulate and alter the extracellular matrix in a way that ultimately results in loss of organ function.

When the researchers applied technology from the lab to fibroblasts, the tissue constructs broke down spontaneously.

As a Ph.D. A student in Morgan’s lab, Wilks found that altering the composition of the nutrients the cells were grown in would help stabilize the formation of the tissue constructs for days, weeks, or even months.

Additionally, Wilks recognized that by adjusting additional parameters such as mold geometry and cell count, he could form stable 3D ring-shaped tissue constructs, or patterns, that facilitated the tension that a caused the fibroblasts to orient and synthesize their own extracellular matrix. .

“That’s when I really got excited: when I saw how the fibroblasts were aligning and synthesizing this beautiful collagen-rich 3D extracellular matrix in a periodic wave pattern that looks like this you see in native connective tissues like ligaments and tendons,” says Wilks. noted. “I had never seen this before in an engineered fabric construction.”

A showdown

Wilks wondered if it was possible to quantify the stiffness and strength of tissue constructs to allow researchers to replicate normal tissue as well as tissue affected by disease.

Using a tensile testing machine called Instron, the team measured the force needed to stretch the fabric until it broke. This type of data can be used to assess mechanical properties such as tissue strength and stiffness, which can then be related to tissues in the human body. It can also be used to measure how adding a drug would change tissue strength and stiffness.

For example, Wilks said, the data can be used to test whether an anti-fibrotic drug candidate stops the stiffening of tissues that is characteristic of fibrotic diseases.

“In this paper, we develop a 3D connective tissue model that allows us to directly quantify how exposing cells in a 3D environment to different nutrients, growth factors, or drug treatments leads to changes in extracellular matrix synthesis and tissue mechanics, which is an important functional measure of tissue and used clinically to monitor disease progression,” Wilks said. “While there is still a lot of work being done, we believe this model shows promise for screening potential anti-fibrotic drugs. This would address a major unmet need as there is currently no treatment available that can completely stop or reverse fibrosis.”

The new model is one of the most advanced constructs for representing the architecture, composition and 3D mechanics of native connective tissues like ligaments and tendons, the researchers said. Animal models are expensive, ethically controversial and not always predictive of human pathophysiology, said Morgan, who directs the Brown University Center for Alternatives to Animals in Testing.

He added that this type of research is a valuable springboard for creating sophisticated models that can replace and surpass the use of animals.

Video: https://youtu.be/IeZwwAkWk-8

About Hector Hedgepeth

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