A team of scientists at Washington University in St. Louis, US, has developed a method for creating tissues of the nucleus pulposus of the spine from induced human pluripotent stem cells (hiPSC). A study, published in Stem Cell Research & Therapy, can help in developing methods for treating chronic pain in the neck and back.
Approximately 80% of people experience debilitating pain in the neck and/or back, often accompanied by destruction of the intervertebral discs.
The intervertebral disc is a cartilaginous layer that provides the mobility of the vertebrae, connecting them together and performing a damping function due to the elastic inner gel-like center, the nucleus pulposus, bounded by a fibrous ring. Nucleus pulposus can degenerate with age, causing the discs to lose their shape and collapse – resulting in pain, among other problems.
Scientists have long been trying to find methods of treatment of degeneration of intervertebral discs at an early stage, including, placing high hopes on the use of stem cells to restore the nucleus pulposus. Previous studies have shown that induced human pluripotent stem cells obtained directly from mature cells can express markers of a wide range of cells, including those from which the intervertebral disc is formed.
“What we did here is study developmental biology first before we designed our experiment”,- said Professor Lori Setton. “That was what was different in this particular study, we created a differentiation protocol to mimic embryonic development.”
In past projects, scientists have tried to get different types of adult cells of bone, nerves and fat tissues – directly from hiPSCs. Setton and her group went the other way having developed the technique allowing to create at first a notochord – a cartilaginous structure that is formed at the earliest stages of development, turning further into a spinal column.
“We know that the intervertebral discs arise from the notochord”, – Setton said. “We decided to go back to the beginning and see if we could convert stem cells to be notochordal cells. Only after passing through the notochordal phase did we take them on to the intervertebral disc phase.”
Setton’s lab exposed the hiPSCs to a variety of different growth factors and culture media to coax them into first developing markers for, and then fully forming into, notochord cells. The researchers observed the differentiation process using the fluorescent cell imaging method, checking for the presence of necessary markers at each stage.
“You can think of it as a push-pull”, – Setton said. “You can push it in one direction, but you have to pull it from the other direction as well. I could push it toward a nerve, but I have to pull it from becoming bone. We didn’t know what combination would work. It’s like cooking in the kitchen, and you have to add things to the gravy. It took us a really long time to figure out that perfect recipe. But now that we did, it’s very repeatable.”
Setton says that the multistep process developed in her laboratory to obtain cells of the nucleus pulposus from human iPSС provides for the necessary quality control, as scientists seek additional opportunities for the clinical application of stem cells.
“If you think about a regulated medical product, we need to categorize it at each stage of its use and at each stage of its development”, – she said. “Just as if you’re manufacturing an automotive part, you do quality control at every stage. That’s what we did here.”
In the following stages of the study, the Setton group plans to study the effects of environmental signals (such as the rigidity of the culture surface, the topography of cells and the way they are attached) on the transformation of the human iPSC.