From skin biopsy to neurons, UCLA scientists use animal origin-free reagents to create cells that may prove clinically usable

Using a specially designed facility, UCLA stem cell scientists have taken human skin cells, reprogrammed them to be pluripotent and then differentiated them into neurons, using animal origin-free reagents and feeder conditions throughout the process.

This is the first time scientists have been able to derive potentially clinically usable induced pluripotent stem (iPS) cells and differentiate them into neurons in animal origin-free derivation and differentiation conditions using commercially available reagents to facilitate broad application, said Saravanan Karumbayaram, study first author and an associate researcher with the Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA.

The Broad center researchers also developed a set of standard operating procedures for the process, so other scientists can benefit from the derivation and differentiation techniques performed under Good Manufacturing Practices (GMP) protocols. GMP protocols are tightly controlled and regulated so the cells created meet all the standards required for use in human beings.

“Developments in stem cell research show that pluripotent stem cells ultimately will be translated into therapies, so we are working to develop the methods and systems needed to make the cells safe for human use,” Karumbayaram said.

The study appears Dec. 7, 2011 in the early online edition of the inaugural issue of the peer-reviewed journal Stem Cells Translational Medicine, a new journal that seeks to bridge stem cell research and clinical trials.

Conventionally, stem cells are grown on mouse fibroblast cells, which provide factors the cells need to flourish and grow. However, because animal products are involved, those cells have to be further tested for any contaminating animal-derived products before they can be used in humans.           

Karumbayaram tried six different animal-free media formulations before arriving at a composition that generated the most robust pluripotent stem cells. He combined two commercial media solutions to create his own mix and tried different concentrations of an important growth factor.

“The colonies we get are of very good quality and are quite stable,” said Karumbayaram, who compared his animal-free colonies to those created conventionally using mouse feeder cells.

Efficiency did suffer. Fewer colonies were created using the animal-free feeders, but the colonies remained stable for at least 20 passages.

The resulting neurons started life as a small skin punch biopsy from a volunteer. Those skin cells were then reprogrammed to become pluripotent stem cells with the ability to make any cell in the human body. The iPS cells were then grown in colonies and later coaxed into becoming neural precursor cells and finally, neurons.

The animal-free cells were compared at every step in the process to cells produced by typical methods, Karumbayaram said, and were found to be of very similar quality.        

“We were very excited when we saw the first colonies growing, because we were not sure it would be possible to derive and grow cells completely animal-free,” he said.

Because the cells were grown in a special facility designed to culture animal-free cells, the testing and examination required to make clinical-grade cells should be much simpler, said William Lowry, study senior author and an assistant professor of molecular, cell and developmental biology in Life Sciences.

To date, at least15 animal-free iPS cell lines have been created at the Broad Stem Cell Research Center.

“It’s critical to note that we are nowhere near ready to use these cells in the clinic,” Lowry said. “We are working to develop methods to make sure these cells are genetically stable and will be as safe as possible for human use. The main goal of this project was to generate a platform that will one day allow clinical translation of stem cells to the clinic.”

The study was supported by the California Institute of Regenerative Medicine, the National Institutes of Health, the Department of Defense, the Foundation to Eradicate Duchenne, the UCLA Muscular Dystrophy Core Center and the Broad Stem Cell Research Center.