Protocol Raises Hopes for Treating Macular Degeneration with Stem Cells

Researchers in Switzerland have developed a protocol for differentiating human embryonic stem cells into retinal pigment epithelium cells that could be used to treat age-related macular degeneration.

The disease affects the macula, which is part of the eye’s retina and provides sharp vision and the ability to distinguish details. One of the most promising treatment strategies for age-related macular degeneration is to replace aging, degenerating retinal pigment epithelium (RPE) cells—which nourish and maintain the eye’s photoreceptor cells—with new ones grown from human embryonic stem cells.

 Scientist have proposed several methods for converting stem cells into RPE, but there is still a gap in knowledge of how cells respond to these stimuli over time. Some protocols take a few months while others can take up to a year.

“None of the differentiation protocols proposed for clinical trials have been scrutinized over time at the single-cell level—we know they can make retinal pigment cells, but how cells evolve to that state remains a mystery,” says Dr. Gioele La Manno, a researcher with Ecole Polytechnique Fédérale de Lausanne’s Life Sciences Independent Research (ELISIR) program. “Overall, the field has been so focused on the product of differentiation, that the path undertaken has been sometimes overlooked. For the field to move forward, it is important to understand aspects of the dynamics of what happens in these protocols.”  The path to maturity, he adds, could be as important as the end state for the safety of treatment or for improving cell purity and reducing production time.

Toward this end, La Manno led a study with Professor Fredrik Lanner at the Karolinska Institute in Sweden that profiles a protocol for differentiating human embryonic stem cells into RPE cells intended for clinical use. Their work, published recently in the journal Stem Cell Reports, shows that the protocol can develop safe and efficient pluripotent stem cell-based therapies for age-related macular degeneration.

“Standard methods such as quantitative PCR and bulk RNA-seq capture the average expression of RNAs from large populations of cells,” says Alex Lederer, a doctoral student at EPFL and one of the study’s lead authors. “In mixed-cell populations, these measurements may obscure critical differences between individual cells that are important for knowing if the process is unfolding correctly.”

Instead, the researchers used a technique called single-cell RNA sequencing (scRNA-seq), which can detect all the active genes in an individual cell at a given time. They were able to study the entire gene expression profile of individual human embryonic stem cells throughout the differentiation protocol, which takes a total of sixty days. This allowed them to map out all the transient states within a population as they grew into retinal pigment cells, but also to optimize the protocol and suppress the growth of non-RPE cells, thus preventing the formation of contaminant cell populations. “The aim is to prevent mixed cell populations at the time of transplantation, and to make sure the cells at the endpoint are similar to original RPE cells from a patient’s eye,” says Lederer.

They found that, that on the way to becoming RPE cells, stem cells go through a process very similar to early embryonic development. During this, the cell culture took up a “rostral embryo patterning,” the process that develops the embryo’s neural tube, which will go on to become its brain and sensory systems for vision, hearing, and taste. After this patterning, the stem cells began to mature into RPE cells.

But the point of the differentiation protocol is to generate a pure population of RPE cells that can be implanted in patients’ retinas to slow down macular degeneration. So the team transplanted their population of cells that had been monitored with scRNA-seq into the subretinal space of two female New Zealand white albino rabbits. The work showed that the protocol not only produces a pure RPE cell population but that those cells can continue maturing even after they have been transplanted in the subretinal space.