The black rat rested calmly in a clear tube, its nose twitching gently as it sniffed the air. Its eyes followed the parade of vertical black and white bars scrolling from left to right across a screen. For a normal rat, this would not be especially impressive; rats — and humans, too — will involuntarily follow these moving lines with their eyes. 

It is, however, quite an impressive feat for this particular rat. This rat, called a Royal College of Surgeons rat, or RCS rat, was born with a genetic mutation that leads to the degeneration of photoreceptors in the retina (1). This mutation should have rendered it completely blind by the time it was 90 days old. Luckily for this rat, it ended up in the laboratory of Magdalene Seiler, an ophthalmology researcher at the University of California, Irvine, who partially restored its vision using retinal organoids grown from human stem cells. 

While the RCS rat isn’t an exact replica of human retinal diseases, it’s nevertheless an important model for researchers seeking ways to treat retinal degeneration. In humans, diseases involving retinal degeneration include age-related macular degeneration (AMD) and the genetic condition retinitis pigmentosa. Retinal degeneration diseases affect hundreds of millions of people worldwide and have enormous impacts on daily functioning and quality of life (2,3). For most retinal degeneration disorders, there are no treatments to halt the progress of disease, let alone restore vision that has been lost. Now, researchers like Seiler are seeking to cure these debilitating diseases by replacing lost cells with lab grown stem cell-derived retinal cells. 

Saving support cells

While the light sensing photoreceptors are crucial for vision, they don’t function in isolation. A tightly packed layer of cells called the retinal pigment epithelium, or RPE, performs essential supportive functions such as recycling photoreceptor debris and mediating the flow of nutrients from the blood to the rest of the retina (4). AMD is primarily a disease of RPE dysfunction, but once the RPE is destroyed, the photoreceptors begin to die as well, leading to vision loss (5). If scientists replace the RPE layer in time, they might be able to halt vision loss in AMD.

This is precisely what Kapil Bharti, an ocular and stem cell researcher at the National Eye Institute, is attempting to do. One benefit of this approach is that RPE cells are relatively easy to derive from stem cells (6). Unlike photoreceptors, they don’t require finely tuned light sensing machinery, and they don’t need to form precise synapses with other cells. 

In order to achieve this goal, Bharti and his team first optimized the process of obtaining cells from patients’ blood, converting them into induced pluripotent stem cells, then coaxing them to become RPE cells. Using patients’ own cells eliminates the risk of rejection and the need for immunosuppressant drugs, but it is a slow and laborious process. “We needed to design a protocol that was very efficient and very reproducible since we have to make the cell therapy for every patient,” said Bharti. 

Previous research showed that simply injecting RPE cells into the back of the eye did not result in an appropriately structured single cell thick RPE layer, so the research team created a patch with a single layer of properly organized RPE cells on a biodegradable scaffold (7).

Next, they tested the patch in animal models. Although a modified version of the patch showed success in a rat model, a rat eye is much smaller than a human eye. “We needed an animal model where we could test the entire patch that will go into patients, which is eight square millimeters,” said Bharti. “We also needed to develop the surgical procedure and optimize a tool to deliver the patch to the back of the eye.”

For this, the researchers turned to pigs. They used a micropulse laser to damage just the pig RPE cells, then transplanted the human RPE patch into the damaged area. Over the following ten weeks, researchers found that the majority of the transplanted cells survived, integrated into the pigs’ retinas, and, most importantly, protected their photoreceptors from degeneration (7). 

We hope to transplant our patch in at an earlier stage of AMD where we can still halt significant vision loss. 
- Kapil Bharti, the National Eye Institute

The success of this preclinical work prompted researchers to launch a clinical trial, and in August 2022, the trial’s first patient received an RPE patch derived from his or her own stem cells (5). At this early phase, researchers are largely seeking to determine safety and feasibility of this treatment and therefore are recruiting patients who are already in fairly late stages of the disease. In future trials, said Bharti, “We hope to transplant our patch in at an earlier stage of AMD where we can still halt significant vision loss.”

If successful, this treatment could be life changing for patients, ensuring that AMD is no longer a diagnosis that condemns patients to losing sight. However, the RPE patch cannot bring back photoreceptors that are already dead, nor can it treat disorders such as retinitis pigmentosa, in which photoreceptors are the primarily affected cells. In these cases, researchers must surmount the trickier task of making and installing photoreceptors.

Steps toward photoreceptor replacement 

David Gamm, now a vision researcher at the University of Wisconsin-Madison (UW-Madison), first became fascinated with photoreceptors while pursuing his MD/PhD degree at the University of Michigan. This fascination led him to choose ophthalmology as his medical specialty. 

“As a pediatric ophthalmologist, one of the things that I do — unfortunately, too often — is diagnose retinitis pigmentosa,” said Gamm. “Unlike a lot of other things that I can fix in the clinic or the operating room, that’s one thing that I can’t fix.”

Retinitis pigmentosa affects about one in four thousand people. It is caused by a mutation in any one of dozens of different genes and leads to photoreceptor death and substantial vision loss relatively early in life. 

“I started thinking about how we can keep the photoreceptors healthier. How can we keep them alive? And how can we replace them when they’re gone?”

During Gamm’s ophthalmology residency at UW-Madison, the university became a hub for stem cell research in the wake of biologist James Thomson’s first successful isolation of human embryonic stem cells in 1998 (8). Gamm dove into stem cell research, beginning the long journey towards coaxing stem cells into functional photoreceptors that could one day replace those lost to disease. This was no easy task. Photoreceptors require complex intracellular machinery to transform light into signals that can be understood by the nervous system.  

In 2012, Gamm’s research team successfully used induced pluripotent stem cells derived from human blood to create structures similar to optic vesicles, the pouches observed in early embryos that develop into eyes (9). Four years later, Gamm and FUJIFILM Cellular Dynamics cofounded Opsis Therapeutics to develop this research into cell replacement therapies to treat retinal degeneration disorders. 

Eventually, the UW-Madison research team created photoreceptors that appeared to have all the correct machinery, but, said Gamm, “We didn’t know if they could actually do the job of detecting light and converting that into an electrical signal.” To find out, Gamm teamed up with Raunak Sinha, a neuroscientist at UW-Madison who specializes in visual processing. The team found that about 35 percent of their stem cell derived cone cells, the photoreceptors responsible for color vision, behaved remarkably similarly to cone cells from nonhuman primates by responding to light — and lack of light — in appropriate ways (10). Although the team is still working on making these responses more consistent across the entire population of cones, “From a proof-of-concept standpoint, this shows that these cells aren’t just statues of cones; they can actually do the job of a cone,” said Gamm. 

Seiler has also been working on growing retinal cells from stem cells; she’s currently exploring how these may be able to restore vision in rat models of retinal degeneration. “Because there are diseases that affect the retinal pigment epithelium as well as the photoreceptors, we wanted to try to replace both of these cell types,” said Seiler. To accomplish this, Seiler’s research team transplanted pieces of retinal organoids into the backs of the eyes of retinal degenerate rats. In addition, in collaboration with research ophthalmologist Biju Thomas’ team at the University of Southern California, her team has used a patch containing a layer of RPE cells glued to a retinal organoid sheet containing photoreceptor progenitor cells (1). The patch is supported by an artificial Bruch’s membrane, the thin layer of connective tissue that sits just behind the RPE in the eye.

Seiler’s team demonstrated that these grafts not only survived long term in the rat eye, they also integrated into the host retina. Using the moving black and white bars to test visual acuity, the team found that rats that received the treatment had significantly better vision than those that received a sham surgery. The team also recorded brain activity and found that the treated rats’ brains responded to light, whereas untreated rats’ brains did not, further supporting improved visual function (1). 

Unlike Bharti’s autologous RPE trial, both Gamm and Seiler are working on a one-size-fits-all approach: a ready made patch that could be transplanted into anyone. Using a single product rather than trying to make custom photoreceptors for each patient has the advantage of being faster and more cost effective. 

Great expectations 

“Ultimately, you do not know if anything works in a human until it is actually tested in a human,” Gamm said. He believes that it won’t be long before the team is ready to begin clinical trials.

Gamm cautions against expecting too much from these early trials. “In the beginning, we’re just hoping to make measurable improvements. That might be taking a patient who can’t see light and giving them the ability to see light, so they can tell if it’s day or night. Or giving a patient the ability to see motion, so they can tell if a car is approaching,” said Gamm. “We're not looking to take somebody who's blind and give them their driver's license back. That's an unrealistic expectation for these early days.”

Yet, there is still much to be learned from these trials. “We’ve come a long way in terms of cell manufacture — what we’re putting in the eye. But what is still a big black box is where we’re putting them,” said Gamm. Unlike a standardized animal model, every patient is different. Even with a perfect product, its not clear how it will function when it is placed into a patient’s eye, especially when ongoing disease makes the eye a potentially hostile environment for the new cells. Gamm likens this to placing a perfectly functioning engine into a completely rusted car. It will be important to choose the right patients and to carefully assess which patients respond well and which do not. Gamm’s research group, as well as others, are examining strategies that could make the host retina a more hospitable environment for the transplanted cells, but it will be a while before these strategies are ready for implementation in patients.

Seiler said that it’s also an open question how long these transplanted cells will last. The patch she created can last for a year in rats, but since rats only live about two years, it’s not possible to test how the patches perform over several years in this species. 

While there are still questions to be answered and problems to be solved, the advances that have been made in creating lab grown retinal cells are truly remarkable. Slowly but surely, researchers are edging closer to halting and even reversing vision loss in retinal degeneration disorders.

References

1. Thomas, B. B. et al. Co-grafts of Human Embryonic Stem Cell Derived Retina Organoids and Retinal Pigment Epithelium for Retinal Reconstruction in Immunodeficient Retinal Degenerate Royal College of Surgeons Rats. Front Neurosci  15, 752958 (2021).
2. Vyawahare, H. & Shinde, P. Age-Related Macular Degeneration: Epidemiology, Pathophysiology, Diagnosis, and Treatment. Cureus  14, e29583
3. O’Neal, T. B. & Luther, E. E. in StatPearls (StatPearls Publishing, 2023). 
4. UCL. The retina and retinal pigment epithelium (RPE). UCL Institute of Ophthalmology (2020). 
5. First U.S. patient receives autologous stem cell therapy to treat dry AMD. National Institutes of Health (NIH) (2022).
6. fb_canada. Restoring Vision with Stem Cells. Fighting Blindness Canada (FBC) (2019). 
7. Sharma, R. et al. Clinical-grade stem cell-derived retinal pigment epithelium patch rescues retinal degeneration in rodents and pigs. Sci Transl Med  11, eaat5580 (2019).
8. Thomson, J. A. et al. Embryonic Stem Cell Lines Derived from Human Blastocysts. Science 282, 1145–1147 (1998).
9. Phillips, M. J. et al. Blood-derived human iPS cells generate optic vesicle-like structures with the capacity to form retinal laminae and develop synapses. Invest Ophthalmol Vis Sci 53, 2007–2019 (2012).
10. Saha, A. et al. Cone photoreceptors in human stem cell-derived retinal organoids demonstrate intrinsic light responses that mimic those of primate fovea. Cell Stem Cell 29, 460-471.e3 (2022).