As an ophthalmology resident at Wilmer Eye Institute, Johns Hopkins Medicine, Thomas Johnson encountered what all ophthalmology residents eventually encounter: patients with irreversible vision loss. “I would see patients with glaucoma or ischemic optic neuropathy — diseases in which the optic nerve is damaged — and they had no vision or very poor vision, and I had to say I’m sorry, there’s really not much I can do to make you see better. When you experience that, you just wish you had some way to reverse it.”
Today, as an assistant professor of ophthalmology at Wilmer, Johnson is co-leading a Johns Hopkins team that’s seeking to do just that.
Johnson has joined forces with neuroscientist Alex Kolodkin and Don Zack, a molecular and stem cell biologist and director of Wilmer’s Center for Stem Cells and Ocular Regenerative Medicine (STORM), to form the Optic Nerve Regeneration Initiative (ONRI). Their goal? To determine what it will take to regenerate the optic nerve in human patients — and then to actually do it.
Zack, the Guerrieri Professor of Genetic Engineering and Molecular Ophthalmology at Wilmer, says that in some ways, optic nerve regeneration is the “holy grail” of ophthalmology. Jeremy Nathans, a professor of molecular biology and genetics, neuroscience and ophthalmology at Johns Hopkins, said it is comparable to that of the original moon mission, except that when the astronauts headed to the moon, they knew what they needed to do to get there. “They knew it was going to be a monumental task,” says Nathans, “but they understood the physics of how it needed to be done. In regenerating the optic nerve, we still have a lot of questions about how the underlying physiology works.”
Like a Fiber-Optic Cable with a Million Fibers
The optic nerve, which has been likened to a fiber-optic cable, is actually a cluster of nerve cells that carries visual information from the retina to the brain. It begins in the retina as the axons of specialized nerve cells, called retinal ganglion cells (RGCs). We’re born with approximately a million RGCs per eye, and when they die, they’re gone forever. In glaucoma — the second leading cause of blindness worldwide — there is an accelerated loss of the axons, and at some point, there aren’t enough to support vision. “Considering that the human optic nerve doesn’t have the ability to heal itself, to help someone with advanced glaucoma to see again, we would have to regenerate the optic nerve,” explains Johnson.
Over the past two years, the ONRI group has been working to assemble a multidisciplinary team of researchers to determine the steps this would involve. One thing they know for sure is that it would require repopulating the retina’s RGCs. To do that, they would need the expertise of stem cell biologists to generate new RGCs; neuroscientists who understand the physiology and circuitry of the cells; ophthalmologists who understand how the eye, retina, and optic nerve work together to communicate with the brain; and neurologists who understand the inner connectivity of the brain and how the brain processes visual information.
In addition, biomedical engineers and biomaterial scientists are needed to generate methods and materials that support the development of new nerve fibers. To date, the ONRI team includes 16 principal investigators or labs involved in the effort — and the group is still recruiting.
Determining a Recipe for RGCs
In advanced glaucoma, it’s not only the fibers that die; the RGCs that live inside the eye actually die. That means regenerating the fibers in the nerve alone is not enough to restore vision. It also requires replacing those entire cells. As Johnson explains, one of the fundamental problems researchers face in stem cell research has been what to replace those cells with. “We have stem cells that are capable of becoming any cell in the body, but you have to develop precise recipes to direct them to become something specific,” he says.
Over the past several years, Zack’s lab, along with a handful of other labs around the country, has achieved this feat, albeit on a rudimentary level. “They essentially figured out how to modify undifferentiated stem cells in a dish to become RGCs,” says Johnson. Researchers have since become increasingly sophisticated at creating cells in a dish that better model a true RGC. Only now, Johnson says, do we have something we might be able to transplant into an eye that has the potential to regenerate the optic nerve. But it’s complicated.
The complexity of the group’s task becomes more apparent when one considers that there are different kinds of RGCs — and that each kind has a different job. “We’re talking about RGCs as if they’re one thing, but in humans, there are at least 30 subtypes of RGCs,” Johnson says. “Some of them are important for conveying information about fine details and color; others, shape and directionality; and still others, movement. Some will communicate to the brain only when something is moving up to down in the visual field. Others might fire when you’re looking at something right on the edge of an object.” To make things even more complicated, he notes, different RGCs wire to different parts of the brain.
While it’s possible that one overall developmental plan governs the behavior of these subtypes, it’s also conceivable that the different subtypes will require different approaches. Kolodkin, the Charles J. Homcy and Simeon G. Margolis Professor at the Department of Neuroscience, and Deputy Director of the Institute for Basic Biomedical Sciences at the Johns Hopkins School of Medicine, is studying how the different subtypes develop normally. Knowing this will allow the group to try to repurpose those same pathways for transplanted cells.
The Sum of the Parts
Once the team is able to generate the right combination of stem cell-derived RGC subtypes, they’ll need special tools to transplant them into the eye: nanocarriers, to carry, protect and nourish the cells; scaffolds, to align and position them to direct them to the parts of the brain where they actually need to grow; and surgical instruments, to manipulate the product into the eye. Zack and biomedical engineer Hai-Quan Mao have already generated a nanofiber scaffold in a dish.
Eventually, the group hopes to make implantable devices out of the same materials that confer directional guidance to the cells. Johnson envisions a two-dimensional scaffold — like a patch — where they will seed the RGCs made from stem cells. “This will probably be something that’s rolled up like a scroll that we can inject into the eye and then unroll,” he says. For that, they’ll need instruments to manipulate it in the eye and get it to lie atop the retina.
In his lab, Johnson is exploring ways to circumvent natural barriers that hinder the migration of RGCs from the transplant site into the retina. One such barrier is a membrane that exists at the interface between the vitreous and the retina, called the internal limiting membrane (ILM). Johnson has found that using enzymes to digest the membrane in mice and rats makes the ILM more porous, allowing more of the transplanted cells to get into the retina, where they can grow dendrites and begin to form crucial connections with retinal neurons.
Assuming the group learns how to repopulate RGCs and transplant them so they can find their way into the retina, the next challenge will be to get them “talking” to the brain. To do so, RGCs must grow their own axon fibers through the optic nerve and into the brain. In turn, the axon fibers need to create molecular communication stations, called synapses, with brain neurons, in order to successfully communicate visual information from the eye. So, while Zack’s lab is working on stem cell engineering, and Johnson’s lab is working on transplantation and incorporation of stem cells into the retina, Kolodkin’s lab is working on methods to promote axon growth through the optic nerve.
“We are making progress,” says Johnson, “but the more people we can have thinking about the problem, collecting the data and generating the tools we need, the faster the process will go.” In May, the group hopes to host a virtual gathering of researchers from around the world to try to form a better consensus about the problems researchers are encountering, the tools and experiments needed, and the roadblocks to progress.
“There are a lot of people who are working on similar questions in slightly different ways, and by getting everyone together to try to figure out the best way to tackle problems, we can be more efficient with our time and resources,” says Johnson. Working together on some of these problems, he adds, could also generate new and improved ideas and collaborations. Says Zack, “This is an exciting time, in which vision restoration through optic nerve regeneration is rapidly transitioning from a dream to a foreseeable reality.”