Several members of the University of Pittsburgh Department of Ophthalmology are working on optic nerve regeneration research. In the Eye & Ear Foundation’s June 13th webinar, “Current Updates in Optic Nerve Regeneration Research,” Drs. Peter Mortensen, Larry Benowitz, and Takaaki Kuwajima talked about the current landscape.
Optic Neuropathy
Dr. Peter Mortensen, MD – who joined UPMC as an Assistant Professor of Ophthalmology in May 2025 – provided a clinical overview of optic neuropathies. Optic neuropathies are diseases that damage the optic nerve, leading to vision loss. What is the optic nerve? It is the interface between the eyes and brain, technically part of the central nervous system. Optic nerves lack the ability to regenerate. A “sick” nerve can sometimes heal, but dead nerves do not regenerate.
There are numerous causes for optic neuropathy, like toxins, nutritional deficiencies, brain tumors, inflammatory disease, infections, genetic conditions, intracranial pressure elevations, and glaucoma. The current focus for diseases of the optic nerve is on maintaining the remaining optic nerve rather than regeneration/regrowth.
Glaucoma
Glaucoma is the most common optic neuropathy and is defined as “optic neuropathy with characteristic visual field defects.” It is often associated with high intraocular pressure. The most common form – primary open angle glaucoma – results in progressive vision loss, often over many years. The optic nerve progressively thins over time. Patients are unaware of visual field loss until late in the disease. Treatments are focused on maintaining rather than restoring both visual fields and optic nerve thickness.
Traumatic Optic Neuropathy
Traumatic optic neuropathy can occur with direct trauma to the eye or periorbital region (particularly the frontal bone). Treatment is controversial, because improvement is sometimes observed without treatment. Steroids may have some benefit, though this is not proven. Vitamins/supplements have unclear benefit. Optic nerve decompression can occur if there is a fracture or hematoma.
Traumatic optic neuropathy is frequently seen with traumatic brain injury (TBI). A CRASH (Corticosteroid Randomization After Significant Head Injury) study found that there is a higher risk of death in patients with TBI following high-dose steroids.
Autosomal Dominant Optic Atrophy
The most common inherited optic neuropathy is autosomal dominant optic atrophy (ADOA), which – as its name implies – is inherited in an autosomal dominant manner. Only one copy of the gene is required. It is most often associated with an OPA1 mutation, and generally presents with painless, progressive, bilateral visual loss through life. Visual acuity is usually better than 20/200. There is variable expressivity, or a wide range of visual impairments that are sometimes very minimal but can be very severe. ADOA is associated with mitochondrial dysfunction. There is no established treatment for ADOA, but it is sometimes treated with vitamin B12, coenzyme Q-10, and idebenone, though there are no proven benefits.
Leber Hereditary Optic Neuropathy
Leber hereditary optic neuropathy (LHON) is the second most common inherited optic neuropathy and the most common inherited mitochondrial disorder. It is passed down as a mitochondrial gene, i.e. a mother to all children; it is not passed down from the father. It generally presents as a bilateral and sequential vision loss and is most common in young males, presenting in their 20s to 40s. There is incomplete penetrance, with 50% of males and 10% of females who have the gene present with vision loss.
The three most common genes are:
- 11778 – most common, minimal chance of visual recovery
- 14484 – 2nd most common, sometimes presents with spontaneous recovery
- 3460 – 3rd most common, minimal chance of visual recovery
Treatments include coenzyme Q-10, idebenone (which seems to stabilize vision loss based on recent studies), avoiding mitochondrial stimulants (e.g. alcohol, caffeine, and certain medications) to prevent worsening, and gene therapy trials (lenadogene nolparvovec). These trials are currently under investigation to deliver a gene for the ND4 enzyme and restore function in retinal ganglion cells (RGCs). Studies demonstrate improvement in vision with this therapy.
NAION
NAION stands for nonarteritic anterior ischemic optic neuropathy, the most common cause of acute, unilateral vision loss in a patient above 50. Calling it a stroke of the optic nerve is a misnomer, because technically it is small vessel disease of the nerve. It is partially due to vascular risk factors like hypertension, hyperlipidemia, diabetes, and obstructive sleep apnea. A nonmodifiable risk factor is the congenital component of a crowded optic nerve.
NAION presents initially with optic disc edema and progresses to optic atrophy. It is often nonprogressive in the affected eye and results in altitudinal field defects. There is minimal improvement in vision after vision loss, with 15% chance of fellow eye involvement in five years. Unfortunately, there is no cure. The focus is on prevention by modifying vascular risk factors.
Giant Cell Arteritis (GCA)
GCA can cause vision loss due to multiple mechanisms, but most commonly arteritic anterior ischemic optic neuropathy, or AAION. This generally affects individuals older than 50. Other associated symptoms include:
- Headache and scalp tenderness
- Polymyalgia rheumatica
- Jaw and tongue claudication
- Transient or permanent vision loss
- Diplopia
- Unintentional weight loss
- Fever of unknown origin
Lab markers of inflammation (ESR, CRP, platelets) are elevated in 80% of cases. The gold standard test is a temporal artery biopsy. The condition is treated with long-term high-dose steroids and steroid-sparing therapies.
With GCA, once vision loss occurs, there is rarely significant improvement, even if treatment is started early. Vision loss can still progress with high-dose steroids. Some patients end up with irreversible vision loss to the no light perception level in both eyes.
Challenges with GCA are:
- Laboratory markers are nonspecific and unreliable
- Variable presentation – sometimes vision loss is initial symptom
- Diagnosis can be unclear, even with labs, biopsy, and additional testing
- Steroid treatment and steroid-sparing regimens can also cause significant harm, up to and including death in rare cases
The current treatment goal is to prevent further vision loss rather than recover vision.
Optic Neuritis
The most common presentation of optic neuritis is unilateral or bilateral, with frequently painful vision loss over several hours. The MRI will show optic nerve enhancement. Common causes are MS, neuromyelitis optic (NMO), or myelin oligodendrocyte glycoprotein associated disease (MOGAD).
Variable presentation/prognosis depends on the cause:
- MS – usually unilateral, moderate response to steroids, typically good recovery
- NMO – unilateral or bilateral, poor response to steroids (unless started early), often poor recovery
- MOG – unilateral or bilateral, rapid response to steroids, typically good recovery
Optic nerve atrophy is almost always present, regardless of cause, and is permanent. With NMO, there is a high risk of permanent blindness if untreated. Similar to GCA, the goal is often to retain current vision with high-dose steroids rather than visual recovery.
Optic Neuritis Treatment Trial
This was a randomized, controlled clinical trial investigating efficacy of different corticosteroid regiments for treatment of optic neuritis. Treatment groups are high dose IV methylprednisolone, lower dose oral prednisone, and a placebo. They found that high dose steroids promote faster recovery of vision, but there is no difference in visual recovery by six months compared to the placebo. High dose steroids temporarily reduce the risk of MS, but the effect lasts two years. Oral steroids are associated with a higher risk of relapse and progression to MS.
Optic Neuropathy – Key Points
Optic neuropathy has multiple causes: Infection, inflammation, demyelination, compression, hereditary, demyelination, trauma, glaucoma, nutrition, toxins, etc. Optic nerves are part of the central nervous system and have limited potential for recovery. “Sick” nerves can show improvement in function, but dead nerves cannot. Current clinical treatments are focused on preserving the remaining optic nerve and preventing further optic nerve fiber loss. Current research aims to eventually restore and regenerate optic nerves.
Optic Nerve Regeneration: Basic Research
Dr. Larry Benowitz, PhD, Professor of Ophthalmology at the University of Pittsburgh and Professor of Neurosurgery and Ophthalmology Emeritus, Harvard Medical School, and Co-Director of the Fox Center for Vision Restoration, said that studies are done in small animals, because from extensive studies of genes and anatomy, neurotransmitters and types of cells are remarkably similar to humans. The experimental paradigm is that the optic nerve is injured a little distance behind the eye.
Dr. Benowitz described the retina as the business part of the eye where the nerve cells are located that gather information. “It’s a remarkable computer,” he said. A huge amount of information is processed right in the retina, and then that information converges on the RGCs, which are the integrators of information in the eye. In turn, they send information back to the brain.
Damage to the optic nerve damages the nerve fibers that originate in the RGCs, and those cells cannot regenerate their axons. They begin to die after their axons are injured, with no hope of recovery.
Surprising Discovery
Years ago, Dr. Benowitz and Dr. Yuqin Yin made an accidental discovery. They accidentally nicked the lens, which caused inflammation. They stopped doing that and used other agents to promote an inflammatory reaction and discovered that intraocular inflammation causes the RGCs to transform back into an early developmental state and turn on a whole host of genes. Many of the RGCs do not die and start regenerating axons down the optic nerve.
A few years later, Dr. Benowitz’s neighbor at Boston Children’s Hospital, Harvard Medical School, Dr. Takuji Kurimoto made a complementary discovery. He found another way to get RGCs to regenerate axons by removing what essentially amounts to a break. In nerve cells, a protein called P10 suppresses the growth ability of nerve cells. Dr. Kurimoto’s lab discovered that if you delete that gene using gene therapy, regeneration occurs.
“Then we simply asked the question, what happens if you put these two things together?” Dr. Benowitz recalled. “That is, cause inflammation and knock out the gene for that suppressor. Do those two have complementary effects? The answer was, they sure do.”
Putting together the stimulus and knocking out the suppressor of regeneration provided around 10 times as much regeneration as either treatment alone. “This was far and away the most regeneration anyone had seen up to this point,” Dr. Benowitz said.
If it is extended, and this is now being done by Dr. Silmara de Lima, PhD, who is now a member of the Department at Pitt, axons grow all the way down the length of the optic nerve. In Dr. de Lima’s studies, she found that some of these nerve fibers find the proper places in the brain to form connections with a very modest level of visual recovery. This hopeful finding stimulated the National Eye Institute, part of NIH, to being to develop what they call their audacious goals initiative, to restore vision to the blind. Pitt is the recipient of a grant, which has helped continue this work. Another federal impetus has added to the effort.
Mediators of Inflammation-Induced Regeneration
What are the molecules that cause this regeneration to occur? There must be molecules made by inflammatory cells. Prior work in the field had studied some of the well-known growth factors. Turns out, it was none of the usual suspects. One of the molecules causing this regenerative growth is a protein called oncomodulin. Cells of the immune system that come into the eye when inflammation is caused make lots of this protein. This led to a major publication from Dr. Benowitz’s lab, where they showed if oncomodulin is packaged into slow-release polymer beads, the optic nerve regenerates considerable distances.
Then they discovered another molecule being made by inflammatory cells called SDF-1. When put together with oncomodulin, they realized they did not have to cause inflammation anymore. Virally mediated gene therapy is used to keep producing the proteins in the eye, getting more regeneration than from causing inflammation. If antagonists are used, if these proteins are prevented from binding to their receptors on RGCs, they pretty much eliminate the regeneration caused by inflammation.
To summarize, inflammatory agents, like Zymosan, activate immune cells, macrophages, and neutrophils. These cells, surprisingly, turn out to make what are called growth factors, or proteins that stimulate nerve cells to regenerate their axons. Now instead of intentionally causing inflammation, these defined molecules can be used to cause significant levels of axon regeneration.
Search for Additional Pro-Regenerative Factors
Having found these strange molecules that were not really on anyone’s radar screen, it occurred to them that maybe there were more remaining to be discovered. Using modern methods of seeing what all the genes are that cells express, Dr. Yin did an experiment. She dissected the retina from a mouse, dissociated the cells, and separated the RGCs. They genetically labeled the RGCs with a fluorescent red protein to make it easy to separate them from the other cell types in the retina. RGCs actually represent less than 1% of all cells in the retina.
The other cells in the retina are things like photoreceptors or interneurons that mediate signal transmission from the photoreceptors to the RGCs. Using modern tricks, they separated the RGCs, examined all the genes they are expressing, and studied what proteins the genes code. They discovered that these genes code about 70 different receptors, growth factors they never knew about.
Dr. Yin screened many of these and struck gold. She discovered several previously unknown growth factors that cause RGCs to regenerate their axons. Just putting IL12 into the eye produces this effect. Dr. Benowitz emphasized that normally there is zero regeneration, but with these molecules, they are able to get more and more and more.
Dr. Kun-Che Chang at Pitt has been working with Dr. Benowitz and Dr. Yin. Dr. Chang has been able to use organoids developed from human stem cells to find out how human RGCs respond through the growth factors they are finding. When he treats these organoids with the factor that Dr. Yin discovered, he can get human RGCs to extend axons. “This is the beginning of studies that are telling us that these molecules that we’re discovering in lower animals are directly relevant to improving axon regeneration for human RGCs,” Dr. Benowitz said.
Lens Injury Pre-Conditioning
One of the strange things that causes regeneration is injury to the lens. Dr. Qian Feng, a postdoctoral fellow in Dr. Benowitz’s lab, helped them make a strange discovery. If the lens is injured two weeks prior to optic nerve damage, remarkable regeneration occurs. This is called conditioning lens injury. Now they are trying to find out why this happens. It is not any of the factors they discovered – something else is causing it. There are multiple leads.
Receptor X-R
Dr. Lili Xie, who used to be a postdoctoral fellow in Dr. Benowitz’s lab and who is now back in China as an associate professor, made an astonishing discovery. The way growth factors work is they bind to what are called receptors on the cell that responds to them. By binding to these receptors, a cascade of molecular events is initiated. Dr. Xie found that one particular receptor is enough just by expressing it; the protein that binds to it is not even needed.
If this receptor is increased and treated with any growth factor, the amount of regeneration is doubled or tripled. On the other hand, if they knock out the gene that encodes this receptor, nothing else can stimulate regeneration. “This discovery is really remarkable,” Dr. Benowitz said. “It tells us that this kind of receptor that’s been hiding in the background is the key to regeneration.”
Summary: Pro-Regenerative Therapies
- Oncomodulin, SDF-1 mediate the effects of intraocular inflammation (via ArmC10 and CXCR4)
- CCL5 mediates the effects of CNTF gene therapy (via CCR5: involves neutrophil activation)
- Conditioning lens injury leads to extensive regeneration (mediators yet unknown)
- IL12 and other previously unexplored ligands to RGC receptors
- Receptor “X-R” overexpression induces regeneration independent of its cognate ligand and augments the effects of multiple unrelated ligands, whereas deletion abrogates positive effects of other treatments (e.g. Pten deletion)
- Others: IGF-1, osteopontin, Lin28/let-7, c-a B-raf; proteins, peptides discovered via transcriptomics (Gal, CRH, Wnt1, Anxa2, Plin2, Mpp1…); blocking receptors to cell-extrinsic growth suppressors (NgR1, 2, 3; PTP); Zn2+ chelation; transcription factors (KLF-7, Elk1, c-Myc; knock-down of REST, Klf-4, -9); epigenetic factors (DNMT3, CTCF); blocking cell-intrinsic growth suppressors (PTEN, SOCS-3, GSK3; RhoA); physiological activity; mitochondrial, ER proteins (ArmX, protrudin; M1)
The challenges are to improve RGC survival in ways that also enable axon regeneration, increase the number of RGCs that regenerate axons to the brain, especially the homologues of human “midget” and “parasol” RGCs, and seeing whether regenerating axons can navigate to their appropriate target areas.
ARPA-H
Pittsburgh was fortunate enough to receive an ARPA grant from the federal government, the Advanced Research program for health initiated by President Biden. Restoring vision to the blind is part of the program. The team at Pitt is comprised of Takaaki Kuwajima, Kun-Che Chang, Silmara de Lima, Boris Rosin, John Ash, and Larry Benowitz. The strategy is to work simultaneously on mouse, rat, and organoid models to optimize regenerative therapies, develop a transplantation model, and identify chemoattractants.
The idea is to first use the mouse model for quick throughput to test the number of treatments. They already know that in the mouse they can injure the optic nerve, give various kinds of treatments, gene therapy, and get significant regeneration. The next step is moving up to a larger animal model, the rat. From there, they are going to small pigs, and then hopefully non-human primates.
They hope to completely transect the optic nerve. Studies done by a neuro-ophthalmologist years ago, Alberto Aguayo, showed that they can attach a piece of sciatic nerve, peripheral nerve, to the cut end of the optic nerve, and get a lot of axons regenerating through that peripheral nerve.
The group at Pitt is combining their treatments that promote regeneration with what is being discovered through combinatorial treatments to get remarkable levels of regeneration in animal models that more and more closely approximate the human visual system.
Optic Nerve Growth and Regeneration Lab
Dr. Takaaki Kuwajima, PhD, Assistant Professor of Ophthalmology, has a lab that is focused on understanding the cellular and molecular mechanisms of RGC protection and optical regeneration after optic nerve injury.
A lot of axon regeneration inhibitory molecules exist in the environment. When Dr. Kuwajima was a postdoc at Columbia University, they set up drug screening using a neuron and in molecules to mimic this inhibitor environment. They tested 50,000 chemical compounds to identify which drugs could overcome this inhibition and promote axon outgrowth. They identified that FDA approved statins are the best candidates.
The mouse optical crush model is one of the well-established injury animal models. Using this, they found that after opting out of an injection of the most effective statin, cerivastatin, into the eye after optic nerve crush, axon regeneration was observed. Several issues were still observed: regenerative ability is still limited, RGC protection is weak, and cerivastatin is not available on the market because of strong side effects.
A current study with Dr. Steve Badylak, DVM, MD, PhD, a professor at Pitt, involves matrix-bound nanovesicles (MBV), which are critical components of FDA-approved extracellular matrix (ECM) biomaterials and healthy tissues. They are neuroprotective reagents in the mouse model of glaucoma. The study combines fluvastatin, which is still available on the market, and MBV to investigate the effects on axon regeneration and RGC protection after optic nerve injury.
Greg Campbell in Dr. Kuwajima’s lab just found that this new combination therapy using MBV and fluvastatin more strongly promoted RGC protection and optic regeneration compared to either alone. They also found some interesting cellular mechanisms. They checked immune responses from the combination therapies and found that they increased the number of specific immune cells, such as neutrophils and monocytes, but not by either alone.
An additional collaboration was set up with Dr. Vijay Gorantla and his surgical team at Wake Forest School of Medicine. They investigated MVB and a fluvastatin based therapy in pigs after optic nerve crush. As observed in the mouse, they found that these combination therapies showed a high RGC production and optic nerve regeneration. This treatment also reduced optic nerve degeneration and minimized defects in the visual functions after injury.
They hope to get the same results in monkeys after injury and then use those therapies for humans in the near future.
An ongoing project is ARPA-H, researching how to establish robust regenerative therapies after optic nerve transection and nerve connections. Using validated single treatments, like modulation of epigenetic factors, intrinsic signals, cytoskeletal and axonal structural regulators, and clinically relevant and/or non-viral reagents, currently they have been testing 45 new combinations and trying to identify novel and robust combination therapies.