ocular surface & dry eye
Linking inflammation and dry eye

Recent studies confirm inflammatory connection, boost therapeutic possibilities

Impression cytology images taken of a normal eye showing many magenta stained goblet cells in the bulbar conjunctiva and of a dry eye with a paucity of goblet cells
Source: Stephen C. Pflugfelder, M.D.


Immediate post inferior punctal cautery for severe dry eye. Dry eye lab studies are discovering inflammatory causes for the condition


Meibomitis associated with evaporative dry eye
Source: John D. Sheppard, M.D.

Some new lab studies further support the connection between dry eye and inflammation, and the research may pave the way for more specific therapeutic options for dry eye patients.

“Until a couple of years ago, we had no biologic targets; now we do, and we have specific candidate molecules,” said Reza Dana, M.D., M.Sc., M.P.H., professor of ophthalmology, Harvard Medical School, Boston; director, Cornea Service, Massachusetts Eye and Ear Infirmary, Boston; and senior scientist, Schepens Eye Research Institute, Boston. Dr. Dana leads a team of researchers that is conducting numerous investigations into dry eye biomarkers.
Research into inflammatory-specific molecules and dry eye could lead to future diagnostic markers and treatments, said Stephen C. Pflugfelder M.D., professor of ophthalmology, Baylor College of Medicine, Houston. Dr. Pflugfelder also has led a number of studies related to inflammation and dry eye and has made similar conclusions as Dr. Dana’s team.
Although some ophthalmologists were initially skeptical about inflammation as the cause for dry eye, the pendulum has moved in favor of inflammation as a cause because of growing research findings, Dr. Dana said. “Since the Dry Eye Workshop, there has been an explosion of new information, and if you look for inflammatory mediators at the cellular level, you universally see them,” Dr. Dana said.

What recent research shows

Recent studies on biomarkers provide a window into dry eye causes. One recent study, “Autoimmunity in dry eye is due to resistance of Th17 to Treg suppression,” was published in the January issue of The Journal of Immunology and was led by Dr. Dana and co-investigators.
Investigators noted that although other researchers have reported evidence for T-cell involvement as the cause of dry eye, most of the previous work has focused on the effector T-cell responses.
“The concept that inefficient functioning of regulatory T-cells (Tregs) may cause unrestrained generation of pathogenic T-cells in DED [dry eye disease] has not yet been systematically investigated,” the investigators wrote. As the investigators explained, Tregs are a “naturally occurring suppressor T-cell population” that inhibits the activity of cells thought to induce inflammation and tissue destruction.
One of the cells thought to induce inflammation is Th17, or interleukin (IL)-17+. IL-17 is undergoing extensive research for its connection to autoimmune diseases such as rheumatoid arthritis and multiple sclerosis, Dr. Dana said.
The investigators theorized that defective Treg homeostasis leads to a dysregulated activation of self-reactive T-cells. To test their theory, dry eye was induced in mice in a controlled-environment chamber; there was also a control group. Investigators performed cell isolation, a FACS analysis, suppression assay, real-time polymerase chain reaction, immunohistochemical staining, and in vivo IL-17 neutralization.
“Tregs isolated from DED mice showed a significantly reduced potential in suppressing proliferation of both naïve T-cells and primed T-cells compared with those from normal mice. In addition, Tregs of DED mice were significantly less effective in suppressing the proliferation of primed T-cells than those of naïve T-cells,” they wrote.
The research also included an analysis of the effects of an IL-17 blockade on Th17 cell frequency and Treg function, which suggested a clear inverse relationship between Th17 cells and Treg function in dry eye.
“The induction of Th17 cells and the lack of Treg control over pathogenic T effectors, in combination, lead to the induction of autoimmunity and sustained inflammation in DED,” investigators concluded.
A study from Dr. Pflugfelder and co-investigators, published online in Immunology in February, also focused on the Th17 response and its role in corneal epithelial barrier disruption in dry eye in a human and experimental mouse dry eye.
“Gene expression in the human dry eye conjunctiva showed increased levels of the Th-17 inducers, IL-23, IL-17A, and interferon-gamma,” they wrote. They also noted that antibody neutralization of IL-17 eliminated experimental dry eye-induced corneal epithelial barrier dysfunction and decreased the expression of matrix metalloproteinases (MMP) -3 and -9.
Dr. Pflugfelder’s team took a closer look at MMP-9 in a study published online in February in the Investigative Ophthalmology and Visual Science. That study evaluated the production and activity of MMP-9 on the ocular surface of 46 patients with dry eye and 18 control subjects. The mean MMP-9 activity was higher in each of the four dry eye severity-based groups than in the control group; the patients with the most severe dry eye had significantly higher levels of MMP-9 activity; dry eye patients had significantly higher levels of IL-1, IL-6, tumor necrosis factor-alpha, and tumor growth factor-1 mRNA transcripts in their conjunctival epithelia compared with controls. “MMP-9 appears to be a potentially useful biomarker for diagnosing, classifying, and monitoring [dysfunctional tear syndrome],” the investigators concluded.

A turning point

Ophthalmologists believe that dry eye research is at a critical turning point right now, similar to arthritis research and treatment about 15 years ago. “Fifteen years ago, steroids and cyclosporine were used for arthritis. Since then, there have been great advances in autoimmune disease with tumor necrosis factor-inhibitors. Understanding the biochemical pathophysiology of certain kinds of arthritis has revolutionized its treatment,” said John D. Sheppard, M.D., professor of ophthalmology, microbiology and immunology, Eastern Virginia Medical School, Norfolk, Va.
With target-specific diagnostic markers and treatments, treatment is more efficient, and there are fewer side effects than with more general treatments such as steroids, he said.
A deeper research-based connection between dry eye and inflammation also confirms the proper use of Restasis (ophthalmic cyclosporine, Allergan, Irvine, Calif.)—the current go-to prescription medication for dry eye, said Esen K. Akpek, M.D., associate professor of ophthalmology, and director, Ocular Surface Diseases and Dry Eye Clinic, Wilmer Eye Institute, Johns Hopkins University, Baltimore.
“Most people don’t start Restasis until the patient has already fibrosed,” she said. Instead, this research backs the idea of starting cyclosporine even with earlier signs of dry eye, she said.
It has taken dry-eye research a longer time to find connections between inflammation and disease than it has in other specialties, Dr. Akpek said. “Maybe there are more advances in other specialties because of their association with morbidity and mortality,” she said. “Dry eye may not kill you, but it can make you miserable.”
Dr. Dana believes that it has taken some time to study biomarkers for dry eye because there was lack of a good model to study the disease. “When we didn’t have good models, we couldn’t really study it,” he said.
Compounding this has been the difficulty of tracking dry eye endpoints in humans for the purpose of new drug approval, said Michael A. Lemp, M.D., clinical professor, Georgetown and George Washington universities, Washington, D.C. Environmental factors often play a role in this difficulty; Dr. Sheppard said that dry eye studies in humans often need at least a year-long course to rule out environmental factors that occur during pollen and ragweed season.
Dr. Lemp also points out that dry eye induced in a lab does not mimic dry eye in systemic autoimmune disease patients.
Although human research is ongoing with potential dry eye drugs, mouse models in which dry eye is induced are common in the lab studies. In fact, one study published in the April 2009 issue of Archives of Ophthalmology tested the lubricant chitosan-N-acetylcysteine conjugate in mice who had dry eye induced by botulinum toxin B injection.
“This allows us to screen many candidate drugs for the differing severity of dry eye in a high throughput fashion,” said study co-investigator Roy S. Chuck, M.D., Ph.D., Tom Clancy Professorship in Ophthalmology, and director of refractive surgery, Wilmer Eye Institute, Johns Hopkins University. Although the model is an aqueous-deficient one, investigators observed secondary inflammatory changes on the ocular surface, Dr. Chuck added.

Looking forward to treatment

The big question—one that’s hard to place a timeline on—is how soon the identification of dry eye inflammatory biomarkers will lead to more specific treatment and which treatment will be most effective.
“There are a lot of different ways it could be approached, and I’m not sure which will be the best way. It’s going to be an active area,” Dr. Pflugfelder said. Some of the lab findings, such as the presence of MMP-9, may also provide strong diagnostic insights.
“We don’t know yet which one or two or three will be the most important in treating the disease process,” Dr. Pflugfelder said.
“We can target specific molecules in the generation of dry eye or the immune expression of dry eye, and we can inhibit the disease,” Dr. Dana said.
Although a number of drugs are under investigation, the U.S. Food and Drug Administration is approving a smaller number of drugs each year, Dr. Sheppard said.
One possibility from Dr. Dana’s team, published in the December 2008 Archives of Ophthalmology investigated the topical application of very late antigen 4 (VLA-4) small-molecule antagonist (anti-VLA-4 sm) in a mouse model involving dry eye. VLA-4 has been shown to serve as a target for treating inflammatory disorders, including relapses of multiple sclerosis, suppression of endotoxin-induced uveitis, and inhibiting eosinophil infiltration in a guinea pig model of allergic conjunctivitis, according to the study.
“Although inflammation has been shown to be related to the pathogenesis of dry eye disease, topical blockade of VLA-4 or other integrins has not been investigated in DED, to our knowledge,” investigators wrote. The VLA-4 blockade significantly decreased staining compared with untreated dry eye and the control group. The VLA-4 blockade also was associated with a significant decrease in conjunctival T-cell numbers and tumor necrosis factor-alpha transcript levels in the cornea and conjunctiva.
In addition to treatments for specific inflammatory biomarkers, secretagogues such as diquafosol tetrasodium (Inspire Pharmaceuticals, Durham, N.C.), which is completing final Phase III trials, remain another therapeutic avenue, Dr. Sheppard said. Dr. Sheppard’s lab is also investigating the recombinant application of lacritin, a naturally occurring protein in the tear film that is believed to be particularly potent. This research, funded by the National Institutes of Health, is beginning Phase IA studies, Dr. Sheppard said.

Editors’ note: The Archives of Ophthalmology study was supported by a research grant from Biogen Idec (Cambridge, Mass.) to Dr. Dana. Dr. Pflugfelder has no financial interests related to his comments. Dr. Sheppard has financial interests with Alcon (Fort Worth, Texas), Allergan (Irvine, Calif.), and Inspire Pharmaceuticals (Durham, N.C.), among other ophthalmic companies. Dr. Akpek has no financial interests related to her comments. Dr. Lemp has financial interests with OcuSense (San Diego). Dr. Chuck’s study was supported by an unrestricted grant from Croma-Pharma (Komeuburg, Austria).

Contact information

Akpek: 410-955-5494, esakpek@jhmi.edu
Chuck: 410-583-2802, rchuck1@jhmi.edu
Dana: 617-912-7401, reza.dana@schepens.harvard.edu
Lemp: 202-338-6424, malemp@lempdc.com
Pflugfelder: 713-798-4944, stevenp@bcm.tmc.edu
Sheppard: 757-622-2200, docshep@hotmail.com