A Novel Uveolymphatic Drainage Pathway—Possible New Target for Glaucoma Treatment

Introduction

Glaucoma is a heterogeneous group of ophthalmic diseases leading to irreversible damage to the optic nerve. According to the World Health Organization, glaucoma is the second leading cause of blindness in the world. ¹ It is estimated to affect ∼70 million people worldwide. ² While the overall mechanism responsible for glaucoma remains obscure, the most important risk factor is elevated intraocular pressure (IOP).^3,4

Despite much advancement in medicine and the dedicated research efforts, the etiology and causes of glaucoma are still unknown. Glaucoma is known to be multi-factorial in origin with some established risk factors predisposing patients to the disease. The risk factors include: age over 40 years, family history of glaucoma, peripheral circulatory blood disorders (e.g., reduced flow in carotid arteries), steroid and other drug therapy, hypertension, significant degree of vision defects (myopia), and diseases of the eye (pseudoexfoliation syndrome, diabetic retinopathy, uveitis, injury).^3,5–7

Pseudoexfoliation syndrome is an age-related elastic microfibrillopathy, in which material is produced, aggregated, accumulated, and is not degraded in vivo in the anterior segment of the eye. This material is produced primarily by the nonpigmented epithelium of the ciliary body, the posterior iris pigment epithelium, and the preequatorial lens epithelium. Pseudoexfoliation glaucoma is associated with poorer response to treatment. It is determined by blocking the trabecular meshwork (TM) both by pigment grains released from the iris during mydriasis, and the pseudoexfoliation material.^8,9

Therapy, Treatment, Medications

The current therapies, whether pharmacological or surgical, are primarily symptomatic with the aim to lower the IOP. If the disease progresses slowly, the initial treatment is pharmacological. In more advanced cases (with a significantly increased IOP not responsive to multiple pharmacological treatments), surgical intervention is needed. It is possible that permanently lowering IOP halts further loss of vision.^10,11

There are several classes of pharmacological agents that lead to a reduction in the level of IOP. In general, these medications work to either increase the outflow of aqueous humor or reduce its production. The surgical procedures used to treat glaucoma focus on the same principles. Trabeculectomy, trabeculectomy with the artificial draining implant, ExPress, canaloplasty, and laser or surgical iridectomy, all facilitate the outflow of aqueous humor. Other modalities such as cryotherapy, cyclophotocoagulation, and endoscopic cyclophotocoagulation focus on cyclodestruction (damage of secretory epithelium of the ciliary body) to decrease the production of aqueous humor.^12–14

If IOP is left untreated, glaucoma can lead to blindness. Unfortunately, despite our best efforts, 10% of people who receive treatment for glaucoma still experience loss of vision. ¹⁵ Thus, there is still a need for advancements in glaucoma therapy, especially with new targets of treatment. A novel uveolymphatic outflow pathway can be a new unifying concept of disease mechanisms and treatment.

Drainage of Aqueous Humor

In the healthy eye, aqueous humor generates an IOP of ∼15 mmHg and is regulated both by its secretion for ciliary body and outflow from the eye. Aqueous humor is drained from the eye by two main pathways: conventional outflow through the TM and Schlemm's canal; and unconventional outflow through the ciliary body through uveal tissue. Both routes are well studied.^16,17

In 2009, Yucel et al. described a third pathway for aqueous humor drainage. This newly discovered route creates a possibility to improve current treatments, improve our understanding of glaucoma, and serve as a target for new treatment strategies. ¹⁸

Lymphatic System of the Eye

While lymphatic vessels have an established role in draining extracellular fluid and solutes, it was considered to be absent in the eye. ¹⁹ Lymphatics are highly permeable to large macromolecules and migrating cells. ²⁰ The system drains extracellular fluids, solutes, and proteins and transport them to lymph nodes and then the blood circulation.^21–23

Before the discovery of specific lymphatic markers, different methods were used to visualize lymphatic tissue in choroid tissue—radioactive, using of electron microscopy, and histological. ²⁴

In 1979, Grüntzig et al. ²⁵ discovered lymphatic pathways in rabbits by introducing a radioactive substance into the aqueous humor and detecting the radioactive signal from superficial cervical lymph nodes. However, Bradbury and Cole ²⁶ described no radioactivity in cats' cervical lymph in a similar study.

In 1988, Krebs and Krebs showed evidence of choroidal lymphatic vessels by using electron microscopy. ²⁷ They described characteristic lack of a continuous external basement lamina and the presence of anchoring filaments.

In 1997, Stefano and Mugnaini used histological examination of animal choroid to show the existence of lymphatic vessels in the choroid. ²⁸ The authors observed that short lymphatic vessels reached the choriocapillaris and proposed that the fluids from the eye is drained directly into the venous system of the eye.

In 2010, Yücel ¹⁸ used two specific lymphatic markers: podoplanin, a transmembrane mucin-type glycoprotein, and lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) to identify lymphatic channels in the human ciliary body using immunofluorescence with a specific D2-40 antibody for podoplanin. Those channels were negative for blood vessel endothelial cell marker CD34, had a distinct lumen, and were surrounded by either discontinuous or no collagen IV-positive basement membrane. The authors confirmed the presence of lymphatic endothelium in human ciliary body by using cryo-immunogold electron microscopy.

To show the lymphatic drainage from the eye, authors injected fluorescent nanospheres to ovine anterior chambers and visualized it in LYVE-1-positive channels 15, 30, and 45 minutes postinjection. Furthermore, intracamerally injected tracer could be detected in several lymph nodes (e.g., cervical, retropharyngeal, and submandibular lymph node). These findings confirm the presence of distinct lymphatic channels in the human ciliary body, and that fluid and solutes flow at least partially through this system.

To measure the percentage of aqueous outflow, Kim et al. ²⁹ reported a model to quantify TM, uveoscleral and lymphatic drainage from the sheep eye. They showed that TM outflow increased rapidly after injection and reached a plateau after 30 minutes, whereas lymphatic drainage increased slowly over a 3-hour period.

Guignier et al. ³⁰ demonstrated in a dynamic in vivo fashion, the existence of lymphatic drainage in the mouse eye. Nanomolecules, marked by technitium-99m, were detected in lymphoscintigraphy by a specially built experimental gamma camera after injection into the anterior chamber of a mouse.

Novel Target for Glaucoma Treatment

The discovery of a new uveolymphatic pathway for aqueous humor drainage identifies a novel and highly relevant target for IOP-lowering therapies in patients suffering from glaucoma. The medications currently in use serve to stimulate fluid drainage through the TM and uveoscleral outflow. It is still not known whether any medication can increase the lymphatic drainage within the eye, but many studies show that lymphatic vessels respond to a wide variety of substances, both biochemical and pharmacological. ³¹

The most promising group are prostaglandins, which are widely prescribed in glaucoma. Prostaglandin analogues increase the aqueous outflow by stimulating PG F2 alpha receptors in the ciliary body and also increase uveoscleral drainage.^32–36

Tam et al. ³⁷ studied the influence of latanoprost on lymphatic drainage. An in vivo nanotracer study showed a greater increase in lymphatic outflow in 11 latanoprost-treated mice versus 11 control animals using hyperspectral imaging methods. The study showed that the lymphatic drainage rate into the submandibular lymph node was 1.23 ± 1.06/h versus 0.30 ± 0.17/h (p < 0.02). This was the first piece of evidence showing that a topical prostaglandin stimulates lymphatic drainage.

Some growth factors are critical for lymphangiogenesis and maintenance of existing lymphatic vessels, ³⁸ for example, VEGF C and its receptor VEGFR3 described by Jeltsch et al. and Wu and Liu.^39,40

To reveal the pathogenic and molecular basis for glaucoma, Thomson et al. measured murine lymphatic defects, which can cause ocular hypertension and glaucoma. They described the genetic disruption of two proteins: Angiopoietin 1 and the orphan receptor TIE2 and also Angiopoietin 2 and the orphan receptor TIE1 in signaling pathways, which increase IOP, cause buphthalmos and other changes seen in glaucoma. They have also linked VEGFR3 and FOXCs to lymphatic disorders and FOXCs to glaucomatic changes. In mice with induced deletion of Angpt1 and Angpt2, the complete absence of LYVE-1-positive ocular lymphatic vessels was observed. Surprisingly, nonocular lymphatic endothelium was shown, although patterning in some organs was abnormal. ⁴¹ This shows that ocular lymphatic drainage is more sensitive to described gene deletion, or lymphatic ocular vessels are a hybrid of both blood and lymphatic vessels as Kitzhatil suggests when describing Schlemm's canal. ⁴²

The lack of Angpt1 and Angpt 2 genes lead to dramatic increase in IOP and glaucoma. This research opens an intriguing new possibility in glaucoma therapy by using ANGPT agonist.

Standard treatment to help drain the aqueous humor from the eye or reduce its production does not solve the main problem of PEX, that is, the accumulation of pathological material in the filtration angle plugging any outflow.

The Use of the New Drainage Pathway in Pseudoexfoliation Glaucoma

Pseudoexfoliation syndrome is an important cause of secondary open-angle glaucoma, usually detected at the stage of significant optic nerve loss. In this form of glaucoma, due to blockage of the pigment grain and exfoliation material, intraocular pressure typically rises in a short time and its fluctuations are very high, leading to rapid damage of the optic nerve.^9,43

Conservative and surgical treatment is more difficult and has a greater risk of failure, because standard glaucoma treatment (increase the aqueous humor drainage from the eye or reduce its production) does not solve the main problem of pseudoexfoliation syndrome (blockage of the TM both by pigment grains released from the iris during mydriasis, and the pseudoexfoliation material). ⁴³ Lymphatics are highly permeable to large macromolecules and drain extracellular fluids, solutes, and proteins. In this way, with the recent discovery of lymphatic vessels in the tissues of the eye, and the demonstration of a new aqueous drainage path, perhaps the use of drugs to increase the flow of this drainage path will be very beneficial for those patients.

However, it is still not known if the lymphatic drainage in glaucoma is decreased or dysfunctional and whether lymphatic stimulation can help in removing the improperly accumulated substances, as is seen in pseudoexfoliation syndrome and in pseudoexfoliation glaucoma.

Conclusion

These studies revealed that lymphatics contribute to the ocular homeostasis and may play important roles in eye disorders. This article reviews current knowledge on lymphatics tissue in the eye and discusses the possibility of lymphatic-targeting therapy.

A novel goal in glaucoma treatment is to decrease IOP by targeting ciliary body lymphatic vessels and stimulate lymphatic outflow. Due to our inability to noninvasively measure lymphatic drainage, and an insufficient understanding of the pathophysiology of glaucomatic changes, further research is required on this topic. However, this new target for glaucoma treatment appears very promising.