The Fibroblast and Glaucoma Surgery

John R. Samples and Paul A. Knepper

Current glaucoma surgery involves an incision in the conjunctiva and the meshwork such as by trabeculectomy or a draining device or a tube placed into the suprachoroidal space. Future possibilities include tubes or shunts placed into other spaces, such as Schlemm’s canal or the suprachoroidal space.

The wound-healing response is an important determinant of the success of glaucoma surgery. It is a multistep process: First, platelets respond, fibrin is deposited, and clotting occurs. Second, inflammation removes the damaged tissues and cells, and platelet-derived growth factor (PDGF) released from α-granules facilitates tissue healing. Third, proliferation occurs, including angiogenesis and collagen deposition. Finally, remodeling occurs, with collagen and extracellular matrix responses.

Fibroblasts are the most common cells in connective tissues. The fibroblast is a type of cell that synthesizes collagen and other components of the extracellular matrix, the main structural framework (stroma) for tissues. Thus, fibroblasts play a critical role in wound healing. They are attracted to wounds by chemicals such as chemokines, cytokines, and inflammatory mediators released at the site of injury. We often tell patients that, under the microscope, these cells look like little snails that migrate to the site of a wound and create a scar; it is always important to point out that the snails that are depositing scar-making material are “made out of you,” that is, of their own cells.

Fibroblasts likely exist in a variety of subtypes and may arrive at the wound in a pluripotent state. At present, there is limited information on how fibroblasts differentiate in the wound-healing process. Myofibroblasts, a fibroblast subtype, are found in fibrovascular membranes in the eye. They are widely distributed in some normal ocular tissues, in which they possibly evolved from quiescent fibroblasts.1 The glaucoma surgeon may be vexed by the adherent membranes and peripheral anterior synechiae they instigate. With all of the glaucoma devices, irrespective of which spaces they connect, it is important that the devices be biocompatible to prevent them from attracting the cells that lead to scar formation. Strategies to enhance biocompatibility include the use of heparin coating, biocompatible hydrogels, and a variety of other bio-friendly materials.

The wounding response specifically associated with the placement of a stent or device through tissue and the subsequent release of small molecules and peptides is very complex and involves mediators in varying amounts. Laser treatment to almost any ocular tissue likely also evokes these wound-healing responses, as we have shown with interleukin-1β and tumor necrosis factor-α (TNF-α).² The notion that these cytokines are ubiquitous and seem to lack specificity is somewhat counterintuitive, as they likely have specific roles in each situation. These cytokines in the trabecular meshwork induce mitosis and alter the trabecular outflow pathways in a positive manner by secondary effects on matrix metalloproteinases. Most wound and inflammatory responses also evoke a macrophage response. The macrophage-like response of trabecular cells is an often-overlooked quality. Fibroblasts interact with macrophages and may be removed by them. Although stem cells have successfully been used to regulate outflow in the meshwork, the therapeutic effect of their phagocytic response, once the cells have settled on trabecular beams, is yet to be fully considered.³ Moreover, stimulation of trabecular cell phagocytic activity may be a means of modifying fibroblast-driven scarring in the eye.

Not much is known about the subtypes of fibroblasts and scar-forming cells in the eye and the degree to which a fibroblast derived from Tenon’s capsule is different from a fibroblast from the conjunctiva. It is likely that these cells vary in their ability to respond to a variety of stimuli and have varying degrees of specialization.

Variation in response to chemotherapy among the cell types seems likely. Wound-healing responses in the eye have traditionally been addressed by antimetabolites such as 5-fluorouracil and mitomycin C. Attempts to manipulate growth factors associated with wound healing have not yet produced success. Yet there are many other molecular mechanisms that can modulate a response to wounding in glaucoma surgical therapy. A multitude of strategies now exist to address fibroblast behavior, including RNA interference therapy; antibodies against growth factors, such as new-generation anti–vascular endothelial growth factor (VEGF), anti-PDGF, and other anti–growth factor treatments; as well as numerous small molecule inhibitors. A better understanding of fibroblasts will likely take us far beyond the present use of corticosteroids to modulate wound healing in our surgeries.

Hyaluronic acid (HA) is just one of many key factors in the eye that may affect fibroblast motility, proliferation and cellular differentiation. CD44, the principal receptor for HA, is abnormal in its soluble form in the glaucoma eye and may offer one of many explanations of why scar formation and fibroblast proliferation may be abnormal in the glaucomatous eye.⁴ Notably, the aqueous humor of primary open-angle glaucoma (POAG) has a higher content of low molecular weight HA (Fig. 20.1). Low molecular weight HA signals through the innate immune receptor complex CD44-Toll-4-MD-2 to cause an upregulated proinflammatory response. Significantly, low molecular weight HA effects can be blocked by the administration of naloxone, a toll-like receptor 4 antagonist.⁵ The use of naloxone or other antagonist of the innate immune system may be a future consideration to prevent excessive wound-healing responses and improve outcomes of glaucoma surgery.

There are additional approaches to controlling fibroblasts. We are working on specific mixtures of collagen gels, with or without additional small molecules to repel fibroblasts from almost any active wound site (in development, Eyegenetix, Columbia, SC). Sustained release reservoirs embedded in the sclera may gradually elute wound-modifying peptides, antibodies, or small molecules such as naloxone. These delivery devices have wide applicability to glaucoma and may also be used to administer neuroprotectants that are otherwise hard to deliver due to molecule size or other pharmacological properties (in development, Refocus, Fort Worth, TX). A better understanding of the interaction of lymphatic channels and fibroblast interactions may yield surprising results because lymphatics may play an important role in conjunctival filtration. Eventually these approaches and others are likely to make the present concerns about bleb failure and scarring at other wound sites a matter of historical interest. New approaches and methods for modulating fibroblasts appear annually at the American Society for Cell Biology conference. Additional insights may help with understanding healing elsewhere, including in the retina, vitreous, and cornea.