Ординатура / Офтальмология / Английские материалы / Glaucoma Surgery_Bettin, Khaw_2012

is at a low BAK exposure rate. Nevertheless, in healthy volunteers Ishibashi et al. [27] demonstrated that preserved timolol caused significantly higher tear film instability and disruption of corneal barrier function than preservative-free timolol. Very similar results were found in healthy volunteers when comparing preservative-free and BAK-containing carteolol [28], with a significantly lower tear break-up time in eyes exposed to BAK. When considering that both studies were conducted in young subjects with a fully normal ocular surface, these results may help better understand the high rate of dry eye symptoms and signs in glaucomatous patients who combine a long duration of treatment, a higher number of medications, and an increased prevalence of impaired ocular surface. Another prospective study was conducted in healthy volunteers evaluating various concentrations of BAK in tear substitutes given eight times a day for 7 days. Even a low concentration of BAK induced goblet cell loss and increased the cytoplasmic/nucleus ratio, two characteristics of dry eye disease [29]. A subsequent study prospectively assigned 132 subjects to one or two BAK-containing eye drops, BAK twice daily, or no treatment. All groups receiving BAK showed significantly decreased Schirmer test values compared with subjects not receiving therapy [30].

Another prospective study using impression cytology and the technique of in vivo confocal microscopy (IVCM), performed in 27 patients before and after 6 months of therapy, showed significant differences between eyes treated with a BAK-containing vs. BAK-free beta-blocker. The IVCM analysis showed 61 and 17% of goblet cell density reduction from baseline, in these two groups, respectively, and using Nelson’s score, the grading of impression cytology parameters was significantly higher in the BAK group. Moreover, these conjunctival epithelial changes were observed only after 6 months of therapy suggesting that preservatives exert their toxic effects in a rather short period of time at least at a subclinical level [31].

Additionally, several comparative observational surveys have in recent years compared the ocular surface parameters in patients receiving preserved and unpreserved eyedrops. A total of 84 patients were evaluated using the sophisticated method of IVCM with respect to their treatment. Significant differences were found between groups on topical BAK-containing IOP-lowering eyedrops, namely preserved betablocker, preserved prostaglandin, fixed or unfixed combinations of both drugs, and a preservative-free beta-blocker group. In particular, the density of superficial epithelial cells and the number of sub-basal nerves were reduced in all preservative-containing groups. In contrast, the density of basal epithelial cells, stromal keratocyte activation, and bead-like nerve shaping were higher in the BAK groups than the control and preservative-free groups [32]. Most importantly, this study pointed out a significantly decreased corneal sensitivity of about 10–30% in all preserved groups compared to control or unpreserved eyedrops, which further demonstrates that BAK may cause major ocular surface impairment despite an apparently good tolerance.

At a larger scale, in an epidemiological survey conducted in 4,107 glaucoma patients to assess the effects of preserved and preservative-free eye drops on ocular

surface-related symptoms and signs, all were significantly more prevalent (about twice as much) in patients using preserved drops compared with those on preservative-free treatment [6]. Likewise, a similarly conducted even larger European survey also demonstrated that the incidence of ocular signs and symptoms was higher in patients receiving preserved eye drops [7].

Possible Impact of Benzalkonium Chloride in Deep Ocular Structures

From all the above-mentioned studies and surveys in humans, as well as in many reliable and consistent experimental in vitro and animal models [5], broad consensus has been reached that BAK causes a variety of ocular surface changes, including dry eye as well as allergic and immunoinflammatory reactions, with overexpression and/ or synthesis of class II antigens, adhesion molecules, chemokines, chemokine receptors, interleukins, or cell death markers and mediators, destruction of goblet cells and inflammatory cell infiltration. Additionally, the substantia propria of the conjunctiva has been shown to be infiltrated by inflammatory cells and fibroblasts [3, 4, 11, 12]. Other reports have hypothesized that BAK plays a role in cataract development [33] and the higher incidence of angiographic cystoid macular edema after cataract surgery. Based on the findings that the preservative causes increased synthesis of proinflammatory mediators and intensified postoperative inflammation, the term ‘pseudophakic preservative maculopathy’ was even proposed [34].

Little is known on BAK pharmacokinetics, but this compound was shown to accumulate in the conjunctiva since only a single drop of BAK could cause measurable amounts in the conjunctiva up to 7 days after instillation [35]. It is highly likely that BAK accumulates in deep tissues after prolonged administration. In addition, the impact of inflammatory and toxic reactions in the ocular surface following chronic treatments with BAK-containing eyedrops over the long-term, and clearly extending to deep conjunctiva and conjunctiva-associated lymphoid tissues, remains to be elucidated. Therefore, the consequences of high rates of cell death and inflammatory signals on deeper but still very close structures, such as Tenon’s capsule and subconjunctival space and even the trabecular meshwork, are most likely and may participate in postoperative inflammation and fibrosis, with subsequent IOP rise.

Increased Rate of Glaucoma Surgery Failure

The failure of glaucoma surgery in most cases originates from a fibroblastic conjunctival response at the bleb level in the early postoperative months. An excessive wound healing process characterized by inflammation and fibroblast proliferation and extracellular matrix deposition results in blockade of aqueous outflow in the subconjunctival space (fig. 4). Increased postoperative fibrosis may be enhanced in populations

Fig. 4. Association of flat and encapsulated blebs in a refractory glaucoma having undergone three failed trabeculectomies.

of patients or conditions known to be at higher risk of surgical failure, but several reports have pointed out the role played by preoperative inflammation of the conjunctiva caused by antiglaucomatous drugs used over the long term [36].

The success rate of trabeculectomy was thus found negatively correlated with long-term topical antiglaucoma therapy [37]. Broadway and colleagues further studied the effect of different long-term antiglaucoma treatments on conjunctival changes and the result of glaucoma filtration surgery, and found a significant relationship between the number of drugs used and duration of treatment, inflammatory cell and fibroblast infiltration in the conjunctiva, and the risk of failure of filtering surgery [3, 4]. Although these results were collected at a time when new more powerful drugs had not yet been developed, approximately 50% of glaucomatous patients nowadays require two or more drugs to control IOP, and are most often operated on after prolonged medical treatment with multiple drugs. Despite significant progress made in glaucoma therapy, filtration surgery success rates thus do not appear to have been improved in the recent years [38]. Indeed, the same mechanisms are still involved, as very recently Helin et al. [39] consistently found that densities of inflammatory cells, namely T-lymphocytes, T-helper lymphocytes, T-cytotoxic lymphocytes, B cells, plasma cells, and macrophages, measured at time of surgery, were significantly higher in patients in whom surgery failed than those with surgical success, during a long follow-up of 2.5 years.

Preoperative Assessment of Failure Risk

After evaluation of the risk factors for failure based on demographic criteria, ocular history, current therapy, a detailed preoperative examination of the patient is also important. The conjunctiva, the subconjunctival tissues, the vascularity and signs of inflammation should be analyzed precisely. Examination of the fellow eye is equally important and particularly if this eye had already undergone filtering surgery. The

evolution of this eye in terms of IOP control and bleb appearance may often be the best witness to judge the efficacy of a previous wound healing modulation therapy. However, there is no clear prognostic marker for surgery success or failure. Broadway et al. [40] used conjunctival biopsies to assess preoperative inflammation, and found fluorometholone efficient one month prior to surgery to decrease inflammatory infiltrates and improve surgical outcome, but this technique remains difficult to propose on a routine basis. Baudouin et al. [41] used impression cytology specimens to measure inflammation in the conjunctival epithelium and also found that a preservativefree nonsteroidal anti-inflammatory agent could decrease inflammation prior to surgery but no correlation was made with success rates. Nevertheless, both preoperative anti-inflammatory strategies were found similarly effective in improving the prognosis of glaucoma surgery [42]. Most interestingly, using impression cytology, a higher MUC5AC expression and a lower HLA-DR expression were observed in patients with further successful glaucoma surgeries than in failures. Surface marker expression could therefore become a predictive factor of successful filtering surgery [24].

Postoperative Changes in the Ocular Surface

Several studies have characterized the histopathology of the conjunctiva at the bleb level. The conjunctival epithelium was described as irregular in thickness, containing numerous intraepithelial microcysts, especially in blebs with adjunctive MMC [43]. Conjunctival epithelial spongiosis associated with inflammatory changes was reported [44], with focally acantholytic epithelial cells separated by clear spaces [45]. The epithelial intercellular spaces observed in the latter study provided evidence that aqueous humor could move transconjunctivally. Microcysts are assumed to be a positive predictive factor for further bleb efficacy [46–48]. They are described as being channels for the passage of aqueous humor [48]. Using impression cytology with an immunofluorescence technique under a confocal microscope [49], presence of numerous goblet cells with typical morphological features but low or no content of the soluble mucin MUC5AC was found at the surface of the functioning blebs (fig. 5), contrasting with highly and homogeneously stained goblet cells outside the limit of the blebs. These ‘empty’ cells seemed to correspond to the microcysts observed clinically or with IVCM, and they were observed only in very low numbers at the surface of nonfunctioning blebs. The absence of goblet cells and/or microcysts thus appeared as factors of poor prognosis after filtering surgery.

Soluble mucins stained with anti-MUC5AC gene-related proteins are highly hydrophilic glycoproteins. A goblet cell measures approximately 20 μm, whereas a microcyst observed in vivo measures between 20 and 150 μm [50]. Some microcysts might directly correspond to goblet cells filled with aqueous humor or highly hydrated, with

Fig. 5. Impression cytology at the level of a functional bleb showing numerous goblet cells with weak irregular mucin staining and most likely transepithelial aqueous humor.

therefore lower mucin content. Larger microcysts observed with IVCM could result from the confluence of adjacent empty or degenerating goblet cells, resulting in large clear spaces that could no longer be identified as individual mucus-secreting cells (fig. 6a).

In the case of mitomycin C (MMC) use, which is commonly administered during filtration surgery to enhance the success of the procedure, thin extremely cystic blebs are often observed. Using impression cytology, this type of bleb showed results similar to those found for functioning blebs, with many large microcysts and confluent nonsecreting goblet cells [49]; however, numerous inflammatory cells were also observed, at higher densities than in non-MMC blebs. These patients did not have inflammatory or infectious signs observed biomicroscopically. A hypothesis is that the fragility of the bleb makes it more sensitive to microtraumatisms and infectious risks.

Postoperative Evaluation of the Bleb

The long-term success of these filtering procedures is mainly dependent on the development of a functioning bleb. The formation and the maintenance of this functioning bleb, with regard to wound healing and conjunctival scarring, are therefore of primary importance. Picht and Grehn [51] classified the developing filtering bleb, showing that favorable bleb development was characterized by microcysts of the conjunctiva, paucity of vessels, diffuse bleb and moderate elevation of the bleb. In contrast, they observed that unfavorable bleb development was characterized by increased vascularization, ‘corkscrew vessels’, encapsulation of the bleb and high-domed appearance. Because in some cases the appearance of the bleb is not

correlated to IOP and because the reason of failure is often unclear, some authors have looked for new in vivo evaluation techniques in order to understand bleb failure mechanisms. Ultrasound biomicroscopy and IVCM (fig. 6) have been used for evaluating bleb functionality [49, 50, 52, 53]. Filtering blebs usually show a normal conjunctival epithelium with numerous microcysts and a subepithelial tissue arranged loosely and hyporeflective, with a high number of optically clear spaces. In contrast, nonfiltering blebs usually show none or very few microcysts (fig. 6a) and a subepithelial tissue hyperreflective with dense collagenous connective tissue (fig. 6b) and sometimes vessels. The presence and the number of microcysts are assumed to be a positive predictive factor for functioning blebs. IVCM examination of filtering blebs seemed to confirm these microcysts as channels for the transconjunctival passage of aqueous humor [48, 49].

a

b

c

Fig. 7. OCT imaging of postoperative blebs. a Functional, mildly elevated with heterogeneous material. b Nonfunctional flat bleb. c Nonfunctional encapsulated bleb.

Although not sufficient for providing histological-like images of the conjunctiva, OCT techniques adapted to the anterior segment may also provide valuable information at the tissue level, showing flat or encapsulated blebs in a totally noninvasive way (fig. 7). This technique is therefore very useful for the follow-up of glaucoma surgery, offering numerous criteria for evaluating bleb functioning, like length and height of the internal fluid-filled cavity, maximum and minimum bleb wall thickness, total bleb height, volumes of the internal fluid-filled cavity and hyporeflective area, and number of microcysts [53, 54]. Nakano et al. [55] even found that heterogeneity of bleb walls 2 weeks after surgery might predict the outcome of trabeculectomy at 6 months.

Conclusion

In conclusion, the ocular surface, especially the conjunctiva, is a major component of glaucoma surgery, both influenced by preoperative topical treatments and interacting with bleb function and scarring formation. Long-term use of topical treatments prior to surgery deeply modifies the ocular surface, in particular the tear film, conjunctival epithelium and subepithelial space, even on a subclinical mode. Chronic, low-grade inflammatory stimulation increases over time and directly enhances the wound healing process when surgery is performed, often resulting in enhanced collagen deposition and fibroblastic response. As the preservative is at least in part responsible for such ocular surface changes, this compound should be avoided or reduced as much as possible in patients with severe glaucoma or requiring multiple therapy, most susceptible to undergo surgery, and those with clinically impaired ocular surface, like dry eye, allergic reactions, or blepharitis. BAK-free compounds are progressively developing owing to a better awareness of the toxic potential of preservatives; it will therefore be of interest to evaluate in the future whether inflammatory reactions are effectively reduced and further surgery is more efficient.

References