INDICATIONS, OUTCOMES, AND COMPLICATIONS – VITAL DYES IN CHROMOVITRECTOMY

chromovitrectomy. IfCG is produced in a synthetic manner without sodium iodine, and high-dose topical or intraocular iodine has been shown to induce severe corneal and retinal damage. As a second difference, iodine-free IfCG is dissolved in 5% glucose, giving an isoosmotic solution of around 310 mmol/kg. In contrast, ICG is usually dissolved in water before further dilution in balanced salt solution (BSS), which produces a hypo-osmolar solution. Indeed, osmolarity changes at the vitreoretinal interface or subretinal space induced by various types of solutions, including ICG dye, have been shown to promote remarkable histological toxicity to retinal cells.2

Several recent clinical investigations have shown positive results with IfCG application without signs of retinal toxicity. In the clinical setting, IfCG-assisted ILM peeling for macular hole surgery demonstrated high closure rates of macular holes in over 90% of eyes and enhanced visual acuity. Conversely, one recent investigation showed the presence of remnants of footplates from Müller or glial cells and neural or ganglion cells respectively after analysis of the ILM peeled with IfCG during macular hole surgery. Therefore, despite that IfCG and ICG may facilitate ILM removal, unwanted retinal alterations may be generated, such as neurosensory RPE and visual field defects. In summary, IfCG with a presumably safer profile may represent an outstanding alternative to ICG use for ILM peeling during chromovitrectomy in humans. IfCG at a concentration of 0.5 mg/ml results in adequate ILM identification and less toxic effects.10

TRYPAN BLUE

TB has been proposed to stain preretinal tissues such as ILM and ERM in chromovitrectomy (Figure 48.2). Although the use of TB for ILM staining may also be feasible, in our experience ILM visualization is much more difficult than with the ICGor BriB-guided procedure. Currently, state-of-the-art TB usage recommends blue dye application mainly for ERM staining. TB exhibits outstanding affinity for ERM because of the strong presence of dead glial cells within those membranes. TB staining of the ERM may minimize mechanical trauma to the retina during ERM removal and should allow the recognition of the whole extent of the ERM, which has led to good surgical outcomes and minimization of the recurrence of ERM in several clinical studies.11

The usefulness of intracameral or intravitreal injection of TB to highlight vitreous gel has been recently proposed. The blue dye in various doses may enhance the ability to detect both the prolapsed vitreous to the anterior chamber and the posterior vitreous remaining in the vitre-

ous cavity. However, in one comparative analysis TB demonstrated an inferior ability to stain the vitreous in comparison to TA and FS.12

TB in vitreoretinal surgery was initially proposed for injection after an air–fluid exchange, allowing for better dye dropping on to the retinal surface. To enhance dye penetration on to retinal surface in an air–fluid exchange, TB may be mixed with glucose at higher than 5%, for instance 10% or 25%, thereby creating a “heavy” TB, denser than water. However, high glucose concentrations should be avoided; for instance, glucose50%hasahighlytoxicosmolarityof2020mOsm/l.Experimental injection of 0.05 ml of a 1000 mOsm solution caused rapid whitening of the posterior retina followed by the development of a large detachment and permanent retinal degeneration. Indeed, osmolarity should be taken into consideration in planning the amount and location of any vitreous injection of dyes and drugs.

Consecutive clinical studies have revealed that TB exerts little or no toxic effect on the retina. For ERM surgery, TB caused no RPE defects or signs of retinal toxicity in most studies in the literature until now. Histopathological analysis of excised ERM showed no retinal cells on the retina side of the ERM or signs of apoptosis, while functional analysis by multifocal ERG also showed no signs of retinal toxicity. Comparative studies evaluated the anatomical and visual outcomes after vitrectomy and ILM peeling for treatment of patients with macular hole using ICG or TB. Their success rate for MH closure was the same; however, visual recovery has been better in the TB group.

In animal experiments, TB has demonstrated a reasonably good retinal biocompatibility in most investigations at the concentration proposed for intravitreal application from 0.06% to 0.15%. Grisanti et al. used fresh hemisected porcine eyes and applied TB 0.15% to the posterior pole after vitreous removal followed by illumination with a standard surgical light probe and source at maximum power for 10 minutes; the procedure caused no histologically detectable damage. These results imply that TB at concentrations of 0.15% or lower represents a safe adjuvant in vitreoretinal surgery.13

Various laboratory studies have evaluated the retina biocompatibility of TB alone or in comparison to ICG. In cultured human RPE and Müller cells, some authors have reported that exposure to TB at concentrations up to 0.3% in vitro induced no toxic effect.14 In accordance with these studies, recent investigations revealed that TB is toxic to cultured RPE cells at concentrations higher than 0.5%. However, ICG may cause more toxicity to the retina, especially to human RPE cell cultures, than TB, independent of any phototoxic potentiating effect of fiberoptic light or solvent toxicity. Our research group showed that subretinal injection of 0.05% ICG results in more substantial retinal damage than that associated with subretinal injection of 0.15% TB.15

Figure 48.2  In porcine eyes, the internal limiting membrane is fine-colored with trypan blue. However, intraoperatively the blue azo agent stains rather the glial epiretinal membranes.

PATENT BLUE

PB has been certified for capsule staining during cataract surgery at a concentration of 0.24%. Animal studies and preliminary clinical data demonstrate moderate affinity of PB for ERM and vitreous, but a poor affinity for the ILM. Our recent clinical data revealed PB as an appropriate vital dye for staining the glial ERM from various causes in a similar manner to TB.16

Toxicology studies revealed conflicting data regarding retinal toxicity of PB. In one study PB was found to induce only mild and reversible retinal toxicity, whereas RPE cells exposed in vitro to PB showed no toxicity.15 Our rabbit subretinal toxicity model demonstrated clinically on FA examination no RPE defects in positions related to the subretinal PB injection in rabbits as seen in the control BSS group. Histologically, subretinal injection of PB resulted only in mild ultrastructural retinal damage during follow-up; the histological damage induced by TB and ICG was more severe than that by PB. The exact safe dosage for intravitreal PB injection remains uncertain.15

BRILLIANT BLUE

In the last few years, the stain BriB has been introduced as a surgical adjuvant for chromovitrectomy and anterior lens capsule staining. In order to enable anterior capsule visualization for capsulorrhexis, BriB

may be applied in an iso-osmolar solution at a concentration of 0.25 mg/ ml or higher. In humans, BriB also produced adequate ILM staining when using an iso-osmolar solution of 0.25 mg/ml during macular hole surgery, while no signs of damage were noticed. Cervera et al. showed similar outcomes with good ILM staining and clinical results and no signs of toxicity in multifocal ERG. In brief, BriB represents a great alternative for ICG and IfCG in chromovitrectomy due to its suitable affinity for ILM.17

Experimental investigation in rat eyes revealed no damage of corneal endothelial cells in the long term after BriB exposure. In rat and primate eyes, no significant retinal pathologic changes were observed with light microscopy and electron microscopy after low-dose BriB injection; there was also no reduction in the amplitude of the ERG waves.18 Our workgroup performed experiments with intravitreal injection of two doses of BriB and five other dyes in rabbit eyes. We found that BriB at the lower dose induced no major retinal changes, while at the higher dose some alterations of the photoreceptors were encountered. Collectively, these data imply that BriB represents an adequate and safe staining agent for clinical evaluation in humans.2

SODIUM FLUORESCeIN (SF)

SF has been found to be highly safe for fundus angiography at concentrations of 5–25%, even when leakage through the retina occurred. Because of its hydrophilic properties, SF is highly absorbed by the vitreous gel, as most available data indicate that intravitreal 0.20% SF dye improves the visualization of clear vitreous fibers through a green staining during chromovitrectomy. In addition, there are so far no sideeffects reported in regard to the use of SF in chromovitrectomy. While so far the main indication of SF in chromovitrectomy remains the vitreous staining, further clinical experience should determine whether there is any role for SF application in the visualization of preretinal membranes.12

TRIAMCINOLONE ACETONIDE

Various clinical investigations have shown that TA produces the best vitreous visibility in comparison to other stains. The crystals of the crystalline steroid adhere avidly to the acellular tissue, thereby enabling a clear contrast between the empty vitreous cavity compared to areas where the vitreous fibers are still present (Figure 48.3). The currently reported surgical technique for TA application consists of a simple injection of the agent into the vitreous cavity directed toward the area of visualization. In addition to its effect on vitreous visualization, a TA

injection during vitrectomy has been proposed to prevent fibrin reaction and postoperative proliferative vitreoretinopathy. Following the initial report, a list of consecutive clinical studies showed the efficacy of TA for staining the transparent vitreous, whereas anatomic or functional signs of complications have rarely been observed. A recent multicenter controlled clinical trial performed in Japan showed decreased risk of postoperative retinal detachment but increased need for postoperative antiglaucoma eye drops with intravitreal TA application.19

In the clinical setting, TA arose as an alternative stain for ILM peeling, since the white specks and crystals may deposit over the ILM, thereby facilitating ILM removal. Afterward, a few clinicopathologic studies disclosed the presence of ILM ultrastructures after TA-assisted peeling. Clinical studies have shown that TA-assisted ILM removal enables good clinical results, with no adverse effects noticeable in the follow-up evaluation period. However, recent data raised concern regarding the use of TA for MH surgery. Crystals of TA have been detected up to 40 days postsurgery with chromovitrectomy for MH surgery. Some authors suggest that postoperative residual TA could diminish the healing process necessary for MH closure.20 Currently available preser- vative-free TA may be better for vitreous staining as a deposit to enhance visualization.

Contradicting results from a considerable number of animal experiments have recently been released regarding retinal toxicity after intravitreal TA injection. Various studies of intravitreal TA injection at concentrations varying from 4 to 30 mg TA have normal morphologic and ERG retinal findings up to 6 months’ follow-up. In contrast to those reports, others examined escalating doses from 0.5 to 20 mg of suspended preservative-free TA in rabbits, and found prominent retinal damage manifested by destruction of photoreceptor outer segments and RPE/photoreceptor interdigitation at a dosage of 4 mg or higher.

There is still uncertainty as to whether TA itself or the vehicle plays the more significant role in retinal damage. In order to elucidate this question, one study compared a group of TA with vehicle to another with preservative-free TA and demonstrated severe damage to photoreceptor only in the group with vehicle.21 In contrast to those outcomes, other investigators revealed a safer profile of the vehicle only: they described normal retinal structure after intravitreal injection of vehicle only in animals. Our investigation disclosed disturbance to photoreceptor segments after subretinal injection of preservative-free TA; however, no clinical abnormality on fundoscopy or fluorescein angiography was detected.20 Future studies should clarify if the vehicle alone, TA alone, or both chemicals produce retinal damage in humans.

Consecutive laboratory studies examined the effects of TA on various types of retinal cells including ARPE19, human glial cells, neurosensory cells, ganglion cells, and choroidal fibroblasts.21 A few studies have indicated that TA may cause severe toxicity to chorioretinal cells. In contrast, some investigators proposed a safe in vitro profile of the white steroid to RPE cells. These conflicting results may highlight the importance of many variables in chemical-induced retina toxicity in the clinical setting.

Figure 48.3  The steroid triamcinolone acetonide stains the acellular vitreous strongly and thereby ensures that the whole posterior vitreous cortex is removed during vitreoretinal surgery.

OPERATIVE TECHNIQUES AND

MODIFICATIONS

DYE INJECTION

Several different surgical approaches have been used to inject vital dyes in the vitreous cavity in chromovitrectomy. One technique has been named the “dry method” or “air-filled technique.” By either name, this technique consists of removing the liquid in the vitreous cavity by a fluid–air exchange before dye injection. Although the technique has the advantage of concentrating the dye in the posterior pole and avoiding contact at the posterior capsule of the lens, it may expose the retinal surface to the injected concentration of the dye. The second proposed technique to inject dyes is called the “wet method” or “fluid-filled technique.” In this method, the intravitreal fluid is left inside the vitreous cavity while the surgeon infuses the dye. The amount of dye in