METHODS

Participants

Enrolment was conducted between 15 March 2005 and 1 February 2007. During this period, 173 patients were referred to the Department of Ophthalmology at Glostrup Hospital, University of Copenhagen for MHS. Patients were examined to confirm the diagnosis of full-thickness stage 2 or 3 idiopathic macular hole according to the classification by Gass5455 and were considered for inclusion according to the inclusion criteria (idiopathic macular hole stage 2 or 3, symptom duration ⩽12 months, visual acuity ⩾34 ETDRS letters, intraocular pressure ⩽23 mm Hg, informed consent) and exclusion criteria (ophthalmoscopically visible epiretinal fibrosis, previous ocular surgery (except cataract surgery), any eye- or systemic disease affecting visual function). Eighty-nine eligible patients were scheduled for cataract surgery with subsequent MHS after 4–6 weeks. At the baseline examinations, the day before MHS, 78 patients still fulfilled the inclusion criteria and subsequently were included in the study for randomised MHS. Approval was obtained from the local Committee on Biomedical Research Ethics, and informed consent according to the tenets of the Declaration of Helsinki was obtained from all subjects before entering the study.

Interventions

Of the 89 eligible patients, all phakic patients had standard phacoemulsification surgery with intraocular lens implantation prior to MHS. Postoperatively anti-inflammatory treatment with dexamethasone eye-drops was administered six times daily for 2 weeks and then tapered slowly with one drop per week until MHS after 4–6 weeks. The cataract operations were performed by two experienced surgeons.

Baseline examinations were performed the day before MHS with scheduled follow-ups 3, 6 and 12 months after surgery. Examinations included: registration of symptom duration, best-corrected ETDRS visual acuity (BCVA) with an initial testing distance of 4 m, slit-lamp- and stereoscopic biomicroscopy, intraocular pressure (IOP) measurement, fundus photography, visual-field testing using 30-2 automated perimetry (Humphrey Field Analyzer 750, Humphrey Instruments, San Leandro, California), OCT classification of the macular hole according to Gass5455 at baseline visits and evaluation of hole closure at follow-up visits (Stratus model 3000; Carl Zeiss Meditec, Humphrey Division, Dublin, California). At baseline and 12 months’ follow-up, registration of fundus autofluorescence was performed (Heidelberg Retina Angiograph II, Heidelberg Engineering GmbH, Heidelberg, Germany).

The surgical intervention for macular hole consisted of (1) vitrectomy alone (non-peeling) or (2) vitrectomy plus ILM peeling with either Indocyanine Green- (ICG) or (3) Trypan Blue (TB) staining according to randomisation. The standard three-port pars plana vitrectomy included verified separation of the posterior vitreous cortex with direct aspiration over the posterior pole. A careful examination of the retinal periphery was then performed after indentation, and any iatrogenic retinal breaks found were treated by laser- or cryopexy. No touch of the retinal surface was performed in the non-peeling group (25 patients). In the ICG-assisted ILM peeling group (35 patients), 25 mg/ml of ICG (ICG pulsion, PULSION Medical Systems, Münich, Germany) was dissolved in 5 ml of isotonic glucose and diluted with 45 ml of balanced saline solution (BSS) to obtain a final isotonic solution of 0.05% (0.5 mg/ml) ICG. In the TB peeling group (18 patients), 0.1 ml of 0.15% TB (Membrane Blue, DORC international, Zuitland, The Netherlands) was used. The ICG solution was applied in a BSS filled eye, and TB was applied in an air-filled eye. Both agents were washed out after an exposure time of 15 s. In cases with insufficient staining after TB exposure, a second application of 15 s was allowed. Peeling of the ILM was performed to cover an area of approximately two disc diameters around the macular hole. Finally, a complete fluid–air exchange and intravitreal tamponade with 10–15% perflouropropane (C₃F₈) was performed. After surgery, patients were instructed to keep a face-down position for at least 10 h per day for 5 days. Patients were provided a registration sheet for help and documentation of face-down time. Postoperatively a combination of dexamethasone- and chloramphenicol eye-drops was administered four times daily for 3 weeks. In cases without primary hole closure, subsequent macular hole surgery was offered as soon as possible with additional 0.05% ICG-assisted ILM peeling and gas tamponade. Breaking the randomisation code in these cases was not necessary because the surgeons were not masked to the intervention.

Outcomes

Primary outcome measure was closure of the macular hole 3 and 12 months after surgery, defined by complete adaptation of the hole margins as assessed by OCT. The secondary outcome measure was best-corrected visual acuity (BCVA) 12 months after anatomical successful surgery. Other key secondary outcome measures were visual-field defects after 12 months as assessed by 30-2 automated perimetry, retinal pigment epitheliopathy (atrophy or hypertrophy) as assessed qualitatively as white-yellow or reddish lesions on fundus photography or hyper-fluorescent areas on fundus autofluorescence images, and optic nerve fibre layer dehiscence defined as arcuate retinal stria running along the optic nerve fibres in the macular region. Intraoperative and postoperative complications were systematically registered.

All examinations were performed according to prospectively defined standard operating procedures by examiners who were masked to allocation status.

Sample size

The trial was initially designed as a two-arm interventional study comparing anatomical and functional results after vitrectomy alone (non-peeling) and vitrectomy plus ICG-assisted ILM peeling for stage 2 and 3 idiopathic macular hole without any visible epiretinal fibrosis. A standard power calculation was performed in order to calculate sample sizes based on a priori assumptions of an anatomical success of 85% after vitrectomy alone171819 and of 100% after vitrectomy plus ICG-assisted ILM peeling.1718192022232425 With a power of 90%, it was calculated that a sample size of 40 eyes per surgical arm (N = 80) was required in order to detect a difference at the 5% significance level.

Participants were to be followed for 12 months after anatomical successful MHS, and when all participants had reached this point, the randomisation code would be broken and statistical analyses performed. An interim analysis was planned when half of the patients had been enrolled, and in the event of significant differences in favour of one of the interventions in anatomical or functional outcome, the inferior arm would be terminated and a third surgical arm with TB-assisted ILM peeling would be introduced. In this case, TB should serve as the presumed non-toxic control dye.

Randomisation

Generation of the random allocation sequence was performed based on a list of random numbers in blocks of four, with eyes stratified according to macular hole size in stage 2 and 3. The two highest numbers allocated patients to MHS with vitrectomy alone and lowest numbers allocated patients to MHS with ICG-assisted ILM peeling. Generation of the allocations list was performed by an independent person in the department, and the list was kept in a locker at the operating room. Final assignment to intervention was performed on the day of surgery by the assisting operating nurse as consecutive patients on the allocation list. The day of operation was decided by an independent visitation office to avoid allocation bias.

Based on the interim analysis performed on 15 May 2006 after enrolment of 40 patients, we concluded that continuing the study with the planned 80 patients randomised 1:1 to peeling and non-peeling was unethical because the non-peeled patients with stage 3 macular holes had a significantly increased risk of lack of primary hole closure. According to our stopping rules, the stage 3 vitrectomy (non-peeling) arm was terminated, and from 16 May 2006 stage 3 holes were randomised 2:1 to surgery with either TB-assisted ILM peeling or ICG-assisted ILM peeling. A new allocation list for stage 3 holes was produced for this purpose. The outcomes after stage 2 MHS were not significantly different at this time, and no changes in the study protocol for stage 2 macular holes were made.

Masking

Investigators and patients were masked to intervention, and only the surgeons were aware of the surgical method used. All examinations through baseline and follow-up visits (BCVA at 3 months, OCT, fundus photography, automated perimetry, fundus autofluorescence imaging) were performed by to examiners (UCC and KK) who were masked to the allocation status. Assessment of 6 and 12 months’ BCVA was performed by a certified masked person (BS) in case accidental demasking had occurred during the funduscopic or photographic evaluations 3 months postoperatively. Evaluation of visual fields, fundus photographs and fundus autofluorescence images was performed by UCC before breaking the randomisation code.

Statistical methods

Comparisons of baseline characteristics among the three groups were performed to confirm successful randomisation. Statistical analyses of rates and proportions between the groups were made for primary and secondary outcomes. According to the study design, only comparisons between the non-peeling group and the ICG peeling group were performed when analysing the overall results and the results for stage 2 holes. All three treatment groups were compared in the analysis of stage 3 holes. Associations between categorical variables were analysed using the χ²- or Fischer exact test. Continuous data were analysed using the one-way analysis of variance. In the event of obvious non-normality or unequal variance which could not be overcome by data transformation, non-parametric tests were used. Multiple comparison analyses (Dunn test) were performed in case of significant differences in means between the three groups (see table 1, fig 3 and “Functional outcome” section). Multiple comparisons of categorical data were performed between each pair of groups, and in case of significant differences in proportions between the groups, the Bonferroni adjustment to the α-value was performed (see table 2). Univariate analyses of grouped pre- and peroperative variables were performed using the χ² test in order to identify factors associated with primary anatomical hole closure. Linear regression analyses were used to identify preoperative factors associated with a favourable functional outcome at 12 months. Statistical analyses were performed using the Sigma Stat 3.5 software package (Systat Software, Richmond, CA).

RESULTS

Participant flow

Between 15 March 2005 and 1 February 2007, 78 eyes (in 75 patients) with stage 2 and 3 idiopathic macular hole without any visible epiretinal fibrosis were randomised. Twenty-five were allocated to vitrectomy alone (non-peeling), 35 to vitrectomy plus ICG-assisted ILM peeling and 18 to vitrectomy plus TB-assisted ILM peeling. All eyes received the intended treatment according to randomisation and were followed for a mean of 12.0 months after surgery (SD = 0.4). One patient (ICG peeling group) was lost to follow-up due to a disseminated cancer disease, leaving 77 eyes for the analyses; 25 in the non-peeling group, 34 in the ICG peeling group and 18 in the TB peeling group. The study flow is summarised in fig 1.

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Figure 1

Flow of participants from allocation to follow-up 12 months postoperatively. *Stage 3 vitrectomy alone (non-peeling) arm terminated 15 May 2006 following planned interim analysis. **Patient 83 excluded from analysis of surgical effect on functional outcome in primary closed holes due to a macula off-retinal detachment with formation of proliferative vitreoretinopathy at last follow-up. ICG, Indocyanine Green; ILM, internal limiting membrane; TB, Trypan Blue.

Baseline data

The baseline patient- and ocular characteristics are summarised in table 1. All registered variables were balanced between the three groups except that the TB peeling group, according to the study design, included only stage 3 holes which thus were expectedly larger.

Anatomical outcome

The anatomical results are presented in table 2. Primary anatomical macular hole closure was obtained in 11 of 25 eyes (44%) in the non-peeling group, 32 of 34 (94%) in the ICG peeling group and 16 of 18 (89%) in the TB peeling group. In the non-peeling group the primary closure rate was six of 11 (55%) for stage 2 holes, and five of 14 (36%) for stage 3 holes. In the ICG peeling group the primary closure rate was 12 of 12 (100%) for stage 2 holes and 20 of 22 (91%) for stage 3 holes. This difference is statistically significant for both stage 2 (p = 0.014) and stage 3 holes (p<0.001). For stage 3 holes, there was no significant difference in primary hole closure rate between ICG-peeled eyes and TB-peeled eyes (p = 1.0).

Sixteen of the 18 patients without primary macular hole closure underwent a second surgery with additional ICG-assisted ILM peeling, one patient (ICG peeling group) declined reoperation, and one patient (TB peeling group) experienced late primary closure of the macular hole between 3 and 6 months of follow-up. None of the closed holes reopened during the 12-month follow-up period. The final anatomical success at 12 months was 24 of 25 (96%) in the non-peeling group, 33 of 34 (97%) in the ICG peeling group and 18 of 18 (100%) in the TB peeling group, p>0.55.

Univariate analyses did not find any statistically significant associations between preoperative variables (gender, age, baseline BCVA, hole aperture diameter, hole base diameter, symptom duration, axial length, self-reported face-down time) and primary macular hole closure (p = 0.08 to 0.98). The only significant factor associated with primary hole closure was whether an ILM peeling operation was performed or not (p<0.001), confirming the highly significant outcome effect of ILM peeling on anatomical closure.

Functional outcome

The visual acuity results of the 77 patients 12 months after macular hole surgery are summarised in fig 2. The overall mean BCVA at 12 months was 70.4 ETDRS letters (SD = 9.8), median 72 letters, range 38–87 letters. Fifty-one of 77 eyes (66.2%) reached BCVA ⩾69 ETDRS letters (corresponding to ⩾20/40 Snellen fraction), and 60 of 77 (77.9%) gained three lines or more on the ETDRS chart (corresponding to a halving of the visual angle).

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Figure 2

Visual acuity before and 12 months after macular hole surgery (n = 77). Horizontal line at 69 Early Treatment of Diabetic Retinopathy Study (ETDRS) letters (corresponding to ⩾20/40 Snellen fraction). Oblique lines above and below the line of equality correspond to gaining or losing three lines or more on the ETDRS chart. (1) Patient #17, macular hole not closed after two operations. (2) Patient #89, macular hole not closed after one operation, declined reoperation. (3) Patient #83, macular hole closed but developed macula-off retinal detachment. ICG, Indocyanine Green; TB, Trypan Blue.

The overall mean BCVA in the non-peeling group was 68.8 letters (SD = 11.4), and the mean BCVA gain was 17.7 ETDRS letters (SD = 8.9). In the ICG peeling group, the mean BCVA was 71.5 letters (SD = 7.9), and the mean BCVA gain was 21.0 ETDRS letters (SD = 8.3). The values are not significantly different (p = 0.45; p = 0.16). For the stage 2 subgroup, there was no significant difference in mean BCVA (73.1 letters vs 70.9 letters) or mean BCVA gain (20.1 vs 16.9 letters) between the non-peeling group and the ICG peeling group (p = 0.59; p = 0.35). For the stage 3 subgroup, there was no significant difference in mean BCVA between the non-peeling group (65.4 letters), ICG peeling group (71.8 letters) or TB peeling group (72.2 letters), p = 0.09. Stage 3 eyes on average gained a mean of 15.8 letters (non-peeling group), 23.2 letters (ICG peeling group) and 22.3 letters (TB peeling group) from baseline until 12 months of follow-up. This difference was significantly different (p = 0.02), and a pairwise multiple comparison analysis confirmed a significant difference between the non-peeling group and the ICG peeling group (p = 0.01) and between the non-peeling group and the TB peeling group (p = 0.02). There was no difference between ICG-peeled and TB-peeled eyes (p = 0.69). Also, the proportion of three-line gaining stage 3 holes were significantly higher in the ICG peeling group (18 of 21) and TB peeling group (13 of 18) as compared with the non-peeling group (eight of 14), p = 0.02. We bear in mind that 14 holes in the non-peeling group, one hole in the ICG peeling group and one hole in the TB peeling group underwent repeat surgery.

When looking at all finally closed holes regardless of treatment regimen (n = 75), there was a significant visual acuity gain from baseline until 3, 6 and 12 months of follow-up (p<0.001; fig 3). There was a trend of continuous visual acuity gain from 3 to 6 months and from 6 to 12 months of follow-up (p>0.05). There was no significant difference in mean visual acuity gain between eyes in the stage 2- and 3 subgroup at any of the follow-up visits (3 months p = 0.34; 6 months: p = 0.24; 12 months: p = 0.19).

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Figure 3

Visual acuity (VA) gain during study, all closed holes (n = 75). ETDRS, Early Treatment of Diabetic Retinopathy Study.

Linear regression analyses found a statistically significant correlation between baseline BCVA and 12 months BCVA (fig 2, r = 0.53, p<0.0001). There was also a statistically significant correlation between baseline hole aperture diameter, baseline hole base diameter and 12-month BCVA (r = 0.31, p = 0.007 and r = 0.32, p = 0.005). Interestingly, there was no correlation between baseline BCVA and 12-month visual acuity gain (r = 0.15, p = 0.19) or between baseline hole aperture diameter and 12-month visual acuity gain (r = 0.004, p = 0.97).

Eyes with primary hole closure

In order to answer the question whether ICG staining and ILM peeling negatively affected the functional outcome, we compared the visual results of primary closed holes (n = 59). Holes in the non-peeling group theoretically would be closed with the minimal amount of retinal trauma, whereas holes in the ICG peeling group would be closed with a possible risk of mechanical retinal damage by ILM peeling and toxic damage by ICG staining. Holes in the TB peeling group would be closed with ILM peeling but with a reduced risk of dye-related toxicity, assuming that Trypan Blue is the “safe” dye. The visual outcome in eyes with primary hole closure is presented in table 3. One patient with obvious visual loss due to a complicated macula-off retinal detachment with formation of proliferative vitreoretinopathy (PVR) and posterior capsular opacification (PCO) was excluded from these analyses.

Overall, eyes with primary macular hole closure reached a mean of 72.8 ETDRS letters (SD = 7.6) and gained a mean of 21.6 letters (SD = 7.1) 12 months after surgery. As illustrated in fig 4, this is significantly better than the 15 eyes that needed a second surgery to close the macular hole (mean BCVA: 66.4 letters (SD = 8.6), p = 0.007; mean visual acuity gain: 17.4 letters (SD = 7.7), p = 0.05). There were no significant differences in baseline ocular characteristics between primary and secondary closed holes (p>0.16). Reoperations were performed a mean of 3.3 months (SD = 1.6 months) after primary surgery.

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Figure 4

(A) Visual acuity (VA) at 12 months in holes with primary and secondary closure. (B) VA gain at 12 months in holes with primary and secondary closure. *One patient (Trypan Blue peeling group) who experienced late primary closure of the macular hole between 3 and 6 months of follow-up is included in the analysis. ETDRS, Early Treatment of Diabetic Retinopathy Study.

The non-peeling group reached a mean of 74.9 letters and gained a mean of 20.7 letters from baseline. The ICG peeling group reached a mean of 72.4 letters and gained a mean of 21.5 letters. The difference was not significantly different (p = 0.32). Considering the stage 2 subgroup with primary macular hole closure, there was a trend of a better mean BCVA in the non-peeling group (78.2 letters) as compared with the ICG peeling group (70.9 letters), p = 0.06. Also, the mean visual acuity gain in the stage 2 subgroup was slightly better in the non-peeling group (21.8 letters) than in the ICG peeling group (16.9 letters) but not statistically significantly so (p = 0.2). In addition, stage 2 holes in the non-peeling group increased significantly more than the ICG peeling group from 3 to 12 months after surgery (6.9 letters vs 1.8 letters; p = 0.02). There were no significant differences in visual results between the non-peeling group, ICG peeling group and TB peeling group for eyes in the stage 3 subgroup with primary macular hole closure (fig 5).

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Figure 5

(A) Visual acuity (VA) gain in primary closed stage 2 holes. (B) VA gain in primary closed stage 3 holes. EDTRS, Early Treatment of Diabetic Retinopathy Study; ICG, Indocyanine Green; TB, Trypan Blue.

Adverse events

The incidence of recorded peroperative and postoperative adverse events is summarised in table 4. There were no differences between the three treatment groups, except that macular RPE changes and dissociation of the optic nerve fibre layer (DONFL) were seen only in the two ILM peeling groups. Iatrogenic retinal tears induced during posterior vitreous cortex separation or ILM peeling were found only in eyes with ICG-assisted ILM peeling (four of 34, 11.8%). Overall iatrogenic retinal tears occurred in five of 93 surgeries (5.4%), and all tears received intraoperative retinopexy. Retinal detachment (RD) occurred in two eyes of 93 vitrectomies (2.2%). One detachment (ICG peeling group) was peripheral and was treated successfully without complications by means of an encircling band, revitrectomy and silicone oil tamponade. The second case (ICG peeling group) developed a macula-off detachment 7 weeks after MHS. At the final follow-up visit, the retina was completely attached after silicone oil instillation and removal, unfortunately with progressive development of moderate to severe PVR and dense posterior capsular opacification.

Visual-field defects were detected by 30-2 automated perimetry in two of 77 eyes (2.6%). One patient (TB peeling group) had a nasally located peripheral wedge shaped absolute scotoma, and the patient (ICG peeling group) who developed a macula-off retinal detachment had an extensive visual-field defect involving the two upper quadrants. No relative defects, defined as loss of more than 10 dB in threshold luminance in two neighbouring stimulus points, were detected. The mean total deviation of sensitivity in the central 10° (16 stimulus points) was manually calculated from the 30-2 total deviation maps. There was no significant difference in mean total deviation of sensitivity in the central 10° between non-peeled eyes (−16.1 dB, SD = 28.9), ICG-peeled eyes (−24.1 dB, SD = 18.9) and TB-peeled eyes (−30.1, SD = 35.8) with primary macular hole closure (p = 0.37).

Retinal pigment epitheliopathy was systematically looked for on fundus photographs in the area of the previous macular hole, in the macular region corresponding to the ILM peeled area (but outside fovea), and peripherally. In total, atrophic or hypertrophic RPE changes were observed as white-yellow or reddish lesions at the final follow-up visit in 36 of 77 eyes (46.8%). Nine of 77 eyes (11.7%) had RPE changes in the fovea at the location of the previous macular hole, equally distributed between the three treatment groups. Macular RPE changes outside fovea were observed only in eyes which had undergone ILM peeling (p = 0.002). In eyes with only one vitrectomy, 33.3% (11 of 33) in the ICG peeling group and 64.7% (11 of 17) in the TB peeling group had macular RPE changes (p = 0.08). Corresponding areas of induced fundus autofluorescence in the macula (iatrogenic punctate chorioretinopathy)37 were seen in only four of 77 eyes (5.2%). All four eyes were primary closed holes from the TB peeling group. There was no difference in final mean BCVA of primary closed holes with macular RPE changes (72.6 letters, SD = 7.8) and without macular RPE changes (72.9 letters, SD = 7.6), p = 0.89. Two patients (one in the non-peeling group and one in the TB peeling group) had peripherally located RPE changes with a corresponding area of induced fundus autofluorescence.

A dissociated optic nerve fibre layer3536 (DONFL) was observed in the macular region on the colour fundus photographs in 33 of 77 eyes (42.9%) 12 months after surgery. DONFL was seen only in eyes which had the ILM peeled. There was no difference in the incidence of DONFL in ICG-peeled (18 of 33 eyes, 54.6%) and TB-peeled eyes (seven of 17 eyes, 41.2%) with only one vitrectomy (p = 0.56). There was no difference in final mean BCVA of primary closed holes with DONFL (72.2 letters, SD = 6.5) and without DONFL (73.3 letters, SD = 8.3), p = 0.62; nor was there any significant difference in 12-month mean total deviation of sensitivity in the central 10°, as obtained from the 30-2 automated perimetry, in primary closed holes with DONFL (−29.4 dB, SD = 22.2) and without DONFL (−20.5 dB, SD = 28.3), p = 0.10. DONFL was not associated with development of absolute or relative visual-field defect in any of the patients.

The most prevalent postoperative complication was formation of posterior iris synechiae, which were present in 24 of 77 eyes (31.2%) at the final follow-up visit with no difference between the three groups after primary surgery (p = 0.30). Posterior capsule opacification occurred in 10 of 77 eyes (13.0%), eight of which underwent YAG-laser capsulotomy. The remaining two eyes did not receive YAG-laser capsulotomy (postponed in one patient with RD sequelae and PVR formation; one patient declined further treatment). Intraocular pressure (IOP) spikes (⩾30 mm Hg) on the first postoperative day were seen after 19 of 93 surgeries (20.4%). The mean IOP values 3, 6 and 12 months after surgery had returned to the baseline level (p = 0.18). There was no significant difference in IOP at any time between the three groups in eyes with only primary vitreous surgery (p-week 1 = 0.29). There was no significant correlation between the used C₃F₈ gas concentrations (10–15%) and postoperative IOP at day 1 (r = 0.13, p = 0.34) or week 1 (r = 0.06, p = 0.70).

DISCUSSION

This randomised clinical trial aimed at examining the value of ILM peeling in stage 2 and 3 idiopathic macular hole surgery. The results show that surgery with ILM peeling, for both stage 2 and 3 macular hole, is associated with a significantly higher closure rate than surgery without ILM peeling (stage 2: 100% vs 55%; stage 3: 90% vs 36%). Visual outcomes in eyes with primary hole closure were not significantly different between the groups, indicating that surgery with short exposure 0.05% (isotonic) ICG-assisted ILM peeling does not significantly affect the functional outcome negatively. Our study is the first randomised clinical trial in Caucasian patients comparing anatomical and functional outcomes after MHS with or without ILM peeling. We have conducted and reported the study according to the CONSORT53 statement in order to obtain complete transparency of study design, conduct and analysis.

The primary closure rate of >90% after ILM peeling (using either ICG- or TB staining) is similar to results from previous studies,1718192022232425 but the primary closure rates after surgery without ILM peeling is lover in this study than previously reported.171819 This is probably attributed to our radical protocol where removal of epiretinal tissue as well as any instrument manipulation with the retinal surface was forbidden in the non-peeling group. Even though our study only included holes with short symptom duration and no visible epiretinal fibrosis at the baseline examination, the anatomical results may indicate that some invisible or fine cellophane maculopathy, exerting tangential traction on the internal limiting membrane, may have been present in some of the patients. Thus, our study shows that ILM removal and/or epiretinal tissue dissection seems to be a necessary manoeuvre in the majority of macular hole surgeries in order to obtain acceptable closure rates. Additionally we cannot exclude the theoretical possibility that missed residual prefoveal vitreous from a split in the posterior vitreous cortex (vitreoscisis) may have exerted tangential traction on the retinal surface.

Besides these anatomical considerations, one also has to appreciate that our a priori assumptions of an 80–90% closure rate without ILM peeling are based entirely on retrospective studies and have newer been shown in randomised trials. The discrepancy may partly be attributed to the well-known risk of exaggerated treatment effects when reporting non-randomised trials.56 The present study’s primary closure rate of 45% without peeling is comparable with the only randomised trial to date on the subject by Kwok et al,21 who found a 32% closure rate without ILM peeling in an Asian population with a high proportion of chronic macular holes.

The overall functional results confirm that MHS generally leads to favourable visual results, with 66.2% of eyes achieving visual acuity of ⩾69 ETDRS letters (⩾20/40) and 77.9% improving more than three lines ( = halving of the visual angle) in this study. These results are in the better end of previously published data,18192022232457 and may be attributed partly to the fact that this study is the first where all included eyes were pseudophakic at baseline, thus eliminating cataract as a confounder for functional outcome, partly to the fact that only stage 2 and 3 lesions with a relatively short symptom duration (<12 months) were included. Stage 4 macular holes were not included in this study, as we presumed these lesions to be of a longer duration and having a higher prevalence of epiretinal membranes.5859 Additionally posterior vitreous cortex separation has already occurred in stage 4 holes, indicating that ILM peeling would be necessary in order to alter the mobility of the macular hole edges and induce macular hole closure.

Posterior capsule opacification (PCO) was systematically looked for and was treated when present in eight of 10 eyes. Of the two remaining eyes with PCO, one was excluded from visual outcome analyses due to a complicated macula-off retinal detachment, thus limiting PCO as a confounder for visual outcome to one patient who declined further treatment (non-peeling group). The 12 months of follow-up in this study may also partly explain the favourable results, because we confirm previous results60 that the visual outcome continues to improve for at least 1 year after surgery, indicating continuous functional recovery of foveal photoreceptors.

In order to answer the question whether ICG-assisted ILM peeling negatively affected the functional outcome, we compared the visual results of primary closed holes in the three groups. One patient with obvious visual loss due to a complicated macula-off retinal detachment was not included in these analyses. We did not find any significant differences in mean BCVA, mean visual acuity gain, percentage of eyes reaching 20/40 or better, or percentage of eyes with halving of the visual angle between the non-peeling group, the ICG peeling group and the TB peeling group. Additionally we found no significant difference in the mean total deviation of sensitivity in the central 10° between non-peeled eyes, ICG-peeled eyes and TB-peeled eyes.

Some indications that ICG peeling may compromise functional outcome slightly were found in the subgroup of primary closed stage 2 holes, where we found a significant difference in mean visual acuity gain from 3 to 12 months postoperatively and a trend of a better mean BCVA between non-peeled eyes and ICG-peeled eyes. Given the change in study protocol after the interim analysis which led to termination of the non-peeling stage 3 arm, the statistical power of the performed analyses for final visual outcome in primary closed holes was lower than desired. However, the study had sufficient power (0.77 and 0.83) to detect differences of ⩾10 ETDRS letters in final mean BCVA in primary closed stage 2 and stage 3 holes. Our observation that reoperated eyes performed significantly worse in all visual outcome parameters, as compared with holes that closed after a single surgery, indicates that the primary focus should be on closing the hole in one procedure, rather than seeking a 50:50 chance of closing the hole without ILM peeling in the search for a (non-significant) slightly better functional outcome.

For stage 3 holes, ICG-peeled- and TB-peeled eyes had identical closure rates (91% vs 89%), and the visual outcomes for primary closed holes in the non-peeling group, ICG peeling group and the TB peeling group were identical (p = 0.96), indicating that ICG staining of the ILM, as performed in this study, can be performed in stage 3 MHS without any detrimental effects on functional outcome. We used Trypan Blue as the “safe” control dye for staining the ILM,45 assuming it to be non-toxic or at least minimally toxic to retinal cells, even though experimental studies recently have also reported possible Trypan Blue toxicity in RPE cell cultures.616263 Our findings, however, are consistent with the only RCT to date comparing outcomes after TB-assisted and ICG-assisted ILM peeling in MHS, where no adverse observations or differences in functional outcome were found.57

MHS, using any of the techniques described in this study, is a safe procedure with a low incidence of sight-threatening adverse events. Iatrogenic retinal tears were observed in 5.4% of the surgeries, and the retinal detachment rate was 2.2%; thus our results are similar to previously published data.2125 Postoperative visual-field defects which previously in part have been attributed to the use of ICG-assisted ILM peeling were also rare. In only two patients (2.6%), visual-field defects were found at the last follow-up. One defect is probably due to a phototoxic effect from the light probe due to prolonged operation time (TB peeling group), and the second visual-field defect was secondary to development of a postoperative retinal detachment. These data thus support that visual-field defect is a complication with low incidence64 and that ILM peeling or staining does not increase the risk of field defects. Some RPE changes, both in the fovea at the location of the previous macular hole and in the macular area at locations corresponding to grasping the ILM were noted. Foveal RPE changes were equally distributed between the three groups and therefore are unlikely to be specific for ICG-assisted ILM peeling. Macular RPE changes were found in 41.6% of all eyes at the final visit, but they were only seen in eyes which had undergone ILM peeling and are likely to correspond to ILM “grasping marks.” There seemed to be an over-representation of macular RPE changes (64.7% vs 33.3%) and macular iatrogenic punctate chorioretinopathy (four cases vs 0 cases) in the TB peeling group. This adheres to our clinical experience of a more challenging ILM peeling when staining with Trypan Blue than with ICG, especially in eyes without epiretinal membranes as in this study. The macular RPE changes did not affect the visual outcome.

A dehiscence of the retinal nerve fibre layer (DONFL) was found on colour fundus photographs in 50.0% (25 of 50) of the ILM peeled eyes with only one vitrectomy. The small nerve fibre layer irregularities were also to be seen in the corresponding OCT scans. DONFL was observed with the same incidence in ICG-peeled- and TB-peeled eyes, indicating that the ILM peeling procedure itself rather than ICG staining was responsible for the optic nerve layer dehiscence. Visual outcomes were identical in ILM peeled eyes with and without DONFL, and no differences in central retinal sensitivity were detected in eyes with and without DONFL, indicating that DONFL appearance is not detrimental to functional outcome. This adheres to previous reports.3536

All phakic patients underwent cataract surgery 4–6 weeks prior to MHS. Besides giving us a unique opportunity to follow functional outcome prospectively without the confounding effect of inevitable development of cataract, we prevent patients from having to return for potentially difficult cataract surgery with unstabile posterior capsules, loose zonules and a higher risk of posterior capsular rupture due to reduced vitreous gel support.6566 The risk of inducing late reopening of a previously closed macular hole is also reduced.67 Unfourtunately, we observed a high incidence of posterior iris synechiea (31.2%). This is higher than that previously reported in pseudophakic eyes after combined phaco-vitrectomy and face-down positioning (∼10%);6869 however in this study we registered all degress of posterior synechiae (from single strands to sectoral synechiae). Incomplete face-down positioning could be a disposing factor for posterior synechiae development due to forward compression of the IOL by the gas bubble in an upright position. All patients stayed in our department for at least 24 h after surgery for instruction and observation of keeping a complete face-down position. The high incidence of posterior synechiae is probably due to an unresolved inflammatory response present after the cataract surgery which was performed only 4 weeks prior to MHS. All patients received anti-inflammatory treatment with dexamethasone eye-drops from the day after cataract surgery until 3 weeks after MHS. The high incidence of posterior iris synechiea indicates the need for increased anti-inflammatory treatment and/or periodic pupil dilatation in pseudophakic eyes during face-down positioning. Retrospectively, our study design with 100% pseudophakic patients would have allowed an intraocular gas bubble that completely filled the vitreous cavity suggesting that prone posturing might not have been required for closing the macular holes. The study was however not designed to test the effect of face-down positioning and gas tamponades.

The major strengths of the present study are the randomised design minimising selection bias and increasing generalizability, the strict clinical criteria for evaluating preoperative hole size and postoperative hole status, and the fact that all included patients were pseudophakic at baseline eliminating the effect of cataract on functional outcome. The main methodological limitation of our study was the change in study protocol made after the interim analysis, which showed unexpecded poor closure rates after surgery without ILM-peeling. By terminating the non-peeling stage 3 arm and instead introducing the TB peeling arm, the conditions for the initial sample size calculation were changed.

In summary, this article described a randomised clinical trial with masked observers to evaluate the clinical effect, and possible negative influence, of ICG-assisted ILM peeling on retinal function and morphology. ILM peeling was found to induce significantly higher closure rates than non-peeling for both stage 2 and 3 idiopathic macular holes. The use of 0.05% isotonic ICG-assisted ILM peeling with short exposure time did not have any significant detrimental effects on functional outcome, whereas performing a reoperation did. We therefore conclude that 0.05% isotonic ICG-assisted ILM peeling is a safe alternative and can be recommended in all cases of full-thickness macular hole surgery.