Inverted internal limiting membrane-flap technique for large macular hole: a microperimetric study

Background Inverted Internal Limiting Membrane (ILM)-flap technique would seem to lead to higher closure rate and better visual acuity than traditional procedure with ILM peeling for the treatment of large macular hole (LMH). Visual acuity recovery does not reveal many other functional changes related to surgical approach. Our purpose was to evaluate macular function and morphology over a 1-year follow-up after inverted ILM-flap technique for LMH by using microperimetry in order to predict visual prognosis. Methods This study was a prospective unrandomized single-center study. 23 eyes of 22 patients with idiopathic LMH, with a minimum diameter ranging from 400 to 1000 μm, were included. All patients underwent vitrectomy with inverted ILM-flap technique and gas tamponade. We analyzed macular hole closure rate and functional outcomes including best-corrected visual acuity (BCVA), macular sensitivity (MS) at central 12° and central macular sensitivity (CMS) at central 4°, and fixation behavior as bivariate contour ellipse area (BCEA, degrees2) at 68%, 95%, and 99% of fixation points measured by microperimeter, over a follow-up of 12 months. Results The macular hole closure rate was 98%. The BCVA improved from 20/230 (Logmar, 1.06 ± 0.34) to 20/59 (logMar, 0.47 ± 0.45) at last follow-up (p < 0.001). Retinal sensitivity and BCEA significantly improved (MS, p = 0.001; CMS, p < 0.0001; BCEA: 68%, p < 0.01; 95%, p < 0.01; 99%, p = 0.001). Multiple stepwise regression analysis showed the final BCVA was significantly associated with macular hole size (β = 0.002, p = 0.03), preoperative MS (β = − 0.06, p = 0.001) and BCEA at 95% and 99% of fixation points (β = − 0.12, p = 0.01; β = 0.06, p = 0.01). Conclusions Inverted ILM-flap technique for LMH results in good morphologic and functional outcomes. Macular hole size and microperimetric parameters as preoperative MS and BCEA have a predictive role on post-surgical visual acuity.


Background
Idiopathic full-thickness macular hole is an important cause of central vision impairment and metamorphopsia in elderly people [1]. The conventional procedure for macular hole surgery consists of internal limiting membrane (ILM) peeling and air or gas tamponade [2][3][4][5]. In cases with large macular hole (LMH) the anatomical success rate, as the closing of the hole, is as low as 40% to 80% using conventional procedure [6][7][8]. Michalewska et al., in a prospective trial, found that anatomical closure rate was higher using a novel technique of inverted ILMflap technique (98%) compared to traditional procedure with ILM peeling (88%) for the treatment of LMH. In the same study post-operative visual acuity was significantly higher using the new technique [9]. Several papers have

Open Access
International Journal of Retina and Vitreous *Correspondence: alfred.nir@tiscali.it Sborgia et al. Int J Retin Vitr (2019) 5:44 suggested that inverted ILM-flap technique may be better for the treatment of LMHs considering morphologic and functional outcomes [10][11][12][13]. Despite high closure rate and visual acuity recovery, functional changes after macular hole surgery are really complex. Visual acuity recovery does not reveal many other functional changes related to macular pathologies and surgical manipulations. So central retinal sensitivity and fixation behavior analysis using microperimeter can provide further objective and quantitative informations about macular function, enabling to analyse exact correlation between macular disease, as age-related maculopathy, diabetic macular oedema, macular oedema after vein occlusion, idiopathic epiretinal membrane, central serous chorioretinopathy, inflammatory macular oedema and macular dystrophy, and corresponding function, in assessment of natural history and treatment outcomes [14][15][16][17][18][19][20][21][22]. Previous papers reported a significant improvement of retinal sensitivity and fixation properties after traditional ILM peeling for macular hole [23][24][25][26][27], revealing a predictive role of preoperative macular hole feature and microperimetric parameters on visual recovery [27][28][29][30]. The aim of this study was to evaluate morphologic and functional outcomes after vitrectomy with inverted ILM-flap technique for LMH by using microperimetry in order to predict visual prognosis.

Methods
In this prospective study we analysed 22 eyes of 23 positioning for 3 days postoperatively. The study followed the tenets of the Declaration of Helsinki and was approved by the institution's review board. Statistical analysis Statistical analysis was based on all patients included in the study. No formal sample size calculation was performed. Mean and standard deviations were used for continuous variables. A t test was performed on the changes from baseline in BCVA, MS, CMS and BCEA. All statistical tests were performed at the p < 0.05 significance level. Simple linear regression model was performed to assess the relationship between BCVA at 12 months and each indipendent variable. The independent variables included age, sex, lens status, axial length, baseline macular hole size, baseline BCVA, MS, CMS, and BCEA 68%, 95% and 99%. Multiple linear regression model in backward with stepwise method was performed to assess any predictive factors associated with postoperative visual acuity at 12 months (cut-off removal variable, p ≥ 0.10). The factors with a p value < 0.05 in the multiple model were considered as potential baseline predictors. Statistical analysis was made using STATA 12.1 Statistical Software (StataCorp), 2014, release 12 (College Station, TX).

Discussion
The inverted ILM-Flap technique has been reported to have a high closure rate of LMH [7][8][9][10][11]. The ILM-flap might work as a scaffold for the proliferation and migration of activated Müller cells that promote the closure of macular hole producing neurotrophic factors [32]. This technique leads to an acceleration of the wound healing processes at the macula. We reported a high anatomical closure rate of 98%, in line with previous reports. In our experience, we found that some modifications of this technique, as lowering the perfusion pressure when inverting the flap and during air-fluid exchange, and the use of gas tamponade might help to avoid the dislocation of the ILM-flap during and after surgery increasing the closure rate, as confirmed by the results of previous papers [7,8,[10][11][12][13]. The influence of flap inversion on functional recovery has been analysed in several papers to understand whether visual prognosis is affected by the presence of the flap over the hole. Indeed, the flap, working as a scaffold and basement membrane for tissue proliferation, should provide an environment to instruct the photoreceptors to assume correct position during the reconstruction process and finally to improve the postoperative visual acuity [9]. In line with previous studies, we found that postoperative visual acuity significantly improves after ILM-flap inversion [7][8][9][10][11][12]. In particular, major visual acuity improvement was achieved as early as one and 3 months after surgery, and no patients had clinically significant visual acuity improvement beyond this period. This trend in visual improvement could be achieved regardless the recovery of outer retinal layers at the foveal site that would seem occur after at least 3 months from surgery [33]. To date there are no definite conclusions on the role of inverted ILM-flap technique on the outer retinal layers changes and we did not analyse this point. However, our visual outcomes could be explained by the integrity of the detached photoreceptor layer in the perifoveal area or at the edges of the hole, and by the choice to cover rather than to fill the hole with the ILM-flap to avoid that glial tissue proliferation may mechanically obstruct the recovery of outer retinal layers, as previously suggested [34]. However, the functional effects of LMH and surgical technique could be underestimate by BCVA changes so microperimetry, a point to point measurement of retinal sensitivity, may better  reflect the functional status and recovery after the hole has been repaired [23-26, 28, 35]. In this study, we recorded retinal sensitivity and fixation behavior as BCEA, before and after inverted ILM-flap technique for LMH over one-year follow-up. We analysed the changes in MS, which represents the general sensitivity within the central 12°, and CMS, indicating the retinal sensitivity within the central 4° of the macula. Previous studies suggested the necessity of investigating MS and CMS separately when evaluating central retinal sensitivity, considering that the sensitivity of the parafoveal retina area is higher than that at the central 0° in normal individuals, in part explained by the "masking effect" caused by the fixation target during examination [36,37], and the age-related decrease of sensitivity in the perifoveal area than in the center of macula [38]. Moreover, our aim was to evaluate functional recovery at different retinal sites, at central 4° (corresponding to an area of 1200 µm in diameter at the foveal site) where the gliosis process, promoted by the inverted ILM-flap and controlled by Müller cells, could influence inner and outer retinal layers integrity with functional effects [9,39,40], and at central 12° where ILM peeling, inducing a temporary swelling of the arcuate nerve fiber layer (SANFL) as the earliest manifestation of dissociated nerve fiber layer (DONFL), leads to a reduced retinal sensitivity and paracentral scotomata in the peeled area [41]. At baseline, microperimetry demonstrated a lower sensitivity at central 4° (CMS) revealing an absolute scotoma, which corresponds to the neurosensory defect, with surrounding higher sensitivity at 12° (MS) defining a concentric relative scotoma in the region of the retina around the hole [42]. Our results showed that both CMS and MS significantly improved after surgery. CMS improved incrementally from the first follow-up, mainly at month 1 and 3 after surgery, probably related to the viability of detached photoreceptors at the edges of the hole [28,29]. Instead, MS significantly improved only after 6 months from surgery, in particular nine patients showed an early reduction or unchanging of sensitivity at month 1 and 3, probably influenced by early functional damages on inner retinal layers due to ILM peeling, damages that are reversible as confirmed by our results and previous papers [41,43]. Furthermore, Baba et al. observed that Brillant blue G-assisted vitrectomy could guarantee a faster restoration of IS/OS junction and a better  Table 3 Linear

regression model of best corrected visual acuity (BCVA) at 12 months on single variables (A); multiple linear regression model on all variables together in the model (B); final multiple linear regression model in backward with stepwise method (C)
β coefficient, se(β) standard error of coefficient, AL axial length, MH macular hole, BCVA pre best corrected visual acuity preoperative, Logmar logarithm of minimum angle of resolution, MS pre retinal sensitivity preoperative, dB decibel, CMS pre central macular sensitivity preoperative, BCEA 68%, 95%, 99% bivariate contour ellipse area at 68%, 95%, and 99% of fixation points, respectively a All variables included in the model were considered as continuous  postoperative BCVA and retinal sensitivity in the central degrees [44]. Fixation stability is another important functional parameter to consider in the treatment of macular diseases, probably more than fixation location in macular hole condition where the locus of fixation could already be naturally relocated out of the foveal site. We analysed the effect of inverted ILM-flap on a quantitative parameter of fixation behavior, as BCEA. In our series BCEA improvement had a similar trend at 68%, 95% and 99% of fixation points at all follow-up. We observed a reduction in dimension of the cloud of the fixation points at month 1 and 3, followed by a mild increase at 6 months and a new mild reduction of all ellipses after 12 months. Tarita-Nistor et al. reported the same improvement in fixation stability (BCEA) at 1 and 3 months after traditional ILM peeling [27]. As they suggested, the closure of the macular hole could lead to a complex reorganization of fixation behavior. Moreover, we can argue that changes in fixation stability would occur regardless the surgical technique used. In our experience, preoperative functional parameters evaluated as BCVA, MS, CMS and BCEA were individually correlated with final visual acuity. After that, a multiple stepwise linear regression analysis revealed that preoperative MS, BCEA at 95% and 99% of the fixation points and macular hole size have a predictive role on final visual acuity at 12 months. We suggest that MS has a significant influence on postsurgical visual acuity, probably because the inverted ILM-flap technique not always leads to photoreceptor reconstitution, and retinal sensitivity at 12° (MS) is less influenced by foveal microstructure recovery after macular hole closure than CMS at central 4°. The predictive role of a quantitative parameter of fixation behavior as BCEA on postsurgical visual acuity was already reported [27,30], confirming that a smaller ellipsoid area correlates to more stable fixation and better visual performance. Also macular hole size was revealed as an independent factor predicting 12-month postoperative BCVA. Ota et al. observed the same predictive role of hole diameter for 6-month visual acuity in patients underwent different surgical techniques for LMH [45]. This could be because the recovery in foveal structure after inverted ILM-flap not always correspond to a complete reconstitution of outer retinal layers, not differently from traditional ILM peeling [45,46]. Limitations of the study include the small sample size, the absence of a control group, the absence of analysis of outer retinal layers features at OCT scans related to functional changes, and the measurement error or intrinsic variability of microperimetric test. Factors acting on test variability are related to patient's compliance and its anatomical and functional condition, and to the examiner and/or instrument. About patient condition, it should be mentioned the "learning factor" which can justify a certain degree of improvement during follow-up. Regards to the instrument, it should be mentioned the eye-tracker system, not able to ensure the same accuracy of analysis between the posterior pole and peripheral retina, the "point to point" overlapping error (0.5° to 2°) when "Follow-up" program is used, the "4-2 strategy" which can extend the duration of the test, the "ceeling effect" of MP-1, meaning the tendency to accumulate responses at the highest limit of the sensitivity threshold, and thesize of the given stimulus (Goldmann III, 4 mm 2 area, 26 min diameter of arc, or 0.4°) that, because of "spatial summation", involves more photoreceptors which converge centrally on a single ganglional cell. As points of strength of this study we highlight the prospective nature of the study, the long term follow-up of 12 months and to our knowledge, the first microperimetric analysis of inverted ILM-flap technique involving different functional parameters.

Conclusions
High closure rate and visual acuity improvement supported the effectiveness of the inverted ILM-flap technique for LMH. Macular sensitivity at central 12° and fixation analysis, detectable by microperimetry, give new informations on functional recovery, also revealing their predictive role on visual acuity after inverted ILM-flap technique.