Preliminary report of minimally invasive corneal neurotization in patients with neurotrophic keratopathy in Southern China

Abstract

Purpose

To describe the clinical features and outcomes of neurotrophic keratopathy (NK) patients treated with minimally invasive corneal neurotization (MICN).

Methods

Medical records of patients with NK who were treated with neurotization surgery between January 2022 and December 2023 were retrospectively reviewed. Eleven patients underwent neurotization surgery via sural nerve grafts from the supraorbital nerves to the affected contralateral cornea. Ocular examinations as well as Cochet Bonnet esthesiometry (CBA) of the affected cornea were performed.

Results

The baseline corneal conditions were as follows: Mackie stage 2 (3, 27%) and Mackie stage 3 (8, 63%). All the patients demonstrated improvements in corneal sensibility and corneal epithelial healing postoperatively. The CBA measurements improved from 1.8 ± 4.0 mm at baseline to 46.4 ± 13.4 mm at 12 months after surgery (P < 0.05). The mean best-corrected visual acuity of 11 patients improved from logMAR values of 1.8 ± 0.3 at baseline to 0.8 ± 0.7 at 12 months postoperatively (p < 0.05). Before surgery, corneal nerves at subbasal levels were completely absent at IVCM in all the patients. Corneal nerve morphology improved and the mean corneal nerve fibre density (CNFD) was 14.1 ± 11.8 n/mm² 12 months postoperatively. The limbal vascular density decreased from 34.1 ± 4.6% at baseline to 6.4 ± 1.7% at 12 months after surgery (p < 0.05). Two patients underwent successful penetrating keratoplasty after MICN due to corneal leucoma.

Conclusion

MICN is a useful surgical treatment for facilitating corneal epithelium healing and improving corneal sensitivity and visual acuity in patients with severe NK.

Trial registration

This study protocol was reviewed and approved by the Ethics Committee of Zhongshan Ophthalmic Center (no. 2024KYPJ129).

Background

In addition to the detection of sensory input, normal corneal innervation is necessary for corneal epithelial homeostasis, proliferation, and wound healing. The cornea is a richly innervated organ that is supplied by the long ciliary nerve and derived via the ophthalmic branch of the trigeminal nerve. Nerves enter the cornea in the middle third of the stroma and are distributed anteriorly towards the center, where they lose their myelin sheath approximately 1 mm from the limbus. Disease that leads to corneal sensory denervation has been shown to decrease the production of several neuromediators [1]. These neuropeptides include substance P, calcitonin gene-related peptide, neuropeptide Y, and nerve growth factors [2]. Deficiencies in these mediators result in corneal epithelial thinning, surface breakdown, and persistent epithelial defects. Deficiencies in these mediators also result in a reduction in microvilli of the corneal epithelium, leading to poor tear adherence to the cornea [3]. In severe cases, persistent sterile ulceration and melting of the corneal stroma may ultimately lead to perforation. The aim of therapy is to protect the corneal surface and damage the basement membrane. Traditionally, the approaches that are used include punctual plugs, unpreserved artificial tears, autologous serum drops, lateral tarsorrhaphy, and amniotic membrane transplant [4, 5]. These treatments may achieve temporary corneal epithelial healing. However, none of these modalities address the underlying anaesthesia in neurotrophic keratopathy (NK).

Recently, several corneal neurotization techniques have been used to treat patients with unilateral corneal anaesthesia [6,7,8]. These techniques have yielded encouraging results for improving ocular surface homeostasis, corneal sensation, and visual acuity. As a peripheral sensory nerve, the favourable aspect of corneal neurotization is the fact that, unlike motor nerves, a sensory target can tolerate denervation for an extended time period; thus, surgical nerve repair can be performed several years after initial damage, maintaining good success rates [9]. Neurotization may be performed by direct local nerve transfer or indirectly through the use of a nerve graft [10]. Recently, a minimally invasive approach to corneal neurotization (MICN) with small blepharoplasty-style incisions was reported, which used sural nerve grafts to connect axons from the contralateral supratrochlear nerve to the affected cornea. The advantages of this approach are small surgical incisions and less invasive to surrounding structures compared to bicoronal approach [11, 12]. Management of NK is challenging and only a few cases using a nerve graft to connect the source of innervation to the affected cornea are available in China [13]. To evaluate the safety and efficacy of this method, we report the clinical outcomes of 11 patients with NK who were treated with MICN.

Methods

This case series included 11 patients who underwent MICN between 2022 and 2023. Approval for this retrospective study was obtained from the Institutional Review Board of the Zhongshan Ophthalmic Center (no. 2024KYPJ129). Informed consent was obtained from all patients, and the study conformed to the tenets of the Declaration of Helsinki.

All the patients underwent a standardized preoperative evaluation. This included medical and surgical history; best-corrected visual acuity (BCVA); examination of the corneal patch staining with the aid of fluorescein stain (CORE staining scale) [14]; Schirmer’s test; limbal vascularization; assessment of average corneal sensation (central, superior, inferior, nasal, and temporal) via Cochet-Bonnet Aesthesiometry (CBA, Luneau Ophthalmologie); and examination of the subbasal nerve of the cornea via in vivo confocal microscopy (IVCM) with the Rostock Cornea Module of the Heidelberg Retina Tomograph II (HRT IIRCM; Heidelberg Engineering GmbH). IVCM images of corneal nerves at the subbasal level were analysed with ACCMetric on the basis of their technical quality and analysed with ACCMetrics (MA Dabbah, Imaging Science and Biomedical Engineering, Manchester, UK). The following parameters were obtained with ACCMetrics: corneal nerve fibre density (CNFD), corneal nerve branch density (CNBD), corneal nerve fibre length (CNFL), corneal nerve total branch density (CTBD), corneal nerve fibre area (CNFA), corneal nerve fibre width (CNFW), and corneal nerve fractal dimension (CNFrD). Corneal limbal vascularization was captured by an independent technician at the same slit lamp and under identical lighting conditions. Four quadrants of the limbal area were selected to evaluate limbal vascular density (vessel area/total area * 100%). ImageJ software version 1.53 (NIH, Bethesda, Maryland) with vessel analysis plugins was used to trace and analyse limbal vessels in standardized slit lamp photographs. Preoperative evaluation of supraorbital and supratrochlear nerve functions bilaterally via a light touch with a wisp of cotton on forehead skin was performed to determine viable donor nerve sites. Schirmer’s test without topical anesthesia was performed with a strip of filter paper inserted into the lower lid at the junction of the middle and outer thirds of the lower lid without anesthesia. The patient is asked to keep their eyes closed for the duration of the test. After 5 min, the filter paper is removed and the amount of wetting is measured in millimeters. Corneal sensation was measured by a Cochet-Bonnet esthesiometer, which is a handheld device that utilizes a nylon monofilament adjustable from 5 to 60 mm in length. It was directly applied to the central and four peripheral quadrants of the cornea (i.e., superiorly, nasally, inferiorly, and temporally) to determine sensitivity. The monofilament is extended to its full length of 60 mm and retracted incrementally in 5 mm steps until the patient can feel its contact. Fluorescein staining was performed with a fluorescein-impregnated strip wetted with a single drop of sterile saline. The excess drop was shaken and removed. The lower eyelid was pulled down, and the strip was gently touched onto the lower tarsal conjunctiva. The patient was asked to blink gently to distribute the dye across the corneal surface. A photograph of the entire cornea was obtained immediately after staining with a blue excitation filter and a diffusion lens at a 10- to 30- degree oblique angle. Patch staining (Grade 100) of the CORE corneal staining scale was recorded. Visual acuity was measured with a Snellen chart at every visit. Count finger (CF) and hand motion (HM) visual acuity were converted to logMAR 2 and logMAR 3, respectively.

Surgical technique

The surgery was performed under general anaesthesia. The sural nerve was chosen as the source of the nerve graft. A longitudinal incision was made posterior to the lateral malleolus to identify and expose the sural nerve. The nerve was traced upward to obtain the desired length of 11–13 cm (Fig. 1A). The sural nerve was maintained in moist gauze (Fig. 1B). All the surgeries were contralateral transfers. The supraorbital nerve (SON), which acts as the donor nerve, was accessed on the unaffected side via a transverse subbrow incision. A subcutaneous tunnel between the upper lids of the right and left eyes was prepared. The sural nerve was crossed over the galbella until it reached the subconjunctiva of the affected eye, which was secured by a hollow polythene introducer (Fig. 1C). After an epineurectomy, the distal end of the nerve graft was divided into 4–5 fascicles (Fig. 1D). A crescent blade is used to create corneoscleral tunnels through the corneal limbus and into the cornea at 2, 4,8 and 10 o’clock. The end of the nerve graft is inserted into corneoscleral tunnels with tying forceps. The nerve fascicles are sutured to the sclera 2 mm posterior to the limbus with 10 − 0 nylon sutures (Fig. 1E). The incision of conjunctiva is closed with 8 − 0 Vicryl suture. The proximal end of the nerve graft was coapted to the SON in an end-to-end configuration with 10 − 0 nylon sutures (Fig. 1F).

Fig. 1

Surgical steps of corneal neurotization. (A, B) Preparation of sural nerve graft from right lower limb. (C) Stretching tube for manipulation of sural nerve graft in the subcutaneous tunnel between upper lids of right and left eye. (D) The fascicles are separated under the operating microscope (E) The fascicles are sutured with 10 − 0 nylon sutures on sclera 2 mm posterior to the limbus. (F) Sural nerve graft end-to-end coaptation to the donor supraorbital nerve is performed

Postoperatively, patients were prescribed antibiotic eye drops and artificial tears three times daily for the first month after MICN. Patients were followed up postoperatively at week 1; months 1, 3, and 6; and every 3 months thereafter. The primary outcome of this study was postoperative corneal sensibility at 12 months of follow-up. The secondary outcomes included postoperative best-corrected visual acuity (BCVA), the state of cornea healing (defined as 0 mm of lesion staining and no other residual staining) and subbasal nerves of the cornea.

Statistical analysis was performed via SPSS 25.0 for Windows (IBM SPSS Statistics for Windows, Version 25.0; IBM Corp, Armonk, NY, USA). The values are expressed as the means ± SDs. The Mann‒Whitney U test was performed to compare nonnormally distributed variables. Fisher’s exact test was used to compare categorical variables. Analysis of the change in CBA over time was performed with one-way analysis of variance with repeated measures. A paired samples Z test was performed to compare the proportions of the two groups. Statistical significance was defined as P < 0.05.

Results

Eleven eyes of 11 patients were included in the study. Two males and 9 females were included in the study population. The average denervation time was 23.4 ± 26.5 months (range: 7–96 months), with Mackie stages 2 (3, 27%) and Mackie stages 3 (8, 73%). The cause of corneal anaesthesia was surgery after cranial tumor in 7 patients, traumatic brain injury in 2 patients, and prior ocular herpes simplex virus infection in 2 patients. The denervation time was recorded from the time of cranial tumor surgery or traumatic brain injury to the time of MICN. In patients with ocular herpes simplex virus infection, the denervation time was recoded from the examination date of impaired corneal sensation of the affected eye to the time of MICN. The baseline characteristics of the patients are summarized in Table 1. The mean follow-up for the corneal neurotization time was 19.5 ± 6.4 months (range: 12–27 months). In two patients, keratoplasty surgery was eventually performed.

Table 1 Patient demographics and clinical feature

The mean CBA of 11 patients improved from 1.8 ± 4.0 mm at baseline to 20.5 ± 9.1 mm at 3 months, 37.3 ± 14.9 mm at 6 months, 45.0 ± 14.9 mm at 9 months, and 46.4 ± 13.4 mm at 12 months (p < 0.05) (Fig. 2), with 4 patients achieving 60/60 in all quadrants including the centre by postoperative month 12.

Fig. 2

Boxplot of mean Cochet − Bonnet aesthesiometry (CBA) over time after minimally invasive corneal neurotization. Significant changes in CBA were observed 12 months post- surgery (P < 0.05)

Nine patients showed an improvement in BCVA. The mean BCVA of 11 patients improved from logMAR values of 1.8 ± 0.3 at baseline to 0.8 ± 0.7 at 12 months postoperatively (p < 0.05) (Fig. 3). Two patients with corneal leucoma did not exhibit significant improvement in BCVA after corneal neurotization. One year after MICN, penetrating keratoplasty was performed in these two patients. Six months after keratoplasty, BCVA improved from 20/400 and FC preoperatively to 20/100 and 20/50, respectively.

Fig. 3

Boxplot of best-corrected visual acuity (BCVA) in patients over time after minimally invasive corneal neurotization. Significant changes in BCVA were observed 12 months post- surgery (P < 0.05)

Before surgery, nine patients demonstrated more than 0.5 mm of corneal patch staining. The average area of corneal fluorescein patch staining was 14.3 ± 6.1 mm [2] at baseline. At month 1, corneal healing was achieved in 5 of the 9 patients who received corneal neurotization. At month 3, all 9 patients achieved 0 mm of lesion staining and no other residual patch staining (Fig. 4). Total corneal epithelium healing was achieved in 5 eyes of 5 patients with single trigeminal nerve injury and in 1 eye of 4 patients with trigeminal and facial nerve injury at 1 month (p < 0.05).

Fig. 4

Clinical picture of cornea and subbasal nerve plexus of neurotrophic keratopathy owing to a traumatic brain injury, before and after corneal neurotization. (A) pre-operative clinical photograph demonstrating a large central epithelial defect with underlying stromal oedema at baseline. (B) Confocal microscopy images showing the absence of corneal nerves before surgery. (C) One year after surgery, slit-lamp photograph revealed an increase in optical clarity with best corrected visual acuity 20/20. Red arrow shows perilimbal transplanted nerve fascicles. (D) The regenerated corneal sub-basal plexus exhibited a near-normal morphology at 12 months after the surgery. ACCMetrics analysis calculated: CNFD = 31.24 n/mm²; CNBD = 74.99 n/mm²; CNFL = 19.01 mm/ mm²; corneal nerve total branch density (CTBD) = 81.24 n/mm²; CNFA = 0.0098 mm2/mm²; CNFW = 0.021 mm/mm²; CNFrD = 1.52

IVCM was assessed before surgery and at 3, 6, and 12 months after surgery. Before surgery, corneal nerves at subbasal levels were completely absent at IVCM in all patients. After corneal neurotization, corneal nerves were successfully identified via IVCM 3 months postoperatively in all patients. Corneal nerve morphology improved, and the mean CNFD was 16.1 ± 9.1 n/mm² (Table 2) 12 months postoperatively. Representative IVCM images obtained at the subbasal level for patient 4 are shown in Fig. 3. Among these patients, six had single trigeminal nerve dysfunction, with a CNFD of 22.2 ± 7.1 n/mm². Another 5 patients experienced multiple nerve damage events with a CNFD of 8.7 ± 4.5 n/mm² (p = 0.02) 12 months postoperatively. The mean CNFD of contralateral eye was 33.7 ± 4.8 n/mm².

Table 2 IVCM parameters examined before and after MICN

Corneal limbal vascularization was assessed before surgery and 12 months after surgery. Before surgery, the limbal vascular density was 34.1 ± 4.6%. Twelve months after surgery, the limbal vascular density was 6.4 ± 1.7% (p < 0.05)(Figure 5).

Fig. 5

Limbal vascularization change before and after minimally invasive corneal neurotization (MICN). (A) Slit-lamp photograph showing inferior corneal epithelium defect and circumferential (360 degrees) limbal vascularization at baseline. (B) One year after MICN, slit-lamp photograph showing limbal vascularization had decreased significantly. (C) Superior quadrant limbal vascularization at baseline. (D) Superior quadrant limbal vascularization one year postoperatively. (E) Binary version of superior quadrant limbal vascularization at baseline with vascular density 34.2%. (F) One year after MICN, binary version of superior quadrant limbal vascularization with vascular density 7.1%

One year after MICN, the postoperative Schirmer test result did not significantly differ from the result before surgery (9.9 ± 1.8 vs. 9.1 ± 2.7; P = 0.38).

Nine months after MICN, patient 6 presented with a conjunctival cyst, 5 × 5 mm, at 12 o’clock limbus. The cyst was surgically excised. Histopathology of the excised cyst showed a conjunctival cyst lined by the stratified squamous epithelium, with amorphous material in the cyst cavity.

Discussion

The cornea is one of the most richly innervated organs. Normal innervation is vital for maintaining epithelial integrity in the cornea and wound healing. NK is a degenerative disease of the cornea characterized by the absence of corneal sensitivity and impaired epithelial healing. The early signs of NK are increased viscosity of tear mucus and punctate fluorescein staining of the corneal epithelium, which constitute the extent of Mackie’s stage I of NK. Mackie’s stage II is characterized by the acute loss of epithelium, with the defect surrounded by loose epithelium that forms a smooth edge. Folds develop in Descemet’s membrane as the stroma begins to swell. Stage III involves stromal lysis and eventually corneal perforation.

The causes of NK include various ocular and systemic conditions. The most common causes are herpes simplex and herpes zoster virus infections [15]. Intracranial space-occupying tumors and neurosurgical procedures that damage the ophthalmic branch of the trigeminal nerve can also result in this refractory disease. Chemical injuries, especially those caused by alkalis, lead to the most severe damage to the trophic cord. If the process reaches the deeper stroma, the corneal nerves are damaged, and the sensitivity may be markedly affected. Various treatments for NK (including preservative-free lubricants, autologous serum eye drops, and therapeutic contact lenses) can promote corneal epithelial regrowth over a neurotrophic corneal defect. However, conventional treatments for NK may increase the risk of disease recurrence because these treatments do not address the underlying injury of corneal innervation. Recently, the surgical procedure of corneal neurotization has been introduced as a surgical replacement for dysfunctional nerves in cases of unilateral NK. This can be either by direct nerve transfer or by indirect corneal neurotization, which involves the interposition of a nerve graft between a healthy donor nerve and the affected cornea.

In this study, nine patients with active NK underwent surgery. Preoperatively, the baseline volume of the corneal epithelial defect was 14.3 ± 6.1 mm [2]. The ulcers of all 9 patients healed within 3 months after surgery, which was maintained throughout the follow-up period. Compared with that in patients with multiple cranial nerve palsy after surgery, the epithelial defect in 6 patients with single fifth cranial nerve palsy was reduced substantially from baseline. Eyelid dysfunction in conjunction with neurotrophic corneal epithelium defects renders treatment more difficult. Abnormal eyelid blinking inhibits corneal epithelial growth in concert with suboptimal sensory innervation. Changes in eyelid structure and function need to be carefully assessed and treated to prevent exposure keratopathy and vision deterioration.

In this study, an intact epithelium was restored, and corneal transparency was maintained, with an improvement in visual acuity in some of our patients. Most of our patients with visual improvement were in the active stage. Yen reported that there was no significant difference in visual acuity after neurotization in their series. The reason was that most of their patients had corneal scars [16]. Other factors affecting postoperative visual acuity include amblyopia, large-angle esotropia, cataracts, and retinal detachment [17, 18]. In contrast, 3 of the 4 patients demonstrated significant improvement in visual acuity postoperatively in another study [19]. All of our patients were adults, and none of them had amblyopia or posterior ocular disease. Timely surgical intervention to stabilize the ocular surface to control the development of corneal scars is helpful for achieving better postoperative visual acuity. In patients whose deep stromal scars remain, keratoplasty secondary to corneal neurotization surgery is needed to restore corneal clarity and improve visual acuity.

IVCM is a useful clinical diagnostic tool for evaluating regenerated nerves during corneal neurotization. Preoperatively, the subbasal neural plexus was not visible by IVCM in any of the patients. After surgical neurotization, a small number of attenuated subbasal nerve fibres were present as early as 3 months after surgery. One year after surgery, corneal nerves in the subbasal layer were detectable in all patients; however, the quantity and quality of these nerves were poorer than those in the contralateral unaffected eye. This finding suggests that the recovery process of the regenerated nerves was not yet complete. The result of slower subbasal nerve recovery after multiple nerve injury than after single trigeminal injury is not clear. The higher prevalence of abnormal blinking, dry eyes, and altered tear film after multiple nerve injury than after single trigeminal nerve injury may be attributed to the slow recovery of corneal nerves. However, complete recovery of subbasal nerves does not seem to be necessary for corneal epithelial healing, and the limbal vessel subsides. All our patients had the corneal epithelium defects healed 3 months postoperatively, which suggests that only partial recovery of subbasal nerves is sufficient for maintaining homeostasis of the ocular surface and preventing the deterioration of corneal ulcers.

Vessels may invade the cornea from the limbal vascular plexus under pathological conditions. Corneal neovascularization may affect visual acuity via corneal scarring, oedema and lipid deprivation. It also alters the immune privilege of the cornea, thereby increasing the risk of graft rejection resulting from penetrating kertoplasty [20]. Corneal neovascularization occurs secondarily to postinfection, inflammation, keratoplasty, loss of the limbal stem cell barrier, and corneal denervation. Ferrari suggested that corneal nerves and vessels inhibit one another by reducing the number of angiostatic molecules expressed by the cornea, including pigment epithelial-derived factor (PEDF) and epithelial VEGFR3 [21]. In this study, corneal neurotization surgery was not only useful in facilitating corneal epithelial healing and halting stromal melts but also effective in occluding actively growing corneal blood vessels. Improving vascularized corneal recipient beds can reduce graft rejection by blocking the trafficking of graft-derived antigens to regional lymph nodes in subsequent keratoplasty [22, 23]. Thus, corneal neurotization may be helpful for long-term corneal graft survival in NK patients with penetrating keratoplasty.

Various techniques are employed in corneal neurotization. The advantage of direct corneal neurotization is the shorter regeneration distance could result in higher corneal sensation compared with indirect at early postoperative time points. With the use of a more proximal section of the donor nerve, indirect corneal neurotization potentially leading to increased innervation and enhanced corneal sensation. However, the long-term outcomes of direct and indirect neurotization are the same [10]. The drawback of direct corneal neurotization lies in the requirement for a wide facial dissection and extensive nerve manipulation. The sural nerve is a cutaneous sensory nerve of the posterolateral leg to the lateral ankle. The plastic surgeons are familiar with the technique to harvest the sural nerve graft. The use of a sural nerve graft as a conduit allows for the surgical flexibility to use a variety of donor sensory nerve, including the supraorbital nerve, supratrochlear nerve, great auricular nerve, and occipital nerves [24]. In our study, this procedure allowed the healing of NK and recovery of corneal sensations in all patients. In consideration of experience of the surgeon, the surgical technique of MICN is now considered an efficient and safe procedure in resource-limited hospitals.

One of the primary limitations of this study was it was a small retrospective study of 11 patients and short-term follow-up. In addition, the sensation of conjunctiva of the affected eye was not tested. In eyes with partial trigeminal nerve function, a topical recombinant human nerve growth factor, cenegermin (Oxervate, Dompe, Milan) may be an option to promote corneal nerve health and restore the ocular surface homeostasis. A major limitation to the use of the cenegermin in clinical practice is the cost of therapy. Another limitation is cenegermin did not demonstrate a significant improvement in central corneal sensation or bestcorrected visual acuity in clinical trials [25, 26].

In conclusion, this series of NK patients presented with a very harmful corneal condition with a poor prognosis for permanent corneal epithelium healing and ocular surface homeostasis with conventional management techniques. We observed the absence of the corneal nerve plexus preoperatively in all our patients. After surgery, favorable trophic changes, particularly in the subbasal nerve plexus, were observed in our patients. However, the nerve fibers are less in number as compared to a normal eye. Combination of MICN with cenegermin may enhance corneal lesion healing. We suggest that MICN is a useful alternative treatment for this difficult corneal degenerative disease.