Ocular surface reconstruction using novel regenerative medicine
Cultivated Limbal Epithelial Transplantation (CLET) for Limbal Stem Cell Deficiency
Case 1: Treatment of Unilateral Stem Cell Deficiency with Auto-CLET
Autologous cultivated limbal epithelial transplantation (i.e., auto-CLET) is an ideal surgical modality used for avoiding the risk of immunological reaction when treating unilateral cases . In such cases, a 2 × 3 mm limbal biopsy is performed, with the biopsy specimen then cultured on AM to obtain the cultivated sheet. The cultured sheet is then transplanted over the corneal surface after the scarred tissue is removed. Systemic steroids are then used for a maximum of 1–3 weeks postoperative for quick reduction of the surgically induced inflammation. Antibiotics and 0.1% betamethasone are used for 1–3 months postoperative and then tapered and switched to 0.1% fluorometholone to maintain the stable reconstructed ocular surface. A medical-use soft contact lens is used for 3–6 months postoperative to promote complete graft attachment and to prevent any physiological damage caused by a scarred lid margin or dry eye. For long-term postoperative maintenance a minimum dose of 0.1% fluorometholone and protective agents such as 0.1% sodium hyaluronate are used to protect the ocular surface.
The case shown in Fig. 23.2 was a 53-year-old male who suffered from unilateral chemical burn. For treatment, he underwent auto-CLET using a healthy limbal biopsy from his fellow eye. His corneal surface was successfully reconstructed via the cultivated autologous corneal epithelium, and his visual acuity (VA) recovered to 0.7. Although mild conjunctival invasion was observed at 14-years  postoperative, a clear optical zone was maintained in the transplanted corneal epithelium, thus suggesting a long-term supply of regenerative epithelial progenitor cells in the transplanted cell sheet.
Auto-CLET in a case of unilateral chemical burn. (a, b) presurgery; (c, d) 4 years post-CLET; (e, f) 14 years post-CLET
Case 2: Allogeneic Cultivated Limbal Epithelial Transplantation (Allo-CLET)
Allo-CLET is an effective procedure that utilizes donor corneal epithelial progenitor cells to enable reconstruction of the diseased host eye without the need of a biopsy [6, 7]. However, long-term intensive immunosuppression post surgery is essential to prevent immunological rejection. Acute-phase corneal-stem-cell depletion, such as in cases of severe chemical burn with persistent corneal epithelial defect that extends over a long period of time and without wound healing, should be considered the indication for allo-CLET . Moreover, allo-CLET. is sometimes indicated in bilateral cases, in which the most intensive treatment is required to obtain visual recovery, as well as unilateral cases, in which preservation of the healthy fellow eye is vital.
General preoperative examination is essential in order to exclude the risk of activation of any possible infections, such as viral hepatitis and obsolete pulmonary tuberculosis, by immunosuppression. Moreover, regular follow-up of the patient’s general condition via blood tests is required in order to uncover any drug-related complications such as hepatic and renal toxicity and cytopenia. Monitoring of the drug concentration in plasma is also important in order to adjust the amount of drug needed to maintain an effective level in the blood (Fig. 23.3).
Schema illustrating the standard postoperative treatment protocol for allo-CLET
The case of a 58-year-old male who sustained bilateral chemical burns is shown in Fig. 23.4 . Prior to surgery, his left eye showed total conjunctival invasion with minimum stromal scarring. No severe cicatrization was observed, and the tear film was well preserved. After undergoing allo-CLET, his VA recovered to 0.8. Following our standard protocol for allo-CLET, intensive immunosuppression was applied both topically and systemically. As a local treatment in such cases, 0.1% betamethasone should be used for as long as possible, except in cases with steroid-associated complications. The additional use of tacrolimus eye drops should be considered for cases with severe inflammation or steroid-induced high intraocular pressure (IOP). After sufficient stabilization, 0.1% betamethasone can be tapered and switched to 0.1% fluorometholone to reduce the risk of complications associated with long-term steroid use. It is vitally important to apply sufficient systemic immunosuppression for at least 1.5-years postoperative to prevent the rejection. For intensive immunosuppression, 125 mg of methylprednisolone is administered intravenously on the day of surgery and at 3-days postoperative. A combination of three immunosuppressive medications (i.e., steroid, cyclosporine, and mycophenolate mofetil) is systemically used. Daily administration of 1–2 mg of betamethasone is initially used for 2-weeks postoperative, and then switched to 5–15 mg prednisolone. Moreover, 2–3 mg/kg of body weight (BW) of cyclosporin is initially used until the ocular inflammation is sufficiently stabilized. The level of cyclosporin in the plasma should be monitored to prevent excessive absorption. The trough value should be adjusted to between 150 and 200 ng/mL to obtain effective immunosuppression. After reduction of the initial inflammation, 25–50 mg of low-dose cyclosporin is administered to the patient for a long-term immunosuppression of over 1.5 years. Blood tests should be routinely performed to monitor the liver and renal functions. Mycophenolate mofetil is an effective immunosuppressive agent that is suitable for preventing epithelial rejection, and it should be started at 100 mg per day, and then tapered to 500 mg following stabilization of the ocular inflammation. Since allo-CLET requires long-term immunosuppression post surgery, close monitoring of any systemic complications is essential. In the above-described case, the patient’s cornea remained clear at over 17-years postoperative (Fig. 23.4). There were no episodes of epithelial rejection, and his VA was maintained at 0.8 via a twice-daily topical application of 0.1% fluorometholone.
Long-term results post allo-CLET for bilateral chemical burn. (a) pre surgery (b) 4-years postoperative; (c) 12-years postoperative; (d) 17-years postoperative
Major Complications of Allo-CLET and Associated Solutions
Since allo-CLET requires long-term intensive immunosuppression post surgery, it is important to understand and prepare for any possible major complications that might occur. The primary disease, combined with the immunocompromised condition of the host, is associated with the risk of infection. Close monitoring of the bacterial flora in the conjunctiva is helpful for the selection of the proper antibiotics to successfully treat any growth of bacteria that might occur. Preexisting dry eye is one of the risk factors for epithelia-related problems, and strict adherence to the use of preservative-free artificial tears is crucial for proper maintenance of the epithelium.
Persistent Epithelial Defect
Epithelial defect is the most common problem when performing ocular surface reconstruction, and it has the possibility of developing into severe complications such as corneal ulceration, perforation, and infection. Prompt modification of the local treatment is crucial in order to promote epithelial healing and prevent further progression. To promote epithelial proliferation and migration, it is essential to minimize any possible drug-related toxicity. Switching to preservation-free eye drops and oral medication is a first step in the solution. A sufficient amount of artificial tears without preservatives should be supplied, and punctal plugs or occlusion should be considered. The use of an eye patch and temporal tarsorrhaphy is necessary for severe cases that are not effectively treated by local medications.