Limbal stem cell deficiency secondary to vitrectomy in the context of an autoimmune polyendocrine syndrome type

Purpose: The objective of this study was to describe the short-term results of allogenic transplantation of limbal stem cells expanded on amniotic membrane for the ocular surface reconstruction. Methods: Prospective nonrandomized, nonmasked study in a single ophthalmological center. Ten patients with bilateral total limbal stem cell deficiency (LSCD) were included. Expression and presence of ABCB5 and Δp63α in amniotic membrane-cultured limbal epithelial stem cells were analyzed, in relationship with clinical changes after allogenic transplantation. An objective evaluation was performed to determine corneal transparency and superficial vascularization. Results: In a median follow-up time of 11.6 months, 7 patients (70%) were considered as failure compared with the preoperative status. ABCB5 and Δp63α are expressed in similar amount in the limbal epithelial cells expanded in vitro and transplanted in patients with bilateral LSCD. Conclusions: Transplantation of allogenic epithelial limbal cells expanded in amniotic membrane could be considered in patients with LSCD due to burns or congenital etiologies such as aniridia, but its benefit is limited for patients with immunologic diseases.

Limbal stem cell deficiency (LSCD) is one of the leading causes of corneal damage. Injury or inflammation in the cornea causes LSCD, which may be unilateral or bilateral depending upon the cause. Limbal epithelial cell implants successfully improve vision in patients with chemical injury‑induced LSCD. Transplantation of cultured epithelial stem cells has become a treatment of choice for numerous patients with LSCD. Bilateral LSCD is frequently observed in the general population, where no residual stem cells are available for exvivo culture. Allografts are associated with a high risk of rejection, neoplasia, and disease transmission. In this respect, allogenic cell populations from other regions in the patient may substitute for allogenic material. In the present study, dental pulp stem cells were cultured in limbal stem cell media and these cells were characterized against limbal stem cells, revealing the significance of using dental pulp stem cell treatment in bilateral LSCD. The morphology and culture pattern of both limbal and dental pulp stem cells grown in limbal stem‑specific media were similar. Polymerase chain reaction analysis revealed that stem cell markers were highly expressed in limbal stem cells compared to in dental pulp stem cells, regardless of the medium and scaffold in which they were grown. Although dental pulp stem cell molecular expression is quite low at the transcript level, the functional protein level according to immunocytochemistry and western blot analyses demonstrated that stem cells and corneal differentiation molecule levels were quite high, indicating their potential as limbal stem cells in the respective microenvironment.

Limbal Stem Cell Deficiency—Demography and Underlying Causes

The review presents an analysis of clinical trials results for autologous cultured oral mucosal epithelium transplantation (COMET) in patients with bilateral corneal limbal stem cell deficiency (LSCD) over the past 15 years. Detailed characteristics and evaluation are given for anatomical outcomes, visual acuity changes, and complication rates. The results obtained during the analysis confirm the consistency of the concept of corneal re-epithelization by means of COMET. COMET promoted persistent corneal re-epithelization in 81.5% of cases, and visual acuity improvement in 78.8% of patients with LSCD. COMET does not require systemic immunosuppression, and it is accompanied by much smaller numbers and significantly lower grades of complications compared with keratoprosthesis. About 15% of patients experienced developing superficial peripheral corneal neovascularization regressed spontaneously by 12 months of observation. Based on the COMET clinical trials results, the management of patients with bilateral LSCD is under optimization by reference to the pathogenesis of the underlying disease. Thus, autologous cultured oral mucosal epithelium transplantation seems promising for further studies and introduction into routine clinical practice. Key words: limbal stem cell deficiency, ocular surface reconstruction, corneal epithelium, oral mucosal epithelium, transplantation.

Objective: To report a female patient with bilateral limbal stem cell deficiency (LSCD) due to primary adrenocortical insufficiency (PAI).Methods: Case report Results: A 40-year-old female patient had blurry vision, foreign body sensation, tearing, and photophobia for several years. On examination, corneal epithelial haze, surface irregularity, and superficial neovascularization were observed. There was a dull and irregular reflex from the conjunctivalized corneal surface. Medical history revealed that she had a diagnosis of PAI for 11 years and received hormone replacement (fludrocortisone acetate) therapy. With the clinical presentation and examination, the diagnosis was compatible with LSCD. Frequent ocular lubricant and topical steroid drops were initially started and topical cyclosporine treatment was planned for the long term. After 3 weeks, there was no corneal superficial neovascularization and epithelial haze, peripheral stromal haze was still observed. Conclusion: LSCD may rarely be associated with PAI. In patients with LSCD, systemic evaluation should be made to rule out PAI.

To evaluate the midterm outcomes of penetrating keratoplasty (PK) following allogeneic cultivated limbal epithelial transplantation (CLET) for bilateral total limbal stem cell deficiency (LSCD). Ten patients (10 eyes) with bilateral LSCD were enrolled in this prospective noncomparative case series study. Each participant underwent PK approximately 6mo after a CLET. Topical tacrolimus, topical and systemic steroids, and oral ciclosporin were administered postoperatively. Best-corrected visual acuity (BCVA), intraocular pressure (IOP), ocular surface grading scores (OSS), corneal graft epithelial rehabilitation, persistent epithelial defect (PED), immunological rejection, and graft survival rate were assessed. The time interval between PK and allogeneic CLET was 6.90±1.29 (6-10)mo. BCVA improved from 2.46±0.32 logMAR preoperatively to 0.77±0.55 logMAR post-PK (P<0.001). Kaplan-Meier analysis of mean graft survival revealed graft survival rates of 100% at 12 and 24mo and 80.0% at 36mo. PEDs appeared in 5 eyes at different periods post-PK, and graft rejection occurred in 4 eyes. The total OSS decreased from 12.4±4.4 before allogeneic CLET to 1.4±1.51 after PK. A sequential therapy design of PK following allogeneic CLET can maintain a stable ocular surface with improved BCVA despite the relatively high graft rejection rate.

Post-chemotherapy bilateral limbal stem cell deficiency

Introduction and importanceThis report describes a new technique of deep anterior lamellar limbo-keratoplasty for the management of bilateral limbal stem cell deficiency (LSCD) with corneal scarring.Presentation of casesA 45-year-old male presented with chronic sequelae of ocular chemical injury and had bilateral total LSCD with corneal scarring. The visual acuity (VA) in the right eye was counting fingers. A large diameter deep anterior lamellar limbo-keratoplasty (DAL-LK) was carried out and the donor cornea and limbus were sourced from a single tissue. The VA at the last visit, 2.5 years after the surgery was 20/80. A similar presentation was seen in a 31-year-old male with a VA of 20/320 in the right eye. He underwent a DAL-LK and 3 years after the procedure, the VA was 20/60. Both grafts remained clear with no episodes of rejection until the last follow up visit.DiscussionLimbal stem cell transplantation with keratoplasty or a keratoprosthesis is required to manage bilateral LSCD with stromal scarring. The former necessitates multiple interventions while the latter is associated with several globe threatening complications. DAL-LK was devised to overcome these disadvantages and offers a simple, single staged technique of simultaneously transplanting the corneal stroma with the limbal stem cells. As the host endothelium is preserved, there is no risk of rejection episodes.ConclusionDAL-LK can successfully restore stability of the ocular surface and visually rehabilitate cases with bilateral LSCD and stromal scarring. The procedure has stable long-term outcomes with a good safety profile.

Ocular surface reconstruction with ex vivo expanded limbal stem cells (LSCs) is a widely used clinical treatment for patients with limbal stem cell deficiency (LSCD). This is not applicable to patients with bilateral LSCD where there are no remaining LSCs. Cultivated oral mucosa epithelium (OME) has been used as an alternative source of autologous epithelial stem cells for ocular reconstruction in few clinical trials. However, successful generation of stratified OME epithelium has only been achieved in the presence of animal feeder cells and/or animal-derived products in the culture media, likely to contribute to increased risk of pathogen transmission and graft rejection. In this study, we report generation of multilayered OME epithelium that shares many of the characteristics of corneal epithelium using a fully compliant good manufacturing practice, feeder- and animal product-free method. Proof of concept was achieved by transplantation of autologous ex vivo expanded OME in two patients with histologically confirmed bilateral total LSCD that resulted in successful reversal of LSCD in the treated eye up to 24 months.

A 35-year-old woman who had undergone laser-assisted in situ keratomileusis in both eyes experienced bilateral total limbal stem cell deficiency (LSCD) due to chemical burns. Due to bilateral severe LSCD, allogenic simple limbal epithelial transplantation (SLET) from a human leukocyte antigen (HLA)-matched living related donor was the first choice of treatment for her left eye. We report the first case of HLA or ABO matching living related allogenic SLET for permanent restoration of the cornea for bilateral LSCD treatment. Our ABO-HLA-matched living related allogenic SLET alleviation of the systemic immunosuppressant to topical corticosteroids alone. It also came the limitations of prolonged systemic immunosuppressant usage in conjunctival-limbal allografts and keratolimbal allograft.

PurposeTo report the clinical outcomes of allogeneic cell-based therapy for bilateral corneal blindness due to limbal stem cell deficiency (LSCD).MethodsThis retrospective study included 28 eyes of 21 patients, at least...

Limbal stem cell (LSC) transplantation is the only efficient treatment for patients affected by LSC deficiency (LSCD). Allogeneic LSC transplantation is one of the most successful alternative for patients with bilateral LSCD. Nevertheless, the high variability of the human leukocyte antigens (HLA) remains a relevant obstacle to long-term allogeneic graft survival. This study characterized the immunologic properties of LSCs and proposed a genetic engineering strategy to reduce the immunogenicity of LSCs and of their derivatives. Hence, LSC HLA expression was silenced using lentiviral vectors encoding for short hairpin (sh) RNAs targeting β2-microglobulin (β2M) or class II major histocompatibility complex transactivator (CIITA) to silence HLA class I and II respectively. Beside the constitutive expression of HLA class I, LSCs showed the capability to upregulate HLA class II expression under inflammatory conditions. Furthermore, LSCs demonstrated the capability to induce T-cell mediated immune responses. LSCs phenotypical and functional characteristics are not disturbed after genetic modification. However, HLA silenced LSC showed to prevent T cell activation, proliferation and cytotoxicity in comparison to fully HLA-expressing LSCs. Additionally; HLA-silenced LSCs were protected against antibody-mediated cellular-dependent cytotoxicity. Our data is a proof-of-concept of the feasibility to generate low immunogenic human LSCs without affecting their typical features. The use of low immunogenic LSCs may support for long-term survival of LSCs and their derivatives after allogeneic transplantation.

Limbal stem cell (LSC) transplantation is the only efficient treatment for patients affected by LSC deficiency (LSCD). Allogeneic LSC transplantation is one of the most successful alternative for patients with bilateral LSCD. Nevertheless, the high variability of the human leukocyte antigens (HLA) remains a relevant obstacle to long-term allogeneic graft survival. This study characterized the immunologic properties of LSCs and proposed a genetic engineering strategy to reduce the immunogenicity of LSCs and of their derivatives. Hence, LSC HLA expression was silenced using lentiviral vectors encoding for short hairpin (sh) RNAs targeting β2-microglobulin (β2M) or class II major histocompatibility complex transactivator (CIITA) to silence HLA class I and II respectively. Beside the constitutive expression of HLA class I, LSCs showed the capability to upregulate HLA class II expression under inflammatory conditions. Furthermore, LSCs demonstrated the capability to induce T-cell mediated immune responses. LSCs phenotypical and functional characteristics are not disturbed after genetic modification. However, HLA silenced LSC showed to prevent T cell activation, proliferation and cytotoxicity in comparison to HLA-expressing LSCs. Additionally, HLA-silenced LSCs were protected against antibody-mediated cellular-dependent cytotoxicity. Our data is a proof-of-concept of the feasibility to generate low immunogenic human LSCs without affecting their typical features. The use of low immunogenic LSCs may offer support for long-term survival of LSCs and their derivatives after allogeneic transplantation..

Long-term Outcomes of Autolimbal and Allolimbal Transplants

Background. Bilateral limbal stem cell deficiency (LSCD) leads to visual impairment due to corneal epithelial deficiency and conjunctivalization. Penetrating keratoplasty (PKP) is often ineffective in LSCD, necessitating alternative epithelial reconstruction strategies. Autologous labial mucosal epithelium, sharing characteristics with corneal epithelium, presents a promising approach. Purpose. To evaluate the efficacy of a two-stage surgical approach for LSCD: paralimbal oral mucosal epithelial transplantation followed by PKP. Material and methods. Two patients with bilateral LSCD were evaluated and surgically treated at the S.N. Fedorov NMRC «MNTK «Eye Microsurgery». The first stage involved direct transplantation of autologous oral mucosal epithelial cells. The second stage comprised PKP in both clinical cases. Results. Complete epithelialization of the corneal surface after keratoplasty was observed 2-3 weeks after surgery and remained stable during the observation period. Conclusion. The two-stage approach (labial mucosal epithelial transplantation and PKP) can be considered as an alternative to keratoprosthesis in carefully selected patients with bilateral LSCD. Further studies are needed to evaluate the long-term efficacy and safety of this method.