角膜缘niche细胞与基质细胞维持角膜缘干细胞功能的对比研究

目的　比较角膜缘niche细胞（limbal niche cells，LNCs）与角膜缘基质细胞（limbal stromal cells，LSCs）在维持角膜缘干细胞功能上的不同特性。方法　将LNCs和LSCs分别从6个角膜缘组织分离，并在相同的条件下培养、传代。LNCs与LSCs经丝裂霉素C（mitomycin C,MMC）处理后分为LNCs组与LSCs组作为饲养细胞分别与角膜缘干细胞共培养，比较两组角膜缘干细胞克隆形成率（colony-forming efficiency,CFE）、上皮细胞复层化以及细胞标志物和部分基因的表达。结果　LNCs组角膜缘干细胞CFE（6.57±1.54）%高于LSCs组（1.43±0.47）%。 LNCs组细胞复层上皮数（4~5层）多于LSCs组（2~3层）。角膜缘干细胞克隆与免疫荧光染色及mRNA半定量分析结果显示，LNCs组比LSCs组表达了更多干细胞标志物ΔNp63，能更有效地维持角膜缘干细胞的细胞特性。逆转录PCR分析结果显示，LNCs组与LSCs组都分泌了一些维持角膜缘干细胞生长的生长因子，但LNCs组比LSCs组高表达上皮型钙黏蛋白（E-cadherin），低表达营养神经素3（NT3），能更好地支持角膜上皮增殖。结论　LNCs比LSCs能更好地支持角膜缘干细胞的生长及维持其干细胞特性。     

Cryopreserved limbal lamellar keratoplasty for peripheral corneal and limbal reconstruction

This study aimed to evaluate the outcomes and described the recovery process of cryopreserved limbal lamellar keratoplasty (CLLK) for peripheral corneal and limbal diseases. Thirteen eyes of 12 patients with a mean age of 41±23.9y were included. The average follow-up was 12.1±5.6mo. Stable ocular surface was achieved in all eyes at last follow-up. Epithelialization originated from both recipient and graft in 9 eyes. We conclude that CLLK compensates for the shortage of donor corneas and cryopreserved limbal grafts provide epithelialization sources in ocular surface reconstruction.     

Microkeratome-based limbal harvester for limbal stem cell transplantation: preliminary studies

PURPOSE: To develop a mechanical device to perform limbal transplantation. This procedure is a valuable surgical technique for management of limbal stem cell deficiency. However, the freehand dissection to obtain donor tissue is laborious and time-consuming. METHODS: A manual microkeratome (LSK One; Moria/Microtech, Doylestown, Pennsylvania) using a redesigned head (200 microm thickness, 16-mm blade) was used to create partial thickness corneoscleral caps from human donor globes. RESULTS: Corneoscleral caps obtained from human donor globes included a ring of approximately 1 mm of perilimbal sclera in contiguity to the cornea. Scanning electron microscopy showed a smooth cut surface with some chatter at the margins. CONCLUSION: This instrument ensures an effective and straightforward method to dissect the area where stem cells have been indirectly localized.     

Surgical treatment of limbal vernal keratoconjunctivitis by resection of a limbal lesion

PURPOSE: To report our experience managing a limbal mass associated with limbal vernal keratoconjunctivitis (VKC) by surgical excision. METHODS: A 16-year-old girl was referred to our hospital with the chief complaints of itching and photophobia in both eyes. She had a 2-year history of gradually enlarging limbal gelatinous masses on the temporal limbus in both eyes, which were diagnosed as limbal papillae of VKC. As the symptoms in her left eye were more severe, surgical resection of the limbal mass in her left eye was performed to relieve mechanical stress on the eyelid, followed by a free conjunctival autograft taken from the supranasal bulbar conjunctiva. RESULTS: After excision of the mass, ocular inflammation and other symptoms gradually disappeared. At 12-month follow-up, there was no recurrence of the mass. Findings in a histological examination of the excised limbal lesion were consistent with those of limbal papilla of VKC. CONCLUSION: In recalcitrant cases of limbal VKC, in which the symptoms are apparently caused by an elevated limbal mass, surgical excision of the limbal mass can be one of the therapeutic modalities, and it may facilitate resolution of the symptoms caused by chronic limbal VKC.     

Treatment of unilateral limbal stem cell deficiency syndrome by limbal autograft]

PURPOSE: To evaluate the improvement of the ocular surface after limbal autograft in patients with unilateral limbal stem cell deficiency related to chemical burns. MATERIALS AND METHODS: Limbal autograft was performed in five patients with unilateral limbal stem cell deficiency related to chemical burns. Thereafter, four patients underwent penetrating keratoplasty. The limbal graft was obtained from the fellow eye, and was secured with interrupted sutures. Patient follow-up ranged from 10 to 47 months. Limbus and corneas were studied by means of light microscopy. RESULTS: All five patients reported subjective improvement. Vascularization decreased in one cornea. Visual acuity improved in one eye and did not change in the remaining four eyes. After penetrating keratoplasty, graft reepithelialization was achieved after respectively 3, 4, 21, and 30 days. Light microscopy showed the presence of goblet cells in the limbal epithelium in four cases. After limbal autograft, the corneal epithelium was devoid of goblet cells in three out of four cases. CONCLUSION: Limbal autograft improves the ocular surface and the prognosis of subsequent penetrating keratoplasty in patients with unilateral limbal stem cell deficiency related to chemical burn.     

Adult limbal xanthogranuloma.

An 18-year-old white man presented with a non-painful yellow raised swelling on the inferior limbus of his right eye. Systemic and ocular examination revealed no other abnormalities. The lesion was dealt with by simple excision, but when it recurred fairly soon it was removed in toto and replaced by a lamellar graft, without recurrence. Histological examination revealed a typical xanthogranuloma. The question is, why should a healthy male with no other manifestations develop a lesion like this on his limbus?     

Conjunctival and limbal tumours

One hundred cases of histologically proven corneal and conjunctival tumours were extracted from the records at the Princess Alexandra Hospital and from the senior author's private practice in the last ten years. The tumours were analysed according to Duke-Elder's classification. There were 78 epithelial tumours, 15 pigmented naevi, four melanomas, two dermolipomas and one limbal dermoid. Fourteen of the 41 epithelial carcinomas were followed for three years or more. None of the 14 recurred.     

Amniotic membrane transplantation with conjunctival limbal autograft for total limbal stem cell deficiency

PURPOSE: To evaluate the outcomes of corneal surface reconstruction with conjunctival limbal autograft when combined with amniotic membrane transplantation on both the donor and recipient eyes. DESIGN: Retrospective, noncomparative, interventional small case series. PARTICIPANTS: Five eyes of five patients with total limbal stem cell deficiency (LSCD) resulting from pseudopemphigoid (n = 1), chemical burns (n = 3), and extensive removal of conjunctival intraepithelial neoplasia (n = 1) were operated on by one surgeon (SCGT). INTERVENTION: After the removal of fibrovascular pannus from the corneal surface, two conjunctival limbal free grafts were harvested from the fellow eyes in all five patients with unilateral LSCD. Amniotic membrane, with the basement membrane side up, was grafted onto the defect created at the donor site and onto the recipient corneal and limbal sclera before placement of conjunctival limbal grafts. MAIN OUTCOME MEASURES: Symptomatic relief, improvement in visual acuity, fornix deepening, and rapid healing and restoration of normal cornea and limbus in the recipient and donor eyes were assessed. RESULTS: During the mean follow-up of 22 months (range, 11-48 months), all eyes experienced symptomatic relief. All recipient eyes had a mean improvement in visual acuity of nine lines (range, 7-12). The three eyes with stromal vascularization showed regression, and all recipient eyes had marked improvement in corneal clarity. Three eyes receiving simultaneous symblepharon lysis and fornix reconstruction successfully regained deep, stable fornices. The donor eyes showed rapid healing and restoration of the normal limbal landmark, even in one eye where nearly the entire limbus was removed. CONCLUSIONS: Limbal conjunctival transplantation is an effective procedure for restoring the corneal surface integrity in eyes with total LSCD. The additional use of amniotic membrane may contribute to a higher rate of success in the recipient eye and a lower rate of complications in the donor eye, as well as allow the simultaneous correction of concomitant cicatricial abnormalities.     

Evaluation of a microkeratome-based limbal harvester device for limbal stem cell transplantation.

PURPOSE: To assess the cut quality and reproducibility using a novel microkeratome-based limbal harvester. METHODS: An enlarged microkeratome head and stainless steel blades were coupled with a nitrogen gas-driven turbine (15,000 blade oscillations/min) of a microkeratome. A large, 16-mm-diameter suction ring was attached to the globe. A lamellar sclerokeratectomy using head depths of 170 and 200 microm was performed in human donor research eyes. Obtained lenticule thickness was measured by ultrasound pachymetry and the bed size by planimetry. Histologic and scanning electron microscopy (SEM) analyses of the samples were performed. RESULTS: Central lenticule thickness was 294 microm (standard deviation [SD] 37) for the 170 head and 277 microm (SD 91) for the 200 head (p = 0.720). Lenticule diameter was larger in the horizontal meridian using the 170 head (12.8 mm [SD 0.8] vs. 11.9 mm [SD 0.7], p = 0.028), but similar in vertical meridian (12.0 [SD 0.6] versus 11.4 mm [SD 0.6], p = 0.093). Histology showed a multilayer epithelial cell pattern at the lenticule periphery. The SEM showed a smooth cut surface in both the stromal bed and the lenticule. CONCLUSION: Cut reproducibility and quality are similar to those found using standard microkeratomes for corneal lamellar cuts. This system ensures, in a straightforward way, the presence of epithelial cells in the edges of a mechanical sclerokeratectomy for limbal stem cell transplantation.     

Simultaneous deep anterior lamellar keratoplasty and limbal allograft in bilateral limbal stem cell deficiency

Purpose  

To report the efficacy of simultaneous keratolimbal allograft (KLAL) surgery and deep anterior lamellar keratoplasty (DALK) for limbal stem cell deficiency (LSCD).     

角膜缘干细胞理论的近期进展

本文从四个方面对角膜缘干细胞理论及干细胞理论的应用进行较为详细的阐述:(1)角膜缘干细胞理论的起源和确立;(2)角膜缘干细胞理论的实验、生理、病理以及临床证据;(3)角膜缘干细胞理论的应用;(4)角膜缘干细胞理论在应用方面存在的问题及应用前景.     

自体角膜缘移植术治疗角膜缘缺陷症的实验与临床研究

目的探讨自体角膜缘移植术在局限性或全角膜缘缺陷症治疗中的作用。方法动物实验以兔眼前节碱烧伤制作全角膜缘缺陷症模型,烧伤后实施自体角膜缘移植术治疗。结果动物手术组角膜上皮愈合快,角膜新生血管及炎细胞浸润均轻于对照组。临床研究中所有翼状胬肉均未见复发。结论自体角膜缘移植术治疗角膜缘缺陷症效果显著,是一种符合生理机制的、安全的手术方式。     

Ipsilateral limbal translocation for treatment of partial limbal deficiency secondary to ocular alkali burn

BACKGROUND/AIM: Chemical ocular burn, especially by alkali, may result in damage to the limbal stem cells, fundamental in maintaining the integrity of ocular surface. Clinically, patients manifest abnormal conjunctival-corneal healing with peripheral neovessels, chronic epithelial defect, severe stromal inflammation, and conjunctivalisation of the corneal, which is characteristic of limbal deficiency. Penetrating keratoplasty has a poor prognosis in these cases. Available therapeutic options included limbal autograft and allograft transplantation associated or not with human amniotic membrane transplant. The authors propose an alternative treatment with ipsilateral autologous limbal transplantation, transferring the healthy portion of the limbus from the partially burned eye to the deficient area in patients with partial limbal deficiency secondary to alkali burn. METHODS: Ipsilateral limbal autografts were performed in five patients with partial limbal deficiency secondary to alkali burn, transferring the healthy portion of the limbus to the burned area of the same eye, without intervening in the contralateral eye. RESULTS: All cases had regression of the limbal deficiency with improvement of visual acuity to a minimum of 20/60-20/20 (partial), without complications during the follow up period (range 7-11 months). CONCLUSION: Ipsilateral autologous limbal autograft is a therapeutic option for partial limbal deficient eyes.