The effect of allograft rejection after penetrating keratoplasty on central endothelial cell density

We identified all patients who had undergone penetrating keratoplasty for keratoconus and were being observed longitudinally (N = 174). During the follow-up period, 57 of 174 patients (33%) showed evidence of an allograft rejection episode, which occurred at an average of eight months after the operation. We analyzed specular photographs of the corneal endothelium, taken before and after the first allograft rejection episode. A significant decrease in endothelial cell density was observed (11.8%, P less than .0001). As a control, we analyzed all available specular photographs from patients with keratoconus who showed no evidence of allograft rejection after penetrating keratoplasty. The observed endothelial cell density decrease (11.8%) in patients with keratoconus undergoing allograft rejection exceeded that found in the control subjects (6.8%) during a comparable time period (P = .06). Severe allograft rejection episodes resulted in a decrease in endothelial cell density that exceeded expected loss significantly (14.8% compared with 6.9%, P = .01), whereas mild allograft rejection episodes were not associated with a loss in endothelial cell density exceeding that expected (1.8% compared with 6.5%, P = .34).     

穿透性角膜移植术后内皮型免疫排斥反应的临床研究

目的探讨穿透性角膜移植术(PKP)后内皮型免疫排斥反应发生的动态变化,及对角膜植片透明性的影响。方法对1994年1月至1998年12月在我院行PKP术并有完整记录的患者648例(648只眼),男444例(444只眼)、女204例(204只眼)进行随访,统计术后05、1、3、6、9、12、18、24、36、48个月等10个不同时间内皮型免疫排斥的发生率,以及植片混浊的发生率,比较两者动态变化的相关性,制成动态变化曲线,并对术前的病因与术后内皮型免疫排斥反应发生之间的关系进行分析。结果植片混浊与内皮型免疫排斥反应密切相关(线性相关回归分析,P<001);术后3年,内皮型免疫排斥反应的发生率仍有一定比例。术前不同病因及炎性背景的眼病,因内皮型免疫排斥反应发生率不同,而影响植片透明性。结论PKP术后,植片的透明性主要受内皮型免疫排斥反应的影响,二者密切相关;各种角膜疾病因炎性反应不同,其内皮型免疫排斥反应的发生率也不同,重视术后患者的长期随访,对维持术后角膜植片的透明性,保证手术成功率有重要意义。(中华眼科杂志,2005,41145149)     

环孢霉素A滴眼预防高危角膜移植免疫排斥的效果

目标 探讨环孢霉素Ａ（ＣｓＡ）联合地塞米松局部滴眼预防高危穿透角膜移植（ＰＫＰ）免疫排斥反应的效果。方法 将７７眼高危ＰＫＰ分为ＣｓＡ组（３７眼）;２％ＣｓＡ蓖麻油溶液联合地塞米松滴眼;对照组（４０眼）单独使用地塞米松滴眼。采用生存曲线进行比较。结果 术后２年,两组植片存活率分别为８９％和６０％,差异有显著性（Ｐ〈０．０５）。结论 ２％ＣｓＡ蓖麻油溶液联合地塞米松溶液局部滴眼是预防高危ＰＫＰ免疫排     

Hydrolase participation in allograft rejection in rat penetrating keratoplasty

Background: A rat model of orthotopic corneal graft rejection was used to investigate the alterations in hydrolase activity within the corneal graft or within cellular infiltrates during acute rejection. Methods: The distribution of the lysosomal enzymes [acid phosphatase (AP), N-acetyl-ß-D-glucosaminidase (NAG), ß-glucuronidase (R-Glue), ß-galactosidase (ß-Gal), dipeptidylpeptidase II (DPPII)] and of the membrane-bound proteases [aminopeptidase M (APM), aminopeptidase A (APA), -glutamyltransferase (GGT), alkaline phosphatase (ALP), dipeptidylpeptidase IV (DPPIV)] were investigated by histochemical methods in the grafts at 3, 5, 8, 10 and 12 days following allogeneic transplantation. Serial sections of the grafts were also examined for RT1b, CD4, CD4+, CD8, CD11b/c and CD45, in order to determine hydrolase activity within infiltrating cells. Results: Allogeneic grafts were invaded by macrophages, CD4- and CD8-positive lymphocytes. In contrast, syngeneic grafts, performed as a control, contained occasional lymphocytes and focal aggregations of macrophages around suture sites. The allogeneic cellular infiltrate stained intensely for AP and ALP; moderately for ß-Gluc, NAG and ß-Gal; and mildly for GGT, DPPII and APM in grafts at all postoperative times. Serial sectioning indicated that the majority of the lysosomal hydrolases were located in macrophages; AP, APM and GGT were, however, observed in lymphocytes. Vessel ingrowth could be observed with enzyme staining for AP, ß-Gluc, NAG, ALP, APA and APM. Hydrolase activity in the corneal endothelium served as an indicator of endothelial function during the rejection process. Conclusion: Changes in normal hydrolase activities in corneal grafts in the rat model indicate decreasing corneal function during the rejection process. Hydrolases released from infiltrating cells contribute to the morphological disruption and, possibly, to graft rejection.     

穿透性角膜移植术后慢性失功移植片的免疫学研究

目的检测穿透性角膜移植术后慢性失功移植片的免疫与非免疫相关细胞因子表达的变化,探讨免疫与非免疫因素在角膜植片慢性失功中的作用机制。设计临床实验研究。研究对象分为两组,角膜植片慢性失功（CCAD）组：穿透性角膜移植（PK）术后因CCAD导致移植失败的8例患者的角膜植片;正常对照组：由山东跟库提供的正常角膜供体3只。方法免疫组化法检测两组角膜片免疫相关细胞因子表达,结合患者的临床资料进行综合分析。主要指标角膜中CD4^＋、CD8^＋、F4／80、TGF-β、bFGF、α-SMA的表达。结果免疫组化检测显示正常对照组：角膜各层组织 、α-SMA、bFGF表达阴性,F4／80角膜基质偶见表达,TGF-β在角膜上皮层有表达。CCAD组：所有角膜植片全层均术见CD4^＋及CD8^＋T淋巴细胞浸润;5例失功植片基质层F4／80阳性表达,其余3例角膜植片全层表达阴性;所有角膜植片上皮层可见TGF-β阳性表达,基质层内可见TGF-β表达。所有失功角膜片基质层bFGF表达呈弱阳性;所有患者角膜基质内α-SMA表达阳性,后弹力层附近可见较强表达。结论穿透性角膜移植术后慢性失功移植片免疫组化检查未发现支持临床急性免疫排斥反应的证据,抗原递呈细胞及非免疫特异相关细胞因子的异常表达提示免疫与非免疫因素参与了CCAD的发生和发展。     

穿透性角膜移植排斥反应的临床分析

目的探讨导致穿透性角膜移植术后排斥反应的高危病种及危险因素。方法对86例90眼穿透性角膜移植患者术后排斥反应发生率进行回顾性分析。结果各病种排斥反应发生率不同，角膜溃疡为42．86％(9／21)；粘连性角膜白斑为39．13％(9／23)；单纯性角膜白斑为28．57％(8／28)；角膜变性或营养不良为11．11％(1／9)；先天性角膜白斑为100．00％(1／1)；圆锥角膜5眼及大泡性角膜病变3眼无排斥反应发生。术前角膜新生血管形成、虹膜前粘连、术前高眼压、术后高眼压、无晶状体状态、术式增多、大植片使排斥反应发生率增高。结论高危病种有角膜溃疡、粘连性角膜白斑、单疱病毒性角膜炎及化学伤导致的单纯性角膜白斑。术前角膜新生血管形成，虹膜前粘连，术前、术后高眼压，无晶状体，多术式联合，大植片是排斥反应发生的危险因素。     

Immunologic rejection of the central graft after limbal allograft transplantation combined with penetrating keratoplasty

PURPOSE: To study the incidence and prognosis of immunologic rejection of the central graft after limbal allograft transplantation (keratolimbal allograft transplantation [KLAT]) combined with penetrating keratoplasty (PKP). METHODS: Endothelial rejection in central penetrating graft after simultaneous KLAT and PKP using the same donor cornea was retrospectively studied. Incidence, reversibility, prognosis, and changes in limbal grafts were examined. RESULTS: Forty-five eyes underwent simultaneous PKP and KLAT. Endothelial rejection of the central graft was found in 16 eyes (35.6%). At last examination, 10 grafts (62.5%) restored clarity after immunosuppressive therapy. During rejection episodes, four eyes showed engorgement of vessels in limbal grafts, which existed before the episodes. Only one eye developed neovascularization with mild edema of the limbal grafts; however, no other limbal grafts showed abnormalities on biomicroscopy. No epithelial changes were noted, and 14 grafts (87.5%) were covered by corneal epithelium after the rejection. CONCLUSION: Approximately one third of eyes had endothelial rejection in the central graft after simultaneous KLAT and PKP. Abnormalities suggestive of rejection in the limbal grafts were seldom observed in these eyes, suggesting that immunologic response was different in central and limbal grafts.     

高危角膜移植术后免疫排斥反应规律的临床研究

目的 探讨不同病因的高危移植患者行穿透性角膜移植术后免疫排斥反应发生的规律。方法 对６８０例行穿透性角膜移植术中的１２４例１３４只眼高危移植患者术后排斥反应时间、排斥反应发生率及反应类型进行观察。结果 １３４只眼高危角膜移植术后免疫排斥反应发生率在１２％～７５％不等,其中角膜植床严重新生血管化者的排斥率最高。排斥发生时间最早为术后１３天,术后３～６个月为排斥反应发生高峰。排斥反应类型以内皮型和上皮型最多。结论 不同高危因素的穿透性角膜移植术后排斥反应发生率不同,排斥反应发生的时间和类型也不同。     

Risk of endothelial rejection after bilateral penetrating keratoplasty

Using univariate and multivariate survival analyses, the risk of endothelial rejection (ER) was compared between a group of patients undergoing penetrating keratoplasty (PK) in one eye only (unilateral PK) and a concurrent group who underwent PK in both eyes (bilateral PK). There was no significant difference in the time-related risk of ER between the bilateral and unilateral PK groups, after controlling for the increased risk of ER associated with younger age and preoperative stromal vascularity. Further analysis showed no increased ER risk to the first eye after the second eye's PK in the bilateral PK group. The documentation of simultaneous ER episodes in both eyes of two bilateral PK patients, however, may be evidence of an uncommon sharing of histocompatibility antigen(s) in the donor tissue used in these eyes.     

Clinical presentation, risk factors and treatment outcomes of first allograft rejection after penetrating keratoplasty in early and late postoperative period

Purpose

Corneal graft rejection is the most common reason for the failure of an allograft corneal transplant. We undertook this study to identify and compare risk factors and treatment outcomes for early and late corneal graft rejections after optical penetrating keratoplasty.

Methods

A retrospective case file analysis of 880 primary penetrating keratoplasties was performed at a tertiary ophthalmic care centre. Patients were divided into early rejectors (rejection episode within 6 months postoperatively) and late rejectors (rejection episode after 6 months postoperatively). Main parameters evaluated were demographics, preoperative diagnosis and clinical signs, donor tissue details, surgical technique, details of rejection episode, treatment, and outcome information.

Results

A total of 156 patients with rejection episodes were identified. Of these, 42 (26.9%) patients experienced early rejection episodes and 114 (73.1%) patients experienced late rejection episodes. Preoperative donor and recipient characteristics, surgical technique, and clinical presentation of graft rejection were found to be similar between both groups after a Bonferroni correction was applied (P>0.005). Treatment outcomes of graft rejections were not significantly different (P=0.46) between early and late rejectors, with 83% of patients responding to rejection treatment (80% early rejectors; 85% late rejectors).

Conclusion

Patients with early and late graft rejection have similar characteristics and both groups respond to treatment equally.     

建立小鼠角膜植片慢性失功模型的初步探讨

目的探讨建立小鼠角膜植片慢性失功(CCAD)的模型.方法 实验研究.将雄性C57BL/6小鼠与雌性BALB/c小鼠杂交,获得抗原半相合CB6FI代(即F1代)小鼠.分别以C57BL/6(异系)、F1代、BALB/c小鼠(同系)为供体,以BALB/c小鼠为受体,建立免疫背景渐同的小鼠穿透性角膜移植(PK)模型,BALB/c小鼠为正常对照组.PK术后体外对植片进行观察;采用茜素红联合碘化丙啶(PI)/Hoechst双标法观察角膜内皮细胞形态、凋亡与坏死情况;免疫组织化学法检测各组植片CD4+、CD8+T淋巴细胞浸润情况;电镜观察各组角膜片超微结构改变,筛选与临床接近的CCAD模型.生存曲线之间比较采用Log-rank检验.结果 (1)角膜植片观察:异系、F1代、同系和对照组小鼠发生混浊时间的中位数分别为17.0 d、85.5 d、＞100 d及＞100 d(F=344.0,p＜0.01).(2)免疫组织化学检查:异系移植组植片基质中见大量CD4+T淋巴细胞浸润,较多CD8+T淋巴细胞浸润;F1代移植组和同系移植组术后偶见CD4+T淋巴细胞浸润,未见CD8+T淋巴细胞浸润.(3)茜素红联合PI/Hoechst检查:发现异系移植组有较多坏死和凋亡的内皮细胞;F1代移植组内皮细胞减少,可见散在凋亡内皮细胞,未见坏死内皮细胞;同系移植组角膜内皮细胞较正常减少,偶见凋亡内皮细胞,未见坏死内皮细胞.(4)透射电镜检查:各移植组小鼠角膜植片内皮细胞均存在萎缩性改变,异系移植组角膜基质内可见较多炎性细胞;F1代和同系移植组未见明显炎性细胞浸润.结论 PK术后F1代移植组与同系移植组植片出现与临床慢性失功相似的变化过程,可作为研究CCAD的动物模型.

Abstract：

Objective To establish a murine model of chronic corneal allograft dysfunction (CCAD) after penetrating keratoplasty (PK). Methods Experimental study. PK model in mice:Semiallogeneic CB6F1 mice were obtained from matching of female BALB/c and male C57BL/6 mice.C57BL/6 (al logeneic group), CB6F1 (semiallogeneic group) and BALB/c (syngeneic group) grafts were transplanted orthotopically to BALB/c recipients respectively, and BALB/c mice as a control group. The follow-up time was more than 100 d, and graft survival time and corneal opacity score were monitored, and corneal endothelium were examined by alizarin red and PI/Hoechst stain. CD4 + and CD8 + T lymphocytes were examined by immunohistochemistry. Ultrastructure changes of the grafts were examined by electromicroscopy. Log-rank test were used to compare survival curves. Results ( 1 ) Graft examination:Median graft survival times were 17.0 d, 85.5 d, ＞ 100 d and ＞ 100 d in allogeneic, semiallogeneic,syngeneic and control groups, respectively ( F = 344. 0, P ＜ 0. 01 ) . ( 2 ) Immunohistochemistry examination: There were large amount of CD4 + and CD8 + T lymphocyte infiltration in allografts in allogeneic group at 3 weeks after PK; Few CD4 + and CD8 + T lymphocytes were observed in semiallogeneic group and syngeneic goups at 3 weeks after PK; CD4+ and CD8+ T lymphocyte infiltration was not observed in the control group. (3) Endothelium examination: The endothelium can not be counted because the blurred image after the alizarin red combined PI/Hoechst stain and apoptotic and necrotic cells can be seen in allogeneic group; the endothelial cell density decreased and few apoptosis can be detected in semiallogeneic and syngeneic groups; no apoptotic and necrotic endothelial cells were found in the control group.(4) Ultrastructural characteristic changes mainly include fibrosis formation and endothelium atrophy and degeneration in failed grafts in all transplanted groups by electron microscopy examination. Inflammation cells can only be found in the allogeneic group. Conclusions Semiallogeneic and syngeneic transplantation groups present the changes similar to CCAD in clinical study, and both can be regarded as the model that permits molecular evaluation of CCAD.     

Risk factors of rejection after penetrating keratoplasty: a retrospective monocentric study

Graefe's Archive for Clinical and Experimental Ophthalmology - To assess risk factors of rejection after penetrating keratoplasty (PKP). This retrospective monocentric study assessed risk...     

穿透性角膜移植术后角膜植片内皮慢性失功的临床分析

目的探讨圆锥角膜行穿透性角膜移植（PK）术后角膜植片内皮慢性失功的临床特点。设计回顾性病例系列。研究对象圆锥角膜患者PK术后角膜植片内皮慢性失功148例（163眼）。方法所有患者实施PK术,采用非接触式角膜内皮显微镜、A型超声测厚仪和眼前节OCT扫描系统对角膜植片进行评价。主要指标中央角膜内皮细胞密度（ECD）、六边形细胞百分率、内皮细胞丢失率、内皮细胞形态、中央角膜厚度（CCT）及最佳矫正视力。结果PK术后3年内患者植片ECD呈下降趋势,内皮细胞年丢失率为14．2％±10．9％。术后1．5年较术后1年的ECD有明显下降（舟O．002）。术后不同时间点的植片内皮六边形细胞百分率均在50％以上,CCT在500μm以上,各组间比较无统计学差异（辟10．869）。PK术后5年以上复查,患者角膜植片ECD仍呈下降趋势,但个体差异性较大,多数患者的六边形细胞百分率仍在50％以上,内皮细胞面积增大是局部性的,CCT保持在500μm以上。150例未发生排斥的角膜植片除2例发生植片混浊外,其余均保持透明,患者术后最佳矫正视力提高。结论ECD的超生理下降和局部内皮细胞面积增大是PK术后角膜植片内皮慢性失功的主要临床表现。内皮慢性失功的发展存在个体差异性,且在同一个体不同时期发展程度亦有不同。     

Mooren's ulcer and evidence of stromal graft rejection after penetrating keratoplasty.

A Mooren's ulcer developed in a 6-year-old girl after a penetrating keratoplasty for Peters' anomaly. A destructive, circumferential, and centripetal stromal ulceration recurred despite conjunctival resection, corneal gluing, topical and systemic administration of corticosteroids, and repeat graftings. Results of a systemic and a rheumatologic examination were unremarkable. Histopathologic evaluations of donor grafts consistently disclosed a lymphocytic and a plasma cell infiltrate. Indirect immunofluorescent staining using a normal donor cornea substrate was positive at the level of Bowman's membrane and stroma. The patient's lymphocytes were stimulated by a partially purified bovine corneal antigen and a positive antibody titer was found in the patient's sera to the same corneal antigen. These results suggest humoral and cell-mediated immune mechanisms can be involved in the initiation and perpetuation of a stromal rejection process after penetrating keratoplasty.     

角膜移植术后角膜免疫排斥反应的临床探讨

目的 探讨角膜移植术后角膜免疫排斥反应的有效防治方法。方法 对穿透角膜移植术后发生角膜免疫排斥反应的68例(68眼)进行回顾性总结。 结果 经统计学分析结果表明:角膜化学烧伤、角膜新生血管、大植片移植、术前活动性角膜炎症、再次移植等均为穿透角膜移植术后免疫排斥反应的主要因素。 结论 免疫排斥反应的发生与术前高危角膜移植有着非常密切的关系。     

Corneal astigmatism after penetrating keratoplasty. The role of suture technique

A randomized clinical trial was conducted to contrast two techniques of suturing in penetrating keratoplasty (PK) surgery: double running 10-0 and 11-0 sutures (DR), and a combination of 12 interrupted 10-0 sutures with a single running 11-0 suture (IR), followed by selective suture removal. The primary outcome evaluated in the 60 patients within each group was keratometric astigmatism. A decreasing trend in astigmatism over postoperative year 1 was observed only in the IR group (from 4.00 diopters [D] at 3 months to 2.50 D at 12 months). The difference in median astigmatism at 1 year (IR, 2.50 D; DR, 4.00 D) approached statistical significance (P = 0.06, Mann-Whitney U test). Both groups showed comparable steepening of almost 1 D during postoperative year 1. Assessment of the rate of visual rehabilitation was limited by a greater proportion of IR patients showing cystoid macular edema (CME) after surgery. These results, while favorable toward the IR/selective suture removal technique, must be substantiated by a final assessment after all sutures have been removed.     

穿透性角膜移植术后慢性失功移植片的超微结构观察

目的探讨穿透性角膜移植术后慢性失功能移植片的超微结构改变及发生机制。方法角膜植片慢性失功（CCAD）组：穿透性角膜移植（PK）术后因CCAD导致移植失败的12只（12例患者）角膜植片;12例患者两次PK手术的平均间隔时间为69个月,10例患者初次PK术后曾发生1次或以上免疫排斥反应,2例患者未发现免疫排斥反应;正常对照组：由山东省眼库提供的5只角膜植片作为正常供体。对两组角膜片行组织病理学、透射电镜及扫描电镜检查,结合患者的病史进行综合分析。结果透射电镜观察发现CCAD组较正常对照组角膜上皮层变薄,可见大空泡形成,角膜基质层纤维排列紊乱,无明显炎性细胞浸润;后弹力层与角膜内皮细胞之间可见异常的间隙及纤维增生;角膜内皮层萎缩变薄,细胞变形、核染色质浓缩,偶见炎性细胞与角膜内皮细胞黏附。扫描电镜观察发现CCAD组较正常对照组角膜上皮细胞微绒毛数量明显减少,暗细胞增多;角膜内皮细胞数量减少,存在缺失区,内皮细胞可见凋亡小体。结论CCAD植片特征性超微结构改变是内皮细胞的萎缩性改变和非炎性细胞成分的纤维增生。慢性亚临床的抗原依赖与非抗原依赖因素可能共同参与了CCAD的发生,免疫排斥反应可能诱导和促进了CCAD的发生、发展。