Small-diameter acellular porcine corneal stroma for peripheral corneal ulceration treatment

AIM:To evaluate the clinical efficacy of small-diameter acellular porcine corneal stroma(SAPS)for the treatment of peripheral corneal ulceration(PCU).METHODS:This retrospective clinical study included 18 patients(18 eyes)with PCU between April 2018 and December 2020.All patients had PCU and underwent lamellar keratoplasty with SAPS.Observation indicators included preoperative and postoperative best-corrected visual acuity(BCVA)and transparency of SAPS.The infection control rate in the surgical eye-lesion area was also calculated.RESULTS:Eighteen patients underwent lamellar keratoplasty with SAPS to treat PCU.None of the patients experienced rejection after 6mo(18/18)and 12mo(16/16)of follow-up.The BCVA(0.47±0.30)at the 6mo followup after operation was significantly improved compared with the baseline(0.99±0.80),and the difference was statistically significant(Z=-3.415,P<0.05).The BCVA at the 12mo follow-up after operation was not statistically significant compared to the 6mo(Z=0,P=1).With time,the SAPS graft gradually became transparent.At the 6mo(18/18)and 12mo(16/16)follow-up,none of the patients had recurrent corneal infection.CONCLUSION:SAPS is clinically effective in the treatment of PCU,improving the patient’s BCVA and reducing the incidence of rejection after keratoplasty.

Establishment of an untransfected human corneal stromal cell line and its biocompatibility to acellular porcine corneal stroma

AIM: To establish an untransfected human corneal stromal (HCS) cell line and characterize its biocompatibility to acellular porcine corneal stoma (aPCS). METHODS: Primary culture was initiated with a pure population of HCS cells in DMEM/F12 media (pH 7.2) containing 20% fetal bovine serum and various necessary growth factors. The established cell line was characterized by growth property, chromosome analysis, tumorigenicity assay, expression of marker proteins and functional proteins. Furthermore, the biocompatibility of HCS cells with aPCS was examined through histological and immunocytochemistry analyses and with light, electron microscopies. RESULTS: HCS cells proliferated to confluence 2 weeks later in primary culture and have been subcultured to passage 140 so far. A continuous untransfected HCS cell line with a population doubling time of 41.44 hours at passage 80 has been determined. Results of chromosome analysis, morphology, combined with the results of expression of marker protein and functional proteins suggested that the cells retained HCS cell properties. Furthermore, HCS cells have no tumorigenicity, and with excellent biocompatibility to aPCS. CONCLUSION: An untransfected and non-tumorigenic HCS cell line has been established, and the cells maintained positive expression of marker proteins and functional proteins. The cell line, with excellent biocompatibility to aPCS, might be used for in vitroreconstruction of tissue-engineered HCS.

Acellular ostrich corneal stroma used as scaffold for construction of tissue-engineered cornea

AIM： To assess acellular ostrich corneal matrix used as a scaffold to reconstruct a damaged cornea. METHODS： A hypertonic saline solution combined with a digestion method was used to decellularize the ostrich cornea. The microstructure of the acellular corneal matrix was observed by transmission electron microscopy （TEM） and hematoxylin and eosin （H＆E） staining. The mechanical properties were detected by a rheometer and a tension machine. The acellular corneal matrix was also transplanted into a rabbit cornea and cytokeratin 3 was used to check the immune phenotype, RESULTS： The microstructure and mechanical properties of the ostrich cornea were well preserved after the decellularization process, in vitro, the methyl thiazolyl tetrazoUum results revealed that extracts of the acellular ostrich corneas （AOCs） had no inhibitory effects on the proliferation of the corneal epithelial or endothelial cells or on the keratocytes, The rabbit lamellar keratoplasty showed that the transplanted AOCs were transparent and completely incorporated into the host cornea while corneal turbidity and graft dissolution occurred in the acellular porcine cornea （APC） transplantation, The phenotype of the reconstructed cornea was similar to a normal rabbit cornea with a high expression of cytokeratin 3 in the superficial epithelial cell layer, CONCLUSION： We first used AOCs as scaffolds to reconstruct damaged corneas. Compared with porcine corneas, the anatomical structures of ostrich corneas are closer to those of human corneas. In accordance with the principle that structure determines function, a xenograft lamellar keratoplasty also confirmed that the AOC transplantation generated a superior outcome compared to that of the APC graft.

Anti-inflammatory potential of human corneal stroma-derived stem cells determined by a novel in vitro corneal epithelial injury model

BACKGROUND An in vitro injury model mimicking a corneal surface injury was optimised using human corneal epithelial cells(hCEC).AIM To investigate whether corneal-stroma derived stem cells(CSSC) seeded on an amniotic membrane(AM) construct manifests an anti-inflammatory, healing response.METHODS Treatment of hCEC with ethanol and pro-inflammatory cytokines were compared in terms of viability loss, cytotoxicity, and pro-inflammatory cytokine release, in order to generate the in vitro injury. This resulted in an optimal injury of 20%(v/v) ethanol for 30 s with 1 ng/mL interleukin-1(IL-1) beta. Co-culture experiments were performed with CSSC alone and with CSSC-AM constructs.The effect of injury and co-culture on viability, cytotoxicity, IL-6 and IL-8 production, and IL1 B, TNF, IL6, and CXCL8 mRNA expression were assessed.RESULTS Co-culture with CSSC inhibited loss of hCEC viability caused by injury. Enzyme linked immunosorbent assay and polymerase chain reaction showed a significant reduction in the production of IL-6 and IL-8 pro-inflammatory cytokines, and reduction in pro-inflammatory cytokine mRNA expression during co-culture with CSSC alone and with the AM construct. These results confirmed the therapeutic potential of the CSSC and the possible use of AM as a cell carrier for application to the ocular surface.CONCLUSION CSSC were shown to have a potentially therapeutic anti-inflammatory effectwhen treating injured hCEC, demonstrating an important role in corneal regeneration and wound healing, leading to an improved knowledge of their potential use for research and therapeutic purposes.

Murine corneal stroma cells suppress bone marrow-derived dendritic cells maturation in vitro

Background Prostaglandin E2 （PGE2） is a key modulator of dendritic cells （DCs） function, and cornea-derived transforming growth factor beta 2 （TGF-β2） promotes the generation of phenotypically and functionally immature DCs. Therefore, this study was carried out to investigate whether PGE2 is involved in the suppressive effect on DCs maturation mediated by corneal stroma cells （CSCs） and whether PGE2 and TGF-β2 have additive effects in this immunosuppressive mechanism. Methods Bone marrow-derived DCs （BM-DCs）, splenic T cells and CSCs culture supernatant were obtained from mice via various protocols. After that, the level of PGE2 in CSCs culture supernatant was analyzed by enzyme-linked immunosorbent assay. Then, immature BM-DCs pretreated by E-prostanoid 2 receptor antagonist AH6809 or dimethyl sulfoxide were induced to mature in the presence of lipopolysaccharide, with or without CSCs culture supernatant. In parallel experiments, neutralizing TGF-β2 antibody or normal goat IgG was added into the supernatant. Next, the cellular surface markers for DCs maturation, including CD80, CD86, and major histocompatibility complex class Ⅱ （MHC Ⅱ）, were analyzed by flow cytometry; the capability of stimulating the proliferation of T lymphocytes was evaluated by allogeneic mixed lymphocyte reactions and the function of endocytosis was assessed by fluorescein isothiocyanate-dextran uptake. Results Higher concentration of PGE2 was detected in CSCs culture supernatant than in the fresh medium. In addition, compared with control group, after treated with the supernatant in the mature stage, BM-DCs displayed lower expression of CD80, CD86 and MHC Ⅱ, lower T cell stimulatory capacity and higher endocytosis function. However, after the application of AH6809, BM-DCs partially regained T cell stimulatory capacity and expression of CD86 and MHC Ⅱ, but partially lost endocytic activity. Moreover, after the application of AH6809 and neutralizing TGF-β2 antibody, the result of statistical analysis indicated that there was a statistical difference of interaction in the expression of MHC Ⅱ and T cell stimulatory capacity. Conclusions PGE2 contributes to the suppressive effect on BM-DCs maturation mediated by CSCs in vitro, and PGE2 and TGF-β2 have additive effects on the immunosuppression of BM-DCs.

Cellular therapy of the corneal stroma: a new type of corneal surgery for keratoconus and corneal dystrophies

Cellular therapy of the corneal stroma,with either ocular or extraocular stem cells,has been gaining a lot of interest over the last decade.Multiple publications from different research groups are showing its potential benefits in relation to its capacity to improve or alleviate corneal scars,improve corneal transparency in metabolic diseases by enhancing the catabolism of the accumulated molecules,generate new organized collagen within the host stroma,and its immunosuppressive and immunomodulatory properties.Autologous extraocular stem cells do not require a healthy contralateral eye and they do not involve any ophthalmic procedures for their isolation.Mesenchymal stem cells have been the most widely assayed and have the best potential to differentiate into functional adult keratocytes in vivo and in vitro.While embryonic stem cells have been partially abandoned due to ethical implications,the discovery of the induced pluripotent stem cells(iPSC)has opened a new and very promising field for future research as they are pluripotent cells with the capacity to theoretically differentiate into any cell type,with the special advantage that they are obtained from adult differentiated cells.Cellular delivery into the corneal stroma has been experimentally assayed in vivo in multiple ways:systemic versus local injections with or without a carrier.Encouraging preliminary human clinical data is already available although still very limited,and further research is necessary in order to consolidate the clinical applications of this novel therapeutic line.

辐射合成的温敏材料上兔角膜基质细胞的培养

目的:探索以电子加速器合成具有温度敏感特性的聚异丙基丙烯酰胺凝胶(PNIPAAm)及接枝有PNIPAAm水凝胶的培养皿的方法,及兔角膜基质细胞在温敏材料PNIPAAm水凝胶上的生长条件及特性,以及利用PNIPAAm水凝胶获得的细胞片。方法:55%N-异丙基丙烯酰胺(NIPAAm)单体和0.5%的N,N’-亚甲基双丙烯酰胺的异丙醇溶液70μL,加入到35 mm培养皿上,电子加速器下辐射合成温敏材料(PNIPAAm),后续处理后,接种兔角膜基质细胞体外培养。结果:按本实验中的单体配方以及辐射合成方案,能够在培养皿表面合成PNIPAAm,角膜基质细胞能在部分接枝了PNIPAAm的培养皿上生长,并能成片脱离。结论:使用辐射合成温敏培养皿能够获得单层及多层无载体的角膜基质细胞片。

角膜基质再生与修复的研究进展

角膜在眼球屈光系统中起着重要作用,角膜基质是角膜的主要组成部分,角膜基质的损伤可能导致永久性的视力损害,角膜移植术是目前治疗角膜基质疾病最有效的方法,但供体缺乏,长期免疫抑制治疗以防止排斥反应,以及移植物存活的限制阻碍了其进一步发展。此外,还可利用角膜基质内源性再生能力使角膜基质的胶原细胞外基质在合适的条件下能自我更新。然而,角膜基质复杂的超微结构难以在体外模拟。因此,再生医学被应用来克服这些挑战。这些方法涉及多个领域,包括干细胞诱导分化、组织工程技术、基因编辑等。本文就相关的角膜基质再生与修复技术、研究进展和存在的问题进行了归纳总结,并为未来角膜基质再生与修复的临床应用提供可能的途径。

角膜基质修复的机制和调控因素研究进展

角膜基质是维持角膜透明度的重要结构。外伤、感染、手术等可造成角膜基质损伤,引起修复的过程包括基质细胞表型改变、细胞外基质重塑、免疫细胞迁移。当基质严重受损,肌成纤维细胞增多和细胞外基质沉积发生基质纤维化反应,形成角膜瘢痕,是全球致盲的主要原因。目前治疗方式主要是角膜移植手术,因角膜供体资源短缺、手术技巧要求和术后移植排斥风险等治疗效果不佳。近年来,各种分子、细胞和组织对角膜基质损伤修复的调控机制取得一定研究进展。本文就角膜基质损伤修复的机制和角膜损伤原因、角膜结构、分子因素对角膜基质损伤修复的调控进行综述,为探索促进角膜基质修复和再生的途径提供新思路,希望帮助临床预防角膜瘢痕的发生。

新冠肺炎疫情常态化下SMILE围手术期管理策略

目的探讨新冠肺炎常态化疫情防控下,眼科门诊飞秒激光小切口角膜基质透镜取出术围手术期管理和疫情防控效果。方法选取2021年11月~2022年2月期间在我院门诊实施SMILE患者601例(1178眼)作为研究对象,结合疫情管控措施做好患者围手术期护理、检查、健康宣教等工作。观察患者手术配合度,术后效果、并发症、疫情防控效果。结果所有患者手术配合度高,术后视力恢复好,对医疗护理服务满意,无医源性和交叉感染及手术并发症,无新冠肺炎疫情发生。结论新冠肺炎疫情防控期间,严格执行防疫规定及预防措施,严格遵守眼科诊疗防控措施,可有效保障SMILE围手术期医护人员及患者的安全及手术顺利实施。

SMILE与FS—LASIK对近视眼角膜基质切削深度可预测性的比较

背景全飞秒激光小切口角膜基质透镜取出术（SMILE）已越来越多地用于近视的矫治，其手术的安全性、可预测性也受到关注。目的分析和比较SMILE与飞秒激光角膜原位磨镶术（FS—LASIK）对中央角膜组织切削深度的可预测性。方法采用非随机对照研究方法，选取于2015年10月至2016年5月在北京同仁医院拟行角膜屈光手术的近视患者135例270眼，根据患者的选择分为SMILE组69例138眼和Fs—LASIK组66例132眼，组间患者人口基线特征匹配，分别接受SMILE和FS．LASIK，分别于术前及术后1周采用RTVueFD—OCT测量术眼中央角膜厚度值，观察指标包括术中角膜实际切削深度、切削误差和术眼术后屈光度变化，术前与术后1周中央角膜厚度的差值为实际切削深度，预测切削深度与实际切削深度间的偏差值为切削误差，比较2个组间各项测量指标的差异，探讨预测切削深度与实际切削深度值之间的关系。结果SMILE组和FS—LASIK组术眼术后球镜度、柱镜度及等效球镜度的差异均无统计学意义（t=－1．826、－1．405、－1．420，均P〉0．05）。SMILE组患者术后角膜实际切削深度为（76．96±15．27）μm，低于预测切削深度的（96．76±16．52）μm，差异有统计学意义（t=－23．016，P〈0．01）；FS—LASIK组术眼实际切削深度与预测切削深度分别为（77．92±18．69）μm和（77．42±15．60）μm，差异无统计学意义（t=－0．604，P=0．547）。SMILE组术后切削误差量平均为（20．55±8．51）μm，大于FS—LASIK的（7．17±5．97）μm，组问差异有统计学意义（t=14．950，P〈0．01）。2个组术眼角膜预测切削深度与实际切削深度值问均呈线性正相关（r=0．799、0．867，均P〈0．01），SMILE组与FS—LASIK组术眼实际切削深度均随着预测切削深度的增加而增加，直线回归方程分别为l，：3．892±0．749X和Y=3．443±0．957X。结论SMILE术中实际切削深度低于预测切削深度，而FS—LASIK术中2者无明显差异，FS—LASIK可预测性好于SMILE。

飞秒激光小切口基质透镜取出术矫正近视及近视散光2年效果分析

背景已有研究表明飞秒激光小切口基质透镜取出术（SMILE）短期疗效好，术后视力恢复快，并发症少，因此已广泛用于临床，但目前仍缺乏其术后长期疗效和安全性评价研究。目的评估SMILE矫正近视及近视散光的远期有效性、安全性、可预测性、稳定性及相关手术并发症。方法采用系列病例观察研究方法，纳入2013年1—6月在河南省眼科研究所就诊的近视及近视散光患者34例67眼，所有患者均由同一有角膜屈光手术经验的医师行SMILE，分别于术前，术后1d、1周、1个月、3个月、1年及2年时进行随访观察，检查患者裸眼视力（UCVA）、最佳矫正视力（BCVA）、电脑验光、主觉验光、眼压及角膜地形图等参数。SMILE的效果评价指标主要包括有效性指数（术前BCVA／术后UCVA值）、安全性指数（术后BCVA／术前BCVA值）、可预测性[对术眼实际等效球镜度（SE）与预矫正SE进行线性回归分析]和术后屈光稳定性（术后不同时间点SE变化）。结果术前术眼BCVA≥20／20者60眼，占89．55％；术后3个月及2年分别有61跟和60眼UCVA≥20／20，分别占91．04％和89．55％。术后3个月和2年时SMILE有效性指数分别为1．038±0．182和1．029±0.231，差异无统计学意义（t=0．400，P〉0．05）。与术前BCVA相比，术后3个月及2年分别有8．96％和10．45％术眼BCVA降低1行，未发现BCVA降低2行及以上者；术后3个月和2年SMILE安全性指数分别为1．141±0.193和1．312±0．242，差异有统计学意义（t=0．414，P〉0．05）。术后3个月及2年术眼实际SE与预矫正SE均呈明显线性关系，回归方程分别为Y=0．8971X-0．4408（R^2=0．9142，P〈0．05）和Y=0．8937X-0．3823（R^2=0．9157，P〈0．05）。术后1d、1周、1个月、3个月、1年和2年术眼SE分别为（0．013±0．578）、（-0．033±0．489）、（-0．106±0．508）、（-0．103±0．375）、（-0．154±0．518）和（-0．147±0．366）D，总体比较差异无统计学意义（F=0．185，P=0．176）。术中12例18眼出现轻度弥散型不透明气泡（OBL），4例6眼发生弥漫性板层角膜炎（DLK），1例2眼出现点状角膜炎，药物治疗后症状消失。结论SMILE治疗近视及近视散光并发症少，术后2年随访证实其有效性、安全性、可预测性和稳定性较好。

角膜体外重建异种生物载体材料生物相容性研究

目的 研究异种 (猪 )角膜基质的生物相容性 ,评价其作为角膜体外重建载体的可行性。方法 将新鲜、脱水两种猪角膜基质分别植入新西兰白兔角膜层间 ,定期临床观察植片愈合情况 ,并于术后 2周、1、2、4、8个月取兔角膜进行组织学观察。结果 临床观察 :全部植片存活 ,12只术眼未见角膜水肿混浊、角膜新生血管、排斥反应发生 ,新鲜植片在 2个月左右已透明 ,脱水植片在 6个月后透明。组织学观察 :新鲜植片 4个月时与兔角膜基质相融愈合 ,脱水植片经角膜细胞再分布、胶原纤维改建重塑 ,于 8个月后与兔角膜基质相融愈合 ,2组植片愈合过程中未见有淋巴细胞浸润及新生血管生成。结论 异种 (猪 )角膜基质具有良好的生物相容性 ,是一种理想的角膜体外重建载体材料。

准分子激光角膜上皮瓣下磨镶术对兔角膜基质细胞凋亡的影响

目的观察兔行不同切削深度的准分子激光角膜上皮瓣下磨镶术(LASEK)后角膜基质细胞凋亡的变化。方法健康新西兰白兔随机分为3组,随机选择一只眼行LASEK术,分别按近视-3.00、-9.00和-15.00D设计切削,另一只眼为对照眼。24h和7d后常规取角膜制成石蜡切片,采用核苷酸末端转移酶介导的dUTP缺口标记(TUNEL)法检测基质细胞凋亡,计数凋亡细胞,计算凋亡指数进行分析。结果凋亡细胞呈棕黄色,体积缩小,核固缩成团。术后24h-3.00、-9.00和-15.00D三组激光切削眼凋亡指数分别为0.39±0.10、0.55±0.08和0.54±0.12,均分别高于对照眼;-9.00D的切削较-3.00D组凋亡细胞增多,-15.00D组凋亡指数与-9.00D无显著性差异;术后7d激光切削组的凋亡指数与对照眼无显著性差异,且较24h减轻。结论LASEK术可导致兔角膜基质细胞凋亡,且随切削深度的增加,细胞凋亡呈增加趋势。

自体血浆角膜基质内注射病理改变的实验研究

目的 研究角膜基质内注射自体血浆治疗大泡性角膜病变的机制及组织学改变。方法 正常兔角膜 ,基质内注射自体血浆 0 5ml。结果 术后 72h血浆全部吸收 ,角膜水肿消退上皮光滑。通过HE及Pollak染色检查提示 :角膜固有层明显增厚 ,基质内有不同程度的结缔组织增生。结论 结缔组织增生可形成一层纤维屏障 ,阻止水分向前渗透至上皮或上皮下 ,致使大泡难以形成。

应用组织工程技术构建角膜基质组织(英文)

目的应用角膜基质细胞和聚羟基乙酸(PGA)多聚体复合物构建角膜基质,为组织工程技术构建角膜提供依据。方法将培养后的角膜基质细胞-生物材料复合物植入balb/c裸鼠皮下。6周后,对新生组织进行组织学和透射电镜检测,并测定胶原纤维直径。结果石蜡包埋切片显示新生角膜组织具有似正常角膜基质层波浪交叉编织结构。组织工程化的角膜基质胶原纤维直径(27.7±6.2)nm,与正常角膜基质胶原纤维直径相似。结论组织工程技术重建的角膜基质已经具备角膜基质层组织形态学和组织学特征。

人眼角膜基质生物力学特性研究

目的了解正常人眼角膜基质的生物力学特性,初步获得相对正常角膜组织生物力学相关参数。方法使用全飞秒激光小切口角膜基质透镜取出术术中获取的角膜基质透镜,根据实验需要裁切成角膜基质组织试件,在常温和水浴条件下,分别进行应力-应变实验、蠕变实验和应力松弛实验,初步得到新鲜角膜基质组织的应力-应变曲线、蠕变曲线和应力松弛曲线,以评估人眼角膜基质的生物力学特性。结果低应变区切线模量为(1.32±0.48)MPa,高应变区切线模量为(51.26±8.23)MPa,断裂点应变为(0.52±0.06)mm·mm-1,断裂点应力为(13.00±2.51)MPa。在应力为0.01 MPa、0.02 MPa、0.03 MPa时,角膜基质试件弹性模量分别为(0.89±0.25)MPa、(1.28±0.21)MPa、(1.69±0.18)MPa。在1.00 N恒定载荷下,角膜基质试件形变随时间延长而增大,早期变化较快,后期逐渐变缓,600 s时蠕变率为(4.57±1.67)%。维持角膜基质试件长度至初始长度的1.5倍,应力随着时间的延长逐渐衰减,早期衰减较快,随着时间的延长衰减趋于平缓,1 h时松弛率为(28.49±0.04)%。结论初步得出正常角膜基质组织的生物力学参数,可为进一步深入研究角膜的生物力学特性提供一定的理论参考。

角膜体外重构异种生物载体材料植入性实验研究

目的:分析测定异种(猪)角膜基质组织免疫原性,观察其角膜层间植入后与受体角膜愈合情况,以评价该材料生物相容性,并在此载体上体外重构角膜内皮组织,探讨其作为角膜重建载体材料可行性。方法:(1)将新鲜、脱水两种猪角膜基质植片分别植入F344大鼠角膜基质层间,术后12、90d对外周血进行CD25和CD4/CD8双色免疫荧光标记,流式细胞仪测定分析。(2)猪角膜基质植入新西兰白兔角膜层间,定期临床观察植片愈合情况,并取受体兔角膜进行组织学观察。(3)将猫角膜内皮细胞接种于保留后弹力层的脱水猪角膜基质上,加入培养液培养7d,组织学观察体外重构的角膜内皮组织形态结构。结果:(1)测得新鲜组、脱水组大鼠外周血T淋巴细胞CD4+CD25+、CD8+CD25+双阳性表达率及CD4+/CD8+比值与同基因移植组、阴性对照组比较无显著差异(P>0.05)。(2)兔眼临床观察:全部植片存活,12只术眼未见有角膜水肿混浊、角膜新生血管、排斥反应发生,新鲜植片在2个月左右已透明,脱水植片在6个月后透明。兔角膜组织学观察:新鲜植片4个月时与兔角膜基质相融愈合,脱水植片经角膜细胞再分布、胶原纤维改建重塑于8个月后与兔角膜基质相融愈合,两组植片愈合过程中未见有淋巴细胞浸润及新生血管生成。(3)体外重构的内皮组织形态结构与正常内皮层相似。结论:异种(猪)角膜基质免疫原性低,具有良好的生物相容性,是目前较为理想的角膜体外重构载体材料。

异种角膜基质作为生物角膜载体的免疫学研究

目的探讨异种(猪)角膜基质组织免疫原性,以评价其作为角膜重建载体材料的可行性。方法将新鲜、脱水2种猪角膜基质植片分别植入F344大鼠角膜基质层间,术后12d、90d抽取外周血,行抗CD25·FITC和抗CD4/CD8·PE双色免疫荧光标记,流式细胞仪测定分析。结果测得新鲜组、脱水组大鼠术后12d外周血T淋巴细胞表面抗原CD4+和CD25+、CD8+和CD25+双阳性表达率(1.53%±0.47%,2.13%±0.53%与0·57%±0.20%,0.67%±0.18%)及CD4+/CD8+比值(1.43±0.28,1.69±0.18)与同基因移植组(1.78%±0.36%,1.50±0.19)、阴性对照组(2.06%±0.52%,1.44±0.32)比较无显著性差异(P>0.05)。术后90d,新鲜组、脱水组大鼠外周血T淋巴细胞表面抗原CD4+和CD25+、CD8+和CD25+双阳性表达率(1.48%±0.39%,1.50%±0.46%与0·55%±0.15%,0·58%±0·11%)及CD4+/CD8+比值(1.43±0.51,1.36±0.59)与同基因移植组(1.32%±0·75%,1.31±0.29)、阴性对照组(1.34%±0.18%,1.44±0.42)比较,也无显著性差异(P>0.05)。结论异种(猪)角膜基质免疫原性低,是一种理想的角膜体外重建载体材料。

组织工程化角膜基质的构建与移植实验

目的探讨以可降解高分子材料聚羟基乙醇(PLGA)为支架体外构建组织工程化角膜基质的可行性.方法将培养的角膜基质细胞接种于具微孔的可降解高分子材料PLGA内,构建成组织工程化细胞-PLGA复合物,将该复合物移植到兔眼角膜基质层间.分别于移植后1、2周,1、2、3、4个月取材,光学显微镜、电镜观察细胞与PLGA材料的结合情况.裂隙灯显微镜和组织切片检查组织工程化角膜基质的透明性、可降解性以及生物反应性.结果角膜基质细胞能与PLGA高分子材料较好地结合,移植到兔眼角膜层间的组织工程化角膜基质7 d后呈半透明,2周见新生血管从角膜缘向植片方向生长,1个月开始材料降解,3～4个月时大部分材料降解,且新生血管减少,但是移植物呈半透明状.结论PLGA虽然具有组织相容性好、无毒、无免疫性和可降解的特性,但不适合作为构建要求具有很好透明性的角膜组织的支架材料.