前房相关性免疫偏诱导及预防角膜移植排斥反应实验研究

目的 研究前房相关性免疫偏离（ＡＣＡＩＤ）诱导及预防角膜移植排斥反应的作用。方法 采用甘氨酸法制备可溶性角膜分类抗原,分组行兔前房注射诱导ＡＣＡＩＤ,并观察各组部分耳试验和穿透角膜移植术（ＰＫＰ）术后排斥反应。结果 角膜上皮、内皮、基质及全角膜可溶性抗原皆可诱导ＡＣＡＩＤ,诱导质量浓度为２．０ｍｇ／ｍｌ。成功率分别为９０％,１００％,８０％和１００％。同期解膜移植免疫排斥率降低,植片存活时间延长。     

碱烧伤对眼前房免疫赦免状态的削弱和破坏

目的观察碱烧伤后眼免疫赦免状态的改变。方法使用０．５ｍｏｌ／ＬＮａＯＨ在兔眼建立碱烧伤模型。通过胸腺细胞刺激反应检测碱烧伤后不同时间房水的免疫抑制水平。另外，观察了碱烧伤对前房相关免疫偏离（ＡＣＡＩＤ）的支持能力。结果碱烧伤后眼房水的正常免疫抑制效应显著下降，即使在碱烧伤后２个月。烧伤眼接种抗原后ＡＣＡＩＤ诱导失败。结论碱烧伤后角膜移植免疫排斥反应的高发率除与植床的高度血管化和炎症背景外，眼前房免疫赦免机制的破坏或削弱也可能起着重要的作用     

减少房水分泌对眼前房免疫赦免和角膜移植免疫排斥反应的影响

本实验在兔眼建立睫状体冷冻术模型，观察了睫状破坏和减少房水分地眼前房免疫郝免状态，以及角膜移植存活的影响，结果显示：冷冻伤术后６周眼房水的免疫制活性显著下降；ＡＣＡＩＤ现象消失，与正常对照组相比，免疫排斥反应加速植片存活时间缩短，结果提示：正常睫状体的组织结构及分泌的房水及对于维持和介导前房免疫郝免，以及角膜移植的长期存活起着重要作用。这为进一步阐明角膜移植免疫排斥反应机制，并制定相应预防措施提供     

角膜移植免疫排斥反应防治研究进展

角膜移植术后发生免疫排斥反应是导致移植手术失败的主要原因。角膜移植与其他器官移植相比，不易发生免疫排斥反应，主要原因：前房的相关免疫偏离状态；血．房水屏障；角膜缺乏血管、淋巴组织；角膜中央区不含成熟抗原提呈细胞；房水中有大量免疫调节分子；眼前节有Fas配基表达等。角膜移植免疫排斥反应是一个复杂的反应过程，一般包括宿主对异体组织抗原的致敏和宿主对异体组织抗原的反应两方面。相应的治疗措施应围绕三方面：（1）阻断宿主对异源组织抗原的敏感性；（2）诱导免疫耐受，使淋巴不激活或活性减低；（3）降低或阻断免疫效应因素对角膜植片的破坏。随着对角膜移植排斥反应机制的深入研究，将有新的治疗措施出现，特别是诱导免疫耐受防治角膜移植排斥反应的研究前景乐观。     

脾脏在不同种类动物前房相关免疫偏离诱导和维持中的作用

目的 脾脏在小鼠前房相关免疫偏离（ａｎｊｔｅｒｉｏｒ ｃｈａｍｂｅｒ－ａｓｓｏｃｉａｔｅｄ ｉｍｍｕｎｅ ｄｅｖｉａｔｉｏｎ,ＡＣＡＩＤ）的诱导中起着重要的作用。在前房接种抗原４天内摘除脾脏可以消除免疫偏离而导致免疫反应。本研究旨在观察脾脏在其他种类动物ＡＣＡＩＤ的诱导和维持中的作用。方法 在脾脏切除和假性脾脏切除后,使用牛血清白蛋白（ｂｏｖｉｎｅ ｓｅｒｕｍ ａｌｂｕｍｉｎ,ＢＳＡ）作为可溶性抗     

在正常妊娠期间兔眼前房相关免疫偏离的变化

目的 许多研究发现妊娠对眼具有影响。该研究旨在观察正常妊娠期间前房接种可溶性蛋白抗原能否诱导免疫反应的主动下调。方法 标准交配１５天后通过胚胎种植部位确立新西兰兔的中期妊娠。将牛血清白蛋白（ＢＳＡ）分别接种于雄性,非妊娠雌性和中期妊娠雌性兔眼前房,接受附加佐剂的ＢＳＡ免疫方案后,评价诱导前房相关免疫偏离（ＡＣＡＩＤ）的能力。结果 在雄性和非妊娠雌性兔眼前房接种ＢＳＡ诱导了ＢＳＡ特异性的迟发型超敏反     

CD86在排斥反应和前房相关免疫偏离过程中的作用

目的检测大鼠角膜共刺激分子CD86（B7—2）的原位表达,探讨CD86分子与角膜移植排斥反应和前房相关免疫偏离（ACAID）过程的关系。方法制作穿透角膜移植排斥反应和同一供体前房内注射脾细胞诱导ACAID的大鼠模型;角膜移植组进行排斥反应指数（RI）评分;ACAID组进行迟发型超敏反应（DTH）评价;采用免疫组织化学的方法检测CD86在角膜中的原位表达（以脾脏的表达为阳性对照）。结果角膜移植组植片均出现不同程度的新生血管、角膜水肿、混浊、增厚;ACAID组角膜透明,房水清,DTH评价术后2周及4周诱导成功率100％;免疫组织化学检测CD86在正常大鼠角膜组织全层无阳性表达,在移植后出现急性排斥反应的大鼠角膜上皮层中有大量阳性细胞表达,在ACAID诱导成功的大鼠角膜中未见阳性细胞表达。结论共刺激分子CD86参与移植后的急性排斥反应,但可能不参与免疫赦免过程。     

CTLA4-Ig保护同种异体角膜植片

目的:ＣＴＬＡ4(对可溶性抗原递呈细胞Ｂ7共同配体具有高亲和性)通过抑制ＣＤ28细胞和Ｂ7细胞之间相互作用而阻止抗原诱导Ｔ细胞的激活,进而防止免疫移植排斥反应的发生。本研究检测可溶性ＣＴＬＡ4Ｉｇ单独或联合ＵＶＢ照射对兔角膜移植排斥反应治疗效果。方法:将荷兰兔?..     

灵长目前房相关免疫偏离的诱导和维持不需要脾脏的完整性

目的 观察脾脏在灵长目前房相关免疫偏离（ＡＣＡＩＤ）诱导和维持中的作用。方法 按照经典方法在猴眼建立ＡＣＡＩＤ模型。在前房抗原接种的切除脾脉。通过对原特异性迟发型超敏反应（ＤＴＨ）的抑制来珠诱导和维持。结果 在单纯脾脏切除组动物均显示阳怀ＤＴＨ反应;但在单纯前房怕接种组、脾脏切除加前房怕接种组均显示阴性ＤＴＨ反应,且维持时间无显著差异。结论 灵长目ＡＣＡＩＤ的诱导和维持不需要功能性脾脏的存在。实验     

FK506缓释系统前房植入抑制兔高危角膜移植术后的免疫排斥反应

目的探讨前房植入FK506药物缓释系统（DDS）对兔高危角膜移植术后免疫排斥反应的抑制作用和FK506房水药物浓度与免疫排斥反应的关系。方法107只新西兰白兔中随机数字法选取73只兔进行角膜新生血管化模型的制作,其中68只兔作为受体成功建立高危角膜移植动物模型,随机数字法分为对照组、空白DDS前房植入组、环孢素A（CsA）DDS前房植入组（含CsA 1mg）、0．1％FK506眼液滴眼组及FK506 DDS前房植入组（含FK5060．5mg）。角膜移植术后观察各组角膜植片排斥发生的时间,移植术后1周取各组实验兔眼房水和静脉血进行FK506药物浓度检测。0．1％FKS06眼液滴眼组和FKS06 DDS前房植入组在移植术后的不同时间点抽取实验兔眼房水和静脉血,进行FK506药物浓度的检测。观察各组兔移植术后4周和观察期结束时角膜植片的病理变化,同时应用原位杂交的方法检测各组角膜植片内白细胞介素2受体a（IL-2Bot）、单核细胞趋化蛋白1（MCP-1）、Fas及FasL mRNA的表达。结果FK506 DDS前房植入组角膜植片存活时间超过180d,明显优于其他各组（F=926．37,P=0．0000）,其房水和角膜组织中的FK506药物浓度明显高于FKS06眼液滴眼组（T=21．00,P=0．0022）。FKS06 DDS前房植入组在术后24周内均能在房水中检测出FK506。术后4周对照组和空白DDS前房植入组有大量的炎性细胞浸润,并有明显的IL．2Bet和MCP-1mRNA的表达,而CsADDS前房植入组、FK506眼液滴眼组及FK506 DDS植入组角膜未见明显的炎性细胞浸润,未见IL-2Pux和MCP-1mRNA的表达。各组均未见明显的Fas和FasL mRNA的表达。结论前房植入FK506 DDS可有效地抑制高危角膜移植术后免疫排斥反应的发生,房水中较高的FK506药物浓度是防治术后发生免疫排斥反应的重要因素。     

诱导供体特异性前房相关免疫偏离对高危角膜移植排斥反应的影响

目的:探讨诱导供体特异性的前房相关免疫偏离(anterior chamber-associated immune deviation,ACAID)对高危角膜移植排斥反应的影响。方法:采用新西兰白兔眼角膜建立碱烧伤眼模型,实验动物随机分为4组,A:正常角膜常规角膜移植组(正常对照组);B:碱烧伤常规角膜移植组(碱烧伤对照组);C:正常角膜诱导ACAID的角膜移植组(正常诱导组)。D:碱烧伤后诱导ACAID的角膜移植组(碱烧伤诱导组)。B,D组进行左眼碱烧伤,烧伤后1mo进行角膜移植。C,D组在角膜移植前2wk于右眼前房注入可溶性抗原以预先诱导供体特异性ACAID。A,B组右眼前房注等量平衡盐溶液。术后记录植片存活时间;对角膜新生血管(corneal neovascularization,CNV)生长情况进行评分;记录移植排斥指数(rejection index,RI);角膜固定、包埋后制作切片行HE染色。结果:A、C组角膜植片长期存活,碱烧伤对照组植片平均存活25.13±0.64d,碱烧伤诱导组角膜植片平均存活时间为38.25±1.28d。与碱烧伤对照组相比,碱烧伤诱导ACAID组角膜植片平均存活时间显著延长,两组植片存活时间的差异具有统计学意义(P=0.00)。结论:诱导供体特异性ACAID可以延长碱烧伤高危眼角膜植片的存活时间。     

Effect of local macrophage depletion on cellular immunity and tolerance evoked by corneal allografts

Corneal graft rejection can be prevented by local macrophage depletion, via subconjunctival injections with clodronate liposomes. To unravel the underlying immunological mechanism responsible for prolonged graft survival in this circumstance, the effect of this regimen on induction of donor-specific delayed type hypersensitivity (DTH) and anterior chamber associated immune deviation (ACAID) was determined. The study showed that although subconjunctival clodronate liposome-treatment failed to alter systemically induced DTH and ACAID, both types of immune response were absent in clodronate liposome-treated rats after corneal transplantation. Thus, elimination of macrophages from the corneal transplant site renders corneal allografts immunologically "invisible" to the recipient. PURPOSE: To determine whether local macrophage depletion, via administration of clodronate liposomes, alters delayed type hypersensitivity (DTH) responses and induction of anterior chamber associated immune deviation (ACAID) after corneal allotransplantation. METHODS: Clodronate liposome-treated and untreated rats received orthotopic corneal allografts and were tested for DTH responses and induction of ACAID towards donor antigens. Also in subconjunctivally treated and untreated rats, DTH responses were measured after subcutaneous immunization or induction of ACAID with allogeneic spleen cells. RESULTS: Subconjunctival injected clodronate liposomes prevented grafted rats from developing donor-specific DTH as well as ACAID. By contrast, subconjunctivally injected clodronate liposomes had no effect on donor-specific DTH responses after systemic immunization or on the induction of ACAID with allogeneic cells. CONCLUSIONS: Depletion of macrophages at the time of corneal allografting seems to render the grafts immunologically invisible to the recipients. This could explain why these grafts survive "indefinitely" without any other form of therapy.     

Role of tumor necrosis factor receptor expression in anterior chamber-associated immune deviation (ACAID) and corneal allograft survival

PURPOSE: To determine the role of tumor necrosis factor receptors (TNFRs) in corneal allograft rejection. METHODS: Corneal epithelial and endothelial cells were examined by flow cytometry for the expression of TNFRI and TNFRII and their susceptibility to TNF-alpha-induced apoptosis. Corneal allografts from normal and TNFRI and TNFRII knockout (KO) C57BL/6 mice were transplanted to BALB/c hosts, and the fate of the allografts was monitored. C57BL/6 spleen cells were injected into the anterior chamber (AC) of BALB/c mice to induce anterior chamber-associated immune deviation (ACAID) and promote corneal allograft survival. The presence of ACAID suppressor cells in corneal allograft recipients was tested using a local adoptive transfer (LAT) assay. RESULTS: Murine corneal epithelial and endothelial cells expressed TNFRI and TNFRII and were susceptible to TNF-alpha-induced apoptosis, yet corneal allografts from either TNFRI or TNFRII donors did not enjoy a lower incidence of rejection or a prolongation in survival time compared to corneal allografts from normal C57BL/6 donors. Moreover, all 31 of the TNFRII KO corneal grafts were rejected by na?ve BALB/c hosts. Rejection of TNFRII KO corneal grafts occurred even though suppressor cells developed in the hosts and inhibited the expression of delayed-type hypersensitivity to donor alloantigens. CONCLUSIONS: Expression of TNFRII on corneal cells conveys a degree of protection against immune rejection of corneal allografts by a mechanism that is independent of ACAID. Moreover, induction of ACAID before the application of TNFRII KO corneal allografts fails to improve survival and does not replace the TNFRII-dependent protective mechanism.     

Effect of anterior chamber-associated immune deviation (ACAID) on rat islet allograft rejection

Anterior chamber-associated immune deviation (ACAID) is characterized by the systemic inhibition of delayed type hypersensitivity (DTH) reactions to antigens which have previously been placed into the anterior chamber of the eye. Since its initial characterization, ACAID has been elicited to a wide variety of antigens, including alloantigens, and has been shown to be due to the immune deviating effects of factors such as transforming growth factor beta (TGF-beta) in the aqueous humor on ocular antigen-presenting cells (APCs). ACAID can also be induced by injecting animals with nonocular APCs, such as peritoneal exudate cells (PECs), which have been precultured with TGF-beta and antigen in vitro. The objective of this study was to determine whether alloantigenic ACAID can be effective in preventing the rejection of rat islet allografts. The notion that islet allograft rejection can be inhibited by ACAID stems from an early study showing an ACAID-induced delay in the rejection of skin grafts. Furthermore, the immune cells mediating a DTH reaction are similar to those implicated in islet allograft rejection, suggesting that they, too, may be subject to inhibition by ACAID. Our results showed that in spite of successful ACAID induction to islet allografts, recipient rats consistently rejected their grafts. Cytotoxic T cell activity (which is not inhibited by ACAID) directed against donor alloantigens was high in these animals and may have accounted, in part, for graft failure.     

Corneal allograft rejection: risk factors, diagnosis, prevention, and treatment

Recent advances in corneal graft technology, including donor tissue retrieval, storage and surgical techniques, have greatly improved the clinical outcome of corneal grafts. Despite these advances, immune mediated corneal graft rejection remains the single most important cause of corneal graft failure. Several host factors have been identified as conferring a "high risk" status to the host. These include: more than two quadrant vascularisation, with associated lymphatics, which augment the afferent and efferent arc of the immune response; herpes simplex keratitis; uveitis; silicone oil keratopathy; previous failed (rejected) grafts; "hot eyes"; young recipient age; and multiple surgical procedures at the time of grafting. Large grafts, by virtue of being closer to the host limbus, with its complement of vessels and antigen-presenting Langerhans cells, also are more susceptible to rejection. The diagnosis of graft rejection is entirely clinical and in its early stages the clinical signs could be subtle. Graft rejection is largely mediated by the major histocompatibility antigens, minor antigens and perhaps blood group ABO antigens and some cornea-specific antigens. Just as rejection is mediated by active immune mediated events, the lack of rejection (tolerance) is also sustained by active immune regulatory mechanisms. The anterior chamber associated immune deviation (ACAID) and probably, conjunctiva associated lymphoid tissue (CALT) induced mucosal tolerance, besides others, play an important role. Although graft rejection can lead to graft failure, most rejections can be readily controlled if appropriate management is commenced at the proper time. Topical steroids are the mainstay of graft rejection management. In the high-risk situations however, systemic steroids, and other immunosuppressive drugs such as cyclosporin and tacrolimus (FK506) are of proven benefit, both for treatment and prevention of rejection.     

Interleukin-1 receptor antagonist therapy and induction of anterior chamber-associated immune deviation-type tolerance after corneal transplantation

PURPOSE: Topical treatment with interleukin 1 receptor antagonist (IL-1ra) can promote corneal allograft survival by suppressing induction of allodestructive immunity. The purpose of these experiments was to determine whether IL-1ra could also promote induction of allo-protective tolerogenic pathways, including anterior chamber-associated immune deviation (ACAID), which has been shown to participate in long-term survival of corneal transplants. METHODS: Corneal buttons from BALB/c (syngeneic) or C57BL/6 (fully mismatched allogeneic) mice were orthotopically grafted onto BALB/c recipients. Topical IL-1ra or vehicle alone was applied to grafts three times daily. Donor-specific ACAID was measured in allogeneic grafted mice at 4 and 8 weeks after transplantation by ear-challenging grafted hosts with donor-derived splenocytes 1 week after SC immunization. In separate experiments, grafted mice were treated for 4 weeks before injecting ovalbumin (OVA) into their anterior chambers to determine their capacity to induce antigen-specific ACAID. RESULTS: Treatment with IL-1ra did not promote, or inhibit, induction of donor-specific ACAID compared with vehicle-treated controls at either the early or late time points studied. However, IL-1ra treatment after transplantation led to significantly earlier restoration of the grafted eyes' capacity for inducing ACAID to soluble antigen (OVA). CONCLUSIONS: Promotion of OVA-specific ACAID by IL-1ra suggests that suppression of IL-1-mediated mechanisms contributes to recovery of the anterior segment's immunosuppressive microenvironment at least 1 month earlier than would otherwise be seen after corneal transplantation. However, IL-1ra treatment does not alter induction of donor-specific ACAID after transplantation, suggesting that its anti-inflammatory activities do not lead to an ACAID-inducing signal per se. This suggests that IL-1ra promotes graft survival almost exclusively by virtue of suppressing inflammation and not by directly promoting tolerance or antigen-specific regulatory pathways.     

Oversized grafts in children

OBJECTIVE: To evaluate the efficacy of oversized corneal grafts in the pediatric age group. DESIGN: Prospective, nonrandomized clinical trial. PARTICIPANTS AND INTERVENTION: Forty pediatric patients with unilateral or bilateral corneal opacification of congenital or acquired origin underwent corneal grafting surgery over a period of 2 years using donor corneal buttons oversized by 1 mm. MAIN OUTCOME MEASURES: The parameters evaluated were indications for keratoplasty, graft clarity, visual acuity, keratometry, spherical equivalent, anterior chamber depth, and complications. RESULTS: Corneal ulceration was the most common cause of corneal opacification (25%), followed by trauma (20%) and sclerocornea (20%). At 1 year, clear grafts were achieved in 85% of the cohort. The average keratometry at the end of 1 year was 43.28 +/- 1.65 diopters (D) in the congenital opacity group and 43.04 +/- 2.20 D in the acquired group. The keratometric astigmatism was 3.60 +/- 2.60 D in the congenital group and 2.52 +/- 2.20 D in the acquired group. Oversized grafts provided an adequate anterior chamber depth of 2.20 +/- 0.612 mm in the congenital group and 2.36 +/- 0.302 mm in the acquired group. Visual acuity of 20/80 or better was recorded in only 30% of cases in the congenital group as opposed to 47% with acquired opacities. Nine cases had episodes of graft rejection. CONCLUSION: Oversizing donor buttons by 1 mm provides adequate anterior chamber depth and increases the morphologic success of corneal grafting in children.     

Immunobiology of Langerhans cells on the ocular surface. I. Langerhans cells within the central cornea interfere with induction of anterior chamber associated immune deviation

Inoculation of P815 tumor cells into the anterior chamber of eyes of BALB/c mice produces Anterior Chamber Associated Immune Deviation (ACAID) whereby delayed hypersensitivity responses to the minor H antigens of the tumor cells are suppressed. In our laboratory, impaired delayed hypersensitivity is obtained in the majority, but not all normal BALB/c recipients. In the minority, typcial delayed hypersensitivity responses, similar to those induced by subcutaneous immunization with tumor cells, are observed. Retrospective examination of eyes of animals that develop delayed hypersensitivity following intracameral tumor cell injection revealed a high incidence of a corneal disorder in which significant numbers of Langerhans cells are found within the corneal epithelium. BALB/c mice with corneal lesions characterized by opacity and calcium deposition, or whose corneas had become infiltrated with Langerhans cells induced to emigrate by a superficial cautery wound, received intracameral injections of P815 cells. Minimal or no significant suppression of delayed hypersensitivity was obtained, suggesting that ACAID induction had not taken place. These results suggest that the presence of Langerhans cells in the central corneal epithelium mitigates against ACAID induction.     

Oversized corneal grafts for corneal opacities with iridocorneal adhesions.

OBJECTIVE: To evaluate the efficacy of 1-mm oversized corneal grafts in patients with acquired corneal opacities and extensive peripheral iridocorneal adhesions. DESIGN: Prospective noncomparative case series. PARTICIPANTS: Twenty patients (20 eyes) aged 15 years or older with unilateral or bilateral corneal opacification and a shallow anterior chamber. INTERVENTION: Penetrating keratoplasty was performed with donor corneal buttons oversized by 1 mm. MAIN OUTCOME MEASURES: The various parameters evaluated were visual acuity, graft clarity, keratometry, anterior chamber depth, intraocular pressure, and spherical equivalent refraction 12 months after surgery. RESULTS: The keratoplasties were performed in 15 eyes with a corneo-iridic scar after infectious keratitis (75%) and 5 eyes with failed graft (25%). At the final follow-up, a clear graft was achieved in 17 eyes (85%), and 14 eyes (70%) achieved a best-corrected visual acuity of 6/12 or better. Three of the grafts failed because of rejection. The average keratometry was 44.1 +/- 1.0 diopters (D), and the mean spherical equivalent was -3.23 +/- 2.86 D. The oversized grafts provided a mean anterior chamber depth of 2.36 +/- 0.42 mm, and the mean intraocular pressure at the 12 month follow-up was 16.38 +/- 2.09 mmHg. CONCLUSIONS: Corneal grafts oversized by 1 mm provide adequate anterior chamber depth and reduce the risk of peripheral anterior synechiae and secondary glaucoma in patients with corneal opacities and extensive peripheral iridocorneal adhesions.