Fas/FasL抗原在Behcet病患者外周血T淋巴细胞的表达

目的研究Fas/FasL抗原在Behcet病患者外周血T淋巴细胞的表达及其可能的意义。方法采用流式细胞仪(flowcytometry)及三色荧光双标记细胞免疫荧光技术对26例Behcet病患者和43例正常人外周血T淋巴细胞表面Fas和FasL抗原的表达进行检测。结果Behcet病CD4+T细胞Fas表达阳性率为25.70%±7.32%,对照组为14.02%±6.30%,二者有显著性差异(P＜0.01);但CD4+T细胞的FasL表达在2组间无显著性差异。CD8+T细胞Fas表达阳性率为9.47%±6.97%,对照组为3.47%±2.75%,二者有显著性差异(P＜0.01),但FasL表达阳性率二者无显著性差异。结论Behcet病患者外周血CD4+和CD8+T细胞Fas的表达阳性率高于正常对照组,FasL的表达阳性率相对不足,提示这些活化的Fas+T淋巴细胞不能有效地通过Fas/FasL系统介导的凋亡途径被清除,此可能是Behcet病患者体内大量活化的淋巴细胞长期存在和炎症持续存在及复发的重要原因之一。     

Fas／FasL抗原在葡萄膜炎患者外周血T淋巴细胞上的表达

目的评价和探讨Fas/FasL抗原在葡萄膜炎患者外周血T淋巴细胞上的表达及其可能存在的意义.方法采用流式细胞术(flow cytometry,FCM)及三色荧光双标记细胞免疫荧光染色技术对26例Behcet病、17例Vogt-小柳原田综合征患者、25例特发性前葡萄膜炎患者及43例正常人外周血CD4+T细胞和CD8+T细胞表面Fas和FasL的表达进行检测.结果Behcet病、Vogt-小柳原田综合征和特发性前葡萄膜炎患者CD4+T细胞Fas的表达阳性率分别为(25.70±7.32)%、(19.60±11.02)%、(20.81±7.40)%,均显著高于正常对照组[(14.02±6.30)%],而FasL的表达阳性率与正常对照组比较,差异无显著性意义.Behcet病和Vogt-小柳原田综合征患者CD8+T细胞Fas的表达阳性率分别为(9.47±6.97)%,(6.84±5.53)%,均显著高于正常对照组[(3.47±2.75)%];特发性前葡萄膜炎患者CD8+T细胞Fas的表达阳性率[(5.34±3.08)%]与正常对照组[(3.47±2.75)%]比较,差异无显著性意义.Behcet病和Vogt-小柳原田综合征患者CD8+T细胞FasL的表达阳性率分别为(3.66±2.16)%、(4.05±1.38)%,与正常对照组[(3.09±2.04)%]比较,差异无显著性意义;特发性前葡萄膜炎患者CD8+T细胞FasL的表达阳性率[(1.58±0.93)%]低于正常对照组[(3.09±2.04)%].结论葡萄膜炎患者CD4+T细胞和CD8+T细胞Fas的表达阳性率高于正常对照组,而FasL的表达阳性率相对不足,提示这些活化的Fas+T淋巴细胞不能通过Fas/FasL系统介导的凋亡途径被清除,此可能是葡萄膜炎患者体内大量活化的淋巴细胞长期存在和炎症持续存在及复发的重要原因.     

Insufficient expression of Fas antigen on helper T cells in Behçet's disease.

AIMS: To investigate the lack of equilibrium in the regulatory mechanism of the immune system in Behçet's disease (BD), the expression of Fas antigen, an apoptosis related antigen, on peripheral blood lymphocytes from BD patients was analysed. METHODS: Twenty one BD patients were the subjects in this study. Ten healthy adults were examined as controls. Cell surface antigens of lymphocytes were analysed with flow cytometry. RESULTS: There was a significant (p < 0.01) difference in the proportion of CD4 positive cells with CD25 between BD patients with active uveoretinitis (27.6% (SD 8.4%)) and the controls (14.7% (2.3%)), but no significant difference in the proportion of CD4 or CD45RO positive cells with Fas. On the other hand, the proportion of CD8 positive cells with Fas was significantly (p < 0.01) higher in BD patients with active uveoretinitis (45.6% (11.6%)) than in those with inactive uveoretinitis (23.8% (8.1%)) or in the controls (24.4% (2.5%)). The proportion of CD19 positive cells with Fas was also significantly (p < 0.01) higher in BD patients with active uveoretinitis (13.0% (5.0%)) than in the controls (5.1% (2.1%)). CONCLUSION: The insufficient expression of Fas on activated CD4 positive T cells and its high expression on CD8 positive T cells seem to play an important role in the chronic inflammation in BD.     

Bcl-2 expression by CD4 T lymphocytes in Vogt-Koyanagi-Harada disease

Purpose: To determine whether Bcl-2 is expressed on CD 4 + lymphocytes in the aqueous humor (AH) and cerebrospinal fluid (CSF) of patients with Vogt-Koyanagi-Harada (VKH) disease, and to determine whether Fas will induce apoptosis of lymphocytes in the CSF. Methods: The percentages of CD4, CD8, CD45RO, Fas, and Bcl-2 positive T lymphocytes in the AH and CSF of eight patients with active VKH and five healthy controls were determined by flow cytometry. Soluble Fas ligand (sFasL) in the CSF was measured by ELISA. Freshly isolated cells from the CSF were cultured with anti-Fas antibody (Ab) and apoptosis was assessed by the TUNEL method. Results: Fas + CD4 + lymphocytes were the predominant lymphocytes in the AH and CSF of VKH patients. Bcl-2 was strongly expressed in these cells. Soluble FasL was also detected in the CSF. The number of apoptotic cells detected by anti-Fas Ab was not significantly increased in the CSF of VKH patients. Conclusions: In spite of the high expression of Fas antigen on CD4 + cells and the presence of sFasL in the CSF, apoptosis was not observed. Bcl-2 expression may contribute to the regulation of apoptosis of inflammatory cells in the CSF of VKH patients.     

Behcet病周围血淋巴细胞Fas和FasL的表达

目的 观察Behcet病周围血淋巴细胞Fas和FasL的表达及其意义。方法 应用Oligod(T)18和M-MuLv逆反录酶方法，观察19例Behect病患者和19个正常人周围血淋巴细胞的Fas和FasL mRNA表达，并用流动细胞计和免疫荧光法进一步观察T淋巴细胞的Fas和FasL的表达。结果 Behcet病组Fas和FasL的表达明显高于对照组。Fas抗原在CD4^ 和CD86 的表达明显高于对照组，但FasL的表达与对照组无明显表达明显增设，但其分子表达不平衡，此可能是Behcet病慢性和复发性炎症的一个重要机制。     

伏格特-小柳-原田综合征患者外周血淋巴细胞Fas和FasL mRNA的表达

目的 探讨Fas、FasL mRNA在伏格特-烛柳-原田(Vogt-Koyanagi-Harada,VKH)综合征患者外周血淋巴细胞的表达。方法 于2000年1～6月抽取16例VKH综合征患者和19例正常人的外周血,提取淋巴细胞的总RNA,将RNA逆转录为cDNA,然后用荧光定量聚合酶链反应(fluorescent quantitative polymerase chain reaction,FQ-PCR)的方法实时检测Fas、FasL PCR扩增的全过程,计算样本中Fas、FasL mRNA的表达量。结果 VKH综合征患者外周血淋巴细胞的Fas mRNA的表达量[(1.6±2.0)×106]显著高于正常对照组[(5.7±2.0)×105](t=4.50,P<0.05),FasLmRNA的表达量[(1.8±1.5)×106]也显著高于正常对照组[(4.8±3.5)×105](t=9.57,P<0.05)。结论 VKH综合征患者外周血中长期存在的大量Fas、FasL mRNA高表达的活化自身免疫性淋巴细胞,可能是其炎性反应反复发作、病情迁延不愈的重要原因之一。(中华眼科杂志,2004,40:507-509)     

Immunologic analysis of cerebrospinal fluid lymphocytes in Vogt-Koyanagi-Harada disease

In this study, we focused upon the immunologic aspects of Vogt-Koyanagi-Harada disease (VKH) by comparing the cytotoxic activity of peripheral blood leukocytes (PBL) to that of cerebrospinal fluid leukocytes (CSFL) against the human melanoma cell line (P-36) and the human cervical carcinoma cell line (HeLa-S3). The PBL from patients with VKH showed significant cytotoxic activity against P-36 (P less than 0.01), but did not show cytotoxic activity against HeLa-S3. The CSFL showed significantly weaker cytotoxic activity against P-36 compared to that of PBL (P less than 0.02). We also analyzed the cell membrane surface markers applying monoclonal antibodies on PBL and CSFL. The percentage of OKT8+ (CD8: T cytotoxic/suppressor lymphocytes) cells was significantly lower in CSFL than in PBL (P less than 0.05). There was a tendency toward a higher percentage of HLA-DR+ cells (B lymphocytes, monocytes, macrophages, and activated T lymphocytes) and a higher ratio of OKT4+/8+ cells (CD4/CD8: T helper/inducer lymphocytes/T cytotoxic/suppressor lymphocytes) in CSFL from patients with VKH than in their PBL (P less than 0.1).     

Identification of autoreactive T cells in Vogt-Koyanagi-Harada disease

PURPOSE: To determine the finer specificity and immunologic features of autoreactive T cells in Vogt-Koyanagi-Harada (VKH) disease. METHODS: T-cell clones (TCCs ) specific to tyrosinase family proteins were raised from the peripheral blood mononuclear cells (PBMCs) of patients with VKH disease, and the response of the TCCs to 30-mer peptides was determined. The TCCs that were reactive to the peptides with strong binding sites for HLA DRB1*0405 were initially tested. Then, a finer specificity of these TCCs against 12- to 14-mer peptides was determined. The cytokine production of these clones was measured by ELISA. RESULTS: A total of 62 stable TCCs were established from the PBMCs of five patients with VKH (28 clones against tyrosinase, 34 clones against tyrosinase-related protein [TRP]1). Five of 28 TCCs for tyrosinase and 2 of 34 for TRP1 were reactive to the 30-mer peptides with strong binding sites for HLA DRB1*0405. These seven clones showed proliferative responses to one or more of the 12- to 14-mer peptides that match the motif of the strong binding site for HLADRB1*0405. Five of seven of the TCCs may be T-helper (Th) type 1, one of the remaining TCCs may be Th0, and the other may be Th2. CONCLUSIONS: The autoreactive T cells against tyrosinase and/or TRP1 may contribute to the development of VKH disease.     

Enumeration of autoreactive helper T lymphocytes in uveitis.

In a one-stage, interleukin-2 (IL-2), limiting-dilution analysis, peripheral blood mononuclear cells from patients with uveitis and normal control subjects were assayed for S-antigen specific, tetanus-specific, and in vivo activated helper T cells. Controls subjects consistently demonstrated tetanus-specific responses, but neither in vivo activation nor S-antigen specific helper T cell responses were seen. Patients with active forms of diffuse, posterior, and anterior uveitis were found to have significant frequencies of both in vivo activated and S-antigen specific helper T cells in their peripheral blood. These data show that patients with certain forms of uveitis have a measurable frequency of lymphocytes in the peripheral immunologic compartment capable of secreting IL-2 in response to autologous presentation of ocular autoantigen (S-antigen). Limiting-dilution analysis techniques, generating minimal responder cell frequency estimates and distinct IL secretion patterns, may provide an index of disease activity and critical information about the mechanism(s) of ocular inflammation.     

High inducibility of Epstein-Barr virus replication in B lymphocytes in Vogt-Koyanagi-Harada disease]

PURPOSE: The role of Epstein-Barr virus (EBV) reactivation in the pathogenesis of Vogt-Koyanagi-Harada (VKH) disease was examined. MATERIAL AND METHODS: Using B lymphocytes obtained from 8 patients with VKH disease and 10 patients with other types of uveitis, immortarized lymphoblast lines were established and infected with EBV. The degree of EBV activation in each lymphoblast line, in the presence and absence of various stimuli, was assessed by measuring the expression of 3 different antigens involved in replication by immunofluorescent staining and western blot analysis. Quantification of EBV DNA in cell culture supernatants was done by polymerase chain reaction. RESULT: Cell lines established from VKH patients expressed more viral antigens that those established from patients with other types of uveitis. There were greater amounts of EBV DNA in the VKH cell lines. CONCLUSION: B lymphocytes from VKH patients may be more susceptible to EBV activation, and the reactivation of EBV may be involved in the pathogenesis of VKH.     

Thymus-derived lymphocytes in the Vogt-Koyanagi-Harada syndrome.

Previous investigations have suggested that immunologic factors may play a role in the Vogt-Koyanagi-Harada syndrome, a disease involving the eye, central nervous system, and dermis. Among patients with this disease we have observed a decrease in their peripheral blood lymphocytes, as measured by two types of rosette assays. These results strengthen the concept that an altered immunologic status may be important in the pathophysiology of this disease.     

Behcet病患者外周血淋巴细胞Fas、FasL mRNA的表达及意义

目的 :探讨Fas、FasLmRNA在Behcet病患者外周血淋巴细胞的表达及可能的意义。方法 :抽取 19例Behcet病患者和 19个正常人的外周血 ,提取淋巴细胞的总RNA ,然后用荧光定量聚合酶链反应 (fluorescentquantitativepolymerasechainreaction ,FQ -PCR)的方法实时检测Fas、FasLPCR扩增的全过程 ,计算样本中Fas、FasLmRNA的表达量。结果 :Behcet病患者外周血淋巴细胞的FasmRNA的表达量 ( 1 5 0× 10 6± 0 77× 10 6)显著高于正常对照组 ( 0 5 7× 10 6± 0 2 0×10 6) (P <0 0 5 ) ,FasLmRNA的表达量 ( 2 0 0× 10 6± 2 2 5× 10 6)也显著高于正常对照组 ( 0 48× 10 6± 0 35× 10 6) (P <0 0 1)。结论 :Behcet病患者外周血中存在大量Fas、FasLmRNA高表达的活化的自身免疫性淋巴细胞 ,这些细胞的长期存在可能是患者炎症反复发作、病情迁延不愈的重要原因之一。     

Studies on corticosteroid therapy in Vogt-Koyanagi-Harada disease

We evaluated the significance of corticosteroid therapy on 47 new patients with Vogt-Koyanagi-Harada disease examined in the Uveitis Survey Clinic of the Hokkaido University Hospital. All patients were treated with topical corticosteroids, with or without systemic corticosteroids. 18 patients received systemic corticosteroid as pulse therapy, 20 patients received high-dose corticosteroid starting with prednisolone 200 mg, 2 patients received conventional-dose corticosteroid and 7 patients received no systemic corticosteroid therapy. When we evaluated the results 6 months after the initiation of treatment, anterior chamber inflammation was significantly less in patients with pulse and high-dose corticosteroid therapy than in those without systemic corticosteroid therapy. Furthermore, final visual acuity was significantly better in patients with pulse and high-dose corticosteroid than in those without them. However, there was no significant difference between patients with pulse therapy and those with high-dose corticosteroid therapy. The findings are in support of systemic corticosteroid therapy (pulse and high-dose) in the treatment of Vogt-Koyanagi-Harada disease.     

Choroidal folds in Vogt-Koyanagi-Harada disease

PURPOSE: To analyze choroidal folds in Vogt-Koyanagi-Harada (VKH) disease by fluorescein angiography (FA), indocyanine green angiography (ICGA), and optical coherence tomography (OCT). DESIGN: Retrospective, consecutive case series. METHODS: Records of 95 patients diagnosed with VKH disease from October 2001 to July 2006 were reviewed. All patients underwent FA, 17 patients underwent ICGA, and 20 underwent OCT. RESULTS: Of the 95 patients, 11 (12.0%) had choroidal folds and showed 10 to 15 hypofluorescent bands radiating from the optic disk that were similar to the large retinal vessels in shape and number on FA. On ICGA, the choroidal folds showed hyperfluorescence at the late stage. OCT showed clear folds but the number of folds was larger than on FA and ICGA. CONCLUSIONS: Choroidal folds are not uncommon in VKH disease. FA, ICGA, and OCT can help to identify their pathogenesis.