Sarkhosh Kourosh, Tredget Edward E, Li Yunyuan, Kilani Ruhangiz T, Uludag Hasan, Ghahary Aziz
Wound Healing Research Group, Department of Surgery, University of Alberta, Edmonton, Alberta, Canada T6G 2E1.
Wound Repair Regen. 2003 Sep-Oct;11(5):337-45. doi: 10.1046/j.1524-475x.2003.11505.x.
Indoleamine 2,3-dioxygenase (IDO) is an intracellular tryptophan-oxidizing enzyme possessing various immunosuppressive characteristics. In this study, we report the possible use of this enzyme in an allogenic skin substitute to suppress the proliferation of immune cells. Human fetal skin fibroblasts and keratinocytes were treated with the cytokine interferon-gamma to induce expression of IDO mRNA and protein. IDO enzyme activity was evaluated by measurement of kynurenine levels in the interferon-gamma-treated and -untreated cells. Results of Northern analysis showed a dose-dependent response in expression of IDO mRNA to the various concentrations of interferon-gamma used. Northern blot analysis also showed a time-dependent expression of IDO in response to different durations of interferon-gamma treatment. The level of kynurenine measured, as the bioactivity of IDO enzyme, was significantly higher in the interferon-gamma-treated fibroblasts and keratinocytes compared to those of controls (p < 0.001). To illustrate the immunosuppressive effects of IDO on immune cell proliferation, IDO-expressing fibroblasts were cocultured with human peripheral blood mononuclear cells for a period of 5 days. Results of 3H-thymidine incorporation assays showed a significant reduction in proliferation of the mononuclear cells cocultured with IDO-expressing skin cells compared to monocytes cocultured with control (non-IDO-expressing) skin cells (p < 0.001). Furthermore, addition of the IDO-inhibitor (1-methyl-D-tryptophan) significantly reversed the immunosuppressive effects of IDO on monocyte proliferation (p < 0.001). In conclusion, suppression of peripheral blood mononuclear cell proliferation due to interferon-gamma-induced IDO-expression in allogenic human skin cells might shed new light on developing a nonrejectable allogenic skin substitute.
吲哚胺2,3-双加氧酶（IDO）是一种具有多种免疫抑制特性的细胞内色氨酸氧化酶。在本研究中，我们报道了该酶在同种异体皮肤替代物中可能用于抑制免疫细胞增殖的情况。用人细胞因子γ干扰素处理人胎儿皮肤成纤维细胞和角质形成细胞，以诱导IDO mRNA和蛋白的表达。通过测量γ干扰素处理和未处理细胞中的犬尿氨酸水平来评估IDO酶活性。Northern分析结果显示，IDO mRNA的表达对所用不同浓度的γ干扰素呈剂量依赖性反应。Northern印迹分析还显示，IDO的表达对γ干扰素不同处理持续时间呈时间依赖性。作为IDO酶生物活性所测量的犬尿氨酸水平，在γ干扰素处理的成纤维细胞和角质形成细胞中显著高于对照组（p<0.001）。为了说明IDO对免疫细胞增殖的免疫抑制作用，将表达IDO的成纤维细胞与人外周血单个核细胞共培养5天。3H-胸腺嘧啶核苷掺入试验结果显示，与与对照（不表达IDO）皮肤细胞共培养的单核细胞相比，与表达IDO的皮肤细胞共培养的单核细胞增殖显著降低（p<0.001）。此外，添加IDO抑制剂（1-甲基-D-色氨酸）可显著逆转IDO对单核细胞增殖的免疫抑制作用（p<0.001）。总之，同种异体人皮肤细胞中γ干扰素诱导的IDO表达导致外周血单个核细胞增殖受到抑制，这可能为开发不可排斥的同种异体皮肤替代物提供新的思路。
