Babcock T A, Carlin J M
Department of Microbiology, Miami University, Oxford, Ohio, 45056, USA.
Cytokine. 2000 Jun;12(6):588-94. doi: 10.1006/cyto.1999.0661.
Interferon (IFN)-gamma-induced indoleamine 2,3-dioxygenase (IDO) activity is enhanced synergistically by interleukin (IL-)1, tumor necrosis factor-alpha (TNF-alpha) and LPS in IFN-treated macrophages by increasing IDO mRNA concentration. These studies demonstrate that IFN-treated HeLa cells also exhibit dose-dependent enhancement of IDO induction by TNF-alpha and IL-1, with maximal effects at concentrations of 5 ng/ml and 3 ng/ml, respectively. Furthermore, with sub-optimal IFN concentrations, cells treated with maximally effective concentrations of TNF-alpha or IL-1alpha required 3-5 times less IFN to induce the same level of IDO activity as that observed with IFN alone. To detect changes in transcriptional activation of the IDO gene, HeLa cells were transfected with a plasmid containing the IDO 5' regulatory region upstream of a green fluorescent protein (GFP) reporter gene. In transfected cells, IFN induced both IDO activity and GFP that was detected by flow cytometry. When cell-sorted, transfected cells were stimulated with IFN in combination with TNF-alpha or IL-1 but not LPS, increased GFP was detected in comparison to transfected cells treated with IFN alone. Furthermore, increases in GFP expression correlated with IDO enzymatic activity, indicating that combinations of IFN with IL-1 or TNF-alpha increase the transcriptional activity of the IDO promoter region.
在经干扰素处理的巨噬细胞中，干扰素（IFN）-γ诱导的吲哚胺2,3-双加氧酶（IDO）活性通过增加IDO mRNA浓度，被白细胞介素（IL-）1、肿瘤坏死因子-α（TNF-α）和脂多糖协同增强。这些研究表明，经干扰素处理的HeLa细胞也表现出TNF-α和IL-1对IDO诱导的剂量依赖性增强，分别在浓度为5 ng/ml和3 ng/ml时达到最大效应。此外，在次优干扰素浓度下，用最大有效浓度的TNF-α或IL-1α处理的细胞诱导相同水平的IDO活性所需的干扰素比单独使用干扰素时少3至5倍。为了检测IDO基因转录激活的变化，用含有绿色荧光蛋白（GFP）报告基因上游IDO 5'调控区的质粒转染HeLa细胞。在转染细胞中，干扰素诱导IDO活性和GFP，通过流式细胞术检测。当对细胞进行分选后，将转染细胞与干扰素联合TNF-α或IL-1刺激，但不与脂多糖刺激，与单独用干扰素处理的转染细胞相比，检测到GFP增加。此外，GFP表达的增加与IDO酶活性相关，表明干扰素与IL-1或TNF-α的组合增加了IDO启动子区域的转录活性。
