Hughes C C, Pober J S
Molecular Cardiobiology Program, Boyer Center for Molecular Medicine, Yale University School of Medicine, New Haven, CT 06536.
J Immunol. 1993 Apr 15;150(8 Pt 1):3148-60.
Endothelial cells (EC) act as APC for resting PBL in vitro, and may have important roles in vivo in the pathogenesis of allograft rejection and delayed hypersensitivity. We previously reported that human umbilical vein EC provide costimulatory signals to PHA-stimulated PBL via CD2:lymphocyte function-associated Ag-3 and an unidentified ligand pair, resulting in a three- to eight-fold enhancement of IL-2 production. The physiologic relevance of this increase was demonstrated by the proliferative advantage provided by EC to PBL suboptimally stimulated with mAb OKT3. We now report that EC costimulation causes increased levels of IL-2 mRNA as a result of increased IL-2 transcription in PBL. We therefore examined the effects of EC on T cell nuclear factors known to regulate IL-2 transcription, including c-jun and c-fos-two components of the transcription factor AP-1, NFAT, and others. PBL constitutively express c-jun transcripts, and the level of c-jun mRNA is not altered by PHA activation in the absence or presence of EC. In contrast, c-fos mRNA is absent from resting T cells and is induced on PHA activation. EC alone do not induce c-fos mRNA but augment the level of c-fos mRNA in PHA-activated T cells by 3- to 10-fold. This effect is largely independent of the CD2:lymphocyte function-associated Ag-3 pathway. Gel-shift analysis reveals the constitutive presence of nuclear factors in resting PBL that bind to the proximal AP-1 site of the IL-2 promoter and that contain immunoreactive c-Jun but not c-Fos protein. In contrast, AP-1 from PHA-activated cells contains c-Jun and low levels of c-Fos. Strikingly, costimulation with EC results in a dramatic increase (up to 15-fold) in the c-Fos content of AP-1. Levels of other nuclear factors involved in IL-2 regulation were not altered by EC, although NFAT-DNA complexes migrated at a slightly different mobility. In summary, our data suggest that changes in the composition of transcription factor AP-1 is a key molecular mechanism for increasing IL-2 transcription and may underlie the phenomenon of costimulation by EC.
内皮细胞（EC）在体外可作为静息外周血淋巴细胞（PBL）的抗原呈递细胞，在体内同种异体移植排斥反应和迟发型超敏反应的发病机制中可能发挥重要作用。我们之前报道过，人脐静脉内皮细胞通过CD2：淋巴细胞功能相关抗原-3和一对未明确的配体对外周血淋巴细胞（PBL）提供共刺激信号，从而使白细胞介素-2（IL-2）的产生增加三到八倍。内皮细胞对次优剂量抗CD3单克隆抗体（mAb OKT3）刺激的外周血淋巴细胞（PBL）所提供的增殖优势，证明了这种增加的生理相关性。我们现在报道，内皮细胞共刺激导致外周血淋巴细胞（PBL）中IL-2转录增加，从而使IL-2信使核糖核酸（mRNA）水平升高。因此，我们研究了内皮细胞对已知调节IL-2转录的T细胞核因子的影响，这些因子包括转录因子激活蛋白-1（AP-1）的两个组成部分c-jun和c-fos、活化T细胞核因子（NFAT）等。外周血淋巴细胞（PBL）组成性表达c-jun转录本，在有无内皮细胞的情况下，PHA激活均不会改变其信使核糖核酸（mRNA）水平。相反，静息T细胞中不存在c-fos信使核糖核酸（mRNA），PHA激活可诱导其产生。单独的内皮细胞不会诱导c-fos信使核糖核酸（mRNA），但可使PHA激活的T细胞中c-fos信使核糖核酸（mRNA）水平提高3至10倍。这种效应在很大程度上不依赖于CD2：淋巴细胞功能相关抗原-3途径。凝胶迁移分析显示，静息外周血淋巴细胞（PBL）中存在与IL-2启动子近端AP-1位点结合的核因子，这些核因子含有免疫反应性c-Jun但不含c-Fos蛋白。相反，PHA激活细胞中的AP-1含有c-Jun和低水平的c-Fos。引人注目的是，内皮细胞共刺激导致AP-1中c-Fos含量显著增加（高达15倍）。尽管NFAT-DNA复合物迁移时迁移率略有不同，但内皮细胞不会改变其他参与IL-2调节的核因子水平。总之，我们的数据表明，转录因子AP-1组成的变化是增加IL-2转录的关键分子机制，可能是内皮细胞共刺激现象的基础。
