Ahn Soon Cheol, Oh Won Keun, Kim Bo Yeon, Kang Dae Ook, Kim Min Soo, Heo Gun Young, Ahn Jong Seog
Korea Research Institute of Bioscience and Biotechnology, Yusong, Taejon, Korea.
Planta Med. 2003 Jul;69(7):642-6. doi: 10.1055/s-2003-41111.
In the course of screening inhibitors from the methanol (MeOH) extracts of 168 medicinal plants against lymphocyte cell-specific kinase (Lck) Src -homology 2 (SH2) binding to a synthetic phosphotyrosine-containing peptide (phosphopeptide), we isolated rosmarinic acid from the MeOH extract of Prunella vulgaris, which showed specific inhibitory activity. The IC 50 value for Lck SH2 binding to phosphopeptide (SGSGEEPQpYEEIPI) of hamster polyomavirus middle-sized tumor (hmT pY324) was 7 microM. However, even at concentrations of 0.1 to 1000 microM, no significant inhibitions were observed against other SH2 domains binding such as the growth factor receptor binding protein 2 (Grb2) SH2 domain to phosphopeptide of Shc and phospholipase Cgamma1 (PLCgamma1) SH2 domain to translational elongation factor 1alpha (EF1alpha) C-terminal. Rosmarinic acid inhibited interleukin-2 (IL-2) gene expression by 50 % at a concentration of 8 microM in Jurkat cells stimulated with anti-CD3 and anti-CD4 antibodies. FK506 and cyclosporin A (CsA) employed as positive controls showed less than 30 % inhibition at the same concentration. In addition, rosmarinic acid inhibited the intracellular [Ca 2+] i increase in Jurkat cells after T cell activation in a dose-dependent manner at concentrations of 1.4 to 140 microM of rosmarinic acid, which is one of the earliest responses of antigen-specific T cell receptor (TCR) and of the upstream pathway of IL-2 expression. Taken together, these results suggest that rosmarinic acid has the potential to specifically inhibit Lck SH2 domain binding to its cognate ligand, including ZAP-70, Cbl, HS-1, and PLCgamma1, and Lck-dependent Ca 2+ signaling pathway of its downstream effector and finally to modulate IL-2 gene expression after T cell activation.
在从168种药用植物的甲醇提取物中筛选针对淋巴细胞特异性激酶（Lck）Src同源2（SH2）结构域与合成的含磷酸酪氨酸肽（磷酸肽）结合的抑制剂的过程中，我们从夏枯草的甲醇提取物中分离出了迷迭香酸，其显示出特异性抑制活性。Lck SH2与仓鼠多瘤病毒中型肿瘤（hmT pY324）的磷酸肽（SGSGEEPQpYEEIPI）结合的IC50值为7 microM。然而，即使在0.1至1000 microM的浓度下，对于其他SH2结构域的结合也未观察到显著抑制，例如生长因子受体结合蛋白2（Grb2）SH2结构域与Shc的磷酸肽结合以及磷脂酶Cγ1（PLCγ1）SH2结构域与翻译延伸因子1α（EF1α）C末端结合。在抗CD3和抗CD4抗体刺激的Jurkat细胞中，迷迭香酸在8 microM的浓度下可抑制白细胞介素-2（IL-2）基因表达50%。用作阳性对照的FK506和环孢素A（CsA）在相同浓度下显示出不到30%的抑制作用。此外，在1.4至140 microM的迷迭香酸浓度下，迷迭香酸以剂量依赖性方式抑制T细胞活化后Jurkat细胞内[Ca2+]i的增加，这是抗原特异性T细胞受体（TCR）最早的反应之一以及IL-2表达上游途径的反应之一。综上所述，这些结果表明迷迭香酸有可能特异性抑制Lck SH2结构域与其同源配体（包括ZAP-70、Cbl、HS-1和PLCγ1）的结合，以及其下游效应器的Lck依赖性Ca2+信号通路，并最终调节T细胞活化后的IL-2基因表达。
