姜黄素阻断人T细胞增殖中对环孢素A耐药的CD28共刺激途径。

Curcumin blocks cyclosporine A-resistant CD28 costimulatory pathway of human T-cell proliferation.

作者信息

Ranjan D, Johnston T D, Wu G, Elliott L, Bondada S, Nagabhushan M

机构信息

Department of Surgery, University of Kentucky, Lexington, Kentucky, 40536, USA.

出版信息

J Surg Res. 1998 Jul 1;77(2):174-8. doi: 10.1006/jsre.1998.5374.

DOI:10.1006/jsre.1998.5374

PMID:9733605

Abstract

INTRODUCTION

Curcumin (Cur) is a phenolic component of common spice, turmeric. We have reported earlier that it possesses antineoplastic and immunosuppressive properties in vitro. It has been reported that cyclosporine A (CyA), a commonly used immunosuppressant does not inhibit CD28 costimulatory pathway of T-cell activation. We hypothesized that Cur, a tyrosine kinase inhibitor, would block CyA-resistant CD28 costimulatory pathway of human T cell proliferation.

MATERIALS AND METHODS

Human T-lymphocytes were isolated from healthy donors using gradient centrifugation and rosetting techniques. In four separate experiments T-cells were plated in triplicate in 96-well plates at a density of 2X105 cells/well. These cells were stimulated with 0.5 ng/ml phorbol myristate acetate (PMA) + 0.5 (g/ml anti-CD28 antibody (PMA-CD28 group) or 2.5 microgram/ml PHA (PHA group). Cur or CyA at varying concentrations (0.31, 0.625, 1.25, 2.5, 5, or 10 microgram/ml and 1.25, 2.5, 5, 10, 20, or 250 ng/ml, respectively) was added and cellular proliferation was measured by the uptake of [3H]thymidine and is reported (mean cpm/well(SD). Cells from the PMA-CD28 group that were treated with either curcumin or 0.4% DMSO (vehicle control for curcumin) were studied for evidence of apoptosis by staining with viable dyes MC540 and Hoechst 33342 and subsequently analyzed in the cell sorter.

RESULTS

Cur caused a concentration-dependent inhibition of T-cell proliferation in the PMA-CD28 group (from 32775 +/- 3084 to 66 +/- 42 at 5.0 microgram/ml of cur) and PHA group (from 50956 +/- 5747 to 24 +/- 12 at 5.0 microgram/ml) with a calculated ED50 of 3.5 and 7.7, microM respectively. CyA inhibited T-cell proliferation in the PHA group with a calculated ED50 of 2.7 ng/ml but failed to block PMA + anti-CD28-stimulated T-cell proliferation even at 250 ng/ml. PMA-CD28 group cells treated with 10 microgram/ml curcumin showed a significantly increased apoptosis as compared to control (0.4% DMSO).

CONCLUSION

Since Cur blocks the CyA-resistant PMA + anti-CD28 pathway of T-cell proliferation, it may have novel adjuvant immunosuppressive properties.

摘要

引言

姜黄素（Cur）是常见香料姜黄中的一种酚类成分。我们之前报道过它在体外具有抗肿瘤和免疫抑制特性。据报道，常用的免疫抑制剂环孢素A（CyA）不会抑制T细胞活化的CD28共刺激途径。我们推测，作为一种酪氨酸激酶抑制剂的Cur会阻断人T细胞增殖中对CyA耐药的CD28共刺激途径。

材料与方法

使用梯度离心和红细胞吸附技术从健康供体中分离出人T淋巴细胞。在四个独立实验中，将T细胞以2×105个细胞/孔的密度一式三份接种于96孔板中。这些细胞用0.5 ng/ml佛波醇肉豆蔻酸酯乙酸酯（PMA）+0.5μg/ml抗CD28抗体（PMA-CD28组）或2.5μg/ml植物血凝素（PHA）刺激。加入不同浓度的Cur或CyA（分别为0.31、0.625、1.25、2.5、5或10μg/ml和1.25、2.5、5、10、20或250 ng/ml），通过[3H]胸腺嘧啶核苷摄取量测量细胞增殖，并报告结果（平均cpm/孔（标准差））。用活细胞染料MC540和Hoechst 33342染色，研究PMA-CD28组中用姜黄素或0.4%二甲基亚砜（姜黄素的溶剂对照）处理的细胞的凋亡证据，随后在细胞分选仪中进行分析。

结果

Cur对PMA-CD28组（在5.0μg/ml Cur时从32775±3084降至66±42）和PHA组（在5.0μg/ml时从50956±5747降至24±12）的T细胞增殖产生浓度依赖性抑制，计算得出的半数有效剂量（ED50）分别为3.5和7.7μM。CyA对PHA组的T细胞增殖有抑制作用，计算得出的ED50为2.7 ng/ml，但即使在250 ng/ml时也未能阻断PMA+抗CD28刺激的T细胞增殖。与对照组（0.4%二甲基亚砜）相比，用10μg/ml姜黄素处理的PMA-CD28组细胞凋亡显著增加。